A great numbers of cardiovascular disease patients all over the world are suffering in the poor outcomes. Under this situation, cardiac regeneration therapy to reorganize the postnatal heart that is defined as a terminal differentiated-organ is a very important theme and mission for human beings. However, the temporary success of several clinical trials using usual cell types with uncertain cell numbers has provided the transient effect of cell therapy to these patients. We therefore should r...
Lykke, Jacob Alexander; Langhoff-Roos, Jens; Lockwood, Charles J;
The combined effects of preterm delivery, small-for-gestational-age offspring, hypertensive disorders of pregnancy, placental abruption and stillbirth on early maternal death from cardiovascular causes have not previously been described in a large cohort. We investigated the effects of pregnancy ...
Lykke, Jacob Alexander; Langhoff-Roos, Jens; Lockwood, Charles J; Triche, Elizabeth W; Paidas, Michael J
small-for-gestational-age offspring 3.30 [2.25, 4.84]; preterm delivery, small-for-gestational-age offspring and pre-eclampsia 3.85 [2.07, 7.19]. Thus, we conclude that, separately and combined, preterm delivery and small-for-gestational-age are strong markers of early maternal death from both......The combined effects of preterm delivery, small-for-gestational-age offspring, hypertensive disorders of pregnancy, placental abruption and stillbirth on early maternal death from cardiovascular causes have not previously been described in a large cohort. We investigated the effects of pregnancy...... complications on early maternal death in a registry-based retrospective cohort study of 782 287 women with a first singleton delivery in Denmark 1978-2007, followed for a median of 14.8 years (range 0.25-30.2) accruing 11.6 million person-years. We employed Cox proportional hazard models of early death from...
Castle, Jason; Feinstein, Steven B
Using the improvements made in diagnostic contrast enhanced ultrasound, researchers have made significant progress in the field of ultrasound-mediated sonoporation for drug delivery. Many programs take advantage of commercial products; both ultrasound imaging systems and contrast agents to better enable translation from preclinical to first-in-man studies (Kotopoulis et al., Med Phys 40:072902, 2013). Particularly well-suited targets for this novel therapy are diseases of the cardiovascular system. This chapter will highlight several recent studies addressing treatment of both acute and chronic conditions. PMID:26486346
Lykke, J A; Paidas, M J; Damm, P; Triche, E W; Kuczynski, E; Langhoff-Roos, J
Preterm delivery has been shown to be associated with subsequent maternal cardiovascular morbidity. However, the impact of the severity and recurrence of preterm delivery on the risk of specific cardiovascular events and the metabolic syndrome in the mother, have not been investigated.......Preterm delivery has been shown to be associated with subsequent maternal cardiovascular morbidity. However, the impact of the severity and recurrence of preterm delivery on the risk of specific cardiovascular events and the metabolic syndrome in the mother, have not been investigated....
Takahashi, Hironobu; Letourneur, Didier; Grainger, David W.
This review focuses on the new and emerging large-molecule bioactive agents delivered from stent surfaces in drug-eluting stents (DES) to inhibit vascular restenosis in the context of interventional cardiology. New therapeutic agents representing proteins, nucleic acids (small interfering RNAs and large DNA plasmids), viral delivery vectors and even engineered cell therapies require specific delivery designs distinct from traditional smaller molecule approaches on DES. While small molecules a...
Full Text Available Rakesh K Sharma, Donald J Voelker, Roma Sharma, Hanumanth K ReddyUniversity of Arkansas for Medical Sciences, Medical Center of South Arkansas, El Dorado, AR, USAAbstract: Throughout their lifetime, an individual may sustain many injuries and recover spontaneously over a period of time, without even realizing the injury in the first place. Wound healing occurs due to a proliferation of stem cells capable of restoring the injured tissue. The ability of adult stem cells to repair tissue is dependent upon the intrinsic ability of tissues to proliferate. The amazing capacity of embryonic stem cells to give rise to virtually any type of tissue has intensified the search for similar cell lineage in adults to treat various diseases including cardiovascular diseases. The ability to convert adult stem cells into pluripotent cells that resemble embryonic cells, and to transplant those in the desired organ for regenerative therapy is very attractive, and may offer the possibility of treating harmful disease-causing mutations. The race is on to find the best cells for treatment of cardiovascular disease. There is a need for the ideal stem cell, delivery strategies, myocardial retention, and time of administration in the ideal patient population. There are multiple modes of stem cell delivery to the heart with different cell retention rates that vary depending upon method and site of injection, such as intra coronary, intramyocardial or via coronary sinus. While there are crucial issues such as retention of stem cells, microvascular plugging, biodistribution, homing to myocardium, and various proapoptotic factors in the ischemic myocardium, the regenerative potential of stem cells offers an enormous impact on clinical applications in the management of cardiovascular diseases.Keywords: stem cell therapy, stem cell delivery, cardiovascular diseases, myocardial infarction, cardiomyopathy
Sheng, Calvin C.; Li Zhou; Jijun Hao
Heart failure commonly results from an irreparable damage due to cardiovascular diseases (CVDs), the leading cause of morbidity and mortality in the United States. In recent years, the rapid advancements in stem cell research have garnered much praise for paving the way to novel therapies in reversing myocardial injuries. Cell types currently investigated for cellular delivery include embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), and adult stem cell lineages such as ske...
Poay; Sian; Sabrina; Lee; Kian; Keong; Poh
Endothelial dysfunction has been associated with the development of atherosclerosis and cardiovascular diseases. Adult endothelial progenitor cells(EPCs) are derived from hematopoietic stem cells and are capable of forming new blood vessels through a process of vas-culogenesis. There are studies which report correlations between circulating EPCs and cardiovascular risk fac-tors. There are also studies on how pharmacotherapies may influence levels of circulating EPCs. In this review, we discuss the potential role of endothelial progenitor cells as both diagnostic and prognostic biomarkers. In addition, we look at the interaction between cardio-vascular pharmacotherapies and endothelial progenitor cells. We also discuss how EPCs can be used directly and indirectly as a therapeutic agent. Finally, we evalu-ate the challenges facing EPC research and how these may be overcome.
Yu, Hee Tae; Park, Sungha; Shin, Eui-Cheol; Lee, Won-Woo
Age-related changes in the immune system, commonly termed "immunosenescence," contribute to deterioration of the immune response and fundamentally impact the health and survival of elderly individuals. Immunosenescence affects both the innate and adaptive immune systems; however, the most notable changes are in T cell immunity and include thymic involution, the collapse of T cell receptor (TCR) diversity, an imbalance in T cell populations, and the clonal expansion of senescent T cells. Senescent T cells have the ability to produce large quantities of proinflammatory cytokines and cytotoxic mediators; thus, they have been implicated in the pathogenesis of many chronic inflammatory diseases. Recently, an increasing body of evidence has suggested that senescent T cells also have pathogenic potential in cardiovascular diseases, such as hypertension, atherosclerosis, and myocardial infarction, underscoring the detrimental roles of these cells in various chronic inflammatory responses. Given that cardiovascular disease is the number one cause of death worldwide, there is great interest in understanding the contribution of age-related immunological changes to its pathogenesis. In this review, we discuss general features of age-related alterations in T cell immunity and the possible roles of senescent T cells in the pathogenesis of cardiovascular disease. PMID:26188489
Boos, Christopher J; Lip, Gregory Y H; Blann, Andrew D
Quantification of circulating endothelial cells (CECs) in peripheral blood is developing as a novel and reproducible method of assessing endothelial damage/dysfunction. The CECs are thought to be mature cells that have detached from the intimal monolayer in response to endothelial injury and are a different cell population to endothelial progenitor cells (EPCs). The EPCs are nonleukocytes derived from the bone marrow that are believed to have proliferative potential and may be important in vascular regeneration. Currently accepted methods of CEC quantification include the use of immunomagnetic bead separation (with cell counting under fluorescence microscopy) and flow cytometry. Several recent studies have shown increased numbers of CECs in cardiovascular disease and its risk factors, such as unstable angina, acute myocardial infarction, stroke, diabetes mellitus, and critical limb ischemia, but no change in stable intermittent claudication, essential hypertension, or atrial fibrillation. Furthermore, CEC quantification at 48 h after acute myocardial infarction has been shown to be an accurate predictor of major adverse coronary events and death at both 1 month and 1 year. This article presents an overview of the pathophysiology of CECs in the setting of cardiovascular disease and a brief comparison with EPCs. PMID:17045885
Full Text Available Korff Krause, Carsten Schneider, Kai Jaquet, Karl-Heinz KuckHanseatic Heart Center Hamburg, Department of Cardiology, Asklepios Hospital St. Georg, Hamburg, GermanyAbstract: The recent identification of bone marrow-derived adult stem cells and other types of stem cells that could improve heart function after transplantation have raised high expectations. The basic mechanisms have been studied mostly in murine models. However, these experiments revealed controversial results on transdifferentiation vs transfusion of adult stem cells vs paracrine effects of these cells, which is still being debated. Moreover, the reproducibility of these results in precisely translated large animal models is still less well investigated. Despite these weaknesses results of several clinical trials including several hundreds of patients with ischemic heart disease have been published. However, there are no solid data showing that any of these approaches can regenerate human myocardium. Even the effectiveness of cell therapy in these approaches is doubtful. In future we need in this important field of regenerative medicine: i more experimental data in large animals that are closer to the anatomy and physiology of humans, including data on dose effects, comparison of different cell types and different delivery routes; ii a better understanding of the molecular mechanisms involved in the fate of transplanted cells; iii more intensive research on genuine regenerative medicine, applying genetic regulation and cell engineering.Keywords: stem cells, cardiovascular disease
Sheng, Calvin C; Zhou, Li; Hao, Jijun
Heart failure commonly results from an irreparable damage due to cardiovascular diseases (CVDs), the leading cause of morbidity and mortality in the United States. In recent years, the rapid advancements in stem cell research have garnered much praise for paving the way to novel therapies in reversing myocardial injuries. Cell types currently investigated for cellular delivery include embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs), and adult stem cell lineages such as skeletal myoblasts, bone-marrow-derived stem cells (BMSCs), mesenchymal stem cells (MSCs), and cardiac stem cells (CSCs). To engraft these cells into patients' damaged myocardium, a variety of approaches (intramyocardial, transendocardial, transcoronary, venous, intravenous, intracoronary artery and retrograde venous administrations and bioengineered tissue transplantation) have been developed and explored. In this paper, we will discuss the pros and cons of these delivery modalities, the current state of their therapeutic potentials, and a multifaceted evaluation of their reported clinical feasibility, safety, and efficacy. While the issues of optimal delivery approach, the best progenitor stem cell type, the most effective dose, and timing of administration remain to be addressed, we are highly optimistic that stem cell therapy will provide a clinically viable option for myocardial regeneration. PMID:23509740
Calvin C. Sheng
Full Text Available Heart failure commonly results from an irreparable damage due to cardiovascular diseases (CVDs, the leading cause of morbidity and mortality in the United States. In recent years, the rapid advancements in stem cell research have garnered much praise for paving the way to novel therapies in reversing myocardial injuries. Cell types currently investigated for cellular delivery include embryonic stem cells (ESCs, induced pluripotent stem cells (iPSCs, and adult stem cell lineages such as skeletal myoblasts, bone-marrow-derived stem cells (BMSCs, mesenchymal stem cells (MSCs, and cardiac stem cells (CSCs. To engraft these cells into patients’ damaged myocardium, a variety of approaches (intramyocardial, transendocardial, transcoronary, venous, intravenous, intracoronary artery and retrograde venous administrations and bioengineered tissue transplantation have been developed and explored. In this paper, we will discuss the pros and cons of these delivery modalities, the current state of their therapeutic potentials, and a multifaceted evaluation of their reported clinical feasibility, safety, and efficacy. While the issues of optimal delivery approach, the best progenitor stem cell type, the most effective dose, and timing of administration remain to be addressed, we are highly optimistic that stem cell therapy will provide a clinically viable option for myocardial regeneration.
Full Text Available The present paper aims to review the main pathophysiological links between red blood cell disorders and cardiovascular diseases, provides a brief description of the latest studies in this area, and considers implications for clinical practice and therapy. Anemia is associated with a special risk in proatherosclerotic conditions and heart disease and became a new therapeutic target. Guidelines must be updated for the management of patients with red blood cell disorders and cardiovascular diseases, and targets for hemoglobin level should be established. Risk scores in several cardiovascular diseases should include red blood cell count and RDW. Complete blood count and hemorheological parameters represent useful, inexpensive, widely available tools for the management and prognosis of patients with coronary heart disease, heart failure, hypertension, arrhythmias, and stroke. Hypoxia and iron accumulation cause the most important cardiovascular effects of sickle cell disease and thalassemia. Patients with congenital chronic hemolytic anemia undergoing splenectomy should be monitored, considering thromboembolic and cardiovascular risk.
Kritikou, Eva; Kuiper, Johan; Kovanen, Petri T; Bot, Ilze
Mast cells comprise an innate immune cell population, which accumulates in tissues proximal to the outside environment and, upon activation, augments the progression of immunological reactions through the release and diffusion of either pre-formed or newly generated mediators. The released products of mast cells include histamine, proteases, as well as a variety of cytokines, chemokines and growth factors, which act on the surrounding microenvironment thereby shaping the immune responses triggered in various diseased states. Mast cells have also been detected in the arterial wall and are implicated in the onset and progression of numerous cardiovascular diseases. Notably, modulation of distinct mast cell actions using genetic and pharmacological approaches highlights the crucial role of this cell type in cardiovascular syndromes. The acquired evidence renders mast cells and their mediators as potential prognostic markers and therapeutic targets in a broad spectrum of pathophysiological conditions related to cardiovascular diseases. PMID:25959384
Stamm, Christof; Klose, Kristin; Choi, Yeong-Hoon
Regenerative medicine encompasses "tissue engineering" - the in vitro fabrication of tissues and/or organs using scaffold material and viable cells - and "cell therapy" - the transplantation or manipulation of cells in diseased tissue in vivo. In the cardiovascular system, tissue engineering strategies are being pursued for the development of viable replacement blood vessels, heart valves, patch material, cardiac pacemakers and contractile myocardium. Anecdotal clinical applications of such vessels, valves and patches have been described, but information on systematic studies of the performance of such implants is not available, yet. Cell therapy for cardiovascular regeneration, however, has been performed in large series of patients, and numerous clinical studies have produced sometimes conflicting results. The purpose of this chapter is to summarize the clinical experience with cell therapy for diseases of the cardiovascular system, and to analyse possible factors that may influence its outcome.
Patients suffering from myocardial infarction (MI), atherosclerosis and Hereditary Hemorrhagic Telangiectasia type 1 (HHT-1) all have diseased and dysfunctional blood vessels. Cardiovascular repair in these diseases occurs not only locally, but also peripheral blood (progenitor) cells and cytokines/growth factors positively contribute to repair of malfunctioning tissue. In this thesis several aspects of cardiovascular repair have been explored. First, we show that in MI patients relatively la...
Regenerative medicine is an exciting field that aims to create regenerative alternatives to harvest tissues for transplantation. In this approach, the delivery of cells and biological molecules plays a central role. The scaffold (synthetic temporary extracellular matrix) delivers cells to the regenerative site and provides three-dimensional environments for the cells. To fulfil these functions, we design biodegradable polymer scaffolds with structural features on multiple size scales. To enhance positive cell-material interactions, we design nano-sized structural features in the scaffolds to mimic the natural extracellular matrix. We also integrate micro-sized pore networks to facilitate mass transport and neo tissue regeneration. We also design novel polymer devices and self-assembled nanospheres for biomolecule delivery to recapitulate key events in developmental and wound healing processes. Herein, we present recent work in biomedical polymer synthesis, novel processing techniques, surface engineering and biologic delivery. Examples of enhanced cellular/tissue function and regenerative outcomes of these approaches are discussed to demonstrate the excitement of the biomimetic scaffold design and biologic delivery in regenerative medicine. (topical review)
Ian O Smith and Peter X Ma
Full Text Available Regenerative medicine is an exciting field that aims to create regenerative alternatives to harvest tissues for transplantation. In this approach, the delivery of cells and biological molecules plays a central role. The scaffold (synthetic temporary extracellular matrix delivers cells to the regenerative site and provides three-dimensional environments for the cells. To fulfil these functions, we design biodegradable polymer scaffolds with structural features on multiple size scales. To enhance positive cell–material interactions, we design nano-sized structural features in the scaffolds to mimic the natural extracellular matrix. We also integrate micro-sized pore networks to facilitate mass transport and neo tissue regeneration. We also design novel polymer devices and self-assembled nanospheres for biomolecule delivery to recapitulate key events in developmental and wound healing processes. Herein, we present recent work in biomedical polymer synthesis, novel processing techniques, surface engineering and biologic delivery. Examples of enhanced cellular/tissue function and regenerative outcomes of these approaches are discussed to demonstrate the excitement of the biomimetic scaffold design and biologic delivery in regenerative medicine.
Gonzales, Christine; Ullrich, Nina D.; Gerber, Stefan; Berthonneche, Corinne; Niggli, Ernst; Pedrazzini, Thierry
Weakening of cardiac function in patients with heart failure results from a loss of cardiomyocytes in the damaged heart. Cell replacement therapies as a way to induce myocardial regeneration in humans could represent attractive alternatives to classical drug-based approaches. However, a suitable source of precursor cells, which could produce a functional myocardium after transplantation, remains to be identified. In the present study, we isolated cardiovascular precursor cells from ventricles...
Mazhari, Ramesh; Hare, Joshua M.
The possibility of using stem cells to regenerate damaged myocardium has been actively investigated since the late 1990s. Consistent with the traditional view that the heart is a “post-mitotic” organ that possesses minimal capacity for self-repair, much of the preclinical and clinical work has focused exclusively on introducing stem cells into the heart, with the hope of differentiation of these cells into functioning cardiomyocytes. This approach is ongoing and retains promise but to date ha...
Clarke, Murray; Bennett, Martin; Littlewood, Trevor
Cell death is important for both development and tissue homeostasis in the adult. As such, it is tightly controlled and deregulation is associated with diverse pathologies; for example, regulated cell death is involved in vessel remodelling during development or following injury, but deregulated death is implicated in pathologies such as atherosclerosis, aneurysm formation, ischaemic and dilated cardiomyopathies and infarction. We describe the mechanisms of cell death and its role in the norm...
Levade, T; Augé, N; Veldman, R J; Cuvillier, O; Nègre-Salvayre, A; Salvayre, R
Sphingolipids have emerged as a new class of lipid mediators. In response to various extracellular stimuli, sphingolipid turnover can be stimulated in vascular cells and cardiac myocytes. Subsequent generation of sphingolipid molecules such as ceramide, sphingosine, and sphingosine-1-phosphate, is followed by regulation of ion fluxes and activation of various signaling pathways leading to smooth muscle cell proliferation, endothelial cell differentiation or apoptotic cell death, cell contraction, retraction, or migration. The importance of sphingolipids in cardiovascular signaling is illustrated by recent observations implicating them in physiological processes such as vasculogenesis as well as in frequent pathological conditions, including atherosclerosis and its complications. PMID:11717151
Yang, Ningning; Singh, Saurabh; Mahato, Ram I
Triplex-forming oligonucleotides (TFOs) represent an antigene approach for gene regulation through direct interaction with genomic DNA. While this strategy holds great promise owing to the fact that only two alleles need silencing to impact gene regulation, delivering TFOs to target cells in vivo is still a challenge. Our recent efforts have focused on conjugating TFOs to carrier molecules like cholesterol to enhance their cellular uptake and mannose-6-phosphate-bovine serum albumin (M6P-BSA) to target TFO delivery to hepatic stellate cells (HSCs) for treating liver fibrosis. These approaches however are rendered less effective owing to a lack of targeted delivery, as seen with lipid-conjugates, and the potential immune reactions due to repeated dosing with high molecular weight BSA conjugated TFO. In this review, we discuss our latest efforts to enhance the effectiveness of TFO for treating liver fibrosis. We have shown that conjugation of TFOs to M6P-HPMA can enhance TFO delivery to HSCs and has the potential to treat liver fibrosis by inhibiting collagen synthesis. This TFO conjugate shows negligible immunogenicity owing to the use of HPMA, one of the least immunogenic copolymers, thereby making it a suitable and more effective candidate for antifibrotic therapy. PMID:21763370
Full Text Available Patients with end stage heart failure have very few treatment options. The long waiting times for transplant and the complications associated with immunosuppression has led to the search for alternatives. Subsequent to the isolation and characterization of stem cells, tremendous advances have been made and the safety and feasibility of autologous bone marrow derived stem cells has been proven in preclinical studies. Clinical studies have also shown mobilized cells repair the infracted heart, improving function and survival. We have started a clinical study to evaluate the efficacy of bone marrow derived stem cells. Bone-marrow was aspirated from the right iliac crest and the stem cells were isolated by density gradient method and suspended according to the mode of delivery.From Jan 2007 till date 10 patients (8 adults, 2 children, age with end stage cardiovascular disorder of varied etiology (Ischemic left ventricular dysfunction - 6 patients, Primary pulmonary hypertension - 2 patients, Dilated cardiomyopathy -1 patient, Biventricular non-compaction -1 patient underwent stem cell therapy. All patients were evaluated and cardiac function was measured by using echocardiography and thallium scintigraphy. There were no procedure related complications. These patients are being regularly followed-up and one patient who has completed 6-month follow-up has shown improvement in perfusion as well as increase in ejection fraction of 10%. Stem cell therapy in patients with end-stage cardiovascular disorder might be a promising tool by means of angiogenesis and other paracrine mechanisms.
Jiang, Wenjian; Lan, Feng; Zhang, Hongjia
Cardiovascular cells derived from patient specific induced Pluripotent Stem Cell (iPSC) harbor gene mutations associated with the pathogenesis of inherited cardiac diseases and congenital heart diseases (CHD). Numerous reports have demonstrated the utilization of human induced Pluripotent Stem Cell (hiPSC) to model cardiac diseases as a means of investigating their underlying mechanisms. So far, they have been shown to investigate the molecular mechanisms of many cardiac disorders, such as long-QT syndrome (LQT), catecholaminergic polymorphic ventricular tachycardia (CPVT), dilated cardiomyopathy (DCM), hypertrophic cardiomyopathy (HCM), LEOPARD syndrome (LS), arrhythmogenic cardiomyopathy (ACM), Friedreich ataxia (FRDA), Barth syndrome (BTHS), hypoplastic left heart syndrome (HLHS), Marfan syndrome (MFS) and other CHD. This article summarizes the growing body of research related to modeling various cardiac diseases using hiPSCs. Moreover, by reviewing the methods used in previous studies, we propose multiple novel applications of hiPSCs to investigate comprehensive cardiovascular disorders and facilitate drug discovery. PMID:25322695
Full Text Available Intracellular delivery of biomolecules, such as proteins and siRNAs, into primary immune cells, especially resting lymphocytes, is a challenge. Here we describe the design and testing of microfluidic intracellular delivery systems that cause temporary membrane disruption by rapid mechanical deformation of human and mouse immune cells. Dextran, antibody and siRNA delivery performance is measured in multiple immune cell types and the approach's potential to engineer cell function is demonstrated in HIV infection studies.
Colone, Marisa; Kaliappan, Subramanian; Calcabrini, Annarica; Tortora, Mariarosaria; Cavalieri, Francesca; Stringaro, Annarita
Recently, nanomedicine has received increasing attention for its ability to improve the efficacy of cancer therapeutics. Nanosized polymer therapeutic agents offer the advantage of prolonged circulation in the blood stream, targeting to specific sites, improved efficacy and reduced side effects. In this way, local, controlled delivery of the drug will be achieved with the advantage of a high concentration of drug release at the target site while keeping the systemic concentration of the drug low, thus reducing side effects due to bioaccumulation. Various drug delivery systems such as nanoparticles, liposomes, microparticles and implants have been demonstrated to significantly enhance the preventive/therapeutic efficacy of many drugs by increasing their bioavailability and targetability. As these carriers significantly increase the therapeutic effect of drugs, their administration would become less cost effective in the near future. The purpose of our research work is to develop a delivery system for breast cancer cells using a microvector of drugs. These results highlight the potential uses of these responsive platforms suited for biomedical and pharmaceutical applications. At the request of all authors of the paper an updated version was published on 12 July 2016. The manuscript was prepared and submitted without Dr. Francesca Cavalieri's contribution and her name was added without her consent. Her name has been removed in the updated and re-published article.
Full Text Available Guilei Ma,1,# Yong Wang,1,# Ilia Fishbein,2 Mei Yu,1 Linhua Zhang,1 Ivan S Alferiev,2 Jing Yang,1 Cunxian Song,1 Robert J Levy2 1Institute of Biomedical Engineering, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin, People's Republic of China; 2Children's Hospital of Philadelphia, Abramson Research Building, Philadelphia, PA, USA #These authors contributed equally to this work Purpose: To investigate the anchoring of plasmid DNA/anti-DNA antibody/cationic lipid tri-complex (DAC micelles onto bisphosphonate-modified 316 L coronary stents for cardiovascular site-specific gene delivery. Methods: Stents were first modified with polyallylamine bisphosphonate (PAA-BP, thereby enabling the retention of a PAA-BP molecular monolayer that permits the anchoring (via vector-binding molecules of DAC micelles. DAC micelles were then chemically linked onto the PAA-BP-modified stents by using N-succinimidyl-3-(2-pyridyldithiol-propionate (SPDP as a crosslinker. Rhodamine-labeled DNA was used to assess the anchoring of DAC micelles, and radioactive-labeled antibody was used to evaluate binding capacity and stability. DAC micelles (encoding green fluorescent protein were tethered onto the PAA-BP-modified stents, which were assessed in cell culture. The presence of a PAA-BP molecular monolayer on the steel surface was confirmed by X-ray photoelectron spectroscopy and atomic force microscope analysis. Results: The anchoring of DAC micelles was generally uniform and devoid of large-scale patches of defects. Isotopic quantification confirmed that the amount of antibody chemically linked on the stents was 17-fold higher than that of the physical adsorbed control stents and its retention time was also significantly longer. In cell culture, numerous green fluorescent protein-positive cells were found on the PAA-BP modified stents, which demonstrated high localization and efficiency of gene delivery. Conclusion: The DAC micelle
Faiella, Whitney; Atoui, Rony
Stem cell treatments are a desirable therapeutic option to regenerate myocardium and improve cardiac function after myocardial infarction. Several different types of cells have been explored, each with their own benefits and limitations. Induced pluripotent stem cells possess an embryonic-like state and therefore have a high proliferative capacity, but they also pose a risk of teratoma formation. Mesenchymal stem cells have been investigated from both bone marrow and adipose tissue. Their immunomodulatory characteristics may permit the use of allogeneic cells as universal donor cells in the future. Lastly, studies have consistently shown that cardiac stem cells are better able to express markers of cardiogenesis compared to other cell types, as well improve cardiac function. The ideal source of stem cells depends on multiple factors such as the ease of extraction/isolation, effectiveness of engraftment, ability to differentiate into cardiac lineages and effect on cardiac function. Although multiple studies highlight the benefits and limitations of each cell type and reinforce the successful potential use of these cells to regenerate damaged myocardium, more studies are needed to directly compare cells from various sources. It is interesting to note that research using stem cell therapies is also expanding to treat other cardiovascular diseases including non-ischemic cardiomyopathies. PMID:27539581
Bietenbeck, Michael; Florian, Anca; Faber, Cornelius; Sechtem, Udo; Yilmaz, Ali
Magnetic resonance imaging (MRI) allows for an accurate assessment of both functional and structural cardiac parameters, and thereby appropriate diagnosis and validation of cardiovascular diseases. The diagnostic yield of cardiovascular MRI examinations is often increased by the use of contrast agents that are almost exclusively based on gadolinium compounds. Another clinically approved contrast medium is composed of superparamagnetic iron oxide nanoparticles (IONs). These particles may expand the field of contrast-enhanced cardiovascular MRI as recently shown in clinical studies focusing on acute myocardial infarction (AMI) and atherosclerosis. Furthermore, IONs open up new research opportunities such as remote magnetic drug targeting (MDT). The approach of MDT relies on the coupling of bioactive molecules and magnetic nanoparticles to form an injectable complex. This complex, in turn, can be attracted to and retained at a desired target inside the body with the help of applied magnetic fields. In comparison to common systemic drug applications, MDT techniques promise both higher concentrations at the target site and lower concentrations elsewhere in the body. Moreover, concurrent or subsequent MRI can be used for noninvasive monitoring of drug distribution and successful delivery to the desired organ in vivo. This review does not only illustrate the basic conceptual and biophysical principles of IONs, but also focuses on new research activities and achievements in the cardiovascular field, mainly in the management of AMI. Based on the presentation of successful MDT applications in preclinical models of AMI, novel approaches and the translational potential of MDT are discussed. PMID:27486321
Bogt, Koen Elzert Adriaan van der
Stem cell therapy has raised enthusiasm as a potential treatment for cardiovascular diseases. However, questions remain about the in vivo behavior of the cells after transplantation and the mechanism of action with which the cells could potentially alleviate disease symptoms. The objective of the re
Byrne, W L; Murphy, C T; Cronin, M; Wirth, T; Tangney, M
Phagocytic cells including macrophages, dendritic cells and neutrophils are now recognised as playing a negative role in many disease settings including cancer. In particular, macrophages are known to play a pathophysiological role in multiple diseases and present a valid and ubiquitous therapeutic target. The technology to target these phagocytic cells in situ, both selectively and efficiently, is required in order to translate novel therapeutic modalities into clinical reality. We present a novel delivery strategy using non-pathogenic bacteria to effect gene delivery specifically to tumour-associated phagocytic cells. Non-invasive bacteria lack the ability to actively enter host cells, except for phagocytic cells. We exploit this natural property to effect 'passive transfection' of tumour-associated phagocytic cells following direct administration of transgene-loaded bacteria to tumour regions. Using an in vitro-differentiated human monocyte cell line and two in vivo mouse models (an ovarian cancer ascites and a solid colon tumour model) proof of delivery is demonstrated with bacteria carrying reporter constructs. The results confirm that the delivery strategy is specific for phagocytic cells and that the bacterial vector itself recruits more phagocytic cells to the tumour. While proof of delivery to phagocytic cells is demonstrated in vivo for solid and ascites tumour models, this strategy may be applied to other settings, including non-cancer related disease. PMID:25466954
Tao WANG; Ling-bo HOU; Zhen-jun LIU; Yan WANG; Chun-lian CHEN; Xiao XIAO; Dao-wen WANG
Aim: The overexpression of the human tissue kallikrein (HK) gene can reduce blood pressure and ameliorate the secondary syndromes associated with hyper- tension in animal models. The current study was designed to investigate hy- potensive effect of intramuscular delivery of HK gene. Methods: We generated an recombinant adeno-associated virus (rAAV) vector expressing human tissue kallikrein under the control of a cytomegalovirus promoter and administered the rAAV-HK vector to a spontaneously hypertensive rat model at a dose of 1× 1010 virons/rat through intramuscular injection. Results: A persistent, high-level ex- pression of HK post-gene delivery was confirmed by ELISA. The systolic blood pressure in the rats receiving rAAV-LacZ and saline increased from 171.3 mmHg to 182.3 mmHg 28 weeks' post injection. In contrast, the delivery of the HK gene by AAV vectors attenuated the increase of the systolic blood pressure in the treated group. The systolic blood pressure was only slightly lowered (from a level of 174 mmHg to 170.5 mmHg) post-vector administration. The difference in blood pres- sure between the treated group and the control groups is statistically significant at 12.6 mmHg. The hypotensive effect of rAAV-HK persisted until the end of the testing period. In addition, a significant amelioration of cardiovascular hypertrophy, renal injury, and collagen depositions in the rAAV-HK-treated ani- mals were also observed. Conclusion: All the effects are comparable with those of intravenous delivery. Therefore, the intramuscular administration of rAAV-HK may be used in gene therapy for hypertension.
