Inhibition of protein kinase CK2 reduces CYP24A1 expression and enhances 1,25-dihydroxyvitamin D3 anti-tumor activity in human prostate cancer cells

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Luo, Wei; Yu, Wei-Dong; Ma, Yingyu; Chernov, Mikhail; Trump, Donald L.; Johnson, Candace S.

2013-01-01

Vitamin D has broad range of physiological functions and anti-tumor effects. 24-hydroxylase, encoded by the CYP24A1 gene, is the key enzyme for degrading many forms of vitamin D including the most active form, 1,25D3. Inhibition of CYP24A1 enhances 1,25D3 anti-tumor activity. In order to isolate regulators of CYP24A1 expression in prostate cancer cells, we established a stable prostate cancer cell line PC3 with CYP24A1 promoter driving luciferase expression to screen a small molecular library for compounds that inhibit CYP24A1 promoter activity. From this screening, we identified, 4,5,6,7-tetrabromobenzimidazole (TBBz), a protein kinase CK2 selective inhibitor as a disruptor of CYP24A1 promoter activity. We show that TBBz inhibits CYP24A1 promoter activity induced by 1,25D3 in prostate cancer cells. In addition, TBBz downregulates endogenous CYP24A1 mRNA level in TBBz treated PC3 cells. Furthermore, siRNA-mediated CK2 knockdown reduces 1,25D3 induced CYP24A1 mRNA expression in PC3 cells. These results suggest that CK2 contributes to 1,25D3 mediated target gene expression. Lastly, inhibition of CK2 by TBBz or CK2 siRNA significantly enhanced 1,25D3 mediated anti-proliferative effect in vitro and in vivo in a xenograft model. In summary, our findings reveal that protein kinase CK2 is involved in the regulation of CYP24A1 expression by 1,25D3 and CK2 inhibitor enhances 1,25D3 mediated anti-tumor effect. PMID:23358686

A Comparative Analysis of Cytokeratin 18 and 19 Expressions in Odontogenic Keratocyst, Dentigerous Cyst and Radicular Cyst with a Review of Literature

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Shah, Vandana Sandip; Ghanchi, Mohsin Jiva; Gosavi, Sandesh Sachchidanand; Srivastava, Himanshu Mahesh; Pachore, Nivedita Javahir

2016-01-01

Introduction Odontogenic cysts viz Odontogenic Keratocyst (OKC), Dentigerous Cyst (DC) and Radicular Cyst (RC) occur commonly in the oral and maxillofacial region. Cytokeratin (CK) expression studies have been done to evaluate diagnostic accuracy, role in pathogenesis, elucidate behaviour and role in treatment protocols. However, variations have been reported in the expression of CK patterns in these odontogenic cysts, which could be due to the lack of standardization of laboratory techniques. The present study has tried to shed light on CK 18 and 19 expression in odontogenic cysts and offer the brief review of previous studies on these CK. Aim The aim of the present study was to evaluate the intensity and expression patterns of CK 18 and 19 in OKCs, DCs and RCs. Materials and Methods A total of 60 cases, 20 each of OKC, DC and RC were confirmed histologically and evaluated for immunohistochemical expression pattern and intensity of CK 18 and 19. Results A focal and variable expression of CK 18 was observed in 25% of OKCs, 15% of DCs and 10% of RCs. CK 19 was expressed in 75% of OKCs and 100% in DCs as well as RCs. Conclusion The intensity and expression of Cytokeratin 19 was more in all three cysts compared to Cytokeratin 18. PMID:27630961

Pressure overload stimulated cardiac hypertrophy leads to a rapid decrease in the mRNA for creatine kinase

International Nuclear Information System (INIS)

Boheler, K.; Popovich, B.; Dillmann, W.H.

1987-01-01

Cardiac hypertrophy (CH) leads to a decrease in creatine kinase (CK) enzymatic activity. To determine if the mRNA for CK also decreases with CH, they performed the following studies. Cardiac RNA was isolated from rats subjected to either abdominal aortic stenosis (AS) or sham surgery. Through Northern blot analysis, total cardiac RNA was quantitated with a CK specific 32 P-labelled cDNA clone. At 3 and 8 days post-constriction, the mRNA for CK decreases by 54.6 +/- 7% and 65.3 +/- 18% respectively, whereas the heart weight increases by 19% and 37% relative to controls. Further studies indicate that CK mRNA also decreases by 41.8% in hypothyroid rats (Tx) but decreases by a total of 68.1% in Tx rats subjected to 8 days of AS. Pressure overload stimulated CH leads to a rapid decrease in CK mRNA in normal and Tx rats. This CK mRNA decrease may account for the decreased efficiency of contraction seen in CH

Impact of gastro-esophageal reflux on mucin mRNA expression in the esophageal mucosa.

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van Roon, Aafke H C; Mayne, George C; Wijnhoven, Bas P L; Watson, David I; Leong, Mary P; Neijman, Gabriëlle E; Michael, Michael Z; McKay, Andrew R; Astill, David; Hussey, Damian J

2008-08-01

Changes in the expression of mucin genes in the esophageal mucosa associated with uncomplicated gastro-esophageal reflux disease have not been evaluated even though such changes could be associated with reflux-induced mucosal damage. We therefore sought to identify reflux-induced changes in mucin gene expression using a cell line and biopsies from the esophageal mucosa in patients with and without reflux. MUC-1, MUC-3, MUC-4, and MUC-5AC gene expressions were investigated in the HET-1A cell line following exposure to acid (pH 4) and/or bile (120 muM of a bile salt milieu), and in esophageal mucosal biopsies from controls, subjects with non-erosive gastro-esophageal reflux, and subjects with reflux associated with ulcerative esophagitis (erosive). The mucosal biopsies were also evaluated for IL-6 mRNA expression (inflammatory marker) and CK-14 mRNA expression (mucosal basal cell layer marker). Gene expression was determined using real-time reverse transcriptase-polymerase chain reaction analysis. In the cell line studies, there were differences in mRNA levels for all of the evaluated mucins following treatment with either acid or the acid and bile combination. In the studies which evaluated tissue specimens, IL-6 and CK-14 mRNA levels increased according to degree of reflux pathology. The expression of MUC-1 and MUC-4 in mucosa from patients with erosive reflux was lower than in subjects without reflux and in patients with non-erosive reflux, whereas the expression of MUC-3 and MUC-5AC was increased (although these differences did not reach significance at p reflux groups. The correlation between IL-6 and MUC-3 was significant within the control and erosive reflux groups, and the correlation between MUC-1 and MUC-5AC was significant within the erosive reflux group. The results of this study suggest that the profile of mucin expression in the esophageal mucosa is influenced by the pH and composition of the gastro-esophageal reflux. Further work should explore the

Protein Kinase CK2 Expression Predicts Relapse Survival in ERα Dependent Breast Cancer, and Modulates ERα Expression in Vitro

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Marlon D. Williams

2015-12-01

Full Text Available The heterotetrameric protein kinase CK2 has been associated with oncogenic transformation, and our previous studies have shown that it may affect estrogenic signaling. Here, we investigate the role of the protein kinase CK2 in regulating ERα (estrogen receptor α signaling in breast cancer. We determined the correlation of CK2α expression with relapse free breast cancer patient survival utilizing Kaplan Meier Plotter (kmplot.com/analysis/ to mine breast cancer microarrays repositories. Patients were stratified according to ERα status, histological grade, and hormonal therapy. Luciferase reporter assays and flow cytometry were implemented to determine the impact of CK2 inhibition on ERE-mediated gene expression and expression of ERα protein. CK2α expression is associated with shorter relapse free survival among ERα (+ patients with grade 1 or 2 tumors, as well as among those patients receiving hormonal therapy. Biochemical inhibition of CK2 activity results in increased ER-transactivation as well as increased expression among ERα (+ and ERα (− breast cancer cell lines. These findings suggest that CK2 may contribute to estrogen-independent cell proliferation and breast tumor progression, and may potentially serve as a biomarker and pharmacological target in breast cancer.

Expression and characterization of a recombinant maize CK-2 alpha subunit

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Boldyreff, B; Meggio, F; Dobrowolska, G

1993-01-01

to support the immunological data also by biochemical and biophysical experiments the availability of a recombinant CK-2 alpha from maize was a prerequisite. A maize cDNA clone of maize CK-2 alpha was expressed in the bacterial strain BL21 (DE3). The recombinant protein was purified to homogeneity; its......CKIIB, one of the CK-2 like enzymes which have been isolated from maize, has been shown to be a monomeric enzyme that cross-reacts with anti CK-2 alpha specific antibodies suggesting a possible relationship between the two proteins (Dobrowolska et al. (1992) Eur. J. Biochem. 204, 299-303). In order...... molecular mass on one-dimensional SDS PAGE was estimated to be 36.5 kDa. The calculated molecular mass according to the amino acid composition is 39,228 Da (332 amino acids). The recombinant maize CK-2 alpha (rmCK-2 alpha) exhibited mostly the same properties as the recombinant human CK-2 alpha (rhCK-2...

Fluorescent metal nanoshell and CK19 detection on single cell image

International Nuclear Information System (INIS)

Zhang, Jian; Fu, Yi; Li, Ge; Lakowicz, Joseph R.; Zhao, Richard Y.

2011-01-01

Highlights: → Novel metal nanoshell as fluorescence imaging agent. → Fluorescent mAb-metal complex with enhanced intensity and shortened lifetime. → Immuno-interactions of mAb-metal complexes with CK19 molecules on CNCAP and HeLa cell surfaces. → Isolation of conjugated mAb-metal complexes from cellular autofluorescence on cell image. -- Abstract: In this article, we report the synthesis strategy and optical properties of a novel type of fluorescence metal nanoshell when it was used as imaging agent for fluorescence cell imaging. The metal nanoshells were made with 40 nm silica cores and 10 nm silver shells. Unlike typical fluorescence metal nanoshells which contain the organic dyes in the cores, novel metal nanoshells were composed of Cy5-labelled monoclonal anti-CK19 antibodies (mAbs) on the external surfaces of shells. Optical measurements to the single nanoparticles showed that in comparison with the metal free labelled mAbs, the mAb-Ag complexes displayed significantly enhanced emission intensity and dramatically shortened lifetime due to near-field interactions of fluorophores with metal. These metal nanoshells were found to be able to immunoreact with target cytokeratin 19 (CK19) molecules on the surfaces of LNCAP and HeLa cells. Fluorescence cell images were recorded on a time-resolved confocal microscope. The emissions from the metal nanoprobes could be clearly isolated from the cellular autofluorescence backgrounds on the cell images as either individuals or small clusters due to their stronger emission intensities and shorter lifetimes. These emission signals could also be precisely counted on single cell images. The count number may provide an approach for quantifying the target molecules in the cells.

Nuclear localization of the CK2α-subunit correlates with poor prognosis in Clear Cell Renal Cell Carcinoma

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Rabjerg, Maj; Guerra, Barbara; Oliván-Viguera, Aida

2017-01-01

Protein kinase CK2a, one of the two catalytic isoforms of the protein kinase CK2 has been shown to contribute to tumor development, tumor proliferation and suppression of apoptosis in various malignancies. We conducted this study to investigate CK2 expression in different subtypes of Renal Cell...... Carcinoma (RCC) and in the benign oncocytoma. qRT-PCR, immunohistochemistry and Western blot analyses revealed that CK2a expression was significantly increased at the mRNA and protein levels in clear cell RCC (ccRCC). Also the kinase activity of CK2 was significantly increased in ccRCC compared to normal...... renal cortex. Nuclear protein expression of CK2a correlated in univariate analysis with poor Progression Free Survival (HR = 8.11, p = 0.016). Functional analyses (cell proliferation assay) revealed an inhibitory effect of Caki-2 cell growth following CK2 inhibition with CX-4945. Our results suggest...

Protein kinase CK2 modulates IL-6 expression in inflammatory breast cancer

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Drygin, Denis, E-mail: ddrygin@cylenepharma.com; Ho, Caroline B.; Omori, Mayuko; Bliesath, Joshua; Proffitt, Chris; Rice, Rachel; Siddiqui-Jain, Adam; O' Brien, Sean; Padgett, Claire; Lim, John K.C.; Anderes, Kenna; Rice, William G.; Ryckman, David

2011-11-11

Highlights: Black-Right-Pointing-Pointer We examine the potential cross-talk between CK2 and IL-6. Black-Right-Pointing-Pointer Inhibition of CK2 by siRNA or CX-4945 inhibits expression of IL-6 in models of IBC. Black-Right-Pointing-Pointer Treatment of IBC patient in the clinic with CX-4945 reduces her IL-6 plasma levels. Black-Right-Pointing-Pointer We demonstrate that CK2 is a potential therapeutic target for IL-6 driven diseases. -- Abstract: Inflammatory breast cancer is driven by pro-angiogenic and pro-inflammatory cytokines. One of them Interleukin-6 (IL-6) is implicated in cancer cell proliferation and survival, and promotes angiogenesis, inflammation and metastasis. While IL-6 has been shown to be upregulated by several oncogenes, the mechanism behind this phenomenon is not well characterized. Here we demonstrate that the pleotropic Serine/Threonine kinase CK2 is implicated in the regulation of IL-6 expression in a model of inflammatory breast cancer. We used siRNAs targeted toward CK2 and a selective small molecule inhibitor of CK2, CX-4945, to inhibit the expression and thus suppress the secretion of IL-6 in in vitro as well as in vivo models. Moreover, we report that in a clinical trial, CX-4945 was able to dramatically reduce IL-6 levels in plasma of an inflammatory breast cancer patient. Our data shed a new light on the regulation of IL-6 expression and position CX-4945 and potentially other inhibitors of CK2, for the treatment of IL-6-driven cancers and possibly other diseases where IL-6 is instrumental, including rheumatoid arthritis.

Expression of casein kinase 2 during mouse embryogenesis

DEFF Research Database (Denmark)

Mestres, P; Boldyreff, B; Ebensperger, C

1994-01-01

This paper deals with the expression and distribution of casein kinase 2 (CK-2) subunits in mouse embryos at different developmental stages. Expression was investigated at the mRNA level of CK-2 alpha- and beta-subunits by in situ hybridization and distribution at the protein level by immunohisto......This paper deals with the expression and distribution of casein kinase 2 (CK-2) subunits in mouse embryos at different developmental stages. Expression was investigated at the mRNA level of CK-2 alpha- and beta-subunits by in situ hybridization and distribution at the protein level...

Differential expression of galectin-3, CK19, HBME1, and Ret oncoprotein in the diagnosis of thyroid neoplasms by fine needle aspiration biopsy

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Saleh Husain

2009-01-01

Full Text Available Background: Fine needle aspiration biopsy (FNAB is a common and excellent procedure for the evaluation of thyroid lesions that require surgical resection. At times, the FNAB diagnosis can be difficult, particularly of follicular-patterned lesions. Previous studies have shown that some immunohistochemical (IHC markers may be helpful in establishing more accurate diagnosis. In this study, our goal was to evaluate four of the recently investigated markers in differentiating benign from malignant thyroid nodules on FNABs. Materials and Methods: We performed IHC staining of galectin-3, Ret oncoprotein (Ret, HBME-1, and cytokeratin 19 (CK19, on cell block sections of thyroid FNAB cases that had corresponding surgical resections. They included 44 benign lesions (37 hyperplastic or cellular nodules, HN; and 7 follicular adenomas, FA and 27 malignant tumors (6 follicular carcinoma, FC; 19 classic papillary carcinoma, PTC; and 2 follicular variant of papillary carcinoma, FVPC. The stains were done according to the standard avidin-biotin-peroxidase method. Results: Statistical analysis showed that immunoexpression was significantly higher in the malignant group for all four markers. The sensitivity for positive expression for all benign lesions versus malignant tumors was as follows: 10/44 (22.7% versus 25/27 (92.6% for galectin-3; 14/44 (31.8% versus 23/27 (85% for Ret; 12/44 (27.3% versus 24/27 (88.8% for HBME-1; and 13/44 (29.5% versus 23/27 (85% for CK19. The sensitivity and specificity was highest for galectin-3 (92.6% and 77.3%, respectively followed by HMBE-1 (88.9% and 72.7%, respectively. When combining the markers′ expressions, the panel of galectin-3 + HBME-1 showed the highest sensitivity and specificity (90.7% and 75%, respectively, but this was, however, lower than galectin-3 alone (92.3% and 77.3%, respectively. Conclusion: We conclude that galectin-3 is the best single marker in differentiating benign from malignant thyroid lesions with

The effects of pilates exercise on lipid metabolism and inflammatory cytokines mRNA expression in female undergraduates.

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Kim, Hyo-Jin; Kim, Jiyeon; Kim, Chang-Sun

2014-09-01

The purpose of the study was to verify the effects of Pilates exercise by observing the impact of 8 weeks of Pilates exercise on lipid metabolism and inflammatory cytokine mRNA expression in female undergraduates in their 20s who had no prior experience in Pilates exercise and had not exercised in the previous 6 months. There were 18 subjects with no prior experience in Pilates exercise. The subjects were separated into the Pilates exercise group (n = 9) and the non-exercise control group (n = 9). The former performed Pilates exercise for 60-70 minutes over 8 weeks with a gradual strength increase of 9-16 in the Rating of Perceived Exercise (RPE). The body composition, creatine kinase in the bloodstream and lipid metabolism (TC, LDL-C, HDL-C, TG) were measured before and after the experiment and Real-Time PCR was used to investigate the mRNA expression of the inflammatory cytokines IL-6 and TNF-⍺. The creatine kinase (CK) in the blood had significant differences between the groups. The test group showed significant increase compared to the control group after 8 weeks of Pilates exercise (p = 0.007). Lipid analysis showed that the level of high-density lipoprotein cholesterol (HDL-C) was significantly different in the two groups (p = 0.049), with the Pilates exercise group exhibiting significantly higher levels compared to the control group. No significant differences were observed in the levels of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and triglyceride (TG). IL-6 mRNA expression did not show significant differences between the groups either. Timing and TNF-α mRNA expression showed significant effect in both the exercise and the control groups (p = 0.013) but no correlation. It was found from the study that Pilates exercise for 8 weeks affected CK expression (the muscle damage marker) and induced positive changes in the levels of high-density lipoprotein.

Absence of the BRAF mutation in HBME1+ and CK19+ atypical cell clusters in Hashimoto thyroiditis: supportive evidence against preneoplastic change.

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Nasr, Michel R; Mukhopadhyay, Sanjay; Zhang, Shengle; Katzenstein, Anna-Luise A

2009-12-01

An association between Hashimoto thyroiditis and papillary thyroid carcinoma has been postulated for decades. We undertook this study to identify potential precursors of papillary thyroid carcinoma in Hashimoto thyroiditis using a combination of morphologic, immunohistochemical, and molecular techniques. For the study, samples from 59 cases of Hashimoto thyroiditis were stained with antibodies to HBME1 and cytokeratin (CK)19. Tiny HBME1+ and CK19+ atypical cell clusters were identified and analyzed for the BRAF mutation by the colorimetric Mutector assay and allele-specific polymerase chain reaction. HBME1+ and CK19+ atypical cell clusters were identified in 12 (20%) of 59 cases. The minute size (Hashimoto thyroiditis.

Stem cell markers (cytokeratin 17 and cytokeratin 19 in scarring and nonscarring alopecia

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Dalia El Sakka

2016-01-01

Full Text Available Background: Alopecia is one of the most important hair follicle (HF disorders, which is divided into scarring (cicatricial and nonscarring (noncicatricial types. Objective: The aim of this study is to investigate the expression of stem cell (SC markers such as cytokeratin (CK 17 and CK19 in scarring and nonscarring alopecia. Materials and Methods: Thirty patients with scalp alopecia (15 with scarring alopecia and 15 without together with ten healthy volunteers were included in this study. Biopsies were taken from all participants and stained for CK17 and CK19 using immunohistochemistry. Results: There was a statistically significant difference between the nonscarring group and the control group with regard to CK17 expression in the outer layers of the HFs (P = 0.00 and CK19 staining of the inner layers of the HFs (P = 0.008. There was a statistically significant difference between the scarring and the control groups regarding CK17 expression in the outer (P = 0.00 and the inner layers (P = 0.00 of the HFs and CK19 expression in the inner layers of the HFs (P = 0.00. CK17 expression in the outer layers (P = 0.02 and the inner layers of the HFs (P = 0.00 together with CK19 expression in the inner layers of the HFs (P = 0.00 showed statistically significant differences between scarring and nonscarring alopecia groups. Conclusions: The presence of SC markers (CK17 and CK19 in the HFs was affected in both scarring and nonscarring alopecia, but the defect in scarring alopecia is more evident than that of nonscarring alopecia. The persistence of SC markers in some types of scarring alopecia could give a hope for the recovery of these lesions. Further studies are recommended to clarify the benefit from using HF SCs in the treatment of alopecia.

Stem Cell Markers (Cytokeratin 17 and Cytokeratin 19) in Scarring and Nonscarring Alopecia

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El Sakka, Dalia; Gaber, Mohamed Abdel Wahed; Abdou, Asmaa Gaber; Wahed, Moshira Abdel; Saleh, Ahmed Abdel-Wahab; Shehata, Walla

2016-01-01

Background: Alopecia is one of the most important hair follicle (HF) disorders, which is divided into scarring (cicatricial) and nonscarring (noncicatricial) types. Objective: The aim of this study is to investigate the expression of stem cell (SC) markers such as cytokeratin (CK) 17 and CK19 in scarring and nonscarring alopecia. Materials and Methods: Thirty patients with scalp alopecia (15 with scarring alopecia and 15 without) together with ten healthy volunteers were included in this study. Biopsies were taken from all participants and stained for CK17 and CK19 using immunohistochemistry. Results: There was a statistically significant difference between the nonscarring group and the control group with regard to CK17 expression in the outer layers of the HFs (P = 0.00) and CK19 staining of the inner layers of the HFs (P = 0.008). There was a statistically significant difference between the scarring and the control groups regarding CK17 expression in the outer (P = 0.00) and the inner layers (P = 0.00) of the HFs and CK19 expression in the inner layers of the HFs (P = 0.00). CK17 expression in the outer layers (P = 0.02) and the inner layers of the HFs (P = 0.00) together with CK19 expression in the inner layers of the HFs (P = 0.00) showed statistically significant differences between scarring and nonscarring alopecia groups. Conclusions: The presence of SC markers (CK17 and CK19) in the HFs was affected in both scarring and nonscarring alopecia, but the defect in scarring alopecia is more evident than that of nonscarring alopecia. The persistence of SC markers in some types of scarring alopecia could give a hope for the recovery of these lesions. Further studies are recommended to clarify the benefit from using HF SCs in the treatment of alopecia. PMID:27761086

Arsenite induced oxidative damage in mouse liver is associated with increased cytokeratin 18 expression

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Gonsebatt, M.E. [UNAM, Ciudad Universitaria, Dept. Medicina Genomica y Toxicologia Ambiental, Instituto de Investigaciones Biomedicas, Mexico (Mexico); Razo, L.M. del; Sanchez-Pena, L.C. [Seccion de Toxicologia, CINVESTAV, Mexico (Mexico); Cerbon, M.A. [Facultad de Quimica, UNAM, Departamento de Biologia, Mexico (Mexico); Zuniga, O.; Ramirez, P. [Facultad de Estudios Superiores Cuautitlan, UNAM, Laboratorio de Toxicologia Celular, Coordinacion General de Estudios de Posgrado e Investigacion, Cuautitlan Izcalli, Estado de Mexico (Mexico)

2007-09-15

Cytokeratins (CK) constitute a family of cytoskeletal intermediate filament proteins that are typically expressed in epithelial cells. An abnormal structure and function are effects that are clearly related to liver diseases as non-alcoholic steatohepatitis, cirrhosis and hepatocellular carcinoma. We have previously observed that sodium arsenite (SA) induced the synthesis of CK18 protein and promotes a dose-related disruption of cytoplasmic CK18 filaments in a human hepatic cell line. Both abnormal gene expression and disturbance of structural organization are toxic effects that are likely to cause liver disease by interfering with normal hepatocyte function. To investigate if a disruption in the CK18 expression pattern is associated with arsenite liver damage, we investigated CK18 mRNA and protein levels in liver slices treated with low levels of SA. Organotypic cultures were incubated with 0.01, 1 and 10 {mu}M of SA in the absence and presence of N-acetyl cysteine (NAC). Cell viability and inorganic arsenic metabolism were determined. Increased expression of CK18 was observed after exposure to SA. The addition of NAC impeded the oxidative effects of SA exposure, decreasing the production of thiobarbituric acid-reactive substances and significantly diminishing the up regulation of CK18 mRNA and protein. Liver arsenic levels correlated with increased levels of mRNA. Mice treated with intragastric single doses of 2.5 and 5 mg/kg of SA showed an increased expression of CK18. Results suggest that CK18 expression may be a sensible early biomarker of oxidative stress and damage induced by arsenite in vitro and in vivo. Then, during SA exposure, altered CK expression may compromise liver function. (orig.)

Increased IL-10 mRNA and IL-23 mRNA expression in multiple sclerosis: interferon-beta treatment increases IL-10 mRNA expression while reducing IL-23 mRNA expression

DEFF Research Database (Denmark)

Krakauer, M.; Sorensen, P.; Khademi, M.

2008-01-01

volunteers served to confirm initial findings. mRNA was analyzed by real-time reverse transcriptase polymerase chain reaction (PCR). RESULTS: We found elevated expression of interleukin (IL)-23 and IL-10 in untreated MS patients. IFN-beta therapy increased IL-10 and decreased IL-23 expression independently...... of the regulatory cytokine IL-10. The elevated IL-23 mRNA levels in MS patients are noteworthy in view of the newly discovered IL-23-driven Th17 T-cell subset, which is crucial in animal models of MS. Since IFN-beta therapy resulted in decreased IL-23 mRNA levels, the Th17 axis could be another target of IFN...

Purification and characterization of recombinant protein kinase CK2 from Zea mays expressed in Escherichia coli

DEFF Research Database (Denmark)

Riera, Marta; Pages, Montserrat; Issinger, Olaf Georg

2003-01-01

Recombinant protein kinase subunits rmCK2alpha-1 and rmCK2beta-1 from Zea mays were expressed separately in Escherichia coli and assembled to a fully active tetrameric holoenzyme complex in vitro. The obtained maize holoenzyme was purified to homogeneity, biochemically characterized, and compared...... to CK2 from human. Kinetic measurements of the recombinant maize holoenzyme (rmCK2) revealed k(cat) values for ATP and GTP of 4 and 2s(-1), respectively; whereas the recombinant maize catalytic subunit showed almost equal values for ATP and GTP, i.e., ca. 0.8s(-1). A comparison of the k(cat)/K(m) ratio...

Creatine kinase and alpha-actin mRNA levels decrease in diabetic rat hearts

International Nuclear Information System (INIS)

Popovich, B.; Barrieux, A.; Dillmann, W.H.

1987-01-01

Diabetic cardiomyopathy is associated with cardiac atrophy and isoenzyme redistribution. To determine if tissue specific changes occur in mRNAs coding for α-actin and creatine kinase (CK), they performed RNA blot analysis. Total ventricular RNA from control (C) and 4 wk old diabetic (D) rats were hybridized with 32 P cDNA probes for α-actin and CK. A tissue independent cDNA probe, CHOA was also used. Signal intensity was quantified by photodensitometry. D CK mRNA was 47 +/- 16% lower in D vs C. Insulin increases CK mRNA by 20% at 1.5 hs, and completely reverses the deficit after 4 wks. D α-actin mRNA is 66 +/- 18% lower in D vs C. Insulin normalized α-actin mRNA by 5 hs. CHOA mRNA is unchanged in D vs C, but D + insulin CHOA mRNA is 30 +/- 2% lower than C. In rats with diabetic cardiomyopathy, muscle specific CK and α-actin mRNAs are decreased. Insulin treatment reverses these changes

Protein Kinase CK2 Content in GL261 Mouse Glioblastoma.

Science.gov (United States)

Ferrer-Font, Laura; Alcaraz, Estefania; Plana, Maria; Candiota, Ana Paula; Itarte, Emilio; Arús, Carles

2016-07-01

Glioblastoma (GBM) is the most prevalent and aggressive human glial tumour with a median survival of 14-15 months. Temozolomide (TMZ) is the standard chemotherapeutic choice for GBM treatment. Unfortunately, chemoresistence always ensues with concomitant tumour regrowth. Protein kinase CK2 (CK2) contributes to tumour development, proliferation, and suppression of apoptosis in cancer and it is overexpressed in human GBM. Targeting CK2 in GBM treatment may benefit patients. With this translational perspective in mind, we have studied the CK2 expression level by Western blot analysis in a preclinical model of GBM: GL261 cells growing orthotopically in C57BL/6 mice. The expression level of the CK2 catalytic subunit (CK2α) was higher in tumour (about 4-fold) and in contralateral brain parenchyma (more than 2-fold) than in normal brain parenchyma (p < 0.05). In contrast, no significant changes were found in CK2 regulatory subunit (CK2β) expression, suggesting an increased unbalance of CK2α/CK2β in GL261 tumours with respect to normal brain parenchyma, in agreement with a differential role of these two subunits in tumours.

CK8 correlates with malignancy in leukoplakia and carcinomas of the head and neck

International Nuclear Information System (INIS)

Gires, Olivier; Mack, Brigitte; Rauch, Jens; Matthias, Christoph

2006-01-01

Screening of head and neck carcinoma patients with the proteomics-based AMIDA technology yielded a set of tumour-associated antigens, including the intermediate filament protein cytokeratin 8 (CK8). The expression pattern and specificity of CK8 was compared with those of the established markers pan-cytokeratins and CK13, and with that of the proliferation marker Ki67. Expression of CK8 correlated positively with malignancies of the head and neck areas. CK8 was not expressed in healthy epithelium, except for some rare cases of cells of the basal layer and laryngeal tissue. In contrast, the vast majority of head and neck squamous cell carcinomas and metastases strongly expressed CK8. Interestingly, CK8 de novo expression correlated with dysplastic areas of oral leukoplakic lesions, while hyperplastic leukoplakia remained CK8-negative but strongly panCK and CK13 positive. Thus, CK8 is an attractive marker molecule for a differentiated diagnosis of leukoplakia and head and neck carcinomas, which possesses notedly improved specificity as compared with panCK and CK13

Predictive value of BRCA1/2 mRNA expression for response to neoadjuvant chemotherapy in BRCA-negative breast cancers.

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Xu, Ye; Ouyang, Tao; Li, Jinfeng; Wang, Tianfeng; Fan, Zhaoqing; Fan, Tie; Lin, Benyao; Xie, Yuntao

2018-01-01

It is well known that BRCA1 and BRCA2 play a central role in DNA repair, but the relationship between BRCA1 and BRCA2 mRNA expression and response to neoadjuvant chemotherapy in sporadic breast cancer patients has not been well established. Here, we investigate the association between BRCA1 or BRCA2 mRNA expression levels and pathological response in 674 BRCA1/2 mutation-negative breast cancer patients who received neoadjuvant chemotherapy. BRCA1 and BRCA2 mRNA expression were assessed using quantitative real-time polymerase chain reaction in core biopsy breast cancer tissue obtained prior to the initiation of neoadjuvant chemotherapy. A total 129 patients (19.1%) achieved pathological complete response (pCR) after neoadjuvant chemotherapy. Among patients treated with anthracycline-based chemotherapy (n = 531), BRCA1 mRNA low expression patients had a significantly higher pCR rate than intermediate or high BRCA1 mRNA expression groups (24.6% vs 16.8% or 14.0%, P = .031) and retained borderline significance (OR = 1.54, 95% CI = 0.93-2.56, P = .094) in multivariate analysis. Among the 129 patients who received a taxane-based regimen, pCR rate showed no differences in BRCA1 low, intermediate, and high mRNA level subgroups (19.6%, 26.8% and 21.4%, respectively; P = .71). BRCA2 mRNA level was not associated with pCR rate in the anthracyline-based treated subgroup (P = .60) or the taxane-based regimen subgroup (P = .82). Taken together, our findings suggested that BRCA1 mRNA expression could be used as a predictive marker in BRCA1/2 mutation-negative breast cancer patients who received neoadjuvant anthracycline-based treatment. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

Expression of calmodulin mRNA in rat olfactory neuroepithelium.

Science.gov (United States)

Biffo, S; Goren, T; Khew-Goodall, Y S; Miara, J; Margolis, F L

1991-04-01

A calmodulin (CaM) cDNA was isolated by differential hybridization screening of a lambda gt10 library prepared from rat olfactory mucosa. This cDNA fragment, containing most of the open reading frame of the rat CaMI gene, was subcloned and used to characterize steady-state expression of CaM mRNA in rat olfactory neuroepithelium and bulb. Within the bulb mitral cells are the primary neuronal population expressing CaM mRNA. The major CaM mRNA expressed in the olfactory mucosa is 1.7 kb with smaller contributions from mRNAs of 4.0 and 1.4 kb. CaM mRNA was primarily associated with the olfactory neurons and, despite the cellular complexity of the tissue and the known involvement of CaM in diverse cellular processes, was only minimally evident in sustentacular cells, gland cells or respiratory epithelium. Following bulbectomy CaM mRNA declines in the olfactory neuroepithelium as does olfactory marker protein (OMP) mRNA. In contrast to the latter, CaM mRNA makes a partial recovery by one month after surgery. These results, coupled with those from in situ hybridization, indicate that CaM mRNA is expressed in both mature and immature olfactory neurons. The program regulating CaM gene expression in olfactory neurons is distinct from those controlling expression of B50/GAP43 in immature, or OMP in mature, neurons respectively.

Protein kinase CK2 structure-function relationship

DEFF Research Database (Denmark)

Boldyreff, B; Meggio, F; Pinna, L A

1994-01-01

Protein kinase CK2 subunits alpha and beta were expressed either separately or together in a bacterial expression system (pT7-7/BL21(DE3)) and purified to homogeneity. After mixing the subunits, a CK2 holoenzyme (alpha 2 beta 2) was spontaneously reconstituted, which displays identical features...... subunit have been prepared and assayed for their ability to assemble with the catalytic alpha subunit to give a fully competent CK2 holoenzyme. The beta subunit contains an acidic stretch (amino acid 55-64), which is obviously responsible for a negative control of enzyme activity since mutations...

T-lymphocyte cytokine mRNA expression in cystic echinococcosis.

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Fauser, S; Kern, P

1997-04-01

In the present study we investigated cytokine mRNA expression by peripheral blood mononuclear cells (PBMC) from patients with cystic echinococcosis (CE) after stimulation with different antigens. By using reverse transcriptase polymerase chain reaction (RT-PCR) we could demonstrate that restimulation with crude Echinococcus granulosus antigen (Eg-Ag) induced or enhanced Th2 cytokine mRNA expression, especially IL-5 (by using antigen from sheep cyst fluid) in 23 out of 26 investigated CE patients and IL-10 (by using antigen from camel cyst fluid) in 10 out of 10 investigated CE patients. In contrast, IL-5 mRNA expression was absent in PBMC of healthy controls after Eg-Ag stimulation. To determine the specificity of this reaction we stimulated PBMC from 11 CE patients with crude Echinococcus multilocularis antigen (Em-Ag) and PBMC from 8 CE patients with Toxocara canis antigen (Tc-Ag). We found that the PBMC of patients showed a similar mRNA cytokine pattern on stimulation with Em-Ag when compared with Eg-Ag stimulation. The cytokine mRNA pattern on stimulation with Tc-Ag, however, resembled the cytokine mRNA pattern of unstimulated PBMC. Furthermore, the stimulation of PBMC with crude Mycobacterium tuberculosis antigen (H37Ra) and purified protein derivative (PPD) of M. tuberculosis revealed distinct IL-5 mRNA expression in all investigated CE patients, whereas in healthy controls IL-5 mRNA expression was very weak or totally absent. Thus, our results indicate an induction of Th2 cytokine mRNA expression in CE patients, which is frequently observed in parasite infections. Interestingly, this response persists after stimulation with tuberculosis antigens, which normally induce Th1 response.

Ddx19 links mRNA nuclear export with progression of transcription and replication and suppresses genomic instability upon DNA damage in proliferating cells.

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Hodroj, Dana; Serhal, Kamar; Maiorano, Domenico

2017-09-03

The DEAD-box Helicase 19 (Ddx19) gene codes for an RNA helicase involved in both mRNA (mRNA) export from the nucleus into the cytoplasm and in mRNA translation. In unperturbed cells, Ddx19 localizes in the cytoplasm and at the cytoplasmic face of the nuclear pore. Here we review recent findings related to an additional Ddx19 function in the nucleus in resolving RNA:DNA hybrids (R-loops) generated during collision between transcription and replication, and upon DNA damage. Activation of a DNA damage response pathway dependent upon the ATR kinase, a major regulator of replication fork progression, stimulates translocation of the Ddx19 protein from the cytoplasm into the nucleus. Only nuclear Ddx19 is competent to resolve R-loops, and down regulation of Ddx19 expression induces DNA double strand breaks only in proliferating cells. Overall these observations put forward Ddx19 as an important novel mediator of the crosstalk between transcription and replication.

Cytokeratin 8/18 expression indicates a poor prognosis in squamous cell carcinomas of the oral cavity

International Nuclear Information System (INIS)

Fillies, Thomas; Werkmeister, Richard; Packeisen, Jens; Brandt, Burkhard; Morin, Philippe; Weingart, Dieter; Joos, Ulrich; Buerger, Horst

2006-01-01

Intermediary filaments are involved in cell motility and cancer progression. In a variety of organs, the expression of distinct intermediary filaments are associated with patient prognosis. In this study, we seeked to define the prognostic potential of cytokeratin and vimentin expression patterns in squamous cell carcinomas (SCC's) of the oral cavity. 308 patients with histologically proven and surgically treated squamous cell carcinomas of the oral cavity were investigated for the immunohistochemical expression of a variety of intermediary filaments including high- and low-molecular weight cytokeratins (Ck's), such as Ck 5/6, Ck 8/18, Ck 1, CK 10, Ck 14, Ck 19 and vimentin, using the tissue microarray technique. Correlations between clinical features and the expression of Cytokeratins and vimentin were evaluated statistically by Kaplan-Meier curves and multivariate Cox regression analysis. The expression of Ck 8/18 and Ck 19 were overall significantly correlated with a poor clinical prognosis (Ck 8/18 p = 0.04; Ck19 p < 0.01). These findings could also be reproduced for Ck 8/18 in primary nodal-negative SCC's and held true in multivariate-analysis. No significant correlation with patient prognosis could be found for the expression of the other cytokeratins and for vimentin. The expression of Ck 8/18 in SCC's of the oral cavity is an independent prognostic marker and indicates a decreased overall and progression free survival. These results provide an extended knowledge about the role of intermediary filament expression patterns in SCC's

The mRNA expression of SETD2 in human breast cancer: correlation with clinico-pathological parameters

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Al Sarakbi, W; Sasi, W [St George' s University of London, Blackshaw Road, Tooting, London, SW17 OQT (United Kingdom); Jiang, WG [University Department of Surgery, Wales College of Medicine, Cardiff University, CF14 4XN (United Kingdom); Roberts, T; Newbold, RF [Institute of Cancer Genetics and Pharmacogenomics, Brunel University, Uxbridge, Middlesex, UB8 3PH (United Kingdom); Mokbel, K [St George' s University of London, Blackshaw Road, Tooting, London, SW17 OQT (United Kingdom); Institute of Cancer Genetics and Pharmacogenomics, Brunel University, Uxbridge, Middlesex, UB8 3PH (United Kingdom)

2009-08-21

SET domain containing protein 2 (SETD2) is a histone methyltransferase that is involved in transcriptional elongation. There is evidence that SETD2 interacts with p53 and selectively regulates its downstream genes. Therefore, it could be implicated in the process of carcinogenesis. Furthermore, this gene is located on the short arm of chromosome 3p and we previously demonstrated that the 3p21.31 region of chromosome 3 was associated with permanent growth arrest of breast cancer cells. This region includes closely related genes namely: MYL3, CCDC12, KIF9, KLHL18 and SETD2. Based on the biological function of these genes, SETD2 is the most likely gene to play a tumour suppressor role and explain our previous findings. Our objective was to determine, using quantitative PCR, whether the mRNA expression levels of SETD2 were consistent with a tumour suppressive function in breast cancer. This is the first study in the literature to examine the direct relationship between SETD2 and breast cancer. A total of 153 samples were analysed. The levels of transcription of SETD2 were determined using quantitative PCR and normalized against (CK19). Transcript levels within breast cancer specimens were compared to normal background tissues and analyzed against conventional pathological parameters and clinical outcome over a 10 year follow-up period. The levels of SETD2 mRNA were significantly lower in malignant samples (p = 0.0345) and decreased with increasing tumour stage. SETD2 expression levels were significantly lower in samples from patients who developed metastasis, local recurrence, or died of breast cancer when compared to those who were disease free for > 10 years (p = 0.041). This study demonstrates a compelling trend for SETD2 transcription levels to be lower in cancerous tissues and in patients who developed progressive disease. These findings are consistent with a possible tumour suppressor function of this gene in breast cancer.

Biochemical characterization of CK2alpha and alpha' paralogues and their derived holoenzymes: evidence for the existence of a heterotrimeric CK2alpha'-holoenzyme forming trimeric complexes

DEFF Research Database (Denmark)

Olsen, Birgitte; Rasmussen, Tine; Niefind, Karsten

2008-01-01

Altogether 2 holoenzymes and 4 catalytic CK2 constructs were expressed and characterized i.e. CK2alpha (2) (1-335) beta(2); CK2alpha'-derived holoenzyme; CK2alpha(1-335); MBP-CK2alpha'; His-tagged CK2alpha and His-tagged CK2alpha'. The two His-tagged catalytic subunits were expressed in insect...... cells, all others in Escherichia coli. IC(50) studies involving the established CK2 inhibitors DMAT, TBBt, TBBz, apigenin and emodin were carried out and the K(i) values calculated. Although the differences in the K(i) values found were modest, there was a general tendency showing that the CK2...... holoenzymes were more sensitive towards the inhibitors than the free catalytic subunits. Thermal inactivation experiments involving the individual catalytic subunits showed an almost complete loss of activity after only 2 min at 45 degrees C. In the case of the two holoenzymes, the CK2alpha...

Differential expression of CK20, β-catenin, and MUC2/5AC/6 in Lynch syndrome and familial colorectal cancer type X

DEFF Research Database (Denmark)

Haraldsson, Stefan; Klarskov, Louise; Nilbert, Mef

2017-01-01

BACKGROUND: Hereditary non-polyposis colorectal cancer comprises Lynch syndrome and familial colorectal cancer type X (FCCTX). Differences in genetics, demographics and histopathology have been extensively studied. The purpose of this study is to characterize their immunoprofile of markers other...... than MMR proteins. METHODS: We compared the expression patterns of cytokeratins (CK7 and CK20), mucins (MUC2/5 AC/6), CDX2 and β-catenin in Lynch syndrome and FCCTX. RESULTS: Differences were identified for CK20 and nuclear β-catenin, which were significantly more often expressed in FCCTX than in Lynch...... syndrome (p Lynch syndrome tumors compared with FCCTX tumors (p = 0.001,

Level of CYP4G19 Expression Is Associated with Pyrethroid Resistance in Blattella germanica

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Guang-zhou Guo

2010-01-01

Full Text Available German cockroaches have become a large problem in the Shenzhen area because of their pesticide resistance, especially to pyrethroid. A pyrethroid called “Jia Chong Qing” to prevent pests for a long time were found to be resistant to “Jia Chong Qing” with resistance index of 3.88 measured using RT-PCR and immunohistochemistry analysis showed that both CYP4G19 mRNA and CYP4G19 protein expression levels in the wild strain were substantially higher than that of a sensitive strain. dsRNA segments derived from the target gene CYP4G19 were prepared using in vitro transcription and were microinjected into abdomens of the wild strain. Two to eight days after injection, the result showed that CYP4G19 mRNA expressions were significantly reduced in the groups injected with dsRNAs.

Asymmetric expression of protein kinase CK2 subunits in human kidney tumors

DEFF Research Database (Denmark)

Stalter, G; Siemer, S; Becht, E

1994-01-01

of protein kinase CK2 alpha in tumors/normal tissue (T/N) was 1.58 and that of the protein kinase CK2 beta (T/N) was 2.65. The data suggest that the generally described increase in protein kinase CK2 activity in tumor cells may to some extent result from a deregulation in subunit biosynthesis or degradation...

Fluorescence in situ hybridization techniques (FISH) to detect changes in CYP19a gene expression of Japanese medaka (Oryzias latipes)

International Nuclear Information System (INIS)

Park, June-Woo; Tompsett, Amber; Zhang, Xiaowei; Newsted, John L.; Jones, Paul D.; Au, Doris; Kong, Richard; Wu, Rudolf S.S.; Giesy, John P.; Hecker, Markus

2008-01-01

The aim of this study was to develop a sensitive in situ hybridization methodology using fluorescence-labeled riboprobes (FISH) that allows for the evaluation of gene expression profiles simultaneously in multiple target tissues of whole fish sections of Japanese medaka (Oryzias latipes). To date FISH methods have been limited in their application due to autofluorescence of tissues, fixatives or other components of the hybridization procedure. An optimized FISH method, based on confocal fluorescence microscopy was developed to reduce the autofluorescence signal. Because of its tissue- and gender-specific expression and relevance in studies of endocrine disruption, gonadal aromatase (CYP19a) was used as a model gene. The in situ hybridization (ISH) system was validated in a test exposure with the aromatase inhibitor fadrozole. The optimized FISH method revealed tissue-specific expression of the CYP19a gene. Furthermore, the assay could differentiate the abundance of CYP19a mRNA among cell types. Expression of CYP19a was primarily associated with early stage oocytes, and expression gradually decreased with increasing maturation. No expression of CYP19a mRNA was observed in other tissues such as brain, liver, or testes. Fadrozole (100 μg/L) caused up-regulation of CYP19a expression, a trend that was confirmed by RT-PCR analysis on excised tissues. In a combination approach with gonad histology, it could be shown that the increase in CYP19a expression as measured by RT-PCR on a whole tissue basis was due to a combination of both increases in numbers of CYP19a-containing cells and an increase in the amount of CYP19a mRNA present in the cells

Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?

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Edgar Corneille Ontsouka

Full Text Available BACKGROUND: Mammary cell cultures are convenient tools for in vitro studies of mammary gland biology. However, the heterogeneity of mammary cell types, e.g., glandular milk secretory epithelial or myoepithelial cells, often complicates the interpretation of cell-based data. The present study was undertaken to determine the relevance of bovine primary mammary epithelial cells isolated from American Holstein (bMEC US or Swiss Holstein-Friesian (bMEC CH cows, and of primary bovine mammary alveolar epithelial cells stably transfected with simian virus-40 (SV-40 large T-antigen (MAC-T for in vitro analyses. This was evaluated by testing their expression pattern of cytokeratin (CK 7, 18, 19, vimentin, and α-smooth muscle actin (α-SMA. RESULTS: The expression of the listed markers was assessed using real-time quantitative PCR, flow cytometry and immunofluorescence microscopy. Characteristic markers of the mesenchymal (vimentin, myoepithelial (α-SMA and glandular secretory cells (CKs showed differential expression among the studied cell cultures, partly depending on the analytical method used. The relative mRNA expression of vimentin, CK7 and CK19, respectively, was lower (P < 0.05 in immortalized than in primary mammary cell cultures. The stain index (based on flow cytometry of CK7 and CK19 protein was lower (P < 0.05 in MAC-T than in bMECs, while the expression of α-SMA and CK18 showed an inverse pattern. Immunofluorescence microscopy analysis mostly confirmed the mRNA data, while partly disagreed with flow cytometry data (e.g., vimentin level in MAC-T. The differential expression of CK7 and CK19 allowed discriminating between immortal and primary mammary cultures. CONCLUSIONS: The expression of the selected widely used cell type markers in primary and immortalized MEC cells did not allow a clear preference between these two cell models for in vitro analyses studying aspects of milk composition. All tested cell models exhibited to a variable

Tissue-specific mRNA expression profiling in grape berry tissues

Science.gov (United States)

Grimplet, Jerome; Deluc, Laurent G; Tillett, Richard L; Wheatley, Matthew D; Schlauch, Karen A; Cramer, Grant R; Cushman, John C

2007-01-01

Background Berries of grape (Vitis vinifera) contain three major tissue types (skin, pulp and seed) all of which contribute to the aroma, color, and flavor characters of wine. The pericarp, which is composed of the exocarp (skin) and mesocarp (pulp), not only functions to protect and feed the developing seed, but also to assist in the dispersal of the mature seed by avian and mammalian vectors. The skin provides volatile and nonvolatile aroma and color compounds, the pulp contributes organic acids and sugars, and the seeds provide condensed tannins, all of which are important to the formation of organoleptic characteristics of wine. In order to understand the transcriptional network responsible for controlling tissue-specific mRNA expression patterns, mRNA expression profiling was conducted on each tissue of mature berries of V. vinifera Cabernet Sauvignon using the Affymetrix GeneChip® Vitis oligonucleotide microarray ver. 1.0. In order to monitor the influence of water-deficit stress on tissue-specific expression patterns, mRNA expression profiles were also compared from mature berries harvested from vines subjected to well-watered or water-deficit conditions. Results Overall, berry tissues were found to express approximately 76% of genes represented on the Vitis microarray. Approximately 60% of these genes exhibited significant differential expression in one or more of the three major tissue types with more than 28% of genes showing pronounced (2-fold or greater) differences in mRNA expression. The largest difference in tissue-specific expression was observed between the seed and pulp/skin. Exocarp tissue, which is involved in pathogen defense and pigment production, showed higher mRNA abundance relative to other berry tissues for genes involved with flavonoid biosynthesis, pathogen resistance, and cell wall modification. Mesocarp tissue, which is considered a nutritive tissue, exhibited a higher mRNA abundance of genes involved in cell wall function and

Tissue-specific mRNA expression profiling in grape berry tissues

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Cramer Grant R

2007-06-01

Full Text Available Abstract Background Berries of grape (Vitis vinifera contain three major tissue types (skin, pulp and seed all of which contribute to the aroma, color, and flavor characters of wine. The pericarp, which is composed of the exocarp (skin and mesocarp (pulp, not only functions to protect and feed the developing seed, but also to assist in the dispersal of the mature seed by avian and mammalian vectors. The skin provides volatile and nonvolatile aroma and color compounds, the pulp contributes organic acids and sugars, and the seeds provide condensed tannins, all of which are important to the formation of organoleptic characteristics of wine. In order to understand the transcriptional network responsible for controlling tissue-specific mRNA expression patterns, mRNA expression profiling was conducted on each tissue of mature berries of V. vinifera Cabernet Sauvignon using the Affymetrix GeneChip® Vitis oligonucleotide microarray ver. 1.0. In order to monitor the influence of water-deficit stress on tissue-specific expression patterns, mRNA expression profiles were also compared from mature berries harvested from vines subjected to well-watered or water-deficit conditions. Results Overall, berry tissues were found to express approximately 76% of genes represented on the Vitis microarray. Approximately 60% of these genes exhibited significant differential expression in one or more of the three major tissue types with more than 28% of genes showing pronounced (2-fold or greater differences in mRNA expression. The largest difference in tissue-specific expression was observed between the seed and pulp/skin. Exocarp tissue, which is involved in pathogen defense and pigment production, showed higher mRNA abundance relative to other berry tissues for genes involved with flavonoid biosynthesis, pathogen resistance, and cell wall modification. Mesocarp tissue, which is considered a nutritive tissue, exhibited a higher mRNA abundance of genes involved in cell

Effects of Exposure to Two Fragrances on the Gene Expression of Ckm and Ckmt2 and Total CK Activity in the Hearts of Wistar Rats

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Gbenga Anthony Adefolaju

2016-01-01

Full Text Available Background Some of the most commercially used compounds in fragrances have been associated with various adverse effects in various experimental in vivo and in vitro models and are still being used promiscuously in perfumes and as additives in other household products. Objectives This study sought to determine the effects of exposing wistar rats to two locally made Nigerian perfumes on some cardiac performance enzyme and genes. Materials and Methods In this experimental study, 18 animals were allocated into three groups (A, B and C of six each. Groups B and C animals were exposed (by inhalation to the first and second perfumes (designated F1 and F2 respectively for 77 days, while animals in group A were unexposed control. The rats were sacrificed at the end of the exposure period after which heart tissue was excised for creatine kinase enzyme assay and formalin fixed, paraffin embedded heart tissues were processed for RNA extraction and analyzed by quantitative real time polymerase chain reaction for the mRNA expression of creatine kinase genes Ckm and Ckmt2. Results The results showed that animals in both exposure groups demonstrated significantly (P < 0.05 increased expression of striated muscle associated creatine kinase and sarcomeric mitochondria Ck genes as well as the increased release of the cardiomyocyte enzyme CK in the hearts of Wistar rats. Conclusions These results suggest that exposure to these two locally made fragrances contributes to cardiomyocyte stress.

MUC19 expression in human ocular surface and lacrimal gland and its alteration in Sjögren syndrome patients.

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Yu, D F; Chen, Y; Han, J M; Zhang, H; Chen, X P; Zou, W J; Liang, L Y; Xu, C C; Liu, Z G

2008-02-01

This study investigated the expression of MUC19, a newly discovered gel-forming mucin gene, in normal human lacrimal functional unit components and its alteration in Sjögren syndrome patients. Real-time PCR and immunohistochemistry were performed to determine the expression of MUC19 and MUC5AC in human cornea, conjunctiva, and lacrimal gland tissues. Conjunctival impression cytology specimens were collected from normal control subjects and Sjögren syndrome patients for Real-time PCR, PAS staining, and immunohistochemistry assays. In addition, conjunctiva biopsy specimens from both groups were examined for the expression differences of MUC19 and MUC5AC at both mRNA and protein level. The MUC19 mRNA was found to be present in cornea, conjunctiva and lacrimal gland tissues. The immunohistochemical staining of mucins showed that MUC19 was expressed in epithelial cells from corneal, conjunctival, and lacrimal gland tissues. In contrast, MUC5AC mRNA was only present in conjunctiva and lacrimal gland tissues, but not in cornea. Immunostaining demonstrates the co-staining of MUC19 and MUC5AC in conjunctival goblet cells. Consistent with the significant decrease of mucous secretion, both MUC19 and MUC5AC were decreased in conjunctiva of Sjögren syndrome patients compared to normal subjects. Considering the contribution of gel-forming mucins to the homeostasis of the ocular surface, the decreased expression of MUC19 and MUC5AC in Sjögren syndrome patients suggested that these mucins may be involved in the disruption of the ocular surface homeostasis in this disease.

Expression of galectin-9 mRNA in obese children with polymorphism of the lactase gene

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A.E. Abaturov

2018-02-01

.73 ± 52.60 RU DmRNA Gal-9/mRNA actin, which exceeded the expression level of Gal-9 mRNA in children without obesity and lactose malabsorption (313.34 ± 19.70 RU DmRNA Gal-9 / mRNA actin, p < 0.01. After the use of exogenous lactase preparations in children with genotype C/C 13910 and lactose malabsorption for a month, the level of expression of Gal-9 mRNA approximated to the level of 246.21 ± 15.70 RU DmRNA Gal-9/mRNA actin, whereas when used only the low-lactose diet the level increased insignificantly to 58.72 ± 21.10 VD DmRNA Gal-9/mRNA actin, p < 0.01. It is also interesting that in children with genotype C/C 13910 without lactose malabsorption against the background of a low-lactose diet, the level of expression of Gal-9 mRNA decreased to 388.38 ± 20.40 RU DmRNA Gal-9/mRNA actin, approaching the level of healthy children without obesity. Conclusions. In children with genotype C/C 13910 the level of expression of Gal-9 mRNA depends on the lactose malabsorption, which requires replacement therapy that combines a low-lactose diet and the use of exogenous lactase preparations. In the absence of lactose malabsorption, using only the low-lactose diet in children with genotype C/C 13910 approximates the level of expression of Gal-9 mRNA to the level of healthy children without obesity.

Whole Blood mRNA Expression-Based Prognosis of Metastatic Renal Cell Carcinoma.

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Giridhar, Karthik V; Sosa, Carlos P; Hillman, David W; Sanhueza, Cristobal; Dalpiaz, Candace L; Costello, Brian A; Quevedo, Fernando J; Pitot, Henry C; Dronca, Roxana S; Ertz, Donna; Cheville, John C; Donkena, Krishna Vanaja; Kohli, Manish

2017-11-03

The Memorial Sloan Kettering Cancer Center (MSKCC) prognostic score is based on clinical parameters. We analyzed whole blood mRNA expression in metastatic clear cell renal cell carcinoma (mCCRCC) patients and compared it to the MSKCC score for predicting overall survival. In a discovery set of 19 patients with mRCC, we performed whole transcriptome RNA sequencing and selected eighteen candidate genes for further evaluation based on associations with overall survival and statistical significance. In an independent validation of set of 47 patients with mCCRCC, transcript expression of the 18 candidate genes were quantified using a customized NanoString probeset. Cox regression multivariate analysis confirmed that two of the candidate genes were significantly associated with overall survival. Higher expression of BAG1 [hazard ratio (HR) of 0.14, p < 0.0001, 95% confidence interval (CI) 0.04-0.36] and NOP56 (HR 0.13, p < 0.0001, 95% CI 0.05-0.34) were associated with better prognosis. A prognostic model incorporating expression of BAG1 and NOP56 into the MSKCC score improved prognostication significantly over a model using the MSKCC prognostic score only ( p < 0.0001). Prognostic value of using whole blood mRNA gene profiling in mCCRCC is feasible and should be prospectively confirmed in larger studies.

Modulation of fibroblast growth factor 19 expression by bile acids, meal replacement and energy drinks, milk, and coffee.

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Styer, Amanda M; Roesch, Stephen L; Argyropoulos, George

2014-01-01

The enterohepatic pathway involving the fibroblast growth factor 19 (FGF19) and bile acids (BA) has been linked with the etiology and remission of type 2 diabetes (T2D) following Roux-en-Y gastric bypass (RYGB) surgery. Specifically, diabetic patients had lower FGF19 circulating levels but postoperative FGF19 and BA levels were higher in diabetic patients that experience remission of T2D, as compared to non-diabetic patients and diabetic patients that do not experience remission. It has been proposed that this may be due to the direct flow of digestate-free bile acids into the ileum benefiting mostly T2D patients without severe diabetes. We used a human colorectal cell line (LS174T) that endogenously expresses FGF19, real time PCR, and Elisas for precise quantitation of FGF19 mRNA and secreted protein levels. We report here that BA and fractions of BA stimulated FGF19 in vitro but this effect was partially blocked when BA were pre-incubated with a lipoprotein mix which emulates digested food. In addition, we show that FGF19 mRNA was stimulated by meal replacement drinks (Ensure, Glucerna, SlimFast), non-fat milk, and coffee which has been linked with reduced risk for developing diabetes. Pure caffeine and the 5-hour Energy drink, on the other hand, decreased FGF19 mRNA. In summary, FGF19 expression in vitro is modifiable by popular drinks suggesting that such approaches could potentially be used for modulating FGF19 expression in humans.

Modulation of fibroblast growth factor 19 expression by bile acids, meal replacement and energy drinks, milk, and coffee.

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Amanda M Styer

Full Text Available BACKGROUND: The enterohepatic pathway involving the fibroblast growth factor 19 (FGF19 and bile acids (BA has been linked with the etiology and remission of type 2 diabetes (T2D following Roux-en-Y gastric bypass (RYGB surgery. Specifically, diabetic patients had lower FGF19 circulating levels but postoperative FGF19 and BA levels were higher in diabetic patients that experience remission of T2D, as compared to non-diabetic patients and diabetic patients that do not experience remission. It has been proposed that this may be due to the direct flow of digestate-free bile acids into the ileum benefiting mostly T2D patients without severe diabetes. METHODS/RESULTS: We used a human colorectal cell line (LS174T that endogenously expresses FGF19, real time PCR, and Elisas for precise quantitation of FGF19 mRNA and secreted protein levels. We report here that BA and fractions of BA stimulated FGF19 in vitro but this effect was partially blocked when BA were pre-incubated with a lipoprotein mix which emulates digested food. In addition, we show that FGF19 mRNA was stimulated by meal replacement drinks (Ensure, Glucerna, SlimFast, non-fat milk, and coffee which has been linked with reduced risk for developing diabetes. Pure caffeine and the 5-hour Energy drink, on the other hand, decreased FGF19 mRNA. CONCLUSIONS: In summary, FGF19 expression in vitro is modifiable by popular drinks suggesting that such approaches could potentially be used for modulating FGF19 expression in humans.

Differential expression of CK20, β-catenin, and MUC2/5AC/6 in Lynch syndrome and familial colorectal cancer type X.

Science.gov (United States)

Haraldsson, Stefan; Klarskov, Louise; Nilbert, Mef; Bernstein, Inge; Bonde, Jesper; Holck, Susanne

2017-01-01

Hereditary non-polyposis colorectal cancer comprises Lynch syndrome and familial colorectal cancer type X (FCCTX). Differences in genetics, demographics and histopathology have been extensively studied. The purpose of this study is to characterize their immunoprofile of markers other than MMR proteins. We compared the expression patterns of cytokeratins (CK7 and CK20), mucins (MUC2/5 AC/6), CDX2 and β-catenin in Lynch syndrome and FCCTX. Differences were identified for CK20 and nuclear β-catenin, which were significantly more often expressed in FCCTX than in Lynch syndrome ( p  Lynch syndrome tumors compared with FCCTX tumors ( p  = 0.001, Lynch syndrome with a more sporadic-like profile in the former group and a more distinct profile with frequent MUC6 positivity in the latter group.

Involvement of ERK1/2 signaling pathway in atrazine action on FSH-stimulated LHR and CYP19A1 expression in rat granulosa cells

International Nuclear Information System (INIS)

Fa, Svetlana; Pogrmic-Majkic, Kristina; Samardzija, Dragana; Glisic, Branka; Kaisarevic, Sonja; Kovacevic, Radmila; Andric, Nebojsa

2013-01-01

Worldwide used herbicide atrazine is linked to reproductive dysfunction in females. In this study, we investigated the effects and the mechanism of atrazine action in the ovary using a primary culture of immature granulosa cells. In granulosa cells, follicle-stimulating hormone (FSH) activates both cyclic adenosine monophosphate (cAMP) and extracellular-regulated kinase 1/2 (ERK1/2) cascades, with cAMP pathway being more important for luteinizing hormone receptor (LHR) and aromatase (CYP19A1) mRNA expression. We report that 48 h after atrazine exposure the FSH-stimulated LHR and CYP19A1 mRNA expression and estradiol synthesis were decreased, with LHR mRNA being more sensitive to atrazine than CYP19A1 mRNA. Inadequate acquisition of LHR in the FSH-stimulated and atrazine-exposed granulosa cells renders human chorionic gonadotropin (hCG) ineffective to stimulate amphiregulin (Areg), epiregulin (Ereg), and progesterone receptor (Pgr) mRNA expression, suggesting anti-ovulatory effect of atrazine. To dissect the signaling cascade involved in atrazine action in granulosa cells, we used U0126, a pharmacological inhibitor of ERK1/2. U0126 prevents atrazine-induced decrease in LHR and CYP19A1 mRNA levels and estradiol production in the FSH-stimulated granulosa cells. ERK1/2 inactivation restores the ability of hCG to induce expression of the ovulatory genes in atrazine-exposed granulosa cells. Cell-based ELISA assay revealed that atrazine does not change the FSH-stimulated ERK1/2 phosphorylation in granulosa cells. The results from this study reveal that atrazine does not affect but requires ERK1/2 phosphorylation to cause decrease in the FSH-induced LHR and CYP19A1 mRNA levels and estradiol production in immature granulosa cells, thus compromising ovulation and female fertility. - Highlights: • Atrazine inhibits estradiol production in FSH-stimulated granulosa cells. • Atrazine inhibits LHR and Cyp19a1 mRNA expression in FSH-stimulated granulosa cells. • Atrazine

Involvement of ERK1/2 signaling pathway in atrazine action on FSH-stimulated LHR and CYP19A1 expression in rat granulosa cells

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Fa, Svetlana; Pogrmic-Majkic, Kristina; Samardzija, Dragana; Glisic, Branka; Kaisarevic, Sonja; Kovacevic, Radmila; Andric, Nebojsa, E-mail: nebojsa.andric@dbe.uns.ac.rs

2013-07-01

Worldwide used herbicide atrazine is linked to reproductive dysfunction in females. In this study, we investigated the effects and the mechanism of atrazine action in the ovary using a primary culture of immature granulosa cells. In granulosa cells, follicle-stimulating hormone (FSH) activates both cyclic adenosine monophosphate (cAMP) and extracellular-regulated kinase 1/2 (ERK1/2) cascades, with cAMP pathway being more important for luteinizing hormone receptor (LHR) and aromatase (CYP19A1) mRNA expression. We report that 48 h after atrazine exposure the FSH-stimulated LHR and CYP19A1 mRNA expression and estradiol synthesis were decreased, with LHR mRNA being more sensitive to atrazine than CYP19A1 mRNA. Inadequate acquisition of LHR in the FSH-stimulated and atrazine-exposed granulosa cells renders human chorionic gonadotropin (hCG) ineffective to stimulate amphiregulin (Areg), epiregulin (Ereg), and progesterone receptor (Pgr) mRNA expression, suggesting anti-ovulatory effect of atrazine. To dissect the signaling cascade involved in atrazine action in granulosa cells, we used U0126, a pharmacological inhibitor of ERK1/2. U0126 prevents atrazine-induced decrease in LHR and CYP19A1 mRNA levels and estradiol production in the FSH-stimulated granulosa cells. ERK1/2 inactivation restores the ability of hCG to induce expression of the ovulatory genes in atrazine-exposed granulosa cells. Cell-based ELISA assay revealed that atrazine does not change the FSH-stimulated ERK1/2 phosphorylation in granulosa cells. The results from this study reveal that atrazine does not affect but requires ERK1/2 phosphorylation to cause decrease in the FSH-induced LHR and CYP19A1 mRNA levels and estradiol production in immature granulosa cells, thus compromising ovulation and female fertility. - Highlights: • Atrazine inhibits estradiol production in FSH-stimulated granulosa cells. • Atrazine inhibits LHR and Cyp19a1 mRNA expression in FSH-stimulated granulosa cells. • Atrazine

High ALK mRNA expression has a negative prognostic significance in rhabdomyosarcoma

Science.gov (United States)

Bonvini, P; Zin, A; Alaggio, R; Pawel, B; Bisogno, G; Rosolen, A

2013-01-01

Background: Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase aberrantly expressed in cancer, but its clinical and functional importance remain controversial. Mutation or amplification of ALK, as well as its expression levels assessed by conventional immunohistochemistry methods, has been linked to prognosis in cancer, although with potential bias because of the semi-quantitative approaches. Herein, we measured ALK mRNA expression in rhabdomyosarcoma (RMS) and determined its clinical impact on patients' stratification and outcome. Methods: Specimens were obtained from RMS patients and cell lines, and ALK expression was analysed by quantitative RT–PCR, western blotting, IHC, and copy number analysis. Results: High ALK mRNA expression was detected in the vast majority of PAX3/7-FOXO1-positive tumours, whereas PAX3/7-FOXO1-negative RMS displayed considerably lower amounts of both mRNA and protein. Notably, ALK mRNA distinguished unfavourable PAX3/7-FOXO1-positive tumours from PAX3/7-FOXO1-negative RMS (Ptumour size (PALK mRNA levels were of prognostic relevance by Cox univariate regression analysis and correlated with increased risk of relapse (P=0.001) and survival (P=0.01), whereas by multivariate analysis elevated ALK mRNA expression resulted a negative prognostic marker when clinical stage was not included. Conclusion: Quantitative assessment of ALK mRNA expression helps to improve risk stratification of RMS patients and identifies tumours with adverse biological characteristics and aggressive behaviour. PMID:24149177

Whole Blood mRNA Expression-Based Prognosis of Metastatic Renal Cell Carcinoma

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Karthik V. Giridhar

2017-11-01

Full Text Available The Memorial Sloan Kettering Cancer Center (MSKCC prognostic score is based on clinical parameters. We analyzed whole blood mRNA expression in metastatic clear cell renal cell carcinoma (mCCRCC patients and compared it to the MSKCC score for predicting overall survival. In a discovery set of 19 patients with mRCC, we performed whole transcriptome RNA sequencing and selected eighteen candidate genes for further evaluation based on associations with overall survival and statistical significance. In an independent validation of set of 47 patients with mCCRCC, transcript expression of the 18 candidate genes were quantified using a customized NanoString probeset. Cox regression multivariate analysis confirmed that two of the candidate genes were significantly associated with overall survival. Higher expression of BAG1 [hazard ratio (HR of 0.14, p < 0.0001, 95% confidence interval (CI 0.04–0.36] and NOP56 (HR 0.13, p < 0.0001, 95% CI 0.05–0.34 were associated with better prognosis. A prognostic model incorporating expression of BAG1 and NOP56 into the MSKCC score improved prognostication significantly over a model using the MSKCC prognostic score only (p < 0.0001. Prognostic value of using whole blood mRNA gene profiling in mCCRCC is feasible and should be prospectively confirmed in larger studies.

Astrocyte cultures derived from human brain tissue express angiotensinogen mRNA

International Nuclear Information System (INIS)

Milsted, A.; Barna, B.P.; Ransohoff, R.M.; Brosnihan, K.B.; Ferrario, C.M.

1990-01-01

The authors have identified human cultured cell lines that are useful for studying angiotensinogen gene expression and its regulation in the central nervous system. A model cell system of human central nervous system origin expressing angiotensinogen has not previously been available. Expression of angiotensinogen mRNA appears to be a basal property of noninduced human astrocytes, since astrocytic cell lines derived from human glioblastomas or nonneoplastic human brain tissue invariably produced angiotensinogen mRNA. In situ hybridization histochemistry revealed that angiotensinogen mRNA production was not limited to a subpopulation of astrocytes because >99% of cells in these cultures contained angiotensinogen mRNA. These cell lines will be useful in studies of the molecular mechanisms controlling angiotensin synthesis and the role of biologically active angiotensin in the human brain by allowing the authors to examine regulation of expression of the renin-angiotensin system in human astrocyte cultures

[Effects of lipopolysaccharides extracted from Porphyromonas endodontalis on the expression of IL-1beta mRNA and IL-6 mRNA in osteoblasts].

Science.gov (United States)

Yang, Di; Li, Ren; Qiu, Li-Hong; Li, Chen

2009-04-01

To quantify the IL-1 beta mRNA and IL-6 mRNA expression induced by lipopolysaccharides (LPS)extracted from Porphyromonas endodontalis(P.e) in osteoblasts, and to relate P.e-LPS to bone absorption pathogenesis in lesions of chronical apical periodontitis. MG63 was treated with different concentrations of P.e-LPS(0-50 microg/mL) for different hours(0-24h). The expression of IL-1 beta mRNA and IL-6 mRNA was detected by reverse transcription polymerase chain reaction (RT-PCR).Statistical analysis was performed using one- way ANOVA and Dunnett t test with SPSS11.0 software package. The level of IL-1 beta mRNA and IL-6 mRNA increased significantly after treatment with P.e-LPS at more than 5 microg/mL (P<0.01)and for more than 1 hour (P<0.01), which indicated that P.e-LPS induced osteoblasts to express IL-1 beta mRNA and IL-6 mRNA in dose and time dependent manners. P.e-LPS may promote bone resorption in lesions of chronical apical periodontitis by inducing IL-1 beta mRNA and IL-6 mRNA expression in osteoblasts.

Expression study of CYP19A1 gene in a cohort of Iranian leiomyoma patients

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Leila Emrahi

2018-07-01

Full Text Available Background: CYP19A1 gene encodes aromatase, a microsomal key enzyme that catalyzes the synthesis of estrogens from androgens. Accumulating evidence has revealed that aromatase plays an important role in the pathogenesis of leiomyoma through increasing local concentration of estrogens. In this study, we examined the levels of CYP19A1 mRNA to determine the impact of aromatase overexpression in uterine leiomyoma growth. Subjects and methods: Tissues were obtained via myomectomy or hysterectomy from 30 patients. Total RNA was extracted and cDNA was synthesized from each frozen sample. Using SYBR Green dye, Real-time PCR assay was performed by sequence-specific primers. Relative mRNA expression was normalized to the mean of the Ct values determined for HPRT1. Gene expression ratio in each sample was determined relative to the mean ΔCt value of tumor-free margin samples. Results: PCR efficiencies for amplification reactions of HPRT1, and CYP19A genes were calculated as 0.93 and 0.96, respectively. Regression coefficients (R for standard curves were above 0.90. The obtained data revealed that the mean fold increase of CYP19A1 gene expression in leiomyoma samples relative to normal samples was 3.551 (95% CI: 0.04–6.64, S.E., 0.29–5.35. Conclusions: Our results were in accordance with previous studies and imply that up-regulation of CYP19A1 is correlated with the pathogenesis of leiomyoma tumors. We also observed that expression level of CYP19A1 was not linked to the tumor size or localization. It can be concluded that; up-regulation of aromatase is a key factor in the initiation of tumor development as well as tumor growth. Keywords: Leiomyoma, CYP19A1, Real-time PCR, Gene expression study

Exogenous mRNA encoding tetanus or botulinum neurotoxins expressed in Aplysia neurons

NARCIS (Netherlands)

Mochida, Sumiko; Poulain, Bernard; Eisel, Ulrich; Binz, Thomas; Kurazono, Hisao; Niemann, Heiner; Tauc, Ladislav; Bullock, Theodore H.

1990-01-01

Injection of exogenous mRNA purified from various tissue preparations into cellular translation systems such as Xenopus oocytes has allowed expression of complex proteins (e.g., receptors for neurotransmitters). No evidence for expression of injected exogenous mRNA, however, has been reported in

The interaction of CK2alpha and CK2beta, the subunits of protein kinase CK2, requires CK2beta in a preformed conformation and is enthalpically driven

DEFF Research Database (Denmark)

Raaf, Jennifer; Brunstein, Elena; Issinger, Olaf-Georg

2008-01-01

. In contrast to the cyclins in the case of the cyclin-dependent kinases CK2beta is no on-switch of CK2alpha; rather the formation of the CK2 holoenzyme is accompanied with an overall change of the enzyme's profile including a modulation of the substrate specificity, an increase of the thermostability......, and an allocation of docking sites for membranes and other proteins. In this study we used C-terminal deletion variants of human CK2alpha and CK2beta that were enzymologically fully competent and in particular able to form a heterotetrameric holoenzyme. With differential scanning calorimetry (DSC) we confirmed...

Expression of RFC/SLC19A1 is associated with tumor type in bladder cancer patients.

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Alyaa M Abdel-Haleem

Full Text Available Urinary bladder cancer (UBC ranks ninth in worldwide cancer. In Egypt, the pattern of bladder cancer is unique in that both the transitional and squamous cell types prevail. Despite much research on the topic, it is still difficult to predict tumor progression, optimal therapy and clinical outcome. The reduced folate carrier (RFC/SLC19A1 is the major transport system for folates in mammalian cells and tissues. RFC is also the primary means of cellular uptake for antifolate cancer chemotherapeutic drugs, however, membrane transport of antifolates by RFC is considered as limiting to antitumor activity. The purpose of this study was to compare the mRNA expression level of RFC/SLC19A1 in urothelial and non-urothelial variants of bladder carcinomas. Quantification of RFC mRNA in the mucosa of 41 untreated bladder cancer patients was performed using RT-qPCR. RFC mRNA steady-state levels were ∼9-fold higher (N = 39; P<0.0001 in bladder tumor specimens relative to normal bladder mRNA. RFC upregulation was strongly correlated with tumor type (urothelial vs. non-urothelial; p<0.05 where median RFC mRNA expression was significantly (p<0.05 higher in the urothelial (∼14-fold compared to the non-urothelial (∼4-fold variant. This may account for the variation in response to antifolate-containing regimens used in the treatment of either type. RFC mRNA levels were not associated with tumor grade (I, II and III or stage (muscle-invasive vs. non-muscle invasive implying that RFC cannot be used for prognostic purposes in bladder carcinomas and its increased expression is an early event in human bladder tumors pathogenesis. Further, RFC can be considered as a potential marker for predicting response to antifolate chemotherapy in urothelial carcinomas.

Regulation of taurine homeostasis by protein kinase CK2 in mouse fibroblasts

DEFF Research Database (Denmark)

Hansen, Daniel Bloch; Guerra, Barbara; Jacobsen, Jack Hummeland

2011-01-01

Increased expression of the ubiquitous serine/threonine protein kinase CK2 has been associated with increased proliferative capacity and increased resistance towards apoptosis. Taurine is the primary organic osmolyte involved in cell volume control in mammalian cells, and shift in cell volume...... is a critical step in cell proliferation, differentiation and induction of apoptosis. In the present study, we use mouse NIH3T3 fibroblasts and Ehrlich Lettré ascites tumour cells with different CK2 expression levels. Taurine uptake via the Na(+) dependent transporter TauT and taurine release are increased...... and reduced, respectively, following pharmacological CK2 inhibition. The effect of CK2 inhibition on TauT involves modulation of transport kinetics, whereas the effect on the taurine release pathway involves reduction in the open-probability of the efflux pathway. Stimulation of PLA(2) activity, exposure...

Lower FOXO3 mRNA expression in granulosa cells is involved in unexplained infertility.

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Yamamoto, Hikaru; Yamashita, Yoshiki; Saito, Natsuho; Hayashi, Atsushi; Hayashi, Masami; Terai, Yoshito; Ohmichi, Masahide

2017-06-01

The aim of this study was to investigate whether FOXO1 and FOXO3 mRNA expression in granulosa cells is the cause of unexplained infertility. Thirty-one patients aged infertility and 18 with male partner infertility as a control group) whose serum anti-Müllerian hormone level was >0.5 ng/μL were enrolled in the study. All patients underwent oocyte retrieval under a short protocol from June 2012 to October 2013. Real-time PCR was carried out using mRNA extracted from granulosa cells retrieved from mature follicles. We compared FOXO1 and FOXO3 mRNA expression ratios in granulosa cells between the unexplained infertility group and the male infertility group. The relation between FOXO1 and FOXO3 mRNA expression ratios in granulosa cells and assisted reproduction technology clinical outcome was also examined. FOXO3 mRNA expression ratio was significantly lower in the unexplained infertility group than in the male infertility group. Moreover, FOXO3 mRNA expression ratio showed a positive correlation with both the number of retrieved oocytes and serum anti-Müllerian hormone level. A positive correlation was also identified between FOXO1 mRNA expression and total dose of hMG. As well, the number of retrieved oocytes in the unexplained infertility group was statistically lower than that in the male infertility group. A lower FOXO3 mRNA expression in granulosa cells leads to poor oocyte development in patients with unexplained infertility undergoing controlled ovarian stimulation for in vitro fertilization-embryo transfer. © 2017 Japan Society of Obstetrics and Gynecology.

CYP19 gene expression in subcutaneous adipose tissue is associated with blood pressure in women with polycystic ovary syndrome.

Science.gov (United States)

Lecke, Sheila B; Morsch, Débora M; Spritzer, Poli M

2011-11-01

In polycystic ovary syndrome (PCOS), hypertension has been linked to androgen excess and insulin resistance. Aromatase, an enzyme encoded by the CYP19 gene, affects androgen metabolism and estrogen synthesis, influencing the androgen to estrogen balance. We characterized CYP19 gene expression in subcutaneous adipose tissue of women with PCOS and normal controls and evaluated the association between subcutaneous fat CYP19 mRNA, circulating hormone levels, and blood pressure. This case-control study was carried out with 31 PCOS patients and 27 BMI-matched normotensive non-hirsute women with regular cycles. Participants underwent anthropometric measurements, collection of blood samples, and adipose tissue biopsy (28 PCOS and 19 controls). Hypertension was defined as systolic blood pressure ≥ 130 mmHg and/or diastolic blood pressure ≥ 85 mmHg. PCOS patients were divided into normotensive and hypertensive. Main outcome measures were serum estrogen and androgen levels, estrogen-to-androgen ratio, and CYP19 gene expression in subcutaneous fat. Subcutaneous CYP19 mRNA was higher in hypertensive PCOS than in control and normotensive PCOS women (p = 0.014). Estrogen-to-androgen ratio was lower in hypertensive PCOS than controls (p androgen ratio ≤ 0.06 (median for the three groups) was observed in 91% of hypertensive PCOS women, vs. 37% and 61% in the control and normotensive PCOS groups (p = 0.011). CYP19 gene expression in subcutaneous fat of PCOS patient correlated positively with systolic (p = 0.006) and diastolic blood pressure (p = 0.009). Androgen excess and hyperinsulinemia may play a role in the molecular mechanisms that activate aromatase mRNA transcription in abdominal fat tissue. Copyright © 2011 Elsevier Inc. All rights reserved.

The mRNA expression of XRCC repair genes in mice after γ-ray radiation

International Nuclear Information System (INIS)

Wang Qin; Yue Jingyin; Li Jin; Mu Chuanjie; Fan Feiyue

2006-01-01

Objective: To investigate the role of XRCC repair genes in radioresistance of IRM-2 inbred mice. Methods: Northern hybridization was used to measure mRNA expression of XRCC1 and XRCC5 genes in IRM-2 inbred mice. ICR/JCL and 615 after exposure to different doses of γ-ray radiation at different postirradiation time. Results: The levels of XRCC1 and XRCC5 mRNA expression in control IRM-2 mice were higher significantly than those in their control parental mice (P<0.01 and P<0.05). The mRNA expression of XRCC genes in ICR/JCL and 615 mice all increased to some extent after exposure 1, 2 and 4 Gy radiation. But the levels were significantly higher at 2h postirradiation (P<0.05) . The levels of XRCC mRNA expression in IRM-2 mice did not increase significnatly compared with the control mice after exposure 1 and 2 Gy radiation. But the levels of XRCC1 and XRCC5 mRNA expression increased markedly at 4Gy 1h postirradiation (P<0.05 and P<0.01). Conclusion: The basal levels of XRCC1 and XRCC5 mRNA expression in IRM-2 mice were high. The high level of XRCC5 mRNA expression was involved in the repair of DNA double strand breaks induced by higher dose radiation, which perhaps was one of radioresistance causes of IRM-2 mice. (authors)

[Expression of heat shock protein 70 and its mRNA in career exposure to manganese].

Science.gov (United States)

Chen, Wenwen; Shao, Hua; Chi, Mingfeng; Zhang, Zhihu; Shan, Yongle; Zou, Wei

2015-10-01

To analyze the expression levels of heat shock protein70 (HSPs70) and HSPs70 mRNA in different exposure to manganese, and research the neuroprotective effect on the career exposure to manganese. From 2008 to 2009, with cross-sectional study design, and in a locomotive and rolling stock works, by stratified random sampling method, the exposed sample consisted of 180 welders from different welding shops and 100 unexposed in the last three years, non-welder controls with age-matched workers of similar socioeconomic status from the same industry. The control workers had not been exposed to neurotoxic chemicals. The mRNA expressions of four different metabolic enzyme were detected by SYBR Green I quantitative real-time polymerase chain reaction. The expression levels of the two enzymes mRNA in different exposure to manganese were analyzed. The expressions of HSPs70 were detected by Western blot. The concentration of air manganese was determined by GFAAS. The average concentration of 8 h time (8h-TWA) was used to express the level of individual exposure to manganese, according to the air manganese workplace occupational exposure limit (8h-TWA=0.15 mg/m3), the exposed group is divided into high exposed group (>0.15 mg/m3) and low exposure group (<0.15 mg/m3). The individuals exposed to manganese dose of exposed group ((0.25±0.31) mg/m3) was higher than the control group ((0.06±0.02) mg/m3) (t=6.15, P=0.001); individuals exposed to manganese dose of high exposure group for (0.42±0.34) mg/m3, which was higher than low exposure group (0.09±0.07) mg/m3 (t=9.80, P=0.001). HSPs70 mRNA and protein of exposure group (5.65±0.21, 3.26±0.15) were higher than the reference group (0.41±0.03, 1.32±0.12) (t=18.91, t=8.68, P=0.001). HSP70 mRNA and protein of high exposure group (6.48±0.37, 3.67±0.26) were higher than the low exposure group (5.15±0.23, 3.02±0.19) (t=3.24, t=2.01, P=0.003, P=0.043). The expression of peripheral blood lymphocytes HSPs70 level and HSPs70 mRNA

Diagnostic utility of LunX mRNA in peripheral blood and pleural fluid in patients with primary non-small cell lung cancer

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Tian Zhigang

2008-05-01

Full Text Available Abstract Background Progress in lung cancer is hampered by the lack of clinically useful diagnostic markers. The goal of this study was to provide a detailed evaluation of lung cancer tumor markers indicative of molecular abnormalities and to assess their diagnostic utility in non-small cell lung cancer (NSCLC patients. Methods Quantitative real-time RT-PCR was used to determine LunX, CK19, CEA, VEGF-C and hnRNP A2/B1 mRNA levels in peripheral blood and pleural fluid from NSCLC patients, compared with those from patients with other epithelial cancer (esophagus cancer and breast cancer, benign lung disease (pneumonia and tuberculo pleurisy and from healthy volunteers. Results In peripheral blood LunX mRNA was detectable in 75.0% (33/44 of patients with NSCLC, but not in patients with other epithelial cancer (0/28, benign lung disease (0/10 or in healthy volunteers (0/15. In contrast, all other genetic markers were detected in patients with either NSCLC, other epithelia cancer or benign lung disease, and in healthy volunteers. The expression level and positive rate of LunX mRNA in peripheral blood correlated with the pathologic stage of NSCLC (P LunX mRNA was detected in 92.9% (13/14 of malignant pleural fluid samples and was the only marker whose expression level was significantly different between malignant and benign pleural fluid (P LunX mRNA in the peripheral blood of NSCLC patients decreased shortly after clinical treatment (P = 0.005. Conclusion Of several commonly used genetic markers, LunX mRNA is the most specific gene marker for lung cancer and has potential diagnostic utility when measured in the peripheral blood and pleural fluid of NSCLC patients.

CK2—An Emerging Target for Neurological and Psychiatric Disorders

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Julia Castello

2017-01-01

Full Text Available Protein kinase CK2 has received a surge of attention in recent years due to the evidence of its overexpression in a variety of solid tumors and multiple myelomas as well as its participation in cell survival pathways. CK2 is also upregulated in the most prevalent and aggressive cancer of brain tissue, glioblastoma multiforme, and in preclinical models, pharmacological inhibition of the kinase has proven successful in reducing tumor size and animal mortality. CK2 is highly expressed in the mammalian brain and has many bona fide substrates that are crucial in neuronal or glial homeostasis and signaling processes across synapses. Full and conditional CK2 knockout mice have further elucidated the importance of CK2 in brain development, neuronal activity, and behavior. This review will discuss recent advances in the field that point to CK2 as a regulator of neuronal functions and as a potential novel target to treat neurological and psychiatric disorders.

Negative regulation of neuromedin U mRNA expression in the rat pars tuberalis by melatonin.

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Sayaka Aizawa

Full Text Available The pars tuberalis (PT is part of the anterior pituitary gland surrounding the median eminence as a thin cell layer. The characteristics of PT differ from those of the pars distalis (PD, such as cell composition and gene expression, suggesting that the PT has a unique physiological function compared to the PD. Because the PT highly expresses melatonin receptor type 1, it is considered a mediator of seasonal and/or circadian signals of melatonin. Expression of neuromedin U (NMU that is known to regulate energy balance has been previously reported in the rat PT; however, the regulatory mechanism of NMU mRNA expression and secretion in the PT are still obscure. In this study, we examined both the diurnal change of NMU mRNA expression in the rat PT and the effects of melatonin on NMU in vivo. In situ hybridization and quantitative PCR analysis of laser microdissected PT samples revealed that NMU mRNA expression in the PT has diurnal variation that is high during the light phase and low during the dark phase. Furthermore, melatonin administration significantly suppressed NMU mRNA expression in the PT in vivo. On the other hand, 48 h fasting did not have an effect on PT-NMU mRNA expression, and the diurnal change of NMU mRNA expression was maintained. We also found the highest expression of neuromedin U receptor type 2 (NMUR2 mRNA in the third ventricle ependymal cell layer, followed by the arcuate nucleus and the spinal cord. These results suggest that NMU mRNA expression in the PT is downregulated by melatonin during the dark phase and shows diurnal change. Considering that NMU mRNA in the PT showed the highest expression level in the brain, PT-NMU may act on NMUR2 in the brain, especially in the third ventricle ependymal cell layer, with a circadian rhythm.

NONOates regulate KCl cotransporter-1 and -3 mRNA expression in vascular smooth muscle cells.

Science.gov (United States)

Di Fulvio, Mauricio; Lauf, Peter K; Shah, Shalin; Adragna, Norma C

2003-05-01

Nitric oxide (NO) donors regulate KCl cotransport (KCC) activity and cotransporter-1 and -3 (KCC1 and KCC3) mRNA expression in sheep erythrocytes and in primary cultures of rat vascular smooth muscle cells (VSMCs), respectively. In this study, we used NONOates as rapid and slow NO releasers to provide direct evidence implicating NO as a regulator of KCC3 gene expression at the mRNA level. In addition, we used the expression of KCC3 mRNA to further investigate the mechanism of action of these NO donors at the cellular level. Treatment of VSMCs with rapid NO releasers, like NOC-5 and NOC-9, as well as with the direct NO-independent soluble guanylyl cyclase (sGC) stimulator YC-1, acutely increased KCC3 mRNA expression in a concentration- and time-dependent manner. The slow NO releaser NOC-18 had no effect on KCC3 gene expression. A specific NO scavenger completely prevented the NONOate-induced KCC3 mRNA expression. Inhibition of sGC with LY-83583 blocked the NONOate- and YC-1-induced KCC3 mRNA expression. This study shows that in primary cultures of rat VSMCs, the fast NO releasers NOC-9 and NOC-5, but not the slow NO releaser NOC-18, acutely upregulate KCC3 mRNA expression in a NO/sGC-dependent manner.

RNA Binding Protein RBM38 Regulates Expression of the 11-Kilodalton Protein of Parvovirus B19, Which Facilitates Viral DNA Replication.

Science.gov (United States)

Ganaie, Safder S; Chen, Aaron Yun; Huang, Chun; Xu, Peng; Kleiboeker, Steve; Du, Aifang; Qiu, Jianming

2018-04-15

Human parvovirus B19 (B19V) expresses a single precursor mRNA (pre-mRNA), which undergoes alternative splicing and alternative polyadenylation to generate 12 viral mRNA transcripts that encode two structural proteins (VP1 and VP2) and three nonstructural proteins (NS1, 7.5-kDa protein, and 11-kDa protein). Splicing at the second 5' donor site (D2 site) of the B19V pre-mRNA is essential for the expression of VP2 and the 11-kDa protein. We previously identified that cis -acting intronic splicing enhancer 2 (ISE2) that lies immediately after the D2 site facilitates the recognition of the D2 donor for its efficient splicing. In this study, we report that ISE2 is critical for the expression of the 11-kDa viral nonstructural protein. We found that ISE2 harbors a consensus RNA binding motif protein 38 (RBM38) binding sequence, 5'-UGUGUG-3'. RBM38 is expressed during the middle stage of erythropoiesis. We first confirmed that RBM38 binds specifically with the ISE2 element in vitro The knockdown of RBM38 significantly decreases the level of spliced mRNA at D2 that encodes the 11-kDa protein but not that of the D2-spliced mRNA that encodes VP2. Importantly, we found that the 11-kDa protein enhances viral DNA replication and virion release. Accordingly, the knockdown of RBM38 decreases virus replication via downregulating 11-kDa protein expression. Taken together, these results suggest that the 11-kDa protein facilitates B19V DNA replication and that RBM38 is an essential host factor for B19V pre-mRNA splicing and for the expression of the 11-kDa protein. IMPORTANCE B19V is a human pathogen that can cause fifth disease, arthropathy, anemia in immunocompromised patients and sickle cell disease patients, myocarditis, and hydrops fetalis in pregnant women. Human erythroid progenitor cells (EPCs) are most susceptible to B19V infection and fully support viral DNA replication. The exclusive tropism of B19V for erythroid-lineage cells is dependent not only on the expression of viral

Low ERCC1 mRNA and protein expression are associated with worse survival in cervical cancer patients treated with radiation alone

International Nuclear Information System (INIS)

Doll, Corinne M.; Prystajecky, Michael; Eliasziw, Misha; Klimowicz, Alexander C.; Petrillo, Stephanie K.; Craighead, Peter S.; Hao, Desiree; Diaz, Roman; Lees-Miller, Susan P.; Magliocco, Anthony M.

2010-01-01

Purpose: To evaluate the association of excision repair cross-complementation group 1 (ERCC1) expression, using both mRNA and protein expression analysis, with clinical outcome in cervical cancer patients treated with radical radiation therapy (RT). Experimental design: Patients (n = 186) with locally advanced cervical cancer, treated with radical RT alone from a single institution were evaluated. Pre-treatment FFPE biopsy specimens were retrieved from 112 patients. ERCC1 mRNA level was determined by real-time PCR, and ERCC1 protein expression (FL297, 8F1) was measured using quantitative immunohistochemistry (AQUA (registered) ). The association of ERCC1 status with local response, 10-year disease-free (DFS) and overall survival (OS) was analyzed. Results: ERCC1 protein expression levels using both FL297 and 8F1 antibodies were determined for 112 patients; mRNA analysis was additionally performed in 32 patients. Clinical and outcome factors were comparable between the training and validation sets. Low ERCC1 mRNA expression status was associated with worse OS (17.9% vs 50.1%, p = 0.046). ERCC1 protein expression using the FL297 antibody, but not the 8F1 antibody, was significantly associated with both OS (p = 0.002) and DFS (p = 0.010). After adjusting for pre-treatment hemoglobin in a multivariate analysis, ERCC1 FL297 expression status remained statistically significant for OS [HR 1.9 (1.1-3.3), p = 0.031]. Conclusions: Pre-treatment tumoral ERCC1 mRNA and protein expression, using the FL297 antibody, are predictive factors for survival in cervical cancer patients treated with RT, with ERCC1 FL297 expression independently associated with survival. These results identify a subset of patients who may derive the greatest benefit from the addition of cisplatin chemotherapy.

[Impacts of the formula of Suoquanwan(SQW) on expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency].

Science.gov (United States)

Cao, Hong-Ying; Wu, Qing-He; Huang, Ping; He, Jin-Yang

2009-06-01

To observe the impacts of the formula of Suoquanwan (SQW) on the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency. The model rats were induced by adenine (250 mg/kg) for 4 weeks, then treated respectively with SQW or dDAVP. The expression of AQP-2 mRNA and AVPR-V2 mRNA in kidney of Yang-deficiency model by realtime fluorescence quantitative PCR method were investigated. In model rats, the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney decreased, dDAVP and SQW high dose could increased the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney. The others had no influence on the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney. SQW can increase the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency.

1. EXPRESSION OF CK 5 and 6 in breast cancer patients at UTH.cdr

African Journals Online (AJOL)

ESEM

Indigenous Black Zambians at the University Teaching. Hospital, Lusaka ... Key words: Breast cancer, CK 5/6, Basal-like, Triple Negative 1 ... growth rate appear to increase breast cancer risk [6]. ... stresses that result in cell death []. CK 5/6 ...

Regulation of mouse hepatic CYP2D9 mRNA expression by growth and adrenal hormones.

Science.gov (United States)

Jarukamjorn, Kanokwan; Sakuma, Tsutomu; Jaruchotikamol, Atika; Oguro, Miki; Nemoto, Nobuo

2006-02-01

The constitutive expression of CYP2D9 is sexually dimorphic, namely, strong in males, but diminutive in females. Repetition of mimic growth hormone (GH) secretion pattern impressively returned the mRNA expression level to that in intact mice: the GH secretion pattern's regulation of CYP2D9 mRNA expression has been predominantly disrupted by exogenous GH-administration. The extensive decline of CYP2D9 mRNA expression becoming a sexually non-specific P450 in 9-week-old male mice exposed as neonates to monosodium L-glutamate (MSG) suggested that the male GH secretion pattern is a key to the regulation of male-specific CYP2D9 mRNA expression in adult mice. Dexamethasone (Dex) showed possibility to induce CYP2D9 mRNA expression in adult MSG-neonatally treated mice of either sex. However, the antagonism was observed by co-administration of Dex and GH in the males. Dex-administration in adrenalectomized mice significantly elevated CYP2D9 mRNA expression levels. These findings suggest that an adrenal hormone participates in the regulatory mechanism of CYP2D9 mRNA expression in association with GH.

Keratinocyte growth factor mRNA expression in periodontal ligament fibroblasts

DEFF Research Database (Denmark)

Dabelsteen, S; Wandall, H H; Grøn, B

1997-01-01

Keratinocyte growth factor (KGF) is a fibroblast growth factor which mediates epithelial growth and differentiation. KGF is expressed in subepithelial fibroblasts, but generally not in fibroblasts of deep connective tissue, such as fascia and ligaments. Here we demonstrate that KGF mRNA is expres......Keratinocyte growth factor (KGF) is a fibroblast growth factor which mediates epithelial growth and differentiation. KGF is expressed in subepithelial fibroblasts, but generally not in fibroblasts of deep connective tissue, such as fascia and ligaments. Here we demonstrate that KGF m......RNA is expressed in periodontal ligament fibroblasts, and that the expression is increased upon serum stimulation. Fibroblasts from human periodontal ligament, from buccal mucosa, from gingiva, and from skin were established from explants. Alkaline phosphatase activity was used as an indicator of the periodontal...

Bioinspired nanocomplex for spatiotemporal imaging of sequential mRNA expression in differentiating neural stem cells.

Science.gov (United States)

Wang, Zhe; Zhang, Ruili; Wang, Zhongliang; Wang, He-Fang; Wang, Yu; Zhao, Jun; Wang, Fu; Li, Weitao; Niu, Gang; Kiesewetter, Dale O; Chen, Xiaoyuan

2014-12-23

Messenger RNA plays a pivotal role in regulating cellular activities. The expression dynamics of specific mRNA contains substantial information on the intracellular milieu. Unlike the imaging of stationary mRNAs, real-time intracellular imaging of the dynamics of mRNA expression is of great value for investigating mRNA biology and exploring specific cellular cascades. In addition to advanced imaging methods, timely extracellular stimulation is another key factor in regulating the mRNA expression repertoire. The integration of effective stimulation and imaging into a single robust system would significantly improve stimulation efficiency and imaging accuracy, producing fewer unwanted artifacts. In this study, we developed a multifunctional nanocomplex to enable self-activating and spatiotemporal imaging of the dynamics of mRNA sequential expression during the neural stem cell differentiation process. This nanocomplex showed improved enzymatic stability, fast recognition kinetics, and high specificity. With a mechanism regulated by endogenous cell machinery, this nanocomplex realized the successive stimulating motif release and the dynamic imaging of chronological mRNA expression during neural stem cell differentiation without the use of transgenetic manipulation. The dynamic imaging montage of mRNA expression ultimately facilitated genetic heterogeneity analysis. In vivo lateral ventricle injection of this nanocomplex enabled endogenous neural stem cell activation and labeling at their specific differentiation stages. This nanocomplex is highly amenable as an alternative tool to explore the dynamics of intricate mRNA activities in various physiological and pathological conditions.

Parvovirus B19 Replication and Expression in Differentiating Erythroid Progenitor Cells.

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Gloria Bua

Full Text Available The pathogenic Parvovirus B19 (B19V is characterized by a strict adaptation to erythroid progenitor cells (EPCs, a heterogeneous population of differentiating cells with diverse phenotypic and functional properties. In our work, we studied the dynamics of B19V infection in EPCs in dependence on the cell differentiation stage, in terms of distribution of infected cells, synthesis of viral nucleic acids and production of infectious virus. EPCs at early differentiation stage led to an abortive infection, without viral genome replication and a very low transcriptional activity. EPCs at later stages were permissive, with highest levels of viral replicative activity at day 9 (+3.0 Log from 2 to 48 hpi and lower levels at day 18 (+1.5 Log from 2 to 48 hpi. B19V DNA increment was in accordance with the percentage of cells positive to flow-FISH assay (41.4% at day 9, 1.1% at day 18. Quantitation of total RNA indicated a close association of genome replication and transcription with viral RNA accumulation within infected cells related to viral DNA increase during the course of infection. Analysis of the different classes of mRNAs revealed two distinct pattern of genome expression profile with a fine regulation in the frequency utilization of RNA processing signals: an early phase, when cleavage at the proximal site leading to a higher relative production of mRNA for NS protein, and a late phase, when cleavage at the distal site was more frequent leading to higher relative abundance of mRNA for VP and 11 kDA proteins. Infectious virus was released from cells at day 6-15, but not at day 18. Our results, providing a detailed description of B19V replication and expression profile in differentiating EPCs, highlight the very tight adaptation of B19V to a specific cellular target defined both by its erythroid lineage and its differentiation stage.

Parvovirus B19 Replication and Expression in Differentiating Erythroid Progenitor Cells

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Bua, Gloria; Manaresi, Elisabetta; Bonvicini, Francesca; Gallinella, Giorgio

2016-01-01

The pathogenic Parvovirus B19 (B19V) is characterized by a strict adaptation to erythroid progenitor cells (EPCs), a heterogeneous population of differentiating cells with diverse phenotypic and functional properties. In our work, we studied the dynamics of B19V infection in EPCs in dependence on the cell differentiation stage, in terms of distribution of infected cells, synthesis of viral nucleic acids and production of infectious virus. EPCs at early differentiation stage led to an abortive infection, without viral genome replication and a very low transcriptional activity. EPCs at later stages were permissive, with highest levels of viral replicative activity at day 9 (+3.0 Log from 2 to 48 hpi) and lower levels at day 18 (+1.5 Log from 2 to 48 hpi). B19V DNA increment was in accordance with the percentage of cells positive to flow-FISH assay (41.4% at day 9, 1.1% at day 18). Quantitation of total RNA indicated a close association of genome replication and transcription with viral RNA accumulation within infected cells related to viral DNA increase during the course of infection. Analysis of the different classes of mRNAs revealed two distinct pattern of genome expression profile with a fine regulation in the frequency utilization of RNA processing signals: an early phase, when cleavage at the proximal site leading to a higher relative production of mRNA for NS protein, and a late phase, when cleavage at the distal site was more frequent leading to higher relative abundance of mRNA for VP and 11 kDA proteins. Infectious virus was released from cells at day 6–15, but not at day 18. Our results, providing a detailed description of B19V replication and expression profile in differentiating EPCs, highlight the very tight adaptation of B19V to a specific cellular target defined both by its erythroid lineage and its differentiation stage. PMID:26845771

Evaluation of a UCMK/dCK fusion enzyme for gemcitabine-mediated cytotoxicity

International Nuclear Information System (INIS)

Johnson, Adam J.; Brown, Melissa N.; Black, Margaret E.

2011-01-01

Highlights: ► Goal was to enhance dFdC cytotoxicity by the creation of a UCMK/dCK fusion enzyme. ► The UCMK/dCK fusion enzyme possesses both native activities. ► The fusion renders cells equally sensitive to dFdC relative to dCK expression alone. ► Dual activities of fusion not sufficient to augment cell dFdC sensitivity in vitro. ► Data may warrant the implementation of UCMK mutagenesis studies. -- Abstract: While gemcitabine (2′-2′-difluoro-2′-deoxycytidine, dFdC) displays wide-ranging antineoplastic activity as a single agent, variable response rates and poor intracellular metabolism often limit its clinical efficacy. In an effort to enhance dFdC cytotoxicity and help normalize response rates, we created a bifunctional fusion enzyme that combines the enzymatic activities of deoxycytidine kinase (dCK) and uridine/cytidine monophosphate kinase (UCMK) in a single polypeptide. Our goal was to evaluate whether the created fusion could induce beneficial, functional changes toward dFdC, expedite dFdC conversion to its active antimetabolites and consequently amplify cell dFdC sensitivity. While kinetic analyses revealed the UCMK/dCK fusion enzyme to possess both native activities, the fusion rendered cells sensitive to the cytotoxic effects of dFdC at the same level as dCK expression alone. These results suggest that increased wild-type UCMK expression does not provide a significant enhancement in dFdC-mediated cytotoxicity and may warrant the implementation of studies aimed at engineering UCMK variants with improved activity toward gemcitabine monophosphate.

Expression and clinicopathological significance of Mel-18 and Bmi-1 mRNA in gastric carcinoma.

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Lu, You-Wei; Li, Jin; Guo, Wei-Jian

2010-11-08

The Polycomb group (PcG) genes are a class of regulators responsible for maintaining homeotic gene expression throughout cell division. PcG expression is deregulated in some types of human cancer. Both Bmi-1 and Mel-18 are of the key PcG proteins. We investigate the expression and clinicopathological roles of Mel-18 and Bmi-1 mRNA in gastric cancer. The expression of Mel-18 and Bmi-1 in a series of 71 gastric cancer tissues and paired normal mucosal tissues distant from the tumorous lesion was assayed by quantitative real time RT-PCR. The correlation between Mel-18 and Bmi-1 mRNA expression, and between Mel-18 or Bmi-1 mRNA level and clinicopathological characteristics were analyzed. Expression of Mel-18 and Bmi-1 genes was variably detected, but overexpression of Bmi-1 mRNA and decreased expression of Mel-18 mRNA were the most frequent alteration. In addition, the expression of Bmi-1 and Mel-18 mRNA inversely correlates in gastric tumors. Moreover, a significant positive correlation between Bmi-1 overexpression and tumor size, depth of invasion, or lymph node metastasis, and a significant negative correlation between Mel-18 low-expression with lymph node metastasis or the clinical stage were observed. Our data suggest that Mel-18 and Bmi-1 may play crucial but opposite roles in gastric cancer. Decreased Mel-18 and increased Bmi-1 mRNA expression was associated with the carcinogenesis and progression of gastric cancer. It is possible to list Bmi-1 and Mel-18 as biomarkers for predicting the prognosis of gastric cancer.

Responses of mRNA expression of PepT1 in small intestine to ...

African Journals Online (AJOL)

To study the effect of circulation small peptides concentration on mRNA expression in small intestine, graded amount of soybean small peptides (SSP) were infused into lactating goats through duodenal fistulas. Peptide-bound amino acid (PBAA) concentration in arterial plasma and the mRNA expression of PepT1 was ...

Time-course of 5-HT(6) receptor mRNA expression during memory consolidation and amnesia.

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Huerta-Rivas, A; Pérez-García, G; González-Espinosa, C; Meneses, A

2010-01-01

Growing evidence indicates that antagonists of the 5-hydroxytryptamine (serotonin) receptor(6) (5-HT(6)) improve memory and reverse amnesia although the mechanisms involved are poorly understood. Hence, in this paper RT-PCR was used to evaluate changes in mRNA expression of 5-HT(6) receptor in trained and untrained rats treated with the 5-HT(6) receptor antagonist SB-399885 and amnesic drugs scopolamine or dizocilpine. Changes in mRNA expression of 5-HT(6) receptor were investigated at different times in prefrontal cortex, hippocampus and striatum. Data indicated that memory in the Pavlovian/instrumental autoshaping task was a progressive process associated to reduced mRNA expression of 5-HT(6) receptor in the three structures examined. SB-399885 improved long-term memory at 48h, while the muscarinic receptor antagonist scopolamine or the non-competitive NMDA receptor antagonist dizocilpine impaired it at 24h. Autoshaping training and treatment with SB-399885 increased 5-HT(6) receptor mRNA expression in (maximum increase) prefrontal cortex and striatum, 24 or 48h. The scopolamine-induced amnesia suppressed 5-HT(6) receptor mRNA expression while the dizocilpine-induced amnesia did not modify 5-HT(6) receptor mRNA expression. SB-399885 and scopolamine or dizocilpine were able to reestablish memory and 5-HT(6) receptor mRNA expression. These data confirmed previous memory evidence and of more interest is the observation that training, SB-399885 and amnesic drugs modulated 5-HT(6) receptor mRNA expression in prefrontal cortex, hippocampus and striatum. Further investigation in different memory tasks, times and amnesia models together with more complex control groups might provide further clues. Copyright 2009 Elsevier Inc. All rights reserved.

60Co γ-irradiation enhances expression of GAP-43 mRNA in rat brain

International Nuclear Information System (INIS)

Su Bingyin; Cai Wenqin; Zhang Chenggang

2001-01-01

Objective: To study the relationship between the expression of GAP-43 mRNA and nerve regeneration in rat brain after 60 Co γ-irradiation. Methods: Wistar rats were subjected to whole-body irradiation with 8 Gy 60 Co γ-rays. The expression of GAP-43 was detected by in situ hybridization histochemistry using Dig-cRNA probe. Results: It was found that the expression of GAP-43 mRNA increased in the cerebral cortex, caudate, putamen, globus pallidum, thalamus and hypothalamus one week after 8 Gy 60 Co γ-irradiation. The peak of GAP-43 mRNA expression was observed in the fourth week and then began to decrease but still remained at a higher than normal level. However, it decreased to a low level after 7 weeks. Conclusion: Enhanced expression of GAP-43 mRNA after 60 Co γ-irradiation in rat brain is associated with nerve regeneration and reconstruction of synapse

Partial Oxygen Pressure Affects the Expression of Prognostic Biomarkers HIF-1 Alpha, Ki67, and CK20 in the Microenvironment of Colorectal Cancer Tissue.

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Zhang, Lirong; Hu, Yu; Xi, Ning; Song, Jie; Huang, Wenjing; Song, Shanshan; Liu, Yiting; Liu, Xianying; Xie, Yingjun

2016-01-01

Hypoxia is prognostically important in colorectal cancer (CRC) therapy. Partial oxygen pressure (pO 2 ) is an important parameter of hypoxia. The correlation between pO 2 levels and expression levels of prognostic biomarkers was measured in CRC tissues. Human CRC tissues were collected and pO 2 levels were measured by OxyLite. Three methods for tissue fixation were compared, including formalin, Finefix, and Finefix-plus-microwave. Immunohistochemistry (IHC) staining was conducted by using the avidin-biotin complex technique for detecting the antibodies to hypoxia inducible factor-1 (HIF-1) alpha, cytokeratin 20 (CK20), and cell proliferation factor Ki67. The levels of pO 2 were negatively associated with the size of CRC tissues. Finefix-plus-microwave fixation has the potential to replace formalin. Additionally, microwave treatment improved Finefix performance in tissue fixation and protein preservation. The percentage of positive cells and gray values of HIF-1 alpha, CK20, and Ki67 were associated with CRC development ( P < 0.05). The levels of pO 2 were positively related with the gray values of Ki67 and negatively related with the values of HIF-1 alpha and CK20 ( P < 0.05). Thus, the levels of microenvironmental pO 2 affect the expression of predictive biomarkers HIF-1 alpha, CK20, and Ki67 in the development of CRC tissues.

Partial Oxygen Pressure Affects the Expression of Prognostic Biomarkers HIF-1 Alpha, Ki67, and CK20 in the Microenvironment of Colorectal Cancer Tissue

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Lirong Zhang

2016-01-01

Full Text Available Hypoxia is prognostically important in colorectal cancer (CRC therapy. Partial oxygen pressure (pO2 is an important parameter of hypoxia. The correlation between pO2 levels and expression levels of prognostic biomarkers was measured in CRC tissues. Human CRC tissues were collected and pO2 levels were measured by OxyLite. Three methods for tissue fixation were compared, including formalin, Finefix, and Finefix-plus-microwave. Immunohistochemistry (IHC staining was conducted by using the avidin-biotin complex technique for detecting the antibodies to hypoxia inducible factor-1 (HIF-1 alpha, cytokeratin 20 (CK20, and cell proliferation factor Ki67. The levels of pO2 were negatively associated with the size of CRC tissues. Finefix-plus-microwave fixation has the potential to replace formalin. Additionally, microwave treatment improved Finefix performance in tissue fixation and protein preservation. The percentage of positive cells and gray values of HIF-1 alpha, CK20, and Ki67 were associated with CRC development (P<0.05. The levels of pO2 were positively related with the gray values of Ki67 and negatively related with the values of HIF-1 alpha and CK20 (P<0.05. Thus, the levels of microenvironmental pO2 affect the expression of predictive biomarkers HIF-1 alpha, CK20, and Ki67 in the development of CRC tissues.

Protein kinase CK2 in health and disease: Protein kinase CK2: from structures to insights

DEFF Research Database (Denmark)

Niefind, K; Raaf, J; Issinger, Olaf-Georg

2009-01-01

the critical region of CK2alpha recruitment is pre-formed in the unbound state. In CK2alpha the activation segment - a key element of protein kinase regulation - adapts invariably the typical conformation of the active enzymes. Recent structures of human CK2alpha revealed a surprising plasticity in the ATP......Within the last decade, 40 crystal structures corresponding to protein kinase CK2 (former name 'casein kinase 2'), to its catalytic subunit CK2alpha and to its regulatory subunit CK2beta were published. Together they provide a valuable, yet by far not complete basis to rationalize the biochemical...

mRNA Expression of Ovine Angiopoietin-like Protein 4 Gene in Adipose Tissues

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Jing Zhang

2016-05-01

Full Text Available Angiopoietin-like protein 4 (ANGPTL4 is involved in a variety of functions, including lipoprotein metabolism and angiogenesis. To reveal the role of ANGPTL4 in fat metabolism of sheep, ovine ANGPTL4 mRNA expression was analyzed in seven adipose tissues from two breeds with distinct tail types. Forty-eight animals with the gender ratio of 1:1 for both Guangling Large Tailed (GLT and Small Tailed Han (STH sheep were slaughtered at 2, 4, 6, 8, 10, and 12 months of age, respectively. Adipose tissues were collected from greater and lesser omental, subcutaneous, retroperitoneal, perirenal, mesenteric, and tail fats. Ontogenetic mRNA expression of ANGPTL4 in these adipose tissues from GTL and STH was studied by quantitative real time polymerase chain reaction. The results showed that ANGPTL4 mRNA expressed in all adipose tissues studied with the highest in subcutaneous and the lowest in mesenteric fat depots. Months of age, tissue and breed are the main factors that significantly influence the mRNA expression. These results provide new insights into ovine ANGPTL4 gene expression and clues for its function mechanism.

Cyclic-AMP mediated regulation of ABCB mRNA expression in mussel haemocytes.

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Silvia Franzellitti

Full Text Available BACKGROUND: The multixenobiotic resistance system (MXR allows aquatic organisms to cope with their habitat despite high pollution levels by over-expressing membrane and intracellular transporters, including the P-glycoprotein (Pgp. In mammals transcription of the ABCB1 gene encoding Pgp is under cAMP/PKA-mediated regulation; whether this is true in mollusks is not fully clarified. METHODOLOGY/PRINCIPAL FINDINGS: cAMP/PKA regulation and ABCB mRNA expression were assessed in haemocytes from Mediterranean mussels (Mytilus galloprovincialis exposed in vivo for 1 week to 0.3 ng/L fluoxetine (FX alone or in combination with 0.3 ng/L propranolol (PROP. FX significantly decreased cAMP levels and PKA activity, and induced ABCB mRNA down-regulation. FX effects were abolished in the presence of PROP. In vitro experiments using haemocytes treated with physiological agonists (noradrenaline and serotonin and pharmacological modulators (PROP, forskolin, dbcAMP, and H89 of the cAMP/PKA system were performed to obtain clear evidence about the involvement of the signaling pathway in the transcriptional regulation of ABCB. Serotonin (5-HT decreased cAMP levels, PKA activity and ABCB mRNA expression but increased the mRNA levels for a putative 5-HT1 receptor. Interestingly, 5-HT1 was also over-expressed after in vivo exposures to FX. 5-HT effects were counteracted by PROP. Forskolin and dbcAMP increased PKA activity as well as ABCB mRNA expression; the latter effect was abolished in the presence of the PKA inhibitor H89. CONCLUSIONS: This study provides the first direct evidence for the cAMP/PKA-mediated regulation of ABCB transcription in mussels.