Greco, Steven J; Rameshwar, Pranela
Stem cells have been therapeutically utilized in replacement of hematopoetic cells for decades. This is in contrast to the recent emergence of adult stem cells as, perhaps, safe and beneficial therapeutics for multiple diseases and disorders. In particular, mesenchymal stem cells (MSCs) are currently used in multiple human clinical trials. Although MSCs are ubiquitous, bone marrow, umbilical cord and adipose tissue are the sources where MSCs are isolated for research and clinical application. MSCs were thought to be mesodermal due to the initial reports showing their differentiation into specialized mesodermal cells such as chondrocytes. However, it now appears that MSCs might be neuroectodermal in origin. Thus far, there is no evidence of in vivo transformation of MSCs. However, it is too early to prove or disprove that MSCs can be transformed in vivo in clinical trials. MSCs display immunosuppressive properties when placed in a milieu of inflammatory mediators. This phenotype makes MSCs easily available for therapies as 'off-the-shelf cells. Additionally, MSCs express chemotactic receptors, thereby allowing them to migrate to sites of tissue injury. This latter property has proven useful in the embodiment of MSCs as cellular vehicles to deliver targeted therapeutics to precise regions. The MSCs would typically harbor a prodrug or ectopically express a therapeutic gene to be delivered at a targeted site. This approach has been utilized in a number of different indications requiring precise therapeutic delivery, specifically cancer, cardiovascular disorders and neurodegenerative diseases. Combined with their immune-privileged status, safe clinical profile and low tumorigenicity, MSCs offer vast potential to benefit patients with serious diseases, for which limited treatment options exist. PMID:22946432
Thakur, Ashish; Jaiswal, Manish K; Peak, Charles W; Carrow, James K; Gentry, James; Dolatshahi-Pirouz, Alireza; Gaharwar, Akhilesh K
Injectable hydrogels are investigated for cell encapsulation and delivery as they can shield cells from high shear forces. One of the approaches to obtain injectable hydrogels is to reinforce polymeric networks with high aspect ratio nanoparticles such as two-dimensional (2D) nanomaterials. 2D nanomaterials are an emerging class of ultrathin materials with a high degree of anisotropy and they strongly interact with polymers resulting in the formation of shear-thinning hydrogels. Here, we present 2D nanosilicate reinforced kappa-carrageenan (κCA) hydrogels for cellular delivery. κCA is a natural polysaccharide that resembles native glycosaminoglycans and can form brittle hydrogels via ionic crosslinking. The chemical modification of κCA with photocrosslinkable methacrylate groups renders the formation of a covalently crosslinked network (MκCA). Reinforcing the MκCA with 2D nanosilicates results in shear-thinning characteristics, and enhanced mechanical stiffness, elastomeric properties, and physiological stability. The shear-thinning characteristics of nanocomposite hydrogels are investigated for human mesenchymal stem cell (hMSC) delivery. The hMSCs showed high cell viability after injection and encapsulated cells showed a circular morphology. The proposed shear-thinning nanoengineered hydrogels can be used for cell delivery for cartilage tissue regeneration and 3D bioprinting. PMID:27270567
Thakur, Ashish; Jaiswal, Manish K.; Peak, Charles W.; Carrow, James K.; Gentry, James; Dolatshahi-Pirouz, Alireza; Gaharwar, Akhilesh K.
Injectable hydrogels are investigated for cell encapsulation and delivery as they can shield cells from high shear forces. One of the approaches to obtain injectable hydrogels is to reinforce polymeric networks with high aspect ratio nanoparticles such as two-dimensional (2D) nanomaterials. 2D nanomaterials are an emerging class of ultrathin materials with a high degree of anisotropy and they strongly interact with polymers resulting in the formation of shear-thinning hydrogels. Here, we present 2D nanosilicate reinforced kappa-carrageenan (κCA) hydrogels for cellular delivery. κCA is a natural polysaccharide that resembles native glycosaminoglycans and can form brittle hydrogels via ionic crosslinking. The chemical modification of κCA with photocrosslinkable methacrylate groups renders the formation of a covalently crosslinked network (MκCA). Reinforcing the MκCA with 2D nanosilicates results in shear-thinning characteristics, and enhanced mechanical stiffness, elastomeric properties, and physiological stability. The shear-thinning characteristics of nanocomposite hydrogels are investigated for human mesenchymal stem cell (hMSC) delivery. The hMSCs showed high cell viability after injection and encapsulated cells showed a circular morphology. The proposed shear-thinning nanoengineered hydrogels can be used for cell delivery for cartilage tissue regeneration and 3D bioprinting.
Kwon, Kwang-Chul; Daniell, Henry
Plants cells are now approved by the FDA for cost-effective production of protein drugs (PDs) in large-scale current Good Manufacturing Practice (cGMP) hydroponic growth facilities. In lyophilized plant cells, PDs are stable at ambient temperature for several years, maintaining their folding and efficacy. Upon oral delivery, PDs bioencapsulated in plant cells are protected in the stomach from acids and enzymes but are subsequently released into the gut lumen by microbes that digest the plant cell wall. The large mucosal area of the human intestine offers an ideal system for oral drug delivery. When tags (receptor-binding proteins or cell-penetrating peptides) are fused to PDs, they efficiently cross the intestinal epithelium and are delivered to the circulatory or immune system. Unique tags to deliver PDs to human immune or nonimmune cells have been developed recently. After crossing the epithelium, ubiquitous proteases cleave off tags at engineered sites. PDs are also delivered to the brain or retina by crossing the blood-brain or retinal barriers. This review highlights recent advances in PD delivery to treat Alzheimer's disease, diabetes, hypertension, Gaucher's or ocular diseases, as well as the development of affordable drugs by eliminating prohibitively expensive purification, cold chain and sterile delivery. PMID:27378236
Christensen, Jesper F; Bandak, Mikkel; Campbell, Anna;
BACKGROUND: Treatment of testicular germ cell cancer constitutes a major success story in modern oncology. Today, the vast majority of patients are cured by a therapeutic strategy using one or more highly effective components including surgery (orchiectomy), radiotherapy and/or chemotherapy......-induced cardiovascular dysfunction to prevent premature onset of clinical cardiovascular disease in germ cell cancer survivors, with a view towards highlighting future directions of exercise-based survivorship research in the germ cell cancer setting. CONCLUSION: As exercise training may have the potential to ameliorate....... However, the excellent cancer-specific survival comes at considerable costs, as individuals with a history of germ cell cancer experience serious long-term complications, including markedly increased risk of cardiovascular morbidities and premature cardiovascular death. The factors responsible, as well as...
King, Thomas F J
Offspring birthweight is inversely associated with future maternal cardiovascular mortality, a relationship that has yet to be fully elucidated. Endothelial progenitor cells (EPCs) are thought to play a key role in vasculogenesis, and EPC numbers reflect cardiovascular risk.
Hitscherich, Pamela; Wu, Siliang; Gordan, Richard; Xie, Lai-Hua; Arinzeh, Treena; Lee, Eun Jung
Recently, electrospun polyvinylidene fluoride (PVDF) and polyvinylidene fluoride-trifluoroethylene (PVDF-TrFE) scaffolds have been developed for tissue engineering applications. These materials have piezoelectric activity, wherein they can generate electric charge with minute mechanical deformations. Since the myocardium is an electroactive tissue, the unique feature of a piezoelectric scaffold is attractive for cardiovascular tissue engineering applications. In this study, we examined the cytocompatibility and function of pluripotent stem cell derived cardiovascular cells including mouse embryonic stem cell-derived cardiomyocytes (mES-CM) and endothelial cells (mES-EC) on PVDF-TrFE scaffolds. MES-CM and mES-EC adhered well to PVDF-TrFE and became highly aligned along the fibers. When cultured on scaffolds, mES-CM spontaneously contracted, exhibited well-registered sarcomeres and expressed classic cardiac specific markers such as myosin heavy chain, cardiac troponin T, and connexin43. Moreover, mES-CM cultured on PVDF-TrFE scaffolds responded to exogenous electrical pacing and exhibited intracellular calcium handling behavior similar to that of mES-CM cultured in 2D. Similar to cardiomyocytes, mES-EC also demonstrated high viability and maintained a mature phenotype through uptake of low-density lipoprotein and expression of classic endothelial cell markers including platelet endothelial cell adhesion molecule, endothelial nitric oxide synthase, and the arterial specific marker, Notch-1. This study demonstrates the feasibility of PVDF-TrFE scaffold as a candidate material for developing engineered cardiovascular tissues utilizing stem cell-derived cells. Biotechnol. Bioeng. 2016;113: 1577-1585. © 2015 Wiley Periodicals, Inc. PMID:26705272
Full Text Available Stem cells play a special role in the body as agents of self-renewal and auto-reparation for tissues and organs. Stem cell therapies represent a promising alternative strategy to regenerate damaged tissue when natural repairing and conventional pharmacological intervention fail to do so. A fundamental impediment for the evolution of stem cell therapies has been the difficulty of effectively targeting administered stem cells to the disease foci. Biocompatible magnetically responsive nanoparticles are being utilized for the targeted delivery of stem cells in order to enhance their retention in the desired treatment site. This noninvasive treatment-localization strategy has shown promising results and has the potential to mitigate the problem of poor long-term stem cell engraftment in a number of organ systems post-delivery. In addition, these same nanoparticles can be used to track and monitor the cells in vivo, using magnetic resonance imaging. In the present review we underline the principles of magnetic targeting for stem cell delivery, with a look at the logic behind magnetic nanoparticle systems, their manufacturing and design variants, and their applications in various pathological models.
Foged, Camilla; Nielsen, Hanne Moerck
-penetrating peptides as transmembrane drug delivery agents, according to the recent literature, and discusses critical issues and future challenges in relation to fully understanding the fundamental principles of the cell-penetrating peptide-mediated membrane translocation of cargoes and the exploitation of their...
Satya Prakash; Aleksandra Malgorzata Urbanska
Satya Prakash, Aleksandra Malgorzata UrbanskaBiomedical Technology and Cell Therapy Research Laboratory, Departments of Biomedical Engineering and Physiology, Artificial Cells and Organs Research Center, Faculty of Medicine, McGill University, Montreal, Quebec, CanadaAbstract: There has been an ample interest in delivery of therapeutic molecules using live cells. Oral delivery has been stipulated as best way to deliver live cells to humans for therapy. Colon, in particular, is a part of gastr...
Yang, Ningning; Singh, Saurabh; Mahato, Ram I.
Triplex-forming oligonucleotides (TFOs) represent an antigene approach for gene regulation through direct interaction with genomic DNA. While this strategy holds great promise owing to the fact that only two alleles need silencing to impact gene regulation, delivering TFOs to target cells in vivo is still a challenge. Our recent efforts have focused on conjugating TFOs to carrier molecules like cholesterol to enhance their cellular uptake and mannose-6-phosphate-bovine serum albumin (M6P-BSA)...
Eyileten, Ceren; Majchrzak, Kinga; Pilch, Zofia; Tonecka, Katarzyna; Mucha, Joanna; Taciak, Bartlomiej; Ulewicz, Katarzyna; Witt, Katarzyna; Boffi, Alberto; Krol, Magdalena; Rygiel, Tomasz P.
Recent studies indicate the critical role of tumour associated macrophages, tumour associated neutrophils, dendritic cells, T lymphocytes, and natural killer cells in tumourigenesis. These cells can have a significant impact on the tumour microenvironment via their production of cytokines and chemokines. Additionally, products secreted from all these cells have defined specific roles in regulating tumour cell proliferation, angiogenesis, and metastasis. They act in a protumour capacity in vivo as evidenced by the recent studies indicating that macrophages, T cells, and neutrophils may be manipulated to exhibit cytotoxic activity against tumours. Therefore therapy targeting these cells may be promising, or they may constitute drug or anticancer particles delivery systems to the tumours. Herein, we discussed all these possibilities that may be used in cancer treatment. PMID:27212807
Full Text Available Cell therapy is one of the most promising areas within regenerative medicine. However, its full potential is limited by the rapid loss of introduced therapeutic cells before their full effects can be exploited, due in part to anoikis, and in part to the adverse environments often found within the pathologic tissues that the cells have been grafted into. Encapsulation of individual cells has been proposed as a means of increasing cell viability. In this study, we developed a facile, high throughput method for creating temperature responsive microcapsules comprising agarose, gelatin and fibrinogen for delivery and subsequent controlled release of cells. We verified the hypothesis that composite capsules combining agarose and gelatin, which possess different phase transition temperatures from solid to liquid, facilitated the destabilization of the capsules for cell release. Cell encapsulation and controlled release was demonstrated using human fibroblasts as model cells, as well as a therapeutically relevant cell line—human umbilical vein endothelial cells (HUVECs. While such temperature responsive cell microcapsules promise effective, controlled release of potential therapeutic cells at physiological temperatures, further work will be needed to augment the composition of the microcapsules and optimize the numbers of cells per capsule prior to clinical evaluation.
Zimmerlin, Ludovic; Rubin, J. Peter; Pfeifer, Melanie E.; Moore, L.R.; Donnenberg, Vera S.; Donnenberg, Albert D.
Background Adipose tissue represents a practical source of autologous mesenchymal stromal cells (MSC) and vascular-endothelial progenitor cells, available for regenerative therapy without in vitro expansion. One of the problems confronting the therapeutic application of such cells is how to immobilize them at the wound site. Here, we evaluated in vitro the growth and differentiation of human adipose stromal vascular fraction (SVF) cells after delivery using a fibrin spray system. Methods SVF cells were harvested from four human adult patients undergoing elective abdominoplasty using the LipiVage™ system. After collagenase digestion, mesenchymal and endothelial progenitor cells (pericytes, supra-adventitial stromal cells, endothelial progenitors) were quantified by flow cytometry before culture. SVF cells were applied to culture vessels using the Tisseel™ fibrin spray system. SVF cell growth and differentiation was documented by immunofluorescence staining and photomicrography. Results SVF cells remained viable following application and were expanded up to three weeks, when they reached confluence and adipogenic differentiation. Under angiogenic conditions, SVF cells formed endothelial (vWF+, CD31+ and CD34+) tubules surrounded by CD146+ and α-SMA+ perivascular/stromal cells. Discussion Human adipose tissue is a rich source of autologous stem cells, which are readily available for regenerative applications such as wound healing, without in vitro expansion. Our results indicate that mesenchymal and endothelial progenitor cells, prepared in a closed system from unpassaged lipoaspirate samples, retain their growth and differentiation capacity when applied and immobilized on a substrate using a clinically approved fibrin sealant spray system. PMID:23260090
Full Text Available Chuanwei Wang,1,2,* Liping Ning,3,* Hongwei Wang,1,2,* Zaijun Lu,4 Xingang Li,1,2 Xiaoyong Fan,5 Xuping Wang,6 Yuguang Liu1,2 1Department of Neurosurgery, Qilu Hospital of Shandong University, Jinan, People's Republic of China; 2Brain Science Research Institute of Shandong University, Jinan, People's Republic of China; 3Department of Rehabilitation, Provincial Hospital Affiliated to Shandong University, Jinan, People's Republic of China; 4School of Chemistry and Chemical Engineering of Shandong University, Jinan, People's Republic of China; 5Department of Neurosurgery, Shandong Qianfoshan Hospital Affiliated to Shandong University, Jinan, People's Republic of China; 6Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and Chinese Ministry of Public Health, Qilu Hospital of Shandong University, Jinan, People's Republic of China *These authors contributed equally to this work Abstract: Glioblastoma multiforme (GBM is the most common and malignant glioma. Although there has been considerable progress in treatment strategies, the prognosis of many patients with GBM remains poor. In this work, polyethylenimine (PEI and the VTWTPQAWFQWV (VTW peptide were modified and synthesized into GBM-targeting nanoparticles. The transfection efficiency of U-87 (human glioblastoma cells was evaluated using fluorescence microscopy and flow cytometry. Cell internalization was investigated to verify the nanoparticle delivery into the cytoplasm. Results showed that the methods of polymer conjugation and the amount of VTW peptide were important factors to polymer synthesis and transfection. The PEI-VTW20 nanoparticles increased the transfection efficiency significantly. This report describes the use of VTW peptide-based PEI nanoparticles for intracellular gene delivery in a GBM cell-specific manner. Keywords: glioblastoma, polyethylenimine, nanoparticles, drug-delivery systems, gene transfer techniques
Full Text Available Xun Sun, Simu Chen, Jianfeng Han, Zhirong ZhangKey Laboratory of Drug Targeting and Drug Delivery System, Ministry of Education, West China School of Pharmacy, Sichuan University, Chengdu, People’s Republic of ChinaBackground: To establish a potential gene-delivery system with the ability to deliver plasmid DNA to dendritic cells (DCs more efficiently and specifically, we designed and synthesized a low-molecular-weight polyethyleneimine and triethyleneglycol polymer (PEI–TEG and a series of its mannosylated derivatives.Methods: PEI–TEG was synthesized from PEI2000 and PEI600 with TEG as the cross-linker. PEI–TEG was then linked to mannose via a phenylisothiocyanate bridge to obtain man-PEI–TEG conjugates. The DNA conveyance abilities of PEI–TEG, man-PEI–TEG, as well as control PEI25k were evaluated by measuring their zeta potential, particle size, and DNA-binding abilities. The in vitro cytotoxicity, cell uptake, and transfection efficiency of these PEI/DNA complexes were examined on the DC2.4 cell line. Finally, a maturation experiment evaluated the effect of costimulatory molecules CD40, CD80, and CD86 on murine bone marrow-derived DCs (BMDCs using flow cytometry.Results: PEI–TEG and man-PEI–TEG were successfully synthesized and were shown to retain the excellent properties of PEI25k for condensing DNA. Compared with PEI–TEG as well as PEI25k, the man-PEI–TEG had less cytotoxicity and performed better in both cellular uptake and transfection assays in vitro. The results of the maturation experiment showed that all the PEI/DNA complexes induced an adequate upregulation of surface markers for DC maturation.Conclusion: These results demonstrated that man-PEI–TEG can be employed as a DC-targeting gene-delivery system.Keywords: dendritic cells, DCs, mannose, polyethyleneimine, PEI, gene delivery
Carpenter, Cody Westcott
Herein we describe the development of the Red Blood Cell coated nanoparticle, RBC-NP. Purified natural erythrocyte membrane is used to coat drug-loaded poly(lacticco-glycolic acid) (PLGA). Synthetic PLGA co-polymer is biocompatible and biodegradable and has already received US FDA approval for drug-delivery and diagnostics. This work looks specifically at the retention of immunosuppressive proteins on RBC-NPs, right-sidedness of natural RBC membranes interfacing with synthetic polymer nanoparticles, sustained and retarded drug release of RBC-NPs as well as further surface modification of RBC-NPs for increased targeting of model cancer cell lines.
Anselmo, Aaron C.; Mitragotri, Samir
Cellular hitchhiking leverages the use of circulatory cells to enhance the biological outcome of nanoparticle drug delivery systems, which often suffer from poor circulation time and limited targeting. Cellular hitchhiking utilizes the natural abilities of circulatory cells to: (i) navigate the vasculature while avoiding immune system clearance, (ii) remain relatively inert until needed and (iii) perform specific functions, including nutrient delivery to tissues, clearance of pathogens, and i...
Full Text Available Functional nanomaterials have recently attracted strong interest from the biology community, not only as potential drug delivery vehicles or diagnostic tools, but also as optical nanomaterials. This is illustrated by the explosion of publications in the field with more than 2,000 publications in the last 2 years (4,000 papers since 2000; from ISI Web of Knowledge, ‘nanoparticle and cell’ hit. Such a publication boom in this novel interdisciplinary field has resulted in papers of unequal standard, partly because it is challenging to assemble the required expertise in chemistry, physics, and biology in a single team. As an extreme example, several papers published in physical chemistry journals claim intracellular delivery of nanoparticles, but show pictures of cells that are, to the expert biologist, evidently dead (and therefore permeable. To attain proper cellular applications using nanomaterials, it is critical not only to achieve efficient delivery in healthy cells, but also to control the intracellular availability and the fate of the nanomaterial. This is still an open challenge that will only be met by innovative delivery methods combined with rigorous and quantitative characterization of the uptake and the fate of the nanoparticles. This review mainly focuses on gold nanoparticles and discusses the various approaches to nanoparticle delivery, including surface chemical modifications and several methods used to facilitate cellular uptake and endosomal escape. We will also review the main detection methods and how their optimum use can inform about intracellular localization, efficiency of delivery, and integrity of the surface capping. Raphaël Lévy is a BBSRC David Phillips Research Fellow at the University of Liverpool. He graduated in Physics at the University Louis Pasteur in Strasbourg (France. In 2002, after a Master in Soft Condensed Matter Physics, he obtained a PhD in Physics at the University Louis Pasteur. He then moved to
Full Text Available Abstract Background Chromosomal complement, including that provided by the sex chromosomes, influences expression of proteins and molecular signaling in every cell. However, less than 50% of the scientific studies published in 2009 using experimental animals reported sex as a biological variable. Because every cell has a sex, we conducted a literature review to determine the extent to which sex is reported as a variable in cardiovascular studies on cultured cells. Methods Articles from 10 cardiovascular journals with high impact factors (Circulation, J Am Coll Cardiol, Eur Heart J, Circ Res, Arterioscler Thromb Vasc Biol, Cardiovasc Res, J Mol Cell Cardiol, Am J Physiol Heart Circ Physiol, J Heart Lung Transplant and J Cardiovasc Pharmacol and published in 2010 were searched using terms 'cultured' and 'cells' in any order to determine if the sex of those cells was reported. Studies using established cell lines were excluded. Results Using two separate search strategies, we found that only 25 of 90 articles (28% and 20 of 101 articles (19.8% reported the sex of cells. Of those reporting the sex of cells, most (68.9%; n = 31 used only male cells and none used exclusively female cells. In studies reporting the sex of cells of cardiovascular origin, 40% used vascular smooth-muscle cells, and 30% used stem/progenitor cells. In studies using cells of human origin, 35% did not report the sex of those cells. None of the studies using neonatal cardiac myocytes reported the sex of those cells. Conclusions The complement of sex chromosomes in cells studied in culture has the potential to affect expression of proteins and 'mechanistic' signaling pathways. Therefore, consistent with scientific excellence, editorial policies should require reporting sex of cells used in in vitro experiments.
Juan Manuel Senior Sánchez
Full Text Available Although there have been important advances in the field of molecular biology, the mechanisms responsible for nephrogenesis and the factors that modulate the process of development, proliferation, growth, and maturation during fetal and adult life have not been thoroughly explained. Animals, including mammals, share the intrinsic ability to regenerate tissues and organs as an important biological defense mechanism. In the case of the kidney, after tissue damage secondary to injury, anatomical and functional recovery of integrity is achieved, accompanied by the activation of a complex, poorly understood process, leading to the replacement of damaged tubular cells by functional ones that reorganize tubular architecture. This regeneration and repair process is produced by somatic, exogenous, adult stem cells, and probably by intrinsic renal stem cells, that are responsible for maintaining renal homeostasis Aunque se han logrado grandes avances en el campo de la biología molecular, todavía no se han esclarecido completamente los mecanismos responsables de la organogénesis y los factores que modulan el proceso de desarrollo, proliferación, crecimiento y maduración celulares durante la vida fetal y adulta. Los animales comparten la capacidad de regenerar tejidos y órganos, como un mecanismo biológico importante de defensa. En el caso del riñón, luego del daño tisular secundario a una noxa, se produce recuperación anatómica y funcional de la integridad, acompañada por la activación de un proceso sofisticado, mal comprendido, que lleva al reemplazo de las células tubulares dañadas por otras funcionalmente normales que reorganizan la arquitectura tubular. Este fenómeno de recambio se produce gracias a la presencia de células madre adultas somáticas exógenas, responsables del proceso de mantenimiento de la homeostasis renal, y posiblemente por células renales intrínsecas.
Mathiasen, Anders Bruun; Haack-Sørensen, Mandana; Kastrup, Jens
results of most studies are promising, but there are still many unanswered questions. In this review, we explore present preclinical and clinical knowledge regarding the use of stem cells in cardiovascular regenerative medicine, with special focus on mesenchymal stromal cells. We take a closer look at...... for regenerative therapy. Clinical studies on stem cell therapy for cardiac regeneration have shown significant improvements in ventricular pump function, ventricular remodeling, myocardial perfusion, exercise potential and clinical symptoms compared with conventionally treated control groups. The...
Wu, Junru; Pepe, Jason; Rincon, Mercedes
It has been shown experimentally in cell suspensions that pulsed ultrasound (2.0 MHz) could be used to deliver an anti-cancer drug (Adriamycin hydrochloride) into Jurkat lymphocytes and antibodies (goat anti rabbit IgG and anti mouse IgD) into human peripheral blood mononuclear (PBMC) cells and Jurkat lymphocytes assisted by encapsulated microbubbles (Optison). When Adriamycin hydrochloride (ADR) was delivered, the delivery efficiency reached 4.80% and control baseline (no ultrasound and no ADR) was 0.17%. When anti-rabbit IgD was delivered, the efficiencies were 34.90% (control baseline was 1.33%) and 32.50% (control baseline was 1.66%) respectively for Jurkat cells and PBMC. When goat anti rabbit IgG was delivered, the efficiencies were 78.60% (control baseline was 1.60%) and 57.50% (control baseline was 11.30%) respectively for Jurkat cells and PBMC.
Hoop, M.; Mushtaq, F.; Hurter, C.; Chen, X.-Z.; Nelson, B. J.; Pané, S.
most tumors. Approximately a 2.5 times higher drug release from Ni nanotubes at pH = 6 is achieved compared to that at pH = 7.4. The outside of the Ni tube is coated with gold. A fluorescein isothiocyanate (FITC) labeled thiol-ssDNA, a biological marker, was conjugated on its surface by thiol-gold click chemistry, which enables traceability. The Ni nanotube allows the propulsion of the device by means of external magnetic fields. As the proposed nanoarchitecture integrates different functional building blocks, our drug delivery nanoplatform can be employed for carrying molecular drug conjugates and for performing targeted combinatorial therapies, which can provide an alternative and supplementary solution to current drug delivery technologies. Electronic supplementary information (ESI) available: Fig. S1 drug release control experiment; Fig. S2 cell viability assay; video - magnetic manipulation. See DOI: 10.1039/c6nr02228f
Wallrapp, Christine; Thoenes, Eric; Thürmer, Frank;
Glucagon-like peptide-1 (GLP-1) CellBeads are cell-based implants for the sustained local delivery of bioactive factors. They consist of GLP-1 secreting mesenchymal stem cells encapsulated in a spherically shaped immuno-isolating alginate matrix. A highly standardized and reproducible encapsulati...... preclinical studies and an ongoing safety trial in humans but further studies have to prove the overall potential of CellBead technology in cell-based regenerative medicine....
Full Text Available Background: Autologous Bone Marrow stem Cell transplantation is a viable therapeutic option for patients with end stage heart failure due to cardiomyopathy of varied etiology as there are only limited treatment options other than cardiac transplantation. The rationale behind the application of stem cells in these patients include • Stem cells directly replace the affected cells by differentiation into the damaged cell type • Stem cells also exert Paracrine effects by secre tion of growth factors (VGEF,FGF-1to stimu late local cell growth•In addition to the above, stem cells release signaling factors which recruit stem cells from elsewhere by modulating the immune system.Materials and Methods: In this presentation we describe our study on a series of 13 patients who received isolated and expanded CD 34 cells from the bone marrow. Seven had ischemic dysfunction, three had dilated cardiomyopathy and three had primary pulmonary hypertension. Five patients received the stem cells via intracoronary injection, three directly into the myocardium and three intrapulmonary. Results: All patients showed functional improvement of the myocardium recorded by non-invasive investigations and improvement in the quality of life. Follow up period ranged from 6 months to 2 years. Conclusion: Our experience with bone marrow derived stem cells in patients with cardiomyopathy has been encouraging. More studies are planned in the future.
Wang, Junxin; Jokerst, Jesse V.
Stem cell therapy (SCT) has shown very promising preclinical results in a variety of regenerative medicine applications. Nevertheless, the complete utility of this technology remains unrealized. Imaging is a potent tool used in multiple stages of SCT and this review describes the role that imaging plays in cell harvest, cell purification, and cell implantation, as well as a discussion of how imaging can be used to assess outcome in SCT. We close with some perspective on potential growth in the field. PMID:26880997
Haaber, A B; Eidemak, I; Jensen, T;
Cardiovascular risk factors and markers of endothelial cell function were studied in nondiabetic patients with mild to moderate chronic renal failure. The transcapillary escape rate of albumin and the plasma concentrations of von Willebrand factor, fibrinogen, and plasma lipids were measured in 29...
Gopinath, Subash C B; Lakshmipriya, Thangavel; Chen, Yeng; Arshad, M K Md; Kerishnan, Jesinda P; Ruslinda, A R; Al-Douri, Yarub; Voon, C H; Hashim, Uda
Aptamers are single-stranded nucleic acids or peptides identified from a randomized combinatorial library through specific interaction with the target of interest. Targets can be of any size, from small molecules to whole cells, attesting to the versatility of aptamers for binding a wide range of targets. Aptamers show drug properties that are analogous to antibodies, with high specificity and affinity to their target molecules. Aptamers can penetrate disease-causing microbial and mammalian cells. Generated aptamers that target surface biomarkers act as cell-targeting agents and intracellular delivery vehicles. Within this context, the "cell-internalizing aptamers" are widely investigated via the process of cell uptake with selective binding during in vivo systematic evolution of ligands by exponential enrichment (SELEX) or by cell-internalization SELEX, which targets cell surface antigens to be receptors. These internalizing aptamers are highly preferable for the localization and functional analyses of multiple targets. In this overview, we discuss the ways by which internalizing aptamers are generated and their successful applications. Furthermore, theranostic approaches featuring cell-internalized aptamers are discussed with the purpose of analyzing and diagnosing disease-causing pathogens. PMID:27350620
Bundy, David G; Muschelli, John; Clemens, Gwendolyn D; Strouse, John J; Thompson, Richard E; Casella, James F; Miller, Marlene R
Preventive services can reduce the morbidity of sickle cell disease (SCD) in children but are delivered unreliably. We conducted a retrospective cohort study of children aged 2 to 5 years with SCD, evaluating each child for 14 months and expecting that he/she should receive ≥75% of days covered by antibiotic prophylaxis, ≥1 influenza immunization, and ≥1 transcranial Doppler ultrasound (TCD). We used logistic regression to quantify the relationship between ambulatory generalist and hematologist visits and preventive services delivery. Of 266 children meeting the inclusion criteria, 30% consistently filled prophylactic antibiotic prescriptions. Having ≥2 generalist, non-well child care visits or ≥2 hematologist visits was associated with more reliable antibiotic prophylaxis. Forty-one percent of children received ≥1 influenza immunizations. Children with ≥2 hematologist visits were most likely to be immunized (62% vs. 35% among children without a hematologist visit). Only 25% of children received ≥1 TCD. Children most likely to receive a TCD (42%) were those with ≥2 hematologist visits. One in 20 children received all 3 preventive services. Preventive services delivery to young children with SCD was inconsistent but associated with multiple visits to ambulatory providers. Better connecting children with SCD to hematologists and strengthening preventive care delivery by generalists are both essential. PMID:26950087
Full Text Available Different mechanisms in cancer cells become resistant to one or more chemotherapeutics is known as multidrug resistance(MDR which hinders chemotherapy efficacy. Potential factors for MDR includes enhanced drug detoxification, decreased drug uptake, increased intracellular nucleophiles levels, enhanced repair of drug induced DNA damage, overexpression of drug transporter such as P-glycoprotein(P-gp, multidrug resistance-associated proteins(MRP1, MRP2 and breast cancer resistance protein(BCRP. Currently nanoassemblies such as polymeric/solid lipid/inorganic/metal nanoparticles, quantum dots, dendrimers, liposomes, micelles has emerged as an innovative, effective and promising platforms for treatment of drug resistant cancer cells. Nanocarriers have potential to improve drug therapeutic index, ability for multifunctionality, divert ABC-transporter mediated drug efflux mechanism and selective targeting to tumor cells, cancer stem cells, tumor initiating cells or cancer microenvironment. Selective nanocarrier targeting to tumor overcomes dose-limiting side effects, lack of selectivity, tissue toxicity, limited drug access to tumor tissues, high drug doses and emergence of multiple drug resistance with conventional or combination chemotherapy. Current review highlights various nanodrug delivery systems to overcome mechanism of MDR by neutralizing, evading or exploiting the drug efflux pumps and those independent of drug efflux pump mechanism by silencing Bcl-2 and HIF1 gene expressions by siRNA and miRNA, modulating ceramide levels and targeting NF-B. Theragnostics combining a cytotoxic agent, targeting moiety, chemosensitizing agent and diagnostic imaging aid are highlighted as effective and innovative systems for tumor localization and overcoming MDR. Physical approaches such as combination of drug with thermal/ultrasound/photodynamic therapies to overcome MDR are focused. The review focuses on newer drug delivery systems developed to overcome
Giri, Tapan Kumar; Thakur, Deepa; Alexander, Amit; Ajazuddin; Badwaik, Hemant; Tripathi, Dulal Krishna
Alginate is a non-toxic, biocompatible and biodegradable natural polymer with a number of peculiar physicochemical properties for which it has wide applications in drug delivery and cell delivery systems. Hydrogel formation can be obtained by interactions of anionic alginates with multivalent inorganic cations by simple ionotropic gelation method. Hydrophilic polymeric network of three dimensional cross linked structures of hydrogels absorb substantial amount of water or biological fluids. Among the numerous biomaterials used for hydrogel formation alginate has been and will continue to be one of the most important biomaterial. Therefore, in view of the vast literature support, we focus in this review on alginate - based hydrogel as drug delivery and cell delivery carriers for biomedical applications. Various properties of alginates, their hydrogels and also various techniques used for preparing alginate hydrogels have been reviewed. PMID:22998675
Cui, Yuqi; Sun, Qinghua; Liu, Zhenguo
Air pollution is a major challenge to public health. Ambient fine particulate matter (PM) is the key component for air pollution, and associated with significant mortality. The majority of the mortality following PM exposure is related to cardiovascular diseases. However, the mechanisms for the adverse effects of PM exposure on cardiovascular system remain largely unknown and under active investigation. Endothelial dysfunction or injury is considered one of the major factors that contribute to the development of cardiovascular diseases such as atherosclerosis and coronary heart disease. Endothelial progenitor cells (EPCs) play a critical role in maintaining the structural and functional integrity of vasculature. Particulate matter exposure significantly suppressed the number and function of EPCs in animals and humans. However, the mechanisms for the detrimental effects of PM on EPCs remain to be fully defined. One of the important mechanisms might be related to increased level of reactive oxygen species (ROS) and inflammation. Bone marrow (BM) is a major source of EPCs. Thus, the number and function of EPCs could be intimately associated with the population and functional status of stem cells (SCs) in the BM. Bone marrow stem cells and other SCs have the potential for cardiovascular regeneration and repair. The present review is focused on summarizing the detrimental effects of PM exposure on EPCs and SCs, and potential mechanisms including ROS formation as well as clinical implications. PMID:26988063
Currently, the pre-clinical field is rapidly progressing in search of new therapeutic modalities that replace or complement current medication to treat cardiovascular disease. Among these are the single or combined use of stem cells, biomaterials and instructive factors, which together form the triad of tissue engineering and regenerative medicine. Stem cell therapy is a promising approach for repair, remodeling and even regenerate tissue of otherwise irreparable damage, such as after myocard...
Mumcuoglu, Didem; Sardan, Melis; Tekinay, Turgay; Guler, Mustafa O; Tekinay, Ayse B
A drug delivery system designed specifically for oligonucleotide therapeutics can ameliorate the problems associated with the in vivo delivery of these molecules. The internalization of free oligonucleotides is challenging, and cytotoxicity is the main obstacle for current transfection vehicles. To develop nontoxic delivery vehicles for efficient transfection of oligonucleotides, we designed a self-assembling peptide amphiphile (PA) nanosphere delivery system decorated with cell penetrating peptides (CPPs) containing multiple arginine residues (R4 and R8), and a cell surface binding peptide (KRSR), and report the efficiency of this system in delivering G-3129, a Bcl-2 antisense oligonucleotide (AON). PA/AON (peptide amphiphile/antisense oligonucleotide) complexes were characterized with regards to their size and secondary structure, and their cellular internalization efficiencies were evaluated. The effect of the number of arginine residues on the cellular internalization was investigated by both flow cytometry and confocal imaging, and the results revealed that uptake efficiency improved as the number of arginines in the sequence increased. The combined effect of cell penetration and surface binding property on the cellular internalization and its uptake mechanism was also evaluated by mixing R8-PA and KRSR-PA. R8 and R8/KRSR decorated PAs were found to drastically increase the internalization of AONs compared to nonbioactive PA control. Overall, the KRSR-decorated self-assembled PA nanospheres were demonstrated to be noncytotoxic delivery vectors with high transfection rates and may serve as a promising delivery system for AONs. PMID:25828697
Kennedy, Laura Carpin
This thesis reports new gold nanoparticle-based methods to treat chemotherapy-resistant and metastatic tumors that frequently evade conventional cancer therapies. Gold nanoparticles represent an innovative generation of diagnostic and treatment agents due to the ease with which they can be tuned to scatter or absorb a chosen wavelength of light. One area of intensive investigation in recent years is gold nanoparticle photothermal therapy (PTT), in which gold nanoparticles are used to heat and destroy cancer. This work demonstrates the utility of gold nanoparticle PTT against two categories of cancer that are currently a clinical challenge: trastuzumab-resistant breast cancer and metastatic cancer. In addition, this thesis presents a new method of gold nanoparticle delivery using T cells that increases gold nanoparticle tumor accumulation efficiency, a current challenge in the field of PTT. I ablated trastuzumab-resistant breast cancer in vitro for the first time using anti-HER2 labeled silica-gold nanoshells, demonstrating the potential utility of PTT against chemotherapy-resistant cancers. I next established for the first time the use of T cells as gold nanoparticle vehicles in vivo. When incubated with gold nanoparticles in culture, T cells can internalize up to 15000 nanoparticles per cell with no detrimental effects to T cell viability or function (e.g. migration and cytokine secretion). These AuNP-T cells can be systemically administered to tumor-bearing mice and deliver gold nanoparticles four times more efficiently than by injecting free nanoparticles. In addition, the biodistribution of AuNP-T cells correlates with the normal biodistribution of T cell carrier, suggesting the gold nanoparticle biodistribution can be modulated through the choice of nanoparticle vehicle. Finally, I apply gold nanoparticle PTT as an adjuvant treatment for T cell adoptive transfer immunotherapy (Hyperthermia-Enhanced Immunotherapy or HIT) of distant tumors in a melanoma mouse
Wojakowski, W; Landmesser, U.; Bachowski, R; Jadczyk, T; M. Tendera
Circulating bone marrow (BM)-derived stem and progenitor cells (SPCs) participate in turnover of vascular endothelium and myocardial repair after acute coronary syndromes. Acute myocardial infarction (MI) produces a generalized inflammatory reaction, including mobilization of SPCs, increased local production of chemoattractants in the ischemic myocardium, as well as neural and humoral signals activating the SPC egress from the BM. Several types of circulating BM cells were identified in the p...
Llovera Nadal, Laia; Berthold, Peter; Nielsen, Peter E;
have now quantitatively compared the cellular activity (in the pLuc705 HeLa cell splice correction system) of PNA antisense oligomers using lipoplex delivery of cholesterol- and bisphosphonate-PNA conjugates, polyplex delivery via a PNA-polyethyleneimine conjugate and CPP delivery via a PNA...
Zhou, Fang; Jia, Xiaoling; Yang, Qingmao; Yang, Yang; Zhao, Yunhui; Fan, Yubo; Yuan, Xiaoyan
Manipulation of gene expression by means of microRNAs (miRNAs) is one of the emerging strategies to treat cardiovascular and cancer diseases. Nevertheless, efficient delivery of miRNAs to a specific vascular tissue is limited. In this work, a short peptide Arg-Glu-Asp-Val (REDV) was linked to trimethyl chitosan (TMC) via a bifunctional poly(ethylene glycol) (PEG) linker for the targeted delivery of microRNA-126 (miRNA-126) to vascular endothelial cells (VECs). The morphology, serum stability and cytotoxicity of the polyplex/miRNA complexes, namely, TMC/miRNA, TMC-g-PEG/miRNA and TMC-g-PEG-REDV/miRNA, were investigated along with the cellular uptake, proliferation and in vitro miRNA transfection efficiency. By REDV modification, the TMC-g-PEG-REDV/miRNA complex showed negligible cytotoxicity, increased expression of miRNA-126 and enhanced VEC proliferation compared with the TMC/miRNA and TMC-g-PEG/miRNA complexes. In particular, the approaches adopted for the miRNA delivery and targeted peptide REDV modification promote the selective uptake and the growth of VECs over vascular smooth muscle cells. It was suggested that the REDV peptide-modified TMC-g-PEG polyplex could be potentially used as a miRNA carrier in artificial blood vessels for rapid endothelialization. PMID:27055482
Daniel R. Bayzigitov
Full Text Available Fundamental studies of molecular and cellular mechanisms of cardiovascular disease pathogenesis are required to create more effective and safer methods of their therapy. The studies can be carried out only when model systems that fully recapitulate pathological phenotype seen in patients are used. Application of laboratory animals for cardiovascular disease modeling is limited because of physiological differences with humans. Since discovery of induced pluripotency generating induced pluripotent stem cells has become a breakthrough technology in human disease modeling. In this review, we discuss a progress that has been made in modeling inherited arrhythmias and cardiomyopathies, studying molecular mechanisms of the diseases, and searching for and testing drug compounds using patient-specific induced pluripotent stem cell-derived cardiomyocytes.
Full Text Available Cell separation and delivery have recently gained significant attention in biological and biochemical studies. In thiswork, an optical method for separation and controllable delivery of cells by using an abruptly tapered fiber probe is reported. By launching a laser beam at the wavelength of 980 nm into the fiber, a mixture of cells with sizes of ~5 and ~3 μm and poly(methyl methacrylate particles with size of 5 μm are separated into three chains along the direction of propagation of light. The cell and particle chains are delivered in three dimensions over 600 μm distance. Experimental results are interpreted by numerical simulations. Optical forces and forward migration velocities of different particles and cells are calculated and discussed.
McCurley, Amy; McGraw, Adam; Pruthi, Dafina; Jaffe, Iris Z.
The mineralocorticoid receptor (MR), a member of the steroid receptor family, regulates blood pressure by mediating the effects of the hormone aldosterone on renal sodium handling. In recent years, it has become clear that MR is expressed in vascular smooth muscle cells (SMC) and interest has grown in understanding the direct role of SMC MR in regulating vascular function. This interest stems from multiple clinical studies where MR inhibitor treatment reduced the incidence of cardiovascular e...
Nina A Mikirova, Joseph J Casciari, Ronald E Hunninghake, Margaret M Beezley
Full Text Available Being overweight or obese is associated with an increased risk for the development of non-insulin-dependent diabetes mellitus, hypertension, and cardiovascular disease. Dyslipidemia of obesity is characterized by elevated fasting triglycerides and decreased high-density lipoprotein-cholesterol concentrations. Endothelial damage and dysfunction is considered to be a major underlying mechanism for the elevated cardiovascular risk associated with increased adiposity. Alterations in endothelial cells and stem/endothelial progenitor cell function associated with overweight and obesity predispose to atherosclerosis and thrombosis.In our study, we analyzed the effect of a low calorie diet in combination with oral supplementation by vitamins, minerals, probiotics and human chorionic gonadotropin (hCG, 125-180 IUs on the body composition, lipid profile and CD34-positive cells in circulation.During this dieting program, the following parameters were assessed weekly for all participants: fat free mass, body fat, BMI, extracellular/intracellular water, total body water and basal metabolic rate. For part of participants blood chemistry parameters and circulating CD34-positive cells were determined before and after dieting.The data indicated that the treatments not only reduced body fat mass and total mass but also improved the lipid profile. The changes in body composition correlated with the level of lipoproteins responsible for the increased cardiovascular risk factors. These changes in body composition and lipid profile parameters coincided with the improvement of circulatory progenitor cell numbers.As the result of our study, we concluded that the improvement of body composition affects the number of stem/progenitor cells in circulation.
Xiong, Ranhua; Joris, Freya; De Cock, Ine; Demeester, Jo; De Smedt, Stefaan C.; Skirtach, Andre G.; Braeckmans, Kevin
There is considerable interest in using Quantum Dots (QDs) as fluorescent probes such for cellular imaging due to unique advantages in comparison with conventional molecular dyes. However, cytosolic delivery of QDs into live cells remains a major challenge. Here we demonstrate highly efficient delivery of PEG-coated QDs into live cells by means of laser-induced vapour nanobubbles. Using this procedure we succeeded in high-throughput loading of ~80% of cells while maintaining a cell viability of ~85%.
Full Text Available Over the years, cell therapy has become an exciting opportunity to treat human diseases. Early enthusiasm using adult stem cell sources has been tempered in light of preliminary benefits in patients. Considerable efforts have been dedicated, therefore, to explore alternative cells such as those extracted from umbilical cord blood (UCB. In line, UCB banking has become a popular possibility to preserve potentially life-saving cells that are usually discarded after birth, and the number of UCB banks has grown worldwide. Thus, a brief overview on the categories of UCB banks as well as the properties, challenges, and impact of UCB-derived mesenchymal stem cells (MSCs on the area of cardiovascular research is presented. Taken together, the experience recounted here shows that UCBMSCs are envisioned as attractive therapeutic candidates against human disorders arising and/or progressing with vascular deficit.
Full Text Available Adult human cardiac mesenchymal-like stromal cells (CStC represent a relatively accessible cell type useful for therapy. In this light, their conversion into cardiovascular precursors represents a potential successful strategy for cardiac repair. The aim of the present work was to reprogram CStC into functionally competent cardiovascular precursors using epigenetically active small molecules. CStC were exposed to low serum (5% FBS in the presence of 5 µM all-trans Retinoic Acid (ATRA, 5 µM Phenyl Butyrate (PB, and 200 µM diethylenetriamine/nitric oxide (DETA/NO, to create a novel epigenetically active cocktail (EpiC. Upon treatment the expression of markers typical of cardiac resident stem cells such as c-Kit and MDR-1 were up-regulated, together with the expression of a number of cardiovascular-associated genes including KDR, GATA6, Nkx2.5, GATA4, HCN4, NaV1.5, and α-MHC. In addition, profiling analysis revealed that a significant number of microRNA involved in cardiomyocyte biology and cell differentiation/proliferation, including miR 133a, 210 and 34a, were up-regulated. Remarkably, almost 45% of EpiC-treated cells exhibited a TTX-sensitive sodium current and, to a lower extent in a few cells, also the pacemaker I(f current. Mechanistically, the exposure to EpiC treatment introduced global histone modifications, characterized by increased levels of H3K4Me3 and H4K16Ac, as well as reduced H4K20Me3 and H3s10P, a pattern compatible with reduced proliferation and chromatin relaxation. Consistently, ChIP experiments performed with H3K4me3 or H3s10P histone modifications revealed the presence of a specific EpiC-dependent pattern in c-Kit, MDR-1, and Nkx2.5 promoter regions, possibly contributing to their modified expression. Taken together, these data indicate that CStC may be epigenetically reprogrammed to acquire molecular and biological properties associated with competent cardiovascular precursors.
Full Text Available Shantanu Balkundi1, Ari S Nowacek1, Ram S Veerubhotla1, Han Chen2, Andrea Martinez-Skinner1, Upal Roy1, R Lee Mosley1,3, Georgette Kanmogne1, Xinming Liu1,3,4, Alexander V Kabanov3,4, Tatiana Bronich3,4, JoEllyn McMillan1, Howard E Gendelman1,31Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, USA; 2Center for Biotechnology, University of Nebraska-Lincoln, Lincoln, NE, USA; 3Center for Drug Delivery and Nanomedicine, University of Nebraska Medical Center, Omaha, NE, USA; 4Department of Pharmaceutical Sciences, College of Pharmacy, University of Nebraska Medical Center, Omaha, NE, USAAbstract: Nanoformulations of crystalline indinavir, ritonavir, atazanavir, and efavirenz were manufactured by wet milling, homogenization or sonication with a variety of excipients. The chemical, biological, immune, virological, and toxicological properties of these formulations were compared using an established monocyte-derived macrophage scoring indicator system. Measurements of drug uptake, retention, release, and antiretroviral activity demonstrated differences amongst preparation methods. Interestingly, for drug cell targeting and antiretroviral responses the most significant difference among the particles was the drug itself. We posit that the choice of drug and formulation composition may ultimately affect clinical utility.Keywords: human immunodeficiency virus type one, nanotoxicology, monocyte-derived macrophage, nanoformulated antiretroviral therapy, manufacturing techniques
Ramasatyaveni Geesala; Nimai Bar; Dhoke, Neha R.; Pratyay Basak; Amitava Das
Low bioavailability and/or survival at the injury site of transplanted stem cells necessitate its delivery using a biocompatible, biodegradable cell delivery vehicle. In this dataset, we report the application of a porous biocompatible, biodegradable polymer network that successfully delivers bone marrow stem cells (BMSCs) at the wound site of a murine excisional splint wound model. In this data article, we are providing the additional data of the reference article “Porous polymer scaffold fo...
Mathiasen, Anders Bruun; Kastrup, Jens
Stem cell therapy for degenerative diseases, including ischemic heart disease is now a clinical reality. In the search for the optimal cell type for each patient category, many different stem cell subpopulations have been used. In addition, different cell processing procedures and delivery methods...... improvements. To better understand the underlying mechanisms of these results, a reverse translation from bedside to bench has been opened. Non-invasive cell tracking after implantation has a pivotal role in this translation. Imaging based methods can help elucidate important issues such as retention......, migration and efficacy of the transplanted cells. Great effort is being made in finding new and better imaging techniques for different imaging modalities, and much have already been learned. But there are still many unanswered questions. In this review, we give an overview of the imaging modalities used...
Lin, Ming-Yu; Wu, Yi-Chien; Lee, Ji-Ann; Tung, Kuan-Wen; Zhou, Jessica; Teitell, Michael A; Yeh, J Andrew; Chiou, Pei Yu
Introducing functional macromolecules into a variety of living cells is challenging but important for biology research and cell-based therapies. We report a novel cell delivery platform based on rotating shape anisotropic magnetic particles (SAMPs), which make very small cuts on cell membranes for macromolecule delivery with high efficiency and high survivability. SAMP delivery is performed by placing commercially available nickel powder onto cells grown in standard cell culture dishes. Application of a uniform magnetic field causes the magnetic particles to rotate because of mechanical torques induced by shape anisotropic magnetization. Cells touching these rotating particles are nicked, which generates transient membrane pores that enable the delivery of macromolecules into the cytosol of cells. Calcein dye, 3 and 40 kDa dextran polymers, a green fluorescence protein (GFP) plasmid, siRNA, and an enzyme (β-lactamase) were successfully delivered into HeLa cells, primary normal human dermal fibroblasts (NHDFs), and mouse cortical neurons that can be difficult to transfect. The SAMP approach offers several advantages, including easy implementation, low cost, high throughput, and efficient delivery of a broad range of macromolecules. Collectively, SAMP delivery has great potential for a broad range of academic and industrial applications. PMID:26882924
Willis, Monte S.; Townley-Tilson, W.H. Davin; Kang, Eunice Y.; Homeister, Jonathon W.; Patterson, Cam
The ubiquitin proteasome system (UPS) plays a crucial role in biological processes integral to the development of the cardiovascular system and cardiovascular diseases. The UPS prototypically recognizes specific protein substrates and places polyubiquitin chains on them for subsequent destruction by the proteasome. This system is in place to degrade not only misfolded and damaged proteins, but is essential also in regulating a host of cell signaling pathways involved in proliferation, adaptat...
Clayton, Z E; Sadeghipour, S; Patel, S
Standard therapy for atherosclerotic coronary and peripheral arterial disease is insufficient in a significant number of patients because extensive disease often precludes effective revascularization. Stem cell therapy holds promise as a supplementary treatment for these patients, as pre-clinical and clinical research has shown transplanted cells can promote angiogenesis via direct and paracrine mechanisms. Induced pluripotent stem cells (iPSCs) are a novel cell type obtained by reprogramming somatic cells using exogenous transcription factor cocktails, which have been introduced to somatic cells via viral or plasmid constructs, modified mRNA or small molecules. IPSCs are now being used in disease modelling and drug testing and are undergoing their first clinical trial, but despite recent advances, the inefficiency of the reprogramming process remains a major limitation, as does the lack of consensus regarding the optimum transcription factor combination and delivery method and the uncertainty surrounding the genetic and epigenetic stability of iPSCs. IPSCs have been successfully differentiated into vascular endothelial cells (iPSC-ECs) and, more recently, induced endothelial cells (iECs) have also been generated by direct differentiation, which bypasses the pluripotent intermediate. IPSC-ECs and iECs demonstrate endothelial functionality in vitro and have been shown to promote neovessel growth and enhance blood flow recovery in animal models of myocardial infarction and peripheral arterial disease. Challenges remain in optimising the efficiency, safety and fidelity of the reprogramming and endothelial differentiation processes and establishing protocols for large-scale production of clinical-grade, patient-derived cells. PMID:26123569
J.J. García-Vallejo; M. Ambrosini; A. Overbeek; W.E. van Riel; K. Bloem; W.W.J. Unger; F. Chiodo; J.G. Bolscher; K. Nazmi; H. Kalay; Y. van Kooyk
Dendritic cells are the most powerful type of antigen presenting cells. Current immunotherapies targeting dendritic cells have shown a relative degree of success but still require further improvement. One of the most important issues to solve is the efficiency of antigen delivery to dendritic cells
Full Text Available The generation of induced pluripotent stem cells (iPSCs has opened up a new scientific frontier in medicine. This technology has made it possible to obtain pluripotent stem cells from individuals with genetic disorders. Because iPSCs carry the identical genetic anomalies related to those disorders, iPSCs are an ideal platform for medical research. The pathophysiological cellular phenotypes of genetically heritable heart diseases such as arrhythmias and cardiomyopathies, have been modeled on cell culture dishes using disease-specific iPSC-derived cardiomyocytes. These model systems can potentially provide new insights into disease mechanisms and drug discoveries. This review focuses on recent progress in cardiovascular disease modeling using iPSCs, and discusses problems and future perspectives concerning their use.