Region specific regulation of glutamic acid decarboxylase mRNA expression by dopamine neurons in rat brain.

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Lindefors, N; Brene, S; Herrera-Marschitz, M; Persson, H

1989-01-01

In situ hybridization histochemistry and RNA blots were used to study the expression of glutamic acid decarboxylase (GAD) mRNA in rats with or without a unilateral lesion of midbrain dopamine neurons. Two populations of GAD mRNA positive neurons were found in the intact caudate-putamen, substantia nigra and fronto-parietal cortex. In caudate-putamen, only one out of ten of the GAD mRNA positive neurons expressed high levels, while in substantia nigra every second of the positive neurons expressed high levels of GAD mRNA. Relatively few, but intensively labelled neurons were found in the intact fronto-parietal cerebral cortex. In addition, one out of six of the GAD mRNA positive neurons in the fronto-parietal cortex showed a low labeling. On the ipsilateral side, the forebrain dopamine deafferentation induced an increase in the number of neurons expressing high levels of GAD mRNA in caudate-putamen, and a decrease in fronto-parietal cortex. A smaller decrease was also seen in substantia nigra. However, the total number of GAD mRNA positive neurons were not significantly changed in any of these brain regions. The changes in the levels of GAD mRNA after the dopamine lesion were confirmed by RNA blot analysis. Hence, midbrain dopamine neurons appear to control neuronal expression of GAD mRNA by a tonic down-regulation in a fraction of GAD mRNA positive neurons in caudate-putamen, and a tonic up-regulation in a fraction of GAD mRNA positive neurons in fronto-parietal cortex and substantia nigra.

Cytokeratin 19 promoter directs the expression of Cre recombinase in various epithelia of transgenic mice.

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Zhao, Gui-Feng; Zhao, Shuang; Liu, Jia-Jie; Wu, Ji-Cheng; He, Hao-Yu; Ding, Xiao-Qing; Yu, Xue-Wen; Huang, Ke-Qiang; Li, Zhi-Jie; Zheng, Hua-Chuan

2017-03-14

Cytokeratin 19 (K19) is expressed in various differentiated cells, including gastric, intestinal and bronchial epithelial cells, and liver duct cells. Here, we generated a transgenic mouse line, K19-Cre, in which the expression of Cre recombinase was controlled by the promoter of K19. To test the tissue distribution and excision activity of Cre recombinase, K19-Cre transgenic mice were bred with Rosa26 reporter strain and a mouse strain that carries PTEN conditional alleles (PTENLoxp/Loxp). At mRNA level, Cre was strongly expressed in the stomach, lung and intestine, while in stomach, lung, and liver at protein level. The immunoreactivity to Cre was strongly observed the cytoplasm of gastric, bronchial and intestinal epithelial cells. Cre activity was detectable in gastric, bronchial and intestinal epithelial cells, according to LacZ staining. In K19-Cre/PTEN Loxp/Loxp mice, PTEN was abrogated in stomach, intestine, lung, liver and breast, the former two of which were verified by in situ PCR. There appeared breast cancer with PTEN loss. These data suggest that K19 promoter may be a useful tool to study the pathophysiological functions of cytokeratin 19-positive cells, especially gastrointestinal epithelial cells. Cell specificity of neoplasia is not completely attributable to the cell-specific expression of oncogenes and cell-specific loss of tumor suppressor genes.

Expressão do Mg+2, CK, AST e LDH em equinos finalistas de provas de enduro Endurance horses finalists: expression of Mg+2, CK, AST and LDH in horse finalists of endurance race

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Juliana V.F. Sales

2013-01-01

ção de permeabilidade das células musculares estriadas esqueléticas, sugerindo o estabelecimento de um processo inflamatório agudo. Devido à expressão da atividade enzimática da CK (p≤0,001, por sua especificidade em relação à ocorrência de danos na musculatura estriada esquelética, juntamente com o íon magnésio (p=0,0004 que participa de várias reações celulares. Houve alterações na concentração de proteína plasmática total (p=0,0009 e hematócrito (p=0,0001, entre os momentos avaliados. Portanto estes resultados servem como valores de referência de equinos finalistas de provas de enduro de 90 km, auxiliando na prevenção da ocorrência de possíveis danos musculares e processos inflamatórios severos.In recent years, due to rising competitive demands, the equine athlete is being increasingly required. Thus, the demands for high performance have fostered interest in the study of pathophysiology of various horse diseases. The relationship between magnesium and exercise has received significant attention because this ion is closely related with the skeletal muscle tissue. Moreover, among the main strategies for the detection and monitoring of clinical muscle damage, features the evaluation of the activity of the enzymes creatine kinase (CK, lactate dehydrogenase (LDH and aspartate aminotransferase (AST. The search for the establishment of parameters that relate to each other is a determining factor in understanding the physiological changes found on athletic horses in effort. Thus, this study aimed to determine how the blood concentrations of magnesium ion and the enzymatic activities of the enzymes CK, LDH and AST behave in Arabian finalist horses in endurance races of 90km and to relate possible changes to the type of physical effort played by animals. It was evaluated the enzymatic activities of the enzymes CK, LDH, AST e the concentration of the ion magnesium in exercise in relation to the rest state of 14 clinically healthy Arabian horses, 9

TP53 and ATM mRNA expression in skin and skeletal muscle after low-level laser exposure.

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Guedes de Almeida, Luciana; Sergio, Luiz Philippe da Silva; de Paoli, Flavia; Mencalha, Andre Luiz; da Fonseca, Adenilson de Souza

2017-08-01

Low-level lasers are widespread in regenerative medicine, but the molecular mechanisms involved in their biological effects are not fully understood, particularly those on DNA stability. Therefore, this study aimed to investigate mRNA expression of genes related to DNA genomic stability in skin and skeletal muscle tissue from Wistar rats exposed to low-level red and infrared lasers. For this, TP53 (Tumor Protein 53) and ATM (Ataxia Telangiectasia Mutated gene) mRNA expressions were evaluated by real-time quantitative PCR (RT-qPCR) technique 24 hours after low-level red and infrared laser exposure. Our data showed that relative TP53 mRNA expression was not significantly altered in both tissues exposed to lasers. For ATM, relative mRNA expression in skin tissue was not significantly altered, but in muscle tissue, laser exposure increased relative ATM mRNA expression. Low-level red and infrared laser radiations alter ATM mRNA expression related to DNA stability in skeletal muscle tissue.

Survivin gene levels in the peripheral blood of patients with gastric cancer independently predict survival

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Scalerta Romano

2009-12-01

Full Text Available Abstract Background The detection of circulating tumor cells (CTC is considered a promising tool for improving risk stratification in patients with solid tumors. We investigated on whether the expression of CTC related genes adds any prognostic power to the TNM staging system in patients with gastric carcinoma. Methods Seventy patients with TNM stage I to IV gastric carcinoma were retrospectively enrolled. Peripheral blood samples were tested by means of quantitative real time PCR (qrtPCR for the expression of four CTC related genes: carcinoembryonic antigen (CEA, cytokeratin-19 (CK19, vascular endothelial growth factor (VEGF and Survivin (BIRC5. Results Gene expression of Survivin, CK19, CEA and VEGF was higher than in normal controls in 98.6%, 97.1%, 42.9% and 38.6% of cases, respectively, suggesting a potential diagnostic value of both Survivin and CK19. At multivariable survival analysis, TNM staging and Survivin mRNA levels were retained as independent prognostic factors, demonstrating that Survivin expression in the peripheral blood adds prognostic information to the TNM system. In contrast with previously published data, the transcript abundance of CEA, CK19 and VEGF was not associated with patients' clinical outcome. Conclusions Gene expression levels of Survivin add significant prognostic value to the current TNM staging system. The validation of these findings in larger prospective and multicentric series might lead to the implementation of this biomarker in the routine clinical setting in order to optimize risk stratification and ultimately personalize the therapeutic management of these patients.

Calciotrophic hormones and hyperglycemia modulate vitamin D receptor and 25 hydroxyy vitamin D 1-α hydroxylase mRNA expression in human vascular smooth muscle cells.

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Somjen, D; Knoll, E; Sharon, O; Many, A; Stern, N

2015-04-01

Estrogen receptors (ERα and ERβ), the vitamin D receptor (VDR) and 25 hydroxyy vitamin D 1-α hydroxylase (1OHase) mRNA are expressed in vascular smooth muscle cells (VSMC). In these cells estrogenic hormones modulate cell proliferation as measured by DNA synthesis (DNA). In the present study we determined whether or not the calciotrophic hormones PTH 1-34 (PTH) and less- calcemic vitamin D analog QW as well as hyperglycemia can regulate DNA synthesis and CK. E2 had a bimodal effect on VSMC DNA synthesis, such that proliferation was inhibited at 30nM but stimulated at 0.3nM. PTH at 50nM increased, whereas QW at 10nM inhibited DNA synthesis. Hyperglycemia inhibited the effects on high E2, QW and PTH on DNA only. Both QW and PTH increased ERα mRNA expression, but only PTH increased ERβ expression. Likewise, both PTH and QW stimulated VDR and 1OHase expression and activity. ERβ, VDR and 1OHase expression and activity were inhibited by hyperglycemia, but ERα expression was unaffected by hyperglycemia. In conclusion, calcitrophic hormones modify VSMC growth and concomitantly affect ER expression in these cells as well as the endogenous VSMC vitamin D system elements, including VDR and 1OHase. Some of the later changes may likely participate in growth effects. Of importance in the observation is that several regulatory effects are deranged in the presence of hyperglycemia, particularly the PTH- and vitamin D-dependent up regulation of VDR and 1OHase in these cells. The implications of these effects require further studies. This article is part of a Special Issue entitled '17th Vitamin D Workshop'. Copyright © 2014 Elsevier Ltd. All rights reserved.

Correlation of mRNA Expression and Signal Variability in Chronic Intracortical Electrodes.

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Falcone, Jessica D; Carroll, Sheridan L; Saxena, Tarun; Mandavia, Dev; Clark, Alexus; Yarabarla, Varun; Bellamkonda, Ravi V

2018-01-01

The goal for this research was to identify molecular mechanisms that explain animal-to-animal variability in chronic intracortical recordings. Microwire electrodes were implanted into Sprague Dawley rats at an acute (1 week) and a chronic (14 weeks) time point. Weekly recordings were conducted, and action potentials were evoked in the barrel cortex by deflecting the rat's whiskers. At 1 and 14 weeks, tissue was collected, and mRNA was extracted. mRNA expression was compared between 1 and 14 weeks using a high throughput multiplexed qRT-PCR. Pearson correlation coefficients were calculated between mRNA expression and signal-to-noise ratios at 14 weeks. At 14 weeks, a positive correlation between signal-to-noise ratio (SNR) and NeuN and GFAP mRNA expression was observed, indicating a relationship between recording strength and neuronal population, as well as reactive astrocyte activity. The inflammatory state around the electrode interface was evaluated using M1-like and M2-like markers. Expression for both M1-like and M2-like mRNA markers remained steady from 1 to 14 weeks. Anti-inflammatory markers, CD206 and CD163, however, demonstrated a significant positive correlation with SNR quality at 14 weeks. VE-cadherin, a marker for adherens junctions, and PDGFR-β, a marker for pericytes, both partial representatives of blood-brain barrier health, had a positive correlation with SNR at 14 weeks. Endothelial adhesion markers revealed a significant increase in expression at 14 weeks, while CD45, a pan-leukocyte marker, significantly decreased at 14 weeks. No significant correlation was found for either the endothelial adhesion or pan-leukocyte markers. A positive correlation between anti-inflammatory and blood-brain barrier health mRNA markers with electrophysiological efficacy of implanted intracortical electrodes has been demonstrated. These data reveal potential mechanisms for further evaluation to determine potential target mechanisms to improve

Bone marrow-derived mesenchymal stem cells effectively regenerate fibrotic liver in bile duct ligation rat model.

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Mohamed, Hoda E; Elswefy, Sahar E; Rashed, Laila A; Younis, Nahla N; Shaheen, Mohamed A; Ghanim, Amal M H

2016-03-01

Mesenchymal stem cells (MSCs) have attracted lots of attention for the treatment of acute liver failure and end-stage liver diseases. This study aimed at investigating the fundamental mechanism by which bone marrow-derived MSCs (BM-MSCs) induce liver regeneration of fibrotic liver in rats. Rats underwent bile duct ligation (BDL) surgery and four weeks later they were treated with either BM-MSCs (3 × 10(6) cells /rat, once, tail vein injection) or silymarin (100 mg/kg, daily, orally) for four weeks. Liver function tests and hepatic oxidative stress were determined. Hepatic injury and fibrosis were assessed by H and E, Sirus red staining and immunohistochemical expression of α-smooth muscle actin (α-SMA). Hepatocyte growth factor (HGF) and the gene expression of cytokeratin-19 (CK-19) and matrix metalloproteinase-2 (MMP-2) in liver tissue were determined. BDL induced cholestatic liver injury characterized by elevated ALT and AST activities, bilirubin and decreased albumin. The architecture damage was staged as Metavir score: F3, A3. Fibrosis increased around proliferating bile duct as indicated by sirus red staining and α-SMA immunostaining. Fibrogenesis was favored over fibrolysis and confirmed by decreased HGF with increased expression of CK-19, but decreased MMP-2 expression. BM-MSCs treatment restored deteriorated liver functions and restored the histological changes, resolved fibrosis by improving liver regenerative capabilities (P liver regenerative capabilities can be stimulated by BM-MSCs via augmentation of HGF that subsequently up-regulate MMP-2 mRNA while downregulating CK-19 mRNA. © 2016 by the Society for Experimental Biology and Medicine.

Distinction between papillary thyroid hyperplasia and papillary thyroid carcinoma by immunohistochemical staining for cytokeratin 19, galectin-3, and HBME-1.

Science.gov (United States)

Casey, Mary B; Lohse, Christine M; Lloyd, Ricardo V

2003-01-01

The histopathology of papillary thyroid hyperplasia and papillary thyroid carcinoma is similar enough to cause a diagnostic dilemma in a few cases. Both lesions may have papillary fronds with fibrovascular cores, nuclear crowding, and nuclear anisocytosis. Formalin- fixed paraffin-embedded tissues from 30 randomly selected patients with papillary thyroid hyperplasia and an equal number from patients with papillary thyroid carcinoma were analyzed for expression of cytokeratin 19 (CK19), galectin-3, and HBME-1. Cases of papillary thyroid carcinoma had moderate to strong CK19, galectin-3, and HBME-1 reactivity although both CK19 and galectin-3 showed positive staining in a significant number of nonneoplastic thyroid cases. HBME-1 was uncommon in the nonneoplastic cases. These results indicate that HBME-1 may be useful in helping to distinguish papillary thyroid carcinoma from hyperplasia in diagnostically difficult cases.

RP1 is a phosphorylation target of CK2 and is involved in cell adhesion.

Science.gov (United States)

Stenner, Frank; Liewen, Heike; Göttig, Stephan; Henschler, Reinhard; Markuly, Norbert; Kleber, Sascha; Faust, Michael; Mischo, Axel; Bauer, Stefan; Zweifel, Martin; Knuth, Alexander; Renner, Christoph; Wadle, Andreas

2013-01-01

RP1 (synonym: MAPRE2, EB2) is a member of the microtubule binding EB1 protein family, which interacts with APC, a key regulatory molecule in the Wnt signalling pathway. While the other EB1 proteins are well characterized the cellular function and regulation of RP1 remain speculative to date. However, recently RP1 has been implicated in pancreatic cancerogenesis. CK2 is a pleiotropic kinase involved in adhesion, proliferation and anti-apoptosis. Overexpression of protein kinase CK2 is a hallmark of many cancers and supports the malignant phenotype of tumor cells. In this study we investigate the interaction of protein kinase CK2 with RP1 and demonstrate that CK2 phosphorylates RP1 at Ser(236) in vitro. Stable RP1 expression in cell lines leads to a significant cleavage and down-regulation of N-cadherin and impaired adhesion. Cells expressing a Phospho-mimicking point mutant RP1-ASP(236) show a marked decrease of adhesion to endothelial cells under shear stress. Inversely, we found that the cells under shear stress downregulate endogenous RP1, most likely to improve cellular adhesion. Accordingly, when RP1 expression is suppressed by shRNA, cells lacking RP1 display significantly increased cell adherence to surfaces. In summary, RP1 phosphorylation at Ser(236) by CK2 seems to play a significant role in cell adhesion and might initiate new insights in the CK2 and EB1 family protein association.

RP1 is a phosphorylation target of CK2 and is involved in cell adhesion.

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Frank Stenner

Full Text Available RP1 (synonym: MAPRE2, EB2 is a member of the microtubule binding EB1 protein family, which interacts with APC, a key regulatory molecule in the Wnt signalling pathway. While the other EB1 proteins are well characterized the cellular function and regulation of RP1 remain speculative to date. However, recently RP1 has been implicated in pancreatic cancerogenesis. CK2 is a pleiotropic kinase involved in adhesion, proliferation and anti-apoptosis. Overexpression of protein kinase CK2 is a hallmark of many cancers and supports the malignant phenotype of tumor cells. In this study we investigate the interaction of protein kinase CK2 with RP1 and demonstrate that CK2 phosphorylates RP1 at Ser(236 in vitro. Stable RP1 expression in cell lines leads to a significant cleavage and down-regulation of N-cadherin and impaired adhesion. Cells expressing a Phospho-mimicking point mutant RP1-ASP(236 show a marked decrease of adhesion to endothelial cells under shear stress. Inversely, we found that the cells under shear stress downregulate endogenous RP1, most likely to improve cellular adhesion. Accordingly, when RP1 expression is suppressed by shRNA, cells lacking RP1 display significantly increased cell adherence to surfaces. In summary, RP1 phosphorylation at Ser(236 by CK2 seems to play a significant role in cell adhesion and might initiate new insights in the CK2 and EB1 family protein association.

Interaction between CK2α and CK2β, the Subunits of Protein Kinase CK2: Thermodynamic Contributions of Key Residues on the CK2α Surface

DEFF Research Database (Denmark)

Raaf, J; Bischoff, N; Kloppfleisch, K

2011-01-01

that Leu41 or Phe54 single mutations were most disruptive to binding of CK2β. Additionally, these CK2α mutants retained their kinase activity. Furthermore, the substitution of Leu41 in combination with Phe54 showed that the individual mutations were not additive, suggesting that the cooperative action...

Expression and significance of cyclooxygenase-2 mRNA in benign and malignant ascites

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Lu, Jing; Li, Xiao-Feng; Kong, Li-Xia; Ma, Lin; Liao, Su-Huan; Jiang, Chang-You

2013-01-01

AIM: To investigate the mRNA expression of cyclooxygensae-2 (COX-2) in benign and malignant ascites, and to explore the difference in COX-2 mRNA expression among different diseases. METHODS: A total of 36 samples were collected from the Fifth Affiliated Hospital of Sun Yat-Sen University and divided into two experimental groups: benign ascites (n = 21) and malignant ascites (n = 15). Benign ascites included cirrhotic ascites (n = 10) and tuberculous ascites (n = 5). Malignant ascites included oophoroma (n = 7), cancer of colon (n = 5), cancer of the liver (n = 6), gastric cancer (n = 2), and bladder carcinoma (n = 1). The mRNA expression of COX-2 in ascites was examined with reverse transcriptase polymerase chain reaction (RT-PCR) technology, and the positive rate of COX-2 mRNA was compared between different diseases. RESULTS: The positive rate of COX-2 mRNA in malignant ascites was 42.9% (9/21), which was significantly higher than in benign ascites, 6.7% (1/15), difference being significant between these two groups (χ2 = 4.051, P = 0.044). The proportion of the positive rate in the malignant ascites was as follows: ovarian cancers 57.1% (4/7), colon cancer 40.0% (2/5), liver cancer 33.3% (2/6), gastric cancer 50.0% (1/2), and bladder cancer 0.00% (0/1). However, there was no significant difference in COX-2 mRNA expression among various tumors with malignant ascites (χ2 = 1.614, P = 0.806). Among the benign ascites, COX-2 mRNA levels were different between the tuberculous ascites (0/5) and cirrhotic ascites (1/10), but there was no significant difference (P = 1.000). CONCLUSION: COX-2 mRNA, detected by RT-PCR, is useful in the differential diagnosis of benign and malignant ascites, which also has potential value in the clinical diagnosis of tumors. PMID:24187465

Nonparametric testing for DNA copy number induced differential mRNA gene expression

NARCIS (Netherlands)

van Wieringen, W.N.; van de Wiel, M.A.

2009-01-01

The central dogma of molecular biology relates DNA with mRNA. Array CGH measures DNA copy number and gene expression microarrays measure the amount of mRNA. Methods that integrate data from these two platforms may uncover meaningful biological relationships that further our understanding of cancer.

Molecular evolution of adiponectin in Carnivora and its mRNA expression in relation to hepatic lipidosis.

Science.gov (United States)

Nieminen, Petteri; Rouvinen-Watt, Kirsti; Kapiainen, Suvi; Harris, Lora; Mustonen, Anne-Mari

2010-09-15

Adiponectin is a novel adipocyte-derived hormone with low circulating concentrations and/or mRNA expression in obesity and non-alcoholic fatty liver disease (NAFLD). The adiponectin mRNA of several Carnivora species was sequenced to enable further gene expression studies in this clade with potential experimental species to examine the connections of hypoadiponectinemia to hepatic lipidosis. In addition, adiponectin mRNA expression was studied in the retroperitoneal fat of the American mink (Neovison vison), as hepatic lipidosis with close similarities to NAFLD can be rapidly induced to the species by fasting. The mRNA expression was determined after overnight-7d of food deprivation and 28d of re-feeding and correlated to the liver fat %. The homologies between the determined carnivoran mRNA sequences and that of the domestic dog were 92.2-99.1%. As the mRNA expression was not affected by short-term fasting and did not correlate with the liver fat %, there seems to be no clear connection between adiponectin and the development of lipidosis in the American mink. In the future, the obtained sequences can be utilized in further studies of adiponectin expression in comparative endocrinology. Copyright (c) 2010 Elsevier Inc. All rights reserved.

Applying the breaks on gene expression - mRNA deadenylation by Pop2p

DEFF Research Database (Denmark)

Andersen, Kasper Røjkjær; Jonstrup, Anette Thyssen; Van, Lan Bich

When driving a car, control of the brakes is just as important as control of the accelerator pedal. Likewise, in gene expression, regulation of mRNA degradation is as important as regulation of its synthesis (Mühlemann, 2005). The rate-determining step of mRNA decay in eukaryotes seems to be the ......When driving a car, control of the brakes is just as important as control of the accelerator pedal. Likewise, in gene expression, regulation of mRNA degradation is as important as regulation of its synthesis (Mühlemann, 2005). The rate-determining step of mRNA decay in eukaryotes seems...... to be the shortening of the poly(A) tail (deadenylation), as this step is slower than the subsequent decapping and degradation of the mRNA body. The Mega-Dalton Ccr4-Not complex contains two exonucleases, Ccr4p and Pop2p, responsible for this process. It is not known at present why two conserved nucleases are needed...

Rhythmic expression of Nocturnin mRNA in multiple tissues of the mouse

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Green Carla B

2001-05-01

Full Text Available Abstract Background Nocturnin was originally identified by differential display as a circadian clock regulated gene with high expression at night in photoreceptors of the African clawed frog, Xenopus laevis. Although encoding a novel protein, the nocturnin cDNA had strong sequence similarity with a C-terminal domain of the yeast transcription factor CCR4, and with mouse and human ESTs. Since its original identification others have cloned mouse and human homologues of nocturnin/CCR4, and we have cloned a full-length cDNA from mouse retina, along with partial cDNAs from human, cow and chicken. The goal of this study was to determine the temporal pattern of nocturnin mRNA expression in multiple tissues of the mouse. Results cDNA sequence analysis revealed a high degree of conservation among vertebrate nocturnin/CCR4 homologues along with a possible homologue in Drosophila. Northern analysis of mRNA in C3H/He and C57/Bl6 mice revealed that the mNoc gene is expressed in a broad range of tissues, with greatest abundance in liver, kidney and testis. mNoc is also expressed in multiple brain regions including suprachiasmatic nucleus and pineal gland. Furthermore, mNoc exhibits circadian rhythmicity of mRNA abundance with peak levels at the time of light offset in the retina, spleen, heart, kidney and liver. Conclusion The widespread expression and rhythmicity of mNoc mRNA parallels the widespread expression of other circadian clock genes in mammalian tissues, and suggests that nocturnin plays an important role in clock function or as a circadian clock effector.

Combining miRNA and mRNA Expression Profiles in Wilms Tumor Subtypes

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Nicole Ludwig

2016-03-01

Full Text Available Wilms tumor (WT is the most common childhood renal cancer. Recent findings of mutations in microRNA (miRNA processing proteins suggest a pivotal role of miRNAs in WT genesis. We performed miRNA expression profiling of 36 WTs of different subtypes and four normal kidney tissues using microarrays. Additionally, we determined the gene expression profile of 28 of these tumors to identify potentially correlated target genes and affected pathways. We identified 85 miRNAs and 2107 messenger RNAs (mRNA differentially expressed in blastemal WT, and 266 miRNAs and 1267 mRNAs differentially expressed in regressive subtype. The hierarchical clustering of the samples, using either the miRNA or mRNA profile, showed the clear separation of WT from normal kidney samples, but the miRNA pattern yielded better separation of WT subtypes. A correlation analysis of the deregulated miRNA and mRNAs identified 13,026 miRNA/mRNA pairs with inversely correlated expression, of which 2844 are potential interactions of miRNA and their predicted mRNA targets. We found significant upregulation of miRNAs-183, -301a/b and -335 for the blastemal subtype, and miRNAs-181b, -223 and -630 for the regressive subtype. We found marked deregulation of miRNAs regulating epithelial to mesenchymal transition, especially in the blastemal subtype, and miRNAs influencing chemosensitivity, especially in regressive subtypes. Further research is needed to assess the influence of preoperative chemotherapy and tumor infiltrating lymphocytes on the miRNA and mRNA patterns in WT.

Connecting protein and mRNA burst distributions for stochastic models of gene expression

International Nuclear Information System (INIS)

Elgart, Vlad; Jia, Tao; Fenley, Andrew T; Kulkarni, Rahul

2011-01-01

The intrinsic stochasticity of gene expression can lead to large variability in protein levels for genetically identical cells. Such variability in protein levels can arise from infrequent synthesis of mRNAs which in turn give rise to bursts of protein expression. Protein expression occurring in bursts has indeed been observed experimentally and recent studies have also found evidence for transcriptional bursting, i.e. production of mRNAs in bursts. Given that there are distinct experimental techniques for quantifying the noise at different stages of gene expression, it is of interest to derive analytical results connecting experimental observations at different levels. In this work, we consider stochastic models of gene expression for which mRNA and protein production occurs in independent bursts. For such models, we derive analytical expressions connecting protein and mRNA burst distributions which show how the functional form of the mRNA burst distribution can be inferred from the protein burst distribution. Additionally, if gene expression is repressed such that observed protein bursts arise only from single mRNAs, we show how observations of protein burst distributions (repressed and unrepressed) can be used to completely determine the mRNA burst distribution. Assuming independent contributions from individual bursts, we derive analytical expressions connecting means and variances for burst and steady-state protein distributions. Finally, we validate our general analytical results by considering a specific reaction scheme involving regulation of protein bursts by small RNAs. For a range of parameters, we derive analytical expressions for regulated protein distributions that are validated using stochastic simulations. The analytical results obtained in this work can thus serve as useful inputs for a broad range of studies focusing on stochasticity in gene expression

Impaired CK1 delta activity attenuates SV40-induced cellular transformation in vitro and mouse mammary carcinogenesis in vivo.

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Heidrun Hirner

Full Text Available Simian virus 40 (SV40 is a powerful tool to study cellular transformation in vitro, as well as tumor development and progression in vivo. Various cellular kinases, among them members of the CK1 family, play an important role in modulating the transforming activity of SV40, including the transforming activity of T-Ag, the major transforming protein of SV40, itself. Here we characterized the effects of mutant CK1δ variants with impaired kinase activity on SV40-induced cell transformation in vitro, and on SV40-induced mammary carcinogenesis in vivo in a transgenic/bi-transgenic mouse model. CK1δ mutants exhibited a reduced kinase activity compared to wtCK1δ in in vitro kinase assays. Molecular modeling studies suggested that mutation N172D, located within the substrate binding region, is mainly responsible for impaired mutCK1δ activity. When stably over-expressed in maximal transformed SV-52 cells, CK1δ mutants induced reversion to a minimal transformed phenotype by dominant-negative interference with endogenous wtCK1δ. To characterize the effects of CK1δ on SV40-induced mammary carcinogenesis, we generated transgenic mice expressing mutant CK1δ under the control of the whey acidic protein (WAP gene promoter, and crossed them with SV40 transgenic WAP-T-antigen (WAP-T mice. Both WAP-T mice as well as WAP-mutCK1δ/WAP-T bi-transgenic mice developed breast cancer. However, tumor incidence was lower and life span was significantly longer in WAP-mutCK1δ/WAP-T bi-transgenic animals. The reduced CK1δ activity did not affect early lesion formation during tumorigenesis, suggesting that impaired CK1δ activity reduces the probability for outgrowth of in situ carcinomas to invasive carcinomas. The different tumorigenic potential of SV40 in WAP-T and WAP-mutCK1δ/WAP-T tumors was also reflected by a significantly different expression of various genes known to be involved in tumor progression, specifically of those involved in wnt-signaling and DNA

Effect of preoperative oral S-1 combined with regional intra-arterial chemotherapy on malignant molecule expression in locally advanced unresectable gastric cancer tissue

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Lei Liu

2016-11-01

Full Text Available Objective: To study the effect of preoperative oral S-1 combined with regional intra-arterial chemotherapy on malignant molecule expression in locally advanced unresectable gastric cancer tissue. Methods: A total of 144 patients with locally advanced gastric cancer receiving surgical resection after neoadjuvant chemotherapy in our hospital between May 2012 and August 2015 were selected and randomly divided into experimental group who received preoperative oral S-1 combined with regional intra-arterial chemotherapy and control group who received preoperative intravenous systemic chemotherapy. The levels of serum tumor markers were determined after chemotherapy, and the expression levels of tumor suppressor genes and cell cycle-related molecules in tumor tissue were determined after surgical resection. Results: After neoadjuvant chemotherapy, the serum G-17, TK-1, CEA, CA19-9, CA12-5, CA72-4 and CK, CK-MB, ALT, AST levels of experimental group were significantly lower than those of control group; after surgical resection, the p16, p27, PTEN and TXNIP mRNA levels in tumor tissue of experimental group were significantly higher than those of control group while CyclinB2, CyclinD1, CyclinE, CDK1 and CDK2 mRNA levels were significantly lower than those of control group. Conclusions: Preoperative oral S-1 combined with regional intra-arterial chemotherapy can more effectively kill gastric cancer cells, reduce tumor load, inhibit cell cycle and promote cell apoptosis.

Profiles of mRNA expression of genes related to sex differentiation of the gonads in the chicken embryo.

Science.gov (United States)

Yamamoto, I; Tsukada, A; Saito, N; Shimada, K

2003-09-01

Sex is determined genetically in birds. The homogametic sex is male (ZZ), whereas the heterogametic sex is female (ZW). According to the genetic sex, gonads develop into testes or ovary. In this study, we performed experiments to reveal mRNA expression patterns in the gonad between d 5.5 and 8.5 of incubation and examined a possible role of Dss-Ahc critical region on the X chromosome 1 (Dax1), Steroidogenic factor 1 (Sf1), P450aromatase (P450arom), Estrogen receptor alpha (ER alpha), doublesex and mab3 related transcription factor 1 (Dmrt1), Sry-related HMG box gene 9 (Sox9), Gata binding protein 4 (Gata4), and anti-müllerian hormone (Amh) in sex differentiation in chicken embryonic gonads using RNase protection assay. In embryonic chicken gonads, Dax1 mRNA was expressed in both sexes but was higher in females than in males at d 6.5 and 7.5 of incubation. The Sf1 mRNA was expressed in both sexes, but it was expressed more in males at d 5.5 than in females but more in females than in males at d 7.5 and 8.5 of incubation. The P450arom mRNA was expressed only in female gonads from d 5.5 of incubation. The ER alpha mRNA was expressed in both sexes, but it did not show a sex difference. On the other hand, the Dmrt1 mRNA was expressed in both sexes, but it showed a male-specific expression pattern. The male-specific expression pattern was observed in Sox9 mRNA, but it was not expressed in female gonads. The Gata4 mRNA was expressed in both sexes, and sex differences were not revealed throughout the observational period. Amh mRNA was expressed in both sexes, but it had male-specific mRNA expression pattern at d 6.5 to 8.5 of incubation. These results indicate that Dax1, Sf1, and P450arom have possible roles in ovary formation, whereas Dmrt1, Sox9, and Amh are related to testis formation in differentiating chicken gonads at d 5.5 to 8.5 of incubation.

Nitric oxide signaling pathway regulates potassium chloride cotransporter-1 mRNA expression in vascular smooth muscle cells.

Science.gov (United States)

Di Fulvio, M; Lauf, P K; Adragna, N C

2001-11-30

Rat vascular smooth muscle cells (VSMCs) express at least two mRNAs for K-Cl cotransporters (KCC): KCC1 and KCC3. cGMP-dependent protein kinase I regulates KCC3 mRNA expression in these cells. Here, we show evidence implicating the nitric oxide (NO)/cGMP signaling pathway in the expression of KCC1 mRNA, considered to be the major cell volume regulator. VSMCs, expressing soluble guanylyl cyclase (sGC) and PKG-I isoforms showed a time- and concentration-dependent increase in KCC1 mRNA levels after treatment with sodium nitroprusside as demonstrated by semiquantitative RT-PCR. sGC-dependent regulation of KCC1 mRNA expression was confirmed using YC-1, a NO-independent sGC stimulator. The sGC inhibitor LY83583 blocked the effects of sodium nitroprusside and YC-1. Moreover, 8-Br-cGMP increased KCC1 mRNA expression in a concentration- and time-dependent fashion. The 8-Br-cGMP effect was partially blocked by KT5823 but not by actinomycin D. However, actinomycin D and cycloheximide increased basal KCC1 mRNA in an additive manner, suggesting different mechanisms of action for both drugs. These findings suggest that in VSMCs, the NO/cGMP-signaling pathway participates in KCC1 mRNA regulation at the post-transcriptional level.

Local IGFBP-3 mRNA expression, apoptosis and risk of colorectal adenomas

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Omofoye Oluwaseun

2008-05-01

Full Text Available Abstract Background IGF binding protein-3 (IGFBP-3 regulates the bioavailability of insulin-like growth factors I and II, and has both anti-proliferative and pro-apoptotic properties. Elevated plasma IGFBP-3 has been associated with reduced risk of colorectal cancer (CRC, but the role of tissue IGFBP-3 is not well defined. We evaluated the association between tissue or plasma IGFBP-3 and risk of colorectal adenomas or low apoptosis. Methods Subjects were consenting patients who underwent a clinically indicated colonoscopy at UNC Hospitals and provided information on diet and lifestyle. IGFBP-3 mRNA in normal colon was assessed by real time RT-PCR. Plasma IGFBP-3 was measured by ELISA and apoptosis was determined by morphology on H & E slides. Logistic regression was used to compute odds ratio (OR and 95% confidence intervals. Results We observed a modest correlation between plasma IGFBP-3 and tissue IGFBP-3 expression (p = 0.007. There was no significant association between plasma IGFBP-3 and adenomas or apoptosis. Tissue IGFBP-3 mRNA expression was significantly lower in cases than controls. Subjects in the lowest three quartiles of tissue IGFBP-3 gene expression were more likely to have adenomas. Consistent with previous reports, low apoptosis was significantly associated with increased risk of adenomas (p = 0.003. Surprisingly, local IGFBP-3 mRNA expression was inversely associated with apoptosis. Conclusion Low expression of IGFBP-3 mRNA in normal colonic mucosa predicts increased risk of adenomas. Our findings suggest that local IGFBP-3 in the colon may directly increase adenoma risk but IGFBP-3 may act through a pathway other than apoptosis to influence adenoma risk.

Local IGFBP-3 mRNA expression, apoptosis and risk of colorectal adenomas

International Nuclear Information System (INIS)

Keku, Temitope O; Sandler, Robert S; Simmons, James G; Galanko, Joseph; Woosley, John T; Proffitt, Michelle; Omofoye, Oluwaseun; McDoom, Maya; Lund, Pauline K

2008-01-01

IGF binding protein-3 (IGFBP-3) regulates the bioavailability of insulin-like growth factors I and II, and has both anti-proliferative and pro-apoptotic properties. Elevated plasma IGFBP-3 has been associated with reduced risk of colorectal cancer (CRC), but the role of tissue IGFBP-3 is not well defined. We evaluated the association between tissue or plasma IGFBP-3 and risk of colorectal adenomas or low apoptosis. Subjects were consenting patients who underwent a clinically indicated colonoscopy at UNC Hospitals and provided information on diet and lifestyle. IGFBP-3 mRNA in normal colon was assessed by real time RT-PCR. Plasma IGFBP-3 was measured by ELISA and apoptosis was determined by morphology on H & E slides. Logistic regression was used to compute odds ratio (OR) and 95% confidence intervals. We observed a modest correlation between plasma IGFBP-3 and tissue IGFBP-3 expression (p = 0.007). There was no significant association between plasma IGFBP-3 and adenomas or apoptosis. Tissue IGFBP-3 mRNA expression was significantly lower in cases than controls. Subjects in the lowest three quartiles of tissue IGFBP-3 gene expression were more likely to have adenomas. Consistent with previous reports, low apoptosis was significantly associated with increased risk of adenomas (p = 0.003). Surprisingly, local IGFBP-3 mRNA expression was inversely associated with apoptosis. Low expression of IGFBP-3 mRNA in normal colonic mucosa predicts increased risk of adenomas. Our findings suggest that local IGFBP-3 in the colon may directly increase adenoma risk but IGFBP-3 may act through a pathway other than apoptosis to influence adenoma risk

Cloning and mRNA expression pattern analysis under low ...

African Journals Online (AJOL)

This research cloned endochitinase-antifreeze protein precursor (EAPP) gene of Dong-mu 70 rye (Secale cereale) by designing special primers according to Genbank's EAPP gene sequence, and analyzing the influence of low temperature stress on the expression of mRNA with RT-PCR. The results indicated that the ...

Downregulation of TIM-3 mRNA expression in peripheral blood mononuclear cells from patients with systemic lupus erythematosus

Energy Technology Data Exchange (ETDEWEB)

Cai, X.Z. [Central Laboratory, First Affiliated Hospital, China Medical University, Shenyang (China); Department of Immunology, College of Basic Medical Sciences, China Medical University, Shenyang (China); Huang, W.Y.; Qiao, Y.; Chen, Y.; Du, S.Y.; Chen, D.; Yu, S. [Central Laboratory, First Affiliated Hospital, China Medical University, Shenyang (China); Liu, N. [Department of Nephrology, First Affiliated Hospital, China Medical University, Shenyang (China); Dou, L.Y. [Central Laboratory, First Affiliated Hospital, China Medical University, Shenyang (China); Jiang, Y. [Central Laboratory, First Affiliated Hospital, China Medical University, Shenyang (China); Department of Immunology, College of Basic Medical Sciences, China Medical University, Shenyang (China); Department of Dermatology, First Affiliated Hospital, China Medical University, Shenyang (China)

2014-10-17

The T-cell immunoglobulin and mucin domain (TIM) family is associated with autoimmune diseases, but its expression level in the immune cells of systemic lupus erythematosus (SLE) patients is not known. The aim of this study was to investigate whether the expression of TIM-3 mRNA is associated with pathogenesis of SLE. Quantitative real-time reverse transcription-polymerase chain reaction analysis (qRT-PCR) was used to determine TIM-1, TIM-3, and TIM-4 mRNA expression in peripheral blood mononuclear cells (PBMCs) from 132 patients with SLE and 62 healthy controls. The PBMC surface protein expression of TIMs in PBMCs from 20 SLE patients and 15 healthy controls was assayed by flow cytometry. Only TIM-3 mRNA expression decreased significantly in SLE patients compared with healthy controls (P<0.001). No significant differences in TIM family protein expression were observed in leukocytes from SLE patients and healthy controls (P>0.05). SLE patients with lupus nephritis (LN) had a significantly lower expression of TIM-3 mRNA than those without LN (P=0.001). There was no significant difference in the expression of TIM-3 mRNA within different classes of LN (P>0.05). Correlation of TIM-3 mRNA expression with serum IgA was highly significant (r=0.425, P=0.004), but was weakly correlated with total serum protein (r{sub s}=0.283, P=0.049) and serum albumin (r{sub s}=0.297, P=0.047). TIM-3 mRNA expression was weakly correlated with the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI; r{sub s}=-0.272, P=0.032). Our results suggest that below-normal expression of TIM-3 mRNA in PBMC may be involved in the pathogenesis of SLE.

Downregulation of TIM-3 mRNA expression in peripheral blood mononuclear cells from patients with systemic lupus erythematosus

International Nuclear Information System (INIS)

Cai, X.Z.; Huang, W.Y.; Qiao, Y.; Chen, Y.; Du, S.Y.; Chen, D.; Yu, S.; Liu, N.; Dou, L.Y.; Jiang, Y.

2014-01-01

The T-cell immunoglobulin and mucin domain (TIM) family is associated with autoimmune diseases, but its expression level in the immune cells of systemic lupus erythematosus (SLE) patients is not known. The aim of this study was to investigate whether the expression of TIM-3 mRNA is associated with pathogenesis of SLE. Quantitative real-time reverse transcription-polymerase chain reaction analysis (qRT-PCR) was used to determine TIM-1, TIM-3, and TIM-4 mRNA expression in peripheral blood mononuclear cells (PBMCs) from 132 patients with SLE and 62 healthy controls. The PBMC surface protein expression of TIMs in PBMCs from 20 SLE patients and 15 healthy controls was assayed by flow cytometry. Only TIM-3 mRNA expression decreased significantly in SLE patients compared with healthy controls (P<0.001). No significant differences in TIM family protein expression were observed in leukocytes from SLE patients and healthy controls (P>0.05). SLE patients with lupus nephritis (LN) had a significantly lower expression of TIM-3 mRNA than those without LN (P=0.001). There was no significant difference in the expression of TIM-3 mRNA within different classes of LN (P>0.05). Correlation of TIM-3 mRNA expression with serum IgA was highly significant (r=0.425, P=0.004), but was weakly correlated with total serum protein (r s =0.283, P=0.049) and serum albumin (r s =0.297, P=0.047). TIM-3 mRNA expression was weakly correlated with the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI; r s =-0.272, P=0.032). Our results suggest that below-normal expression of TIM-3 mRNA in PBMC may be involved in the pathogenesis of SLE

Visfatin mRNA expression in human subcutaneous adipose tissue is regulated by exercise

DEFF Research Database (Denmark)

Frydelund-Larsen, Lone; Åkerström, Thorbjörn; Nielsen, Søren

2006-01-01

in abdominal subcutaneous adipose tissue and skeletal muscle biopsies obtained from healthy young men at time points 0, 3, 4.5, 6, 9, and 24 h in relation to either 3 h of ergometer cycle exercise at 60% of Vo(2 max) or rest. Adipose tissue visfatin mRNA expression increased threefold at the time points 3, 4......Visfatin [pre-beta-cell colony-enhancing factor (PBEF)] is a novel adipokine that is produced by adipose tissue, skeletal muscle, and liver and has insulin-mimetic actions. Regular exercise enhances insulin sensitivity. In the present study, we therefore examined visfatin mRNA expression.......5, and 6 h in response to exercise (n = 8) compared with preexercise samples and compared with the resting control group (n = 7, P = 0.001). Visfatin mRNA expression in skeletal muscle was not influenced by exercise. The exercise-induced increase in adipose tissue visfatin was, however, not accompanied...

Association of chemerin mRNA expression in human epicardial adipose tissue with coronary atherosclerosis

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Wang Linjie

2011-10-01

Full Text Available Abstract Background Growing evidence suggests that epicardial adipose tissue (EAT may play a key role in the pathogenesis and development of coronary artery disease (CAD by producing several inflammatory adipokines. Chemerin, a novel adipokine, has been reported to be involved in regulating immune responses and glucolipid metabolism. Given these properties, chemerin may provide an interesting link between obesity, inflammation and atherosclerosis. In this study, we sought to determine the relationship of chemerin expression in EAT and the severity of coronary atherosclerosis in Han Chinese patients. Methods Serums and adipose tissue biopsies (epicardial and thoracic subcutaneous were obtained from CAD (n = 37 and NCAD (n = 16 patients undergoing elective cardiac surgery. Gensini score was used to assess the severity of CAD. Serum levels of chemerin, adiponectin and insulin were measured by ELISA. Chemerin protein expression in adipose tissue was detected by immunohistochemistry. The mRNA levels of chemerin, chemR23, adiponectin and TNF-alpha in adipose tissue were detected by RT-PCR. Results We found that EAT of CAD group showed significantly higher levels of chemerin and TNF-alpha mRNA, and significantly lower level of adiponectin mRNA than that of NCAD patients. In CAD group, significantly higher levels of chemerin mRNA and protein were observed in EAT than in paired subcutaneous adipose tissue (SAT, whereas such significant difference was not found in NCAD group. Chemerin mRNA expression in EAT was positively correlated with Gensini score (r = 0.365, P P P P P P P > 0.05. Conclusions The expressions of chemerin mRNA and protein are significantly higher in EAT from patients with CAD in Han Chinese patients. Furthermore, the severity of coronary atherosclerosis is positive correlated with the level of chemerin mRNA in EAT rather than its circulating level.

Expression of Panton-Valentine leukocidin mRNA among Staphylococcus aureus isolates associates with specific clinical presentations.

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Fangyou Yu

Full Text Available Panton-Valentine leukocidin (PVL; gene designation lukF/S-PV is likely an important virulence factor for Staphylococcus aureus (S. aureus, as qualitative expression of the protein correlates with severity for specific clinical presentations, including skin and soft tissue infections (SSTIs. Development of genetic approaches for risk-assessment of patients with S. aureus infections may prove clinically useful, and whether lukF/S-PV gene expression correlates with specific clinical presentations for S. aureus has been largely unexplored. In the present study, we quantified lukS-PV mRNA among 96 S. aureus isolates to determine whether expression levels correlated with specific clinical presentations in adults and children. Expression level of lukS-PV mRNA among isolates from skin and soft tissue infections (SSTIs was significantly greater than among isolates from blood stream infection (BSIs, and expression level of lukS-PV mRNA among BSI isolates from children was significantly greater than for BSI isolates among adults. Moreover, expression level of lukS-PV mRNA among community-acquired (CA isolates was significantly greater than for hospital-acquired (HA isolates. These data justify additional studies to determine the potential clinical utility for lukS-PV mRNA quantification as a predictive tool for severity of S. aureus infection.

Up-Regulation of CYP2C19 Expression by BuChang NaoXinTong via PXR Activation in HepG2 Cells.

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Hong Sun

Full Text Available Cytochrome P450 2C19 (CYP2C19 is an important drug-metabolizing enzyme (DME, which is responsible for the biotransformation of several kinds of drugs such as proton pump inhibitors, platelet aggregation inhibitors and antidepressants. Previous studies showed that Buchang NaoXinTong capsules (NXT increased the CYP2C19 metabolic activity in vitro and enhanced the antiplatelet effect of clopidogrel in vivo. However, the underlying molecular mechanism remained unclear. In the present study, we examined whether Pregnane X receptor (PXR plays a role in NXT-mediated regulation of CYP2C19 expression.We applied luciferase assays, real-time quantitative PCR (qPCR, Western blotting and cell-based analysis of metabolic activity experiments to investigate the NXT regulatory effects on the CYP2C19 promoter activity, the mRNA/ protein expression and the metabolic activity.Our results demonstrated that NXT significantly increased the CYP2C19 promoter activity when co-transfected with PXR in HepG2 cells. Mutations in PXR responsive element abolished the NXT inductive effects on the CYP2C19 promoter transcription. Additionally, NXT incubation (150 and 250μg/mL also markedly up-regulated endogenous CYP2C19 mRNA and protein levels in PXR-transfected HepG2 cells. Correspondingly, NXT leaded to a significant enhancement of the CYP2C19 catalytic activity in PXR-transfected HepG2 cells.In summary, this is the first study to suggest that NXT could induce CYP2C19 expression via PXR activation.

Hepatocellular carcinoma: microstructure and expression features of hepatocyte marker, alphafetoprotein, cytokeratins 7 and 20

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V. A. Tumanskiy

2014-04-01

Full Text Available V. A. Tumanskiy, M. D. Zubko Aims. In order to determine the microscopic features and expression level of HepРar-1, AFP, CK7, CK20 by the cells of hepatocellular carcinoma histopathological, histochemical and immunohistochemical study of 53 liver biopsies from patients with hepatocellular carcinoma was conducted. Methods and results. The photo digital morphometry was used and area of immunopositive cells was calculated. In 92.45% of cases the cytoplasmic expression of HepPar was established. The area of immunopositive cells was 49.35±25.45%. 81.13% of patients with hepatocellular carcinoma had the cytoplasmic and nuclear expression of α- fetoprotein. The area of AFP-immunopositive cells was 37.25±15.47% on the average. There was a direct weak connection between the expression level of HepPar-1 and AFP. CK7 expression was noted in 37.74% of cases and CK20 expression was noted in 30.13% of cases of HCC. CK7+ and CK20+ cells are distributed in the form of focal clusters and take area of 21.08±5.19% and 29.35±17.31% respectively. There was direct medium strength connection between the expression level of AFP and СК7, AFP and СК20 by the tumor cells. There was strong direct correlation between the level of expression of HepPar-1 and CК7, HepPar-1 and CК20.

Effects of exogenous ATM gene on mRNA expression of human telomerase reverse transcriptase in AT cells induced by irradiation

International Nuclear Information System (INIS)

Sheng Fangjun; Cao Jianping; Luo Jialin; Zhu Wei; Liu Fenju; Feng Shuang; Song Jianyuan; Li Chong

2005-01-01

The study is to observe effects of exogenous ATM gene on mRNA expression of hTERT (human telomerase reverse transcriptase) in fibroblast cells (AT5BIVA cells) from skin of Ataxia-telangiectasia (AT) patients and to study the regulation of ATM to hTERT. Using reverse transcription polymerase chain reaction (RT-PCR), mRNA expression of hTERT in AT, PEBS7-AT, ATM + -AT and GM cells irradiated with 0 and 3 Gy of 60 Co γ-rays were examined respectively. The difference of the mRNA expression of hTERT among AT, PEBS7-AT, ATM + -AT and GM cells were analyzed. Difference of the mRNA expression of hTERT between 0 Gy and 3 Gy groups was analyzed, too. The results showed that the mRNA expression of hTERT in GM cells was negative, but positive mRNA expression of hTERT in AT cells. The mRNA expression of hTERT in ATM + -AT cells decreased significantly (p 60 Co γ-rays, the mRNA expression of hTERT in GM cells was positive, and that in AT, PEBS7-AT, ATM + -AT cells was increased (p + -AT cells was lower than that in AT and PEBS7-AT cells respectively (p<0.05). It is postulated that exogenous ATM is able to downregulate the mRNA expression of hTERT in AT cells, ionizing radiation can induce the mRNA expression of hTERT in cells and telomerase anticipates the repair of damaged DNA. (authors)

Expression and autoregulation of transforming growth factor beta receptor mRNA in small-cell lung cancer cell lines

DEFF Research Database (Denmark)

Nørgaard, P; Spang-Thomsen, M; Poulsen, H S

1996-01-01

In small-cell lung cancer cell lines resistance to growth inhibition by transforming growth factor (TGF)-beta 1, was previously shown to correlate with lack of TGF-beta receptor I (RI) and II (RII) proteins. To further investigate the role of these receptors, the expression of mRNA for RI, RII...... and beta-glycan (RIII) was examined. The results showed that loss of RII mRNA correlated with TGF-beta 1 resistance. In contrast, RI-and beta-glycan mRNA was expressed by all cell lines, including those lacking expression of these proteins. According to Southern blot analysis, the loss of type II m......RNA was not due to gross structural changes in the gene. The effect of TGF-beta 1 on expression of TGF-beta receptor mRNA (receptor autoregulation) was examined by quantitative Northern blotting in four cell lines with different expression of TGF-beta receptor proteins. In two cell lines expressing all three TGF...

Group II intron inhibits conjugative relaxase expression in bacteria by mRNA targeting

Science.gov (United States)

Piazza, Carol Lyn; Smith, Dorie

2018-01-01

Group II introns are mobile ribozymes that are rare in bacterial genomes, often cohabiting with various mobile elements, and seldom interrupting housekeeping genes. What accounts for this distribution has not been well understood. Here, we demonstrate that Ll.LtrB, the group II intron residing in a relaxase gene on a conjugative plasmid from Lactococcus lactis, inhibits its host gene expression and restrains the naturally cohabiting mobile element from conjugative horizontal transfer. We show that reduction in gene expression is mainly at the mRNA level, and results from the interaction between exon-binding sequences (EBSs) in the intron and intron-binding sequences (IBSs) in the mRNA. The spliced intron targets the relaxase mRNA and reopens ligated exons, causing major mRNA loss. Taken together, this study provides an explanation for the distribution and paucity of group II introns in bacteria, and suggests a potential force for those introns to evolve into spliceosomal introns. PMID:29905149

Sheep oocyte expresses leptin and functional leptin receptor mRNA

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Seyyed Jalil Taheri

2016-09-01

Conclusions: The result of present study reveals that leptin and its functional receptor (Ob-Rb mRNA are expressed in sheep oocyte and further studies should investigate the role(s of leptin on sheep oocyte physiology and embryo development.

Expression, purification and crystallization of the catalytic subunit of protein kinase CK2 from Zea mays

DEFF Research Database (Denmark)

Guerra, B; Niefind, K; Pinna, L A

1998-01-01

The catalytic (alpha) subunit of protein kinase CK2 (CK2alpha) was originally cloned and overexpressed in the Escherichia coli strain pT7-7/BL21(DE3). The protein has been purified to homogeneity and crystallized. The crystals belong to the monoclinic space group C2, they have unit-cell parameter...

Integrating microRNA and mRNA expression profiles in response to radiation-induced injury in rat lung

International Nuclear Information System (INIS)

Xie, Ling; Zhou, Jundong; Zhang, Shuyu; Chen, Qing; Lai, Rensheng; Ding, Weiqun; Song, ChuanJun; Meng, XingJun; Wu, Jinchang

2014-01-01

Exposure to radiation provokes cellular responses, which are likely regulated by gene expression networks. MicroRNAs are small non-coding RNAs, which regulate gene expression by promoting mRNA degradation or inhibiting protein translation. The expression patterns of both mRNA and miRNA during the radiation-induced lung injury (RILI) remain less characterized and the role of miRNAs in the regulation of this process has not been studied. The present study sought to evaluate miRNA and mRNA expression profiles in the rat lung after irradiation. Male Wistar rats were subjected to single dose irradiation with 20 Gy using 6 MV x-rays to the right lung. (A dose rate of 5 Gy/min was applied). Rats were sacrificed at 3, 12 and 26 weeks after irradiation, and morphological changes in the lung were examined by haematoxylin and eosin. The miRNA and mRNA expression profiles were evaluated by microarrays and followed by quantitative RT-PCR analysis. A cDNA microarray analysis found 2183 transcripts being up-regulated and 2917 transcripts down-regulated (P ≤ 0.05, ≥2.0 fold change) in the lung tissues after irradiation. Likewise, a miRNAs microarray analysis indicated 15 miRNA species being up-regulated and 8 down-regulated (P ≤ 0.05). Subsequent bioinformatics anal -yses of the differentially expressed mRNA and miRNAs revealed that alterations in mRNA expression following irradiation were negatively correlated with miRNAs expression. Our results provide evidence indicating that irradiation induces alterations of mRNA and miRNA expression in rat lung and that there is a negative correlation of mRNA and miRNA expression levels after irradiation. These findings significantly advance our understanding of the regulatory mechanisms underlying the pathophysiology of radiation-induced lung injury. In summary, RILI does not develop gradually in a linear process. In fact, different cell types interact via cytokines in a very complex network. Furthermore, this study suggests that

Aberrant Expression of TNF-α and TGF-β1 mRNA in Spontaneous Abortion

Institute of Scientific and Technical Information of China (English)

Ji-fen HU; Hong-chu BAO; Feng-chuan ZHU; Cai-ling YOU

2004-01-01

Objective To investigate the aberrant expressions of TNF-α and TGF-β1 in peripheral blood mononuclear cells (PBMCs) and placental tissues in patients with early spontaneous abortionMethods Using the technique of semi-quantitative reverse transcript-polymerase chain reaction (RT-PCR), TNF-α mRNA and TGF-β1 mRNA in PBMCs were measured in spontaneous abortion group (30 cases), normal pregnancy group (25 cases) and nonpregnant group (25 cases). The expressive intension of TNF-α protein and TGF-β1 protein in placental tissues was also identified by immunohistochemistry.Results Both levels of TNF-α mRNA and TGF-β1 mRNA expressed in PBMCs were significantly different between the three groups respectively (P＜0. 05). Levels of TNF-α in syncytiotrophoblastic and cytotrophoblastic cells of the two aborted groups were substantially higher than those of the non-pregnant group (P＜0. 01), but the levels of TGF-β1 in syncytiotrophoblastic cells of the two aborted groups were markedly lower than those of the non-pregnant group (P＜0. 01).Conclusion There is potential relation between TGF-β1 at the fetomaternal interface and spontaneous abortion. TGF-β1 may contribute to the maintenance of pregnancy,and low-level expression of TGF-β1 may be associated with pregnancy failure.

CX4945 suppresses the growth of castration-resistant prostate cancer cells by reducing AR-V7 expression.