Michael S. Detamore
Full Text Available Sensory neural hearing loss and vestibular dysfunction have become the most common forms of sensory defects, affecting millions of people worldwide. Developing effective therapies to restore hearing loss is challenging, owing to the limited regenerative capacity of the inner ear hair cells. With recent advances in understanding the developmental biology of mammalian and non-mammalian hair cells a variety of strategies have emerged to restore lost hair cells are being developed. Two predominant strategies have developed to restore hair cells: transfer of genes responsible for hair cell genesis and replacement of missing cells via transfer of stem cells. In this review article, we evaluate the use of several genes involved in hair cell regeneration, the advantages and disadvantages of the different viral vectors employed in inner ear gene delivery and the insights gained from the use of embryonic, adult and induced pluripotent stem cells in generating inner ear hair cells. Understanding the role of genes, vectors and stem cells in therapeutic strategies led us to explore potential solutions to overcome the limitations associated with their use in hair cell regeneration.
Singh, Baljeet; Garg, Tarun; Goyal, Amit K; Rath, Goutam
Cardiovascular disease is the disease that affects the cardiovascular system, vascular diseases of the brain and kidney, and peripheral arterial disease. Despite of all advances in pharmacological and clinical treatment, heart failure is a leading cause of morbidness and mortality worldwide. Many new therapeutic advance strategies, including cell transplantation, gene delivery or therapy, and cytokines or other small molecules, have been research to treat heart failure. The main aim of this review article is to focus on nano carriers advancement and addressing the problems associated with old and modern therapeutics such as nonspecific effects and poor stability. PMID:25046615
Maysam Mansouri, Maysam; Bellon-Echeverria, Itxaso; Rizk, Aurélien; Ehsaei, Zahra; Cianciolo Cosentino, Chiara; Silva, Catarina S.; Berger, Imre
Multigene delivery and subsequent cellular expression is emerging as a key technology required in diverse research fields including, synthetic and structural biology, cellular reprogramming and functional pharmaceutical screening. Current viral delivery systems such as retro- and adenoviruses suffer from limited DNA cargo capacity, thus impeding unrestricted multigene expression. We developed MultiPrime, a modular, non-cytotoxic, non-integrating, baculovirus-based vector system expediting hig...
Full Text Available Jun is a highly conserved member of the multimeric activator protein 1 transcription factor complex and plays an important role in human cancer where it is known to be critical for proliferation, cell cycle regulation, differentiation, and cell death. All of these biological functions are also crucial for embryonic development. Although all Jun null mouse embryos die at mid-gestation with persistent truncus arteriosus, a severe cardiac outflow tract defect also seen in human congenital heart disease, the developmental mechanisms are poorly understood. Here we show that murine Jun is expressed in a restricted pattern in several cell populations important for cardiovascular development, including the second heart field, pharyngeal endoderm, outflow tract and atrioventricular endocardial cushions and post-migratory neural crest derivatives. Several genes, including Isl1, molecularly mark the second heart field. Isl1 lineages include myocardium, smooth muscle, neural crest, endocardium, and endothelium. We demonstrate that conditional knockout mouse embryos lacking Jun in Isl1-expressing progenitors display ventricular septal defects, double outlet right ventricle, semilunar valve hyperplasia and aortic arch artery patterning defects. In contrast, we show that conditional deletion of Jun in Tie2-expressing endothelial and endocardial precursors does not result in aortic arch artery patterning defects or embryonic death, but does result in ventricular septal defects and a low incidence of semilunar valve defects, atrioventricular valve defects and double outlet right ventricle. Our results demonstrate that Jun is required in Isl1-expressing progenitors and, to a lesser extent, in endothelial cells and endothelial-derived endocardium for cardiovascular development but is dispensable in both cell types for embryonic survival. These data provide a cellular framework for understanding the role of Jun in the pathogenesis of congenital heart disease.
Full Text Available Mesenchymal stem cells (MSCs hold a great promise for application in several therapies due to their unique biological characteristics. In order to harness their full potential in cell-or gene-based therapies it might be advantageous to enhance some of their features through gene delivery strategies. Accordingly, we are interested in developing an efficient and safe methodology to genetically engineer human bone marrow MSC (BM MSC, enhancing their therapeutic efficacy in Regenerative Medicine. The plasmid DNA delivery was optimized using a cationic liposome-based reagent. Transfection efficiencies ranged from ~2% to ~35%, resulting from using a Lipid/DNA ratio of 1.25 with a transgene expression of 7 days. Importantly, the number of plasmid copies in different cell passages was quantified for the first time and ~20,000 plasmid copies/cell were obtained independently of cell passage. As transfected MSC have shown high viabilities (>90% and recoveries (>52% while maintaining their multipotency, this might be an advantageous transfection strategy when the goal is to express a therapeutic gene in a safe and transient way.
Many cell types were known to have migratory properties towards tumors and different research groups have shown reliable results regarding cells as delivery vehicles of therapeutics for targeted cancer treatment. Present report discusses proof of concept for 1. Cell mediated delivery of Magnetic nanoparticles (MNPs) and targeted Magnetic hyperthermia (MHT) as a cancer treatment by using in vivo mouse cancer models, 2. Cells surface engineering with chimeric proteins for targeted cancer treatment by using in vitro models. 1. Tumor homing cells can carry MNPs specifically to the tumor site and tumor burden will decrease after alternating magnetic field (AMF) exposure. To test this hypothesis, first we loaded Fe/Fe3O4 bi-magnetic NPs into neural progenitor cells (NPCs), which were previously shown to migrate towards melanoma tumors. We observed that NPCs loaded with MNPs travel to subcutaneous melanoma tumors. After alternating magnetic field (AMF) exposure, the targeted delivery of MNPs by the NPCs resulted in a mild decrease in tumor size (Chapter-2). Monocytes/macrophages (Mo/Ma) are known to infiltrate tumor sites, and also have phagocytic activity which can increase their uptake of MNPs. To test Mo/Ma-mediated MHT we transplanted Mo/Ma loaded with MNPs into a mouse model of pancreatic peritoneal carcinomatosis. We observed that MNP-loaded Mo/Ma infiltrated pancreatic tumors and, after AMF treatment, significantly prolonged the lives of mice bearing disseminated intraperitoneal pancreatic tumors (Chapter-3). 2. Targeted cancer treatment could be achieved by engineering tumor homing cell surfaces with tumor proteases cleavable, cancer cell specific recombinant therapeutic proteins. To test this, Urokinase and Calpain (tumor specific proteases) cleavable; prostate cancer cell (CaP) specific (CaP1 targeting peptide); apoptosis inducible (Caspase3 V266ED3)- rCasp3V266ED3 chimeric protein was designed in silico. Hypothesized membrane anchored chimeric protein (rCasp3V
Ndungu, John M.; Lu, Yang J.; Zhu, Shijun; Yang, Chao; Wang, Xu; Chen, Georgia; Dong M Shin; Snyder, James P.; Shoji, Mamoru; Sun, Aiming
We previously reported a novel drug delivery system, drug-linker-Phe-Phe-Arg-methylketone-(FFR-mk)-factor VIIa (fVIIa). The method utilizes tissue factor (TF), which is aberrantly and abundantly expressed on many cancer cells. The advantage of this delivery system is its ability to furnish a potent anti-cancer drug specifically to the tumor vasculature and cancer cells. In this p...
Willis, Monte S; Townley-Tilson, W H Davin; Kang, Eunice Y; Homeister, Jonathon W; Patterson, Cam
The ubiquitin proteasome system (UPS) plays a crucial role in biological processes integral to the development of the cardiovascular system and cardiovascular diseases. The UPS prototypically recognizes specific protein substrates and places polyubiquitin chains on them for subsequent destruction by the proteasome. This system is in place to degrade not only misfolded and damaged proteins, but is essential also in regulating a host of cell signaling pathways involved in proliferation, adaptation to stress, regulation of cell size, and cell death. During the development of the cardiovascular system, the UPS regulates cell signaling by modifying transcription factors, receptors, and structural proteins. Later, in the event of cardiovascular diseases as diverse as atherosclerosis, cardiac hypertrophy, and ischemia/reperfusion injury, ubiquitin ligases and the proteasome are implicated in protecting and exacerbating clinical outcomes. However, when misfolded and damaged proteins are ubiquitinated by the UPS, their destruction by the proteasome is not always possible because of their aggregated confirmations. Recent studies have discovered how these ubiquitinated misfolded proteins can be destroyed by alternative "specific" mechanisms. The cytosolic receptors p62, NBR, and histone deacetylase 6 recognize aggregated ubiquitinated proteins and target them for autophagy in the process of "selective autophagy." Even the ubiquitination of multiple proteins within whole organelles that drive the more general macro-autophagy may be due, in part, to similar ubiquitin-driven mechanisms. In summary, the crosstalk between the UPS and autophagy highlight the pivotal and diverse roles the UPS plays in maintaining protein quality control and regulating cardiovascular development and disease. PMID:20167943
Biscardi, Monica; Teodori, Elisabetta; Caporale, Roberto;
designed and synthesized to improve their MDR-reverting activity and reduce cardiovascular effects. Cytotoxicity (WST-1 methods) and functional (calcein-acetoxymethyl (Calcein-AM)) assays were performed on a resistant cell line K-562/doxR and on the mononuclear cells (MNCs) of patients with AML...... strongest activity. Results obtained from the MNCs were superimposible to K-562/doxR. Further studies on pump functional analysis confirmed the cytotoxic test results: MM 36, CTS 27 and CTS 41 showed a striking inhibition of P-glycoprotein (Pgp) efflux in K-562/doxR and MNCs. Cardiovascular activity of MM...... 36, CTS 27 and CTS 41, that are the most interesting compounds as MDR inhibitors, followed this course: MM 36>CTS 27>CTS 41, the last one presenting no cardiovascular activity. Chemosensivity to IDA in K-562/doxR cells and AML blasts could be enhanced in vitro by the adjuvant use of the six new VRP...
Chang, Chia Wei; Petrie, Tye; Clark, Alycia; Lin, Xin; Sondergaard, Claus S.; Griffiths, Leigh G.
In this study, we investigate the translational potential of a novel combined construct using an FDA-approved decellularized porcine small intestinal submucosa extracellular matrix (SIS-ECM) seeded with human or porcine mesenchymal stem cells (MSCs) for cardiovascular indications. With the emerging success of individual component in various clinical applications, the combination of SIS-ECM with MSCs could provide additional therapeutic potential compared to individual components alone for cardiovascular repair. We tested the in vitro effects of MSC-seeding on SIS-ECM on resultant construct structure/function properties and MSC phenotypes. Additionally, we evaluated the ability of porcine MSCs to modulate recipient graft-specific response towards SIS-ECM in a porcine cardiac patch in vivo model. Specifically, we determined: 1) in vitro loading-capacity of human MSCs on SIS-ECM, 2) effect of cell seeding on SIS-ECM structure, compositions and mechanical properties, 3) effect of SIS-ECM seeding on human MSC phenotypes and differentiation potential, and 4) optimal orientation and dose of porcine MSCs seeded SIS-ECM for an in vivo cardiac application. In this study, histological structure, biochemical compositions and mechanical properties of the FDA-approved SIS-ECM biomaterial were retained following MSCs repopulation in vitro. Similarly, the cellular phenotypes and differentiation potential of MSCs were preserved following seeding on SIS-ECM. In a porcine in vivo patch study, the presence of porcine MSCs on SIS-ECM significantly reduced adaptive T cell response regardless of cell dose and orientation compared to SIS-ECM alone. These findings substantiate the clinical translational potential of combined SIS-ECM seeded with MSCs as a promising therapeutic candidate for cardiac applications. PMID:27070546
Soh, Boon-Seng; Ng, Shi-Yan; Wu, Hao; Buac, Kristina; Park, Joo-Hye C; Lian, Xiaojun; Xu, Jiejia; Foo, Kylie S; Felldin, Ulrika; He, Xiaobing; Nichane, Massimo; Yang, Henry; Bu, Lei; Li, Ronald A; Lim, Bing; Chien, Kenneth R
Coronary arteriogenesis is a central step in cardiogenesis, requiring coordinated generation and integration of endothelial cell and vascular smooth muscle cells. At present, it is unclear whether the cell fate programme of cardiac progenitors to generate complex muscular or vascular structures is entirely cell autonomous. Here we demonstrate the intrinsic ability of vascular progenitors to develop and self-organize into cardiac tissues by clonally isolating and expanding second heart field cardiovascular progenitors using WNT3A and endothelin-1 (EDN1) human recombinant proteins. Progenitor clones undergo long-term expansion and differentiate primarily into endothelial and smooth muscle cell lineages in vitro, and contribute extensively to coronary-like vessels in vivo, forming a functional human-mouse chimeric circulatory system. Our study identifies EDN1 as a key factor towards the generation and clonal derivation of ISL1(+) vascular intermediates, and demonstrates the intrinsic cell-autonomous nature of these progenitors to differentiate and self-organize into functional vasculatures in vivo. PMID:26952167
Akiyama, Yukinori; Radtke, Christine; HONMOU, OSAMU; Kocsis, Jeffery D.
Bone marrow contains a population of pluripotent cells that can differentiate into a variety of cell lineages, including neural cells. When injected directly into the demyelinated spinal cord they can elicit remyelination. Recent work has shown that following systemic delivery of bone marrow cells functional improvement occurs in contusive spinal cord injury and stroke models in rat. We report here that secondary to intravenous introduction of an acutely isolated bone marrow cell fraction (mo...
Rabussay, Dietmar; Dev, Nagendu B.; Fewell, Jason; Smith, Louis C.; Widera, Georg; Zhang, Lei
The effectiveness of potentially powerful therapeutics, including DNA, is often limited by their inability to permeate the cell membrane efficiently. Electroporation (EP) also referred to as `electropermeabilization' of the outer cell membrane renders this barrier temporarily permeable by inducing `pores' across the lipid bilayer. For in vivo EP, the drug or DNA is delivered into the interstitial space of the target tissue by conventional means, followed by local EP. EP pulses of micro- to millisecond duration and field strengths of 100-1500 V cm-1 generally enhance the delivery of certain chemotherapeutic drugs by three to four orders of magnitude and intracellular delivery of DNA several hundred-fold. We have used EP in clinical studies for human cancer therapy and in animals for gene therapy and DNA vaccination. Late stage squamous cell carcinomas of the head and neck were treated with intratumoural injection of bleomycin and subsequent EP. Of the 69 tumours treated, 25% disappeared completely and another 32% were reduced in volume by more than half. Residence time of bleomycin in electroporated tumours was significantly greater than in non-electroporated lesions. Histological findings and gene expression patterns after bleomycin-EP treatment indicated rapid apoptosis of the majority of tumour cells. In animals, we demonstrated the usefulness of EP for enhanced DNA delivery by achieving normalization of blood clotting times in haemophilic dogs, and by substantially increasing transgene expression in smooth muscle cells of arterial walls using a novel porous balloon EP catheter. Finally, we have found in animal experiments that the immune response to DNA vaccines can be dramatically enhanced and accelerated by EP and co-injection of micron-sized particles. We conclude that EP represents an effective, economical and safe approach to enhance the intracellular delivery, and thus potency, of important drugs and genes for therapeutic purposes. The safety and pharmaco
Ran, Li; Tan, Xiaohua; Li, Yanchun; Zhang, Huafeng; Ma, Ruihua; Ji, Tiantian; Dong, Wenqian; Tong, Tong; Liu, Yuying; Chen, Degao; Yin, Xiaonan; Liang, Xiaoyu; Tang, Ke; Ma, Jingwei; Zhang, Yi; Cao, Xuetao; Hu, Zhuowei; Qin, Xiaofeng; Huang, Bo
Oncolytic viruses have been utilized for the treatment of various cancers. However, delivery of the viral particles to tumor cells remains a major challenge. Microparticles (MP) are vesicle forms of plasma membrane fragments of 0.1-1 μm in size that are shed by cells. We have previously shown the delivery of chemotherapeutic drugs using tumor cell-derived MPs (T-MP). Here we report that T-MPs can be utilized as a unique carrier system to deliver oncolytic adenoviruses to human tumors, leading to highly efficient cytolysis of tumor cells needed for in vivo treatment efficacy. This T-MP-mediated oncolytic virotherapy approach holds multiple advantages, including: 1) delivery of oncolytic adenovirus by T-MPs is able to avoid the antiviral effect of host antibodies; 2) delivery of oncolytic adenovirus by T-MPs is not limited by virus-specific receptor that mediates the entry of virus into tumor cells; 3) T-MPs are apt at delivering oncolytic adenoviruses to the nucleus of tumor cells as well as to stem-like tumor-repopulating cells for the desired purpose of killing them. These findings highlight a novel oncolytic adenovirus delivery system with highly promising clinical applications. PMID:26950165
Kowalski, Piotr S.; Leus, Niek G. J.; Scherphof, Gerrit L.; Ruiters, Marcel H. J.; Kamps, Jan A. A. M.; Molema, Grietje
Increased insight in the role of endothelial cells in the pathophysiology of cancer, inflammatory and cardiovascular diseases, has drawn great interest in pharmacological interventions aiming at the endothelium in diseased sites. Their location in the body makes them suitable targets for therapeutic
Heydarkhan-Hagvall, Sepideh; Nsair, Ali; Beygui, Ramin E.; Shemin, Richard J
The use of living cells as a therapeutic option presents several challenges including identification of a suitable source, development of adequate derivation, maintenance and differentiation methods, and very importantly proof of safety and efficacy. One of the major issues for cardiovascular tissue engineering is determining the ideal cell type for use in regenerative therapies.Many clinical trials have used bone marrow derived mononuclear cells (BM-MNC) (Schächinger V 2006). These clinical ...
Fuster, Valentin; Sanz, Javier
Clinical trials looking at ways to promote myocardial regeneration have reported that the administered therapies have either neutral effects or modest benefits of questionable impact. These somewhat disappointing results should emphasize the need for translational research, with bidirectional feedback between the basic research laboratory and the clinical arena. Such a translational pathway is illustrated by the quest to find an effective therapy for restenosis, which culminated in the development of sirolimus. At this point a move away from the bedside and a return to the bench seems necessary to better understand the mechanisms of action of progenitor cells and stimulating factors. Without such basic knowledge research might be prematurely discouraged and the opportunity to fully understand the true potential of cardiovascular regenerative therapy might be missed. PMID:17230204
Full Text Available The aim of this study was to determine prognostic factors for the risk of new vascular events during the first 6 months after acute myocardial infarction (AMI or atherothrombotic stroke (AS. We were interested in the prognostic role of endothelial progenitor cells (EPC and circulating endothelial cells (CEC.Between February 2009 and July 2012, 100 AMI and 50 AS patients were consecutively studied in three Spanish centres. Patients with previously documented coronary artery disease or ischemic strokes were excluded. Samples were collected within 24h of onset of symptoms. EPC and CEC were studied using flow cytometry and categorized by quartiles. Patients were followed for up to 6 months. NVE was defined as new acute coronary syndrome, transient ischemic attack (TIA, stroke, or any hospitalization or death from cardiovascular causes. The variables included in the analysis included: vascular risk factors, carotid intima-media thickness (IMT, atherosclerotic burden and basal EPC and CEC count. Multivariate survival analysis was performed using Cox regression analysis.During follow-up, 19 patients (12.66% had a new vascular event (5 strokes; 3 TIAs; 4 AMI; 6 hospitalizations; 1 death. Vascular events were associated with age (P = 0.039, carotid IMT≥0.9 (P = 0.044, and EPC count (P = 0.041 in the univariate analysis. Multivariate Cox regression analysis showed an independent association with EPC in the lowest quartile (HR: 10.33, 95%CI (1.22-87.34, P = 0.032] and IMT≥0.9 [HR: 4.12, 95%CI (1.21-13.95, P = 0.023].Basal EPC and IMT≥0.9 can predict future vascular events in patients with AMI and AS, but CEC count does not affect cardiovascular risk.
Tan, Jifu; Thomas, Antony; Liu, Yaling
Multifunctional nanomedicine holds considerable promise as the next generation of medicine that allows for targeted therapy with minimal toxicity. Most current studies on Nanoparticle (NP) drug delivery consider a Newtonian fluid with suspending NPs. However, blood is a complex biological fluid composed of deformable cells, proteins, platelets, and plasma. For blood flow in capillaries, arterioles and venules, the particulate nature of the blood needs to be considered in the delivery process....
Raphaël Lévy; Umbreen Shaheen; Yann Cesbron; Violaine Sée
Functional nanomaterials have recently attracted strong interest from the biology community, not only as potential drug delivery vehicles or diagnostic tools, but also as optical nanomaterials. This is illustrated by the explosion of publications in the field with more than 2,000 publications in the last 2 years (4,000 papers since 2000; from ISI Web of Knowledge, ‘nanoparticle and cell’ hit). Such a publication boom in this novel interdisciplinary field has resulted in papers o...
Zhang, Tao; Wang, Qing-Ming
A fuel cell is a device that can convert chemical energy into electricity directly. Among various types of fuel cells, both polymer electrolyte membrane fuel cells (PEMFCs) and direct methanol fuel cells (DMFCs) can work at low temperature (mini pumps, the size of the piezoelectric micropump is much smaller and the energy consumption is much lower. Thus, it is very viable and effective to use a piezoelectric valveless micropump for fuel delivery in miniaturized DMFC power systems.
Gui, Liqiong; Qian, Hong; Rocco, Kevin A; Grecu, Loreta; Niklason, Laura E
Cellular therapy via direct intratracheal delivery has gained interest as a novel therapeutic strategy for treating various pulmonary diseases including cystic fibrosis lung disease. However, concerns such as insufficient cell engraftment in lungs and lack of large animal model data remain to be resolved. This study aimed to establish a simple method for evaluating cell retention in lungs and to develop reproducible approaches for efficient cell delivery into mouse and pig lungs. Human lung epithelial cells including normal human bronchial/tracheal epithelial (NHBE) cells and human lung epithelial cell line A549 were infected with pSicoR-green fluorescent protein (GFP) lentivirus. GFP-labeled NHBE cells were delivered via a modified intratracheal cell instillation method into the lungs of C57BL/6J mice. Two days following cell delivery, GFP ELISA-based assay revealed a substantial cell-retention efficiency (10.48 ± 2.86%, n = 7) in mouse lungs preinjured with 2% polidocanol. When GFP-labeled A549 cells were transplanted into Yorkshire pig lungs with a tracheal intubation fiberscope, a robust initial cell attachment (22.32% efficiency) was observed at 24 h. In addition, a lentiviral vector was developed to induce the overexpression and apical localization of cystic fibrosis transmembrane conductance regulator (CFTR)-GFP fusion proteins in NHBE cells as a means of ex vivo CFTR gene transfer in nonprogenitor (relatively differentiated) lung epithelial cells. These results have demonstrated the convenience and efficiency of direct delivery of exogenous epithelial cells to lungs in mouse and pig models and provided important background for future preclinical evaluation of intratracheal cell transplantation to treat lung diseases. PMID:25416381
Maharjan, Sushila; Singh, Bijay; Jiang, Tao; Yoon, So-Yeon; Li, Hui-Shan; Kim, Girak; Gu, Min Jeong; Kim, Soo Ji; Park, Ok-Jin; Han, Seung Hyun; Kang, Sang-Kee; Yun, Cheol-Heui; Choi, Yun-Jaie; Cho, Chong-Su
A successful delivery of antigen through oral route requires to overcome several barriers, such as enzymatic barrier of gastrointestinal tract and epithelial barrier that constitutes of microfold cells (M cells) for antigen uptake. Although each barrier represents a critical step in determining the final efficiency of antigen delivery, the transcytosis of antigen by M cells in the follicle-associated epithelium (FAE) to Peyer's patches appears to be a major bottleneck. Considering the systemic administration of receptor activator of nuclear factor (NF)-ĸB ligand (RANKL) induces differentiation of receptor activator of nuclear factor (NF)-ĸB (RANK)-expressing enterocytes into M cells, here, we illustrated a promising approach of antigen delivery using full length transmembrane RANKL (mRANKL). The results showed that the intraperitoneal injection of mRANKL increased the population of dendritic cells and macrophages in mesenteric lymph nodes and spleen. Subsequently, systemic administration of mRANKL resulted in significantly higher number of functional GP2(+) M cells leading higher transcytosis of fluorescent beads through them. To corroborate the effect of mRANKL in antigen delivery through M cells, we orally delivered microparticulate antigen to mice treated with mRANKL. Oral immunization induced strong protective IgA and systemic IgG antibody responses against orally delivered antigen in mRANKL-treated mice. The higher antibody responses are attributed to the higher transcytosis of antigens through M cells. Ultimately, the higher memory B cells and effector memory CD4 T cells after oral immunization in RANKL-treated mice confirmed potency of RANKL-mediated antigen delivery. To the best of our knowledge, this is the first study to demonstrate significant induction of mucosal and humoral immune responses to M cell targeted oral vaccines after the systemic administration of RANKL. PMID:26851393
Full Text Available Diseases such as hypertension, atherosclerosis, hyperlipidemia, and diabetes are associated with vascular functional and structural changes including endothelial dysfunction, altered contractility and vascular remodeling. Cellular events underlying these processes involve changes in vascular smooth muscle cell (VSMC growth, apoptosis/anoikis, cell migration, inflammation, and fibrosis. Many factors influence cellular changes, of which angiotensin II (Ang II appears to be amongst the most important. The physiological and pathophysiological actions of Ang II are mediated primarily via the Ang II type 1 receptor. Growing evidence indicates that Ang II induces its pleiotropic vascular effects through NADPH-driven generation of reactive oxygen species (ROS. ROS function as important intracellular and intercellular second messengers to modulate many downstream signaling molecules, such as protein tyrosine phosphatases, protein tyrosine kinases, transcription factors, mitogen-activated protein kinases, and ion channels. Induction of these signaling cascades leads to VSMC growth and migration, regulation of endothelial function, expression of pro-inflammatory mediators, and modification of extracellular matrix. In addition, ROS increase intracellular free Ca2+ concentration ([Ca2+]i, a major determinant of vascular reactivity. ROS influence signaling molecules by altering the intracellular redox state and by oxidative modification of proteins. In physiological conditions, these events play an important role in maintaining vascular function and integrity. Under pathological conditions ROS contribute to vascular dysfunction and remodeling through oxidative damage. The present review focuses on the biology of ROS in Ang II signaling in vascular cells and discusses how oxidative stress contributes to vascular damage in cardiovascular disease.
Jiang, Xinglu; Wang, Guobao; Liu, Ru; Wang, Yaling; Wang, Yongkui; Qiu, Xiaozhong; Gao, Xueyun
To date, RNase degradation and endosome/lysosome trapping are still serious problems for siRNA-based molecular therapy, although different kinds of delivery formulations have been tried. In this report, a cell penetrating peptide (CPP, including a positively charged segment, a linear segment, and a hydrophobic segment) and a single wall carbon nanotube (SWCNT) are applied together by a simple method to act as a siRNA delivery system. The siRNAs first form a complex with the positively charged segment of CPP via electrostatic forces, and the siRNA-CPP further coats the surface of the SWCNT via hydrophobic interactions. This siRNA delivery system is non-sensitive to RNase and can avoid endosome/lysosome trapping in vitro. When this siRNA delivery system is studied in Hela cells, siRNA uptake was observed in 98% Hela cells, and over 70% mRNA of mammalian target of rapamycin (mTOR) is knocked down, triggering cell apoptosis on a significant scale. Our siRNA delivery system is easy to handle and benign to cultured cells, providing a very efficient approach for the delivery of siRNA into the cell cytosol and cleaving the target mRNA therein.
Lavandero, Sergio; Chiong, Mario; Rothermel, Beverly A.; Hill, Joseph A.
Cardiovascular disease is the leading cause of death worldwide. As such, there is great interest in identifying novel mechanisms that govern the cardiovascular response to disease-related stress. First described in failing hearts, autophagy within the cardiovascular system has been widely characterized in cardiomyocytes, cardiac fibroblasts, endothelial cells, vascular smooth muscle cells, and macrophages. In all cases, a window of optimal autophagic activity appears to be critical to the mai...
Full Text Available Living cells encounter many stimuli from the immediate environment. Receptors recognize these environmental cues and transduce signals to produce cell responses. The frequency of a signal is now emerging as an important factor determining cell responses. As a componentry system in understanding temporal stimulation, microfluidic devices allow the observation of cell behaviour under dynamic stimulation and controllable environment. In this paper we describe the design, construction and characterization of a microfluidic device suitable for cell stimulation studies.
Tan, Jifu; Thomas, Antony; Liu, Yaling
Multifunctional nanomedicine holds considerable promise as the next generation of medicine that allows for targeted therapy with minimal toxicity. Most current studies on Nanoparticle (NP) drug delivery consider a Newtonian fluid with suspending NPs. However, blood is a complex biological fluid composed of deformable cells, proteins, platelets, and plasma. For blood flow in capillaries, arterioles and venules, the particulate nature of the blood needs to be considered in the delivery process. The existence of the cell-free-layer and NP-cell interaction will largely influence both the dispersion and binding rates, thus impact targeted delivery efficacy. In this paper, a particle-cell hybrid model is developed to model NP transport, dispersion, and binding dynamics in blood suspension. The motion and deformation of red blood cells is captured through the Immersed Finite Element Method. The motion and adhesion of individual NPs are tracked through Brownian adhesion dynamics. A mapping algorithm and an interaction potential function are introduced to consider the cell-particle collision. NP dispersion and binding rates are derived from the developed model under various rheology conditions. The influence of red blood cells, vascular flow rate, and particle size on NP distribution and delivery efficacy is characterized. A non-uniform NP distribution profile with higher particle concentration near the vessel wall is observed. Such distribution leads to over 50% higher particle binding rate compared to the case without RBC considered. The tumbling motion of RBCs in the core region of the capillary is found to enhance NP dispersion, with dispersion rate increases as shear rate increases. Results from this study contribute to the fundamental understanding and knowledge on how the particulate nature of blood influences NP delivery, which will provide mechanistic insights on the nanomedicine design for targeted drug delivery applications. PMID:22375153
Progulske-Fox, A; Kozarov, E; Dorn, B; Dunn, W; Burks, J; Wu, Y
Our laboratory is interested in the genes and gene products involved in the interactions between Porphyromonas gingivalis (Pg) and the host. These interactions may occur in either the periodontal tissues or other non-oral host tissues such as those of the cardiovascular system. We have previously reported the cloning of several genes encoding hemagglutinins, surface proteins that interact with the host tissues, and are investigating their roles in the disease process. Primary among these is HagA, a very large protein with multiple functional groups that have significant sequence homology to protease genes of this species. Preliminary evidence indicates that an avirulent Salmonella typhimurium strain containing hagA is virulent in mice. These data indicate that HagA may be a key virulence factor of Pg. Additionally, we are investigating the invasion of primary human coronary artery endothelial cells (HCAEC) by Pg because of the recent epidemiological studies indicating a correlation between periodontal disease (PD) and coronary heart disease (CHD). We found that some, but not all, strains of Pg are able to invade these cells. Scanning electron microsopy of the infected HCAEC demonstrated that the invading organisms initially attached to the host cell surface as aggregates and by a "pedestal"-like structure. By transmission electronmicroscopy it could be seen that internalized bacteria were present within multimembranous compartments localized with rough endoplasmic reticulum. In addition, invasion of the HCAEC by Pg resulted in an increase in the degradation of long-lived cellular proteins. These data indicate that Pg are present within autophagosomes and may use components of the autophagic pathway as a means to survive intracellularly. However, Pg presence within autophagosomes in KB cells could not be observed or detected. It is therefore likely that Pg uses different invasive mechanisms for different host cells. This and the role of HagA in invasion is currently
Shantanu Balkundi1, Ari S Nowacek1, Ram S Veerubhotla1, Han Chen2, Andrea Martinez-Skinner1, Upal Roy1, R Lee Mosley1,3, Georgette Kanmogne1, Xinming Liu1,3,4, Alexander V Kabanov3,4, Tatiana Bronich3,4, JoEllyn McMillan1, Howard E Gendelman1,31Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, USA; 2Center for Biotechnology, University of Nebraska-Lincoln, Lincoln, NE, USA; 3Center for Drug Delivery and Nanomedicine, University of Nebr...
Han, Xin; Liu, Zongbin; Jo, Myeong Chan; Zhang, Kai; Li, Ying; Zeng, Zihua; Li, Nan; Zu, Youli; Qin, Lidong
The CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR-associated) nuclease system represents an efficient tool for genome editing and gene function analysis. It consists of two components: single-guide RNA (sgRNA) and the enzyme Cas9. Typical sgRNA and Cas9 intracellular delivery techniques are limited by their reliance on cell type and exogenous materials as well as their toxic effects on cells (for example, electroporation). We introduce and optimize a microfluidic membrane deformation method to deliver sgRNA and Cas9 into different cell types and achieve successful genome editing. This approach uses rapid cell mechanical deformation to generate transient membrane holes to enable delivery of biomaterials in the medium. We achieved high delivery efficiency of different macromolecules into different cell types, including hard-to-transfect lymphoma cells and embryonic stem cells, while maintaining high cell viability. With the advantages of broad applicability across different cell types, particularly hard-to-transfect cells, and flexibility of application, this method could potentially enable new avenues of biomedical research and gene targeting therapy such as mutation correction of disease genes through combination of the CRISPR-Cas9-mediated knockin system. PMID:26601238
Xiao, Yuhong; Kwon, Kwang-Chul; Hoffman, Brad E; Kamesh, Aditya; Jones, Noah T; Herzog, Roland W; Daniell, Henry
Targeted oral delivery of GFP fused with a GM1 receptor binding protein (CTB) or human cell penetrating peptide (PTD) or dendritic cell peptide (DCpep) was investigated. Presence of GFP(+) intact plant cells between villi of ileum confirm their protection in the digestive system from acids/enzymes. Efficient delivery of GFP to gut-epithelial cells by PTD or CTB and to M cells by all these fusion tags confirm uptake of GFP in the small intestine. PTD fusion delivered GFP more efficiently to most tissues or organs than the other two tags. GFP was efficiently delivered to the liver by all fusion tags, likely through the gut-liver axis. In confocal imaging studies of human cell lines using purified GFP fused with different tags, GFP signal of DCpep-GFP was only detected within dendritic cells. PTD-GFP was only detected within kidney or pancreatic cells but not in immune modulatory cells (macrophages, dendritic, T, B, or mast cells). In contrast, CTB-GFP was detected in all tested cell types, confirming ubiquitous presence of GM1 receptors. Such low-cost oral delivery of protein drugs to sera, immune system or non-immune cells should dramatically lower their cost by elimination of prohibitively expensive fermentation, protein purification cold storage/transportation and increase patient compliance. PMID:26706477
van Rossum Albert C; Raman Subha V; McConnell Michael V; Lawson Mark A; Higgins Charles B; Friedrich Matthias G; Bogaert Jan G; Bluemke David; Hundley W Gregory; Flamm Scott; Kramer Christopher M; Nagel Eike; Neubauer Stefan
Abstract These reporting guidelines are recommended by the Society for Cardiovascular Magnetic Resonance (SCMR) to provide a framework for healthcare delivery systems to disseminate cardiac and vascular imaging findings related to the performance of cardiovascular magnetic resonance (CMR) examinations.
Full Text Available Cheng-Hung Lee,1,2 Yu-Huang Lin,3 Shang-Hung Chang,1 Chun-Der Tai,3 Shih-Jung Liu,2 Yen Chu,4 Chao-Jan Wang,5 Ming-Yi Hsu,5 Hung Chang,6 Gwo-Jyh Chang,7 Kuo-Chun Hung,1 Ming-Jer Hsieh,1 Fen-Chiung Lin,1 I-Chang Hsieh,1 Ming-Shien Wen,1 Yenlin Huang81Division of Cardiology, Department of Internal Medicine, Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Linkou, 2Department of Mechanical Engineering, 3Graduate Institute of Medical Mechatronics, Chang Gung University, 4Laboratory of Cardiovascular Physiology, Division of Thoracic and Cardiovascular Surgery, 5Department of Medical Imaging and Intervention, 6Hematology-Oncology Division, Department of Internal Medicine, Chang Gung Memorial Hospital, Linkou, 7Graduate Institute of Clinical Medicinal Sciences, Chang Gung University College of Medicine, Linkou, 8Department of Anatomical Pathology, Chang Gung Memorial Hospital, Linkou, Tao-Yuan, TaiwanAbstract: Incomplete endothelialization, blood cell adhesion to vascular stents, and inflammation of arteries can result in acute stent thromboses. The systemic administration of acetylsalicylic acid decreases endothelial dysfunction, potentially reducing thrombus, enhancing vasodilatation, and inhibiting the progression of atherosclerosis; but, this is weakened by upper gastrointestinal bleeding. This study proposes a hybrid stent with biodegradable nanofibers, for the local, sustained delivery of acetylsalicylic acid to injured artery walls. Biodegradable nanofibers are prepared by first dissolving poly(D,L-lactide-co-glycolide and acetylsalicylic acid in 1,1,1,3,3,3-hexafluoro-2-propanol. The solution is then electrospun into nanofibrous tubes, which are then mounted onto commercially available bare-metal stents. In vitro release rates of pharmaceuticals from nanofibers are characterized using an elution method, and a high-performance liquid chromatography assay. The experimental results suggest that biodegradable nanofibers
Grześkowiak, Bartosz F; Hryhorowicz, Magdalena; Tuśnio, Karol; Grzeszkowiak, Mikołaj; Załęski, Karol; Lipiński, Daniel; Zeyland, Joanna; Mykhaylyk, Olga; Słomski, Ryszard; Jurga, Stefan; Woźniak, Anna
The transgenic process allows for obtaining genetically modified animals for divers biomedical applications. A number of transgenic animals for xenotransplantation have been generated with the somatic cell nuclear transfer (SCNT) method. Thereby, efficient nucleic acid delivery to donor cells such as fibroblasts is of particular importance. The objective of this study was to establish stable transgene expressing porcine fetal fibroblast cell lines using magnetic nanoparticle-based gene delivery vectors under a gradient magnetic field. Magnetic transfection complexes prepared by self-assembly of suitable magnetic nanoparticles, plasmid DNA, and an enhancer under an inhomogeneous magnetic field enabled the rapid and efficient delivery of a gene construct (pCD59-GFPBsd) into porcine fetal fibroblasts. The applied vector dose was magnetically sedimented on the cell surface within 30 min as visualized by fluorescence microscopy. The PCR and RT-PCR analysis confirmed not only the presence but also the expression of transgene in all magnetofected transgenic fibroblast cell lines which survived antibiotic selection. The cells were characterized by high survival rates and proliferative activities as well as correct chromosome number. The developed nanomagnetic gene delivery formulation proved to be an effective tool for the production of genetically engineered fibroblasts and may be used in future in SCNT techniques for breeding new transgenic animals for the purpose of xenotransplantation. PMID:27048425
Brinker, C Jeffrey; Ashley, Carlee Erin; Jiang, Xingmao; Liu, Juewen; Peabody, David S; Wharton, Walker Richard; Carnes, Eric; Chackerian, Bryce; Willman, Cheryl L
Various embodiments provide materials and methods for synthesizing protocells for use in targeted delivery of cargo components to cancer cells. In one embodiment, the lipid bilayer can be fused to the porous particle core to form a protocell. The lipid bilayer can be modified with targeting ligands or other ligands to achieve targeted delivery of cargo components that are loaded within the protocell to a target cell, e.g., a type of cancer. Shielding materials can be conjugated to the surface of the lipid bilayer to reduce undesired non-specific binding.
Campbell, Richard A.; Watkins, Erik B.; Jagalski, Vivien;
We show that both gravity and electrostatics are key factors regulating interactions between model cell membranes and self-assembled liquid crystalline aggregates of dendrimers and phospholipids. The system is a proxy for the trafficking of reservoirs of therapeutic drugs to cell membranes for slow...... the aggregates to activate endocytosis pathways on specific cell types is discussed in the context of targeted drug delivery applications. © 2014 American Chemical Society....
Culiat, Cymbeline T
It has been identified in accordance with the present invention that Nell1 is essential for normal cardiovascular development by promoting proper formation of the heart and blood vessels. The present invention therefore provides therapeutic methods for treating cardiovascular disorders by employing a Nell1 protein or nucleic acid molecule.
Full Text Available Kun Qian,1,2 Jing Wu,1 Enqi Zhang,1 Yingge Zhang,3 Ailing Fu1 1School of Pharmaceutical Sciences, Southwest University, 2College of Plant Protection, Southwest University, Chongqing, People’s Republic of China; 3Institute of Pharmacology and Toxicology, Key Laboratory of Nanopharmacology and Nanotoxicology, Beijing Academy of Medical Sciences, Beijing, People’s Republic of China Abstract: Highly-efficient delivery of macromolecules into cells for both imaging and therapy (theranostics remains a challenge for the design of a delivery system. Here, we suggested a novel hybrid protein–lipid polymer nanocapsule as an effective and nontoxic drug delivery and imaging carrier. The biodegradable nanocapsules showed the typical double emulsion features, including fluorescently labeled bovine serum albumin shell, oil phase containing poly(lactic-co-glycolic acid and linoleic acid, and inner aqueous phase. The nanocapsules were spherical in shape, with an average size of about 180 nm. Proteins packed into the inner aqueous phase of the nanocapsules could be delivered into cells with high efficiency, and the fluorescence of the fluorescently labeled bovine serum albumin could be used for tracing the protein migration and cellular location. Further studies suggested that the co-delivery of transcription factor p53 and lipophilic drug paclitaxel with the nanocapsules acted synergistically to induce Hela cell apoptosis, and the fluorescence of apoptotic cells was clearly observed under a fluorescence microscope. Such multifunctional delivery system would have great potential applications in drug delivery and theranostic fields. Keywords: emulsion, protein transport, fluorescence labeling, theranostics, cell apoptosis
Philpott, Caroline C.; Ryu, Moon-Suhn
Eukaryotic cells contain hundreds of proteins that require iron cofactors for activity. These iron enzymes are located in essentially every subcellular compartment; thus, iron cofactors must travel to every compartment in the cell. Iron cofactors exist in three basic forms: Heme, iron–sulfur clusters, and simple iron ions (also called non-heme iron). Iron ions taken up by the cell initially enter a kinetically labile, exchangeable pool that is referred to as the labile iron pool. The majority...
A major challenge with the use of quantum dots (QDs) for cellular imaging and biomolecular delivery is the attainment of QDs freely dispersed inside the cells. Conventional methods such as endocytosis, lipids based delivery and electroporation are associated with delivery of QDs in vesicles and/or as aggregates that are not monodispersed. In this study, we demonstrate a new technique for reversible permeabilization of cells to enable the introduction of freely dispersed QDs within the cytoplasm. Our approach combines osmosis driven fluid transport into cells achieved by creating a hypotonic environment and reversible permeabilization using low concentrations of cell permeabilization agents like Saponin. Our results confirm that highly efficient endocytosis-free intracellular delivery of QDs can be accomplished using this method. The best results were obtained when the cells were treated with 50 μg ml−1 Saponin in a hypotonic buffer at a 3:2 physiological buffer:DI water ratio for 5 min at 4 ° C. (paper)
Medepalli, Krishnakiran; Alphenaar, Bruce W.; Keynton, Robert S.; Sethu, Palaniappan
A major challenge with the use of quantum dots (QDs) for cellular imaging and biomolecular delivery is the attainment of QDs freely dispersed inside the cells. Conventional methods such as endocytosis, lipids based delivery and electroporation are associated with delivery of QDs in vesicles and/or as aggregates that are not monodispersed. In this study, we demonstrate a new technique for reversible permeabilization of cells to enable the introduction of freely dispersed QDs within the cytoplasm. Our approach combines osmosis driven fluid transport into cells achieved by creating a hypotonic environment and reversible permeabilization using low concentrations of cell permeabilization agents like Saponin. Our results confirm that highly efficient endocytosis-free intracellular delivery of QDs can be accomplished using this method. The best results were obtained when the cells were treated with 50 μg ml-1 Saponin in a hypotonic buffer at a 3:2 physiological buffer:DI water ratio for 5 min at 4 ° C.
Zhang, Yong-Tai; Shen, Li-Na; Wu, Zhong-Hua; Zhao, Ji-Hui; Feng, Nian-Ping
This study investigated the effect of skin viability on its permeability to psoralen delivered by ethosomes, as compared with liposomes. With decreasing skin viability, the amount of liposome-delivered psoralen that penetrated through the skin increased, whereas skin deposition of psoralen from both ethosomes and liposomes reduced. Psoralen delivery to human-immortalized epidermal cells was more effective using liposomes, whereas delivery to human embryonic skin fibroblast cells was more effective when ethosomes were used. These findings agreed with those of in vivo studies showing that skin psoralen deposition from ethosomes and liposomes first increased and then plateaued overtime, which may indicate gradual saturation of intracellular drug delivery. It also suggested that the reduced deposition of ethosome- or liposome-delivered psoralen in skin with reduced viability may relate to reduced cellular uptake. This work indicated that the effects of skin viability should be taken into account when evaluating nanocarrier-mediated drug skin permeation. PMID:25070929
Full Text Available BACKGROUND: Microscopic techniques enable real-space imaging of complex biological events and processes. They have become an essential tool to confirm and complement hypotheses made by biomedical scientists and also allow the re-examination of existing models, hence influencing future investigations. Particularly imaging live cells is crucial for an improved understanding of dynamic biological processes, however hitherto live cell imaging has been limited by the necessity to introduce probes within a cell without altering its physiological and structural integrity. We demonstrate herein that this hurdle can be overcome by effective cytosolic delivery. PRINCIPAL FINDINGS: We show the delivery within several types of mammalian cells using nanometre-sized biomimetic polymer vesicles (a.k.a. polymersomes that offer both highly efficient cellular uptake and endolysomal escape capability without any effect on the cellular metabolic activity. Such biocompatible polymersomes can encapsulate various types of probes including cell membrane probes and nucleic acid probes as well as labelled nucleic acids, antibodies and quantum dots. SIGNIFICANCE: We show the delivery of sufficient quantities of probes to the cytosol, allowing sustained functional imaging of live cells over time periods of days to weeks. Finally the combination of such effective staining with three-dimensional imaging by confocal laser scanning microscopy allows cell imaging in complex three-dimensional environments under both mono-culture and co-culture conditions. Thus cell migration and proliferation can be studied in models that are much closer to the in vivo situation.
Brahmamdam, Pavan; Watanabe, Eizo; Unsinger, Jacqueline; Chang, Katherine C; Schierding, William; Hoekzema, Andrew S; Zhou, Tony T; McDonough, Jacquelyn S; Holemon, Heather; Heidel, Jeremy D; Coopersmith, Craig M; McDunn, Jonathan E; Hotchkiss, Richard S
Immune suppression is a major cause of morbidity and mortality in the patients with sepsis. Apoptotic loss of immune effector cells such as CD4 T and B cells is a key component in the loss of immune competence in sepsis. Inhibition of lymphocyte apoptosis has led to improved survival in animal models of sepsis. Using quantitative real-time polymerase chain reaction of isolated splenic CD4 T and B cells, we determined that Bim and PUMA, two key cell death proteins, are markedly upregulated during sepsis. Lymphocytes have been notoriously difficult to transfect with small interfering RNA (siRNA). Consequently a novel, cyclodextrin polymer-based, transferrin receptor-targeted, delivery vehicle was used to coadminister siRNA to Bim and PUMA to mice immediately after cecal ligation and puncture. Antiapoptotic siRNA-based therapy markedly decreased lymphocyte apoptosis and prevented the loss of splenic CD4 T and B cells. Flow cytometry confirmed in vivo delivery of siRNA to CD4 T and B cells and also demonstrated decreases in intracellular Bim and PUMA protein. In conclusion, Bim and PUMA are two critical mediators of immune cell death in sepsis. Use of a novel cyclodextrin polymer-based, transferrin receptor-targeted siRNA delivery vehicle enables effective administration of antiapoptotic siRNAs to lymphocytes and reverses the immune cell depletion that is a hallmark of this highly lethal disorder. PMID:19033888
Mastorakos, Panagiotis; Kambhampati, Siva P.; Mishra, Manoj K.; Wu, Tony; Song, Eric; Hanes, Justin; Kannan, Rangaramanujam M.
Ocular gene therapy holds promise for the treatment of numerous blinding disorders. Despite the significant progress in the field of viral and non-viral gene delivery to the eye, significant obstacles remain in the way of achieving high-level transgene expression without adverse effects. The retinal pigment epithelium (RPE) is involved in the pathogenesis of retinal diseases and is a key target for a number of gene-based therapeutics. In this study, we addressed the inherent drawbacks of non-viral gene vectors and combined different approaches to design an efficient and safe dendrimer-based gene-delivery platform for delivery to human RPE cells. We used hydroxyl-terminated polyamidoamine (PAMAM) dendrimers functionalized with various amounts of amine groups to achieve effective plasmid compaction. We further used triamcinolone acetonide (TA) as a nuclear localization enhancer for the dendrimer-gene complex and achieved significant improvement in cell uptake and transfection of hard-to-transfect human RPE cells. To improve colloidal stability, we further shielded the gene vector surface through incorporation of PEGylated dendrimer along with dendrimer-TA for DNA complexation. The resultant complexes showed improved stability while minimally affecting transgene delivery, thus improving the translational relevance of this platform.Ocular gene therapy holds promise for the treatment of numerous blinding disorders. Despite the significant progress in the field of viral and non-viral gene delivery to the eye, significant obstacles remain in the way of achieving high-level transgene expression without adverse effects. The retinal pigment epithelium (RPE) is involved in the pathogenesis of retinal diseases and is a key target for a number of gene-based therapeutics. In this study, we addressed the inherent drawbacks of non-viral gene vectors and combined different approaches to design an efficient and safe dendrimer-based gene-delivery platform for delivery to human RPE
Reeves, Analise Z; Spears, William E; Du, Juan; Tan, Kah Yong; Wagers, Amy J; Lesser, Cammie F
Many Gram-negative pathogens encode type 3 secretion systems, sophisticated nanomachines that deliver proteins directly into the cytoplasm of mammalian cells. These systems present attractive opportunities for therapeutic protein delivery applications; however, their utility has been limited by their inherent pathogenicity. Here, we report the reengineering of a laboratory strain of Escherichia coli with a tunable type 3 secretion system that can efficiently deliver heterologous proteins into mammalian cells, thereby circumventing the need for virulence attenuation. We first introduced a 31 kB region of Shigella flexneri DNA that encodes all of the information needed to form the secretion nanomachine onto a plasmid that can be directly propagated within E. coli or integrated into the E. coli chromosome. To provide flexible control over type 3 secretion and protein delivery, we generated plasmids expressing master regulators of the type 3 system from either constitutive or inducible promoters. We then constructed a Gateway-compatible plasmid library of type 3 secretion sequences to enable rapid screening and identification of sequences that do not perturb function when fused to heterologous protein substrates and optimized their delivery into mammalian cells. Combining these elements, we found that coordinated expression of the type 3 secretion system and modified target protein substrates produces a nonpathogenic strain that expresses, secretes, and delivers heterologous proteins into mammalian cells. This reengineered system thus provides a highly flexible protein delivery platform with potential for future therapeutic applications. PMID:25853840
Luo, Zhong; Cai, Kaiyong; Hu, Yan; Zhang, Beilu; Xu, Dawei
A nanoreservoir for efficient intracellular anticancer drug delivery based on mesoporous silica nanoparticles end-capped with lactobionic acid-grafted bovine serum albumin is fabricated. It demonstrates great potential for both cell-specific endocytosis and intracellular pH-responsive controlled release of drugs. A possible endocytosis pathway/mechanism of the smart controlled drug release system is proposed. PMID:23184747
Ashley, Carlee E.; CARNES, ERIC C.; Phillips, Genevieve K.; Padilla, David; Durfee, Paul N.; Brown, Page A.; Hanna, Tracey N.; Liu, Juewen; Phillips, Brandy; Carter, Mark B.; Carroll, Nick J.; Jiang, Xingmao; Dunphy, Darren R.; Willman, Cheryl L.; Petsev, Dimiter N.
Encapsulation of drugs within nanocarriers that selectively target malignant cells promises to mitigate side effects of conventional chemotherapy and to enable delivery of the unique drug combinations needed for personalized medicine. To realize this potential, however, targeted nanocarriers must simultaneously overcome multiple challenges, including specificity, stability, and a high capacity for disparate cargos. Here we report porous nanoparticle-supported lipid bilayers (protocells) that ...
Wu, Ting-Hsiang; Teslaa, Tara; Kalim, Sheraz; French, Christopher T; Maghadam, Shahriar; Wall, Randolph; Miller, Jeffery F.; Witte, Owen N.; Teitell, Michael A.; Chiou, Pei-Yu
It is difficult to achieve controlled cutting of elastic, mechanically fragile, and rapidly resealing mammalian cell membranes. Here, we report a photothermal nanoblade that utilizes a metallic nanostructure to harvest short laser pulse energy and convert it into a highly localized explosive vapor bubble, which rapidly punctures a lightly-contacting cell membrane via high-speed fluidic flows and induced transient shear stress. The cavitation bubble pattern is controlled by the metallic struct...
Caroline C Philpott
Full Text Available Eukaryotic cells contain hundreds of proteins that require iron cofactors for activity. These iron enzymes are located in essentially every subcellular compartment; thus, iron cofactors must travel to every compartment in the cell. Iron cofactors exist in three basic forms: Heme, iron-sulfur clusters, and simple iron ions (also called non-heme iron. Iron ions taken up by the cell initially enter a kinetically labile, exchangeable pool that is referred to as the labile iron pool. The majority of the iron in this pool is delivered to mitochondria, where it is incorporated into heme and iron-sulfur clusters, as well as non-heme iron enzymes. These cofactors must then be distributed to nascent proteins in the mitochondria, cytosol, and membrane-bound organelles. Emerging evidence suggests that specific systems exist for the distribution of iron cofactors within the cell. These systems include membrane transporters, protein chaperones, specialized carriers, and small molecules. This review focuses on the distribution of iron ions in the cytosol and will highlight differences between the iron distribution systems of simple eukaryotes and mammalian cells.
Knollmann, Björn C.
As part of the series on Controversies in Cardiovascular Research, the article reviews the strengths and limitations of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) as models of cardiac arrhythmias. Specifically, the article attempts to answer the following questions: Which clinical arrhythmias can be modeled by iPSC-CM? How well can iPSC-CM model adult ventricular myocytes? What are the strengths and limitations of published iPSC-CM arrhythmia models? What new mechanistic i...
Jantzen, Alexandra E.; Lane, Whitney O.; Gage, Shawn M.; Haseltine, Justin M; Galinat, Lauren J; Jamiolkowski, Ryan M.; Lin, Fu-Hsiung; Truskey, George A.; Achneck, Hardean E.
Implantable cardiovascular devices are manufactured from artificial materials (e.g. titanium (Ti), expanded polytetrafluoroethylene), which pose the risk of thromboemboli formation1,2,3. We have developed a method to line the inside surface of Ti tubes with autologous blood-derived human or porcine endothelial progenitor cells (EPCs)4. By implanting Ti tubes containing a confluent layer of porcine EPCs in the inferior vena cava (IVC) of pigs, we tested the improved biocompatibility of the cel...
Wei Junchen; Han Zhiqiang; Weng Yanjie; Li Kezhen; Cao Li; Wu Mingfu; Hu Wencheng; Wu Peng; Ma Quanfu; Xu Hongbin; You Lanying; Liao Shujie; Gao Qinglei; Fang Yong; Li Xiao
Abstract Background Mesenchymal stem cells (MSCs) have been considered to be the attractive vehicles for delivering therapeutic agents toward various tumor diseases. This study was to explore the distribution pattern, kinetic delivery of adenovirus, and therapeutic efficacy of the MSC loading of E1A mutant conditionally replicative adenovirus Adv-Stat3(-) which selectively replicated and expressed high levels of anti-sense Stat3 complementary DNA in breast cancer and melanoma cells. Methods W...
Clairambault, Jean; Fercoq, Olivier
In this paper, we introduce a model for drug delivery optimisation in a chronotherapeu-tics framework. We present a pharmacokinetics and pharmacodynamics model for oxaliplatin and 5-Fluorouracil, a classic therapeutic association in the treatment of colorectal cancer. We derive the phar-macokinetics model using law of mass action and enzyme kinetics. We design an age-structured cell cycle PDE model with drug damage and repair phases to account for the effect of the drugs on proliferating cell...
Hirsch, Stephan Tobias Florian; von Peter, Sebastian; Dubin, Grzegorz; Mittler, Dominik; Jacobsen, Frank; Lehnhardt, Marcus; Eriksson, Elof; Steinau, Hans-Ulrich; Steinsträßer, Lars
The adenoviral transfer of therapeutic genes into epidermal and dermal cells is an interesting approach to treat skin diseases and to promote wound healing. The aim of this study was to assess the in vitro and in vivo transfection efficacy in skin and burn wounds after adenoviral gene delivery. Primary keratinocytes (HKC), fibroblasts (HFB), and HaCaT cells were transfected using different concentrations of an adenoviral construct (eGFP). Transfection efficiency and cytotoxicity was determine...