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Deng, Chuangzhong; Chen, Jieping; Guo, Shengjie; Wang, Yanjun; Zhou, Qianghua; Li, Zaishang; Yang, Xingping; Yu, Xingsu; Zhang, Zhenfeng; Zhou, Fangjian; Han, Hui; Yao, Kai

2017-08-01

The aberrant expression of casein kinase 2 (CK2) has been reported to be involved in the tumorigenesis and progression of prostate cancer. The inhibition of CK2 activity represses androgen-dependent prostate cancer cells by attenuating the androgen receptor (AR) signaling pathway. In this study, we examined the effect of CK2 inhibition in castration-resistant prostate cancer (CRPC) cells, in which AR variants (ARVs) play a predominant role. A newly synthetic CK2 selective inhibitor CX4945 was utilized to study the effect of CK2 inhibition in CRPC cells by CCK8 assay and colony formation assay. Protein and mRNA levels of full-length AR (AR-FL) and AR-V7 were determined by qPCR and western blot, respectively. The nuclear translocation of p50 and p65 was assessed to reflect the activity of the NF-κB pathway. CX4945 reduced the proliferation of CRPC cells in a dose-dependent and time-dependent manner. AR-V7 rather than AR-FL was downregulated by CX4945 in both the mRNA and protein level. Furthermore, CX4945 could restore the sensitivity of CRPC cells to bicalutamide. The analysis of possible mechanisms demonstrated that the inhibition of CK2 diminished the phosphorylation of p65 at ser529 and thus attenuated the activity of the NF-κB pathway. The inhibition of CK2 by CX4945 can repress the viability of CRPC cells and restore their sensitivity to anti-androgen therapy by suppressing AR-V7. This finding presents a potential option for the treatment of prostate cancer, especially CRPC.

Development of a high-throughput screening-compatible assay to identify inhibitors of the CK2alpha/CK2beta interaction

DEFF Research Database (Denmark)

Hochscherf, Jennifer; Lindenblatt, Dirk; Steinkrueger, Michaela

2015-01-01

Increased activity of protein kinase CK2 is associated with various types of cancer, neurodegenerative diseases, and chronic inflammation. In the search for CK2 inhibitors, attention has expanded toward compounds disturbing the interaction between CK2alpha and CK2beta in addition to established a...

CD133 expression in well-differentiated pancreatic neuroendocrine tumors: a potential predictor of progressive clinical courses.

Science.gov (United States)

Sakai, Yasuhiro; Hong, Seung-Mo; An, Soyeon; Kim, Joo Young; Corbeil, Denis; Karbanová, Jana; Otani, Kyoko; Fujikura, Kohei; Song, Ki-Byung; Kim, Song Cheol; Akita, Masayuki; Nanno, Yoshihide; Toyama, Hirochika; Fukumoto, Takumi; Ku, Yonson; Hirose, Takanori; Itoh, Tomoo; Zen, Yoh

2017-03-01

The present study aimed to elucidate whether the stemness molecule, CD133, is expressed in well-differentiated pancreatic neuroendocrine tumors (PanNETs; World Health Organization grades 1 and 2) and establish its clinical relevance using 2 separate cohorts. In the first series (n = 178) in which tissue microarrays were available, immunohistochemistry revealed that CD133 was expressed in 14 cases (8%). CD133+ PanNETs had higher TNM stages (P < .01), more frequent lymphovascular invasion (P = .01), and higher recurrence rates (P = .01). In the second cohort (n = 56), the expression of CD133 and CK19 was examined in whole tissue sections. CD133 and CK19 were positive in 10 (18%) and 36 (64%) cases, respectively. CD133 expression correlated with higher pT scores (P < .01), the presence of microscopic venous infiltration (P = .03), and shorter disease-free periods (P < .01). When cases were divided into grade 1 and 2 neoplasms, patients with CD133+ PanNET continued to have shorter disease-free periods than did those with CD133- tumors in both groups (P < .01 and P = .02, respectively). Although CK19+ cases had shorter disease-free periods than did CK19- cases in the whole cohort (P = .02), this difference was less apparent in subanalyses of grade 1 and 2 cases. CD133 expression also appeared to be an independent predictive factor for tumor recurrence in a multivariate analysis (P = .018). The CD133 phenotype was identical between primary and metastatic foci in 17 of 18 cases from which tissues of metastatic deposits were available. In conclusion, the combination of CD133 phenotyping and World Health Organization grading may assist in stratifying patients in terms of the risk of progressive clinical courses. Copyright © 2016 Elsevier Inc. All rights reserved.

BAY11 enhances OCT4 synthetic mRNA expression in adult human skin cells.

Science.gov (United States)

Awe, Jason P; Crespo, Agustin Vega; Li, You; Kiledjian, Megerditch; Byrne, James A

2013-02-06

The OCT4 transcription factor is involved in many cellular processes, including development, reprogramming, maintaining pluripotency and differentiation. Synthetic OCT4 mRNA was recently used (in conjunction with other reprogramming factors) to generate human induced pluripotent stem cells. Here, we discovered that BAY 11-7082 (BAY11), at least partially through an NF-κB-inhibition based mechanism, could significantly increase the expression of OCT4 following transfection of synthetic mRNA (synRNA) into adult human skin cells. We tested various chemical and molecular small molecules on their ability to suppress the innate immune response seen upon synthetic mRNA transfection. Three molecules - B18R, BX795, and BAY11 - were used in immunocytochemical and proliferation-based assays. We also utilized global transcriptional meta-analysis coupled with quantitative PCR to identify relative gene expression downstream of OCT4. We found that human skin cells cultured in the presence of BAY11 resulted in reproducible increased expression of OCT4 that did not inhibit normal cell proliferation. The increased levels of OCT4 resulted in significantly increased expression of genes downstream of OCT4, including the previously identified SPP1, DUSP4 and GADD45G, suggesting the expressed OCT4 was functional. We also discovered a novel OCT4 putative downstream target gene SLC16A9 which demonstrated significantly increased expression following elevation of OCT4 levels. For the first time we have shown that small molecule-based stabilization of synthetic mRNA expression can be achieved with use of BAY11. This small molecule-based inhibition of innate immune responses and subsequent robust expression of transfected synthetic mRNAs may have multiple applications for future cell-based research and therapeutics.

Annexin-1 Mediates Microglial Activation and Migration via the CK2 Pathway during Oxygen–Glucose Deprivation/Reperfusion

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Shuangxi Liu

2016-10-01

Full Text Available Annexin-1 (ANXA1 has shown neuroprotective effects and microglia play significant roles during central nervous system injury, yet the underlying mechanisms remain unclear. This study sought to determine whether ANXA1 regulates microglial response to oxygen–glucose deprivation/reperfusion (OGD/R treatment and to clarify the downstream molecular mechanism. In rat hippocampal slices, OGD/R treatment enhanced the ANXA1 expression in neuron, the formyl peptide receptor (FPRs expression in microglia, and the microglial activation in the CA1 region (cornu ammonis 1. These effects were reversed by the FPRs antagonist Boc1. The cell membrane currents amplitude of BV-2 microglia (the microglial like cell-line was increased when treated with Ac2-26, the N-terminal peptide of ANXA1. Ac2-26 treatment enhanced BV-2 microglial migration whereas Boc1 treatment inhibited the migration. In BV-2 microglia, both the expression of the CK2 target phosphorylated α-E-catenin and the binding of casein kinase II (CK2 with α-E-catenin were elevated by Ac2-26, these effects were counteracted by the CK2 inhibitor TBB and small interfering (si RNA directed against transcripts of CK2 and FPRs. Moreover, both TBB and siRNA-mediated inhibition of CK2 blocked Ac2-26-mediated BV-2 microglia migration. Our findings indicate that ANXA1 promotes microglial activation and migration during OGD/R via FPRs, and CK2 target α-E-catenin phosphorylation is involved in this process.

Peripheral mononuclear cell resistin mRNA expression is increased in type 2 diabetic women.

Science.gov (United States)

Tsiotra, Panayoula C; Tsigos, Constantine; Anastasiou, Eleni; Yfanti, Eleni; Boutati, Eleni; Souvatzoglou, Emmanouil; Kyrou, Ioannis; Raptis, Sotirios A

2008-01-01

Resistin has been shown to cause insulin resistance and to impair glucose tolerance in rodents, but in humans its physiological role still remains elusive. The aim of this study was to examine whether resistin mRNA expression in human peripheral mononuclear cells (PBMCs) and its corresponding plasma levels are altered in type 2 diabetes. Resistin mRNA levels were easily detectable in human PBMC, and found to be higher in DM2 compared to healthy women (P = .05). Similarly, mononuclear mRNA levels of the proinflammatory cytokines IL-1beta, TNF-alpha, and IL-6 were all significantly higher in DM2 compared to control women (P DM2 women (P = .051), and overall, they correlated significantly with BMI (r = 0.406, P = .010) and waist circumference (r = 0.516, P = .003), but not with fasting insulin levels or HOMA-IR. Resistin mRNA expression is increased in PBMC from DM2 women, together with increased expression of the inflammatory cytokines IL-1beta, TNF-alpha, and IL-6, independent of obesity. These results suggest that resistin and cytokines might contribute to the low-grade inflammation and the increased atherogenic risk observed in these patients.

Expressions of interferon-inducible genes IFIT1 and IFIT4 mRNA in PBMCs of patients with systemic lupus erythematosus

International Nuclear Information System (INIS)

Liu Chunyan; Chen Xingguo; Wang Zizheng

2009-01-01

To investigate the expression levels of interferon-inducible genes (IFIT1, IFIT4) in the peripheral blood mononuclear cells (PBMCs) of patients with systemic lupus erythematosus (SLE), and the relations between these genes expression levels and disease activity, the expression levels of IFIT1 and IFIT4 mRNA in the 95 patients with SLE and 48 normal controls were detected by Sybr green dye based real-time quantitative PCR method, and these genes expression levels were compared with anti-double strand DNA antibody. The associations between the expression levels of IFIT1, IFIT4 mRNA, anti-double strand DNA antibody and SLEDAI scores in patients with SLE were analyzed. The results showed that the expression levels of IFIT1, IFIT4 mRNA in the SLE patients were significantly higher than those of the normal controls (P<0.01). The expression levels of IFIT1, IFIT4 mRNA in the active SLE patients were higher than those of the inactive SLE patients (P<0.05). The real time expression levels of IFIT1 and IFIT4 mRNA showed positive correlations with each other (P<0.05) in patients with SLE. There was positively correlation between the expression levels of IFIT1, IFIT4 mRNA and the anti-double strand DNA antibody (P<0.05). The expression levels of IFIT1, IFIT4 mRNA in patients with SLE were significantly higher than those of the normal controls, and positively associated with SLEDAI scores, so they were helpful in evaluating SLE disease activity and severity. To inhibit the expressions of IFIT1, IFIT4 mRNA may provide a novel target for SLE treatment. (authors)

mRNA expression of genes regulating lipid metabolism in ringed seals (Pusa hispida) from differently polluted areas

International Nuclear Information System (INIS)

Castelli, Martina Galatea; Rusten, Marte; Goksøyr, Anders; Routti, Heli

2014-01-01

Highlights: •Genes regulating lipid metabolism were studied in ringed seals. •We compared highly contaminated Baltic seals and less contaminated Svalbard seals. •mRNA expression of hepatic PPARγ was higher in the Baltic seals. •mRNA expression of adipose PPARγ target genes was higher in the Baltic seals. •Contaminant exposure may affect lipid metabolism in the Baltic ringed seals. -- Abstract: There is a growing concern about the ability of persistent organic pollutants (POPs) to influence lipid metabolism. Although POPs are found at high concentrations in some populations of marine mammals, for example in the ringed seal (Pusa hispida) from the Baltic Sea, little is known about the effects of POPs on their lipid metabolism. An optimal regulation of lipid metabolism is crucial for ringed seals during the fasting/molting season. This is a physiologically stressful period, during which they rely on the energy stored in their fat reserves. The mRNA expression levels for seven genes involved in lipid metabolism were analyzed in liver and/or blubber tissue from molting ringed seals from the polluted Baltic Sea and a less polluted reference location, Svalbard (Norway). mRNA expression of genes encoding peroxisome proliferator-activated receptors (PPAR) α and γ and their target genes acyl-coenzyme A oxidase 1 (ACOX1) and cluster of differentiation 36 (CD36) were analyzed in liver. mRNA expression level of genes encoding PPARβ, PPARγ and their target genes encoding fatty acid binding protein 4 (FABP4) and adiponectin (ADIPOQ) were measured in inner and middle blubber layers. In addition, we evaluated the influence of molting status on hepatic mRNA expression of genes encoding PPARs and their target genes in ringed seals from Svalbard. Our results show higher mRNA expression of genes encoding hepatic PPARγ and adipose PPARβ, FABP4, and ADIPOQ in the Baltic seals compared to the Svalbard seals. A positive relationship between mRNA expressions of genes

mRNA expression of genes regulating lipid metabolism in ringed seals (Pusa hispida) from differently polluted areas

Energy Technology Data Exchange (ETDEWEB)

Castelli, Martina Galatea [Norwegian Polar Institute, Fram Centre, 9296 Tromsø (Norway); University of Bergen, Department of Biology, 5020 Bergen (Norway); Rusten, Marte; Goksøyr, Anders [University of Bergen, Department of Biology, 5020 Bergen (Norway); Routti, Heli, E-mail: heli.routti@npolar.no [Norwegian Polar Institute, Fram Centre, 9296 Tromsø (Norway)

2014-01-15

Highlights: •Genes regulating lipid metabolism were studied in ringed seals. •We compared highly contaminated Baltic seals and less contaminated Svalbard seals. •mRNA expression of hepatic PPARγ was higher in the Baltic seals. •mRNA expression of adipose PPARγ target genes was higher in the Baltic seals. •Contaminant exposure may affect lipid metabolism in the Baltic ringed seals. -- Abstract: There is a growing concern about the ability of persistent organic pollutants (POPs) to influence lipid metabolism. Although POPs are found at high concentrations in some populations of marine mammals, for example in the ringed seal (Pusa hispida) from the Baltic Sea, little is known about the effects of POPs on their lipid metabolism. An optimal regulation of lipid metabolism is crucial for ringed seals during the fasting/molting season. This is a physiologically stressful period, during which they rely on the energy stored in their fat reserves. The mRNA expression levels for seven genes involved in lipid metabolism were analyzed in liver and/or blubber tissue from molting ringed seals from the polluted Baltic Sea and a less polluted reference location, Svalbard (Norway). mRNA expression of genes encoding peroxisome proliferator-activated receptors (PPAR) α and γ and their target genes acyl-coenzyme A oxidase 1 (ACOX1) and cluster of differentiation 36 (CD36) were analyzed in liver. mRNA expression level of genes encoding PPARβ, PPARγ and their target genes encoding fatty acid binding protein 4 (FABP4) and adiponectin (ADIPOQ) were measured in inner and middle blubber layers. In addition, we evaluated the influence of molting status on hepatic mRNA expression of genes encoding PPARs and their target genes in ringed seals from Svalbard. Our results show higher mRNA expression of genes encoding hepatic PPARγ and adipose PPARβ, FABP4, and ADIPOQ in the Baltic seals compared to the Svalbard seals. A positive relationship between mRNA expressions of genes

CK2(beta)tes gene encodes a testis-specific isoform of the regulatory subunit of casein kinase 2 in Drosophila melanogaster

DEFF Research Database (Denmark)

Kalmykova, Alla I; Shevelyov, Yuri Y; Polesskaya, Oksana O

2002-01-01

An earlier described CK2(beta)tes gene of Drosophila melanogaster is shown to encode a male germline specific isoform of regulatory beta subunit of casein kinase 2. Western-analysis using anti-CK2(beta)tes Ig revealed CK2(beta)tes protein in Drosophila testes extract. Expression of a CK2(beta...... and coimmunoprecipitation analysis of protein extract from Drosophila testes, we demonstrated an association between CK2(beta)tes and CK2alpha. Northern-analysis has shown that another regulatory (beta') subunit found recently in D. melanogaster genome is also testis-specific. Thus, we describe the first example of two...

Lipoprotein Lipase mRNA expression in different tissues of farm ...

African Journals Online (AJOL)

Lipoprotein lipase (LPL) controls triacylglycerol partitioning between adipose tissues and muscles, so it is important enzyme for fattening of animals .The present work was planned to clarify the use of polymerase chain reaction (PCR) for detection of LPL mRNA expression in different tissues representing internal organs of ...

GABAergic Neurons in the Rat Medial Septal Complex Express Relaxin-3 Receptor (RXFP3 mRNA

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Hector Albert-Gascó

2018-01-01

Full Text Available The medial septum (MS complex modulates hippocampal function and related behaviors. Septohippocampal projections promote and control different forms of hippocampal synchronization. Specifically, GABAergic and cholinergic projections targeting the hippocampal formation from the MS provide bursting discharges to promote theta rhythm, or tonic activity to promote gamma oscillations. In turn, the MS is targeted by ascending projections from the hypothalamus and brainstem. One of these projections arises from the nucleus incertus in the pontine tegmentum, which contains GABA neurons that co-express the neuropeptide relaxin-3 (Rln3. Both stimulation of the nucleus incertus and septal infusion of Rln3 receptor agonist peptides promotes hippocampal theta rhythm. The Gi/o-protein-coupled receptor, relaxin-family peptide receptor 3 (RXFP3, is the cognate receptor for Rln3 and identification of the transmitter phenotype of neurons expressing RXFP3 in the septohippocampal system can provide further insights into the role of Rln3 transmission in the promotion of septohippocampal theta rhythm. Therefore, we used RNAscope multiplex in situ hybridization to characterize the septal neurons expressing Rxfp3 mRNA in the rat. Our results demonstrate that Rxfp3 mRNA is abundantly expressed in vesicular GABA transporter (vGAT mRNA- and parvalbumin (PV mRNA-positive GABA neurons in MS, whereas ChAT mRNA-positive acetylcholine neurons lack Rxfp3 mRNA. Approximately 75% of Rxfp3 mRNA-positive neurons expressed vGAT mRNA (and 22% were PV mRNA-positive, while the remaining 25% expressed Rxfp3 mRNA only, consistent with a potential glutamatergic phenotype. Similar proportions were observed in the posterior septum. The occurrence of RXFP3 in PV-positive GABAergic neurons gives support to a role for the Rln3-RXFP3 system in septohippocampal theta rhythm.

Évaluation de deux variétés de riz (CK73 et CK90) à la Toxicité ...

African Journals Online (AJOL)

SARAH

30 avr. 2017 ... Evaluation of two rice varieties (CK73 and CK90) on iron toxicity and some biotic stress in. Yamoussoukro ... des parties aériennes (Becker et Asch, 2005 ; ... T0 : témoin, variété WITA 9 ; T1 : variété CK 73 ; T2 : variété CK 90.

Differential expression of PARP1 mRNA in leucocytes of patients ...

Indian Academy of Sciences (India)

P. 2011 Differential expression of PARP1 mRNA in leucocytes of patients with Down's syndrome. J. Genet. ... of Alzheimer disease at an earlier age than subjects with- ... family and personal informed consent. .... In effect, they report that.

Peripheral Mononuclear Cell Resistin mRNA Expression Is Increased in Type 2 Diabetic Women

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Panayoula C. Tsiotra

2008-01-01

Full Text Available Resistin has been shown to cause insulin resistance and to impair glucose tolerance in rodents, but in humans its physiological role still remains elusive. The aim of this study was to examine whether resistin mRNA expression in human peripheral mononuclear cells (PBMCs and its corresponding plasma levels are altered in type 2 diabetes. Resistin mRNA levels were easily detectable in human PBMC, and found to be higher in DM2 compared to healthy women (P=.05. Similarly, mononuclear mRNA levels of the proinflammatory cytokines IL-1β, TNF-α, and IL-6 were all significantly higher in DM2 compared to control women (P<.001. The corresponding plasma resistin levels were slightly, but not significantly, increased in DM2 women (P=.051, and overall, they correlated significantly with BMI (r=0.406, P=.010 and waist circumference (r=0.516, P=.003, but not with fasting insulin levels or HOMA-IR. Resistin mRNA expression is increased in PBMC from DM2 women, together with increased expression of the inflammatory cytokines IL-1β, TNF-α, and IL-6, independent of obesity. These results suggest that resistin and cytokines might contribute to the low-grade inflammation and the increased atherogenic risk observed in these patients.

Endurance exercise induces mRNA expression of oxidative enzymes in human skeletal muscle late in recovery

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Leick, Lotte; Plomgaard, Peter S.; Grønløkke, L.

2010-01-01

exercise. To test the hypothesis that mRNA expression of many oxidative enzymes is up-regulated late in recovery (10-24 h) after exercise, male subjects (n=8) performed a 90-min cycling exercise (70% VO(2-max)), with muscle biopsies obtained before exercise (pre), and after 10, 18 and 24 h of recovery....... The mRNA expression of carnitine-palmitoyltransferase (CPT)I, CD36, 3-hydroxyacyl-CoA-dehydrogenase (HAD), cytochrome (Cyt)c, aminolevulinate-delta-synthase (ALAS)1 and GLUT4 was 100-200% higher at 10-24 h of recovery from exercise than in a control trial. Exercise induced a 100-300% increase...... in peroxisome proliferator-activated receptor gamma co-activator (PGC)-1alpha, citrate synthase (CS), CPTI, CD36, HAD and ALAS1 mRNA contents at 10-24 h of recovery relative to before exercise. No protein changes were detected in Cytc, ALAS1 or GLUT4. This shows that mRNA expression of several training...

Prognostic significance of urokinase plasminogen activator and plasminogen activator inhibitor-1 mRNA expression in lymph node- and hormone receptor-positive breast cancer

International Nuclear Information System (INIS)

Leissner, Philippe; Verjat, Thibault; Bachelot, Thomas; Paye, Malick; Krause, Alexander; Puisieux, Alain; Mougin, Bruno

2006-01-01

One of the most thoroughly studied systems in relation to its prognostic relevance in patients with breast cancer, is the plasminogen activation system that comprises of, among others, the urokinase Plasminogen Activator (uPA) and its main inhibitor, the Plasminogen Activator Inhibitor-1 (PAI-1). In this study, we investigated the prognostic value of uPA and PAI-1 at the mRNA level in lymph node- and hormone receptor-positive breast cancer. The study included a retrospective series of 87 patients with hormone-receptor positive and axillary lymph node-positive breast cancer. All patients received radiotherapy, adjuvant anthracycline-based chemotherapy and five years of tamoxifen treatment. The median patient age was 54 and the median follow-up time was 79 months. Distant relapse occurred in 30 patients and 22 patients died from breast cancer during follow-up. We investigated the prognostic value of uPA and PAI-1 at the mRNA level as measured by real-time quantitative RT-PCR. uPA and PAI-1 gene expression was not found to be correlated with any of the established clinical and pathological factors. Metastasis-free Survival (MFS) and Breast Cancer specific Survival (BCS) were significantly shorter in patients expressing high levels of PAI-1 mRNA (p < 0.0001; p < 0.0001; respectively). In Cox multivariate analysis, the level of PAI-1 mRNA appeared to be the strongest prognostic factor for MFS (Hazard Ratio (HR) = 10.12; p = 0.0002) and for BCS (HR = 13.17; p = 0.0003). Furthermore, uPA gene expression was not significantly associated neither with MFS (p = 0.41) nor with BCS (p = 0.19). In a Cox-multivariate regression analysis, uPA expression did not demonstrate significant independent prognostic value. These findings indicate that high PAI-1 mRNA expression represents a strong and independent unfavorable prognostic factor for the development of metastases and for breast cancer specific survival in a population of hormone receptor- and lymph node-positive breast cancer

Prognostic significance of urokinase plasminogen activator and plasminogen activator inhibitor-1 mRNA expression in lymph node- and hormone receptor-positive breast cancer

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Krause Alexander

2006-08-01

Full Text Available Abstract Background One of the most thoroughly studied systems in relation to its prognostic relevance in patients with breast cancer, is the plasminogen activation system that comprises of, among others, the urokinase Plasminogen Activator (uPA and its main inhibitor, the Plasminogen Activator Inhibitor-1 (PAI-1. In this study, we investigated the prognostic value of uPA and PAI-1 at the mRNA level in lymph node- and hormone receptor-positive breast cancer. Methods The study included a retrospective series of 87 patients with hormone-receptor positive and axillary lymph node-positive breast cancer. All patients received radiotherapy, adjuvant anthracycline-based chemotherapy and five years of tamoxifen treatment. The median patient age was 54 and the median follow-up time was 79 months. Distant relapse occurred in 30 patients and 22 patients died from breast cancer during follow-up. We investigated the prognostic value of uPA and PAI-1 at the mRNA level as measured by real-time quantitative RT-PCR. Results uPA and PAI-1 gene expression was not found to be correlated with any of the established clinical and pathological factors. Metastasis-free Survival (MFS and Breast Cancer specific Survival (BCS were significantly shorter in patients expressing high levels of PAI-1 mRNA (p PAI-1 mRNA appeared to be the strongest prognostic factor for MFS (Hazard Ratio (HR = 10.12; p = 0.0002 and for BCS (HR = 13.17; p = 0.0003. Furthermore, uPA gene expression was not significantly associated neither with MFS (p = 0.41 nor with BCS (p = 0.19. In a Cox-multivariate regression analysis, uPA expression did not demonstrate significant independent prognostic value. Conclusion These findings indicate that high PAI-1 mRNA expression represents a strong and independent unfavorable prognostic factor for the development of metastases and for breast cancer specific survival in a population of hormone receptor- and lymph node-positive breast cancer patients.

High ALK mRNA expression has a negative prognostic significance in rhabdomyosarcoma

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Bonvini, P; Zin, A; Alaggio, R; Pawel, B; Bisogno, G; Rosolen, A

2013-01-01

Background: Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase aberrantly expressed in cancer, but its clinical and functional importance remain controversial. Mutation or amplification of ALK, as well as its expression levels assessed by conventional immunohistochemistry methods, has been linked to prognosis in cancer, although with potential bias because of the semi-quantitative approaches. Herein, we measured ALK mRNA expression in rhabdomyosarcoma (RMS) and determined its clin...

Cytochrome P450-2C11 mRNA is not expressed in endothelial cells dissected from rat renal arterioles.

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Heil, Sandra G; De Vriese, An S; Kluijtmans, Leo A J; Dijkman, Henry; van Strien, Denise; Akkers, Robert; Blom, Henk J

2005-01-01

Cytochrome P450 (CYP) isoenzymes (CYP2C and CYP2J) are involved in the production of epoxyeicosatrienoic acids, which are postulated as endothelium-derived hyperpolarizing factors (EDHFs). We hypothesized that if CYP2C11 is involved in the EDHF-mediated responses, its mRNA should be expressed in endothelial cells. We, therefore, examined the mRNA expression of CYP2C11 in endothelial cells of renal arterioles. Laser microdissection was applied to isolate endothelial cells from the renal arterioles of 4 male and 4 female Wistar rats. As a positive control of CYP2C11 expression, hepatocytes were also dissected from these rats. RNA was isolated and real-time quantitative polymerase chain reaction (Q-PCR) analysis was applied. Q-PCR analysis showed that CYP2C11 mRNA was not expressed in laser microdissected endothelial cells of renal arterioles of male and female rats. CYP2C11 mRNA expression was highly abundant in hepatocytes dissected from male livers, but in female livers hardly any CYP2C11 mRNA was detected. We have shown that endothelial cells can be dissected from small renal arterioles by laser microdissection to study the mRNA expression of specific genes by Q-PCR. Using this novel tool, we demonstrated that the CYP2C11 mRNA was not expressed in the endothelial cells of renal arterioles. Therefore, we speculate that CYP2C11 does not contribute to the EDHF-mediated responses in renal arterioles. Copyright (c) 2005 S. Karger AG, Basel.

Expression and clinicopathological significance of Mel-18 mRNA in colorectal cancer.

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Tao, Ji; Liu, Yan-Long; Zhang, Gan; Ma, Yu-Yan; Cui, Bin-Bin; Yang, Yan-Mei

2014-10-01

Mel-18 is a member of the polycomb group (PcG) of proteins, which are chromatin regulatory factors that play an important role in oncogenesis. This study was designed to investigate the clinical and prognostic significance of Mel-18 in colorectal cancer (CRC) patients. For this purpose, expression of Mel-18 mRNA was evaluated in 82 primary CRC and paired noncancerous mucosa samples by qRT-PCR and Western blotting. We found that overall Mel-18 mRNA expression in the CRC tissue was significantly lower than in the noncancerous mucosal tissue (p = 0.007, Wilcoxon matched-pairs signed-ranks test). Mel-18 was conversely correlated with the pathological classifications (p = 0.003 for T, p Mel-18 showed prolonged disease-free survivals (DFS) (p Mel-18 expression may be a risk factor for the patients' 3-year DFS (HR = 1.895; 95 % CI 1.032, 3.477; p = 0.039). It was therefore concluded that the lower Mel-18 expression might contribute to the CRC development/progression.

Protein phosphatase magnesium-dependent 1δ (PPM1D mRNA expression is a prognosis marker for hepatocellular carcinoma.

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Guang-Bing Li

Full Text Available Protein phosphatase magnesium-dependent 1δ (PPM1D is an oncogene, overexpressed in many solid tumors, including ovarian cancer and breast cancer. The current study examined the expression and the prognostic value of PPM1D mRNA in human hepatocellular carcinoma (HCC.Total RNA was extracted from 86 HCC and paired non-cancerous liver tissues. PPM1D mRNA expression was determined by real-time quantitative reverse transcriptase-polymerase chain reaction (qPCR. Immunohistochemistry assay was used to verify the expression of ppm1d protein in the HCC and non-cancerous liver tissues. HCC patients were grouped according to PPM1D mRNA expression with the average PPM1D mRNA level in non-cancerous liver tissue samples as the cut-off. Correlations between clinicopathologic variables, overall survival and PPM1D mRNA expression were analyzed.PPM1D mRNA was significantly higher in HCC than in the paired non-cancerous tissue (p<0.01. This was confirmed by ppm1d staining. 56 patients were classified as high expression group and the other 30 patients were categorized as low expression group. There were significant differences between the two groups in term of alpha-fetoprotein (α-FP level (p<0.01, tumor size (p<0.01, TNM stage (p<0.01, recurrence incidence (p<0.01 and family history of liver cancer (p<0.01. The current study failed to find significant differences between the two groups in the following clinical characteristics: age, gender, portal vein invasion, lymphnode metastasis, hepatitis B virus (HBV infection and alcohol intake. Survival time of high expression group was significantly shorter than that of low expression group (median survival, 13 months and 32 months, respectively, p<0.01.Up-regulation of PPM1D mRNA was associated with progressive pathological feature and poor prognosis in HCC patients. PPM1D mRNA may serve as a prognostic marker in HCC.

Altered PIWI-LIKE 1 and PIWI-LIKE 2 mRNA expression in ejaculated spermatozoa of men with impaired sperm characteristics.

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Giebler, Maria; Greither, Thomas; Müller, Lisa; Mösinger, Carina; Behre, Hermann M

2018-01-01

In about half the cases of involuntary childlessness, a male infertility factor is involved. The PIWI-LIKE genes, a subclade of the Argonaute protein family, are involved in RNA silencing and transposon control in the germline. Knockout of murine Piwi-like 1 and 2 homologs results in complete infertility in males. The aim of this study was to analyze whether the mRNA expression of human PIWI-LIKE 1-4 genes is altered in ejaculated spermatozoa of men with impaired sperm characteristics. Ninety male participants were included in the study, among which 47 were with normozoospermia, 36 with impaired semen characteristics according to the World Health Organization (WHO) manual, 5 th edition, and 7 with azoospermia serving as negative control for the PIWI-LIKE 1-4 mRNA expression in somatic cells in the ejaculate. PIWI-LIKE 1-4 mRNA expression in the ejaculated spermatozoa of the participants was measured by quantitative real-time PCR. In nonazoospermic men, PIWI-LIKE 1-4 mRNA was measurable in ejaculated spermatozoa in different proportions. PIWI-LIKE 1 (100.0%) and PIWI-LIKE 2 (49.4%) were more frequently expressed than PIWI-LIKE 3 (9.6%) and PIWI-LIKE 4 (15.7%). Furthermore, a decreased PIWI-LIKE 2 mRNA expression showed a significant correlation with a decreased sperm count (P = 0.022) and an increased PIWI-LIKE 1 mRNA expression with a decreased progressive motility (P = 0.048). PIWI-LIKE 1 and PIWI-LIKE 2 mRNA expression exhibited a significant association with impaired sperm characteristics and may be a useful candidate for the evaluation of the impact of PIWI-LIKE 1-4 mRNA expression on male infertility.

UCP2 mRNA expression is dependent on glucose metabolism in pancreatic islets

International Nuclear Information System (INIS)

Dalgaard, Louise T.

2012-01-01

Highlights: ► UCP2 mRNA levels are decreased in islets of Langerhans from glucokinase deficient mice. ► UCP2 mRNA up-regulation by glucose is dependent on glucokinase. ► Absence of UCP2 increases GSIS of glucokinase heterozygous pancreatic islets. ► This may protect glucokinase deficient mice from hyperglycemic damages. -- Abstract: Uncoupling Protein 2 (UCP2) is expressed in the pancreatic β-cell, where it partially uncouples the mitochondrial proton gradient, decreasing both ATP-production and glucose-stimulated insulin secretion (GSIS). Increased glucose levels up-regulate UCP2 mRNA and protein levels, but the mechanism for UCP2 up-regulation in response to increased glucose is unknown. The aim was to examine the effects of glucokinase (GK) deficiency on UCP2 mRNA levels and to characterize the interaction between UCP2 and GK with regard to glucose-stimulated insulin secretion in pancreatic islets. UCP2 mRNA expression was reduced in GK+/− islets and GK heterozygosity prevented glucose-induced up-regulation of islet UCP2 mRNA. In contrast to UCP2 protein function UCP2 mRNA regulation was not dependent on superoxide generation, but rather on products of glucose metabolism, because MnTBAP, a superoxide dismutase mimetic, did not prevent the glucose-induced up-regulation of UCP2. Glucose-stimulated insulin secretion was increased in UCP2−/− and GK+/− islets compared with GK+/− islets and UCP2 deficiency improved glucose tolerance of GK+/− mice. Accordingly, UCP2 deficiency increased ATP-levels of GK+/− mice. Thus, the compensatory down-regulation of UCP2 is involved in preserving the insulin secretory capacity of GK mutant mice and might also be implicated in limiting disease progression in MODY2 patients.

Protein Kinase CK2 Regulates Cytoskeletal Reorganization during Ionizing Radiation-Induced Senescence of Human Mesenchymal Stem Cells

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Wang, Daojing; Jang, Deok-Jin

2009-08-21

Human mesenchymal stem cells (hMSC) are critical for tissue regeneration. How hMSC respond to genotoxic stresses and potentially contribute to aging and cancer remain underexplored. We demonstrated that ionizing radiation induced cellular senescence of hMSC over a period of 10 days, showing a critical transition between day 3 and day 6. This was confirmed by senescence-associated beta-galactosidase (SA-{beta}-gal) staining, protein expression profiles of key cell cycle regulators (retinoblastoma (Rb) protein, p53, p21{sup waf1/Cip1}, and p16{sup INK4A}), and senescence-associated secretory phenotypes (SASPs) (IL-8, IL-12, GRO, and MDC). We observed dramatic cytoskeletal reorganization of hMSC through reduction of myosin-10, redistribution of myosin-9, and secretion of profilin-1. Using a SILAC-based phosphoproteomics method, we detected significant reduction of myosin-9 phosphorylation at Ser1943, coinciding with its redistribution. Importantly, through treatment with cell permeable inhibitors (4,5,6,7-tetrabromo-1H-benzotriazole (TBB) and 2-dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole (DMAT)), and gene knockdown using RNA interference, we identified CK2, a kinase responsible for myosin-9 phosphorylation at Ser1943, as a key factor contributing to the radiation-induced senescence of hMSC. We showed that individual knockdown of CK2 catalytic subunits CK2{alpha} and CK2{alpha}{prime} induced hMSC senescence. However, only knockdown of CK2{alpha} resulted in morphological phenotypes resembling those of radiation-induced senescence. These results suggest that CK2{alpha} and CK2{alpha}{prime} play differential roles in hMSC senescence progression, and their relative expression might represent a novel regulatory mechanism for CK2 activity.

IGF-1R mRNA expression is increased in obese children.

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Ricco, Rafaela Cristina; Ricco, Rubens Garcia; Queluz, Mariangela Carletti; de Paula, Mariana Teresa Sarti; Atique, Patricia Volpon; Custódio, Rodrigo José; Tourinho Filho, Hugo; Del Roio Liberatori, Raphael; Martinelli, Carlos Eduardo

2018-04-01

Obese children are often taller than age-matched subjects. Reports on GH and IGF-I levels in obese individuals are controversial, with normal and reduced GH-IGF-I levels having been reported in this group of patients. Thus, the aim of this study was to analyse insulin-like growth factor type 1 receptor (IGF-IR) mRNA expression in obese children. Forty-seven pre-pubertal children were included in this study: 29 were obese and taller than their target height, and 18 were normal eutrophic controls. Fasting blood samples were collected for IGF-IR mRNA expression in isolated lymphocytes and serum IGF-I, ALS, IGFBP-3, and IGFBP-1 concentration analysis. Relative IGF-IR gene expression (2 -ΔΔCT ) was significantly (P=0.025) higher in obese children (median 1.87) than in controls (1.15). Fourteen of the 29 obese subjects showed 2 -ΔΔCT values greater than or equal to 2, while only 2 individuals in the control group showed values above 2 (P=0.01). Obese children showed significantly (P=0.01) higher IGF-I concentrations than the control group (237ng/ml and 144ng/ml, respectively). Among obese patients, 65.5% had IGF-I values above the 75 percentile of the control group (P=0.02). ALS concentration was significantly (P=0.04) higher in the obese group, while IGFBP-3 levels were similar in obese and control children. IGFBP-1 concentration was lower in obese children, while insulin levels and HOMA-IR index were higher than in controls. The higher IGF-IR mRNA expression observed in obese children, associated with the higher IGF-I and ALS and the lower IGFBP-1 levels, suggest that the higher stature observed in these children may be due to increased IGF-I bioactivity. Copyright © 2017 Elsevier Ltd. All rights reserved.

Endoplasmic reticulum stress increases AT1R mRNA expression via TIA-1-dependent mechanism.

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Backlund, Michael; Paukku, Kirsi; Kontula, Kimmo K; Lehtonen, Jukka Y A

2016-04-20

As the formation of ribonucleoprotein complexes is a major mechanism of angiotensin II type 1 receptor (AT1R) regulation, we sought to identify novel AT1R mRNA binding proteins. By affinity purification and mass spectroscopy, we identified TIA-1. This interaction was confirmed by colocalization of AT1R mRNA and TIA-1 by FISH and immunofluorescence microscopy. In immunoprecipitates of endogenous TIA- 1, reverse transcription-PCR amplified AT1R mRNA. TIA-1 has two binding sites within AT1R 3'-UTR. The binding site proximal to the coding region is glyceraldehyde-3-phosphate dehydrogenase (GAPDH)-dependent whereas the distal binding site is not. TIA-1 functions as a part of endoplasmic reticulum (ER) stress response leading to stress granule (SG) formation and translational silencing. We and others have shown that AT1R expression is increased by ER stress-inducing factors. In unstressed cells, TIA-1 binds to AT1R mRNA and decreases AT1R protein expression. Fluorescence microscopy shows that ER stress induced by thapsigargin leads to the transfer of TIA-1 to SGs. In FISH analysis AT1R mRNA remains in the cytoplasm and no longer colocalizes with TIA-1. Thus, release of TIA-1-mediated suppression by ER stress increases AT1R protein expression. In conclusion, AT1R mRNA is regulated by TIA-1 in a ER stress-dependent manner. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Skeletal muscle myostatin mRNA expression is fiber-type specific and increases during hindlimb unloading

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Carlson, C. J.; Booth, F. W.; Gordon, S. E.

1999-01-01

Transgenic mice lacking a functional myostatin (MSTN) gene demonstrate greater skeletal muscle mass resulting from muscle fiber hypertrophy and hyperplasia (McPherron, A. C., A. M. Lawler, and S. -J. Lee. Nature 387: 83-90, 1997). Therefore, we hypothesized that, in normal mice, MSTN may act as a negative regulator of muscle mass. Specifically, we hypothesized that the predominately slow (type I) soleus muscle, which demonstrates greater atrophy than the fast (type II) gastrocnemius-plantaris complex (Gast/PLT), would show more elevation in MSTN mRNA abundance during hindlimb unloading (HU). Surprisingly, MSTN mRNA was not detectable in weight-bearing or HU soleus muscle, which atrophied 42% by the 7th day of HU in female ICR mice. In contrast, MSTN mRNA was present in weight-bearing Gast/PLT muscle and was significantly elevated (67%) at 1 day but not at 3 or 7 days of HU. However, the Gast/PLT muscle had only atrophied 17% by the 7th day of HU. Because the soleus is composed only of type I and IIa fibers, whereas the Gast/PLT expresses type IId/x and IIb in addition to type I and IIa, it was necessary to perform a more careful analysis of the relationship between MSTN mRNA levels and myosin heavy-chain (MHC) isoform expression (as a marker of fiber type). A significant correlation (r = 0.725, P < 0. 0005) was noted between the percentage of MHC isoform IIb expression and MSTN mRNA abundance in several muscles of the mouse hindlimb. These results indicate that MSTN expression is not strongly associated with muscle atrophy induced by HU; however, it is strongly associated with MHC isoform IIb expression in normal muscle.

Developmental changes in hypothalamic oxytocin and oxytocin receptor mRNA expression and their sensitivity to fasting in male and female rats.

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Matsuzaki, Toshiya; Iwasa, Takeshi; Munkhzaya, Munkhsaikhan; Tungalagsuvd, Altankhuu; Kawami, Takako; Murakami, Masahiro; Yamasaki, Mikio; Yamamoto, Yuri; Kato, Takeshi; Kuwahara, Akira; Yasui, Toshiyuki; Irahara, Minoru

2015-04-01

Oxytocin (OT) affects the central nervous system and is involved in a variety of social and non-social behaviors. Recently, the role played by OT in energy metabolism and its organizational effects on estrogen receptor alpha (ER-α) during the neonatal period have gained attention. In this study, the developmental changes in the hypothalamic mRNA levels of OT, the OT receptor (OTR), and ER-α were evaluated in male and female rats. In addition, the fasting-induced changes in the hypothalamic mRNA levels of OT and the OTR were evaluated. Hypothalamic explants were taken from postnatal day (PND) 10, 20, and 30 rats, and the mRNA level of each molecule was measured. Hypothalamic OT mRNA expression increased throughout the developmental period in both sexes. The rats' hypothalamic OTR mRNA levels were highest on PND 10 and decreased throughout the developmental period. In the male rats, the hypothalamic mRNA levels of ER-α were higher on PND 30 than on PND 10. On the other hand, no significant differences in hypothalamic ER-α mRNA expression were detected among the examined time points in the female rats, although hypothalamic ER-α mRNA expression tended to be higher on PND 30 than on PND 10. Significant positive correlations were detected between hypothalamic OT and ER-α mRNA expression in both the male and female rats. Hypothalamic OT mRNA expression was not affected by fasting at any of the examined time points in either sex. These results indicate that hypothalamic OT expression is not sensitive to fasting during the developmental period. In addition, as a positive correlation was detected between hypothalamic OT and ER-α mRNA expression, these two molecules might interact with each other to induce appropriate neuronal development. Copyright © 2015 Elsevier Ltd. All rights reserved.

Differential between Protein and mRNA Expression of CCR7 and SSTR5 Receptors in Crohn's Disease Patients

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Nathalie Taquet

2009-01-01

Full Text Available Crohn's disease (CD is a multifactorial chronic inflammatory bowel disease of unknown cause. The aim of the present study was to explore if mRNA over-expression of SSTR5 and CCR7 found in CD patients could be correlated to respective protein expression. When compared to healthy donors, SSTR5 was over-expressed 417 ± 71 times in CD peripheral blood mononuclear cells (PBMCs. Flow cytometry experiments showed no correlation between mRNA and protein expression for SSTR5 in PBMCs. In an attempt to find a reason of such a high mRNA expression, SSTR5 present on CD PBMCs were tested and found as biologically active as on healthy cells. In biopsies of CD intestinal tissue, SSTR5 was not over-expressed but CCR7, unchanged in PBMCs, was over-expressed by 10 ± 3 times in the lamina propria. Confocal microscopy showed a good correlation of CCR7 mRNA and protein expression in CD intestinal biopsies. Our data emphasize flow and image cytometry as impossible to circumvent in complement to molecular biology so to avoid false interpretation on receptor expressions. Once confirmed by further large-scale studies, our preliminary results suggest a role for SSTR5 and CCR7 in CD pathogenesis.

Seasonal relationship between gonadotropin, growth hormone, and estrogen receptor mRNA expression in the pituitary gland of largemouth bass.

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Martyniuk, Christopher J; Kroll, Kevin J; Porak, Wesley F; Steward, Cheree; Grier, Harry J; Denslow, Nancy D

2009-09-15

The objectives of this study were to investigate the seasonal changes in pituitary gonadotropins, growth hormone (GH), and estrogen receptor (ER) isoform mRNA in wild female and male largemouth bass (LMB) (Micropterus salmoides) from an unpolluted habitat to better understand reproductive physiology in this ecologically important species. Female pituitary luteinizing hormone (LH) beta subunit and follicle stimulating hormone (FSH) beta subunit mRNA showed significant seasonal variation with levels peaking from January to April and were lowest from May to August. Male LMB showed more variation in gonadotropin subunit expression from month to month. Females had approximately 2-3 times higher gonadotropin mRNA levels in the pituitary when compared to males. All three gonadotropin mRNAs in females were positively correlated to gonadosomatic index (GSI), but only LHbeta mRNA was correlated to GSI in males. Gonadotropin mRNA expression also increased with increasing oocyte and sperm maturation. Gonadotropin beta subunit mRNA expression was positively correlated to GH mRNA in both sexes. The expression of all three ER isoforms was significantly correlated to each other in both sexes. The concurrent increase in all three ER mRNA isoforms with increasing gonadotropin mRNA in females and males suggests a prominent role for E2 feedback on pituitary gonadotropin synthesis in both sexes and that each of the three ER isoforms are likely to play a role in the pituitary during teleost reproduction.

FLT3-ITD and MLL-PTD influence the expression of MDR-1, MRP-1, and BCRP mRNA but not LRP mRNA assessed with RQ-PCR method in adult acute myeloid leukemia.

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Nasilowska-Adamska, Barbara; Solarska, Iwona; Paluszewska, Monika; Malinowska, Iwona; Jedrzejczak, Wieslaw W; Warzocha, Krzysztof

2014-04-01

Fms-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD) and mixed-lineage leukemia gene-partial tandem duplication (MLL-PTD) are aberrations associated with leukemia which indicate unsatisfactory prognosis. Downstream regulatory targets of FLT3-ITD and MLL-PTD are not well defined. We have analyzed the expression of MDR-1, multidrug resistant protein-1 (MRP-1), breast cancer resistance protein (BCRP), and lung resistance protein (LRP) messenger RNA (mRNA) in relation to the mutational status of FLT3-ITD and MLL-PTD in 185 acute myeloid leukemia (AML) adult patients. The real-time quantitative polymerase chain reaction method was performed to assess the expression of the MDR-1, MRP-1, BCRP, and LRP mRNA, and the results were presented as coefficients calculated using an intermediate method according to Pfaffl's rule. Significantly higher expressions of MDR-1 mRNA were found in patients who did not harbor FLT3-ITD (0.20 vs. 0.05; p = 0.0001) and MRP-1 mRNA in patients with this mutation (0.96 vs. 0.70; p = 0.002) and of BCRP mRNA in patients with MLL-PTD (0.61 vs. 0.38; p = 0.03). In univariate analysis, the high expression of MDR-1 mRNA (≥0.1317) negatively influenced the outcome of induction therapy (p = 0.05), whereas the high expression of BCRP mRNA (≥1.1487) was associated with a high relapse rate (RR) (p = 0.013). We found that the high expression of MDR-1 (≥0.1317), MRP-1 (≥0.8409), and BCRP mRNA (≥1.1487) significantly influenced disease-free survival (DFS; p = 0.059, 0.032, and 0.009, respectively) and overall survival (0.048, 0.014, and 0.059, respectively). Moreover, a high expression of BCRP mRNA (≥1.1487) proved to be an independent prognostic factor for RR (p = 0.01) and DFS (p = 0.002) in multivariate analysis. The significant correlation between the expression of MDR-1, MRP-1, and BCRP mRNA and FLT3-ITD or MLL-PTD in AML patients requires further investigation.

Amitriptyline induces brain-derived neurotrophic factor (BDNF) mRNA expression through ERK-dependent modulation of multiple BDNF mRNA variants in primary cultured rat cortical astrocytes and microglia.

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Hisaoka-Nakashima, Kazue; Kajitani, Naoto; Kaneko, Masahiro; Shigetou, Takahiro; Kasai, Miho; Matsumoto, Chie; Yokoe, Toshiki; Azuma, Honami; Takebayashi, Minoru; Morioka, Norimitsu; Nakata, Yoshihiro

2016-03-01

A significant role of brain-derived neurotrophic factor (BDNF) has been previously implicated in the therapeutic effect of antidepressants. To ascertain the contribution of specific cell types in the brain that produce BDNF following antidepressant treatment, the effects of the tricyclic antidepressant amitriptyline on rat primary neuronal, astrocytic and microglial cortical cultures were examined. Amitriptyline increased the expression of BDNF mRNA in astrocytic and microglial cultures but not neuronal cultures. Antidepressants with distinct mechanisms of action, such as clomipramine, duloxetine and fluvoxamine, also increased BDNF mRNA expression in astrocytic and microglial cultures. There are multiple BDNF mRNA variants (exon I, IIA, IV and VI) expressed in astrocytes and microglia and the variant induced by antidepressants has yet to be elaborated. Treatment with antidepressants increased the expression of exon I, IV and VI in astrocyte and microglia. Clomipramine alone significantly upregulated expression of exon IIA. The amitriptyline-induced expression of both total and individual BDNF mRNA variants (exon I, IV and VI) were blocked by MEK inhibitor U0126, indicating MEK/ERK signaling is required in the expression of BDNF. These findings indicate that non-neural cells are a significant target of antidepressants and further support the contention that glial production of BDNF is crucial role in the therapeutic effect of antidepressants. The current data suggest that targeting of glial function could lead to the development of antidepressants with a truly novel mechanism of action. Copyright © 2016 Elsevier B.V. All rights reserved.

SIRT1 and FOXO1 mRNA expression in PBMC correlates to physical activity in COPD patients

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Taka C

2017-11-01

Full Text Available Chihiro Taka, Ryuji Hayashi, Kazuki Shimokawa, Kotaro Tokui, Seisuke Okazawa, Kenta Kambara, Minehiko Inomata, Toru Yamada, Shoko Matsui, Kazuyuki Tobe First Department of Internal Medicine, Faculty of Medicine, University of Toyama, Sugitani, Toyama, Toyama, Japan Background: Physical activity (PA is considered as one of the most important prognostic predictors in chronic obstructive pulmonary disease (COPD patients. Longevity gene, SIRT1, is reported to be involved in the pathogenesis of COPD by regulating the signaling pathways of oxidative stress, inflammation, and aging. We hypothesize that SIRT1 and related genes are also associated with the benefits of PA in COPD patients.Methods: Eighteen COPD outpatients were enrolled in this study, and their PA level was assessed with an accelerometer. We assessed the SIRT1 and related genes mRNA expression levels in the peripheral blood mononuclear cells (PBMCs of the subjects. We carried out respiratory function testing, blood gas analysis, the 6-minute walk test, and measurement of the cross-sectional area of the erector spinae muscles (ESMCSA by chest computed tomography. We analyzed the association of PA with the results of each of the examinations.Results: The mean age was 72±9 years, and the mean forced expiratory volume in 1 second was 1.4±0.56 L (52%±19% predicted. Our findings revealed a correlation between the daily PA and ESMCSA. The SIRT1 and Forkhead box O (FOXO1 mRNA expression levels in PBMCs were positively correlated with moderate-PA time (r=0.60, p=0.008 for SIRT1 and r=0.59, p=0.01 for FOXO1. Keywords: COPD, accelerometer, mRNA, walking, sedentary, moderate

Eradication of damaged keratinocytes in cutaneous lichen planus forms demonstrated by evaluation of epidermal and follicular expression of CK15, indices of apoptosis and regulatory protein S100.

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Upeniece, Ilze; Groma, Valerie; Skuja, Sandra; Cauce, Vinita

The study of cytoskeleton arrangement and its contribution to survival of cell-to-cell contacts appears to be essential for understanding of numerous cellular and tissue processes. Applying CK15, S100 labeling and TUNEL reaction to cutaneous lichen planus subtypes, we found CK15 expression in the outer and inner root sheath of hair follicles, the basal epidermal layer, and eccrine glands. Its follicular expression was decreased in nearby inflammatory infiltrates. The CK15 immunopositivity was mostly described as weak (92.3%) for lichen planus but equally subdivided into weak, moderate and strong in lichen planopilaris (2 = 32.514; df = 4; p lichen planopilaris involving the scalp: 81.2 ±10.7; 87.8 ±10.7 and 88.0 ±10.5 for the basal, spinous and upper epidermal layers, respectively. S100 positive epidermal and follicular cells did not differ in the lesions demonstrated in the study groups; still immunoreactivity was more pronounced in the scalp region of lichen planopilaris. Damage of cell-to-cell contacts was confirmed by electron microscopy. Apart from immunocyte-mediated keratinocyte death, cytoskeleton-based injury and loss of cell-to-cell and matrix contacts may be of great importance, leading to eradication of degrading cells and thus contributing to the pathogenesis of lichen planus.

Rift Valley fever virus NSS gene expression correlates with a defect in nuclear mRNA export.

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Copeland, Anna Maria; Van Deusen, Nicole M; Schmaljohn, Connie S

2015-12-01

We investigated the localization of host mRNA during Rift Valley fever virus (RVFV) infection. Fluorescence in situ hybridization revealed that infection with RVFV altered the localization of host mRNA. mRNA accumulated in the nuclei of RVFV-infected but not mock-infected cells. Further, overexpression of the NSS gene, but not the N, GN or NSM genes correlated with mRNA nuclear accumulation. Nuclear accumulation of host mRNA was not observed in cells infected with a strain of RVFV lacking the gene encoding NSS, confirming that expression of NSS is likely responsible for this phenomenon. Published by Elsevier Inc.

Quantitative tissue-specific dynamics of in vivo GILZ mRNA expression and regulation by endogenous and exogenous glucocorticoids.

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Ayyar, Vivaswath S; Almon, Richard R; Jusko, William J; DuBois, Debra C

2015-06-01

Glucocorticoids (GC) are steroid hormones, which regulate metabolism and immune function. Synthetic GCs, or corticosteroids (CS), have appreciable clinical utility via their ability to suppress inflammation in immune-mediated diseases like asthma and rheumatoid arthritis. Recent work has provided insight to novel GC-induced genes that mediate their anti-inflammatory effects, including glucocorticoid-induced leucine zipper (GILZ). Since GILZ comprises an important part of GC action, its regulation by both drug and hormone will influence CS therapy. In addition, GILZ expression is often employed as a biomarker of GC action, which requires judicious selection of sampling time. Understanding the in vivo regulation of GILZ mRNA expression over time will provide insight into both the physiological regulation of GILZ by endogenous GC and the dynamics of its enhancement by CS. A highly quantitative qRT-PCR assay was developed for measuring GILZ mRNA expression in tissues obtained from normal and CS-treated rats. This assay was applied to measure GILZ mRNA expression in eight tissues; to determine its endogenous regulation over time; and to characterize its dynamics in adipose tissue, muscle, and liver following treatment with CS. We demonstrate that GILZ mRNA is expressed in several tissues. GILZ mRNA expression in adipose tissue displayed a robust circadian rhythm that was entrained with the circadian oscillation of endogenous corticosterone; and is strongly enhanced by acute and chronic dosing. Single dosing also enhanced GILZ mRNA in muscle and liver, but the dynamics varied. In conclusion, GILZ is widely expressed in the rat and highly regulated by endogenous and exogenous GCs. © 2015 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

Colonization by non-pathogenic bacteria alters mRNA expression of cytochromes P450 in originally germ-free mice.

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Jourová, L; Anzenbacher, P; Lišková, B; Matušková, Z; Hermanová, P; Hudcovic, T; Kozáková, H; Hrnčířová, L; Anzenbacherová, E

2017-11-01

Gut microbiota provides a wide range of beneficial function for the host and has an immense effect on the host's health state. It has also been shown that gut microbiome is often involved in the biotransformation of xenobiotics; however, the molecular mechanisms of the interaction between the gut bacteria and the metabolism of drugs by the host are still unclear. To investigate the effect of microbial colonization on messenger RNA (mRNA) expression of liver cytochromes P450 (CYPs), the main drug-metabolizing enzymes, we used germ-free (GF) mice, lacking the intestinal flora and mice monocolonized by non-pathogenic bacteria Lactobacillus plantarum NIZO2877 or probiotic bacteria Escherichia coli Nissle 1917 compared to specific pathogen-free (SPF) mice. Our results show that the mRNA expression of Cyp1a2 and Cyp2e1 was significantly increased, while the expression of Cyp3a11 mRNA was decreased under GF conditions compared to the SPF mice. The both bacteria L. plantarum NIZO2877 and E. coli Nissle 1917 given to the GF mice decreased the level of Cyp1a2 mRNA and normalized it to the control level. On the other hand, the colonization by these bacteria had no effect on the expression of Cyp3a11 mRNA in the liver of the GF mice (which remained decreased). Surprisingly, monocolonization with chosen bacterial strains has shown a different effect on the expression of Cyp2e1 mRNA in GF mice. Increased level of Cyp2e1 expression observed in the GF mice was found also in mice colonized by L. plantarum NIZO2877 ; however, the colonization with probiotic E. coli Nissle 1917 caused a decrease in Cyp2e1 expression and partially restored the SPF mice conditions.