Subramanian, Nithya; Kanwar, Jagat R; Akilandeswari, Balachandran; Kanwar, Rupinder K; Khetan, Vikas; Krishnakumar, Subramanian
A chimeric aptamer-DNAzyme conjugate was generated for the first time using a nucleolin aptamer (NCL-APT) and survivin Dz (Sur_Dz) and exhibited the targeted killing of cancer cells. This proof of concept of using an aptamer for the delivery of DNAzyme can be applied to other cancer types to target survivin in cancer cells in a specific manner. PMID:25797393
The effectiveness of potentially powerful therapeutics, including DNA, is often limited by their inability to permeate the cell membrane efficiently. Electroporation (EP) also referred to as 'electropermeabilization' of the outer cell membrane renders this barrier temporarily permeable by inducing 'pores' across the lipid bilayer. For in vivo EP, the drug or DNA is delivered into the interstitial space of the target tissue by conventional means, followed by local EP. EP pulses of micro- to millisecond duration and field strengths of 100-1500 V cm-1 generally enhance the delivery of certain chemotherapeutic drugs by three to four orders of magnitude and intracellular delivery of DNA several hundred-fold. We have used EP in clinical studies for human cancer therapy and in animals for gene therapy and DNA vaccination. Late stage squamous cell carcinomas of the head and neck were treated with intratumoural injection of bleomycin and subsequent EP. Of the 69 tumours treated, 25% disappeared completely and another 32% were reduced in volume by more than half. Residence time of bleomycin in electroporated tumours was significantly greater than in non-electroporated lesions. Histological findings and gene expression patterns after bleomycin-EP treatment indicated rapid apoptosis of the majority of tumour cells. In animals, we demonstrated the usefulness of EP for enhanced DNA delivery by achieving normalization of blood clotting times in haemophilic dogs, and by substantially increasing transgene expression in smooth muscle cells of arterial walls using a novel porous balloon EP catheter. Finally, we have found in animal experiments that the immune response to DNA vaccines can be dramatically enhanced and accelerated by EP and co-injection of micron-sized particles. We conclude that EP represents an effective, economical and safe approach to enhance the intracellular delivery, and thus potency, of important drugs and genes for therapeutic purposes. The safety and pharmaco
Lee, Dylan E; Ayoub, Nagi; Agrawal, Devendra K
Mesenchymal stem cells (MSCs) (also known as multipotent mesenchymal stromal cells) possess the capacity for self-renewal and multi-lineage differentiation, and their ability to enhance cutaneous wound healing has been well characterized. Acting via paracrine interactions, MSCs accelerate wound closure, increase angiogenesis, promote resolution of wound inflammation, favorably regulate extracellular matrix remodeling, and encourage regeneration of skin with normal architecture and function. A number of studies have employed novel methods to amplify the delivery and efficacy of MSCs. Non-traditional sources of MSCs, including Wharton's jelly and medical waste material, have shown efficacy comparable to that of traditional sources, such as bone marrow and adipose tissue. The potential of alternative methods to both introduce MSCs into wounds and increase migration of MSCs into wound areas has also been demonstrated. Taking advantage of the associations between MSCs with M2 macrophages and microRNA, methods to enhance the immunomodulatory capacity of MSCs have shown success. New measures to enhance angiogenic capabilities have also exhibited effectiveness, often demonstrated by increased levels of proangiogenic vascular endothelial growth factor. Finally, hypoxia has been shown to have strong wound-healing potential in terms of increasing MSC efficacy. We have critically reviewed the results of the novel studies that show promise for the continued development of MSC-based wound-healing therapies and provide direction for continued research in this field. PMID:26960535
Devarajan, Keerthana; Forrest, M Laird; Detamore, Michael S; Staecker, Hinrich
Hearing is one of our main sensory systems and having a hearing disorder can have a significant impact in an individual's quality of life. Sensory neural hearing loss (SNHL) is the most common form of hearing loss; it results from the degeneration of inner ear sensory hair cells and auditory neurons in the cochlea, cells that are terminally differentiated. Stem cell-and gene delivery-based strategies provide an opportunity for the replacement of these cells. In recent years, there has been an increasing interest in gene delivery to mesenchymal stem cells. In this study, we evaluated the potential of human umbilical cord mesenchymal stromal cells (hUCMSCs) as a possible source for regenerating inner ear hair cells. The expression of Atoh1 induced the differentiation of hUCMSCs into cells that resembled inner ear hair cells morphologically and immunocytochemically, evidenced by the expression of hair cell-specific markers. The results demonstrated for the first time that hUCMSCs can differentiate into hair cell-like cells, thus introducing a new potential tissue engineering and cell transplantation approach for the treatment of hearing loss. PMID:23379581
Wu, Fang; Wuensch, Sherry A.; Azadniv, Mitra; Ebrahimkhani, Mohammad R.; Crispe, I. Nicholas
We aim to define the role of Kupffer cells in intrahepatic antigen presentation, using the selective delivery of antigen to Kupffer cells rather than other populations of liver antigen-presenting cells. To achieve this we developed a novel antigen delivery system that can target antigens to macrophages, based on a galactosylated low-density lipoprotein nano-scale platform. Antigen was delivered via the galactose particle receptor (GPr), internalized, degraded and presented to T cells. The con...
We describe biodegradable mesoporous hybrid NPs in the presence of proteins, and its application for drug delivery. We synthesized oxamide-phenylene-based mesoporous organosilica nanoparticles (MON) in the absence of silica source which had a remarkably high organic content with a high surface area. Oxamide functions provided biodegradability in the presence of trypsin model proteins. MON displayed exceptionally high payloads of hydrophilic and hydrophobic drugs (up to 84 wt%), and a unique zero premature leakage without the pore capping, unlike mesoporous silica. MON were biocompatible and internalized into cancer cells for drug delivery.
Oba, Makoto; Kato, Takuma; Furukawa, Kaori; Tanaka, Masakazu
A peptide composed of lysine with a guanidinylethyl (GEt) amine structure in the side chain [Lys(GEt)] was developed as a cell-penetrating peptide directed to plasmid DNA (pDNA) delivery. The GEt amine adopted a diprotonated form at neutral pH, which may have led to the more efficient cellular uptake of a Lys(GEt)-peptide than an arginine-peptide at a low concentration. Lys(GEt)-peptide/pDNA complexes showed the highest transfection efficiency due to efficient endosomal escape without any cytotoxicity. Lys(GEt)-peptide may be a promising candidate as a gene delivery carrier.
Nina A Mikirova, Joseph J Casciari, Ronald E Hunninghake, Margaret M Beezley
Being overweight or obese is associated with an increased risk for the development of non-insulin-dependent diabetes mellitus, hypertension, and cardiovascular disease. Dyslipidemia of obesity is characterized by elevated fasting triglycerides and decreased high-density lipoprotein-cholesterol concentrations. Endothelial damage and dysfunction is considered to be a major underlying mechanism for the elevated cardiovascular risk associated with increased adiposity. Alterations in endothelial c...
Su, Jing; Chen, Feng; Cryns, Vincent L.; Messersmith, Phillip B.
A novel cell-targeting, pH-sensitive polymeric carrier was employed in this study for delivery of the anticancer drug bortezomib (BTZ) to cancer cells. Our strategy is based on facile conjugation of BTZ to catechol-containing polymeric carriers that are designed to be taken up selectively by cancer cells through cell surface receptor-mediated mechanisms. The polymer used as a building block in this study was poly(ethylene glycol), which was chosen for its ability to reduce nonspecific interac...
Orive, Gorka; Cobos, Raquel; Gorriti, Janire; Pedraz, Jose L; Meregalli, Mirella; Torrente, Yvan
In the last few years several technologies are being developed for eventually repairing or replacing damaged or injured tissues and even organs. Some of these emerging technologies include the design and development of new biomaterials, the optimization of nano- and micro-technologies for drug and cell delivery, the use of autologous proteins or the application of stem cells as therapeutics. Thus, several types of stem cells, e.g. ESCs, iPSCs, MSCs, CD133+ stem cells are being evaluated for tissue regeneration purposes. The present review describes some of these emerging technologies and discusses their potential benefits and challenges. PMID:25934974
Chemoresistance is a major obstacle in cancer treatment. Targeted therapies that enhance cancer cell sensitivity to chemotherapeutic agents have the potential to increase drug efficacy while reducing toxic effects on untargeted cells. Targeted cancer therapy by RNA interference (RNAi) is a relatively new approach that can be used to reversibly silence genes in vivo by selectively targeting genes such as the epidermal growth factor receptor (EGFR), which has been shown to increase the sensitivity of cancer cells to taxane chemotherapy. However, delivery represents the main hurdle for the broad development of RNAi therapeutics. We report here the use of core/shell hydrogel nanoparticles (nanogels) functionalized with peptides that specially target the EphA2 receptor to deliver small interfering RNAs (siRNAs) targeting EGFR. Expression of EGFR was determined by immunoblotting, and the effect of decreased EGFR expression on chemosensitization of ovarian cancer cells after siRNA delivery was investigated. Treatment of EphA2 positive Hey cells with siRNA-loaded, peptide-targeted nanogels decreased EGFR expression levels and significantly increased the sensitivity of this cell line to docetaxel (P < 0.05). Nanogel treatment of SK-OV-3 cells, which are negative for EphA2 expression, failed to reduce EGFR levels and did not increase docetaxel sensitivity (P > 0.05). This study suggests that targeted delivery of siRNAs by nanogels may be a promising strategy to increase the efficacy of chemotherapy drugs for the treatment of ovarian cancer. In addition, EphA2 is a viable target for therapeutic delivery, and the siRNAs are effectively protected by the nanogel carrier, overcoming the poor stability and uptake that has hindered clinical advancement of therapeutic siRNAs
Nwokeoha, Sandra; Carlisle, Robert; Cleveland, Robin O
The delivery of genes into cells through the transfer of ribonucleic acids (RNAs) has been found to cause a change in the level of target protein expression. RNA-based transfection is conceptually more efficient than commonly delivered plasmid DNA because it does not require division or damage of the nuclear envelope, thereby increasing the chances of the cell remaining viable. Shock waves (SWs) have been found to induce cellular uptake by transiently altering the permeability of the plasma membrane, thereby overcoming a critical step in gene therapy. However, accompanying SW bio-effects include dose-dependent irreversible cell injury and cytotoxicity. Here, the effect of SWs generated by a clinical lithotripter on the viability and permeabilisation of three different cell lines in vitro was investigated. Comparison of RNA stability before and after SW exposure revealed no statistically significant difference. Optimal SW exposure parameters were identified to minimise cell death and maximise permeabilisation, and applied to enhanced green fluorescent protein (eGFP) messenger RNA (mRNA) or anti-eGFP small interfering RNA delivery. As a result, eGFP mRNA expression levels increased up to 52-fold in CT26 cells, whereas a 2-fold decrease in GFP expression was achieved after anti-eGFP small interfering RNA delivery to MCF-7/GFP cells. These results indicate that SW parameters can be employed to achieve effective nucleotide delivery, laying the foundation for non-invasive and high-tolerability RNA-based gene therapy. PMID:27444864
Nahire, Rahul Rajaram
Lipid and polymeric nanoparticles, although proven to be effective drug delivery systems compared to free drugs, have shown considerable limitations pertaining to their uptake and release at tumor sites. Spatial and temporal control over the delivery of anticancer drugs has always been challenge to drug delivery scientists. Here, we have developed and characterized multifunctional nanoparticles (liposomes and polymersomes) which are targeted specifically to cancer cells, and release their contents with tumor specific internal triggers. To enable these nanoparticles to be tracked in blood circulation, we have imparted them with echogenic characteristic. Echogenicity of nanoparticles is evaluated using ultrasound scattering and imaging experiments. Nanoparticles demonstrated effective release with internal triggers such as elevated levels of MMP-9 enzyme found in the extracellular matrix of tumor cells, decreased pH of lysosome, and differential concentration of reducing agents in cytosol of cancer cells. We have also successfully demonstrated the sensitivity of these particles towards ultrasound to further enhance the release with internal triggers. To ensure the selective uptake by folate receptor- overexpressing cancer cells, we decorated these nanoparticles with folic acid on their surface. Fluorescence microscopic images showed significantly higher uptake of folate-targeted nanoparticles by MCF-7 (breast cancer) and PANC-1 (pancreatic cancer) cells compared to particles without any targeting ligand on their surface. To demonstrate the effectiveness of these nanoparticles to carry the drugs inside and kill cancer cells, we encapsulated doxorubicin and/or gemcitabine employing the pH gradient method. Drug loaded nanoparticles showed significantly higher killing of the cancer cells compared to their non-targeted counterparts and free drugs. With further development, these nanoparticles certainly have potential to be used as a multifunctional nanocarriers for image
Full Text Available Samuel Eisenstein,1 Shu-Hsia Chen,2 Ping-Ying Pan21Department of Surgery, 2Department of Oncological Sciences and Tisch Cancer Institute, The Icahn School of Medicine at Mount Sinai, New York, NY, USAAbstract: Oncolytic virotherapy on its own has numerous drawbacks, including an inability of the virus to actively target tumor cells and systemic toxicities at the high doses necessary to effectively treat tumors. Addition of immune cell-based carriers of oncolytic viruses holds promise as a technique in which oncolytic virus can be delivered directly to tumors in smaller and less toxic doses. Interestingly, the cell carriers themselves have also demonstrated antitumor effects, which can be augmented further by tailoring the appropriate oncolytic virus to the appropriate cell type. This review discusses the multiple factors that go into devising an effective, cell-based delivery system for oncolytic viruses.Keywords: oncolytic virus, cell carrier, immune cells, cancer therapy, myeloid-derived suppressor cells
Beloor, Jagadish; Ramakrishna, Suresh; Nam, Kihoon; Seon Choi, Chang; Kim, Jongkil; Kim, Sung Hwa; Cho, Hyong Jin; Shin, HeungSoo; Kim, Hyongbum; Kim, Sung Wan; Lee, Sang-Kyung; Kumar, Priti
Stem cells are poorly permissive to non-viral gene transfection reagents. In this study, we explored the possibility of improving gene delivery into human embryonic (hESC) and mesenchymal (hMSC) stem cells by synergizing the activity of a cell-binding ligand with a polymer that releases nucleic acids in a cytoplasm-responsive manner. A 29 amino acid long peptide, RVG, targeting the nicotinic acetylcholine receptor (nAchR) was identified to bind both hMSC and H9-derived hESC. Conjugating RVG to a redox-sensitive biodegradable dendrimer-type arginine-grafted polymer (PAM-ABP) enabled nanoparticle formation with plasmid DNA without altering the environment-sensitive DNA release property and favorable toxicity profile of the parent polymer. Importantly, RVG-PAM-ABP quantitatively enhanced transfection into both hMSC and hESC compared to commercial transfection reagents like Lipofectamine 2000 and Fugene. ∼60% and 50% of hMSC and hESC were respectively transfected, and at increased levels on a per cell basis, without affecting pluripotency marker expression. RVG-PAM-ABP is thus a novel bioreducible, biocompatible, non-toxic, synthetic gene delivery system for nAchR-expressing stem cells. Our data also demonstrates that a cell-binding ligand like RVG can cooperate with a gene delivery system like PAM-ABP to enable transfection of poorly-permissive cells. PMID:25515928
Wu, Yu-Wen; Goubran, Hadi; Seghatchian, Jerard; Burnouf, Thierry
Therapeutic and diagnostic applications of nanomedicine are playing increasingly important roles in human health. Various types of synthetic nanoparticles, including liposomes, micelles, and other nanotherapeutic platforms and conjugates, are being engineered to encapsulate or carry drugs for treating diseases such as cancer, cardiovascular disorders, neurodegeneration, and inflammations. Nanocarriers are designed to increase the half-life of drugs, decrease their toxicity and, ideally, target pathological sites. Developing smart carriers with the capacity to deliver drugs specifically to the microenvironment of diseased cells with minimum systemic toxicity is the goal. Blood cells, and potentially also the liposome-like micro- and nano-vesicles they generate, may be regarded as ideally suited to perform such specific targeting with minimum immunogenic risks. Blood cell membranes are "decorated" with complex physiological receptors capable of targeting and communicating with other cells and tissues and delivering their content to the surrounding pathological microenvironment. Blood cells, such as erythrocytes, have been developed as permeable carriers to release drugs to diseased tissues or act as biofactory allowing enzymatic degradation of a pathological substrate. Interestingly, attempts are also being made to improve the targeting capacity of synthetic nanoparticles by "decorating" their surface with blood cell membrane receptor-like biochemical structures. Research is needed to further explore the benefits that blood cell-derived microvesicles, as a Trojan horse delivery systems, can bring to the arsenal of therapeutic micro- and nanotechnologies. This short review focuses on the therapeutic roles that red blood cells and platelets can play as smart drug-delivery systems, and highlights the benefits that blood transfusion expertise can bring to this exciting and novel biomedical engineering field. PMID:27179926
Yu, Meihua; Jambhrunkar, Siddharth; Thorn, Peter; Chen, Jiezhong; Gu, Wenyi; Yu, Chengzhong
In this paper, a targeted drug delivery system has been developed based on hyaluronic acid (HA) modified mesoporous silica nanoparticles (MSNs). HA-MSNs possess a specific affinity to CD44 over-expressed on the surface of a specific cancer cell line, HCT-116 (human colon cancer cells). The cellular uptake performance of fluorescently labelled MSNs with and without HA modification has been evaluated by confocal microscopy and fluorescence-activated cell sorter (FACS) analysis. Compared to bare MSNs, HA-MSNs exhibit a higher cellular uptake via HA receptor mediated endocytosis. An anticancer drug, doxorubicin hydrochloride (Dox), has been loaded into MSNs and HA-MSNs as drug delivery vehicles. Dox loaded HA-MSNs show greater cytotoxicity to HCT-116 cells than free Dox and Dox-MSNs due to the enhanced cell internalization behavior of HA-MSNs. It is expected that HA-MSNs have a great potential in targeted delivery of anticancer drugs to CD44 over-expressing tumors.
Full Text Available Cancer has arisen to be of the most prominent health care issues across the world in recent years. Doctors have used physiological intervention as well as chemical and radioactive therapeutics to treat cancer thus far. As an alternative to current methods, gene delivery systems with high efficiency, specificity, and safety that can reduce side effects such as necrosis of tissue are under development. Although viral vectors are highly efficient, concerns have arisen from the fact that viral vectors are sourced from lethal diseases. With this in mind, rod shaped nano-materials such as carbon nanotubes (CNTs have become an attractive option for drug delivery due to the enhanced permeability and retention effect in tumors as well as the ability to penetrate the cell membrane. Here, we successfully engineered poly (lactic-co-glycolic (PLGA functionalized CNTs to reduce toxicity concerns, provide attachment sites for pro-apoptotic protein caspase-3 (CP3, and tune the temporal release profile of CP3 within bone cancer cells. Our results showed that CP3 was able to attach to functionalized CNTs, forming CNT-PLGA-CP3 conjugates. We show this conjugate can efficiently transduce cells at dosages as low as 0.05 μg/ml and suppress cell proliferation up to a week with no further treatments. These results are essential to showing the capabilities of PLGA functionalized CNTs as a non-viral vector gene delivery technique to tune cell fate.
Brown, M Rowan; Hondow, Nicole; Brydson, Rik; Rees, Paul; Brown, Andrew P; Summers, Huw D
The application of nanoparticles (NPs) within medicine is of great interest; their innate physicochemical characteristics provide the potential to enhance current technology, diagnostics and therapeutics. Recently a number of NP-based diagnostic and therapeutic agents have been developed for treatment of various diseases, where judicious surface functionalization is exploited to increase efficacy of administered therapeutic dose. However, quantification of heterogeneity associated with absolute dose of a nanotherapeutic (NP number), how this is trafficked across biological barriers has proven difficult to achieve. The main issue being the quantitative assessment of NP number at the spatial scale of the individual NP, data which is essential for the continued growth and development of the next generation of nanotherapeutics. Recent advances in sample preparation and the imaging fidelity of transmission electron microscopy (TEM) platforms provide information at the required spatial scale, where individual NPs can be individually identified. High spatial resolution however reduces the sample frequency and as a result dynamic biological features or processes become opaque. However, the combination of TEM data with appropriate probabilistic models provide a means to extract biophysical information that imaging alone cannot. Previously, we demonstrated that limited cell sampling via TEM can be statistically coupled to large population flow cytometry measurements to quantify exact NP dose. Here we extended this concept to link TEM measurements of NP agglomerates in cell culture media to that encapsulated within vesicles in human osteosarcoma cells. By construction and validation of a data-driven transfer function, we are able to investigate the dynamic properties of NP agglomeration through endocytosis. In particular, we statistically predict how NP agglomerates may traverse a biological barrier, detailing inter-agglomerate merging events providing the basis for
Full Text Available Nanotechnology has been integrated into healthcare system in terms of diagnosis as well as therapy. The massive impact of imaging nanotechnology has a deeper intervention in cardiology i.e. as contrast agents , to target vulnerable plaques with site specificity and in a theranostic approach to treat these plaques, stem cell delivery in necrotic myocardium, etc. Thus cardiovascular nanoimaging is not limited to simple diagnosis but also can help real time tracking during therapy as well as surgery. The present review provides a comprehensive description of the molecular imaging techniques for cardiovascular diseases with the help of nanotechnology and the potential clinical implications of nanotechnology for future applications.
Short communication: Effect of commercial or depurinized milk diet on plasma advanced oxidation protein products, cardiovascular markers, and bone marrow CD34+ stem cell potential in rat experimental hyperuricemia.
Kocic, Gordana; Sokolovic, Dusan; Jevtovic, Tatjana; Cvetkovic, Tatjana; Veljkovic, Andrej; Kocic, Hristina; Stojanovic, Svetlana; Jovanovic, Aneta; Jovanovic, Jelena; Zivkovic, Petar
Cardiovascular repair and myocardial contractility may be improved by migration of bone marrow stem cells (BMSC) and their delivery to the site of injury, a process known as BMSC homing. The aim of our study was to examine the dietary effect of a newly patented depurinized milk (DP) that is almost free of uric acid and purine and pyrimidine compounds compared with a standard commercial 1.5% fat UHT milk diet or allopurinol therapy in rat experimental hyperuricemia. Bone marrow stem cell potential (BMCD34(+), CD34-postive bone marrow cells), plasma oxidative stress parameters [advanced oxidation protein products, AOPP) and thiobarbituric acid reactive substances (TBARS)], myocardial damage markers [creatine phosphokinase (CPK), aspartate aminotransferase (AST), and lactate dehydrogenase (LDH)], plasma cholesterol, and high-density lipoprotein cholesterol were investigated. The DP milk diet significantly increased the number of BMCD34(+) stem cells compared with commercial UHT milk. Allopurinol given alone also increased the number of BMCD34(+). Hyperuricemia caused a significant increase in all plasma enzyme markers for myocardial damage (CPK, LDH, and AST). A cardioprotective effect was achieved with allopurinol but almost equally with DP milk and more than with commercial milk. Regarding plasma AOPP, TBARS, and cholesterol levels, the most effective treatment was DP milk. In conclusion, the protective role of a milk diet on cardiovascular function may be enhanced through the new depurinized milk diet, which may improve cardiovascular system function via increased bone marrow stem cell regenerative potential, decreased plasma oxidative stress parameters, and decreased levels of myocardial damage markers and cholesterol. New dairy technology strategies focused on eliminating harmful milk compounds should be completely nontoxic. Novel milk products should be tested for their ability to improve tissue repair and function. PMID:25218755
Das, Soumyashree; Yu, Shiyan; Sakamori, Ryotaro; Vedula, Pavan; Feng, Qiang; Flores, Juan; Hoffman, Andrew; Fu, Jiang; Stypulkowski, Ewa; Rodriguez, Alexis; Dobrowolski, Radek; Harada, Akihiro; Hsu, Wei; Bonder, Edward M; Verzi, Michael P; Gao, Nan
Communication between stem and niche supporting cells maintains the homeostasis of adult tissues. Wnt signaling is a crucial regulator of the stem cell niche, but the mechanism that governs Wnt ligand delivery in this compartment has not been fully investigated. We identified that Wnt secretion is partly dependent on Rab8a-mediated anterograde transport of Gpr177 (wntless), a Wnt-specific transmembrane transporter. Gpr177 binds to Rab8a, depletion of which compromises Gpr177 traffic, thereby weakening the secretion of multiple Wnts. Analyses of generic Wnt/β-catenin targets in Rab8a knockout mouse intestinal crypts indicate reduced signaling activities; maturation of Paneth cells - a Wnt-dependent cell type - is severely affected. Rab8a knockout crypts show an expansion of Lgr5(+) and Hopx(+) cells in vivo. However, in vitro, the knockout enteroids exhibit significantly weakened growth that can be partly restored by exogenous Wnts or Gsk3β inhibitors. Immunogold labeling and surface protein isolation identified decreased plasma membrane localization of Gpr177 in Rab8a knockout Paneth cells and fibroblasts. Upon stimulation by exogenous Wnts, Rab8a-deficient cells show ligand-induced Lrp6 phosphorylation and transcriptional reporter activation. Rab8a thus controls Wnt delivery in producing cells and is crucial for Paneth cell maturation. Our data highlight the profound tissue plasticity that occurs in response to stress induced by depletion of a stem cell niche signal. PMID:26015543
Full Text Available Superparamagnetic nanoparticles are promising candidates for gene delivery into mammalian somatic cells and may be useful for reproductive cloning using the somatic cell nuclear transfer technique. However, limited investigations of their potential applications in animal genetics and breeding, particularly multiple-gene delivery by magnetofection, have been performed. Here, we developed a stable, targetable and convenient system for delivering multiple genes into the nuclei of porcine somatic cells using magnetic Fe3O4 nanoparticles as gene carriers. After surface modification by polyethylenimine, the spherical magnetic Fe3O4 nanoparticles showed strong binding affinity for DNA plasmids expressing the genes encoding a green (DNAGFP or red (DNADsRed fluorescent protein. At weight ratios of DNAGFP or DNADsRed to magnetic nanoparticles lower than or equal to 10∶1 or 5∶1, respectively, the DNA molecules were completely bound by the magnetic nanoparticles. Atomic force microscopy analyses confirmed binding of the spherical magnetic nanoparticles to stretched DNA strands up to several hundred nanometers in length. As a result, stable and efficient co-expression of GFP and DsRed in porcine kidney PK-15 cells was achieved by magnetofection. The results presented here demonstrate the potential application of magnetic nanoparticles as an attractive delivery system for animal genetics and breeding studies.
Clements, Başak Açan; Hsu, Charlie Y M; Kucharski, Cezary; Lin, Xiaoyue; Rose, Laura; Uludağ, Hasan
Basic fibroblast growth factor (bFGF) is capable of stimulating osteogenic differentiation of preosteoblast cells in vitro and new bone tissue deposition in vivo. Delivering the gene for the protein, rather than the protein itself, is considered advantageous for bone repair since gene delivery obviates the need to produce the protein in pharmaceutical quantities. To explore the feasibility of bFGF gene delivery by nonviral methods, we transfected primary rat bone marrow stromal cells (BMSC) using cationic polymers (polyethylenimine and poly(L-lysine)-palmitic acid) in vitro. After delivering a bFGF-expression plasmid (pFGF2-IRES-AcGFP) to BMSC, the presence of bFGF in culture supernatants was detected by a commercial ELISA. As much as 0.3 ng bFGF/10(6) cells/day was obtained from the BMSC under optimal conditions. This secretion rate was approximately 100-fold lower than the secretion obtained from immortal, and easy-to-transfect, human 293T cells. These data suggest the feasibility of modifying BMSC with nonviral delivery systems for bFGF expression, but also highlight the need for substantial improvement in transfection rate for an effective therapy. PMID:19495899
Bellini, Michela; Mazzucchelli, Serena; Galbiati, Elisabetta; Sommaruga, Silvia; Fiandra, Luisa; Truffi, Marta; Rizzuto, Maria A; Colombo, Miriam; Tortora, Paolo; Corsi, Fabio; Prosperi, Davide
A genetically engineered apoferritin variant consisting of 24 heavy-chain subunits (HFn) was produced to achieve a cumulative delivery of an antitumor drug, which exerts its cytotoxic action by targeting the DNA at the nucleus of human cancer cells with subcellular precision. The rationale of our approach is based on exploiting the natural arsenal of defense of cancer cells to stimulate them to recruit large amounts of HFn nanoparticles loaded with doxorubicin inside their nucleus in response to a DNA damage, which leads to a programmed cell death. After demonstrating the selectivity of HFn for representative cancer cells compared to healthy fibroblasts, doxorubicin-loaded HFn was used to treat the cancer cells. The results from confocal microscopy and DNA damage assays proved that loading of doxorubicin in HFn nanoparticles increased the nuclear delivery of the drug, thus enhancing doxorubicin efficacy. Doxorubicin-loaded HFn acts as a "Trojan Horse": HFn was internalized in cancer cells faster and more efficiently compared to free doxorubicin, then promptly translocated into the nucleus following the DNA damage caused by the partial release in the cytoplasm of encapsulated doxorubicin. This self-triggered translocation mechanism allowed the drug to be directly released in the nuclear compartment, where it exerted its toxic action. This approach was reliable and straightforward providing an antiproliferative effect with high reproducibility. The particular self-assembling nature of HFn nanocage makes it a versatile and tunable nanovector for a broad range of molecules suitable both for detection and treatment of cancer cells. PMID:25312541
Wan, Long; Jiao, Jian; Cui, Yu; Guo, Jingwen; Han, Ning; Di, Donghua; Chang, Di; Wang, Pu; Jiang, Tongying; Wang, Siling
In this paper, hyaluronic acid (HA) functionalized uniform mesoporous carbon spheres (UMCS) were synthesized for targeted enzyme responsive drug delivery using a facile electrostatic attraction strategy. This HA modification ensured stable drug encapsulation in mesoporous carbon nanoparticles in an extracellular environment while increasing colloidal stability, biocompatibility, cell-targeting ability, and controlled cargo release. The cellular uptake experiments of fluorescently labeled mesoporous carbon nanoparticles, with or without HA functionalization, demonstrated that HA-UMCS are able to specifically target cancer cells overexpressing CD44 receptors. Moreover, the cargo loaded doxorubicin (DOX) and verapamil (VER) exhibited a dual pH and hyaluronidase-1 responsive release in the tumor microenvironment. In addition, VER/DOX/HA-UMCS exhibited a superior therapeutic effect on an in vivo HCT-116 tumor in BALB/c nude mice. In summary, it is expected that HA-UMCS will offer a new method for targeted co-delivery of drugs to tumors overexpressing CD44 receptors.
state to get a good pharmacological performance. Recombinant antibody engineering with non-natural amino acids, or enzyme-mediated conjugation approaches (transglutaminase, Sortase A or endoglycosidase) have been reported for producing homogeneous antibody conjugates. However, these methods require......-templated organic synthesis due to the wide existence of the 3-histidine cluster in most wild-type proteins. In this thesis, three projects that relate to targeted drug delivery to cancer cells based on the DTPC method is described. The first project was a delivery system which uses transferrin as the targeting...... ligand and saporin (ribosome inactivating protein) as the warhead to achieve enhanced cellular uptake and cytotoxicity of saporin to transferrin receptor overexpressed cancer cell line. The transferrin-saporin conjugate complex are formed by linking the site-selective DNA-transferrin conjugates with mono...