Rev-erb beta regulates the Srebp-1c promoter and mRNA expression in skeletal muscle cells

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Ramakrishnan, Sathiya N.; Lau, Patrick; Crowther, Lisa M. [The University of Queensland, Institute for Molecular Bioscience, St. Lucia, Qld 4072 (Australia); Cleasby, Mark E. [Diabetes and Obesity Research Program, Garvan Institute of Medical Research, St. Vincent' s Hospital, 384 Victoria Street, Darlinghurst, Sydney, NSW 2010 (Australia); Millard, Susan; Leong, Gary M. [The University of Queensland, Institute for Molecular Bioscience, St. Lucia, Qld 4072 (Australia); Cooney, Gregory J. [Diabetes and Obesity Research Program, Garvan Institute of Medical Research, St. Vincent' s Hospital, 384 Victoria Street, Darlinghurst, Sydney, NSW 2010 (Australia); Muscat, George E.O., E-mail: g.muscat@imb.uq.edu.au [The University of Queensland, Institute for Molecular Bioscience, St. Lucia, Qld 4072 (Australia)

2009-10-30

The nuclear hormone receptor, Rev-erb beta operates as a transcriptional silencer. We previously demonstrated that exogenous expression of Rev-erb{beta}{Delta}E in skeletal muscle cells increased Srebp-1c mRNA expression. We validated these in vitro observations by injection of an expression vector driving Rev-erb{beta}{Delta}E expression into mouse tibialis muscle that resulted in increased Srebp-1c mRNA expression. Paradoxically, Rev-erb{beta} siRNA expression in skeletal muscle cells repressed Srebp-1c expression, and indicated that Rev-erb{beta} expression was necessary for Srebp-1c expression. ChIP analysis demonstrated that Rev-erb{beta} was recruited to the Srebp-1c promoter. Moreover, Rev-erb{beta} trans-activated the Srebp-1c promoter, in contrast, Rev-erb{beta} efficiently repressed the Rev-erb{alpha} promoter, a previously characterized target gene. Finally, treatment with the Rev-erb agonist (hemin) (i) increased the trans-activation of the Srebp-1c promoter by Rev-erb{beta}; and (ii) increased Rev-erb{beta} and Srebp-1c mRNA expression. These data suggest that Rev-erb{beta} has the potential to activate gene expression, and is a positive regulator of Srebp-1c, a regulator of lipogenesis.

Correlation between Ahlbäck radiographic classification and anterior cruciate ligament status in primary knee arthrosis

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Glaucus Cajaty Martins

Full Text Available ABSTRACT OBJECTIVE: To correlate the Ahlbäck radiographic classification with the anterior cruciate ligament (ACL status in knee arthritis patients. METHODS: The study evaluated 89 knees of patients who underwent total knee arthroplasty due to primary osteoarthritis: 16 male and 69 females, with mean age 69.79 years (53-87 years. Osteoarthritis was classified radiographically by the Ahlbäck radiographic classification into five grades. The ACL was classified in the surgery as present or absent. The correlation of ACL status and Ahlbäck classification was assessed, as well as those of ACL status and the parameters age, gender, and tibiofemoral angulation (varus-valgus. RESULTS: In cases of varus knees, there was a correlation between grades I to III and ACL presence in 41/47 (86.7% cases and between grades IV and V and ACL absence in 15/17 (88.2% cases (p < 0.0001. In valgus knees, no statistically significant correlation was observed between the ACL status and the Ahlbäck classification. In the present study, absence of the ACL was more common in men (9/17; 52% than in women (19/72; 26%. CONCLUSION: In cases of medial osteoarthritis, the Ahlbäck radiographic classification is a useful parameter to predict ACL status (presence or absence. In gonarthritis in genu valgum, ACL status was not predicted by Ahlbäck's classification.

The potential lipolysis function of musclin and its mRNA expression ...

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Musclin is a newly discovered factor and its functions remain to be defined. This study investigated the tissue expression pattern of musclin gene and its potential effect on lipid metabolism. Musclin mRNA levels in adipose, muscle tissues and primary adipocytes were examined by quantitative PCR. The musclin gene ...

Unexpected Binding Mode of a Potent Indeno[1,2-b]indole-Type Inhibitor of Protein Kinase CK2 Revealed by Complex Structures with the Catalytic Subunit CK2α and Its Paralog CK2α′

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Jennifer Hochscherf

2017-12-01

Full Text Available Protein kinase CK2, a member of the eukaryotic protein kinase superfamily, is associated with cancer and other human pathologies and thus an attractive drug target. The indeno[1,2-b]indole scaffold is a novel lead structure to develop ATP-competitive CK2 inhibitors. Some indeno[1,2-b]indole-based CK2 inhibitors additionally obstruct ABCG2, an ABC half transporter overexpressed in breast cancer and co-responsible for drug efflux and resistance. Comprehensive derivatization studies revealed substitutions of the indeno[1,2-b]indole framework that boost either the CK2 or the ABCG2 selectivity or even support the dual inhibition potential. The best indeno[1,2-b]indole-based CK2 inhibitor described yet (IC50 = 25 nM is 5-isopropyl-4-(3-methylbut-2-enyl-oxy-5,6,7,8-tetrahydroindeno[1,2-b]indole-9,10-dione (4p. Herein, we demonstrate the membrane permeability of 4p and describe co-crystal structures of 4p with CK2α and CK2α′, the paralogs of human CK2 catalytic subunit. As expected, 4p occupies the narrow, hydrophobic ATP site of CK2α/CK2α′, but surprisingly with a unique orientation: its hydrophobic substituents point towards the solvent while its two oxo groups are hydrogen-bonded to a hidden water molecule. An equivalent water molecule was found in many CK2α structures, but never as a critical mediator of ligand binding. This unexpected binding mode is independent of the interdomain hinge/helix αD region conformation and of the salt content in the crystallization medium.

Regulation and function of FTO mRNA expression in human skeletal muscle and subcutaneous adipose tissue

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Grunnet, Louise G; Nilsson, Emma; Ling, Charlotte

2009-01-01

Objective. Common variants in FTO (the fat-mass and obesity-associated gene) associate with obesity and type 2 diabetes. The regulation and biological function of FTO mRNA expression in target tissue is unknown. We investigated the genetic and non-genetic regulation of FTO mRNA in skeletal muscle...... and adipose tissue, and their influence on in vivo glucose and fat metabolism. Research Design and Methods. The FTO rs9939609 polymorphism was genotyped in two twin cohorts: 1) 298 elderly twins aged 62-83 years with glucose tolerance ranging from normal to type 2 diabetes and 2) 196 young (25-32 years......) and elderly (58-66 years) non-diabetic twins examined by a hyperinsulinemic euglycemic clamp including indirect calorimetry. FTO mRNA expression was determined in subcutaneous adipose tissue (n=226) and skeletal muscle biopsies (n=158). Results. Heritability of FTO expression in both tissues was low, and FTO...

A pilot trial assessing urinary gene expression profiling with an mRNA array for diabetic nephropathy.

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Min Zheng

Full Text Available BACKGROUND: The initiation and progression of diabetic nephropathy (DN is complex. Quantification of mRNA expression in urinary sediment has emerged as a novel strategy for studying renal diseases. Considering the numerous molecules involved in DN development, a high-throughput platform with parallel detection of multiple mRNAs is needed. In this study, we constructed a self-assembling mRNA array to analyze urinary mRNAs in DN patients with aims to reveal its potential in searching novel biomarkers. METHODS: mRNA array containing 88 genes were fabricated and its performance was evaluated. A pilot study with 9 subjects including 6 DN patients and 3 normal controls were studied with the array. DN patients were assigned into two groups according to their estimate glomerular rate (eGFR: DNI group (eGFR>60 ml/min/1.73 m(2, n = 3 and DNII group (eGFR<60 ml/min/1.73 m(2, n = 3. Urinary cell pellet was collected from each study participant. Relative abundance of these target mRNAs from urinary pellet was quantified with the array. RESULTS: The array we fabricated displayed high sensitivity and specificity. Moreover, the Cts of Positive PCR Controls in our experiments were 24±0.5 which indicated high repeatability of the array. A total of 29 mRNAs were significantly increased in DN patients compared with controls (p<0.05. Among these genes, α-actinin4, CDH2, ACE, FAT1, synaptopodin, COL4α, twist, NOTCH3 mRNA expression were 15-fold higher than those in normal controls. In contrast, urinary TIMP-1 mRNA was significantly decreased in DN patients (p<0.05. It was shown that CTGF, MCP-1, PAI-1, ACE, CDH1, CDH2 mRNA varied significantly among the 3 study groups, and their mRNA levels increased with DN progression (p<0.05. CONCLUSION: Our pilot study demonstrated that mRNA array might serve as a high-throughput and sensitive tool for detecting mRNA expression in urinary sediment. Thus, this primary study indicated that mRNA array probably could be a

Multiple correlation analyses revealed complex relationship between DNA methylation and mRNA expression in human peripheral blood mononuclear cells.

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Xie, Fang-Fei; Deng, Fei-Yan; Wu, Long-Fei; Mo, Xing-Bo; Zhu, Hong; Wu, Jian; Guo, Yu-Fan; Zeng, Ke-Qin; Wang, Ming-Jun; Zhu, Xiao-Wei; Xia, Wei; Wang, Lan; He, Pei; Bing, Peng-Fei; Lu, Xin; Zhang, Yong-Hong; Lei, Shu-Feng

2018-01-01

DNA methylation is an important regulator on the mRNA expression. However, a genome-wide correlation pattern between DNA methylation and mRNA expression in human peripheral blood mononuclear cells (PBMCs) is largely unknown. The comprehensive relationship between mRNA and DNA methylation was explored by using four types of correlation analyses and a genome-wide methylation-mRNA expression quantitative trait locus (eQTL) analysis in PBMCs in 46 unrelated female subjects. An enrichment analysis was performed to detect biological function for the detected genes. Single pair correlation coefficient (r T1 ) between methylation level and mRNA is moderate (-0.63-0.62) in intensity, and the negative and positive correlations are nearly equal in quantity. Correlation analysis on each gene (T4) found 60.1% genes showed correlations between mRNA and gene-based methylation at P correlation (R T4  > 0.8). Methylation sites have regulation effects on mRNA expression in eQTL analysis, with more often observations in region of transcription start site (TSS). The genes under significant methylation regulation both in correlation analysis and eQTL analysis tend to cluster to the categories (e.g., transcription, translation, regulation of transcription) that are essential for maintaining the basic life activities of cells. Our findings indicated that DNA methylation has predictive regulation effect on mRNA with a very complex pattern in PBMCs. The results increased our understanding on correlation of methylation and mRNA and also provided useful clues for future epigenetic studies in exploring biological and disease-related regulatory mechanisms in PBMC.

Aging alters mRNA expression of amyloid transporter genes at the blood-brain barrier.

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Osgood, Doreen; Miller, Miles C; Messier, Arthur A; Gonzalez, Liliana; Silverberg, Gerald D

2017-09-01

Decreased clearance of potentially toxic metabolites, due to aging changes, likely plays a significant role in the accumulation of amyloid-beta (Aβ) peptides and other macromolecules in the brain of the elderly and in the patients with Alzheimer's disease (AD). Aging is the single most important risk factor for AD development. Aβ transport receptor proteins expressed at the blood-brain barrier are significantly altered with age: the efflux transporters lipoprotein receptor-related protein 1 and P-glycoprotein are reduced, whereas the influx transporter receptor for advanced glycation end products is increased. These receptors play an important role in maintaining brain biochemical homeostasis. We now report that, in a rat model of aging, gene transcription is altered in aging, as measured by Aβ receptor gene messenger RNA (mRNA) at 3, 6, 9, 12, 15, 20, 30, and 36 months. Gene mRNA expression from isolated cerebral microvessels was measured by quantitative polymerase chain reaction. Lipoprotein receptor-related protein 1 and P-glycoprotein mRNA were significantly reduced in aging, and receptor for advanced glycation end products was increased, in parallel with the changes seen in receptor protein expression. Transcriptional changes appear to play a role in aging alterations in blood-brain barrier receptor expression and Aβ accumulation. Copyright © 2017 Elsevier Inc. All rights reserved.

Anesthesia for euthanasia influences mRNA expression in healthy mice and after traumatic brain injury.

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Staib-Lasarzik, Irina; Kriege, Oliver; Timaru-Kast, Ralph; Pieter, Dana; Werner, Christian; Engelhard, Kristin; Thal, Serge C

2014-10-01

Tissue sampling for gene expression analysis is usually performed under general anesthesia. Anesthetics are known to modulate hemodynamics, receptor-mediated signaling cascades, and outcome parameters. The present study determined the influence of anesthetic paradigms typically used for euthanization and tissue sampling on cerebral mRNA expression in mice. Naïve mice and animals with acute traumatic brain injury induced by controlled cortical impact (CCI) were randomized to the following euthanasia protocols (n=10-11/group): no anesthesia (NA), 1 min of 4 vol% isoflurane in room air (ISO), 3 min of a combination of 5 mg/kg midazolam, 0.05 mg/kg fentanyl, and 0.5 mg/kg medetomidine intraperitoneally (COMB), or 3 min of 360 mg/kg chloral hydrate intraperitoneally (CH). mRNA expression of actin-1-related gene (Act1), FBJ murine osteosarcoma viral oncogene homolog B (FosB), tumor necrosis factor alpha (TNFα), heat shock protein beta-1 (HspB1), interleukin (IL)-6, tight junction protein 1 (ZO-1), IL-1ß, cyclophilin A, micro RNA 497 (miR497), and small cajal body-specific RNA 17 were determined by real-time polymerase chain reaction (PCR) in hippocampus samples. In naïve animals, Act1 expression was downregulated in the CH group compared with NA. FosB expression was downregulated in COMB and CH groups compared with NA. CCI reduced Act1 and FosB expression, whereas HspB1 and TNFα expression increased. After CCI, HspB1 expression was significantly higher in ISO, COMB, and CH groups, and TNFα expression was elevated in ISO and COMB groups. MiR497, IL-6, and IL-1ß were upregulated after CCI but not affected by anesthetics. Effects were independent of absolute mRNA copy numbers. The data demonstrate that a few minutes of anesthesia before tissue sampling are sufficient to induce immediate mRNA changes, which seem to predominate in the early-regulated gene cluster. Anesthesia-related effects on gene expression might explain limited reproduciblity of real

Effects of corticosteroid on the expressions of neuropeptide and cytokine mRNA and on tenocyte viability in lateral epicondylitis

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Han Soo

2012-10-01

Full Text Available Abstract Background The purpose of this study was to determine the reaction mechanism of corticosteroid by analyzing the expression patterns of neuropeptides (substance P (SP, calcitonin gene related peptide (CGRP and of cytokines (interleukin (IL-1α, tumor growth factor (TGF-β after corticosteroid treatment in lateral epicondylitis. In addition, we also investigated whether corticosteroid influenced tenocyte viability. Methods The corticosteroid triamcinolone acetonide (TAA was applied to cultured tenocytes of lateral epicondylitis, and the changes in the mRNA expressions of neuropeptides and cytokines and tenocyte viabilities were analyzed at seven time points. Quantitative real-time polymerase chain reaction and an MTT assay were used. Results The expression of SP mRNA was maximally inhibited by TAA at 24 hours but recovered at 72 hours, and the expressions of CGRP mRNA and IL-1α mRNA were inhibited at 24 and 3 hours, respectively. The expression of TGF-β mRNA was not significant. Tenocyte viability was significantly reduced by TAA at 24 hours. Conclusions We postulate that the reaction mechanism predominantly responsible for symptomatic relief after a corticosteroid injection involves the inhibitions of neuropeptides and cytokines, such as, CGRP and IL-1α. However the tenocyte viability was compromised by a corticosteroid.

Promoter Methylation and mRNA Expression of Response Gene to Complement 32 in Breast Carcinoma

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Nasab, E. E.; Nasab, E. E.; Hashemi, M.; Rafighdoost, F.

2016-01-01

Response gene to complement 32 (RGC32), induced by activation of complements, has been characterized as a cell cycle regulator; however, its role in carcinogenesis is still controversial. In the present study we compared RGC32 promoter methylation patterns and mRNA expression in breast cancerous tissues and adjacent normal tissues. Materials and Methods. Sixty-three breast cancer tissues and 63 adjacent non neoplastic tissues were included in our study. Design. Nested methylation-specific polymerase chain reaction (Nested-MSP) and quantitative PCR (qPCR) were used to determine RGC32 promoter methylation status and its mRNA expression levels, respectively. Results. RGC32 methylation pattern was not different between breast cancerous tissue and adjacent non neoplastic tissue (OR=2.30, 95% CI=0.95-5.54). However, qPCR analysis displayed higher levels of RGC32 mRNA in breast cancerous tissues than in noncancerous tissues (1.073 versus 0.959; P=0.001), irrespective of the promoter methylation status. The expression levels and promoter methylation of RGC32 were not correlated with any of patients’ clinical characteristics (P>0.05).

Sex differences in spatiotemporal expression of AR, ERα, and ERβ mRNA in the perinatal mouse brain.

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Mogi, Kazutaka; Takanashi, Haruka; Nagasawa, Miho; Kikusui, Takefumi

2015-01-01

It has been shown that every masculinized function might be organized by a particular contribution of androgens vs. estrogens in a critical time window. Here, we aimed to investigate the sex differences in brain testosterone levels and in the spatiotemporal dynamics of steroid receptor mRNA expression in perinatal mice, by using enzyme immunoassay and real-time PCR, respectively. We found that testosterone levels in the forebrain transiently increased around birth in male mice. During the perinatal period, levels of androgen receptor mRNA in the hypothalamus (hypo) and prefrontal cortex (PFC) were higher in male mice than in female mice. Estrogen receptor α (ERα) mRNA levels in the hypo and hippocampus were higher in male mice than in female mice before birth. In contrast, ERβ mRNA expression in the PFC was higher in female mice immediately after birth. These spatiotemporal sex differences in steroid receptor expression might contribute to organizing sex differences of not only reproductive function, but also anxiety, stress responses, and cognition in mice. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Mesenchymal stem cells cannot affect mRNA expression of toll-like receptors in different tissues during sepsis.

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Pedrazza, Leonardo; Pereira, Talita Carneiro Brandão; Abujamra, Ana Lucia; Nunes, Fernanda Bordignon; Bogo, Maurício Reis; de Oliveira, Jarbas Rodrigues

2017-07-01

Experimental animal models and human clinical studies support a crucial role for TLRs in infectious diseases. The aim of this study was to test the ability of MSCs, which have immunomodulatory effects, of altering the mRNA expression of toll-like receptors during a experimental model of sepsis in different tissues. Three experimental groups (male C57BL/6 mice) were formed for the test: control group, untreated septic group and septic group treated with MSCs (1 × 10 6 cells/animal). Lungs, cortex, kidney, liver and colon tissue were dissected after 12 h of sepsis induction and TLR2/3/4/9 mRNA were evaluated by RT-qPCR. We observed a decrease of TLR2 and 9 mRNA expression in the liver of the sepsis group, while TLR3 was decreased in the lung and liver. No change was found between the sepsis group and the sepsis + MSC group. In this model of experimental sepsis the MSCs were unable to modify the mRNA expression of the different toll-like receptors evaluated.

The alteration of mRNA expression of SOD and GPX genes, and proteins in tomato (Lycopersicon esculentum Mill under stress of NaCl and/or ZnO nanoparticles

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Hesham F. Alharby

2016-11-01

Full Text Available Five cultivars of tomato having different levels of salt stress tolerance were exposed to different treatments of NaCl (0, 3 and 6 g L−1 and ZnO-NPs (0, 15 and 30 mg L−1. Treatments with NaCl at both 3 and 6 g L−1 suppressed the mRNA levels of superoxide dismutase (SOD and glutathione peroxidase (GPX genes in all cultivars while plants treated with ZnO-NPs in the presence of NaCl, showed increments in the mRNA expression levels. This indicated that ZnO-NPs had a positive response on plant metabolism under salt stress. Superior expression levels of mRNA were observed in the salt tolerant cultivars, Sandpoint and Edkawy while the lowest level was detected in the salt sensitive cultivar, Anna Aasa. SDS–PAGE showed clear differences in patterns of protein expression among the cultivars. A negative protein marker for salt sensitivity and ZnO-NPs was detected in cv. Anna Aasa at a molecular weight of 19.162 kDa, while the tolerant cultivar Edkawy had two positive markers at molecular weights of 74.991 and 79.735 kDa. Keywords: Tomato, Salt stress, Nanoparticles, Gene expression, Real-time PCR, Polymorphism

Fas ligand expression in human and mouse cancer cell lines; a caveat on over-reliance on mRNA data

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Ryan Aideen E

2006-02-01

Full Text Available Abstract Background During carcinogenesis, tumors develop multiple mechanisms for evading the immune response, including upregulation of Fas ligand (FasL/CD95L expression. Expression of FasL may help to maintain tumor cells in a state of immune privilege by inducing apoptosis of anti-tumor immune effector cells. Recently this idea has been challenged by studies reporting that tumor cells of varying origin do not express FasL. In the present study, we aimed to comprehensively characterize FasL expression in tumors of both murine and human origin over a 72 hour time period. Methods RNA and protein was extracted from six human (SW620, HT29, SW480, KM12SM, HCT116, Jurkat and three mouse (CMT93, CT26, B16F10 cancer cell lines at regular time intervals over a 72 hour time period. FasL expression was detected at the mRNA level by RT-PCR, using intron spanning primers, and at the protein level by Western Blotting and immunofluorescence, using a polyclonal FasL- specific antibody. Results Expression of FasL mRNA and protein was observed in all cell lines analysed. However, expression of FasL mRNA varied dramatically over time, with cells negative for FasL mRNA at many time points. In contrast, 8 of the 9 cell lines constitutively expressed FasL protein. Thus, cells can abundantly express FasL protein at times when FasL mRNA is absent. Conclusion These findings demonstrate the importance of complete analysis of FasL expression by tumor cells in order to fully characterize its biological function and may help to resolve the discrepancies present in the literature regarding FasL expression and tumor immune privilege.

Associations of ACE Gene Insertion/Deletion Polymorphism, ACE Activity, and ACE mRNA Expression with Hypertension in a Chinese Population

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He, Qingfang; Fan, Chunhong; Yu, Min; Wallar, Gina; Zhang, Zuo-Feng; Wang, Lixin; Zhang, Xinwei; Hu, Ruying

2013-01-01

Background The present study was designed to explore the association of angiotensin converting enzyme (ACE) gene insertion/deletion (I/D, rs4646994) polymorphism, plasma ACE activity, and circulating ACE mRNA expression with essential hypertension (EH) in a Chinese population. In addition, a new detection method for circulating ACE mRNA expression was explored. Methods The research was approved by the ethics committee of Zhejiang Provincial Center for Disease Prevention and Control. Written informed consent was obtained prior to the investigation. 221 hypertensives (cases) and 221 normotensives (controls) were interviewed, subjected to a physical examination, and provided blood for biochemical and genetic tests. The ACE mRNA expression was analyzed by real time fluorescent quantitative Reverse Transcription PCR (FQ-RT-PCR). We performed logistic regression to assess associations of ACE I/D genotypes, ACE activity, and ACE mRNA expression levels with hypertension. Results The results of the multivariate logistic regression analysis showed that the additive model (ID, DD versus II) of the ACE genotype revealed an association with hypertension with adjusted OR of 1.43(95% CI: 1.04-1.97), and ACE ID genotype with adjusted OR of 1.72(95% CI: 1.01-2.92), DD genotype with adjusted OR of 1.94(95% CI: 1.01-3.73), respectively. In addition, our data also indicate that plasma ACE activity (adjusted OR was 1.13(95% CI: 1.08-1.18)) was significantly related to hypertension. However, the plasma ACE mRNA expressions were not different between the cases and controls. Conclusion ACE I/D polymorphism and ACE activity revealed significant influence on hypertension, while circulating ACE mRNA expression was not important factors associated with hypertension in this Chinese population. The detection of circulating ACE mRNA expression by FQ-RT-PCR might be a useful method for early screening and monitoring of EH. PMID:24098401

High BMI levels associate with reduced mRNA expression of IL10 and increased mRNA expression of iNOS (NOS2) in human frontal cortex

DEFF Research Database (Denmark)

Lauridsen, J K; Olesen, R H; Vendelbo, J

2017-01-01

analysis was performed with BMI as variable on data on IL10, IL1β, IL6, PTGS2 (COX2) and NOS2 (iNOS). Increasing BMI is associated with a decrease in the mRNA expression of IL10 (P=0.014) and an increase in the expression of NOS2 (iNOS; P=0.040). Expressions of IL10 and NOS2 (iNOS) were negatively...... correlated (PIL10 was mostly affected by individuals with BMI ⩾40. Multiple linear regression analyses with BMI, age, sex and race as variables were performed in order to identify potential confounders. In conclusion, increasing BMI could affect the IL10-mediated anti...

Development of Pharmacophore Model for Indeno[1,2-b]indoles as Human Protein Kinase CK2 Inhibitors and Database Mining

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Samer Haidar

2017-01-01

Full Text Available Protein kinase CK2, initially designated as casein kinase 2, is an ubiquitously expressed serine/threonine kinase. This enzyme, implicated in many cellular processes, is highly expressed and active in many tumor cells. A large number of compounds has been developed as inhibitors comprising different backbones. Beside others, structures with an indeno[1,2-b]indole scaffold turned out to be potent new leads. With the aim of developing new inhibitors of human protein kinase CK2, we report here on the generation of common feature pharmacophore model to further explain the binding requirements for human CK2 inhibitors. Nine common chemical features of indeno[1,2-b]indole-type CK2 inhibitors were determined using MOE software (Chemical Computing Group, Montreal, Canada. This pharmacophore model was used for database mining with the aim to identify novel scaffolds for developing new potent and selective CK2 inhibitors. Using this strategy several structures were selected by searching inside the ZINC compound database. One of the selected compounds was bikaverin (6,11-dihydroxy-3,8-dimethoxy-1-methylbenzo[b]xanthene-7,10,12-trione, a natural compound which is produced by several kinds of fungi. This compound was tested on human recombinant CK2 and turned out to be an active inhibitor with an IC50 value of 1.24 µM.

DDAH2 mRNA expression is inversely associated with some cardiovascular risk-related features in healthy young adults.

Science.gov (United States)

Puchau, Blanca; Hermsdorff, Helen Hermana M; Zulet, M Angeles; Martínez, J Alfredo

2009-01-01

The purpose of this study was to evaluate whether the mRNA expression profiles of three genes (PRMT1, DDAH2 and NOS3) are related to ADMA metabolism and signalling, and the potential relationships with anthropometrical, biochemical, lifestyle and inflammatory indicators in healthy young adults. An emphasis on the putative effect of different mRNA expression on cardiovascular risk-related features was paid. Anthropometrical measurements as well as lifestyle features were analyzed in 120 healthy young adults. Fasting blood samples were collected for the measurement of glucose and lipid profiles as well as the concentrations of selected inflammatory markers. Profiles of mRNA expression were assessed for PRMT1, DDAH2 and NOS3 genes from peripheral blood mononuclear cells. Regarding inflammatory biomarkers, DDAH2 was inversely associated with IL-6 and TNF-alpha. Moreover, subjects in the highest quintile of DDAH2 mRNA expression showed a reduced risk to have higher values of waist circumference, and to be more prone to show higher values of HDL-c. Interestingly, DDAH2 gene expression seemed to be related with some anthropometrical, biochemical, lifestyle and inflammatory indicators linked to cardiovascular risk in apparently healthy young adults, emerging as a potential disease marker.

DDAH2 mRNA Expression Is Inversely Associated with Some Cardiovascular Risk-Related Features in Healthy Young Adults

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Blanca Puchau

2009-01-01

Full Text Available The purpose of this study was to evaluate whether the mRNA expression profiles of three genes (PRMT1, DDAH2 and NOS3 are related to ADMA metabolism and signalling, and the potential relationships with anthropometrical, biochemical, lifestyle and inflammatory indicators in healthy young adults. An emphasis on the putative effect of different mRNA expression on cardiovascular risk-related features was paid. Anthropometrical measurements as well as lifestyle features were analyzed in 120 healthy young adults. Fasting blood samples were collected for the measurement of glucose and lipid profiles as well as the concentrations of selected inflammatory markers. Profiles of mRNA expression were assessed for PRMT1, DDAH2 and NOS3 genes from peripheral blood mononuclear cells. Regarding inflammatory biomarkers, DDAH2 was inversely associated with IL-6 and TNF-α. Moreover, subjects in the highest quintile of DDAH2 mRNA expression showed a reduced risk to have higher values of waist circumference, and to be more prone to show higher values of HDL-c. Interestingly, DDAH2 gene expression seemed to be related with some anthropometrical, biochemical, lifestyle and inflammatory indicators linked to cardiovascular risk in apparently healthy young adults, emerging as a potential disease marker.

Reconstructing the thermal evolution of the CK chondrite parent body using Northwest Africa 5343, the least metamorphosed CK chondrite

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Dunn, T. L.; Gross, J.; O'Hara, E. J.

2017-12-01

Carbonaceous chondrites (CCs) represent some of the most pristine solar system material, providing constraints on the early formation of planetesimals. The CK chondrites are the only group of CCs to exhibit the full range of thermal metamorphism (petrologic type 3 to 6). Most unequilibrated CK chondrites (CK3s) have been metamorphosed to petrologic subtype 3.8 or higher. However, homogeneity of olivine suggests that CK3 chondrite Northwest Africa (NWA) 5343 is less metamorphosed than the other CK3s. The presence of unrecrystallized matrix indicates that it is less than petrologic type 3.7. To better assess the lower limits of metamorphism on the CK chondrite parent body, we performed a detailed analysis of matrix material in NWA 5343. Ascertaining the lower limit of metamorphism in the CK chondrites is critical when addressing the CK-CV parent body debate (e.g., one vs. two parent bodies), and will shed light onto the evolution of metamorphosed CC parent bodies. We recognize two texturally distinct regions in the matrix of NWA 5343. Both have similar mineralogies (mostly olivine with lesser pyroxene and plagioclase), but differ in grain size, shape, and porosity. The porous region of the sample is characterized by subhedral-rounded olivine grains, typically Skeletal pyroxene is also common. Original pore space is filled with a Ca-rich glass that appears to originate from an unusual vein in this region. Most interestingly, the extent of metamorphism varies within NWA 5343. Larger, anhedral olivine in the glassy region suggest that this region is more metamorphosed than the porous region. Even within the porous region there is a range of metamorphism, with small patches of granoblastic olivine intermixed with the clastic matrix. This suggests that NWA 5343 may represent a metamorphic breccia, a common occurrence in OCs and CCs of lower petrologic types, and provides insight into the evolution of the only completely metamorphosed CC parent body.

EST Table: CK561988 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK561988 rswpb0_004912.y1 10/09/29 64 %/191 aa ref|XP_001605087.1| PREDICTED: similar to fetal alzheimer... PREDICTED: similar to fetal alzheimer antigen, falz [Tribolium castaneum] CK536202 swp ...

Enhancing chemosensitivity to gemcitabine via RNA interference targeting the catalytic subunits of protein kinase CK2 in human pancreatic cancer cells

International Nuclear Information System (INIS)

Kreutzer, Jan N; Ruzzene, Maria; Guerra, Barbara

2010-01-01

Pancreatic cancer is a complex genetic disorder that is characterized by rapid progression, invasiveness, resistance to treatment and high molecular heterogeneity. Various agents have been used in clinical trials showing only modest improvements with respect to gemcitabine-based chemotherapy, which continues to be the standard first-line treatment for this disease. However, owing to the overwhelming molecular alterations that have been reported in pancreatic cancer, there is increasing focus on targeting molecular pathways and networks, rather than individual genes or gene-products with a combination of novel chemotherapeutic agents. Cells were transfected with small interfering RNAs (siRNAs) targeting the individual CK2 subunits. The CK2 protein expression levels were determined and the effect of its down-regulation on chemosensitization of pancreatic cancer cells was investigated. The present study examined the impact on cell death following depletion of the individual protein kinase CK2 catalytic subunits alone or in combination with gemcitabine and the molecular mechanisms by which this effect is achieved. Depletion of the CK2α or -α' subunits in combination with gemcitabine resulted in marked apoptotic and necrotic cell death in PANC-1 cells. We show that the mechanism of cell death is associated with deregulation of distinct survival signaling pathways. Cellular depletion of CK2α leads to phosphorylation and activation of MKK4/JNK while down-regulation of CK2α' exerts major effects on the PI3K/AKT pathway. Results reported here show that the two catalytic subunits of CK2 contribute differently to enhance gemcitabine-induced cell death, the reduced level of CK2α' being the most effective and that simultaneous reduction in the expression of CK2 and other survival factors might be an effective therapeutic strategy for enhancing the sensitivity of human pancreatic cancer towards chemotherapeutic agents

Chaos and Fractals in C-K Map

Science.gov (United States)

Wang, Xing-Yuan; Liang, Qing-Yong; Meng, Juan

The characteristic of the fixed points of the Carotid-Kundalini (C-K) map is investigated and the boundary equation of the first bifurcation of the C-K map in the parameter plane is given. Based on the studies of the phase graph, the power spectrum, the correlation dimension and the Lyapunov exponents, the paper reveals the general features of the C-K map transforming from regularity. Meanwhile, using the periodic scanning technology proposed by Welstead and Cromer, a series of Mandelbrot-Julia (M-J) sets of the complex C-K map are constructed. The symmetry of M-J set and the topological inflexibility of distributing of periodic region in the Mandelbrot set are investigated. By founding the whole portray of Julia sets based on Mandelbrot set qualitatively, we find out that Mandelbrot sets contain abundant information of structure of Julia sets.

EST Table: CK518471 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK518471 rswjb0_007167.y1 10/09/29 80 %/101 aa ref|XP_001605087.1| PREDICTED: similar to fetal alzheimer...11424.1| PREDICTED: similar to fetal alzheimer antigen, falz [Tribolium castaneum] CK536202 swj ...

Expressão do Mg+2, CK, AST e LDH em equinos finalistas de provas de enduro

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Juliana V.F. Sales

2013-01-01

ção de permeabilidade das células musculares estriadas esqueléticas, sugerindo o estabelecimento de um processo inflamatório agudo. Devido à expressão da atividade enzimática da CK (p≤0,001, por sua especificidade em relação à ocorrência de danos na musculatura estriada esquelética, juntamente com o íon magnésio (p=0,0004 que participa de várias reações celulares. Houve alterações na concentração de proteína plasmática total (p=0,0009 e hematócrito (p=0,0001, entre os momentos avaliados. Portanto estes resultados servem como valores de referência de equinos finalistas de provas de enduro de 90 km, auxiliando na prevenção da ocorrência de possíveis danos musculares e processos inflamatórios severos.

Isoenzyme-specific up-regulation of glutathione transferase and aldo-keto reductase mRNA expression by dietary quercetin in rat liver.

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Odbayar, Tseye-Oidov; Kimura, Toshinori; Tsushida, Tojiro; Ide, Takashi

2009-05-01

The impact of quercetin on the mRNA expression of hepatic enzymes involved in drug metabolism was evaluated with a DNA microarray and real-time PCR. Male Sprague-Dawley rats were fed an experimental diet containing either 0, 2.5, 5, 10, or 20 g/kg of quercetin for 15 days. The DNA microarray analysis of the gene expression profile in pooled RNA samples from rats fed diets containing 0, 5, and 20 g/kg of quercetin revealed genes of some isoenzymes of glutathione transferase (Gst) and aldo-keto reductase (Akr) to be activated by this flavonoid. Real-time PCR conducted with RNA samples from individual rats fed varying amounts of quercetin together with the microarray analysis showed that quercetin caused marked dose-dependent increases in the mRNA expression of Gsta3, Gstp1, and Gstt3. Some moderate increases were also noted in the mRNA expression of isoenzymes belonging to the Gstm class. Quercetin also dose-dependently increased the mRNA expression of Akr1b8 and Akr7a3. However, it did not affect the parameters of the other Gst and Akr isoenzymes. It is apparent that quercetin increases the mRNA expression of Gst and Akr involved in drug metabolism in an isoenzyme-specific manner. Inasmuch as Gst and Akr isoenzymes up-regulated in their gene expression are involved in the prevention and attenuation of cancer development, this consequence may account for the chemopreventive propensity of quercetin.

[Expression and significance of P-gp/mdr1 mRNA, MRP and LRP in non-Hodgkin's lymphoma].

Science.gov (United States)

Li, Le; Su, Li-ping; Ma, Li; Zhao, Jin; Zhu, Lei; Zhou, Yong-an

2009-03-01

To explore the expression and clinical significance of P-glycoprotein (P-gp)/mdr1mRNA, multidrug resistance-associated protein (MRP) and lung resistance protein (LRP) in newly diagnosed non-Hodgkin's lymphoma. mdr1 mRNA of in 41 patients with non-Hodgkin's lymphoma was assayed by semi-quantitative RT-PCR. The expressions of P-gp, MRP and LRP proteins in lymph node viable blasts were identified by flow cytometry. The results were compared with those obtained from control cases, and the correlation of the changes with clinical outcomes was analyzed. (1) Among the 41 cases, the positive expression of P-gp protein was detected in 8 cases, MRP in 7 cases, LRP in 15 cases, and mdr 1 mRNA in 11 cases. (2) The P-gp and LRP levels in NHL were significantly higher than those in control group, but MRP wasn't. The P-gp over-expression was significantly associated with mdr1mRNA (r = 0.396, P = 0.01). No correlation was showed among the expressions of P-gp, MRP and LRP. (3) Patients with P-gp expression had a poorer outcome of chemotherapy than those with P-gp-negative (P = 0.005). P-gp expression was significantly associated with higher clinical stage (P = 0.046) and elevated serum lactate dehydrogenase level (P = 0.032), but not associated with malignant degree (P = 0.298). MRP had no impact on the outcome of chemotherapy (P = 0.212), and wasn't significantly associated with higher clinical stage (P = 0.369), elevated LDH (P = 0.762) and higher malignant degree (P = 0.451). Patients with LRP expression had a poorer outcome of chemotherapy than those LRP-negative (P = 0.012). LRP expression was significantly associated with higher clinical stage (P = 0.0019), elevated LDH (P = 0.02) and higher malignant degree (P = 0.01). The data of this study indicate that P-gp and LRP expressions but not MRP expression are important in the mechanism of drug resistance associated with a poor clinical outcome in previously untreated NHL.

High BMI levels associate with reduced mRNA expression of IL10 and increased mRNA expression of iNOS (NOS2) in human frontal cortex

DEFF Research Database (Denmark)

Lauridsen, J K; Olesen, R H; Vendelbo, J

2017-01-01

unknown. Therefore we aim to examine the relationship between BMI and gene expression of central inflammatory markers in the human frontal cortex. Microarray data of 141 neurologically and psychiatrically healthy individuals were obtained through the BrainCloud database. A simple linear regression...... correlated (Plinear regression analyses with BMI, age, sex and race as variables were performed in order to identify potential confounders. In conclusion, increasing BMI could affect the IL10-mediated anti...... analysis was performed with BMI as variable on data on IL10, IL1β, IL6, PTGS2 (COX2) and NOS2 (iNOS). Increasing BMI is associated with a decrease in the mRNA expression of IL10 (P=0.014) and an increase in the expression of NOS2 (iNOS; P=0.040). Expressions of IL10 and NOS2 (iNOS) were negatively...

Arabidopsis mRNA polyadenylation machinery: comprehensive analysis of protein-protein interactions and gene expression profiling

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Mo Min

2008-05-01

Full Text Available Abstract Background The polyadenylation of mRNA is one of the critical processing steps during expression of almost all eukaryotic genes. It is tightly integrated with transcription, particularly its termination, as well as other RNA processing events, i.e. capping and splicing. The poly(A tail protects the mRNA from unregulated degradation, and it is required for nuclear export and translation initiation. In recent years, it has been demonstrated that the polyadenylation process is also involved in the regulation of gene expression. The polyadenylation process requires two components, the cis-elements on the mRNA and a group of protein factors that recognize the cis-elements and produce the poly(A tail. Here we report a comprehensive pairwise protein-protein interaction mapping and gene expression profiling of the mRNA polyadenylation protein machinery in Arabidopsis. Results By protein sequence homology search using human and yeast polyadenylation factors, we identified 28 proteins that may be components of Arabidopsis polyadenylation machinery. To elucidate the protein network and their functions, we first tested their protein-protein interaction profiles. Out of 320 pair-wise protein-protein interaction assays done using the yeast two-hybrid system, 56 (~17% showed positive interactions. 15 of these interactions were further tested, and all were confirmed by co-immunoprecipitation and/or in vitro co-purification. These interactions organize into three distinct hubs involving the Arabidopsis polyadenylation factors. These hubs are centered around AtCPSF100, AtCLPS, and AtFIPS. The first two are similar to complexes seen in mammals, while the third one stands out as unique to plants. When comparing the gene expression profiles extracted from publicly available microarray datasets, some of the polyadenylation related genes showed tissue-specific expression, suggestive of potential different polyadenylation complex configurations. Conclusion An

CRY Drives Cyclic CK2-Mediated BMAL1 Phosphorylation to Control the Mammalian Circadian Clock.

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Teruya Tamaru

Full Text Available Intracellular circadian clocks, composed of clock genes that act in transcription-translation feedback loops, drive global rhythmic expression of the mammalian transcriptome and allow an organism to anticipate to the momentum of the day. Using a novel clock-perturbing peptide, we established a pivotal role for casein kinase (CK-2-mediated circadian BMAL1-Ser90 phosphorylation (BMAL1-P in regulating central and peripheral core clocks. Subsequent analysis of the underlying mechanism showed a novel role of CRY as a repressor for protein kinase. Co-immunoprecipitation experiments and real-time monitoring of protein-protein interactions revealed that CRY-mediated periodic binding of CK2β to BMAL1 inhibits BMAL1-Ser90 phosphorylation by CK2α. The FAD binding domain of CRY1, two C-terminal BMAL1 domains, and particularly BMAL1-Lys537 acetylation/deacetylation by CLOCK/SIRT1, were shown to be critical for CRY-mediated BMAL1-CK2β binding. Reciprocally, BMAL1-Ser90 phosphorylation is prerequisite for BMAL1-Lys537 acetylation. We propose a dual negative-feedback model in which a CRY-dependent CK2-driven posttranslational BMAL1-P-BMAL1 loop is an integral part of the core clock oscillator.

Prolactin suppresses a progestin-induced CK5-positive cell population in luminal breast cancer through inhibition of progestin-driven BCL6 expression

Science.gov (United States)

Sato, Takahiro; Tran, Thai H.; Peck, Amy R.; Girondo, Melanie A.; Liu, Chengbao; Goodman, Chelain R.; Neilson, Lynn M.; Freydin, Boris; Chervoneva, Inna; Hyslop, Terry; Kovatich, Albert J.; Hooke, Jeffrey A.; Shriver, Craig D.; Fuchs, Serge Y.; Rui, Hallgeir

2014-01-01

Prolactin controls the development and function of milk-producing breast epithelia but also supports growth and differentiation of breast cancer, especially luminal subtypes. A principal signaling mediator of prolactin, Stat5, promotes cellular differentiation of breast cancer cells in vitro, and loss of active Stat5 in tumors is associated with anti-estrogen therapy failure in patients. In luminal breast cancer progesterone induces a cytokeratin-5 (CK5)-positive basal cell-like population. This population possesses characteristics of tumor stem cells including quiescence, therapy-resistance, and tumor-initiating capacity. Here we report that prolactin counteracts induction of the CK5-positive population by the synthetic progestin R5020 in luminal breast cancer cells both in vitro and in vivo. CK5-positive cells were chemoresistant as determined by four-fold reduced rate of apoptosis following docetaxel exposure. Progestin-induction of CK5 was preceded by marked up-regulation of BCL6, an oncogene and transcriptional repressor critical for the maintenance of leukemia-initiating cells. Knockdown of BCL6 prevented induction of CK5-positive cell population by progestin. Prolactin suppressed progestin-induced BCL6 through Jak2-Stat5 but not Erk- or Akt-dependent pathways. In premenopausal but not postmenopausal patients with hormone receptor-positive breast cancer, tumor protein levels of CK5 correlated positively with BCL6, and high BCL6 or CK5 protein levels were associated with unfavorable clinical outcome. Suppression of progestin-induction of CK5-positive cells represents a novel pro-differentiation effect of prolactin in breast cancer. The present progress may have direct implications for breast cancer progression and therapy since loss of prolactin receptor-Stat5 signaling occurs frequently and BCL6 inhibitors currently being evaluated for lymphomas may have value for breast cancer. PMID:23708665

Astrophysical applications of Delbrück scattering: Dust scattered gamma radiation from gamma ray bursts

International Nuclear Information System (INIS)

Kunwar, B.; Bhadra, A.; Gupta, S.K. Sen

2014-01-01

A preliminary, and perhaps the first, study of astrophysical applications of Delbrück scattering in a gamma-ray emitting celestial object like a gamma-ray burst (GRB) has been made. At energies≥100 MeV the elastic scattering of gamma-ray photons off the molecular dust surrounding the GRB site is dominated by Delbrück scattering. Expressions for Delbrück-scattered gamma-ray flux as a function of time has been obtained for a few selected energies by assuming a simple model of GRB. These are compared with Compton-scattered flux. At certain situations, interestingly, the former is found to exceed the latter for the first few milliseconds of the burst. The issue of detectability of Delbrück-scattered gamma-ray echo from the cloud of a GRB is discussed. Although it is observed that the detection of such an echo is not within the capability of the presently operating gamma-ray missions such as Fermi LAT, a rough estimate shows that one can be optimistic that future generation gamma-ray telescopes might be able to see such photons' contribution to the total flux. - Highlights: ► Astrophysical application of Delbrück scattering in a GRB has been made. ► Initially, the Delbrück scattering may dominate the scattering of GeV γ-rays. ► The issue of detectability of such radiations is discussed

Assessment of potential biomarkers, metallothionein and vitellogenin mRNA expressions in various chemically exposed benthic Chironomus riparius larvae

Science.gov (United States)

Park, Kiyun; Kwak, Inn-Sil

2012-12-01

The objective of this study was conducted to identify the possibility of using Chironomus metallothionein (MT) and vitellogenin (VTG) as biomarkers of stress caused by endocrinedisrupting chemicals (EDCs), heavy metals, herbicides and veterinary antibiotics. We characterized the MT and VTG cDNA in Chironomus riparius and evaluated their mRNA expression profiles following exposure to different environmental pollutants. The gene expression analysis showed that the MT mRNA levels increased significantly after long-term exposure to cadmium (Cd), copper (Cu), Lead (Pb), di(2-ethylhexyl) phthalate (DEHP), and 2,4-dichlorophenoxyacetic acid (2,4-D). Moreover, the VTG mRNA expression increased significantly in C. riparius larvae exposed to BPA, NP, DEHP, Cd, 2,4-D and fenbendazole. Evaluation of the long-term effects of environmental pollutants revealed up regulation of Chironomus MT mRNA in response to DEHP exposure among EDCs, and the level of the VTG mRNA was increased significantly following treatment with Cd and herbicide 2,4-D at all concentrations in a dose-dependent manner. These results indicate that VTG could be used as a potential biomarker of herbicide and Cd as well as EDCs, while MT was a potential biomarker of heavy metals such as Cd, Cu, and Pb in aquatic environments.

Statins Activate Human PPAR Promoter and Increase PPAR mRNA Expression and Activation in HepG2 Cells

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Makoto Seo

2008-01-01

Full Text Available Statins increase peroxisome proliferator-activated receptor (PPAR mRNA expression, but the mechanism of this increased PPAR production remains elusive. To examine the regulation of PPAR production, we examined the effect of 7 statins (atorvastatin, cerivastatin, fluvastatin, pitavastatin, pravastatin, rosuvastatin, and simvastatin on human PPAR promoter activity, mRNA expression, nuclear protein levels, and transcriptional activity. The main results are as follows. (1 Majority of statins enhanced PPAR promoter activity in a dose-dependent manner in HepG2 cells transfected with the human PPAR promoter. This enhancement may be mediated by statin-induced HNF-4. (2 PPAR mRNA expression was increased by statin treatment. (3 The PPAR levels in nuclear fractions were increased by statin treatment. (4 Simvastatin, pravastatin, and cerivastatin markedly enhanced transcriptional activity in 293T cells cotransfected with acyl-coenzyme A oxidase promoter and PPAR/RXR expression vectors. In summary, these data demonstrate that PPAR production and activation are upregulated through the PPAR promoter activity by statin treatment.

[Differentiation of human umbilical cord derived mesenchymal stem cells into low immunogenic and functional hepatocyte-like cells in vitro].

Science.gov (United States)

Ren, Hong-ying; Zhao, Qin-jun; Xing, Wen; Yang, Shao-guang; Lu, Shi-hong; Ren, Qian; Zhang, Lei; Han, Zhong-chao

2010-04-01

To investigate the biological function of hepatocyte-like cells derived from mesenchymal stem cells that isolated from human umbilical cord UC-MSCs in vitro, and to detect the changes in the immunogenicity of the differentiated hepatocyte-like cells (DHC). Transdifferentiation of UC-MSCs into hepatic lineage in vitro was induced in modified two-step induction medium. The expressions of hepatic specific markers were detected by RT-PCR analysis and immunofluorescence staining at different time points after induction. The levels of albumin and urea in the supernatants of cultures were measured by enzyme-linked immunosorbent assay. Furthermore, the immunosuppressive property of DHC was detected by one-way mixed lymphocyte culture. The mRNA and proteins of alpha fetoprotein (AFP), albumin (ALB),and cytokeratin-19 (CK-19) were expressed in naive UC-MSCs at low levels. DHC highly expressed hepatic markers AFP, ALB, CK-19, and tryptophan 2, 3-dioxygenase 14 and 28 days after hepatic differentiation and were accompanied by an increased production of ALB and urea in supernatant in a time-dependent manner. DHC did not express human leukocyte antigen DR antigen and significantly decreased the lymphocyte proliferation. UC-MSCs are able to differentiate into functional hepatocyte-like cells in vitro, while the immunogenicity of DHC remains low.

Relationship of calcitonin mRNA expression to the differentiation state of HL 60 cells.

Science.gov (United States)

Kiefer, P; Bacher, M; Pflüger, K H

1994-05-01

Raised plasma levels of immunoreactive human calcitonin (ihCT) can be found in patients with myeloid leukemia and seem to indicate a poor prognosis. High levels were found in acute undifferentiated and acute myeloblastic leukemia. To test whether CT expression could be a marker of myeloid differentiation, we used the promyelocytic leukemia cell line HL 60 which also expresses ihCT as a model system for myeloid differentiation. Exponentially growing HL 60 cells as well as differentiation induced HL 60 cells expressed a single 1.0 Kb CT transcript. The induction of HL 60 cell differentiation along the granulocytic lineage by DMSO or HMBA had no effect on the level of CT transcripts. Induction of monocytic/macrophagic differentiation by TPA resulted in a transient, about 10-fold elevated expression of CT steady state mRNA after 24 h. In contrast to TPA, induction of HL 60 cell differentiation along the monocytic pathway by Vit D3 had no detectable effect on the level of the CT in RNA expression at corresponding time points. These findings suggest that the transient induction of CT steady state mRNA expression by TPA is rather a direct effect of the phorbol ester than commitment along the monocytic line of differentiation.

[mRNA expression of dopamine receptor D2 and dopamine transporter in peripheral blood lymphocytes before and after treatment in children with tic disorder].

Science.gov (United States)

Ji, Xiao-Yi; Wu, Min

2016-04-01

To investigate the mRNA expression of dopamine receptor D2 (DRD2) and dopamine transporter (DAT) in peripheral blood lymphocytes before and after treatment in children with tic disorder (TD). RT-PCR was used to measure the mRNA expression of DRD2 and DAT in peripheral blood lymphocytes before and after treatment in 60 children with TD. The correlations between mRNA expression of DRD2 and DAT and the severity of TD were analyzed. Sixty healthy children served as the control group. Before treatment, the children with TD had a significant increase in the mRNA expression of DRD2 and DAT compared with the control group (PTic Severity Scale (YGTSS) score (P<0.05). In the children with moderate TD, the mRNA expression of DAT was positively correlated with YGTSS score (P<0.05). In children with TD, the mRNA expression of DRD2 in peripheral blood lymphocytes can be used as one of the indicators for diagnosing TD, assessing the severity of TD, and evaluating clinical outcomes.

Enrofloxacin and Probiotic Lactobacilli Influence PepT1 and LEAP-2 mRNA Expression in Poultry.

Science.gov (United States)

Pavlova, Ivelina; Milanova, Aneliya; Danova, Svetla; Fink-Gremmels, Johanna

2016-12-01

Expression of peptide transporter 1 (PepT1) and liver-expressed antimicrobial peptide 2 (LEAP-2) in chickens can be influenced by food deprivation, pathological conditions and drug administration. Effect of three putative probiotic Lactobacillus strains and enrofloxacin on the expression of PepT1 and LEAP-2 mRNA was investigated in Ross 308 chickens. One-day-old chicks (n = 24) were allocated to following groups: control (without treatment); group treated with probiotics via feed; group treated with a combination of probiotics and enrofloxacin; and a group given enrofloxacin only. The drug was administered at a dose of 10 mg kg -1 , via drinking water for 5 days. Samples from liver, duodenum and jejunum were collected 126 h after the start of the treatment. Expression levels of PepT1 and LEAP-2 were determined by real-time polymerase chain reaction and were statistically evaluated by Mann-Whitney test. Enrofloxacin administered alone or in combination with probiotics provoked a statistically significant up-regulation of PepT1 mRNA levels in the measured organ sites. These changes can be attributed to a tendency of improvement in utilization of dietary peptide and in body weight gain. LEAP-2 mRNA expression levels did not change significantly in enrofloxacin-treated chickens in comparison with control group.

Increase of CTGF mRNA expression by respiratory syncytial virus infection is abrogated by caffeine in lung epithelial cells.

Science.gov (United States)

Kunzmann, Steffen; Krempl, Christine; Seidenspinner, Silvia; Glaser, Kirsten; Speer, Christian P; Fehrholz, Markus

2018-04-16

Respiratory syncytial virus (RSV) is a leading cause of severe lower respiratory tract infection in early childhood. Underlying pathomechanisms of elevated pulmonary morbidity in later infancy are largely unknown. We found that RSV-infected H441 cells showed increased mRNA expression of connective tissue growth factor (CTGF), a key factor in airway remodeling. Additional dexamethasone treatment led to further elevated mRNA levels, indicating additive effects. Caffeine treatment prevented RSV-mediated increase of CTGF mRNA. RSV may be involved in airway remodeling processes by increasing CTGF mRNA expression. Caffeine might abrogate these negative effects and thereby help to restore lung homeostasis. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Effects of active acromegaly on bone mRNA and microRNA expression patterns.

Science.gov (United States)

Belaya, Zhanna; Grebennikova, Tatiana; Melnichenko, Galina; Nikitin, Alexey; Solodovnikov, Alexander; Brovkina, Olga; Grigoriev, Andrey; Rozhinskaya, Liudmila; Lutsenko, Alexander; Dedov, Ivan

2018-04-01

To evaluate the response of bone to chronic long-term growth hormone (GH) and insulin-like growth factor-1 (IGF1) excess by measuring the expression of selected mRNA and microRNA (miR) in bone tissue samples of patients with active acromegaly. Case-control study. Bone tissue samples were obtained during transsphenoidal adenomectomy from the sphenoid bone (sella turcica) from 14 patients with clinically and biochemically confirmed acromegaly and 10 patients with clinically non-functioning pituitary adenoma (NFPA) matched by sex and age. Expression of genes involved in the regulation of bone remodeling was studied using quantitative polymerase chain reaction (qPCR). Of the genes involved in osteoblast and osteoclast activity, only alkaline phosphatase (ALP) mRNA was 50% downregulated in patients with acromegaly. GH excess caused increased expression of the Wnt signaling antagonists ( DKK1) and agonists ( WNT10B) and changes in the levels of miR involved in mesenchymal stem cell commitment to chondrocytes (miR-199a-5p) or adipocytes (miR-27-5p, miR-125b-5p, miR-34a-5p, miR-188-3p) P  Acromegaly had minimal effects on tested mRNAs specific to osteoblast or osteoclast function except for downregulated ALP expression. The expressions of miR known to be involved in mesenchymal stem cell commitment and downregulated TWIST1 expression suggest acromegaly has a negative effect on osteoblastogenesis. © 2018 European Society of Endocrinology.

The utility of cytokeratins 7 and 20 (CK7/20 immunohistochemistry in the distinction of short-segment Barrett esophagus from gastric intestinal metaplasia: Is it reliable?

Directory of Open Access Journals (Sweden)

Bakirci Nadi

2003-12-01

Full Text Available Abstract Background The purpose of the present correlative immunohistochemical study was to assess the utility of cytokeratin (CK7 and CK20 expression in the diagnosis of short-segment Barrett esophagus, particularly its efficacy in differentiating Barrett mucosa from intestinal metaplasia of the gastric cardia and corpus. Methods Two groups of endoscopic biopsy specimens were examined, including 20 endoscopic biopsy specimens of short-segment Barrett esophagus (Group A and equal number exhibiting Helicobacter pylori associated intestinal metaplasia of the gastric cardia and corpus (Group B. All were investigated by immunohistochemistry using the standard ABC method for CK7 and CK20 expression. Fisher's exact test was used for statistical analysis of Barrett CK7/20 and gastric CK7/20 patterns between the groups. Results The anticipated pattern of reactivity in Barrett mucosa (CK7: strong diffuse positivity in superficial and deep glands; CK20: positivity in surface epithelium and superficial glands was seen in 2 cases of Group A specimens. The expected gastric pattern (CK7: patchy immunostaining with variable involvement of deep glands; CK20: patchy immunostaining of superficial and deep glands in incomplete intestinal metaplasia / absence of CK7 immunoreactivity with strong CK20 staining in superficial and deep glands in complete intestinal metaplasia was seen in 8 cases of Group B specimens. The respective sensitivity and false-negativity values of CK7/20 staining for Barrett pattern in Group A were 10% and 90%, respectively. These values for gastric pattern in Group B were 40% and 60%, respectively. The specificity and false-positivity values of both patterns were same (100% and 0%, respectively. There was no statistically significant difference for Barrett pattern between the two groups (P = 0.487, while the observation of gastric pattern was significantly higher in Group B than in Group A (P = 0.02. Conclusions We concluded that these

Expression of connexin 37, 40, and 43 mRNA and protein in renal preglomerular arterioles

DEFF Research Database (Denmark)

Arensbak, B; Mikkelsen, Hanne Birte; Gustafsson, F

2001-01-01

arterioles in frozen sections was evaluated. SMC were isolated from kidneys using an iron oxide sieve method and explant technique. Total RNA from these cultures was tested by RT-PCR analysis for the expression of the three connexins mRNA. Using immunofluorescence we examined whether the expression pattern...

Effect of electroacupuncture on brain-derived neurotrophic factor mRNA expression in mouse hippocampus following cerebral ischemia-reperfusion injury.

Science.gov (United States)

Zhao, Jianxin; Xu, Huazhou; Tian, Yuanxiang; Hu, Manxiang; Xiao, Hongling

2013-04-01

This work aims to observe the effects of electroacupuncture on brain-derived neurotrophic factor (BDNF) mRNA expression in mouse hippocampus following cerebral ischemia-reperfusion injury. The models of mouse cerebral ischemia-reperfusion injury were established. A total of 96 healthy mice were randomly assigned into 4 groups, namely, the sham surgery, model, model + electroacupuncture, and mode + hydergine groups. Mice in the model + electroacupuncture group were treated through electroacupuncture at the Shenshu (BL 23), Geshu (BL 17), and Baihui (GV 20) acupoints. Mice in the model+hydergine group were intragastrically administered with hydergine (0.77 mg/kg(-1) x day(-1)). The levels of BDNF mRNA expressions in the hippocampus were ana lyzed through a semi-quantitative reverse transcription-polymerase chain reaction assay on days 1 and 7 after the surgeries. BDNF mRNA expressions in the mouse hippocampus of the model group on days 1 and 7 after the surgery were higher than those of the sham surgery group (both P electroacupuncture treatment, BDNF mRNA expression in the mouse hippocampus of the model + electroacupuncture group was significantly elevated compared with the model group (both P 0.05). Electroacupuncture treatment enhances endogenous BDNF expression, which may improve the survival environment for intracerebral neurons and inhibit the apoptosis of hippocampal cells.

The NO signaling pathway differentially regulates KCC3a and KCC3b mRNA expression.

Science.gov (United States)

Di Fulvio, Mauricio; Lauf, Peter K; Adragna, Norma C

2003-11-01

Nitric oxide (NO) donors and protein kinase G (PKG) acutely up-regulate K-Cl cotransporter-1 and -3 (KCC1 and KCC3) mRNA expression in vascular smooth muscle cells (VSMCs). Here, we report the presence, relative abundance, and regulation by sodium nitroprusside (SNP) of the novel KCC3a and KCC3b mRNAs, in primary cultures of rat VSMCs. KCC3a and KCC3b mRNAs were expressed in an approximate 3:1 ratio, as determined by semiquantitative RT-PCR analysis. SNP as well as YC-1 and 8-Br-cGMP, a NO-independent stimulator of soluble guanylyl cyclase (sGC) and PKG, respectively, increased KCC3a and KCC3b mRNA expression by 2.5-fold and 8.1-fold in a time-dependent manner, following a differential kinetics. Stimulation of the NO/sGC/PKG signaling pathway with either SNP, YC-1, or 8-Br-cGMP decreased the KCC3a/KCC3b ratio from 3.0+/-0.4 to 0.9+/-0.1. This is the first report on a differential regulation by the NO/sGC/PKG signaling pathway of a cotransporter and of KCC3a and KCC3b mRNA expression.

Freund's adjuvant-induced inflammation: clinical findings and its effect on hepcidin mRNA expression in horses

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José P. Oliveira-Filho

2014-01-01

Full Text Available Hypoferremia observed during systemic inflammatory disorders is regulated by hepcidin. Hepcidin up-regulation is particularly important during acute inflammation, as it restricts the availability of iron, which is necessary for pathogenic microorganism growth before adaptive immunity occurs. The aim of this study was to evaluate the clinical findings and hepatic hepcidin mRNA expression in horses using a Freund's complete adjuvant (FCA model of inflammation. The expression of hepcidin mRNA in the liver was determined in healthy horses following two intramuscular injections of FCA at 0 h and 12 h. Plasma iron and fibrinogen concentrations were measured at multiple time points between 0 h and 240 h post-FCA injection (PI. Hepcidin mRNA expression was determined by RT-qPCR using liver biopsy samples performed at 0 h (control, 6 h and 18 h PI. The mean plasma fibrinogen level was significantly different from the control values only between 120 and 216 h PI. The mean plasma iron level was significantly lower than the control between 16 and 72 h PI, reaching the lowest levels at 30 h PI (33 % of the initial value, and returned to the reference value from 96 h PI to the end of the experiment. Hepcidin mRNA expression increased at 6 h PI and remained high at 18 h PI. The iron plasma concentration was an earlier indicator of inflammatory processes in horses when compared with fibrinogen and might be useful for the early detection of inflammation in the horse. FCA administration caused the rapid onset of hypoferremia, and this effect was likely the result of up-regulated hepatic hepcidin gene expression. This study emphasizes the importance of hepcidin and iron metabolism during inflammation in horses.

Purification and characterization of a CkTLP protein from Cynanchum komarovii seeds that confers antifungal activity.

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Qinghua Wang

Full Text Available BACKGROUND: Cynanchum komarovii Al Iljinski is a desert plant that has been used as analgesic, anthelminthic and antidiarrheal, but also as a herbal medicine to treat cholecystitis in people. We have found that the protein extractions from C. komarovii seeds have strong antifungal activity. There is strong interest to develop protein medication and antifungal pesticides from C. komarovii for pharmacological or other uses. METHODOLOGY/PRINCIPAL FINDINGS: An antifungal protein with sequence homology to thaumatin-like proteins (TLPs was isolated from C. komarovii seeds and named CkTLP. The three-dimensional structure prediction of CkTLP indicated the protein has an acid cleft and a hydrophobic patch. The protein showed antifungal activity against fungal growth of Verticillium dahliae, Fusarium oxysporum, Rhizoctonia solani, Botrytis cinerea and Valsa mali. The full-length cDNA was cloned by RT-PCR and RACE-PCR according to the partial protein sequences obtained by nanoESI-MS/MS. The real-time PCR showed the transcription level of CkTLP had a significant increase under the stress of abscisic acid (ABA, salicylic acid (SA, methyl jasmonate (MeJA, NaCl and drought, which indicates that CkTLP may play an important role in response to abiotic stresses. Histochemical staining showed GUS activity in almost the whole plant, especially in cotyledons, trichomes and vascular tissues of primary root and inflorescences. The CkTLP protein was located in the extracellular space/cell wall by CkTLP::GFP fusion protein in transgenic Arabidopsis. Furthermore, over-expression of CkTLP significantly enhanced the resistance of Arabidopsis against V. dahliae. CONCLUSIONS/SIGNIFICANCE: The results suggest that the CkTLP is a good candidate protein or gene for contributing to the development of disease-resistant crops.

Differential regulation of amyloid-β-protein mRNA expression within hippocampal neuronal subpopulations in Alzheimer disease

International Nuclear Information System (INIS)

Higgins, G.A.; Lewis, D.A.; Bahmanyar, S.; Goldgaber, D.; Gajdusek, D.C.; Young, W.G.; Morrison, J.H.; Wilson, M.C.

1988-01-01

The authors have mapped the neuroanatomical distribution of amyloid-β-protein mRNA within neuronal subpopulations of the hippocampal formation in the cynomolgus monkey (Macaca fascicularis), normal aged human, and patients with Alzheimer disease. Amyloid-β-protein mRNA appears to be expressed in all hippocampal neurons, but at different levels of abundance. In the central nervous system of monkey and normal aged human, image analysis shows that neurons of the dentate gyrus and cornu Ammonis fields contain a 2.5-times-greater hybridization signal than is present in neurons of the subiculum and entorhinal cortex. In contrast, in the Alzheimer disease hippocampal formation, the levels of amyloid-β-protein mRNA in the cornu Ammonis field 3 and parasubiculum are equivalent. These findings suggest that within certain neuronal subpopulations cell type-specific regulation of amyloid-β-protein gene expression may be altered in Alzheimer disease

68Ga-DOTATOC PET/CT and somatostatin receptor (sst1-sst5) expression in normal human tissue: correlation of sst2 mRNA and SUVmax

International Nuclear Information System (INIS)

Boy, Christian; Poeppel, Thorsten D.; Jentzen, Walter; Brandau, Wolfgang; Bockisch, Andreas; Heusner, Till A.; Antoch, Gerald; Redmann-Bischofs, Anja; Unger, Nicole; Mann, Klaus; Petersenn, Stephan

2011-01-01

By targeting somatostatin receptors (sst) radiopeptides have been established for both diagnosis and therapy. For physiologically normal human tissues the study provides a normative database of maximum standardized uptake value (SUV max ) and sst mRNA. A total of 120 patients were subjected to diagnostic 68 Ga-DOTATOC positron emission tomography (PET)/CT (age range 19-83 years). SUV max values were measured in physiologically normal tissues defined by normal morphology, absence of surgical intervention and absence of metastatic spread during clinical follow-up. Expression of sst subtypes (sst1-sst5) was measured independently in pooled adult normal human tissue by real-time reverse transcriptase polymerase chain reaction (RT-PCR). SUV max revealed a region-specific pattern (e.g., mean ± SD, spleen 31.1 ± 10.9, kidney 16.9 ± 5.3, liver 12.8 ± 3.6, stomach 7.0 ± 3.1, head of pancreas 6.2 ± 2.3, small bowel 4.8 ± 1.8, thyroid 4.7 ± 2.2, bone 3.9 ± 1.3, large bowel 2.9 ± 0.8, muscle 2.1 ± 0.5, parotid gland 1.9 ± 0.6, axillary lymph node 0.8 ± 0.3 and lung 0.7 ± 0.3). SUV max was age independent. Gender differences were evident within the thyroid (female/male: 3.7 ± 1.6/5.5 ± 2.4, p max values exclusively correlated with sst2 expression (r = 0.846, p max with the expression of the other four subtypes. In normal human tissues 68 Ga-DOTATOC imaging has been related to the expression of sst2 at the level of mRNA. The novel normative database may improve diagnostics, monitoring and therapy of sst-expressing tumours or inflammation on a molecular basis. (orig.)

Sodium arsenite-induced inhibition of cell proliferation is related to inhibition of IL-2 mRNA expression in mouse activated T cells

Energy Technology Data Exchange (ETDEWEB)

Conde, Patricia; Acosta-Saavedra, Leonor C.; Calderon-Aranda, Emma S. [Centro de Investigacion y de Estudios Avanzados, CINVESTAV, Seccion Toxicologia, P.O. Box 14-740, Mexico, D.F. (Mexico); Goytia-Acevedo, Raquel C. [Universidad Juarez del Estado de Durango, Facultad de Medicina, Gomez Palacio, Durango (Mexico)

2007-04-15

A proposed mechanism for the As-induced inhibition of cell proliferation is the inhibition of IL-2 secretion. However, the effects of arsenite on IL-2 mRNA expression or on the ERK pathway in activated-T cells have not yet been described. We examined the effect of arsenite on IL-2 mRNA expression, cell activation and proliferation in PHA-stimulated murine lymphocytes. Arsenite (1 and 10 {mu}M) decreased IL-2 mRNA expression, IL-2 secretion and cell proliferation. Arsenite (10 {mu}M) strongly inhibited ERK-phosphorylation. However, the partial inhibition (50%) of IL-2 mRNA produced by 1 {mu}M, consistent with the effects on IL-2 secretion and cell proliferation, could not be explained by the inhibition of ERK-phosphorylation, which was not affected at this concentration. The inhibition of IL-2 mRNA expression caused by 1 {mu}M could be associated to effects on pathways located downstream or parallel to ERK. Arsenite also decreased early activation (surface CD69{sup +} expression) in both CD4{sup +} and CD8{sup +}, and decreased total CD8{sup +} count without significantly affecting CD4{sup +}, supporting that the cellular immune response mediated by cytotoxic T cells is an arsenic target. Thus, our results suggest that arsenite decreases IL-2 mRNA levels and T-cell activation and proliferation. However, further studies on the effects of arsenite on IL-2 gene transcription and IL-2 mRNA stability are needed. (orig.)

The effects of valproic acid on the mRNA expression of Natriuretic ...

African Journals Online (AJOL)

Mona Hajikazemi

2017-04-28

Apr 28, 2017 ... Real Time RT-PCR was used to quantify differential mRNA expression of NPR-A and KCNQ1 genes. Two-way ANOVA and bonferroni post-tests were used to analyze data statistically. Results: We showed that VPA treatment inhibits the growth of SW-480 cells more efficiently compared to. HT-29. NPR-A ...

The Expression of mRNA LMP1 Epstein-Barr Virus from FFPE Tumour Biopsy: a Potential Biomarker of Nasopharyngeal Carcinoma Diagnosis

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Daniel Joko Wahyono

2017-07-01

Full Text Available Nasopharyngeal carcinoma (NPC is a multifactorial disease that is endemic geographically in the world. Indonesian population has a highly incidence rate that is 6.2/100,000 people year. The pathogenesis of NPC is more directly reflected by carcinoma-specific viral transcriptional activity at the site of primary tumour. Epstein-Barr virus (EBV infection in NPC is reflected by the expression of EBV latent and lytic gene. In fact, mRNA Latent Membrane Protein 1 (LMP1 EBV expression was an important latent infection biomarker. The aim of this study was to determine a potential use of relative expression of mRNA LMP1 EBV from formalin-fixed paraffin embedded (FFPE tumour biopsy in NPC as a tumour biomarker. This reseach design was a cross sectional study. The samples were the archived specimens of FFPE tumour biopsy from NPC WHO-3 patient which were collected from untreated patients from 2014 in the Department of Pathology Anatomy, Prof. dr. Margono Soekarjo Hospital, Purwokerto. The expression of mRNA LMP1 EBV expression was determined by RT-PCR technique. The positivity of mRNA LMP1 EBV expression was 51.9%, indicating a moderate positivity. The result proved that the expression of mRNA LMP1 EBV from FFPE NPC WHO-3 tumour biopsy was a potential biomarker of NPC diagnosis. The molecular methods would improved the management of NPC, particularly in the histopathological diagnosis of NPC.

Early-life stress induces persistent alterationsin 5-HT1Areceptor and serotonin transporter mRNA expression in the adultrat brain.

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Javier A. Bravo

2014-04-01

Full Text Available Early-life experience plays a major role in the stress response throughout life. Neonatal maternal separation (MS is an animal model of depression with an altered serotonergic response. We hypothesize that this alteration may be caused by differences in 5-HT1A receptor and serotonin transporter (SERT mRNA expression in brain areas involved in the control of emotions, memory and fear as well as in regions controlling the central serotonergic tone.To test this, Sprague-Dawley rats were subjected to MS for 3h daily during post-natal days 2-12. As control, age matched rats were not separated (NS from their dams. When animals reached adulthood (11-13 weeks brain was extracted and mRNA expression of 5-HT1A receptor in amygdala, hippocampus and dorsal raphé nucleus (DRN and SERT in the DRN was analyzed through in-situ hybridisation.Densitometric analysis revealed that MS increased 5-HT1A receptor mRNA expression in the amygdala, and reduced its expression in the DRN, but no changes were observed in the hippocampus in comparison to NS controls. Also, MS reduced SERT mRNA expression in the DRN when compared to NS rats.These results suggest that early-life stress induces persistent changes in 5-HT1A receptor and SERT mRNA expression in key brain regions involved in the development of stress-related psychiatric disorders. The reduction in SERT mRNA indicates an alteration that is in line with clinical findings such as polymorphic variants in individuals with higher risk of depression. These data may help to understand how early-life stress contributes to the development of mood disorders in adulthood.

CK2 activity is modulated by growth rate in Saccharomyces cerevisiae

International Nuclear Information System (INIS)

Tripodi, Farida; Cirulli, Claudia; Reghellin, Veronica; Marin, Oriano; Brambilla, Luca; Schiappelli, Maria Patrizia; Porro, Danilo; Vanoni, Marco; Alberghina, Lilia; Coccetti, Paola

2010-01-01

Research highlights: → CK2 subunits are nuclear both in glucose and in ethanol growing yeast cells. → CK2 activity is modulated in S. cerevisiae. → CK2 activity is higher in conditions supporting higher growth rates. → V max is higher in faster growing cells, while K m is not affected. -- Abstract: CK2 is a highly conserved protein kinase controlling different cellular processes. It shows a higher activity in proliferating mammalian cells, in various types of cancer cell lines and tumors. The findings presented herein provide the first evidence of an in vivo modulation of CK2 activity, dependent on growth rate, in Saccharomyces cerevisiae. In fact, CK2 activity, assayed on nuclear extracts, is shown to increase in exponential growing batch cultures at faster growth rate, while localization of catalytic and regulatory subunits is not nutritionally modulated. Differences in intracellular CK2 activity of glucose- and ethanol-grown cells appear to depend on both increase in molecule number and k cat . Also in chemostat cultures nuclear CK2 activity is higher in faster growing cells providing the first unequivocal demonstration that growth rate itself can affect CK2 activity in a eukaryotic organism.

Parallel mRNA, proteomics and miRNA expression analysis in cell line models of the intestine.

Science.gov (United States)

O'Sullivan, Finbarr; Keenan, Joanne; Aherne, Sinead; O'Neill, Fiona; Clarke, Colin; Henry, Michael; Meleady, Paula; Breen, Laura; Barron, Niall; Clynes, Martin; Horgan, Karina; Doolan, Padraig; Murphy, Richard

2017-11-07

To identify miRNA-regulated proteins differentially expressed between Caco2 and HT-29: two principal cell line models of the intestine. Exponentially growing Caco-2 and HT-29 cells were harvested and prepared for mRNA, miRNA and proteomic profiling. mRNA microarray profiling analysis was carried out using the Affymetrix GeneChip Human Gene 1.0 ST array. miRNA microarray profiling analysis was carried out using the Affymetrix Genechip miRNA 3.0 array. Quantitative Label-free LC-MS/MS proteomic analysis was performed using a Dionex Ultimate 3000 RSLCnano system coupled to a hybrid linear ion trap/Orbitrap mass spectrometer. Peptide identities were validated in Proteome Discoverer 2.1 and were subsequently imported into Progenesis QI software for further analysis. Hierarchical cluster analysis for all three parallel datasets (miRNA, proteomics, mRNA) was conducted in the R software environment using the Euclidean distance measure and Ward's clustering algorithm. The prediction of miRNA and oppositely correlated protein/mRNA interactions was performed using TargetScan 6.1. GO biological process, molecular function and cellular component enrichment analysis was carried out for the DE miRNA, protein and mRNA lists via the Pathway Studio 11.3 Web interface using their Mammalian database. Differential expression (DE) profiling comparing the intestinal cell lines HT-29 and Caco-2 identified 1795 Genes, 168 Proteins and 160 miRNAs as DE between the two cell lines. At the gene level, 1084 genes were upregulated and 711 were downregulated in the Caco-2 cell line relative to the HT-29 cell line. At the protein level, 57 proteins were found to be upregulated and 111 downregulated in the Caco-2 cell line relative to the HT-29 cell line. Finally, at the miRNAs level, 104 were upregulated and 56 downregulated in the Caco-2 cell line relative to the HT-29 cell line. Gene ontology (GO) analysis of the DE mRNA identified cell adhesion, migration and ECM organization, cellular lipid

Effects of low dose radiation on expressions of ICAM-1 mRNA and protein in kidney of diabetic mice

International Nuclear Information System (INIS)

Zhang Chi; Li Xiaokun; Gong Shouliang; Liu Xiaoju; Zhao Xue; Liu Xiaoju; Zhao Xue; Shen Wenjie; Li Cai; Cai Lu

2010-01-01

Objective: To study the effects of low dose radiation (LDR) on the expressions of intercellular adhesion molecule-1 (ICAM-1) mRNA and protein in kidney of diabetes mellitus (DM) mice and illuminate that anti-inflammation of LDR is a main mechanism for diabetic therapy. Methods: The healthy and right age C57BL/6J mice were divided into 4 groups including control, DM, LDR and DM/LDR. The mice in DM and DM/LDR groups were injected intraperitoneally with streptozocin (STZ) to set up DM models. The mice in DM/LDR and LDR groups were irradiated with 25 mGy every other day for 4 weeks. The expressions of ICAM-1 mRNA and protein in kidney were detected with RT-PCR and Western blotting 2, 4, 8, 12 and 16 weeks after irradiation. Results: The expressions of ICAM-1 mRNA and protein in kidney had no significant difference among 4 groups before LDR (P>0.05). The expressions of ICAM-1 mRNA and protein 2 weeks after irradiation with LDR were higher than those in the other 3 groups (P<0.05). The expressions of ICAM-1 mRNA and protein in the DM/LDR group 4 weeks after irradiation were also significantly higher than those in non-DM groups (P<0.05), but still significantly lower than those in DM group (P<0.05), and the significant differences were kept to 16 weeks after irradiation. But the expressions of ICAM-1 mRNA and protein in LDR group were significantly higher than those in control group (P<0.05). IHC assay showed that the glomerular and tubular in DM and DM/LDR groups were abnormal and the quantities of the positive staining cells were significantly increased compared with non-DM groups. However the damage of glomerular and tubular in DM/LDR was significantly supressed compared with DM group and the positive staining cells were also decreased. Conclusion: Under the circumstance of DM, LDR can significantly decrease the expressions of ICAM-1 mRNA and protein in mouse kidney to relief the inflammation reaction in kidney; but in normal condition, LDR can improve the immunity and

Resorufin: a lead for a new protein kinase CK2 inhibitor

DEFF Research Database (Denmark)

Sandholt, Iben Skjøth; Olsen, Birgitte Brinkmann; Guerra, Barbara

2009-01-01

Screening a natural compound library led to the identification of resorufin as a highly selective and potent inhibitor of protein kinase CK2. Out of 52 kinases tested, only CK2 was inhibited, in contrast to emodin, a structurally related, known CK2 inhibitor that, in addition to CK2, inhibited te...

HFE mRNA expression is responsive to intracellular and extracellular iron loading: short communication.

Science.gov (United States)

Mehta, Kosha J; Farnaud, Sebastien; Patel, Vinood B

2017-10-01

In liver hepatocytes, the HFE gene regulates cellular and systemic iron homeostasis by modulating cellular iron-uptake and producing the iron-hormone hepcidin in response to systemic iron elevation. However, the mechanism of iron-sensing in hepatocytes remain enigmatic. Therefore, to study the effect of iron on HFE and hepcidin (HAMP) expressions under distinct extracellular and intracellular iron-loading, we examined the effect of holotransferrin treatment (1, 2, 5 and 8 g/L for 6 h) on intracellular iron levels, and mRNA expressions of HFE and HAMP in wild-type HepG2 and previously characterized iron-loaded recombinant-TfR1 HepG2 cells. Gene expression was analyzed by real-time PCR and intracellular iron was measured by ferrozine assay. Data showed that in the wild-type cells, where intracellular iron content remained unchanged, HFE expression remained unaltered at low holotransferrin treatments but was upregulated upon 5 g/L (p HFE and HAMP expressions were elevated only at low 1 g/L treatment (p HFE (p HFE mRNA was independently elevated by extracellular and intracellular iron-excess. Thus, it may be involved in sensing both, extracellular and intracellular iron. Repression of HAMP expression under simultaneous intracellular and extracellular iron-loading resembles non-hereditary iron-excess pathologies.

Analysis of mRNA expression of genes related to fatty acids synthesis in goose fatty liver

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Shuxia Xiang

2010-11-01

Full Text Available The aim of our study was to evaluate the effect of overfeeding on mRNA expression levels of genes involved in lipogenesis, in order to understand the mechanism of hepatic stea - tosis in the goose. Using Landes geese (Anser anser and Sichuan White geese (Anser cygnoides as experimental animals, we quantified the mRNA expression of lipogenic genes, acetyl-CoA carboxylase-α (ACCα and fatty acid synthase (FAS, and of two transcription factors, sterol regulatory element-binding proteins- 1 (SREBP-1 and carbohydrate responsive element-binding protein (ChREBP by real-time polymerase chain reaction (RTPCR, and measured the lipid and triglyceride (TG content in the liver and the plasma level of glucose, insulin and TG. Our results indicated that compared to the control group, the overfeeding induced an increase of the lipid and TG content in the liver and also of the plasma insulin and TG concentration in both breeds. However, the plasma glucose level decreased after overfeeding in the Sichuan White goose, and there was no evident change in the Landes goose. Lastly, the mRNA expression of ACCα, FAS, SREBP-1 and ChREBP in the overfed group was lower than in the control group in both breeds. We concluded that the lipogenesis pathway plays a role in overfeeding- induced hepatic steatosis and that the decreased mRNA level of related genes may be the indicator of hepatic steatosis.

The Impact of Ramadan Fasting on SIRT1 mRNA Expression in Peripheral Blood Mononuclear Cells

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Mostafa Haji Molahoseini

2016-11-01

Full Text Available Background:The aim of this study was to evaluate the effect of Ramadan fasting on SIRT1 mRNA expression in healthy men.Islamic Ramadan fasting is a holy religious ceremony that has many spiritual benefits. Additionally, it can be considered as the equivalent of calorie restriction that may affect physical health. The results of previous studies revealed that calorie restriction increases the lifespan in laboratory rodents via increasing the expression of a histone deacetylase named SIRT1. Additionally, SIRT1 is known for its anti-inflammatory properties. Materials and Methods: Overall, 43 men volunteered for participating in this one-group before and after (self-controlled study. Two mL blood samples were taken prior to fasting and at the end of the 30th day of fasting. Routine biochemical tests and SIRT1 mRNA expression analysis were performed. Results: Cholesterol and low-density lipoproteins increase, however, high-density lipoproteins level decreased after Ramadan fasting. The analysis of real-time PCR results revealed that SIRT1 mRNA expression in human peripheral blood mononuclear cells increased 4.63 fold in fasting state in comparison with non-fasting state. Conclusion: Ramadan fasting has a significant effect on SIRT1 gene expression. Considering the immunosuppressive and anti-inflammatory properties of SIRT1, further studies are needed to evaluate the effects of SIRT1 up-regulation on the autoimmune and inflammatory diseases during Ramadan fasting.

PrP mRNA and protein expression in brain and PrP(c) in CSF in Creutzfeldt-Jakob disease MM1 and VV2.

Science.gov (United States)

Llorens, Franc; Ansoleaga, Belén; Garcia-Esparcia, Paula; Zafar, Saima; Grau-Rivera, Oriol; López-González, Irene; Blanco, Rosi; Carmona, Margarita; Yagüe, Jordi; Nos, Carlos; Del Río, José Antonio; Gelpí, Ellen; Zerr, Inga; Ferrer, Isidre

2013-01-01

Creutzfeldt-Jakob disease (CJD) is a heterogenic neurodegenerative disorder associated with abnormal post-translational processing of cellular prion protein (PrP(c)). CJD displays distinctive clinical and pathological features which correlate with the genotype at the codon 129 (methionine or valine: M or V respectively) in the prion protein gene and with size of the protease-resistant core of the abnormal prion protein PrP(sc) (type 1: 20/21 kDa and type 2: 19 kDa). MM1 and VV2 are the most common sporadic CJD (sCJD) subtypes. PrP mRNA expression levels in the frontal cortex and cerebellum are reduced in sCJD in a form subtype-dependent. Total PrP protein levels and PrP(sc) levels in the frontal cortex and cerebellum accumulate differentially in sCJD MM1 and sCJD VV2 with no relation between PrP(sc) deposition and spongiform degeneration and neuron loss, but with microgliosis, and IL6 and TNF-α response. In the CSF, reduced PrP(c), the only form present in this compartment, occurs in sCJD MM1 and VV2. PrP mRNA expression is also reduced in the frontal cortex in advanced stages of Alzheimer disease, Lewy body disease, progressive supranuclear palsy, and frontotemporal lobe degeneration, but PrP(c) levels in brain varies from one disease to another. Reduced PrP(c) levels in CSF correlate with PrP mRNA expression in brain, which in turn reflects severity of degeneration in sCJD.

CK1α ablation in keratinocytes induces p53-dependent, sunburn-protective skin hyperpigmentation.

Science.gov (United States)

Chang, Chung-Hsing; Kuo, Che-Jung; Ito, Takamichi; Su, Yu-Ya; Jiang, Si-Tse; Chiu, Min-Hsi; Lin, Yi-Hsiung; Nist, Andrea; Mernberger, Marco; Stiewe, Thorsten; Ito, Shosuke; Wakamatsu, Kazumasa; Hsueh, Yi-An; Shieh, Sheau-Yann; Snir-Alkalay, Irit; Ben-Neriah, Yinon

2017-09-19

Casein kinase 1α (CK1α), a component of the β-catenin destruction complex, is a critical regulator of Wnt signaling; its ablation induces both Wnt and p53 activation. To characterize the role of CK1α (encoded by Csnk1a1 ) in skin physiology, we crossed mice harboring floxed Csnk1a1 with mice expressing K14-Cre-ER T2 to generate mice in which tamoxifen induces the deletion of Csnk1a1 exclusively in keratinocytes [single-knockout (SKO) mice]. As expected, CK1α loss was accompanied by β-catenin and p53 stabilization, with the preferential induction of p53 target genes, but phenotypically most striking was hyperpigmentation of the skin, importantly without tumorigenesis, for at least 9 mo after Csnk1a1 ablation. The number of epidermal melanocytes and eumelanin levels were dramatically increased in SKO mice. To clarify the putative role of p53 in epidermal hyperpigmentation, we established K14-Cre-ER T2 CK1α/p53 double-knockout (DKO) mice and found that coablation failed to induce epidermal hyperpigmentation, demonstrating that it was p53-dependent. Transcriptome analysis of the epidermis revealed p53-dependent up-regulation of Kit ligand (KitL). SKO mice treated with ACK2 (a Kit-neutralizing antibody) or imatinib (a Kit inhibitor) abrogated the CK1α ablation-induced hyperpigmentation, demonstrating that it requires the KitL/Kit pathway. Pro-opiomelanocortin (POMC), a precursor of α-melanocyte-stimulating hormone (α-MSH), was not activated in the CK1α ablation-induced hyperpigmentation, which is in contrast to the mechanism of p53-dependent UV tanning. Nevertheless, acute sunburn effects were successfully prevented in the hyperpigmented skin of SKO mice. CK1α inhibition induces skin-protective eumelanin but no carcinogenic pheomelanin and may therefore constitute an effective strategy for safely increasing eumelanin via UV-independent pathways, protecting against acute sunburn.

Crystal structure of human protein kinase CK2

DEFF Research Database (Denmark)

Niefind, K; Guerra, B; Ermakowa, I

2001-01-01

The crystal structure of a fully active form of human protein kinase CK2 (casein kinase 2) consisting of two C-terminally truncated catalytic and two regulatory subunits has been determined at 3.1 A resolution. In the CK2 complex the regulatory subunits form a stable dimer linking the two catalyt...... as a docking partner for various protein kinases. Furthermore it shows an inter-domain mobility in the catalytic subunit known to be functionally important in protein kinases and detected here for the first time directly within one crystal structure.......The crystal structure of a fully active form of human protein kinase CK2 (casein kinase 2) consisting of two C-terminally truncated catalytic and two regulatory subunits has been determined at 3.1 A resolution. In the CK2 complex the regulatory subunits form a stable dimer linking the two catalytic...... subunits, which make no direct contact with one another. Each catalytic subunit interacts with both regulatory chains, predominantly via an extended C-terminal tail of the regulatory subunit. The CK2 structure is consistent with its constitutive activity and with a flexible role of the regulatory subunit...

Lipopolysaccharide inhibits colonic biotin uptake via interference with membrane expression of its transporter: a role for a casein kinase 2-mediated pathway.

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Lakhan, Ram; Said, Hamid M

2017-04-01

Biotin (vitamin B7), an essential micronutrient for normal cellular functions, is obtained from both dietary sources as well as gut microbiota. Absorption of biotin in both the small and large intestine is via a carrier-mediated process that involves the sodium-dependent multivitamin transporter (SMVT). Although different physiological and molecular aspects of intestinal biotin uptake have been delineated, nothing is known about the effect of LPS on the process. We addressed this issue using in vitro (human colonic epithelial NCM460 cells) and in vivo (mice) models of LPS exposure. Treating NCM460 cells with LPS was found to lead to a significant inhibition in carrier-mediated biotin uptake. Similarly, administration of LPS to mice led to a significant inhibition in biotin uptake by native colonic tissue. Although no changes in total cellular SMVT protein and mRNA levels were observed, LPS caused a decrease in the fraction of SMVT expressed at the cell surface. A role for casein kinase 2 (CK2) (whose activity was also inhibited by LPS) in mediating the endotoxin effects on biotin uptake and on membrane expression of SMVT was suggested by findings that specific inhibitors of CK2, as well as mutating the putative CK2 phosphorylation site (Thr 78 Ala) in the SMVT protein, led to inhibition in biotin uptake and membrane expression of SMVT. This study shows for the first time that LPS inhibits colonic biotin uptake via decreasing membrane expression of its transporter and that these effects likely involve a CK2-mediated pathway.

Raf-1/CK2 and RhoA/ROCK signaling promote TNF-α-mediated endothelial apoptosis via regulating vimentin cytoskeleton.

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Yang, Lifeng; Tang, Lian; Dai, Fan; Meng, Guoliang; Yin, Runting; Xu, Xiaole; Yao, Wenjuan

2017-08-15

Both RhoA/ROCK and Raf-1/CK2 pathway play essential roles in cell proliferation, apoptosis, differentiation, and multiple other common cellular functions. We previously reported that vimentin is responsible for TNF-α-induced cell apoptosis. Herein, we investigated the regulation of RhoA/ROCK and Raf-1/CK2 signaling on vimentin filaments and endothelial apoptosis mediated by TNF-α. Treatment with TNF-α significantly induced the activation of RhoA and ROCK, and the expression of ROCK1. RhoA deficiency could obviously inhibit ROCK activation and ROCK1 expression induced by TNF-α. Both RhoA deficiency and ROCK activity inhibition (Y-27632) greatly inhibited endothelial apoptosis and preserved cell viability in TNF-α-induced human umbilical vein endothelial cells (HUVECs). Also vimentin phosphorylation and the remodeling of vimentin or phospho-vimentin induced by TNF-α were obviously attenuated by RhoA suppression and ROCK inhibition. TNF-α-mediated vimentin cleavage was significantly inhibited by RhoA suppression and ROCK inhibition through decreasing the activation of caspase3 and 8. Furthermore, TNF-α treatment greatly enhanced the activation of Raf-1. Suppression of Raf-1 or CK2 by its inhibitor (GW5074 or TBB) blocked vimentin phosphorylation, remodeling and endothelial apoptosis, and preserved cell viability in TNF-α-induced HUVECs. However, Raf-1 inhibition showed no significant effect on TNF-α-induced ROCK expression and activation, suggesting that the regulation of Raf-1/CK2 signaling on vimentin was independent of ROCK. Taken together, these results indicate that both RhoA/ROCK and Raf-1/CK2 pathway are responsible for TNF-α-mediated endothelial cytotoxicity via regulating vimentin cytoskeleton. Copyright © 2017 Elsevier B.V. All rights reserved.

EST Table: CK561471 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK561471 rswpb0_004007.y1 10/09/29 77 %/177 aa ref|XP_001656149.1| fetal alzheimer ...antigen, falz [Aedes aegypti] gb|EAT35210.1| fetal alzheimer antigen, falz [Aedes aegypti] 10/09/01 71 %/177... gi|189240808|ref|XP_001811424.1| PREDICTED: similar to fetal alzheimer antigen, falz [Tribolium castaneum] CK536202 swp ...

Expression of a serine protease (motopsin PRSS12) mRNA in the mouse brain: in situ hybridization histochemical study.

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Iijima, N; Tanaka, M; Mitsui, S; Yamamura, Y; Yamaguchi, N; Ibata, Y

1999-03-20

Serine proteases are considered to play several important roles in the brain. In an attempt to find novel brain-specific serine proteases (BSSPs), motopsin (PRSS-12) was cloned from a mouse brain cDNA library by polymerase chain reaction (PCR). Northern blot analysis demonstrated that the postnatal 10-day mouse brain contained the most amount of motopsin mRNA. At this developmental stage, in situ hybridization histochemistry showed that motopsin mRNA was specifically expressed in the following regions: cerebral cortical layers II/III, V and VIb, endopiriform cortex and the limbic system, particularly in the CA1 region of the hippocampal formation. In addition, in the brainstem, the oculomotor nucleus, trochlear nucleus, mecencephalic and motor nuclei of trigeminal nerve (N), abducens nucleus, facial nucleus, nucleus of the raphe pontis, dorsoral motor nucleus of vagal N, hypoglossal nucleus and ambiguus nucleus showed motopsin mRNA expression. Expression was also found in the anterior horn of the spinal cord. The above findings strongly suggest that neurons in almost all motor nuclei, particularly in the brainstem and spinal cord, express motopsin mRNA, and that motopsin seems to have a close relation to the functional role of efferent neurons. Copyright 1999 Elsevier Science B.V.

Small molecules CK-666 and CK-869 inhibit actin-related protein 2/3 complex by blocking an activating conformational change.

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Hetrick, Byron; Han, Min Suk; Helgeson, Luke A; Nolen, Brad J

2013-05-23

Actin-related protein 2/3 (Arp2/3) complex is a seven-subunit assembly that nucleates branched actin filaments. Small molecule inhibitors CK-666 and CK-869 bind to Arp2/3 complex and inhibit nucleation, but their modes of action are unknown. Here, we use biochemical and structural methods to determine the mechanism of each inhibitor. Our data indicate that CK-666 stabilizes the inactive state of the complex, blocking movement of the Arp2 and Arp3 subunits into the activated filament-like (short pitch) conformation, while CK-869 binds to a serendipitous pocket on Arp3 and allosterically destabilizes the short pitch Arp3-Arp2 interface. These results provide key insights into the relationship between conformation and activity in Arp2/3 complex and will be critical for interpreting the influence of the inhibitors on actin filament networks in vivo. Copyright © 2013 Elsevier Ltd. All rights reserved.

Analysis of p130 protein and mRNA expression in ten patients with uterine papillary serous carcinoma

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Shao-ting XU

2011-11-01

Full Text Available Objective To examine p130 protein and mRNA expression in uterine papillary serous carcinoma(UPSC and their clinical and pathologic significance.Methods A total of 10 UPSC patients(Stage I were included,with 10 cases of high-level endometrial carcinoma of the same stage taken as the control group and 10 cases of normal proliferative stage endometrium(EM taken as the disease control group.The level of p130 protein expression was determined by hematoxylin and eosin staining,microscopic observation,and immunohistochemistry,whereas the p130 mRNA levels were examined through real-time quantitative reverse transcriptase polymerase chain reaction.The clinicopathologic analysis was carried out in combination with clinical data.Results The p130 protein and p130 mRNA expression levels in the UPSC group(0.46±0.01 and 0.56±0.06,respectively were apparently less than that of the normal proliferative stage endometrium group(0.91±0.04 and 2.81±0.40,respectively;P < 0.01 and also less than those in high-level endometrial carcinoma(P < 0.05.Clinicopathologic analysis shows that all patients are post-menopausal women with symptoms of irregular vaginal bleeding and the average tumor size was 7.5cm(range: 1.2-14.8cm.The pathologic features are same as that of high-level ovarian papillary serous carcinoma.Conclusion Reduced p130 protein and p130 mRNA expression in UPSC might correlate with poor prognosis in UPSC patients.

A case of spuriously high CK-MB: Contemplate beyond cardiac

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Rishard Abdul

2017-10-01

Full Text Available Creatine kinase-MB (CK-MB is a widely tested enzyme in cardiac disease and thus has important clinical implications. We relate the scenario of a young patient presenting with chest pain whose CK-MB levels remained inordinately elevated despite a normal total CK level, resolution of symptoms and exclusion of coronary artery disease. Further analysis by electrophoresis revealed the presence of a rare molecular variant of creatine kinase: macro-CK type 1. While this may be a benign finding, there are sparse data demonstrating an association with non-cardiac pathologies such as malignancy, endocrinopathies, connective tissue and autoimmune diseases. Thus, after further exclusion of these associated conditions, the patient was reassured on the likely benign nature of macro-CK in this case.

Bile acid flux through portal but not peripheral veins inhibits CYP7A1 expression without involvement of ileal FGF19 in rabbits.

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Shang, Quan; Guo, Grace L; Honda, Akira; Shi, Daniel; Saumoy, Monica; Salen, Gerald; Xu, Guorong

2014-08-15

It was proposed that CYP7A1 expression is suppressed through the gut-hepatic signaling pathway fibroblast growth factor (FGF) 15/19-fibroblast growth factor receptor 4, which is initiated by activation of farnesoid X receptor in the intestine rather than in the liver. The present study tested whether portal bile acid flux alone without ileal FGF19 could downregulate CYP7A1 expression in rabbits. A rabbit model was developed by infusing glycodeoxycholic acid (GDCA) through the splenic vein to bypass ileal FGF19. Study was conducted in four groups of rabbits: control; bile fistula + bovine serum albumin solution perfusion (BF); BF + GDCA (by portal perfusion); and BF + GDCA-f (by femoral perfusion). Compared with only BF, BF + GDCA (6 h portal perfusion) suppressed CYP7A1 mRNA, whereas BF + GDCA-f (via femoral vein) with the same perfusion rate of GDCA did not show inhibitory effects. Meanwhile, there was a decrease in ileal FGF19 expression and portal FGF19 protein levels, but an equivalent increase in biliary bile acid outputs in both GDCA perfusion groups. This study demonstrated that portal bile acid flux alone downregulated CYP7A1 expression with diminished FGF19 expression and protein levels, whereas the same bile acid flux reaching the liver through the hepatic artery via femoral vein had no inhibitory effect on CYP7A1. We propose that bile acid flux through the portal venous system may be a kind of "intestinal factor" that suppresses CYP7A1 expression. Copyright © 2014 the American Physiological Society.

Rift Valley fever virus NS{sub S} gene expression correlates with a defect in nuclear mRNA export

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Copeland, Anna Maria; Van Deusen, Nicole M.; Schmaljohn, Connie S., E-mail: Connie.s.schmaljohn.civ@mail.mil

2015-12-15

We investigated the localization of host mRNA during Rift Valley fever virus (RVFV) infection. Fluorescence in situ hybridization revealed that infection with RVFV altered the localization of host mRNA. mRNA accumulated in the nuclei of RVFV-infected but not mock-infected cells. Further, overexpression of the NS{sub S} gene, but not the N, G{sub N} or NS{sub M} genes correlated with mRNA nuclear accumulation. Nuclear accumulation of host mRNA was not observed in cells infected with a strain of RVFV lacking the gene encoding NS{sub S}, confirming that expression of NS{sub S} is likely responsible for this phenomenon. - Highlights: • Rift Valley fever virus (RVFV) infection alters the localization of host mRNA. • mRNA accumulates in the nuclei of RVFV-infected but not mock-infected cells. • NS{sub S} is likely responsible for mRNA relocalization to the nucleus.

The functional interplay between protein kinase CK2 and CCA1 transcriptional activity is essential for clock temperature compensation in Arabidopsis.

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Sergi Portolés

2010-11-01

Full Text Available Circadian rhythms are daily biological oscillations driven by an endogenous mechanism known as circadian clock. The protein kinase CK2 is one of the few clock components that is evolutionary conserved among different taxonomic groups. CK2 regulates the stability and nuclear localization of essential clock proteins in mammals, fungi, and insects. Two CK2 regulatory subunits, CKB3 and CKB4, have been also linked with the Arabidopsis thaliana circadian system. However, the biological relevance and the precise mechanisms of CK2 function within the plant clockwork are not known. By using ChIP and Double-ChIP experiments together with in vivo luminescence assays at different temperatures, we were able to identify a temperature-dependent function for CK2 modulating circadian period length. Our study uncovers a previously unpredicted mechanism for CK2 antagonizing the key clock regulator CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1. CK2 activity does not alter protein accumulation or subcellular localization but interferes with CCA1 binding affinity to the promoters of the oscillator genes. High temperatures enhance the CCA1 binding activity, which is precisely counterbalanced by the CK2 opposing function. Altering this balance by over-expression, mutation, or pharmacological inhibition affects the temperature compensation profile, providing a mechanism by which plants regulate circadian period at changing temperatures. Therefore, our study establishes a new model demonstrating that two opposing and temperature-dependent activities (CCA1-CK2 are essential for clock temperature compensation in Arabidopsis.

Alterations in Lipoxygenase and Cyclooxygenase-2 Catalytic Activity and mRNA Expression in Prostate Carcinoma

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Scott B. Shappell

2001-01-01

Full Text Available Recent studies in prostate tissues and especially cell lines have suggested roles for arachidonic acid (AA metabolizing enzymes in prostate adenocarcinoma (Pca development or progression. The goal of this study was to more fully characterize lipoxygenase (LOX and cyclooxygenase-2 (COX-2 gene expression and AA metabolism in benign and malignant prostate using snap-frozen tissues obtained intraoperatively and mRNA analyses and enzyme assays. Formation of 15-hydroxyeicosatetraenoic acid (15-HETE was detected in 23/29 benign samples and 15-LOX-2 mRNA was detected in 21/25 benign samples. In pairs of pure benign and Pca from the same patients, 15-HETE production and 15-LOX-2 mRNA were reduced in Pca versus benign in 9/14 (P=.04 and 14/17 (P=.002, respectively. Under the same conditions, neither 5HETE nor 12-HETE formation was detectable in 29 benign and 24 tumor samples; with a more sensitive assay, traces were detected in some samples, but there was no clear association with tumor tissue. COX-2 mRNA was detected by nuclease protection assay in 7/16 benign samples and 5/16 tumors. In benign and tumor pairs from 10 patients, COX-2 was higher in tumor versus benign in only 2, with similar results by in situ hybridization. Paraffin immunoperoxidase for COX2 was performed in whole mount sections from 87 additional radical prostatectomy specimens, with strong expression in ejaculatory duct as a positive control and corroboration with in situ hybridization. No immunostaining was detected in benign prostate or tumor in 45% of cases. Greater immunostaining in tumor versus benign was present in only 17% of cases, and correlated with high tumor grade (Gleason score 8 and 9 vs. 5 to 7. In conclusion, reduced 15-LOX-2 expression and 15-HETE formation is the most characteristic alteration of AA metabolism in Pca. Increased 12-HETE and 5-HETE formation in Pca were not discernible. Increased COX-2 expression is not a typical abnormality in Pca in general, but

HDAC inhibitors repress BARD1 isoform expression in acute myeloid leukemia cells via activation of miR-19a and/or b.

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Ilaria Lepore

Full Text Available Over the past years BARD1 (BRCA1-associated RING domain 1 has been considered as both a BRCA1 (BReast Cancer susceptibility gene 1, early onset interactor and tumor suppressor gene mutated in breast and ovarian cancers. Despite its role as a stable heterodimer with BRCA1, increasing evidence indicates that BARD1 also has BRCA1-independent oncogenic functions. Here, we investigate BARD1 expression and function in human acute myeloid leukemias and its modulation by epigenetic mechanism(s and microRNAs. We show that the HDACi (histone deacetylase inhibitor Vorinostat reduces BARD1 mRNA levels by increasing miR-19a and miR-19b expression levels. Moreover, we identify a specific BARD1 isoform, which might act as tumor diagnostic and prognostic markers.

Integrated Analysis of Long Noncoding RNA and mRNA Expression Profile in Advanced Laryngeal Squamous Cell Carcinoma.

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Ling Feng

Full Text Available Long non-coding RNA (lncRNA plays an important role in tumorigenesis. However, the expression pattern and function of lncRNAs in laryngeal squamous cell carcinoma (LSCC are still unclear. To investigate the aberrantly expressed lncRNAs and mRNAs in advanced LSCC, we screened lncRNA and mRNA expression profiles in 9 pairs of primary Stage IVA LSCC tissues and adjacent non-neoplastic tissues by lncRNA and mRNA integrated microarrays. Gene Ontology and pathway analysis were performed to find out the significant function and pathway of the differentially expressed mRNAs, gene-gene functional interaction network and ceRNA network were constructed to select core mRNAs, and lncRNA-mRNA expression correlation network was built to identify the interactions between lncRNA and mRNA. qRT-PCR was performed to further validate the expressions of selected lncRNAs and mRNAs in advanced LSCC. We found 1459 differentially expressed lncRNAs and 2381 differentially expressed mRNAs, including 846 up-regulated lncRNAs and 613 down-regulated lncRNAs, 1542 up-regulated mRNAs and 839 down-regulated mRNAs. The mRNAs ITGB1, HIF1A, and DDIT4 were selected as core mRNAs, which are mainly involved in biological processes, such as matrix organization, cell cycle, adhesion, and metabolic pathway. LncRNA-mRNA expression correlation network showed LncRNA NR_027340, MIR31HG were positively correlated with ITGB1, HIF1A respectively. LncRNA SOX2-OT was negatively correlated with DDIT4. qRT-PCR further validated the expression of these lncRNAs and mRNAs. The work provides convincing evidence that the identified lncRNAs and mRNAs are potential biomarkers in advanced LSCC for further future studies.

Significance of the BRAF mRNA Expression Level in Papillary Thyroid Carcinoma: An Analysis of The Cancer Genome Atlas Data.

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Young Jun Chai

Full Text Available BRAFV600E is the most common mutation in papillary thyroid carcinoma (PTC, and it is associated with high-risk prognostic factors. However, the significance of the BRAF mRNA level in PTC remains unknown. We evaluated the significance of BRAF mRNA expression level by analyzing PTC data from The Cancer Genome Atlas (TCGA database.Data from 499 patients were downloaded from the TCGA database. After excluding other PTC variants, we selected 353 cases of classic PTC, including 193 cases with BRAFV600E and 160 cases with the wild-type BRAF. mRNA abundances were measured using RNA-Seq with the Expectation Maximization algorithm.The mean BRAF mRNA level was significantly higher in BRAFV600E patients than in patients with wild-type BRAF (197.6 vs. 179.3, p = 0.031. In wild-type BRAF patients, the mean BRAF mRNA level was higher in cases with a tumor > 2 cm than those with a tumor ≤ 2.0 cm (189.4 vs. 163.8, p = 0.046, and was also higher in cases with lymph node metastasis than in those without lymph node metastasis (188.5 vs. 157.9, p = 0.040. Within BRAFV600E patients, higher BRAF mRNA expression was associated with extrathyroidal extension (186.4 vs. 216.4, p = 0.001 and higher T stage (188.1 vs. 210.2, p = 0.016.A higher BRAF mRNA expression level was associated with tumor aggressiveness in classic PTC regardless of BRAF mutational status. Evaluation of BRAF mRNA level may be helpful in prognostic risk stratification of PTC.

Effects of fasting, temperature, and photoperiod on preproghrelin mRNA expression in Chinese perch.

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Song, Yi; Zhao, Cheng; Liang, Xu-Fang; He, Shan; Tian, Changxu; Cheng, Xiaoyan; Yuan, Xiaochen; Lv, Liyuan; Guo, Wenjie; Xue, Min; Tao, Ya-Xiong

2017-06-01

Preproghrelin, a gut/brain peptide, plays an important role in the regulation of food intake and energy homeostasis in teleost and mammals. In the present study, we obtained the full-length preproghrelin cDNA in Chinese perch. The preproghrelin messenger RNA (mRNA) tissue expression showed that level was much higher in stomach and pituitary than in other tissues. The fasting study showed, after gastric emptying (3-6 h), short-term fasting (6-12 h) increased preproghrelin expression in the stomach. While in the pituitary, fasting reduced preproghrelin expression at 1, 3, 12, and 48 h, presenting state fluctuation of self-adjustment. The temperature study showed that the mRNA expression of preproghrelin was the highest in the brain at 26 °C and highest in the stomach at 32 °C, respectively, with different optimum temperature in these two tissues, reflecting spatiotemporal differences of regulation by central nervous system and peripheral organs. The photoperiod study showed that normal light (11 h of lightness and 13 h of darkness) led to highest preproghrelin expression, both in the brain and in the stomach, than continuous light or continuous dark, proving food intake is adapted to natural photoperiod or normal light in this study. These results all indicated that tissue-specific preproghrelin expression of Chinese perch could be significantly affected by environmental factors. Short-term fasting of 6 h after gastric emptying, 26 °C, and normal light led to higher preproghrelin expression, which indicated potential appetite increase in Chinese perch.

Peroxisome proliferator-activated receptor α (PPARα mRNA expression in human hepatocellular carcinoma tissue and non-cancerous liver tissue

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Kurokawa Tsuyoshi

2011-12-01

Full Text Available Abstract Background Peroxisome proliferator-activated receptor α (PPARα regulates lipid metabolism in the liver. It is unclear, however, how this receptor changes in liver cancer tissue. On the other hand, mouse carcinogenicity studies showed that PPARα is necessary for the development of liver cancer induced by peroxisome proliferators, and the relationship between PPARα and the development of liver cancer have been the focus of considerable attention. There have been no reports, however, demonstrating that PPARα is involved in the development of human liver cancer. Methods The subjects were 10 patients who underwent hepatectomy for hepatocellular carcinoma. We assessed the expression of PPARα mRNA in human hepatocellular carcinoma tissue and non-cancerous tissue, as well as the expression of target genes of PPARα, carnitine palmitoyltransferase 1A and cyclin D1 mRNAs. We also evaluated glyceraldehyde 3-phosphate dehydrogenase, a key enzyme in the glycolytic system. Results The amounts of PPARα, carnitine palmitoyltransferase 1A and glyceraldehyde 3-phosphate dehydrogenase mRNA in cancerous sections were significantly increased compared to those in non-cancerous sections. The level of cyclin D1 mRNA tends to be higher in cancerous than non-cancerous sections. Although there was a significant correlation between the levels of PPARα mRNA and cyclin D1 mRNA in both sections, however the correlation was higher in cancerous sections. Conclusion The present investigation indicated increased expression of PPARα mRNA and mRNAs for PPARα target genes in human hepatocellular carcinoma. These results might be associated with its carcinogenesis and characteristic features of energy production.

Effect of electroacupuncture on TRPM7 mRNA expression after cerebral ischemia/reperfusion in rats via TrkA pathway.

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Zhao, Li; Shi, Jing; Sun, Ning; Tian, Shunlian; Meng, Xianfang; Liu, Xiaochun; Li, Lingli

2005-01-01

The effect of electroacupuncture (EA) on TRPM7 mRNA expression of focal cerebral ischemia in rats and further the role of EA in the relationship between TRPM7 and trkA pathway was investigated. Thirty SD rats were randomly divided into 5 groups : normal group, ischemia/reperfusion group, EA treated group (ischemic rats with EA treatment), TE infusion group (ischemic rats with EA treatment and TE buffer infusion), AS-ODN group (ischemic rats with EA treatment and antisense trkA oligonucleotide infusion). The stroke animal model was established by the modified method of middle cerebral artery occlusion. Antisense trkA oligonucleotide that blocked NGFs effects was injected into cerebroventricle before EA. The TRPM7 mRNA was detected by RT-PCR method. The results showed that there were low TRPM7 mRNA levels in cortex and hippocampus in normal group. Compared with normal group, TRPM7 mRNA expression was increased significantly in ischemia/reperfusion group (PPM7 mRNA was found in EA treated group in contrast to ischemia/reperfusion group (P<0.05). The expression of TRPM7 mRNA in AS-ODN group was remarkably increased compared with EA treated group and TE infusion group (P<0.05). The results indicated that TRPM7 channels in the ischemic cortex and hippocampus in rats might play a key role in ischemic brain injury. EA could reverse the overexpression of TRPM7 in cerebral ischemia/reperfusion rats. And the inhibitory effect of EA on TRPM7 channels might be through trkA pathway.

The classification of mRNA expression levels by the phosphorylation state of RNAPII CTD based on a combined genome-wide approach

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Tachibana Taro

2011-10-01

Full Text Available Abstract Background Cellular function is regulated by the balance of stringently regulated amounts of mRNA. Previous reports revealed that RNA polymerase II (RNAPII, which transcribes mRNA, can be classified into the pausing state and the active transcription state according to the phosphorylation state of RPB1, the catalytic subunit of RNAPII. However, genome-wide association between mRNA expression level and the phosphorylation state of RNAPII is unclear. While the functional importance of pausing genes is clear, such as in mouse Embryonic Stem cells for differentiation, understanding this association is critical for distinguishing pausing genes from active transcribing genes in expression profiling data, such as microarrays and RNAseq. Therefore, we examined the correlation between the phosphorylation of RNAPII and mRNA expression levels using a combined analysis by ChIPseq and RNAseq. Results We first performed a precise quantitative measurement of mRNA by performing an optimized calculation in RNAseq. We then visualized the recruitment of various phosphorylated RNAPIIs, such as Ser2P and Ser5P. A combined analysis using optimized RNAseq and ChIPseq for phosphorylated RNAPII revealed that mRNA levels correlate with the various phosphorylation states of RNAPII. Conclusions We demonstrated that the amount of mRNA is precisely reflected by the phased phosphorylation of Ser2 and Ser5. In particular, even the most "pausing" genes, for which only Ser5 is phosphorylated, were detectable at a certain level of mRNA. Our analysis indicated that the complexity of quantitative regulation of mRNA levels could be classified into three categories according to the phosphorylation state of RNAPII.

Integrated Analysis of Dysregulated ncRNA and mRNA Expression Profiles in Humans Exposed to Carbon Nanotubes.