Huang, Sha; Fu, Xiaobing
The objective of regenerative medicine is to provide cells with a local environment of artificial extracellular matrix where they can proliferate and differentiate efficiently and therefore, induce the repair of defective tissues according to the natural healing potential of patients. For this purpose, naturally derived materials are being widely used because of their similarities to the extracellular matrix, typically good biocharacteristics and inherent cellular interaction. Also, natural polymers can be engineered to release growth factors and related agents in response to physiologic signals to imitate the natural healing process and to promote fast tissue regeneration and reduce scarring in wounds. Although synthetic materials have been used extensively in tissue engineering fields, this review illustrates the contribution of natural materials and natural materials-based protein delivery systems to regenerative medicine research, with emphasis on the application of multifunctional vehicles for cell and growth factor delivery in skin regeneration research. PMID:19850093
Kato, Takuma; Yamashita, Hiroko; Misawa, Takashi; Nishida, Koyo; Kurihara, Masaaki; Tanaka, Masakazu; Demizu, Yosuke; Oba, Makoto
Cell-penetrating peptides (CPPs) have been developed as drug, protein, and gene delivery tools. In the present study, arginine (Arg)-rich CPPs containing unnatural amino acids were designed to deliver plasmid DNA (pDNA). The transfection ability of one of the Arg-rich CPPs examined here was more effective than that of the Arg nonapeptide, which is the most frequently used CPP. The transfection efficiencies of Arg-rich CPPs increased with longer post-incubation times and were significantly higher at 48-h and 72-h post-incubation than that of the commercially available transfection reagent TurboFect. These Arg-rich CPPs were complexed with pDNA for a long time in cells and effectively escaped from the late endosomes/lysosomes into the cytoplasm. These results will be helpful for designing novel CPPs for pDNA delivery. PMID:27132868
Hoban, Megan D; Romero, Zulema; Cost, Gregory J; Mendel, Matthew; Holmes, Michael; Kohn, Donald B
This unit describes the protocol for the delivery of reagents for targeted genome editing to CD34(+) hematopoietic stem/progenitor cells (HSPCs). Specifically, this unit focuses on the process of thawing and pre-stimulating CD34(+) HSPCs, as well as the details of their electroporation with in vitro-transcribed mRNA-encoding site-specific nucleases [in this case zinc-finger nucleases (ZFNs)]. In addition, discussed is delivery of a gene editing donor template in the form of an oligonucleotide or integrase-defective lentiviral vector (IDLV). Finally, an analysis of cell survival following treatment and downstream culture conditions are presented. While optimization steps might be needed for each specific application with respect to nuclease and donor template amount, adherence to this protocol will serve as an excellent starting point for this further work. PMID:26840227
Yang, Ningning; Ye, Zhaoyang; Li, Feng; Mahato, Ram I.
The objective was to determine whether bioconjugation of type I collagen specific triplex forming oligonucleotide (TFO) to N-(2-hydroxypropyl) methacrylamide (HPMA) containing tetrapeptide Gly-Phe-Leu-Gly (GFLG) and mannose 6-phosphate (M6P) can provide their targeted delivery to hepatic stellate cells (HSCs). Following bioconjugation, M6P-GFLG-HPMA-GFLG-32P-TFO was characterized by PAGE, HPLC and GPC, and then its biodistribution was determined. TFO was dissociated from the conjugate when in...
Gitanjali Sharma; Sushant Lakkadwala; Amit Modgil; Jagdish Singh
The challenge of effectively delivering therapeutic agents to brain has led to an entire field of active research devoted to overcome the blood brain barrier (BBB) and efficiently deliver drugs to brain. This review focusses on exploring the facets of a novel platform designed for the delivery of drugs to brain. The platform was constructed based on the hypothesis that a combination of receptor-targeting agent, like transferrin protein, and a cell-penetrating peptide (CPP) will enhance the de...
Shahidan, Nur Nabilah
This thesis present a study of thermo-gelling polycaprolactone (PCL) dispersion which consist of a mixture of PCL microspheres (MSs) and thermo-responsive, graft cationic copolymer. The PCL microspheres are solid or colloidosomes (hollow). This study aims toward an injectable colloidal cell delivery system. The thermo-responsive copolymer used in this study is a new family of cationic graft copolymer. The cationic graft copolymer consisted of cationic poly(N,N-dimethylaminoethyl methacrylate)...
Kadonosono, Tetsuya; Yamano, Akihiro; Goto, Toshiki; Tsubaki, Takuya; Niibori, Mizuho; Kuchimaru, Takahiro; Kizaka-Kondoh, Shinae
Cell-penetrating peptides (CPPs), also referred to as protein transduction domains (PTDs), can mediate the cellular uptake of a wide range of macromolecules including peptides, proteins, oligonucleotides, and nanoparticles, and thus have received considerable attention as a promising method for drug delivery in vivo. Here, we report that CPP/PTDs facilitate the extravasation of fused proteins by binding to neuropilin-1 (NRP1), a vascular endothelial growth factor (VEGF) co-receptor expressed on the surface of endothelial and some tumor cells. In this study, we examined the capacity of the amphipathic and cationic CPP/PTDs, PTD-3 and TAT-PTD, respectively, to bind cells in vitro and accumulate in xenograft tumors in vivo. Notably, these functions were significantly suppressed by pre-treatment with NRP1-neutralizing Ab. Furthermore, co-injection of iRGD, a cyclic peptide known to increase NRP1-dependent vascular permeability, significantly reduced CPP/PTD tumor delivery. This data demonstrates a mechanism by which NRP1 promotes the extravasation of CPP/PTDs that may open new avenues for the development of more efficient CPP/PTD delivery systems. PMID:25592386
Inge L. Huibregtse
Full Text Available Interleukin-10 (IL-10 plays an indispensable role in mucosal tolerance by programming dendritic cells (DCs to induce suppressor Th-cells. We have tested the modulating effect of L. lactis secreting human IL-10 (L. lactisIL-10 on DC function in vitro. Monocyte-derived DC incubated with L. lactisIL-10 induced effector Th-cells that markedly suppressed the proliferation of allogenic Th-cells as compared to L. lactis. This suppressive effect was only seen when DC showed increased CD83 and CD86 expression. Furthermore, enhanced production of IL-10 was measured in both L. lactisIL-10-derived DC and Th-cells compared to L. lactis-derived DC and Th-cells. Neutralizing IL-10 during DC-Th-cell interaction and coculturing L. lactisIL-10-derived suppressor Th-cells with allogenic Th-cells in a transwell system prevented the induction of suppressor Th-cells. Only 130 pg/mL of bacterial-derived IL-10 and 40 times more exogenously added recombinant human IL-10 were needed during DC priming for the generation of suppressor Th-cells. The spatially restricted delivery of IL-10 by food-grade bacteria is a promising strategy to induce suppressor Th-cells in vivo and to treat inflammatory diseases.
Xiao-Kang Zheng; Long-Hua Chen; Xiao Yan; Hong-Mei Wang
AIM: To explore the impact of prolonged fraction dosedelivery time modeling intensity-modulated radiation therapy (IMRT) on cell killing of human hepatocellular carcinoma (HCC) HepG2 and Hep3B cell lines.METHODS: The radiobiological characteristics of human HCC HepG2 and Hep3b cell lines were studied with standard clonogenic assays, using standard linear-quadratic model and incomplete repair model to fit the dose-survival curves. The identical methods were also employed to investigate the biological effectiveness of irradiation protocols modeling clinical conventional fractionated external beam radiotherapy (EBRT, fraction delivery time 3 min) and IMRT with different prolonged fraction delivery time (15, 30, and 45 min). The differences of cell surviving fraction irradiated with different fraction delivery time were tested with paired t-test. Factors determining the impact of prolonged fraction delivery time on cell killing were analyzed.RESULTS: The α/β and repair half-time (T1/2) of HepG2and Hep3b were 3.1 and 7.4 Gy, and 22 and 19 min respectively. The surviving fraction of HepG2 irradiated modeling IMRT with different fraction delivery time was significantly higher than irradiated modeling EBRT and the cell survival increased more pronouncedly with the fraction delivery time prolonged from 15 to 45 min,while no significant differences of cell survival in Hep3b were found between different fraction delivery time protocols.CONCLUSION: The prolonged fraction delivery time modeling IMRT significantly decreased the cell killing in HepG2 but not in Hep3b. The capability of sub-lethal damage repair was the predominant factor determining the cell killing decrease. These effects, if confirmed by clinical studies, should be considered in designing IMRT treatments for HCC.
Song, Yanyan; Lou, Bo; Zhao, Peng; Lin, Chao
A folate-decorated, disulfide-based cationic dextran conjugate having dextran as the main chain and disulfide-linked 1,4-bis(3-aminopropyl)piperazine (BAP) residues as the grafts was designed and successfully prepared as a multifunctional gene delivery vector for targeted gene delivery to ovarian cancer SKOV-3 cells in vitro and in vivo. Initially, a new bioreducible cationic polyamide (denoted as pSSBAP) was prepared by polycondensation reaction of bis(p-nitrophenyl)-3,3'-dithiodipropanoate, a disulfide-containing monomer, and BAP. It was found that the pSSBAP was highly efficient for in vitro gene delivery against MCF-7 and SKOV-3 cell lines. Subsequently, two cationic dextran conjugates with different amounts of BAP residues (denoted as Dex-SSBAP6 and Dex-SSBAP30, respectively) were synthesized by coupling BAP to disulfide-linked carboxylated dextran or coupling pSSBAP-oligomer to p-nitrophenyl carbonated dextran. Both two conjugates were able to bind DNA to form nanosized polyplexes with an improved colloidal stability in physiological conditions. The polyplexes, however, were rapidly dissociated to liberate DNA in a reducing environment. In vitro transfection experiments revealed that the polyplexes of Dex-SSBAP30 efficiently transfected SKOV-3 cells, yielding transfection efficiency that is comparable to that of linear polyethylenimine or lipofectamine 2000. AlamarBlue assay showed that the conjugates had low cytotoxicity in vitro at a high concentration of 100 mg/L. Further, Dex-SSBAP30 has primary amine side groups and thus allows for folate (FA) conjugation, yielding FA-coupled Dex-SSBAP30 (Dex-SSBAP30-FA). It was found that Dex-SSBAP30-FA was efficient for targeted gene delivery to SKOV-3 tumor xenografted in a nude mouse model by intravenous injection, inducing a higher level of gene expression in the tumor as compared to Dex-SSBAP30 lacking FA and comparable gene expression to linear polyethylenimine as one of the most efficient polymeric vectors for
Bobba, Samantha; Di Girolamo, Nick
: Worldwide, 45 million people are blind. Corneal blindness is a major cause of visual loss, estimated to affect 10 million. For the most difficult to treat patients, including those with a disease called limbal stem cell deficiency, a donor corneal graft is not a viable option; thus, patients are treated with specialized stem cell grafts, which fail in a significant proportion (30 to 50%) of subjects. This unacceptable failure rate means there is a pressing need to develop minimally invasive, long-lasting, cost-effective therapies to improve patient quality of life and lessen the economic burden. Restoring vision in patients with severe corneal disease is the main focus of our research program; however, to achieve our goals and deliver the best quality stem cell therapy, we must first understand the basic biology of these cells, including their residence, the factors that support their long-term existence, markers to identify and isolate them, and carriers that facilitate expansion, delivery, and protection during engraftment. We recently achieved some of these goals through the discovery of stem cell markers and the development of a novel and innovative contact lens-based cell transfer technique that has been successfully trialed on patients with corneal blindness. Although several popular methodologies are currently available to nurture and transfer stem cells to the patients' ocular surface, contact lenses provide many advantages that will be discussed in this review article. The job for clinician-researchers will be to map precisely how these cells contribute to restoring ocular health and whether improvements in the quality of cells and the cell delivery system can be developed to reduce disease burden. PMID:26390346
In our previous work, it was found that acoustic cavitation might play a role in improving the cell permeability to microparticles when liposomes were used in an in vitro experiment. The purpose of this project is to expand our study and to learn other possible mechanisms by which cells may interact with liposomes under ultrasound (US) excitation and become transiently permeable to microparticles. It is further hypothesized that two possible scenarios may be involved in in vitro experiments: (1) drug-carrying liposomes transiently overcome the cell membrane barrier and enter into a cell while the cell is still viable; (2) the liposomes incorporate with a cell at its membrane through a fusing process. To prove this hypothesis, liposomes of two different structures were synthesized: one has fluorescent molecules encapsulated into liposomes and the other has fluorescent markers incorporated into the shells of liposomes. Liposomes of each kind were mixed with human breast cancer cells (MCF7-cell line) in a suspension at 5 (liposomes) : 1 (cell) ratio and were then exposed to a focused 1 MHz ultrasound beam at its focal region for 40 s. The US signal contained 20 cycles per tone-burst at a pulse-repetition-frequency of 10 kHz; the spatial peak acoustic pressure amplitude was 0.25 MPa. It was found that the possible mechanisms might include the acoustic cavitation, the endocytosis and cell-fusion. Acoustic radiation force might make liposomes collide with cells effectively and facilitate the delivery process
Annexin A5, a 35KDa protein, specifically binds with high affinity to phosphatidylserine (P.S.) which is actively redistributed to the external leaflet of plasmic membranes in apoptotic cells and activated platelets. Annexin A5 radiolabelled with 99mTc(99mTc-ANX5) was developed by Strauss (stanford, Usa) to image apoptosis in vivo: tumours cells apoptosis induced by chemo-radiotherapy, ischemia/reperfusion lesions in animals and patients, graft rejection. Additionally, many in vitro data suggest that annexin A5 also stains necrosis (membrane disruption), which occurs in all types of cell death. This preclinical work aimed to evaluate the potential interest of 99mTc-ANX5 imaging as a clinical tool in cardiovascular diseases. Four studies performed in rat models of myocardial infarction by coronary ligation and ischemia-reperfusion, and in rat models of subacute and acute (isoproterenol-induced) myocarditis show the ability of 99mTc-ANX5 to detect in vivo cardio myocytes death by apoptosis and necrosis. Another study demonstrates that 99mTc-ANX5 is highly accurate to evaluate in vivo the biological activity of parietal thrombus in a rat model of elastase-induced abdominal aortic aneurysm. These results suggest that 99mTc-ANX5 imaging could be used in patients for non invasive diagnosis, prognostic evaluation in acute myocarditis and in various thrombotic cardiovascular diseases. (author)
Piepoli, Massimo F.; Corra, Ugo; Adamopoulos, Stamatis; Benzer, Werner; Bjarnason-Wehrens, Birna; Cupples, Margaret; Dendale, Paul; Doherty, Patrick; Gaita, Dan; Hofer, Stefan; McGee, Hannah; Mendes, Miguel; Niebauer, Josef; Pogosova, Nana; Garcia-Porrero, Esteban
Despite major improvements in diagnostics and interventional therapies, cardiovascular diseases remain a major health care and socio-economic burden both in western and developing countries, in which this burden is increasing in close correlation to economic growth. Health authorities and the general population have started to recognize that the fight against these diseases can only be won if their burden is faced by increasing our investment on interventions in lifestyle changes and preventi...
Li, Ruijing; Chen, Kexin; Li, Geng; Han, Guoxiang; Yu, Sheng; Yao, Juming; Cai, Yurong
Porous microspheres fabricated from bioceramics have great potential for cell delivery in injectable tissue engineering application. The size and structure of pores in the microspheres are important for the effective protection and transportation of cells. In this study, porous hydroxyapatite microspheres are fabricated through the water-in-oil emulsion method followed by a calcination treatment at the high temperature. Both self-made resorcinol-formaldehyde (RF) composite spheres and camphene are used as pore-forming agents to produce big pores corresponding to the size of RF spheres and connected channel among big pores in hydroxyapatite matrix. The properties of the microspheres are characterized using X-ray diffraction, thermogravimetry analysis, universal material machine, field emission scanning electron microscopy. Cell assays are carried out to evaluate the cellular compatibility of the microspheres. The results showed that the hydroxyapatite microspheres with controllable pore structure and high porosity could be fabricated by this method, which have better strength to resist the compressive force. The microspheres are conducive to support adhesion, proliferation and differentiation of MC3T3-E1 cells. The results indicate that the obtained porous hydroxyapatite microspheres can be a permeable microenvironment for cell delivery in injectable tissue engineering.
Gissberg, Olof; Zaghloul, Eman M; Lundin, Karin E; Nguyen, Chi-Hung; Landras-Guetta, Corinne; Wengel, Jesper; Zain, Rula; Smith, C I Edvard
AS1411 is a g-quadruplex-forming aptamer capable of selectively entering cancer cells by nucleolin receptor-mediated uptake. In this study, we investigated the cell internalization properties and plasticity of AS1411 carrying different locked nucleic acid-containing cargo oligonucleotides (ONs) for delivery into A549 and U2OS cells. We found that internalization efficiency is highly governed by ON cargo chemistry and composition since the inherent antitumor properties of AS1411 were lost when attached to a nontoxic ON, noTox. However, a toxic ON, Tox, demonstrated potent cytotoxicity after aptamer-mediated uptake in A549 cells. We also examined the effect of unlocked nucleic acid (UNA) modifications in the loop region of the aptamer, and how the cargo ONs and UNA incorporation affect the secondary structure of AS1411, in the presence or absence of two novel ellipticine derivatives. These findings add new insights to the design and future applications of aptamer-guided delivery of ON cargo to cancer cells. PMID:26859550
Kong, Hyun Joon; Liu, Jodi; Riddle, Kathryn; Matsumoto, Takuya; Leach, Kent; Mooney, David J.
Non-viral gene vectors are commonly used for gene therapy owing to safety concerns with viral vectors. However, non-viral vectors are plagued by low levels of gene transfection and cellular expression. Current efforts to improve the efficiency of non-viral gene delivery are focused on manipulations of the delivery vector, whereas the influence of the cellular environment in DNA uptake is often ignored. The mechanical properties (for example, rigidity) of the substrate to which a cell adheres have been found to mediate many aspects of cell function including proliferation, migration and differentiation, and this suggests that the mechanics of the adhesion substrate may regulate a cell's ability to uptake exogeneous signalling molecules. In this report, we present a critical role for the rigidity of the cell adhesion substrate on the level of gene transfer and expression. The mechanism relates to material control over cell proliferation, and was investigated using a fluorescent resonance energy transfer (FRET) technique. This study provides a new material-based control point for non-viral gene therapy.
Kristensen, Mie; Nielsen, Hanne Mørck
Oral delivery of biopharmaceuticals, for example peptides and proteins, constitutes a great challenge in drug delivery due to their low chemical stability and poor permeation across the intestinal mucosa, to a large extent limiting the mode of administration to injections, which is not favouring patient compliance. Nevertheless, cell-penetrating peptides (CPPs) have shown promising potential as carriers to overcome the epithelium, and this minireview highlights recent knowledge gained within the field of CPP-mediated transepithelial delivery of therapeutic peptides and proteins from the intestine. Two approaches may be pursued: co-administration of the carrier and therapeutic peptide in the form of complexes obtained by simple bulk mixing, or administration of covalent conjugates demanding more advanced production methodologies. These formulation approaches have their pros and cons, and which is to be preferred depends on the physicochemical properties of both the specific CPP and the specific cargo. In addition to the physical epithelial barrier, a metabolic barrier must be overcome in order to obtain CPP-mediated delivery of a cargo drug from the intestine, and a number of strategies have been employed to delay enzymatic degradation of the CPP. The mechanisms by which CPPs translocate across membranes are not fully understood, but possibly involve endocytosis as well as direct translocation, and the CPP-mediated transepithelial delivery of cargo drugs thus likely involves similar mechanisms for the initial membrane interaction and translocation. However, the mechanisms responsible for transcytosis of the cargo drug, if taken up by an endocytic mechanism, or direct translocation across the epithelium are so far not known. PMID:26525297
Frenz, Theresa; Grabski, Elena; Durán, Verónica; Hozsa, Constantin; Stępczyńska, Anna; Furch, Marcus; Gieseler, Robert K; Kalinke, Ulrich
Targeted drug delivery systems hold promise for selective provision of active compounds to distinct tissues or cell subsets. Thus, locally enhanced drug concentrations are obtained that would confer improved efficacy. As a consequence adverse effects should be diminished, as innocent bystander cells are less affected. Currently, several controlled drug delivery systems based on diverse materials are being developed. Some systems exhibit material-associated toxic effects and/or show low drug loading capacity. In contrast, liposomal nanocarriers are particularly favorable because they are well tolerated, poorly immunogenic, can be produced in defined sizes, and offer a reasonable payload capacity. Compared with other immune cells, professional antigen-presenting cells (APCs) demonstrate enhanced liposome uptake mediated by macropinocytosis, phagocytosis and presumably also by clathrin- and caveolae-mediated endocytosis. In order to further enhance the targeting efficacy toward APCs, receptor-mediated uptake appears advisable. Since APC subsets generally do not express single linage-specific receptors, members of the C-type lectin receptor (CLR) family are compelling targets. Examples of CLR expressed by APCs include DEC-205 (CD205) expressed by myeloid dendritic cells (DC) and monocytes, the mannose receptor C type 1 (MR, CD206) expressed by DC, monocytes and macrophages, DC-SIGN (CD209) expressed by DC, and several others. These receptors bind glycans, which are typically displayed by pathogens and thus support pathogen uptake and endocytosis. Further research will elucidate whether glycan-decorated liposomes will not only enhance APCs targeting but also enable preferential delivery of their payload to discrete subcellular compartments. PMID:25701806
Lopes, Ivo; C N Oliveira, Ana; P Sárria, Marisa; P Neves Silva, João; Gonçalves, Odete; Gomes, Andreia C; Real Oliveira, Maria Elisabete C D
We report the development and characterization of a novel nanometric system for specific delivery of therapeutic siRNA for cancer treatment. This vector is based on a binary mixture of the cationic surfactant dioctadecyldimethylammonium chloride (DODAC) and the helper lipid monoolein (MO). These liposomes were previously validated by our research group as promising non-viral vectors for nucleic acid delivery. In this work, the DODAC:MO vesicles were for the first time functionalized with polyethylene glycol and PEG-folate conjugates to achieve both maximal stability in biological fluids and increase selectivity toward folate receptor α expressing cells. The produced DODAC:MO:PEG liposomes were highly effective in RNA complexation (close to 100%), and the resulting lipoplexes also demonstrated high stability in conditions simulating their administration by intravenous injection (physiological pH, high NaCl, heparin and fetal bovine serum concentrations). In addition, cell uptake of the PEG-folate-coated lipoplexes was significantly greater in folate receptor α positive breast cancer cells (39% for 25 µg/mL of lipid and 31% for 40 µg/mL) when compared with folate receptor α negative cells (31% for 25 µg/mL of lipid and 23% for 40 µg/mL) and to systems without PEG-folate (≈13% to 16% for all tested conditions), supporting their selectivity towards the receptor. Overall, the results support these systems as appealing vectors for selective delivery of siRNA to cancer cells by folate receptor α-mediated internalization, aiming at future therapeutic applications of interest. PMID:26340109
Full Text Available BACKGROUND: Gene therapy has tremendous potential for both inherited and acquired diseases. However, delivery problems limited their clinical application, and new gene delivery vehicles with low cytotoxicity and high transfection efficiency are greatly required. METHODS: In this report, we designed and synthesized three amphiphilic molecules (L1-L3 with the structures involving 1, 4, 7, 10-tetraazacyclododecane (cyclen, imidazolium and a hydrophobic dodecyl chain. Their interactions with plasmid DNA were studied via electrophoretic gel retardation assays, fluorescent quenching experiments, dynamic light scattering and transmission electron microscopy. The in vitro gene transfection assay and cytotoxicity assay were conducted in four cell lines. RESULTS: Results indicated that L1 and L3-formed liposomes could effectively bind to DNA to form well-shaped nanoparticles. Combining with neutral lipid DOPE, L3 was found with high efficiency in gene transfer in three tumor cell lines including A549, HepG2 and H460. The optimized gene transfection efficacy of L3 was nearly 5.5 times more efficient than that of the popular commercially available gene delivery agent Lipofectamine 2000™ in human lung carcinoma cells A549. In addition, since L1 and L3 had nearly no gene transfection performance in normal cells HEK293, these cationic lipids showed tumor cell-targeting property to a certain extent. No significant cytotoxicity was found for the lipoplexes formed by L1-L3, and their cytotoxicities were similar to or slightly lower than the lipoplexes prepared from Lipofectamine 2000™. CONCLUSION: Novel cyclen-based cationic lipids for effective in vitro gene transfection were founded, and these studies here may extend the application areas of macrocyclic polyamines, especially for cyclen.
Chi, Lianhua; Na, Moon-Hee; Jung, Hyun-Kyung; Vadevoo, Sri Murugan Poongkavithai; Kim, Cheong-Wun; Padmanaban, Guruprasath; Park, Tae-In; Park, Jae-Yong; Hwang, Ilseon; Park, Keon Uk; Liang, Frank; Lu, Maggie; Park, Jiho; Kim, In-San; Lee, Byung-Heon
A growing body of evidence suggests that pathological lesions express tissue-specific molecular targets or biomarkers within the tissue. Interleukin-4 receptor (IL-4R) is overexpressed in many types of cancer cells, including lung cancer. Here we investigated the properties of IL-4R-binding peptide-1 (IL4RPep-1), a CRKRLDRNC peptide, and its ability to target the delivery of liposomes to lung tumor. IL4RPep-1 preferentially bound to H226 lung tumor cells which express higher levers of IL-4R compared to H460 lung tumor cells which express less IL-4R. Mutational analysis revealed that C1, R2, and R4 residues of IL4RPep-1 were the key binding determinants. IL4RPep-1-labeled liposomes containing doxorubicin were more efficiently internalized in H226 cells and effectively delivered doxorubicin into the cells compared to unlabeled liposomes. In vivo fluorescence imaging of nude mice subcutaneously xenotransplanted with H226 tumor cells indicated that IL4RPep-1-labeled liposomes accumulate more efficiently in the tumor and inhibit tumor growth more effectively compared to unlabeled liposomes. Interestingly, expression of IL-4R was high in vascular endothelial cells of tumor, while little was detected in vascular endothelial cells of control organs including the liver. IL-4R expression in cultured human vascular endothelial cells was also up-regulated when activated by a pro-inflammatory cytokine tumor necrosis factor-α. Moreover, the up-regulation of IL-4R expression was observed in primary human lung cancer tissues. These results indicate that IL-4R-targeting nanocarriers may be a useful strategy to enhance drug delivery through the recognition of IL-4R in both tumor cells and tumor endothelial cells. PMID:25979323
Aires, Antonio; Ocampo, Sandra M; Simões, Bruno M; Josefa Rodríguez, María; Cadenas, Jael F; Couleaud, Pierre; Spence, Katherine; Latorre, Alfonso; Miranda, Rodolfo; Somoza, Álvaro; Clarke, Robert B; Carrascosa, José L; Cortajarena, Aitziber L
Nanomedicine nowadays offers novel solutions in cancer therapy and diagnosis by introducing multimodal treatments and imaging tools in one single formulation. Nanoparticles acting as nanocarriers change the solubility, biodistribution and efficiency of therapeutic molecules, reducing their side effects. In order to successfully apply these novel therapeutic approaches, efforts are focused on the biological functionalization of the nanoparticles to improve the selectivity towards cancer cells. In this work, we present the synthesis and characterization of novel multifunctionalized iron oxide magnetic nanoparticles (MNPs) with antiCD44 antibody and gemcitabine derivatives, and their application for the selective treatment of CD44-positive cancer cells. The lymphocyte homing receptor CD44 is overexpressed in a large variety of cancer cells, but also in cancer stem cells (CSCs) and circulating tumor cells (CTCs). Therefore, targeting CD44-overexpressing cells is a challenging and promising anticancer strategy. Firstly, we demonstrate the targeting of antiCD44 functionalized MNPs to different CD44-positive cancer cell lines using a CD44-negative non-tumorigenic cell line as a control, and verify the specificity by ultrastructural characterization and downregulation of CD44 expression. Finally, we show the selective drug delivery potential of the MNPs by the killing of CD44-positive cancer cells using a CD44-negative non-tumorigenic cell line as a control. In conclusion, the proposed multifunctionalized MNPs represent an excellent biocompatible nanoplatform for selective CD44-positive cancer therapy in vitro. PMID:26754042
Aires, Antonio; Ocampo, Sandra M.; Simões, Bruno M.; Josefa Rodríguez, María; Cadenas, Jael F.; Couleaud, Pierre; Spence, Katherine; Latorre, Alfonso; Miranda, Rodolfo; Somoza, Álvaro; Clarke, Robert B.; Carrascosa, José L.; Cortajarena, Aitziber L.