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Anna A Shvedova

Full Text Available As the application of carbon nanotubes (CNT in consumer products continues to rise, studies have expanded to determine the associated risks of exposure on human and environmental health. In particular, several lines of evidence indicate that exposure to multi-walled carbon nanotubes (MWCNT could pose a carcinogenic risk similar to asbestos fibers. However, to date the potential markers of MWCNT exposure are not yet explored in humans.In the present study, global mRNA and ncRNA expression profiles in the blood of exposed workers, having direct contact with MWCNT aerosol for at least 6 months (n = 8, were compared with expression profiles of non-exposed (n = 7 workers (e.g., professional and/or technical staff from the same manufacturing facility.Significant changes in the ncRNA and mRNA expression profiles were observed between exposed and non-exposed worker groups. An integrative analysis of ncRNA-mRNA correlations was performed to identify target genes, functional relationships, and regulatory networks in MWCNT-exposed workers. The coordinated changes in ncRNA and mRNA expression profiles revealed a set of miRNAs and their target genes with roles in cell cycle regulation/progression/control, apoptosis and proliferation. Further, the identified pathways and signaling networks also revealed MWCNT potential to trigger pulmonary and cardiovascular effects as well as carcinogenic outcomes in humans, similar to those previously described in rodents exposed to MWCNTs.This study is the first to investigate aberrant changes in mRNA and ncRNA expression profiles in the blood of humans exposed to MWCNT. The significant changes in several miRNAs and mRNAs expression as well as their regulatory networks are important for getting molecular insights into the MWCNT-induced toxicity and pathogenesis in humans. Further large-scale prospective studies are necessary to validate the potential applicability of such changes in mRNAs and miRNAs as prognostic markers

Postmortem mRNA expression patterns in left ventricular myocardial tissues and their implications for forensic diagnosis of sudden cardiac death.

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Son, Gi Hoon; Park, Seong Hwan; Kim, Yunmi; Kim, Ji Yeon; Kim, Jin Wook; Chung, Sooyoung; Kim, Yu-Hoon; Kim, Hyun; Hwang, Juck-Joon; Seo, Joong-Seok

2014-03-01

Sudden cardiac death (SCD), which is primarily caused by lethal heart disorders resulting in structural and arrhythmogenic abnormalities, is one of the prevalent modes of death in most developed countries. Myocardial ischemia, mainly due to coronary artery disease, is the most common type of heart disease leading to SCD. However, postmortem diagnosis of SCD is frequently complicated by obscure histological evidence. Here, we show that certain mRNA species, namely those encoding hemoglobin A1/2 and B (Hba1/2 and Hbb, respectively) as well as pyruvate dehydrogenase kinase 4 (Pdk4), exhibit distinct postmortem expression patterns in the left ventricular free wall of SCD subjects when compared with their expression patterns in the corresponding tissues from control subjects with non-cardiac causes of death. Hba1/2 and Hbb mRNA expression levels were higher in ischemic SCD cases with acute myocardial infarction or ischemic heart disease without recent infarction, and even in cardiac death subjects without apparent pathological signs of heart injuries, than control subjects. By contrast, Pdk4 mRNA was expressed at lower levels in SCD subjects. In conclusion, we found that altered myocardial Hba1/2, Hbb, and Pdk4 mRNA expression patterns can be employed as molecular signatures of fatal cardiac dysfunction to forensically implicate SCD as the primary cause of death.

Hedgehog signaling pathway is active in GBM with GLI1 mRNA expression showing a single continuous distribution rather than discrete high/low clusters.

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Chandra, Vikas; Das, Tapojyoti; Gulati, Puneet; Biswas, Nidhan K; Rote, Sarang; Chatterjee, Uttara; Ghosh, Samarendra N; Deb, Sumit; Saha, Suniti K; Chowdhury, Anup K; Ghosh, Subhashish; Rudin, Charles M; Mukherjee, Ankur; Basu, Analabha; Dhara, Surajit

2015-01-01

Hedgehog (Hh) signaling pathway is a valid therapeutic target in a wide range of malignancies. We focus here on glioblastoma multiforme (GBM), a lethal malignancy of the central nervous system (CNS). By analyzing RNA-sequencing based transcriptomics data on 149 clinical cases of TCGA-GBM database we show here a strong correlation (r = 0.7) between GLI1 and PTCH1 mRNA expression--as a hallmark of the canonical Hh-pathway activity in this malignancy. GLI1 mRNA expression varied in 3 orders of magnitude among the GBM patients of the same cohort showing a single continuous distribution-unlike the discrete high/low-GLI1 mRNA expressing clusters of medulloblastoma (MB). When compared with MB as a reference, the median GLI1 mRNA expression in GBM appeared 14.8 fold lower than that of the "high-Hh" cluster of MB but 5.6 fold higher than that of the "low-Hh" cluster of MB. Next, we demonstrated statistically significant up- and down-regulation of GLI1 mRNA expressions in GBM patient-derived low-passage neurospheres in vitro by sonic hedgehog ligand-enriched conditioned media (shh-CM) and by Hh-inhibitor drug vismodegib respectively. We also showed clinically achievable dose (50 μM) of vismodegib alone to be sufficient to induce apoptosis and cell cycle arrest in these low-passage GBM neurospheres in vitro. Vismodegib showed an effect on the neurospheres, both by down-regulating GLI1 mRNA expression and by inducing apoptosis/cell cycle arrest, irrespective of their relative endogenous levels of GLI1 mRNA expression. We conclude from our study that this single continuous distribution pattern of GLI1 mRNA expression technically puts almost all GBM patients in a single group rather than discrete high- or low-clusters in terms of Hh-pathway activity. That is suggestive of therapies with Hh-pathway inhibitor drugs in this malignancy without a need for further stratification of patients on the basis of relative levels of Hh-pathway activity among them.

Small, synthetic, GC-rich mRNA stem-loop modules 5' proximal to the AUG start-codon predictably tune gene expression in yeast.

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Lamping, Erwin; Niimi, Masakazu; Cannon, Richard D

2013-07-29

A large range of genetic tools has been developed for the optimal design and regulation of complex metabolic pathways in bacteria. However, fewer tools exist in yeast that can precisely tune the expression of individual enzymes in novel metabolic pathways suitable for industrial-scale production of non-natural compounds. Tuning expression levels is critical for reducing the metabolic burden of over-expressed proteins, the accumulation of toxic intermediates, and for redirecting metabolic flux from native pathways involving essential enzymes without negatively affecting the viability of the host. We have developed a yeast membrane protein hyper-expression system with critical advantages over conventional, plasmid-based, expression systems. However, expression levels are sometimes so high that they adversely affect protein targeting/folding or the growth and/or phenotype of the host. Here we describe the use of small synthetic mRNA control modules that allowed us to predictably tune protein expression levels to any desired level. Down-regulation of expression was achieved by engineering small GC-rich mRNA stem-loops into the 5' UTR that inhibited translation initiation of the yeast ribosomal 43S preinitiation complex (PIC). Exploiting the fact that the yeast 43S PIC has great difficulty scanning through GC-rich mRNA stem-loops, we created yeast strains containing 17 different RNA stem-loop modules in the 5' UTR that expressed varying amounts of the fungal multidrug efflux pump reporter Cdr1p from Candida albicans. Increasing the length of mRNA stem-loops (that contained only GC-pairs) near the AUG start-codon led to a surprisingly large decrease in Cdr1p expression; ~2.7-fold for every additional GC-pair added to the stem, while the mRNA levels remained largely unaffected. An mRNA stem-loop of seven GC-pairs (∆G = -15.8 kcal/mol) reduced Cdr1p expression levels by >99%, and even the smallest possible stem-loop of only three GC-pairs (∆G = -4.4 kcal/mol) inhibited

Developmental and growth defects in mice with combined deficiency of CK2 catalytic genes.

Science.gov (United States)

Landesman-Bollag, Esther; Belkina, Anna; Hovey, Beth; Connors, Edward; Cox, Charles; Seldin, David C

2011-10-01

The CK2 α and α' catalytic gene products have overlapping biochemical activity, but in vivo, their functions are very different. Deletion of both alleles of CK2α leads to mid-gestational embryonic lethality, while deletion of both alleles of CK2α' does not interfere with viability or development of embryos; however, adult CK2α'-/-males are infertile. To further elucidate developmental roles of CK2, and analyze functional overlap between the two catalytic genes, mice with combined knockouts were bred. Mice bearing any two CK2 catalytic alleles were phenotypically normal. However, inheritance of a single CK2α allele, without either CK2α' allele, resulted in partial embryonic lethality. Such mice that survived through embryogenesis were smaller at birth than littermate controls, and weighed less throughout life. However, their cardiac function and lifespan were normal. Fibroblasts derived from CK2α+/-CK2α'-/- embryos grew poorly in culture. These experiments demonstrate that combined loss of one CK2α allele and both CK2α' alleles leads to unique abnormalities of growth and development.

Complex p63 mRNA isoform expression patterns in squamous cell carcinoma of the head and neck

DEFF Research Database (Denmark)

Thurfjell, N.; Coates, P.J.; Uusitalo, T.

2004-01-01

on the role of p63 expression in human tumours, we used quantitative real-time RT-PCR to study individual p63 isoforms in squamous cell carcinomas of the head and neck (SCCHN). In keeping with previous reports, expression of the deltaN- and p63alpha-isoforms predominated and deltaNp63 mRNA was expressed...

Integrated analysis of miRNA and mRNA expression profiles in tilapia gonads at an early stage of sex differentiation.

Science.gov (United States)

Tao, Wenjing; Sun, Lina; Shi, Hongjuan; Cheng, Yunying; Jiang, Dongneng; Fu, Beide; Conte, Matthew A; Gammerdinger, William J; Kocher, Thomas D; Wang, Deshou

2016-05-04

MicroRNAs (miRNAs) represent a second regulatory network that has important effects on gene expression and protein translation during biological process. However, the possible role of miRNAs in the early stages of fish sex differentiation is not well understood. In this study, we carried an integrated analysis of miRNA and mRNA expression profiles to explore their possibly regulatory patterns at the critical stage of sex differentiation in tilapia. We identified 279 pre-miRNA genes in tilapia genome, which were highly conserved in other fish species. Based on small RNA library sequencing, we identified 635 mature miRNAs in tilapia gonads, in which 62 and 49 miRNAs showed higher expression in XX and XY gonads, respectively. The predicted targets of these sex-biased miRNAs (e.g., miR-9, miR-21, miR-30a, miR-96, miR-200b, miR-212 and miR-7977) included genes encoding key enzymes in steroidogenic pathways (Cyp11a1, Hsd3b, Cyp19a1a, Hsd11b) and key molecules involved in vertebrate sex differentiation (Foxl2, Amh, Star1, Sf1, Dmrt1, and Gsdf). These genes also showed sex-biased expression in tilapia gonads at 5 dah. Some miRNAs (e.g., miR-96 and miR-737) targeted multiple genes involved in steroid synthesis, suggesting a complex miRNA regulatory network during early sex differentiation in this fish. The sequence and expression patterns of most miRNAs in tilapia are conserved in fishes, indicating the basic functions of vertebrate miRNAs might share a common evolutionary origin. This comprehensive analysis of miRNA and mRNA at the early stage of molecular sex differentiation in tilapia XX and XY gonads lead to the discovery of differentially expressed miRNAs and their putative targets, which will facilitate studies of the regulatory network of molecular sex determination and differentiation in fishes.

Local ATP generation by brain-type creatine kinase (CK-B facilitates cell motility.

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Jan W P Kuiper

Full Text Available BACKGROUND: Creatine Kinases (CK catalyze the reversible transfer of high-energy phosphate groups between ATP and phosphocreatine, thereby playing a storage and distribution role in cellular energetics. Brain-type CK (CK-B deficiency is coupled to loss of function in neural cell circuits, altered bone-remodeling by osteoclasts and complement-mediated phagocytotic activity of macrophages, processes sharing dependency on actomyosin dynamics. METHODOLOGY/PRINCIPAL FINDINGS: Here, we provide evidence for direct coupling between CK-B and actomyosin activities in cortical microdomains of astrocytes and fibroblasts during spreading and migration. CK-B transiently accumulates in membrane ruffles and ablation of CK-B activity affects spreading and migration performance. Complementation experiments in CK-B-deficient fibroblasts, using new strategies to force protein relocalization from cytosol to cortical sites at membranes, confirmed the contribution of compartmentalized CK-B to cell morphogenetic dynamics. CONCLUSION/SIGNIFICANCE: Our results provide evidence that local cytoskeletal dynamics during cell motility is coupled to on-site availability of ATP generated by CK-B.

Changes in growth hormone (GH) messenger RNA (GH mRNA) expression in the rat anterior pituitary after single interferon (IFN) alpha administration

International Nuclear Information System (INIS)

Romanowski, W.; Braczkowski, R.; Nowakowska-Zajdel, E.; Muc-Wierzgon, M.; Zubelewicz-Szkodzinska, B.; Kosiewicz, J.; Korzonek, I.

2006-01-01

Introduction: Interferon a (IFN-a) is a cytokine with pleiotropic effects which, via different pathways, influences the secretion of certain cytokines and hormones. Growth hormone (GH) secreted from the pituitary has physiological effects on various target tissues. The question is how IFN-a administered in various types of disease influences GH secretion. This study investigated the acute effect of IFN-a on GH mRNA expression in the rat anterior pituitary. Objective: The aim of the study was to measure the cellular expression of GH mRNA by in situ hybridisation in the anterior pituitary after a single administration of IFN-a. Material and methods: Rats were administered an intraperitoneal injection of IFN-a or saline. The rat pituitaries were taken 2 and 4 hours after IFN/saline administration and kept frozen until in situ hybridisation histochemistry. A 31 - base 35S -labelled oligonucleotide probe complementary to part of the exonic mRNA sequence coding for GH mRNA was used. All control and experimental sections were hybridised in the same hybridisation reaction. Results: Acute administration of interferon a increased GH mRNA expression in the anterior pituitary in the 4-hour group in comparison with the control group, and there was no difference between the control group and the 2-hour rats. Conclusion: A single IFN-a administration was found to exert an influence on anterior pituitary GH mRNA expression. These observations may pave the way for presenting a possible new action of IFN-a. (author) GH mRNA, anterior pituitary, interferon

Gemcitabine-Based Chemogene Therapy for Pancreatic Cancer Using Ad-dCK::UMK GDEPT and TS/RR siRNA Strategies

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Soukaina Réjiba

2009-07-01

Full Text Available Gemcitabine is a first-line agent for advanced pancreatic cancer therapy. However, its efficacy is often limited by its poor intracellular metabolism and chemoresistance. To exert its antitumor activity, gemcitabine requires to be converted to its active triphosphate form. Thus, our aim was to improve gemcitabine activation using gene-directed enzyme prodrug therapy based on gemcitabine association with the deoxycytidine kinase::uridine monophosphate kinase fusion gene (dCK::UMK and small interference RNA directed against ribonucleotide reductase (RRM2 and thymidylate synthase (TS. In vitro, cytotoxicity was assessed by 3-[4,5-dimethylthiazol-2-yl]-3,5-diphenyl tetrazolium bromide and [3H]thymidine assays. Apoptosis-related gene expression and activity were analyzed by reverse transcription-polymerase chain reaction, Western blot, and ELISA. For in vivo studies, the treatment efficacy was evaluated on subcutaneous and orthotopic pancreatic tumor models. Our data indicated that cell exposure to gemcitabine induced a down-regulation of dCK expression and up-regulation of TS and RR expression in Panc1-resistant cells when compared with BxPc3- and HA-hpc2-sensitive cells. The combination of TS/RRM2 small interference RNA with Ad-dCK::UMK induced a 40-fold decrease of gemcitabine IC50 in Panc1 cells. This strong sensitization was associated to apoptosis induction with a remarkable increase in TRAIL expression and a diminution of gemcitabine-induced nuclear factor-κB activity. In vivo, the gemcitabine-based tritherapy strongly reduced tumor volumes and significantly prolonged mice survival. Moreover, we observed an obvious increase of apoptosis and decrease of cell proliferation in tumors receiving the tritherapy regimens. Together, these findings suggest that simultaneous TS/RRM2-gene silencing and dCK::UMK gene overexpression markedly improved gemcitabine's therapeutic activity. Clearly, this combined strategy warrants further investigation.

Expression Profile of IL-35 mRNA in Gingiva of Chronic Periodontitis and Aggressive Periodontitis Patients: A Semiquantitative RT-PCR Study

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Nagaraj B. Kalburgi

2013-01-01

Full Text Available Background. Proinflammatory and anti-inflammatory cytokines play a key role in the pathogenesis of periodontal diseases. Secretion of bioactive IL-35 has been described by T regulatory cells ( and is required for their maximal suppressive activity. are involved in the modulation of local immune response in chronic periodontitis patients. Objective. Hence, the present study was aimed to investigate the expression of IL-35 mRNA in chronic periodontitis and aggressive periodontitis patients. Materials and Methods. The present study was carried out in 60 subjects, which included 20 chronic periodontitis patients, 20 aggressive periodontitis patients, and 20 periodontally healthy controls. IL-35 mRNA expression in gingival tissue samples of all subjects was semiquantitatively analyzed using Reverse Transcriptase Polymerase Chain Reaction (RT-PCR. Results. The present study demonstrated the expression of IL-35 mRNA in gingival tissues of all the three groups. IL-35 mRNA expression was highest in chronic periodontitis subjects ( as compared to the aggressive periodontitis group ( and least seen in healthy patients (. Conclusion. The increased expression of IL-35 in chronic and aggressive periodontitis suggests its possible role in pathogenesis of periodontitis. Future studies done on large samples with intervention will strengthen our result.

Relationship between expression of leptin receptors mRNA in breast tissue, plasma leptin level in breast cancer patients with obesity and clinical pathologic data

International Nuclear Information System (INIS)

Li Chunrui; Liu Wenli; Sun Hanying; Zhou Jianfeng

2007-01-01

In order to investigate the expression of leptin receptors mRNA in breast tissue and plasma leptin levels in breast cancer patients with obesity and their relationship with clinical pathologic data, 124 subjects who were either obesity or had suffered from breast benign disease with obesity, or breast cancer with obesity were entered into this study. The levels of plasma leptin in all subjects were determined and leptin receptors mRNA expression levels were measured by RT-PCR in breast tissue of breast cancer patients with obesity and breast benign disease with obesity. The results showed that plasma leptin levels in breast cancer patients with obesity were significantly higher than those in breast benign disease with obesity and obesity patients alone (P<0.05). The expression of the leptin receptor long form [-Lep-R(L)-] mRNA and the leptin receptor short form [-Lep-R(S)-] mRNA in breast tissue of breast cancer patients with obesity were significantly higher than that in breast tissue of breast benign disease patients with obesity (P<0.05). The plasma leptin level had remarkable positive correlation with the expressions of the Lep-R(L) mRNA and the Lep-R(S) mRNA. The plasma leptin level and leptin receptors mRNA expression levels in patients were not correlated with the axillary node metastasis, menopause, the TNM stage or pathological type. Therefore, leptin may have a promoting effect on the carcinogenesis of breast cancer. (authors)

InterProScan Result: CK520508 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK520508 CK520508_1_ORF1 CAD87D477268C018 PFAM PF04683 Proteasom_Rpn13 NA ? IPR006773 unintegrated Cellular... Component: nucleus (GO:0005634)|Cellular Component: cytoplasm (GO:0005737) ...

Integrated analysis of miRNA and mRNA expression in childhood medulloblastoma compared with neural stem cells.

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Laura A Genovesi

Full Text Available Medulloblastoma (MB is the most common malignant brain tumor in children and a leading cause of cancer-related mortality and morbidity. Several molecular sub-types of MB have been identified, suggesting they may arise from distinct cells of origin. Data from animal models indicate that some MB sub-types arise from multipotent cerebellar neural stem cells (NSCs. Hence, microRNA (miRNA expression profiles of primary MB samples were compared to CD133+ NSCs, aiming to identify deregulated miRNAs involved in MB pathogenesis. Expression profiling of 662 miRNAs in primary MB specimens, MB cell lines, and human CD133+ NSCs and CD133- neural progenitor cells was performed by qRT-PCR. Clustering analysis identified two distinct sub-types of MB primary specimens, reminiscent of sub-types obtained from their mRNA profiles. 21 significantly up-regulated and 12 significantly down-regulated miRNAs were identified in MB primary specimens relative to CD133+ NSCs (p<0.01. The majority of up-regulated miRNAs mapped to chromosomal regions 14q32 and 17q. Integration of the predicted targets of deregulated miRNAs with mRNA expression data from the same specimens revealed enrichment of pathways regulating neuronal migration, nervous system development and cell proliferation. Transient over-expression of a down-regulated miRNA, miR-935, resulted in significant down-regulation of three of the seven predicted miR-935 target genes at the mRNA level in a MB cell line, confirming the validity of this approach. This study represents the first integrated analysis of MB miRNA and mRNA expression profiles and is the first to compare MB miRNA expression profiles to those of CD133+ NSCs. We identified several differentially expressed miRNAs that potentially target networks of genes and signaling pathways that may be involved in the transformation of normal NSCs to brain tumor stem cells. Based on this integrative approach, our data provide an important platform for future

Associations of ACE Gene Insertion/Deletion Polymorphism, ACE Activity, and ACE mRNA Expression with Hypertension in a Chinese Population

OpenAIRE

He, Qingfang; Fan, Chunhong; Yu, Min; Wallar, Gina; Zhang, Zuo-Feng; Wang, Lixin; Zhang, Xinwei; Hu, Ruying

2013-01-01

Background The present study was designed to explore the association of angiotensin converting enzyme (ACE) gene insertion/deletion (I/D, rs4646994) polymorphism, plasma ACE activity, and circulating ACE mRNA expression with essential hypertension (EH) in a Chinese population. In addition, a new detection method for circulating ACE mRNA expression was explored. Methods The research was approved by the ethics committee of Zhejiang Provincial Center for Disease Prevention and Control. Written i...

Sequence, 'subtle' alternative splicing and expression of the CYYR1 (cysteine/tyrosine-rich 1) mRNA in human neuroendocrine tumors

International Nuclear Information System (INIS)

Vitale, Lorenza; Coppola, Domenico; Strippoli, Pierluigi; Frabetti, Flavia; Huntsman, Shane A; Canaider, Silvia; Casadei, Raffaella; Lenzi, Luca; Facchin, Federica; Carinci, Paolo; Zannotti, Maria

2007-01-01

CYYR1 is a recently identified gene located on human chromosome 21 whose product has no similarity to any known protein and is of unknown function. Analysis of expressed sequence tags (ESTs) have revealed high human CYYR1 expression in cells belonging to the diffuse neuroendocrine system (DNES). These cells may be the origin of neuroendocrine (NE) tumors. The aim of this study was to conduct an initial analysis of sequence, splicing and expression of the CYYR1 mRNA in human NE tumors. The CYYR1 mRNA coding sequence (CDS) was studied in 32 NE tumors by RT-PCR and sequence analysis. A subtle alternative splicing was identified generating two isoforms of CYYR1 mRNA differing in terms of the absence (CAG - isoform, the first described mRNA for CYYR1 locus) or the presence (CAG + isoform) of a CAG codon. When present, this specific codon determines the presence of an alanine residue, at the exon 3/exon 4 junction of the CYYR1 mRNA. The two mRNA isoform amounts were determined by quantitative relative RT-PCR in 29 NE tumors, 2 non-neuroendocrine tumors and 10 normal tissues. A bioinformatic analysis was performed to search for the existence of the two CYYR1 isoforms in other species. The CYYR1 CDS did not show differences compared to the reference sequence in any of the samples, with the exception of an NE tumor arising in the neck region. Sequence analysis of this tumor identified a change in the CDS 333 position (T instead of C), leading to the amino acid mutation P111S. NE tumor samples showed no significant difference in either CYYR1 CAG - or CAG + isoform expression compared to control tissues. CYYR1 CAG - isoform was significantly more expressed than CAG + isoform in NE tumors as well as in control samples investigated. Bioinformatic analysis revealed that only the genomic sequence of Pan troglodytes CYYR1 is consistent with the possible existence of the two described mRNA isoforms. A new 'subtle' splicing isoform (CAG + ) of CYYR1 mRNA, the sequence and

In Search of Small Molecule Inhibitors Targeting the Flexible CK2 Subunit Interface

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Benoît Bestgen

2017-02-01

Full Text Available Protein kinase CK2 is a tetrameric holoenzyme composed of two catalytic (α and/or α’ subunits and two regulatory (β subunits. Crystallographic data paired with fluorescence imaging techniques have suggested that the formation of the CK2 holoenzyme complex within cells is a dynamic process. Although the monomeric CK2α subunit is endowed with a constitutive catalytic activity, many of the plethora of CK2 substrates are exclusively phosphorylated by the CK2 holoenzyme. This means that the spatial and high affinity interaction between CK2α and CK2β subunits is critically important and that its disruption may provide a powerful and selective way to block the phosphorylation of substrates requiring the presence of CK2β. In search of compounds inhibiting this critical protein–protein interaction, we previously designed an active cyclic peptide (Pc derived from the CK2β carboxy-terminal domain that can efficiently antagonize the CK2 subunit interaction. To understand the functional significance of this interaction, we generated cell-permeable versions of Pc, exploring its molecular mechanisms of action and the perturbations of the signaling pathways that it induces in intact cells. The identification of small molecules inhibitors of this critical interaction may represent the first-choice approach to manipulate CK2 in an unconventional way.

Utility of CK7 and CK20 immunohistochemistry in the detection of synchronous breast and colon carcinoma in a pleural effusion: a case report and supporting survey of archival material

DEFF Research Database (Denmark)

Stopyra, G A; Warhol, M J; Multhaupt, H A

2001-01-01

, respectively. An immunohistochemical survey of archival breast and colon primary and metastatic carcinomas confirmed the established CK7+/CK20- phenotype of breast and CK7-/CK20+ phenotype of colon primary carcinomas, and the maintenance of this phenotype in metastases thereof. A survey of benign and malignant...

Relationship between PPARα mRNA expression and mitochondrial respiratory function and ultrastructure of the skeletal muscle of patients with COPD.

Science.gov (United States)

Zhang, Jian-Qing; Long, Xiang-Yu; Xie, Yu; Zhao, Zhi-Huan; Fang, Li-Zhou; Liu, Ling; Fu, Wei-Ping; Shu, Jing-Kui; Wu, Jiang-Hai; Dai, Lu-Ming

2017-11-02

Peripheral muscle dysfunction is an important complication in patients with chronic obstructive pulmonary disease (COPD). The objective of this study was to explore the relationship between the levels of peroxisome proliferator-activated receptor α (PPARα) mRNA expression and the respiratory function and ultrastructure of mitochondria in the vastus lateralis of patients with COPD. Vastus lateralis biopsies were performed on 14 patients with COPD and 6 control subjects with normal lung function. PPARα mRNA levels in the muscle tissue were detected by real-time PCR. A Clark oxygen electrode was used to assess mitochondrial respiratory function. Mitochondrial number, fractional area in skeletal muscle cross-sections, and Z-line width were observed via transmission electron microscopy. The PPARα mRNA expression was significantly lower in COPD patients with low body mass index (BMIL) than in both COPD patients with normal body mass index (BMIN) and controls. Mitochondrial respiratory function (assessed by respiratory control ratio) was impaired in COPD patients, particularly in BMIL. Compared with that in the control group, mitochondrial number and fractional area were lower in the BMIL group, but were maintained in the BMIN group. Further, the Z-line became narrow in the BMIL group. PPARα mRNA expression was positively related to mitochondrial respiratory function and volume density. In COPD patients with BMIN, mitochondria volume density was maintained, while respiratory function decreased, whereas both volume density and respiratory function decreased in COPD patients with BMIL. PPARα mRNA expression levels are associated with decreased mitochondrial respiratory function and volume density, which may contribute to muscle dysfunction in COPD patients.

Valsartan Upregulates Kir2.1 in Rats Suffering from Myocardial Infarction via Casein Kinase 2.

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Li, Xinran; Hu, Hesheng; Wang, Ye; Xue, Mei; Li, Xiaolu; Cheng, Wenjuan; Xuan, Yongli; Yin, Jie; Yang, Na; Yan, Suhua

2015-06-01

Myocardial infarction (MI) results in an increased susceptibility to ventricular arrhythmias, due in part to decreased inward-rectifier K+ current (IK1), which is mediated primarily by the Kir2.1 protein. The use of renin-angiotensin-aldosterone system antagonists is associated with a reduced incidence of ventricular arrhythmias. Casein kinase 2 (CK2) binds and phosphorylates SP1, a transcription factor of KCNJ2 that encodes Kir2.1. Whether valsartan represses CK2 activation to ameliorate IK1 remodeling following MI remains unclear. Wistar rats suffering from MI received either valsartan or saline for 7 days. The protein levels of CK2 and Kir2.1 were each detected via a Western blot analysis. The mRNA levels of CK2 and Kir2.1 were each examined via quantitative real-time PCR. CK2 expression was higher at the infarct border; and was accompanied by a depressed IK1/Kir2.1 protein level. Additionally, CK2 overexpression suppressed KCNJ2/Kir2.1 expression. By contrast, CK2 inhibition enhanced KCNJ2/Kir2.1 expression, establishing that CK2 regulates KCNJ2 expression. Among the rats suffering from MI, valsartan reduced CK2 expression and increased Kir2.1 expression compared with the rats that received saline treatment. In vitro, hypoxia increased CK2 expression and valsartan inhibited CK2 expression. The over-expression of CK2 in cells treated with valsartan abrogated its beneficial effect on KCNJ2/Kir2.1. AT1 receptor antagonist valsartan reduces CK2 activation, increases Kir2.1 expression and thereby ameliorates IK1 remodeling after MI in the rat model.

InterProScan Result: CK559994 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK559994 CK559994_4_ORF2 020DFCD801EB69C2 PFAM PF04114 Gaa1 NA ? IPR007246 unintegrated Cellular... Component: integral to membrane (GO:0016021)|Cellular Component: GPI-anchor transamidase complex (GO:0042765) ...

Rainbow trout CK9, a CCL25-like ancient chemokine that attracts and regulates B cells and macrophages, the main antigen presenting cells in fish.

Science.gov (United States)

Aquilino, Carolina; Granja, Aitor G; Castro, Rosario; Wang, Tiehui; Abos, Beatriz; Parra, David; Secombes, Christopher J; Tafalla, Carolina

2016-04-05

CK9 is a rainbow trout (Oncorhynchus mykiss) CC chemokine phylogenetically related to mammalian CCL25. Although CK9 is known to be transcriptionally regulated in response to inflammation particularly in mucosal tissues, its functionality has never been revealed. In the current work, we have demonstrated that CK9 is chemoattractant for antigen presenting cells (APCs) expressing major histocompatibility complex class II (MHC II) on the cell surface. Among these APCs, CK9 has a strong chemotactic capacity for both B cells (IgM+ and IgT+) and macrophages. Along with its chemotactic capacities, CK9 modulated the MHC II turnover of B lymphocytes and up-regulated the phagocytic capacity of both IgM+ cells and macrophages. Although CK9 had no lymphoproliferative effects, it increased the survival of IgT+ lymphocytes. Furthermore, we have established that the chemoattractant capacity of CK9 is strongly increased after pre-incubation of leukocytes with a T-independent antigen, whereas B cell receptor (BCR) cross-linking strongly abrogated their capacity to migrate to CK9, indicating that CK9 preferentially attracts B cells at the steady state or under BCR-independent stimulation. These results point to CK9 being a key regulator of B lymphocyte trafficking in rainbow trout, able to modulate innate functions of teleost B lymphocytes and macrophages.

Variation of stemness markers expression in tumor nodules from synchronous multi-focal hepatocellular carcinoma - an immunohistochemical study.

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Lo, Regina Cheuk-Lam; Leung, Carmen Oi-Ning; Chok, Kenneth Siu-Ho; Ng, Irene Oi-Lin

2017-08-01

Advancing knowledge in molecular pathogenesis of hepatocellular carcinoma (HCC) opens up new horizons in the diagnostic, prognostic and therapeutic perspectives. Assessing the expression of molecular targets prior to definitive treatment is gaining importance in clinical practice. In this study, we investigated the variation in expression pattern of stemness markers in synchronous multi-focal HCC. In the first cohort, 21 liver explants with multi-focal HCC were examined for expression of stemness markers EpCAM, Sox9 and CK19 by immunohistochemistry (IHC). Expression data of 50 tumor nodules were analyzed to determine the concordance of expression among nodules in the same livers. In the second cohort, 14 tumor nodules from 6 multi-focal HCC cases proven as intra-hepatic metastasis were examined for Soc9 immunoexpression. In the first cohort, thirty nodules from 16 cases expressed one or more markers, with Sox9 being most frequently expressed. Complete concordance of expression pattern for all 3 markers was observed in 6 cases. Discrepancy of staining degree was noted in 4 cases for EpCAM, 14 cases for Sox9, and 6 cases for CK19. A two-tier or three-tier difference in staining scores was noted in 5 cases for Sox9 and one case for CK19. With Sox9, identical tumor morphology in terms of Edmondson grading and growth pattern did not infer the same degree of immunoexpression; and the largest tumor nodule was not representative of highest IHC score. In the second cohort of intra-hepatic metastasis, complete concordance of Sox9 expression level was observed in 5 out of 6 cases; while the remaining case showed a 1-tier difference of positive staining. Our findings suggested that clonality of tumor nodules is apparently an important factor to infer immunoexpression pattern. When there is limited information to discern multiple primaries versus intra-hepatic metastasis in multi-focal HCC, discordant degree of stemness markers expression among tumor nodules was commonly

IER5 gene's mRNA expression after irradiation

International Nuclear Information System (INIS)

Ding Kuke; Shen Jingjing; Xu Lili; Li Yanling; Zhou Ping; Ma Binrong; Zhao Zengqiang; Sui Jianli; Zhou Pingkun

2008-01-01

Objective: To explore the effect of irradiation on IER5 gene expression. Methods: Two kinds of cells (AHH-1 and HeLa) and the BALB/c-nu mice inoculated with tumor cells were exposed to 60 Co γ- rays and analyzed by real-time PCR. The above-mentioned irradiated objects were firstly divided into groups by different doses and post-radiation time, then mRNA were extracted and reverse-transcripted to DNA before real-time PCR test. Results: Under the same condition, AHH-1 was more sensitive to radiation than HeLa. The dose level corresponding to the expression peak of AHH-1 was less than that of HeLa. For AHH-1 cells, the response to 2 Gy irradiation was earlier than that to 10 Gy. But there was not remarkable difference for HeLa response between 2 and 10 Gy, and the top transcriptional levels for both cells nearly simultaneously appeared at 2 h after irradiation. In addition, the IER5 gene of human liver tumor was more sensitive than that of lung cancer and brain tumor. Conclusions: IER5 might be a candidate biomarker of radiation injury, and had the potential value in radiation-therapy for liver tumor. (authors)

Basaloid Squamous Cell Carcinoma of the Head and Neck: Subclassification into Basal, Ductal, and Mixed Subtypes Based on Comparison of Clinico-pathologic Features and Expression of p53, Cyclin D1, Epidermal Growth Factor Receptor, p16, and Human Papillomavirus

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Kyung-Ja Cho

2017-07-01

Full Text Available Background Basaloid squamous cell carcinoma (BSCC is a rare variant of squamous cell carcinoma with distinct pathologic characteristics. The histogenesis of BSCC is not fully understood, and the cancer has been suggested to originate from a totipotent primitive cell in the basal cell layer of the surface epithelium or in the proximal duct of secretory glands. Methods Twenty-six cases of head and neck BSCC from Asan Medical Center, Seoul, Korea, reported during a 14-year-period were subclassified into basal, ductal, and mixed subtypes according to the expression of basal (cytokeratin [CK] 5/6, p63 or ductal markers (CK7, CK8/18. The cases were also subject to immunohistochemical study for CK19, p53, cyclin D1, epidermal growth factor receptor (EGFR, and p16 and to in situ hybridization for human papillomavirus (HPV, and the results were clinico-pathologically compared. Results Mixed subtype (12 cases was the most common, and these cases showed hypopharyngeal predilection, older age, and higher expression of CK19, p53, and EGFR than other subtypes. The basal subtype (nine cases showed frequent comedo-necrosis and high expression of cyclin D1. The ductal subtype (five cases showed the lowest expression of p53, cyclin D1, and EGFR. A small number of p16- and/or HPV-positive cases were not restricted to one subtype. BSCC was the cause of death in 19 patients, and the average follow-up period for all patients was 79.5 months. Overall survival among the three subtypes was not significantly different. Conclusions The results of this study suggest a heterogeneous pathogenesis of head and neck BSCC. Each subtype showed variable histology and immunoprofiles, although the clinical implication of heterogeneity was not determined in this study.

PAI-1 mRNA expression and plasma level in rheumatoid arthritis: relationship with 4G/5G PAI-1 polymorphism.

Science.gov (United States)

Muñoz-Valle, José Francisco; Ruiz-Quezada, Sandra Luz; Oregón-Romero, Edith; Navarro-Hernández, Rosa Elena; Castañeda-Saucedo, Eduardo; De la Cruz-Mosso, Ulises; Illades-Aguiar, Berenice; Leyva-Vázquez, Marco Antonio; Castro-Alarcón, Natividad; Parra-Rojas, Isela

2012-12-01

Rheumatoid arthritis (RA) is a chronic inflammatory disease affecting the synovial membrane, cartilage and bone. PAI-1 is a key regulator of the fibrinolytic system through which plasminogen is converted to plasmin. The plasmin activates the matrix metalloproteinase system, which is closely related with the joint damage and bone destruction in RA. The aim of this study was to investigate the relationship between 4G/5G PAI-1 polymorphism with mRNA expression and PAI-1 plasma protein levels in RA patients. 113 RA patients and 123 healthy subjects (HS) were included in the study. The 4G/5G PAI-1 polymorphism was determined by polymerase chain reaction-restriction fragment length polymorphism method; the PAI-1 mRNA expression was determined by real-time PCR; and the soluble PAI-1 (sPAI-1) levels were quantified using an ELISA kit. No significant differences in the genotype and allele frequencies of 4G/5G PAI-1 polymorphism were found between RA patients and HS. However, the 5G/5G genotype was the most frequent in both studied groups: RA (42%) and HS (44%). PAI-1 mRNA expression was slightly increased (0.67 fold) in RA patients with respect to HS (P = 0.0001). In addition, in RA patients, the 4G/4G genotype carriers showed increased PAI-1 mRNA expression (3.82 fold) versus 4G/5G and 5G/5G genotypes (P = 0.0001), whereas the sPAI-1 plasma levels did not show significant differences. Our results indicate that the 4G/5G PAI-1 polymorphism is not a marker of susceptibility in the Western Mexico. However, the 4G/4G genotype is associated with high PAI-1 mRNA expression but not with the sPAI-1 levels in RA patients.

The quantification of COMT mRNA in post mortem cerebellum tissue: diagnosis, genotype, methylation and expression

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Craig Ian W

2006-02-01

Full Text Available Abstract Background The COMT gene is located on chromosome 22q11, a region strongly implicated in the aetiology of several psychiatric disorders, in particular schizophrenia. Previous research has suggested that activity and expression of COMT is altered in schizophrenia, and is mediated by one or more polymorphisms within the gene, including the functional Val158Met polymorphism. Method In this study we examined the expression levels of COMT mRNA using quantitative RT-PCR in 60 post mortem cerebellum samples derived from individuals with schizophrenia, bipolar disorder, depression, and no history of psychopathology. Furthermore, we have examined the methylation status of two CpG sites in the promoter region of the gene. Results We found no evidence of altered COMT expression or methylation in any of the psychiatric diagnoses examined. We did, however, find evidence to suggest that genotype is related to COMT gene expression, replicating the findings of two previous studies. Specifically, val158met (rs165688; Val allele rs737865 (G allele and rs165599 (G allele all showed reduced expression (P COMT expression, with females exhibiting significantly greater levels of COMT mRNA. Conclusion The expression of COMT does not appear to be altered in the cerebellum of individuals suffering from schizophrenia, bipolar disorder or depression, but does appear to be influenced by single nucleotide polymorphisms within the gene.

InterProScan Result: CK524647 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK524647 CK524647_3_ORF1 1A3C8EDABD21145E PROFILE PS51174 BARWIN_3 NA ? IPR001153 unintegrated Biological... Process: defense response to bacterium (GO:0042742)|Biological Process: defense response to fungus (GO:0050832) ...

Collagen V-induced nasal tolerance downregulates pulmonary collagen mRNA gene and TGF-beta expression in experimental systemic sclerosis

Directory of Open Access Journals (Sweden)

Parra Edwin R

2010-01-01

Full Text Available Abstract Background The purpose of this study was to evaluate collagen deposition, mRNA collagen synthesis and TGF-beta expression in the lung tissue in an experimental model of scleroderma after collagen V-induced nasal tolerance. Methods Female New Zealand rabbits (N = 12 were immunized with 1 mg/ml of collagen V in Freund's adjuvant (IM. After 150 days, six immunized animals were tolerated by nasal administration of collagen V (25 μg/day (IM-TOL daily for 60 days. The collagen content was determined by morphometry, and mRNA expressions of types I, III and V collagen were determined by Real-time PCR. The TGF-beta expression was evaluated by immunostaining and quantified by point counting methods. To statistic analysis ANOVA with Bonferroni test were employed for multiple comparison when appropriate and the level of significance was determined to be p Results IM-TOL, when compared to IM, showed significant reduction in total collagen content around the vessels (0.371 ± 0.118 vs. 0.874 ± 0.282, p p p = 0.026. The lung tissue of IM-TOL, when compared to IM, showed decreased immunostaining of types I, III and V collagen, reduced mRNA expression of types I (0.10 ± 0.07 vs. 1.0 ± 0.528, p = 0.002 and V (1.12 ± 0.42 vs. 4.74 ± 2.25, p = 0.009 collagen, in addition to decreased TGF-beta expression (p Conclusions Collagen V-induced nasal tolerance in the experimental model of SSc regulated the pulmonary remodeling process, inhibiting collagen deposition and collagen I and V mRNA synthesis. Additionally, it decreased TGF-beta expression, suggesting a promising therapeutic option for scleroderma treatment.

Increased expression of Th17 cytokines in patients with psoriasis

African Journals Online (AJOL)

AJL

2012-02-16

Feb 16, 2012 ... immune, heredity, psychology and environment factors ... ground with mortar and pestle, cooled by liquid nitrogen of the ground tissue .... Figure 1. Correction between expression of IL-17A mRNA and IL-23P19 mRNA. skin.

mRNA expression profile in DLD-1 and MOLT-4 cancer cell lines cultured under Microgravity

Data.gov (United States)

National Aeronautics and Space Administration — DLD-1 and MOLT-4 cell lines were cultured in a Rotating cell culture system to simulate microgravity and mRNA expression profile was observed in comparison to Static...

Small, synthetic, GC-rich mRNA stem-loop modules 5′ proximal to the AUG start-codon predictably tune gene expression in yeast

Science.gov (United States)

2013-01-01

Background A large range of genetic tools has been developed for the optimal design and regulation of complex metabolic pathways in bacteria. However, fewer tools exist in yeast that can precisely tune the expression of individual enzymes in novel metabolic pathways suitable for industrial-scale production of non-natural compounds. Tuning expression levels is critical for reducing the metabolic burden of over-expressed proteins, the accumulation of toxic intermediates, and for redirecting metabolic flux from native pathways involving essential enzymes without negatively affecting the viability of the host. We have developed a yeast membrane protein hyper-expression system with critical advantages over conventional, plasmid-based, expression systems. However, expression levels are sometimes so high that they adversely affect protein targeting/folding or the growth and/or phenotype of the host. Here we describe the use of small synthetic mRNA control modules that allowed us to predictably tune protein expression levels to any desired level. Down-regulation of expression was achieved by engineering small GC-rich mRNA stem-loops into the 5′ UTR that inhibited translation initiation of the yeast ribosomal 43S preinitiation complex (PIC). Results Exploiting the fact that the yeast 43S PIC has great difficulty scanning through GC-rich mRNA stem-loops, we created yeast strains containing 17 different RNA stem-loop modules in the 5′ UTR that expressed varying amounts of the fungal multidrug efflux pump reporter Cdr1p from Candida albicans. Increasing the length of mRNA stem-loops (that contained only GC-pairs) near the AUG start-codon led to a surprisingly large decrease in Cdr1p expression; ~2.7-fold for every additional GC-pair added to the stem, while the mRNA levels remained largely unaffected. An mRNA stem-loop of seven GC-pairs (∆G = −15.8 kcal/mol) reduced Cdr1p expression levels by >99%, and even the smallest possible stem-loop of only three GC-pairs (�

InterProScan Result: CK554737 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK554737 CK554737_1_ORF2 7FBB5DC6E03E4AAA PANTHER PTHR21347 CLEFT LIP AND PALATE ASSOCIATED TRANSME...MBRANE PROTEIN-RELATED 3.8e-29 T IPR008429 Cleft lip and palate transmembrane 1 ...

InterProScan Result: CK524647 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK524647 CK524647_3_ORF1 1A3C8EDABD21145E PROFILE PS51174 BARWIN_3 8.795 T IPR001153 Barwin Biological... Process: defense response to bacterium (GO:0042742)|Biological Process: defense response to fungus (GO:0050832) ...

Effects of hypoxic preconditioning on the changes of expression of neuroglobin mRNA and labeled positive cells following cerebral ischemia in gerbils

Institute of Scientific and Technical Information of China (English)

Yong Zhang; Yanqun Chang; Zhenfang Liu; Qingxi Fu; Xiao Zhang

2006-01-01

expression of NGB mRNA was detected with RT-PCR, and the number of NGF positive cells was counted with immunohistochemical staining. The NGB mRNA expression and number of NGB positive cells at different time points after ischemia were compared between the gerbils treated with and without hypoxic preconditioning.MAIN OUTCOME MEASURES: NGB mRNA expression and number of NGB positive cells at 1, 5, 10, 30 and 60 minutes after cerebral ischemia.RESULTS: All the 66 Mongolian gerbils were involved in the analysis of results. ① Results of NGB mRNA: In the cerebral ischemia group, the NGB mRNA expression began to increase at 1 minute after cerebral ischemia, but had no obvious difference as compared with that in the sham-operated group (P ＞ 0.05), that at 5 minutes was obviously higher than that in the sham-operated group (0.951±0.034, 0.597±0.008, P ＜ 0.05), it decreased gradually 10, 30 and 60 minutes after cerebral ischemia, but had no obvious difference as compared with that in the sham-operated group (P＞ 0.05). In the hypoxic preconditioning group, the NGB mRNA expression increased rapidly 1 minute after cerebral ischemia, which was obviously higher than that in the cerebral ischemia group (0.641 ±0.010, 0.618±0.015, P ＜ 0.05), and the expressions at 5, 10 and 30 minutes were still obviously higher than those in the cerebral ischemia group (0.995±0.020 vs. 0.951 ±0.034; 0.941 ±0.010 vs. 0.615±0.018; 0.642±0.010 vs. 0.608±0.010, P ＜ 0.05-0.01 ), whereas the expression at 60 minutes were not obviously different from that in the cerebral ischemia group (P＞ 0.05). ② Number of NGB positive cells: The numbers of NGB positive cells at 1, 10 and 30 minutes after cerebral ishcemia in the hypoxic preconditioning group [(50.2±3.3), (67.2±3.3), (35.0±4.3) cells] were obviously more than those in the cerebral ischemia group [(33.0±2.1 ), (60.5± 1.9), (23.3±3.1) cells, P＜ 0.05-0.01], whereas those at 5 and 60 minutes had no obvious differences between the two

InterProScan Result: CK564775 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK564775 CK564775_2_ORF2 26DFD9502C5FAF5B PANTHER PTHR11824 VOLTAGE-DEPENDENT CALCI...UM CHANNEL BETA SUBUNIT 4.6e-14 T IPR000584 Voltage-dependent calcium channel, L-type, beta subunit Molecular Function: volt

InterProScan Result: CK545780 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK545780 CK545780_1_ORF1 ADB621D805C57486 PFAM PF00995 Sec1 2e-26 T IPR001619 Sec1-like protein Biological... Process: vesicle docking involved in exocytosis (GO:0006904)|Biological Process: vesicle-mediated transport (GO:0016192) ...

InterProScan Result: CK500303 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK500303 CK500303_3_ORF1 7378D8878C223668 PFAM PF04192 Utp21 9.7e-29 T IPR007319 Small-subunit process...ome, Utp21 Biological Process: rRNA processing (GO:0006364)|Cellular Component: small-subunit processome (GO:0032040) ...

[Influence of FPS on the expression of LDL-R mRNA in the liver tissues of hyperlipidemic rats].

Science.gov (United States)

Wu, Qing-he; Xing, Yan-hong; Rong, Xiang-lu; Huang, Ping

2007-08-01

To explore the effect of FPS on low-density lipoprotein acceptor (LDL-R) mRNA in the liver tissues of hyperlipidemic rats. Sixty healthy male SD rats were randomly divided into six groups: normal control, model control, Gynostemma pentaphyllum, FPS low dosage, FPS moderate dosage, and FPS high dosage group. Excepting the rats in the normal control group, the ones in other groups were all made rats' hyperlipidemic model by irrigating hyperlipidemic emulsion into the stomach and observed the expression of LDL-R mRNA in the liver tissues of rats of each group. Relative content of LDL-RmRNA in low and moderate dosage groups was notably higher than that inmodel group. The contents's difference was not remarkable between FPS moderate dosage group and Gynostemma pentaphyllum group. FPS can appreciably increase the expression of LDL-R mRNA in the liver tissues of hyperlipidemic rats and promote the elimination ofLDL-C to reduce serum cholesterol notably.

The Expression of Bone Morphogenetic Protein 2 and Matrix Metalloproteinase 2 through Retinoic Acid Receptor Beta Induced by All-Trans Retinoic Acid in Cultured ARPE-19 Cells.

Directory of Open Access Journals (Sweden)

Zhenya Gao

Full Text Available All-trans retinoic acid (ATRA plays an important role in ocular development. Previous studies found that retinoic acid could influence the metabolism of scleral remodeling by promoting retinal pigment epithelium (RPE cells to secrete secondary signaling factors. The purpose of this study was to investigate whether retinoic acid affected secretion of bone morphogenetic protein 2 (BMP-2 and matrix metalloproteinase 2 (MMP-2 and to explore the signaling pathway of retinoic acid in cultured acute retinal pigment epithelial 19 (ARPE-19 cells.The effects of ATRA (concentrations from 10-9 to 10-5 mol/l on the expression of retinoic acid receptors (RARs in ARPE-19 cells were examined at the mRNA and protein levels using reverse transcription-polymerase chain reaction (RT-PCR and western blot assay, respectively. The effects of treating ARPE-19 cells with ATRA concentrations ranging from 10-9 to 10-5 mol/l for 24 h and 48 h or with 10-6mol/l ATRA at different times ranging from 6h to 72h were assessed using real-time quantitative PCR (qPCR and enzyme-linked immunosorbent assay (ELISA. The contribution of RARβ-induced activation of ARPE-19 cells was confirmed using LE135, an antagonist of RARβ.RARβ mRNA levels significantly increased in the ARPE-19 cells treated with ATRA for 24h and 48h. These increases in RARβ mRNA levels were dose dependent (at concentrations of 10-9 to 10-5 mol/l with a maximum effect observed at 10-6 mol/l. There were no significant changes in the mRNA levels of RARα and RARγ. Western blot assay revealed that RARβ protein levels were increased significantly in a time-dependent manner in ARPE-19 cells treated with 10-6 mol/l ATRA from 12 h to 72 h, with a marked increase observed at 24 h and 48 h. The upregulation of RARβ and the ATRA-induced secretion in ARPE-19 cells could be inhibited by the RARβ antagonist LE135.ATRA induced upregulation of RARβ in ARPE-19 cells and stimulated these cells to secrete BMP-2 and MMP-2.

Changes in apoptotic microRNA and mRNA expression profiling in Caenorhabditis elegans during the Shenzhou-8 mission

International Nuclear Information System (INIS)

Gao Ying; Li Shuai; Xu Dan; Wang Junjun; Sun Yeqing

2015-01-01

Radiation and microgravity exposure have been proven to induce abnormal apoptosis in microRNA (miRNA) and mRNA expression, but whether space conditions, including radiation and microgravity, activate miRNAs to regulate the apoptosis is undetermined. For that purpose, we investigated miRNome and mRNA expression in the ced-1 Caenorhabditis elegans mutant vs the wild-type, both of which underwent spaceflight, spaceflight 1g-centrifuge control and ground control conditions during the Shenzhou-8 mission. Results showed that no morphological changes in the worms were detected, but differential miRNA expression increased from 43 (ground control condition) to 57 and 91 in spaceflight and spaceflight control conditions, respectively. Microgravity altered miRNA expression profiling by decreasing the number and significance of differentially expressed miRNA compared with 1 g incubation during spaceflight. Alterations in the miRNAs were involved in alterations in apoptosis, neurogenesis larval development, ATP metabolism and GTPase-mediated signal transduction. Among these, 17 altered miRNAs potentially involved in apoptosis were screened and showed obviously different expression signatures between space conditions. By integrated analysis of miRNA and mRNA, miR-797 and miR-81 may be involved in apoptosis by targeting the genes ced-10 and both drp-1 and hsp-1, respectively. Compared with ground condition, space conditions regulated apoptosis though a different manner on transcription, by altering expression of seven core apoptotic genes in spaceflight condition, and eight in spaceflight control condition. Results indicate that, miRNA of Caenorhabditis elegans probably regulates apoptotic gene expression in response to space environmental stress, and shows different behavior under microgravity condition compared with 1 g condition in the presence of space radiation. (author)

The Drosophila melanogaster DmCK2beta transcription unit encodes for functionally non-redundant protein isoforms.

Science.gov (United States)

Jauch, Eike; Wecklein, Heike; Stark, Felix; Jauch, Mandy; Raabe, Thomas

2006-06-07

Genes encoding for the two evolutionary highly conserved subunits of a heterotetrameric protein kinase CK2 holoenzyme are present in all examined eukaryotic genomes. Depending on the organism, multiple transcription units encoding for a catalytically active CK2alpha subunit and/or a regulatory CK2beta subunit may exist. The phosphotransferase activity of members of the protein kinase CK2alpha family is thought to be independent of second messengers but is modulated by interaction with CK2beta-like proteins. In the genome of Drosophila melanogaster, one gene encoding for a CK2alpha subunit and three genes encoding for CK2beta-like proteins are present. The X-linked DmCK2beta transcription unit encodes for several CK2beta protein isoforms due to alternative splicing of its primary transcript. We addressed the question whether CK2beta-like proteins are redundant in function. Our in vivo experiments show that variations of the very C-terminal tail of CK2beta isoforms encoded by the X-linked DmCK2beta transcription unit influence their functional properties. In addition, we find that CK2beta-like proteins encoded by the autosomal D. melanogaster genes CK2betates and CK2beta' cannot fully substitute for a loss of CK2beta isoforms encoded by DmCK2beta.

Effect of Heat Stress on the Expression of GABA Receptor mRNA in the HPG Axis of Wenchang Chickens

Directory of Open Access Journals (Sweden)

LJ Xie

Full Text Available ABSTRACT We investigated the effect of heat stress (HS on the expression of the GABA receptor in the hypothalamic-pituitary-gonadal (HPG axis of Wenchang chickens. Real-time quantitative RT-PCR (qRT-PCR was used to quantify the GABA receptor mRNA levels along the HPG axis of chickens under HS (40±0.5 °C for 1-6 weeks. Our results showed that the expression of GABAA and GABAB receptor at the mRNAs levels in the tissues of HPG axis exhibited fluctuation and variability. After HS, the mRNA level of GABAA receptor was significantly reduced in the hypothalamus of 1-week-old and in the pituitary of 3-week-old chickens, but significantly increased in the pituitary of 1-, 4-, and 5-week-old chickens. The GABAB receptor mRNA level significantly declined in the hypothalamus of 1-week-old and in the pituitary of 3-week-old chickens, but was significantly upregulated in the pituitary and testis of 1- and 2-week-old chickens. At other time points, the expressions of GABAA receptor and GABAB receptor showed no significant differences compared with control group. These results indicated that the levels of GABAA receptor and GABAB receptor mRNAs varied in different tissues of the HPG axis in chickens of different ages, displaying temporal and spatial variations. GABA receptor behaved as a positively-regulated gene by HS, i.e., its mRNA was increased by HS; similarly, it was a negatively-regulated gene by HS, when its expression was reduced by HS.

[Differential expression of IGF-I and its mRNA in mandibular condylar cartilage of rat--direct evidence for servosystem theory of facial growth].

Science.gov (United States)

Zhou, Z; Luo, S

1998-05-01

It was studied the expression of IGF-I and its mRNA in the condylar cartilage of 10 7-week-old SD male rats by using in situ hybridization and immunohisto-chemistry technique. The results showed both IGF-I and its gene expressed in growing rat condyle. IGF-I peptide was abundant in germinal zone, and positive reaction of its mRNA was strongest in transitional and maturational zones. These indicate that condylar cartilage has the capability of local production and secretion of IGF-I, mediating the command effect of STH, and differential expression of IGF-I and its mRNA might establish the local feedback loop, which supply a direct evidence for servosystem theory of facial growth.

Keratin14 mRNA expression in human pneumocytes during quiescence, repair and disease.

Directory of Open Access Journals (Sweden)

Marco Confalonieri

Full Text Available The lung alveoli slowly self-renew pneumocytes, but their facultative regeneration capacity is rapidly efficient after an injury, so fibrosis infrequently occurs. We recently observed Keratin 14 (KRT14 expression during diffuse alveolar damage (DAD, but not in controls. We wonder if KRT14 may be a marker of pneumocyte transition from quiescence to regeneration. Quantitative PCR and Western blot analyses highlighted the presence of KRT14 (mRNA and protein only in human lung samples with DAD or interstitial lung disease (ILD. In the exponentially growing cell lines A549 and H441, the mRNA and protein levels of KRT14 peaked at day one after cell seeding and decreased at day two, opposite to what observed for the proliferation marker E2F1. The inverse relation of KRT14 versus E2F1 expression holds true also for other proliferative markers, such as cyclin E1 and cyclin D1. Of interest, we also found that E2F1 silencing caused cell cycle arrest and increased KRT14 expression, whilst E2F1 stimulation induced cell cycle progression and decreased KRT14. KRT14 also increased in proliferative pneumocytes (HPAEpiC just before transdifferentiation. Overall, our results suggest that KRT14 is a viable biomarker of pneumocyte activation, and repair/regeneration. The involvement of KRT14 in regenerative process may suggest a novel pharmaceutical target to accelerate lung repair.

Effects of Thermal Stress on the mRNA Expression of SOD, HSP90, and HSP70 in the Spotted Sea Bass ( Lateolabrax maculatus)

Science.gov (United States)

Shin, Moon-Kyeong; Park, Ho-Ra; Yeo, Won-Jun; Han, Kyung-Nam

2018-03-01

The aim of this study was to elucidate the molecular mechanisms underlying the thermal stress response in the spotted sea bass ( Lateolabrax maculatus). Spotted sea basses were exposed to 4 different water temperatures (20, 22, 24, and 28°C) in increasing increments of 2°C/h from 18°C (control) for different time periods (0, 6, 12, 24, 48, 72, and 96 h). Subsequently, 3 tissues (liver, muscle, and gill) were isolated, and the levels of SOD, HSP90, and HSP70 mRNA were assessed. SOD mRNA expression was maintained at baseline levels of control fish at all water temperatures in the liver, while muscle and gill tissue showed an increase followed by a decrease over each certain time with higher water temperature. HSP90 mRNA expression increased in the liver at ≤ 24°C over time, but maintained baseline expression at 28°C. In muscle, HSP90 mRNA expression gradually increased at all water temperatures, but increased and then decreased at ≥ 24°C in gill tissue. HSP70 mRNA expression exhibited an increase and then a decrease in liver tissue at 28°C, but mainly showed similar expression patterns to HSP90 in all tissues. These results suggest the activity of a defense mechanism using SOD, HSP90, and HSP70 in the spotted sea bass upon rapid increases in water temperature, where the expression of these genes indicated differences between tissues in the extent of the defense mechanisms. Also, these results indicate that high water temperature and long-term thermal stress exposure can inhibit physiological defense mechanisms.

Genetic variation in ATP5O is associated with skeletal muscle ATP50 mRNA expression and glucose uptake in young twins.

Directory of Open Access Journals (Sweden)

Tina Rönn

Full Text Available BACKGROUND: Impaired oxidative capacity of skeletal muscle mitochondria contribute to insulin resistance and type 2 diabetes (T2D. Furthermore, mRNA expression of genes involved in oxidative phosphorylation, including ATP5O, is reduced in skeletal muscle from T2D patients. Our aims were to investigate mechanisms regulating ATP5O expression in skeletal muscle and association with glucose metabolism, and the relationship between ATP5O single nucleotide polymorphisms (SNPs and risk of T2D. METHODOLOGY/PRINCIPAL FINDINGS: ATP5O mRNA expression was analyzed in skeletal muscle from young (n = 86 and elderly (n = 68 non-diabetic twins before and after a hyperinsulinemic euglycemic clamp. 11 SNPs from the ATP5O locus were genotyped in the twins and a T2D case-control cohort (n = 1466. DNA methylation of the ATP5O promoter was analyzed in twins (n = 22 using bisulfite sequencing. The mRNA level of ATP5O in skeletal muscle was reduced in elderly compared with young twins, both during basal and insulin-stimulated conditions (p<0.0005. The degree of DNA methylation around the transcription start of ATP5O was <1% in both young and elderly twins and not associated with mRNA expression (p = 0.32. The mRNA level of ATP5O in skeletal muscle was positively related to insulin-stimulated glucose uptake (regression coefficient = 6.6; p = 0.02. Furthermore, two SNPs were associated with both ATP5O mRNA expression (rs12482697: T/T versus T/G; p = 0.02 and rs11088262: A/A versus A/G; p = 0.004 and glucose uptake (rs11088262: A/A versus A/G; p = 0.002 and rs12482697: T/T versus T/G; p = 0.005 in the young twins. However, we could not detect any genetic association with T2D. CONCLUSIONS/SIGNIFICANCE: Genetic variation and age are associated with skeletal muscle ATP5O mRNA expression and glucose disposal rate, suggesting that combinations of genetic and non-genetic factors may cause the reduced expression of ATP5O in T2D muscle. These findings propose a role for ATP5O, in

InterProScan Result: CK504225 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK504225 CK504225_3_ORF1 FC416AA9BCCD39B8 PFAM PF04615 Utp14 8.5e-24 T IPR006709 Small-subunit process...ome, Utp14 Biological Process: rRNA processing (GO:0006364)|Cellular Component: small-subunit processome (GO:0032040) ...

G-cimp status prediction of glioblastoma samples using mRNA expression data.

Science.gov (United States)

Baysan, Mehmet; Bozdag, Serdar; Cam, Margaret C; Kotliarova, Svetlana; Ahn, Susie; Walling, Jennifer; Killian, Jonathan K; Stevenson, Holly; Meltzer, Paul; Fine, Howard A

2012-01-01

Glioblastoma Multiforme (GBM) is a tumor with high mortality and no known cure. The dramatic molecular and clinical heterogeneity seen in this tumor has led to attempts to define genetically similar subgroups of GBM with the hope of developing tumor specific therapies targeted to the unique biology within each of these subgroups. Recently, a subset of relatively favorable prognosis GBMs has been identified. These glioma CpG island methylator phenotype, or G-CIMP tumors, have distinct genomic copy number aberrations, DNA methylation patterns, and (mRNA) expression profiles compared to other GBMs. While the standard method for identifying G-CIMP tumors is based on genome-wide DNA methylation data, such data is often not available compared to the more widely available gene expression data. In this study, we have developed and evaluated a method to predict the G-CIMP status of GBM samples based solely on gene expression data.

Efficacy of Omega Fatty Acid Supplementation on mRNA Expression Level of Tumor Necrosis Factor Alpha in Patients with Gastric Adenocarcinoma.

Science.gov (United States)

Hosseinzadeh, Asghar; Ardebili, Seyed Mojtaba Mohaddes

2016-09-01

Tumor necrosis factor alpha (TNF-α), a multifunctional cytokine, is involved in apoptosis, cell proliferation, cell survival, and inflammation. It plays a dual role in cancer development and progression. It has been revealed that polyunsaturated fatty acids (PUFAs) modulate the production and activity of TNF family cytokines. The objective of the present study was to evaluate the effect of PUFAs on messenger RNA expression levels of TNF-α in patients with gastric adenocarcinoma. Thirty-four chemotherapy-naive patients diagnosed with gastric adenocarcinoma were randomly divided into two groups. The first group (17 individuals) received cisplatin without supplements and the second group (17 individuals) received cisplatin plus orally administered PUFA supplements for 3 weeks, based on treatment strategies. The gastric biopsy samples were obtained from all participants before and after treatment, and TNF-α mRNA expression levels were evaluated by quantitative real-time PCR procedure. Our findings revealed that TNF-α mRNA expression is downregulated in group II, after receiving cisplatin and omega fatty acid supplement for 3 weeks. However, this difference is not statistically significant (p > 0.05). TNF-α mRNA expression did not show significant alteration in group I, after receiving cisplatin alone. Taken together, we concluded that omega fatty acids reduce TNF-α expression at the mRNA level in patients with gastric adenocarcinoma. These data suggest that TNF-α may act as a potential target for the therapy of human gastric adenocarcinoma.

The mRNA level of MLH1 in peripheral blood is a biomarker for the diagnosis of hereditary nonpolyposis colorectal cancer.

Science.gov (United States)

Yu, Hong; Li, Hui; Cui, Yongan; Xiao, Wei; Dai, Guihong; Huang, Junxing; Wang, Chaofu

2016-01-01

Hereditary nonpolyposis colorectal cancer (HNPCC) is caused by functional defects in mismatch repair (MMR) genes, including mutL homolog 1 (MLH1) and mutS homolog 2 (MSH2). This study aimed to assess whether the mRNA expression of MLH1 in peripheral blood could be used as a biomarkers for the diagnosis of HNPCC. The mRNA level of MLH1 was determined in 19 HNPCC families (46 members) using real-time quantitative polymerase chain reaction (qPCR). The mRNA levels of MLH1 in HNPCC were significantly lower than controls (P MLH1 for the diagnosis of HNPCC with the area under curve of 0.858. At an optimal cut-off value (0.511), the mRNA level of MLH1 had a sensitivity of 81.3% and a specificity of 86.7% for distinguishing HNPCC from controls. In conclusion, the mRNA expression of MLH1 in peripheral blood may serve as a biomarker for the diagnosis of HNPCC.

Paraoxonase-2 and paraoxonase-3: comparison of mRNA expressions in the placentae of unexplained intrauterine growth restricted and noncomplicated pregnancies.

Science.gov (United States)

Dikbas, Levent; Yapca, Omer Erkan; Dikbas, Neslihan; Gundogdu, Cemal

2017-05-01

Recent evidence suggests that oxidative stress is involved in the pathophysiology of many human diseases. It has been demonstrated that oxidative stress is associated with intrauterine growth restriction (IUGR), and the depletion of placental antioxidant systems has been suggested as a key factor in this disease. Our aims were to explore the possible role of antioxidant paraoxonase-2 (PON2) and paraoxonase-3 (PON3) in the pathophysiology of unexplained IUGR. We have studied the expression of mRNA for PON2, PON3 in placental tissues by using RT-qPCR. Two groups, consisting of normal (n = 18) and unexplained IUGR pregnancies (n = 20) were compared. Our results demonstrated that there were no significant differences in the mRNA expressions of PON2, PON3 between the two groups (p = 0.28, p = 0.90, respectively). PON2 and PON3 were down-regulated in IUGR. Antenatal steroid therapy had no effect on the expression mRNA in placentae of unexplained IUGR pregnancies compared to non-treated group. These results suggest that PON2, PON3 mRNA levels were not changed significantly in placentae of IUGR when compared to normal pregnant women.

Phosphorylation of the Fas associated factor FAF1 by protein kinase CK2 and identification of serines 289 and 291 as the in vitro phosphorylation sites

DEFF Research Database (Denmark)

Jensen, H H; Hjerrild, M; Guerra, B

2001-01-01

We previously identified the human Fas associated factor (FAF1) as one of the interacting partners of protein kinase CK2 beta subunit. Since FAF1 is a phosphoprotein we investigated whether it is a substrate for CK2. Here, we report the full length human FAF1 cDNA sequence, expression of FAF1...

[Correlation between the mRNA expression of tissue inhibitor of metalloproteinase-1 and apparent diffusion coefficient on diffusion-weighted imaging in rats' liver fibrosis].

Science.gov (United States)

Zhan, Yuefu; Liang, Xianwen; Han, Xiangjun; Chen, Jianqiang; Zhang, Shufang; Tan, Shun; Li, Qun; Wang, Xiong; Liu, Fan

2017-02-28

To explore the correlation between the apparent diffusion coefficient (ADC) and mRNA expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) in different stages of liver fibrosis in rats.
 Methods: A model of liver fibrosis in rats was established by intraperitoneal injection of high-fat diet combined with porcine serum. After drug administration for 4 weeks, 48 rats served as a model group and 12 rats served as a control group, then they underwent diffusion weighted imaging (DWI) scanning. The value of ADC was calculated at b value=800 s/mm2. The rats were sacrificed and carried out pathologic examination after DWI scanning immediately. The mRNA expression of TIMP-1 was detected by real time-polymerase chain reaction (RT-PCR). The rats of hepatic fibrosis were also divided into a S0 group (n=4), a S1 group (n=11), a S2 group (n=12), a S3 group (n=10), and a S4 group (n=9) according to their pathological stage. The value of ADC and the expression of TIMP-1 mRNA among the different stage groups of liver fibrosis were compared, and the correlation between ADC and the TIMP-1 mRNA were analyzed.
 Results: The ADC value and the TIMP-1 mRNA expression were significantly different between the control group and the liver fibrosis group (F=46.54 and 53.87, P0.05). For the comparison of TIMP-1 mRNA, there was no significant difference between the S1 group and the S2 group, the S3 group and the S4 group (both P>0.05). There were significant differences among the rest of the groups (all Pcorrelation analysis showed that there was a negative correlation between the ADC value and the TIMP-1 mRNA expression (r=-0.76, Pcorrelation between them.

Characterization and expression profile of the ovarian cytochrome P-450 aromatase (cyp19A1) gene during thermolabile sex determination in Pejerrey, Odontesthes bonariensis

Science.gov (United States)

Karube, M.; Fernandino, J.I.; Strobl-Mazzulla, P.; Strussmann, C.A.; Yoshizaki, G.; Somoza, G.M.; Patino, R.

2007-01-01

Cytochrome P450 aromatase (cyp19) is an enzyme that catalyzes the conversion of androgens to estrogens and may play a role in temperature- dependent sex determination (TSD) of reptiles, amphibians, and fishes. In this study, the ovarian P450 aromatase form (cyp19A1) of pejerrey Odontesthes bonariensis, a teleost with marked TSD, was cloned and its expression profile evaluated during gonadal differentiation at feminizing (17??C, 100% females), mixed-sex producing (24 and 25??C, 73.3 and 26.7% females, respectively), and masculinizing (29??C, 0% females) temperatures. The deduced cyp19A1 amino acid sequence shared high identity (>77.8%) with that from other teleosts but had low identity (<61.8%) with brain forms (cyp19A2), including that of pejerrey itself. The tissue distribution analysis of cyp19A1 mRNA in adult fish revealed high expression in the ovary. Semi-quantitative reverse transcription polymerase chain reaction analysis of the bodies of larvae revealed that cyp19A1 expression increased before the appearance of the first histological signs of ovarian differentiation at the feminizing temperature but remained low at the masculinizing temperature. The expression levels at mixed-sex producing temperatures were bimodal rather than intermediate, showing low and high modal values similar to those at the feminizing and masculinizing temperatures, respectively. The population percentages of high and low expression levels at intermediate temperatures were proportional to the percentage of females and males, respectively, and high levels were first observed at about the time of sex differentiation of females. These results suggest that cyp19A1 is involved in the process of ovarian formation and possibly also in the TSD of pejerrey. ?? 2007 Wiley-Liss, Inc.

Expression of mRNA for galanin, galanin-like peptide and galanin receptors 1-3 in the ovine hypothalamus and pituitary gland: effects of age and gender.

Science.gov (United States)

Whitelaw, Christine Margaret; Robinson, Jane Elizabeth; Chambers, George Ballantine; Hastie, Peter; Padmanabhan, Vasantha; Thompson, Robert Charles; Evans, Neil Price

2009-01-01

The neurotransmitters/neuromodulators galanin (GAL) and galanin-like peptide (GALP) are known to operate through three G protein-coupled receptors, GALR1, GALR2 and GALR3. The aim of this study was to investigate changes in expression of mRNA for galanin, GALP and GALR1-3 in the hypothalamus and pituitary gland, of male and female sheep, to determine how expression changed in association with growth and the attainment of reproductive competence. Tissue samples from the hypothalami and pituitary glands were analysed from late foetal and pre-pubertal lambs and adult sheep. Although mRNA for galanin and GALR1-3 was present in both tissues, at all ages and in both genders, quantification of GALP mRNA was not possible due to its low levels of expression. mRNA expression for both galanin and its receptors was seen to change significantly in both tissues as a function of age. Specifically, hypothalamic galanin mRNA expression increased with age in the male, but decreased with age in the female pituitary gland. mRNA expression for all receptors increased between foetal and pre-pubertal age groups and decreased significantly between pre-pubertal and adult animals. The results indicate that the expression of mRNA for galanin and its receptors changes dynamically with age and those significant differences exist with regard to tissue type and gender. These changes suggest that galaninergic neuroendocrine systems could be involved in the regulation of ovine growth and or the development of reproductive competence. The roles played by these systems in the sheep, however, may differ from other species, in particular the neuroendocrine link between nutrition and reproduction and GALR1's role in pituitary signalling.

Studies on mRNA expression of the somatostatin receptor family in lung cancer

International Nuclear Information System (INIS)

Wang Jing; Deng Jinglan; Wu Shengxi; Qiao Hongqing

2000-01-01

Objective: To investigate the characteristics of expression and distribution of 5 subtypes of somatostatin receptors (SSTR1∼5) in lung cancer. Methods: With [α- 35 S]dATP labelled oligonucleotides of the 5 SSTR subtypes as probes, using in situ hybridization, patterns of mRNA expression were detected in lung cancer tissue sections of 21 cases which fell in varied pathologic types. Additionally, Leica Q-500 image analyzing device was employed to semi-quantitatively analyze density of the expression. Results: Patterns of SSTR1∼5 expression in lung cancer were as follows: SSTR2 expression was dominant in small cell lung cancer (SCLC) while in non-small cell lung cancer (NSCLC) such as adenous and squamous, SSTR1 expression was stronger than that of the other 4 subtypes, In density of SSTR1∼5 expression in lung cancer, NSCLC was higher than SCLC (P<0.01). Conclusions: even though patterns and density of expression of SSTR subtypes in the lung cancer showed heterogeneity in different histopathologic types, as in SCLC and in NSCLC. Therefore, it has positive prospects for somatostatin analog-oriented agents to be used in treatment of both types of the lung cancers

Expression of Flk-1 and Cyclin D2 mRNA in the Myocardium of Rats with Doxorubicin-Induced Cardiomyopathy and after Treatment with Betulonic Acid Amide.

Science.gov (United States)

Mzhelskaya, M M; Klinnikova, M G; Koldysheva, E V; Lushnikova, E L

2017-10-01

The expression of VEGFR2 (Flk-1, according to immunohistochemistry) and of cyclin D2 mRNA (according to real-time PCR) in the myocardium of rats is studied in doxorubicin-induced cardiomyopathy and in response to betulonic acid amide. Doxorubicin alone and in combination with betulonic acid amide causes after 3 days a manifest reduction of cyclin D2 mRNA expression (by 38 and 63%, respectively), while injection of betulonic acid amide alone causes a 23-fold increase of cyclin D2 mRNA expression. An increase of cyclin D2 mRNA expression has been detected in all experimental groups after 14 days of experiment, the most pronounced in response to betulonic acid amide (63 times). The expression of Flk-1 in cardiomyocytes increases significantly in response to both chemical agents starting from day 3 of experiment. These results indicate that doxorubicin and betulonic acid amide induce cytoprotective reactions in the myocardium, first at the intracellular, then at the cellular levels.

Relationship between serum IGF-1 and skeletal muscle IGF-1 mRNA expression to phosphocreatine recovery after exercise in obese men with reduced GH.

Science.gov (United States)

Hamarneh, Sulaiman R; Murphy, Caitlin A; Shih, Cynthia W; Frontera, Walter; Torriani, Martin; Irazoqui, Javier E; Makimura, Hideo

2015-02-01

GH and IGF-1 are believed to be physiological regulators of skeletal muscle mitochondria. The objective of this study was to examine the relationship between GH/IGF-1 and skeletal muscle mitochondria in obese subjects with reduced GH secretion in more detail. Fifteen abdominally obese men with reduced GH secretion were treated for 12 weeks with recombinant human GH. Subjects underwent (31)P-magnetic resonance spectroscopy to assess phosphocreatine (PCr) recovery as an in vivo measure of skeletal muscle mitochondrial function and percutaneous muscle biopsies to assess mRNA expression of IGF-1 and mitochondrial-related genes at baseline and 12 weeks. At baseline, skeletal muscle IGF-1 mRNA expression was significantly associated with PCr recovery (r = 0.79; P = .01) and nuclear respiratory factor-1 (r = 0.87; P = .001), mitochondrial transcription factor A (r = 0.86; P = .001), peroxisome proliferator-activated receptor (PPAR)γ (r = 0.72; P = .02), and PPARα (r = 0.75; P = .01) mRNA expression, and trended to an association with PPARγ coactivator 1-α (r = 0.59; P = .07) mRNA expression. However, serum IGF-1 concentration was not associated with PCr recovery or any mitochondrial gene expression (all P > .10). Administration of recombinant human GH increased both serum IGF-1 (change, 218 ± 29 μg/L; P IGF-1 mRNA in muscle (fold change, 2.1 ± 0.3; P = .002). Increases in serum IGF-1 were associated with improvements in total body fat (r = -0.53; P = .04), trunk fat (r = -0.55; P = .03), and lean mass (r = 0.58; P = .02), but not with PCr recovery (P > .10). Conversely, increase in muscle IGF-1 mRNA was associated with improvements in PCr recovery (r = 0.74; P = .02), but not with body composition parameters (P > .10). These data demonstrate a novel association of skeletal muscle mitochondria with muscle IGF-1 mRNA expression, but independent of serum IGF-1 concentrations.

Resveratrol induces antioxidant and heat shock protein mRNA expression in response to heat stress in black-boned chickens.

Science.gov (United States)

Liu, L L; He, J H; Xie, H B; Yang, Y S; Li, J C; Zou, Y

2014-01-01

This study investigated the effects of dietary resveratrol at 0, 200, 400, or 600 mg/kg of diet on the performance, immune organ growth index, serum parameters, and expression levels of heat shock protein (Hsp) 27, Hsp70, and Hsp90 mRNA in the bursa of Fabricius, thymus, and spleen of 42-d-old female black-boned chickens exposed to heat stress at 37 ± 2°C for 15 d. The results showed that heat stress reduced daily feed intake and BW gain; decreased serum glutathione (GSH), growth hormone, and insulin-like growth factor-1 levels; and inhibited GSH peroxidase (GSH-Px), superoxide dismutase (SOD), and catalase (CAT) activities compared with birds subjected to thermo-neutral circumstances. Chickens that were fed diets supplemented with resveratrol exhibited a linear increase in feed intake and BW gain (P stress. In contrast, serum malonaldehyde concentrations were decreased (P stress also reduced (P stress and coincided with an increase in supplemental resveratrol levels. The expression of Hsp27, Hsp70, and Hsp90 mRNA in the bursa of Fabricius and spleen were increased (P stress compared with no heat stress. Resveratrol attenuated the heat stress-induced overexpression of Hsp27, Hsp70, and Hsp90 mRNA in the bursa of Fabricius and spleen and increased the low expression of Hsp27 and Hsp90 mRNA in thymus upon heat stress. The results suggest that supplemental resveratrol improves growth performance and reduces oxidative stress in heat-stressed black-boned chickens by increasing serum growth hormone concentrations and modulating the expression of heat shock genes in organs of the immune system.

InterProScan Result: CK503564 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK503564 CK503564_2_ORF2 AE9493CB5CA4BF90 PFAM PF00067 p450 6.3e-21 T IPR001128 Cytochrome P450 Molecular... Function: monooxygenase activity (GO:0004497)|Molecular Function: iron ion binding (GO:0005506)|Molecular... Function: electron carrier activity (GO:0009055)|Molecular Function: heme binding (GO:0020037) ...

InterProScan Result: CK533198 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK533198 CK533198_1_ORF1 249946CC09DDBD81 PRINTS PR00385 P450 6.6e-07 T IPR001128 Cytochrome P450 Molecular... Function: monooxygenase activity (GO:0004497)|Molecular Function: iron ion binding (GO:0005506)|Molecular... Function: electron carrier activity (GO:0009055)|Molecular Function: heme binding (GO:0020037) ...

InterProScan Result: CK534983 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK534983 CK534983_3_ORF1 2D41434E503577EF PRINTS PR00385 P450 2.2e-09 T IPR001128 Cytochrome P450 Molecular... Function: monooxygenase activity (GO:0004497)|Molecular Function: iron ion binding (GO:0005506)|Molecular... Function: electron carrier activity (GO:0009055)|Molecular Function: heme binding (GO:0020037) ...

InterProScan Result: CK534983 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK534983 CK534983_3_ORF1 2D41434E503577EF PFAM PF00067 p450 1.3e-43 T IPR001128 Cytochrome P450 Molecular... Function: monooxygenase activity (GO:0004497)|Molecular Function: iron ion binding (GO:0005506)|Molecular... Function: electron carrier activity (GO:0009055)|Molecular Function: heme binding (GO:0020037) ...

InterProScan Result: CK533198 [KAIKOcDNA[Archive

Lifescience Database Archive (English)

Full Text Available CK533198 CK533198_1_ORF1 249946CC09DDBD81 PFAM PF00067 p450 1.5e-44 T IPR001128 Cytochrome P450 Molecular... Function: monooxygenase activity (GO:0004497)|Molecular Function: iron ion binding (GO:0005506)|Molecular... Function: electron carrier activity (GO:0009055)|Molecular Function: heme binding (GO:0020037) ...

The effects of estrogen receptors α- and β-specific agonists and antagonists on cell proliferation and energy metabolism in human bone cell line.

Science.gov (United States)

Somjen, D; Katzburg, S; Sharon, O; Grafi-Cohen, M; Knoll, E; Stern, N

2011-02-01

In cultured human osteoblasts estradiol-17β (E2) modulated DNA synthesis, the specific activity of creatine kinase BB (CK), 12 and 15 lipoxygenase (LO) mRNA expression and formation of 12- and 15-hydroxyeicosatetraenoic acid (HETE). We now investigate the response of human bone cell line (SaOS2) to phytoestrogens and estrogen receptors (ER)-specific agonists and antagonists. Treatment of SaSO2 with E2, 2,3-bis (4-hydroxyphenyl)-propionitrile (DPN; ERβ-specific agonist), 4,4',4″-[4-propyl-(1H)-pyrazol-1,3,5-triyl] tris-phenol (PPT; ERα-specific agonist), biochainin A (BA), daidzein (D), genistein (G) and raloxifene (Ral) showed increased DNA synthesis and CK. Ral inhibited completely all stimulations except DPN and to some extent D. The ERα-specific antagonist methyl-piperidino-pyrazole (MPP) and the ERβ-specific antagonist 4-[2-phenyl-5,7-bis (tri-fluoro-methyl) pyrazolo [1,5-a]pyrimidin-3-yl] phenol (PTHPP) inhibited DNA synthesis, CK and reactive oxygen species (ROS) formation induced by estrogens according to their receptors affinity. The LO inhibitor baicaleine inhibited only E2, DPN and G's effects. E2 and Ral unlike all other compounds had no effect on ERα mRNA expression, while ERβ mRNA expression was stimulated by all compounds. All compounds modulated the expression of 12LO and 15LO mRNA, except E2, PPT and Ral for 12LO, and 12- and 15-HETE productions and stimulated ROS formation which was inhibited by NADPH oxidase inhibitors diphenyleneiodonium chloride (DPI) and N-acetyl cysteine and the estrogen inhibitor ICI. DPI did not affect hormonal-induced DNA and CK. In conclusion, we provide evidence for the separation of mediation via ERα and ERβ pathways in the effects of estrogenic compounds on osteoblasts, but the role of LO/HETE/ROS is unclear. Copyright © 2010 Wiley-Liss, Inc.

Leptin receptor (Ob-R) mRNA expression and serum leptin concentration in patients with colorectal and metastatic colorectal cancer

Energy Technology Data Exchange (ETDEWEB)

Erkasap, N.; Ozkurt, M. [Department of Physiology, Osmangazi University Medical Faculty, Meselik, Eskisehir (Turkey); Erkasap, S.; Yasar, F. [Department of General Surgery, Osmangazi University Medical Faculty, Meselik, Eskisehir (Turkey); Uzuner, K. [Department of Physiology, Osmangazi University Medical Faculty, Meselik, Eskisehir (Turkey); Ihtiyar, E. [Department of General Surgery, Osmangazi University Medical Faculty, Meselik, Eskisehir (Turkey); Uslu, S.; Kara, M. [Department of Biochemistry, Osmangazi University Medical Faculty, Meselik, Eskisehir (Turkey); Bolluk, O. [Department of Biostatistics, Osmangazi University Medical Faculty, Meselik, Eskisehir (Turkey)

2013-03-19

The objective of the present study was to investigate the effect of leptin on the progression of colorectal carcinoma to metastatic disease by analyzing the serum leptin concentration and Ob-R gene expression in colon cancer tissues. Tissue samples were obtained from 31 patients who underwent surgical resection for colon (18 cases) and metastatic colon (13 cases) cancer. Serum leptin concentration was determined by an enzyme-linked immunosorbent assay (ELISA) and Ob-R mRNA expression by real-time polymerase chain reaction (RT-PCR) for both groups. ELISA data were analyzed by the Student t-test and RT-PCR data were analyzed by the Mann-Whitney U-test. RT-PCR results demonstrated that mRNA expression of Ob-R in human metastatic colorectal cancer was higher than in local colorectal cancer tissues. On the other hand, mean serum leptin concentration was significantly higher in local colorectal cancer patients compared to patients with metastatic colorectal cancer. The results of the present study suggest a role for leptin in the progression of colon cancer to metastatic disease without weight loss. In other words, significantly increased Ob-R mRNA expression and decreased serum leptin concentration in patients with metastatic colon cancer indicate that sensitization to leptin activity may be a major indicator of metastasis to the colon tissue and the determination of leptin concentration and leptin gene expression may be used to aid the diagnosis.

Leptin receptor (Ob-R) mRNA expression and serum leptin concentration in patients with colorectal and metastatic colorectal cancer

International Nuclear Information System (INIS)

Erkasap, N.; Ozkurt, M.; Erkasap, S.; Yasar, F.; Uzuner, K.; Ihtiyar, E.; Uslu, S.; Kara, M.; Bolluk, O.

2013-01-01

The objective of the present study was to investigate the effect of leptin on the progression of colorectal carcinoma to metastatic disease by analyzing the serum leptin concentration and Ob-R gene expression in colon cancer tissues. Tissue samples were obtained from 31 patients who underwent surgical resection for colon (18 cases) and metastatic colon (13 cases) cancer. Serum leptin concentration was determined by an enzyme-linked immunosorbent assay (ELISA) and Ob-R mRNA expression by real-time polymerase chain reaction (RT-PCR) for both groups. ELISA data were analyzed by the Student t-test and RT-PCR data were analyzed by the Mann-Whitney U-test. RT-PCR results demonstrated that mRNA expression of Ob-R in human metastatic colorectal cancer was higher than in local colorectal cancer tissues. On the other hand, mean serum leptin concentration was significantly higher in local colorectal cancer patients compared to patients with metastatic colorectal cancer. The results of the present study suggest a role for leptin in the progression of colon cancer to metastatic disease without weight loss. In other words, significantly increased Ob-R mRNA expression and decreased serum leptin concentration in patients with metastatic colon cancer indicate that sensitization to leptin activity may be a major indicator of metastasis to the colon tissue and the determination of leptin concentration and leptin gene expression may be used to aid the diagnosis

Leptin receptor (Ob-R mRNA expression and serum leptin concentration in patients with colorectal and metastatic colorectal cancer

Directory of Open Access Journals (Sweden)

N. Erkasap

Full Text Available The objective of the present study was to investigate the effect of leptin on the progression of colorectal carcinoma to metastatic disease by analyzing the serum leptin concentration and Ob-R gene expression in colon cancer tissues. Tissue samples were obtained from 31 patients who underwent surgical resection for colon (18 cases and metastatic colon (13 cases cancer. Serum leptin concentration was determined by an enzyme-linked immunosorbent assay (ELISA and Ob-R mRNA expression by real-time polymerase chain reaction (RT-PCR for both groups. ELISA data were analyzed by the Student t-test and RT-PCR data were analyzed by the Mann-Whitney U-test. RT-PCR results demonstrated that mRNA expression of Ob-R in human metastatic colorectal cancer was higher than in local colorectal cancer tissues. On the other hand, mean serum leptin concentration was significantly higher in local colorectal cancer patients compared to patients with metastatic colorectal cancer. The results of the present study suggest a role for leptin in the progression of colon cancer to metastatic disease without weight loss. In other words, significantly increased Ob-R mRNA expression and decreased serum leptin concentration in patients with metastatic colon cancer indicate that sensitization to leptin activity may be a major indicator of metastasis to the colon tissue and the determination of leptin concentration and leptin gene expression may be used to aid the diagnosis.

G-cimp status prediction of glioblastoma samples using mRNA expression data.

Directory of Open Access Journals (Sweden)

Mehmet Baysan

Full Text Available Glioblastoma Multiforme (GBM is a tumor with high mortality and no known cure. The dramatic molecular and clinical heterogeneity seen in this tumor has led to attempts to define genetically similar subgroups of GBM with the hope of developing tumor specific therapies targeted to the unique biology within each of these subgroups. Recently, a subset of relatively favorable prognosis GBMs has been identified. These glioma CpG island methylator phenotype, or G-CIMP tumors, have distinct genomic copy number aberrations, DNA methylation patterns, and (mRNA expression profiles compared to other GBMs. While the standard method for identifying G-CIMP tumors is based on genome-wide DNA methylation data, such data is often not available compared to the more widely available gene expression data. In this study, we have developed and evaluated a method to predict the G-CIMP status of GBM samples based solely on gene expression data.

Effect of Degradation of Zearalenone-Contaminated Feed by Bacillus licheniformis CK1 on Postweaning Female Piglets

Directory of Open Access Journals (Sweden)

Guanhua Fu

2016-10-01

Full Text Available Zearalenone (ZEA, an estrogenic mycotoxin, is mainly produced by Fusarium fungi. In this study, Bacillus licheniformis CK1 isolated from soil with the capability of degrading ZEA was evaluated for its efficacy in reducing the adverse effects of ZEA in piglets. The gilts were fed one of the following three diets for 14 days: a basic diet for the control group; the basic diet supplemented with ZEA-contaminated basic diet for the treatment 1 (T1 group; and the basic diet supplemented with fermented ZEA-contaminated basic diet by CK1 for the treatment 2 (T2 group. The actual ZEA contents (analyzed were 0, 1.20 ± 0.11, 0.47 ± 0.22 mg/kg for the control, T1, and T2 diets, respectively. The results showed that the T1 group had significantly increased the size of vulva and the relative weight of reproductive organs compared to the control group at the end of the trial. The T1 group significantly decreased the concentration of the luteinizing hormone (LH compared with the control and T2 groups. Expression of ERβ was significantly up-regulated in the T2 group compared with the control. In addition, expression of ERβ was not different between the control and the T1 group. In summary, our results suggest that Bacillus licheniformis CK1 could detoxify ZEA in feed and reduce the adverse effects of ZEA in the gilts.

Effect of Degradation of Zearalenone-Contaminated Feed by Bacillus licheniformis CK1 on Postweaning Female Piglets.

Science.gov (United States)

Fu, Guanhua; Ma, Junfei; Wang, Lihong; Yang, Xin; Liu, Jeruei; Zhao, Xin

2016-10-17

Zearalenone (ZEA), an estrogenic mycotoxin, is mainly produced by Fusarium fungi. In this study, Bacillus licheniformis CK1 isolated from soil with the capability of degrading ZEA was evaluated for its efficacy in reducing the adverse effects of ZEA in piglets. The gilts were fed one of the following three diets for 14 days: a basic diet for the control group; the basic diet supplemented with ZEA-contaminated basic diet for the treatment 1 (T1) group; and the basic diet supplemented with fermented ZEA-contaminated basic diet by CK1 for the treatment 2 (T2) group. The actual ZEA contents (analyzed) were 0, 1.20 ± 0.11, 0.47 ± 0.22 mg/kg for the control, T1, and T2 diets, respectively. The results showed that the T1 group had significantly increased the size of vulva and the relative weight of reproductive organs compared to the control group at the end of the trial. The T1 group significantly decreased the concentration of the luteinizing hormone (LH) compared with the control and T2 groups. Expression of ERβ was significantly up-regulated in the T2 group compared with the control. In addition, expression of ERβ was not different between the control and the T1 group. In summary, our results suggest that Bacillus licheniformis CK1 could detoxify ZEA in feed and reduce the adverse effects of ZEA in the gilts.

In human granulosa cells from small antral follicles, androgen receptor mRNA and androgen levels in follicular fluid correlate with FSH receptor mRNA

DEFF Research Database (Denmark)

Nielsen, M. E.; Rasmussen, I. A.; Kristensen, S. G.

2011-01-01

significantly with the expression of AMHRII, but did not correlate with any of the hormones in the follicular fluid. These data demonstrate an intimate association between AR expression in immature granulosa cells, and the expression of FSHR in normal small human antral follicles and between the follicular......Human small antral follicles (diameter 3-9 mm) were obtained from ovaries surgically removed for fertility preservation. From the individual aspirated follicles, granulosa cells and the corresponding follicular fluid were isolated in 64 follicles, of which 55 were available for mRNA analysis (24...... and to the follicular fluid concentrations of AMH, inhibin-B, progesterone and estradiol. AR mRNA expression in granulosa cells and the follicular fluid content of androgens both showed a highly significant positive association with the expression of FSHR mRNA in granulosa cells. AR mRNA expression also correlated...

Hospital-acquired pneumonia after lung resection surgery is associated with characteristic cytokine gene expression.

LENUS (Irish Health Repository)

White, Mary

2012-02-01

BACKGROUND: Infection in humans has been linked with altered cytokine gene transcription. It is unclear whether this phenomenon is a consequence of an established disease process or precedes the infective process. The primary end point of this study was to determine whether hospital-acquired pneumonia (HAP) was associated with differential gene expression of interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, and IL-23p19. The secondary end point was to identify whether alteration in gene expression preceded the clinical onset of infection. METHODS: Sixty consecutive patients undergoing elective thoracic surgery were recruited. HAP was diagnosed as per National Nosocomial Infection Surveillance guidelines. Messenger RNA (mRNA) and protein levels were analyzed preoperatively and 24 h and 5 days postoperatively. RESULTS: Forty-one patients had an uncomplicated recovery. Nineteen patients developed HAP. IL-6, IL-10, IL-12p35, IL-23p19, IL-27p28, TNF-alpha, and IFN-gamma mRNA and protein levels of IL-6, IL-23, and IFN-gamma in peripheral blood leukocytes were analyzed before surgery and 24 h and 5 days postsurgery. IL-23p19 mRNA levels were reduced in the pneumonia group (median, 4.19; 10th-90th centile range, 3.90-4.71) compared with the nonpneumonia group (4.50; 3.85-5.32) day 1 postsurgery (P=02). IFN-gamma mRNA levels were reduced in the pneumonia group (2.48; 1.20-3.20) compared with nonpneumonia group (2.81; 2.10-3.26) (P=03) day 5 postsurgery. Results are expressed as log to base 10 copy numbers of cytokine mRNA per 10 million beta-actin mRNA copy numbers. All values are given as median and 10th to 90th centile range. CONCLUSIONS: Cytokine gene expression is altered immediately following surgery in patients with postoperative HAP.

Effect of hyperthyroidism on circulating prolactin and hypothalamic expression of tyrosine hydroxylase, prolactin signaling cascade members and estrogen and progesterone receptors during late pregnancy and lactation in the rat.

Science.gov (United States)

Pennacchio, Gisela E; Neira, Flavia J; Soaje, Marta; Jahn, Graciela A; Valdez, Susana R

2017-02-15

Hyperthyroidism (HyperT) compromises pregnancy and lactation, hindering suckling-induced PRL release. We studied the effect of HyperT on hypothalamic mRNA (RT-qPCR) and protein (Western blot) expression of tyrosine hydroxylase (TH), PRL receptor (PRLR) and signaling pathway members, estrogen-α (ERα) and progesterone (PR) receptors on late pregnancy (days G19, 20 and 21) and early lactation (L2) in rats. HyperT advanced pre-partum PRL release, reduced circulating PRL on L2 and increased TH mRNA (G21 and L2), p-TH, PRLR mRNA, STAT5 protein (G19 and L2), PRLR protein (G21) and CIS protein (G19). PRs mRNAs and protein decreased on G19 but afterwards PRA mRNA (G20), PRB mRNA (G21) and PRA mRNA and protein (L2) increased. ERα protein increased on G19 and decreased on G20. Thus, the altered hypothalamic PRLR, STAT5, PR and ERα expression in hyperthyroid rats may induce elevated TH expression and activation, that consequently, elevate dopaminergic tone during lactation, blunting suckling-induced PRL release and litter growth. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

L-DOPA decarboxylase mRNA expression is associated with tumor stage and size in head and neck squamous cell carcinoma: a retrospective cohort study

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Geomela Panagiota-Aikaterini

2012-10-01

Full Text Available Abstract Background Head and neck squamous cell carcinoma (HNSCC represents one of the most commonly diagnosed malignancies worldwide. The DDC gene encodes L-DOPA decarboxylase, an enzyme catalyzing the decarboxylation of L-DOPA to dopamine. We have recently shown that DDC mRNA is a significant predictor of patients’ prognosis in colorectal adenocarcinoma and prostate cancer. The aim of the current study was to analyze the DDC mRNA expression in HNSCC patients. Methods 53 malignant tumors were resected from the larynx, pharynx, tongue, buccal mucosa, parotid glands, and nasal cavity, as well as from 34 adjacent non-cancerous tissues of HNSCC patients, and were homogenized. Total RNA was isolated and converted into first-strand cDNA. An ultrasensitive real-time PCR method based on the SYBR Green chemistry was used for DDC mRNA quantification in head and neck tissue specimens. Relative quantification was performed using the comparative Ct (2-ddCt method. Results DDC mRNA levels were lower in squamous cell carcinomas (SCCs of the larynx and tongue than in adjacent non-cancerous tissue specimens. Furthermore, low DDC mRNA expression was noticed in laryngeal and tongue tumors of advanced TNM stage or bigger size, compared to early-stage or smaller tumors, respectively. No statistically significant differences were observed between SCCs resected from pharynx, buccal mucosa, or nasal cavity, and their normal counterparts. Conclusion This is the first study examining the DDC mRNA expression in HNSCC. According to our results, DDC mRNA expression may constitute a potential prognostic biomarker in tongue and/or larynx SCCs, which principally represent the overwhelming majority of HNSCC cases.

L-DOPA decarboxylase mRNA expression is associated with tumor stage and size in head and neck squamous cell carcinoma: a retrospective cohort study

International Nuclear Information System (INIS)

Geomela, Panagiota-Aikaterini; Kontos, Christos K; Yiotakis, Ioannis; Fragoulis, Emmanuel G; Scorilas, Andreas

2012-01-01

Head and neck squamous cell carcinoma (HNSCC) represents one of the most commonly diagnosed malignancies worldwide. The DDC gene encodes L-DOPA decarboxylase, an enzyme catalyzing the decarboxylation of L-DOPA to dopamine. We have recently shown that DDC mRNA is a significant predictor of patients’ prognosis in colorectal adenocarcinoma and prostate cancer. The aim of the current study was to analyze the DDC mRNA expression in HNSCC patients. 53 malignant tumors were resected from the larynx, pharynx, tongue, buccal mucosa, parotid glands, and nasal cavity, as well as from 34 adjacent non-cancerous tissues of HNSCC patients, and were homogenized. Total RNA was isolated and converted into first-strand cDNA. An ultrasensitive real-time PCR method based on the SYBR Green chemistry was used for DDC mRNA quantification in head and neck tissue specimens. Relative quantification was performed using the comparative Ct (2 -ddCt ) method. DDC mRNA levels were lower in squamous cell carcinomas (SCCs) of the larynx and tongue than in adjacent non-cancerous tissue specimens. Furthermore, low DDC mRNA expression was noticed in laryngeal and tongue tumors of advanced TNM stage or bigger size, compared to early-stage or smaller tumors, respectively. No statistically significant differences were observed between SCCs resected from pharynx, buccal mucosa, or nasal cavity, and their normal counterparts. This is the first study examining the DDC mRNA expression in HNSCC. According to our results, DDC mRNA expression may constitute a potential prognostic biomarker in tongue and/or larynx SCCs, which principally represent the overwhelming majority of HNSCC cases

Synthesis and Characterization of a Magnetically Active 19F Molecular Beacon.

Science.gov (United States)

Dempsey, Megan E; Marble, Hetal D; Shen, Tun-Li; Fawzi, Nicolas L; Darling, Eric M

2018-02-21

Gene expression is used extensively to describe cellular characteristics and behaviors; however, most methods of assessing gene expression are unsuitable for living samples, requiring destructive processes such as fixation or lysis. Recently, molecular beacons have become a viable tool for live-cell imaging of mRNA molecules in situ. Historically, beacon-mediated imaging has been limited to fluorescence-based approaches. We propose the design and synthesis of a novel molecular beacon for magnetic resonance detection of any desired target nucleotide sequence. The biologically compatible synthesis incorporates commonly used bioconjugation reactions in aqueous conditions and is accessible for laboratories without extensive synthesis capabilities. The resulting beacon uses fluorine ( 19 F) as a reporter, which is broadened, or turned "off", via paramagnetic relaxation enhancement from a stabilized nitroxide radical spin label when the beacon is not bound to its nucleic acid target. Therefore, the 19 F NMR signal of the beacon is quenched in its hairpin conformation when the spin label and the 19 F substituent are held in proximity, but the signal is recovered upon beacon hybridization to its specific complementary nucleotide sequence by physical separation of the radical from the 19 F reporter. This study establishes a path for magnetic resonance-based assessment of specific mRNA expression, providing new possibilities for applying molecular beacon technology in living systems.

Codon optimization of the HIV-1 vpu and vif genes stabilizes their mRNA and allows for highly efficient Rev-independent expression

International Nuclear Information System (INIS)

Nguyen, Kim-Lien; Llano, Manuel; Akari, Hirofumi; Miyagi, Eri; Poeschla, Eric M.; Strebel, Klaus; Bour, Stephan

2004-01-01

Two HIV-1 accessory proteins, Vpu and Vif, are notoriously difficult to express autonomously in the absence of the viral Tat and Rev proteins. We examined whether the codon bias observed in the vpu and vif genes relative to highly expressed human genes contributes to the Rev dependence and low expression level outside the context of the viral genome. The entire vpu gene as well as the 5' half of the vif gene were codon optimized and the resulting open reading frames (ORFs) (vphu and hvif, respectively) were cloned in autonomous expression vectors under the transcriptional control of the CMV promoter. Codon optimization efficiently removed the expression block observed in the native genes and allowed high levels of Rev- and Tat-independent expression of Vpu and Vif. Most of the higher protein levels detected are accounted for by enhanced steady-state levels of the mRNA encoding the optimized species. Nuclear run-on experiments show for the first time that codon optimization has no effect on the rate of transcriptional initiation or elongation of the vphu mRNA. Instead, optimization of the vpu gene was found to stabilize the vphu mRNA in the nucleus and enhance its export to the cytoplasm. This was achieved by allowing the optimized mRNA to use a new CRM1-independent nuclear export pathway. This work provides a better understanding of the molecular mechanisms underlying the process of codon optimization and introduces novel tools to study the biological functions of the Vpu and Vif proteins independently of other viral proteins

Effect of rat ovary irradiation or OVX on the expression of COLI and TGF-β1 mRNA in the rat bone

International Nuclear Information System (INIS)

Gao Yanhong; Gao Jianjun; Jin Weifang; Wang Hongfu

2003-01-01

To observe the effects of exposure of rat ovary to radiation or OVX on the expression of TGF-β 1 and COLI in the rat bone. The mRNA levels of TGF-β 1 and COLI in rat tibiae were measured with RT-PCR after the rat ovaries were irradiated by 50 Gy of 137 Cs γ-rays or OVX. For both the radiation group and the OVX group, the COLI mRNA level in the rat bone increased, whereas the TGF-β 1 decreased. Irradiation of ovary and OVX affect the expression of COLI and TGF-β 1 mRNA in bone probably in a similar way which is related to estrogen decrease

Elevated expression of H19 and Igf2 in the female mouse eye.

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Björn Reinius

Full Text Available The catalogue of genes expressed at different levels in the two sexes is growing, and the mechanisms underlying sex differences in regulation of the mammalian transcriptomes are being explored. Here we report that the expression of the imprinted non-protein-coding maternally expressed gene H19 was female-biased specifically in the female mouse eye (1.9-fold, p = 3.0E-6 while not being sex-biased in other somatic tissues. The female-to-male expression fold-change of H19 fell in the range expected from an effect of biallelic versus monoallelic expression. Recently, the possibility of sex-specific parent-of-origin allelic expression has been debated. This led us to hypothesize that H19 might express biallelically in the female mouse eye, thus escape its silencing imprint on the paternal allele specifically in this tissue. We therefore performed a sex-specific imprinting assay of H19 in female and male eye derived from a cross between Mus musculus and Mus spretus. However, this analysis demonstrated that H19 was exclusively expressed from the maternal gene copy, disproving the escape hypothesis. Instead, this supports that the female-biased expression of H19 is the result of upregulation of the single maternal. Furthermore, if H19 would have been expressed from both gene copies in the female eye, an associated downregulation of Insulin-like growth factor 2 (Igf2 was expected, since H19 and Igf2 compete for a common enhancer element located in the H19/Igf2 imprinted domain. On the contrary we found that also Igf2 was significantly upregulated in its expression in the female eye (1.2-fold, p = 6.1E-3, in further agreement with the conclusion that H19 is monoallelically elevated in females. The female-biased expression of H19 and Igf2 specifically in the eye may contribute to our understanding of sex differences in normal as well as abnormal eye physiology and processes.

UCP2 mRNA expression is dependent on glucose metabolism in pancreatic islets

DEFF Research Database (Denmark)

Dalgaard, Louise Torp

2012-01-01

Uncoupling Protein 2 (UCP2) is expressed in the pancreatic β-cell, where it partially uncouples the mitochondrial proton gradient, decreasing both ATP-production and glucose-stimulated insulin secretion (GSIS). Increased glucose levels up-regulate UCP2 mRNA and protein levels, but the mechanism f...... down-regulation of UCP2 is involved in preserving the insulin secretory capacity of GK mutant mice and might also be implicated in limiting disease progression in MODY2 patients....

RANKL/RANK/OPG cytokine receptor system: mRNA expression pattern in BPH, primary and metastatic prostate cancer disease.

Science.gov (United States)

Christoph, Frank; König, Frank; Lebentrau, Steffen; Jandrig, Burkhard; Krause, Hans; Strenziok, Romy; Schostak, Martin

2018-02-01

The cytokine system RANKL (receptor activator of NF-κB ligand), its receptor RANK and the antagonist OPG (osteoprotegerin) play a critical role in bone turnover. Our investigation was conducted to describe the gene expression at primary tumour site in prostate cancer patients and correlate the results with Gleason Score and PSA level. Seventy-one samples were obtained from prostate cancer patients at the time of radical prostatectomy and palliative prostate resection (n = 71). Patients with benign prostate hyperplasia served as controls (n = 60). We performed real-time RT-PCR after microdissection of the samples. The mRNA expression of RANK was highest in tumour tissue from patients with bone metastases (p BPH or locally confined tumours, also shown in clinical subgroups distinguished by Gleason Score (BPH tissue but did not exceed as much as in the tumour tissue. We demonstrated that RANK, RANKL and OPG are directly expressed by prostate cancer cells at the primary tumour site and showed a clear correlation with Gleason Score, serum PSA level and advanced disease. In BPH, mRNA expression is also detectable, but RANK expression does not exceed as much as compared to tumour tissue.

[mRNA expression of notch ligand-delta-like-1 and jagged-1 in mesenchymal stem cells of MDS patients].

Science.gov (United States)

Fei, Cheng-Ming; Gu, Shu-Cheng; Zhao, You-Shan; Guo, Juan; Li, Xiao; Chang, Chun-Kang

2014-12-01

This study was aimed to investigated the mRNA expression levels of Notch ligands- Delta-like-1 and Jagged-1 in bone marrow mesenchymal stem cells of patients with myelodysplastic syndrome (MDS), and to explore their relation with onset of MDS. Bone marrow mesenchymal stem cells of 38 patients with MDS and 16 normal subjects as control were collected to detect mRNA expression of Delta-like-1 and Jagged-1 by using real-time quantitative polymerase chain reaction. The results showed that the expression levels of Delta-like-1 and Jagged-1 in mesenchymal stem cells of MDS patients were significantly higher than that in normal controls (P MDS patients (r = 0.502, P MDS patients with abnormal karyotypes were significantly higher than those in MDS patients with normal karyotypes (P 0.05). It is concluded that the changes of Delta-like-1 and Jagged-1 expression level in MSC may play a role in the pathogenesis of myelodysplastic syndrome.

Effect of acute resistance exercise and sex on human patellar tendon structural and regulatory mRNA expression

DEFF Research Database (Denmark)

Sullivan, B.E.; Carroll, C.C.; Jemiolo, B.

2009-01-01

Sullivan BE, Carroll CC, Jemiolo B, Trappe SW, Magnusson SP, Dossing S, Kjaer M, Trappe TA. Effect of acute resistance exercise and sex on human patellar tendon structural and regulatory mRNA expression. J Appl Physiol 106: 468-475, 2009. First published November 20, 2008; doi: 10.1152/japplphysiol.......91341.2008.-Tendon is mainly composed of collagen and an aqueous matrix of proteoglycans that are regulated by enzymes called matrix metalloproteinases ( MMPs) and tissue inhibitors of metalloproteinases (TIMPs). Although it is known that resistance exercise (RE) and sex influence tendon metabolism...... and mechanical properties, it is uncertain what structural and regulatory components contribute to these responses. We measured the mRNA expression of tendon's main fibrillar collagens (type I and type III) and the main proteoglycans (decorin, biglycan, fibromodulin, and versican) and the regulatory enzymes MMP...

mRNA secondary structure at start AUG codon is a key limiting factor for human protein expression in Escherichia coli

International Nuclear Information System (INIS)

Zhang Weici; Xiao Weihua; Wei Haiming; Zhang Jian; Tian Zhigang

2006-01-01

Codon usage and thermodynamic optimization of the 5'-end of mRNA have been applied to improve the efficiency of human protein production in Escherichia coli. However, high level expression of human protein in E. coli is still a challenge that virtually depends upon each individual target genes. Using human interleukin 10 (huIL-10) and interferon α (huIFN-α) coding sequences, we systematically analyzed the influence of several major factors on expression of human protein in E. coli. The results from huIL-10 and reinforced by huIFN-α showed that exposing AUG initiator codon from base-paired structure within mRNA itself significantly improved the translation of target protein, which resulted in a 10-fold higher protein expression than the wild-type genes. It was also noted that translation process was not affected by the retained short-range stem-loop structure at Shine-Dalgarno (SD) sequences. On the other hand, codon-optimized constructs of huIL-10 showed unimproved levels of protein expression, on the contrary, led to a remarkable RNA degradation. Our study demonstrates that exposure of AUG initiator codon from long-range intra-strand secondary structure at 5'-end of mRNA may be used as a general strategy for human protein production in E. coli

From molecule to behavior: Brain aromatase (cyp19a1b) characterization, expression analysis and its relation with social status and male agonistic behavior in a Neotropical cichlid fish.

Science.gov (United States)

Ramallo, Martín R; Morandini, Leonel; Birba, Agustina; Somoza, Gustavo M; Pandolfi, Matías

2017-03-01

The enzyme aromatase, responsible for the conversion of C19 androgens to C18 estrogens, exists as two paralogue copies in teleost fish: Cyp19a1a mostly expressed in the gonads, referred as gonadal aromatase, and Cyp19a1b, mostly expressed in the brain, accordingly known as brain aromatase. The neural localization of Cyp19a1b is greatly contained within the social behavior network and mesolimbic reward system in fish, suggesting a strong role of estrogen synthesis in the regulation of social behavior. In this work we aimed to analyze the variation in cyp19a1b expression in brain and pituitary of males of a highly social cichlid, Cichlasoma dimerus (locally known as chanchita), and its relation with inter-individual variability in agonistic behavior in a communal social environment. We first characterized chanchita's cyp19a1b mRNA and deduced amino acid sequence, which showed a high degree of conservation when compared to other teleost brain aromatase sequences, and its tissue expression patterns. Within the brain, Cyp19a1b was solely detected at putative radial glial cells of the forebrain, close to the brain ventricles. We then studied the relative expression levels of cyp19a1b by Real Time PCR in the brain and pituitary of males of different social status, territorial vs. non-territorial, and its relationship with an index of agonistic behavior. We found that even though, brain aromatase expression did not differ between types of males, pituitary cyp19a1b expression levels positively correlated with the index of agonistic behavior. This suggests a novel role of the pituitary in the regulation of social behavior by local estrogen synthesis. Copyright © 2017 Elsevier Inc. All rights reserved.

Increased IL-17 and 22 mRNA expression in pediatric patients with otitis media with effusion.

Science.gov (United States)

Kwon, Oh Eun; Park, Sang Hyun; Kim, Sung Su; Shim, Haeng Seon; Kim, Min Gyeong; Kim, Young Il; Kim, Sang Hoon; Yeo, Seung Geun

2016-11-01

Middle ear effusion has been reported to be associated with immune responses in patients with otitis media with effusion (OME). Although various cytokines are involved in immunologic responses in patients with OME, no study to date has assessed the involvement of the pro-inflammatory cytokines interleukin (IL)-17 and IL-22. This study analyzed the levels of expression of IL-17 and IL-22 in the middle ear effusion of patients with OME. Patients aged Effusion fluid samples were obtained during surgery and levels of IL-17 and IL-22 mRNAs assessed by real-time PCR. IL-17 and IL-22 mRNA levels were compared in patients with effusion fluid positive and negative for bacteria; in patients with and without accompanying diseases, recurrent disease, and re-operation; and relative to fluid characteristics. The study cohort included 70 pediatric patients, 46 boys and 24 girls, of mean age 4.31 ± 2.11 years. The levels of IL-17 and IL-22 mRNA were higher in patients with than without sinusitis, but only IL-22 mRNA levels differed significantly (p < 0.05). The level of IL-17 mRNA was significantly higher in patients who did than did not undergo T&A (p < 0.05). The level of IL-22 expression was significantly higher in mucoid and purulent middle ear fluid samples than in serous fluid samples (p < 0.05). IL-17 and IL-22 mRNAs are involved in the pathophysiology of OME and are significantly higher in subjects with than without accompanying diseases. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.