Nanomedicine nowadays offers novel solutions in cancer therapy and diagnosis by introducing multimodal treatments and imaging tools in one single formulation. Nanoparticles acting as nanocarriers change the solubility, biodistribution and efficiency of therapeutic molecules, reducing their side effects. In order to successfully apply these novel therapeutic approaches, efforts are focused on the biological functionalization of the nanoparticles to improve the selectivity towards cancer cells. In this work, we present the synthesis and characterization of novel multifunctionalized iron oxide magnetic nanoparticles (MNPs) with antiCD44 antibody and gemcitabine derivatives, and their application for the selective treatment of CD44-positive cancer cells. The lymphocyte homing receptor CD44 is overexpressed in a large variety of cancer cells, but also in cancer stem cells (CSCs) and circulating tumor cells (CTCs). Therefore, targeting CD44-overexpressing cells is a challenging and promising anticancer strategy. Firstly, we demonstrate the targeting of antiCD44 functionalized MNPs to different CD44-positive cancer cell lines using a CD44-negative non-tumorigenic cell line as a control, and verify the specificity by ultrastructural characterization and downregulation of CD44 expression. Finally, we show the selective drug delivery potential of the MNPs by the killing of CD44-positive cancer cells using a CD44-negative non-tumorigenic cell line as a control. In conclusion, the proposed multifunctionalized MNPs represent an excellent biocompatible nanoplatform for selective CD44-positive cancer therapy in vitro.
Jensen, Frank Bo
Vertebrate red blood cells (RBCs) seem to serve tissue oxygen delivery in two distinct ways. Firstly, RBCs enable the adequate transport of O2 between respiratory surfaces and metabolizing tissues by means of their high intracellular concentration of hemoglobin (Hb), appropriate allosteric...... interactions between Hb ligand-binding sites, and an adjustable intracellular chemical environment that allows fine-tuning of Hb O2 affinity. Secondly, RBCs may sense tissue O2 requirements via their degree of deoxygenation when they travel through the microcirculation and release vasodilatory compounds that...... enhance blood flow in hypoxic tissues. This latter function could be important in matching tissue O2 delivery with local O2 demand. Three main mechanisms by which RBCs can regulate their own distribution in the microcirculation have been proposed. These are: (1) deoxygenation-dependent release of ATP from...
Soni, Namrata; Soni, Neetu; Pandey, Himanshu; Maheshwari, Rahul; Kesharwani, Prashant; Tekade, Rakesh Kumar
Gemcitabine (GmcH) is an effective anti-cancer agent used in the chemotherapy of lung cancer. However, the clinical applications of GmcH has been impeded primarily due to its low blood residence time, unfavorable pharmacokinetic and pharmacodynamic (PK/PD) profile, and poor penetration in the complex environment of lung cancer cells. Thus, the present study aims to formulate GmcH loaded mannosylated solid lipid nanoparticles (GmcH-SLNs) for improving its drug uptake into the lung cancer cells. GmcH-SLNs were prepared by emulsification and solvent evaporation process, and surface modification was done with mannose using ring opening technique. The cellular toxicity and cell uptake studies were performed in A549 lung adenocarcinoma cell line. The developed nanoformulation appears to be proficient in targeted delivery of GmcH with improved therapeutic effectiveness and enhanced safety. PMID:27459173
McDonald, Michael A.; Spurlin, Tighe A.; Tona, Alessandro; Elliott, John T.; Halter, Michael; Plant, Anne L.
This paper presents preliminary results on the use of transferrin protein nanospheres (TfpNS) for targeting cancer cells in vitro. Protein nanospheres represent an easily prepared and modifiable nanoplatform for receptor-specific targeting, molecular imaging and gene delivery. Rhodamine B isothiocyanate conjugated TfpNS (RBITC-TfpNS) show significantly enhanced uptake in vitro in SK-MEL-28 human malignant melanoma cells known to overexpress transferrin receptors compared to controls. RBITCTfpNS labeling of the cancer cells is due to transferrin receptor-mediated uptake, as demonstrated by competitive inhibition with native transferrin. Initial fluorescence microscopy studies indicate GFP plasmid can be transfected into melanoma cells via GFP plasmid encapsulated by TfpNS.
Full Text Available PURPOSE: Bone marrow stem cell therapy is a new, attractive therapeutic approach for treatment of intervertebral disc (IVD degeneration; however, leakage and backflow of transplanted cells into the structures surrounding the disc may lead to the formation of undesirable osteophytes. The purpose of this study was to develop a technique for minimally invasive and accurate delivery of stem cells. METHODS: Porcine mesenchymal stem cells (MSCs were labeled with superparamagnetic iron oxide nanoparticles (SPIO, Molday ION rhodamine and first injected into the explanted swine lumbar IVD, followed by ex vivo 3T MRI. After having determined sufficient sensitivity, IVD degeneration was then induced in swine (n=3 by laser-evaporation. 3 x 10(6 SPIO-labeled cells embedded within hydrogel were injected in 2 doses using a transcutaneous cannula and an epidural anesthesia catheter. T2-weighted MR images were obtained at 3T before and immediately after cell infusion. Two weeks after injection, histological examination was performed for detection of transplanted cells. RESULTS: MSCs were efficiently labeled with Molday ION rhodamine. Cells could be readily detected in the injected vertebral tissue explants as distinct hypointensities with sufficient sensitivity. MR monitoring indicated that the MSCs were successfully delivered into the IVD in vivo, which was confirmed by iron-positive Prussian Blue staining of the tissue within the IVD. CONCLUSION: We have developed a technique for non-invasive monitoring of minimally invasive stem delivery into the IVD at 3T. By using a large animal model mimicking the anatomy of IVD in humans, the present results indicate that this procedure may be clinically feasible.
Li, Jing; Ma, Fang-Kui; Dang, Qi-Feng; Liang, Xing-Guo; Chen, Xi-Guang
A novel targeted drug delivery system, glucose-conjugated chitosan nanoparticles (GCNPs), was developed for specific recognition and interaction with glucose transporters (Gluts) over-expressed by tumor cells. GC was synthesized by using succinic acid as a linker between glucosamine and chitosan (CS), and successful synthesis was confirmed by NMR and elemental analysis. GCNPs were prepared by ionic crosslinking method, and characterized in terms of morphology, size, and zeta potential. The optimally prepared nanoparticles showed spherical shapes with an average particle size of (187.9 ± 3.8) nm and a zeta potential of (- 15.43 ± 0.31) mV. The GCNPs showed negligible cytotoxicity to mouse embryo fibroblast and 4T1 cells. Doxorubicin (DOX) could be efficiently entrapped into GCNPs, with a loading capacity and encapsulation efficiency of 20.11% and 64.81%, respectively. DOX-loaded nanoparticles exhibited sustained-release behavior in phosphate buffered saline (pH 7.4). In vitro cellular uptake studies showed that the GCNPs had better endocytosis ability than CSNPs, and the antitumor activity of DOX/GCNPs was 4-5 times effectiveness in 4T1 cell killing than that of DOX/CSNPs. All the results demonstrate that nanoparticles decorated with glucose have specific interactions with cancer cells via the recognition between glucose and Gluts. Therefore, Gluts-targeted GCNPs may be promising delivery agents in cancer therapies.
Zhou, Peixun; Hatziieremia, Sophia; Elliott, Moira A; Scobie, Linda; Crossan, Claire; Michie, Alison M; Holyoake, Tessa L; Halbert, Gavin W; Jørgensen, Heather G
Chronic Myeloid Leukemia (CML) stem/progenitor cells, which over-express Bcr-Abl, respond to imatinib by a reversible block in proliferation without significant apoptosis. As a result, patients are unlikely to be cured owing to the persistence of leukemic quiescent stem cells (QSC) capable of initiating relapse. Previously, we have reported that intracellular levels of imatinib in primary primitive CML cells (CD34+38(lo/⁻)), are significantly lower than in CML progenitor cells (total CD34+) and leukemic cell lines. The aim of this study was to determine if potentially sub-therapeutic intracellular drug concentrations in persistent leukemic QSC may be overcome by targeted drug delivery using synthetic Low Density Lipoprotein (sLDL) particles. As a first step towards this goal, however, the extent of uptake of sLDL by leukemic cell lines and CML patient stem/progenitor cells was investigated. Results with non-drug loaded particles have shown an increased and preferential uptake of sLDL by Bcr-Abl positive cell lines in comparison to Bcr-Abl negative. Furthermore, CML CD34+ and primitive CD34+38(lo/⁻) cells accumulated significantly higher levels of sLDL when compared with non-CML CD34+ cells. Thus, drug-loading the sLDL nanoparticles could potentially enhance intracellular drug concentrations in primitive CML cells and thus aid their eradication. PMID:20869412
Shock waves have non-destructive life science applications in cell transfection and drug delivery. Based on molecular dynamics simulations, the shockwave causes transient compression of the cell membrane, which causes the hydrophobic interior of the lipid bilayer to become thinner. This allows diffusion of water molecules across the membrane. Recently, the nanothermite composition consisting of CuO nanorods and Al nanoparticles was shown to propagate at velocities in the same range as metallic azides and fulminates; however, the CuO/Al mixture produces lower pressure levels. An in vitro testing system was developed to deliver shock waves produced by nanothermites into cell suspensions and/or tissues. The plasmid encoded for production of green-fluorescent protein was delivered into cells including, among other types, chicken cardiomyocytes, cell lines (T47-D and Ins-1), and Arabidopsis plant cells. It was found that the nanothermite pressure impulses induced transfection resulting in production of green fluorescent protein in 99% of the cardiomyocytes. Additionally, transfected cell survival was evaluated, and the pressure impulses did not produce any elevated levels of cell death compared with control cell suspensions.
Lei, Pedro; Padmashali, Roshan M; Andreadis, Stelios T
We investigated fibrin-mediated gene transfer by embedding pDNA within the hydrogel during polymerization and using two modes of gene transfection with cells placed either on the surface (2D transfection) or within the hydrogel (3D transfection). Using this model, we found that cell transfection depended strongly on the local cell-pDNA microenvironment as defined by the 2D vs. 3D context, target cell type and density, as well as fibrinogen and pDNA concentrations. When cells were embedded within the fibrin matrix lipofectamine-induced cell death decreased significantly, especially at low target cell density. Addition of fibrinolytic inhibitors decreased gene transfer in a dose-dependent manner, suggesting that fibrin degradation may be necessary for efficient gene transfer. We also provided proof-of-concept that fibrin-mediated gene transfer can be used for spatially localized gene delivery, which is required in cell-transfection microarrays. When lipoplex-containing hydrogels were spotted in an array format gene transfer was strictly confined to pDNA-containing fibrin spots with no cross-contamination between neighboring sites. Collectively, our data suggest that fibrin may be used as a biomaterial to deliver genes in an efficient, cell-controlled and spatially localized manner for potential applications in vitro or in vivo. PMID:19395019
Seras Franzoso, Joaquin; Peebo, Karl; Garcia Fruitós, Elena; Vázquez Gómez, Esther; Rinas, Ursula; Villaverde Corrales, Antonio
Altres ajuts: We are indebted CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN, Spain) for funding our research on inclusion bodies. Bacterial inclusion bodies (IBs) have recently been used to generate biocompatible cell culture interfaces, with diverse effects on cultured cells such as cell adhesion enhancement, stimulation of cell growth or induction of mesenchymal stem cell differentiation. Additionally, novel applications of IBs as sustained protein delivery systems with...
Full Text Available Margherita Maioli,1–3 Giovanni Contini,1 Sara Santaniello,1,2 Pasquale Bandiera,1 Gianfranco Pigliaru,1,2 Raimonda Sanna,5 Salvatore Rinaldi,3 Alessandro P Delitala,1 Andrea Montella,1,5 Luigi Bagella,1,6 Carlo Ventura2–41Department of Biomedical Sciences, University of Sassari, Sassari, 2Laboratory of Molecular Biology and Stem Cell Engineering, National Institute of Biostructures and Biosystems, Bologna, 3Department of Regenerative Medicine, Rinaldi Fontani Institute, Florence, 4Cardiovascular Department, S Orsola-Malpighi Hospital, University of Bologna, Bologna, 5Facility of Genetic and Developmental Biology, AOU Sassari, Sassari, Italy; 6Sbarro Institute for Cancer Research and Molecular Medicine, Center for Biotechnology, College of Science and Technology, Temple University, Philadelphia, PA, USAAbstract: Mouse embryonic stem cells were previously observed along with mesenchymal stem cells from different sources, after being treated with a mixed ester of hyaluronan with butyric and retinoic acids, to show a significant increase in the yield of cardiogenic and vascular differentiated elements. The aim of the present study was to determine if stem cells derived from primitive fetal cells present in human amniotic fluid (hAFSCs and cultured in the presence of a mixture of hyaluronic (HA, butyric (BU, and retinoic (RA acids show a higher yield of differentiation toward the cardiovascular phenotype as compared with untreated cells. During the differentiation process elicited by exposure to HA + BU + RA, genes controlling pluripotency and plasticity of stem cells, such as Sox2, Nanog, and Oct4, were significantly downregulated at the transcriptional level. At this point, a significant increase in expression of genes controlling the appearance of cardiogenic and vascular lineages in HA + BU + RA-treated cells was observed. The protein expression levels typical of cardiac and vascular phenotypes, evaluated by Western blotting
Yue, Jianhui; Wu, Jun; Liu, Di; Zhao, Xiaoli; Lu, William W.
Nanotechnology has made a significant impact on the development of nanomedicine. Nonviral vectors have been attracting more attention for the advantage of biosafety in gene delivery. Polyethylenimine (PEI)-conjugated chitosan (chitosan-g-PEI) emerged as a promising nonviral vector and has been demonstrated in many tumor cells. However, there is a lack of study focused on the behavior of this vector in stem cells which hold great potential in regenerative medicine. Therefore, in this study, in vitro gene delivering effect of chitosan-g-PEI was investigated in bone marrow stem cells. pIRES2-ZsGreen1-hBMP2 dual expression plasmid containing both the ZsGreen1 GFP reporter gene and the BMP2 functional gene was constructed for monitoring the transgene expression level. Chitosan-g-PEI-mediated gene transfer showed 17.2% of transfection efficiency and more than 80% of cell viability in stem cells. These values were higher than that of PEI. The expression of the delivered BMP2 gene in stem cells enhanced the osteogenic differentiation. These results demonstrated that chitosan-g-PEI is capable of applying in delivering gene to stem cells and providing potential applications in stem cell-based gene therapy.
We have fabricated an electric cell fusion chip with an embedded cell delivery function driven by surface tension and evaluated its performance with several types of plant cells. The chip consists of a polydimethylsiloxane-based microchannel with a fusion chamber and gold–titanium (Au–Ti) electrodes. The velocity profiles of the microfluid in the channel and fusion chamber were calculated to predict cell movement, and the electric field distribution between the electrodes was also calculated in order to determine the appropriate electrode shape. The range of the fluid velocity in the fusion chamber is 20–50 µm s−1 and the measured speed of the cells is approximately 45 µm s−1, which is sufficiently slow for the motion of the cells in the fusion chamber to be monitored and controlled. We measured the variation of the pearl chain ratio with frequency for five kinds of plant cells, and determined that the optimal frequency for pearl chain formation is 1.5 MHz. The electrofusion of cells was successfully carried out under ac field (amplitude: 0.4–0.5 kV cm−1, frequency: 1.5 MHz) and dc pulse (amplitude: 1.0 kV cm−1, duration: 20 ms) conditions
Power, Anthony T; Bell, John C
Recent years have seen tremendous advances in the development of exquisitely targeted replicating virotherapeutics that can safely destroy malignant cells. Despite this promise, clinical advancement of this powerful and unique approach has been hindered by vulnerability to host defenses and inefficient systemic delivery. However, it now appears that delivery of oncolytic viruses within carrier cells may offer one solution to this critical problem. In this review, we compare the advantages and limitations of the numerous cell lineages that have been investigated as delivery platforms for viral therapeutics, and discuss examples showing how combined cell-virus biotherapeutics can be used to achieve synergistic gains in antitumor activity. Finally, we highlight avenues for future preclinical research that might be taken in order to refine cell-virus biotherapeutics in preparation for human trials. PMID:17264852
Rabideau, Amy E; Pentelute, Bradley Lether
The intracellular delivery of peptide and protein therapeutics is a major challenge due to the plasma membrane, which acts as a barrier between the extracellular environment and the intracellular milieu. Over the past two decades, a nontoxic PA/LFN delivery platform derived from anthrax lethal toxin has been developed for the transport of non-native cargo into the cytosol of cells in order to understand the translocation process through a protective antigen (PA) pore and to probe intracellular biological functions. Enzyme-mediated ligation using sortase A and native chemical ligation are two facile methods used to synthesize these non-native conjugates, inaccessible by recombinant technology. Cargo molecules that translocate efficiently include enzymes from protein toxins, antibody mimic proteins, and peptides of varying lengths and non-natural amino acid compositions. The PA pore has been found to effectively convey over 30 known cargos other than native lethal factor (LF; i.e., non-native) with diverse sequences and functionalities on the LFN transporter protein. All together these studies demonstrated that non-native cargos must adopt an unfolded or extended conformation and contain a suitable charge composition in order to efficiently pass through the PA pore. This review provides insight into design parameters for the efficient delivery of new cargos using PA and LFN. PMID:27055654
Betts, Corinne A; Wood, Matthew J A
Duchenne muscular dystrophy is a severe, X-linked muscle wasting disorder caused by the absence of an integral structural protein called dystrophin. This is caused by mutations or deletions in the dystrophin gene which disrupt the reading frame, thereby halting the production of a functional protein. A number of potential therapies have been investigated for the treatment of this disease including utrophin upregulation, 'stop-codon read through' aminoglycosides and adeno-associated virus gene replacement as well as stem cell therapy. However, the most promising treatment to date is the use of antisense oligonucleotides which cause exon skipping by binding to a specific mRNA sequence, skipping the desired exon, thereby restoring the reading frame and producing a truncated yet functional protein. The results from recent 2'OMePS and morpholino clinical trials have renewed hope for Duchenne patients; however in vivo studies in a mouse model, mdx, have revealed low systemic distribution and poor delivery of oligonucleotides to affected tissues such as the brain and heart. However a variety of cell penetrating peptides directly conjugated to antisense oligonucleotides have been shown to enhance delivery in Duchenne model systems with improved systemic distribution and greater efficacy compared to 'naked' antisense oligonucleotides. These cell penetrating peptides, combined with an optimised dose and dosing regimen, as well as thorough toxicity profile have the potential to be developed into a promising treatment which may be progressed to clinical trial. PMID:23140454
Park, Kyung Min; Lee, Sang Young; Joung, Yoon Ki; Na, Jae Sik; Lee, Myung Chul; Park, Ki Dong
Injectable hydrogels have been studied for potential applications for articular cartilage regeneration. In this study, a thermosensitive chitosan-Pluronic (CP) hydrogel was designed as an injectable cell delivery carrier for cartilage regeneration. The CP conjugate was synthesized by grafting Pluronic onto chitosan using EDC/NHS chemistry. The sol-gel phase transition and mechanical properties of the CP hydrogel were examined by rheological experiments. The CP solution underwent a sol-gel transition around 25 degrees C at which the storage modulus (G') approaches 10(4)Pa, highlighting the potential of this material as an injectable scaffold for cartilage regeneration. The CP hydrogel was formed rapidly by increasing the temperature. The morphology of the dried CP hydrogel was observed by scanning electron microscopy. In vitro cell culture was performed using bovine chondrocytes. The proliferation of bovine chondrocytes and the amount of synthesized glycosaminoglycan increased for 28 days. These results suggested that the CP hydrogel has potential as an injectable cell delivery carrier for cartilage regeneration and could serve as a new biomaterial for tissue engineering. PMID:19261553
Full Text Available Abstract Background Central arterial stiffness represents a well-established predictor of cardiovascular disease. Decreased circulating endothelial progenitor cells (EPCs, increased asymmetric dimethyl-arginine (ADMA levels, traditional cardiovascular risk factors and insulin resistance have all been associated with increased arterial stiffness. The correlations of novel and traditional cardiovascular risk factors with central arterial stiffness in prediabetic individuals were investigated in the present study. Methods The study population consisted of 53 prediabetic individuals. Individuals were divided into groups of isolated impaired fasting glucose (IFG, isolated impaired glucose tolerance (IGT and combined IGT-IFG. Age, sex, family history of diabetes, smoking history, body mass index (BMI, waist to hip ratio (WHR, waist circumference (WC, blood pressure, lipid profile, levels of high sensitive C-reactive protein (hsCRP, glomerular filtration rate (GFR, and history of antihypertensive or statin therapy were obtained from all participants. Insulin resistance was evaluated using the Homeostatic Model Assessment (HOMA-IR. Carotid -femoral pulse wave velocity was used as an index of arterial stiffness. Circulating EPC count and ADMA serum levels were also determined. Results Among studied individuals 30 (56.6% subjects were diagnosed with isolated IFG, 9 (17% with isolated IGT (17% and 14 with combined IFG-IGT (26.4%. In univariate analysis age, mean blood pressure, fasting glucose, total cholesterol, LDL cholesterol, and ADMA levels positively correlated with pulse-wave velocity while exercise and GFR correlated negatively. EPC count did not correlate with PWV. In multivariate stepwise regression analysis PWV correlated independently and positively with LDL-Cholesterol (low density lipoprotein and ADMA levels and negatively with exercise. Conclusions Elevated ADMA and LDL-C levels are strongly associated with increased arterial stiffness among
Full Text Available Liang Zhao,1,* Hongdan Li,2,* Yijie Shi,1 Guan Wang,2 Liwei Liu,1 Chang Su,3 Rongjian Su2 1School of Pharmacy, Liaoning Medical University, Jinzhou, People’s Republic of China; 2Central Laboratory of Liaoning Medical University, Jinzhou, People’s Republic of China; 3School of Veterinary Medicine, Liaoning Medical University, Jinzhou, People’s Republic of China *These authors contributed equally to this work Abstract: Nanoparticles (NPs which target specific agents could effectively recognize the target cells and increase the stability of chemical agents by encapsulation. As such, NPs have been widely used in cancer treatment research. Recently, over 90% of treatment failure cases in patients with metastatic cancer were attributed to resistance to chemotherapy. Surface-exposed glucose-regulated protein of 78 kDa (GRP78 is expressed highly on many tumor cell surfaces in many human cancers and is related to the regulation of invasion and metastasis. Herein, we report that NPs conjugated with antibody against GRP78 (mAb GRP78-NPs inhibit the adhesion, invasion, and metastasis of hepatocellular carcinoma (HCC and promote drug delivery of 5-fluorouracil into GRP78 high-expressed human hepatocellular carcinoma cells. Our new findings suggest that mAb GRP78-NPs could enhance drug accumulation by effectively transporting NPs into cell surface GRP78-overexpressed human hepatocellular carcinoma cells and then inhibit cell proliferation and viability and induce cell apoptosis by regulating caspase-3. In brief, mAb GRP78-NPs effectively inhibit cancer cell invasion and enhance antitumor efficiency by targeted drug delivery. Keywords: 5-Fu, apoptosis, HCC, caspase-3
Ergün, Bahar; De Cola, Luisa; Galla, Hans-Joachim; Kehr, Nermin Seda
The alternating layer-by-layer deposition of oppositely charged polyelectrolytes on fluorescence-dye-(Hst)-loaded zeolites L ((Hst) Zeo-PSS/PLL) is described. The arrays and nanocomposite (NC) hydrogels of (Hst) Zeo-PSS/PLL are prepared. The subsequent cell experiments show the potential application of arrays and NC hydrogels of (Hst) Zeo-PSS/PLL as alternative 2D- and 3D-surfaces, respectively, for 2D- and 3D-surface-mediated controlled organic molecules delivery applications. PMID:27114067
Gao, Changyong; Wu, Zhiguang; Lin, Zhihua; Lin, Xiankun; He, Qiang
We report a biomimetic delivery of microsized capsule-cushioned leukocyte membrane vesicles (CLMVs) through the conversion of freshly reassembled leukocyte membrane vesicles (LMVs), including membrane lipids and membrane-bound proteins onto the surface of layer-by-layer assembled polymeric multilayer microcapsules. The leukocyte membrane coating was verified by using electron microscopy, a quartz crystal microbalance, dynamic light scattering, and confocal laser scanning microscopy. The resulting CLMVs have the ability to effectively evade clearance by the immune system and thus prolong the circulation time in mice. Moreover, we also show that the right-side-out leukocyte membrane coating can distinctly improve the accumulation of capsules in tumor sites through the molecular recognition of membrane-bound proteins of CLMVs with those of tumor cells in vitro and in vivo. The natural cell membrane camouflaged polymeric multilayer capsules with the immunosuppressive and tumor-recognition functionalities of natural leukocytes provide a new biomimetic delivery platform for disease therapy.We report a biomimetic delivery of microsized capsule-cushioned leukocyte membrane vesicles (CLMVs) through the conversion of freshly reassembled leukocyte membrane vesicles (LMVs), including membrane lipids and membrane-bound proteins onto the surface of layer-by-layer assembled polymeric multilayer microcapsules. The leukocyte membrane coating was verified by using electron microscopy, a quartz crystal microbalance, dynamic light scattering, and confocal laser scanning microscopy. The resulting CLMVs have the ability to effectively evade clearance by the immune system and thus prolong the circulation time in mice. Moreover, we also show that the right-side-out leukocyte membrane coating can distinctly improve the accumulation of capsules in tumor sites through the molecular recognition of membrane-bound proteins of CLMVs with those of tumor cells in vitro and in vivo. The natural
Nano-diamonds (NDs) are often considered as inert platforms with high interests for biomedical applications. They are well adapted for drug delivery, and may display embedded fluorescence. We report here on a new way to consider these nano-carbon particles, by revealing therapeutic capacities coming from electronic properties of NDs. With an optimized surface chemistry, the generation of Reactive Oxygen Species (ROS) occurs when those hydrogen-terminated NDs are exposed to photon irradiation, thus opening up the field towards the radiosensitization of tumor cells. (authors)
Full Text Available Jurja C Coyuco, Yuanjie Liu, Bee-Jen Tan, Gigi NC ChiuDepartment of Pharmacy, Faculty of Science, National University of Singapore, SingaporeAbstract: Although carbon nanomaterials (CNMs have been increasingly studied for their biomedical applications, there is limited research on these novel materials for oral drug delivery. As such, this study aimed to explore the potential of CNMs in oral drug delivery, and the objectives were to evaluate CNM cytotoxicity and their abilities to modulate paracellular transport and the P-glycoprotein (P-gp efflux pump. Three types of functionalized CNMs were studied, including polyhydroxy small-gap fullerenes (OH-fullerenes, carboxylic acid functionalized single-walled carbon nanotubes (fSWCNT-COOH and poly(ethylene glycol functionalized single-walled carbon nanotubes (fSWCNT-PEG, using the well-established Caco-2 cell monolayer to represent the intestinal epithelium. All three CNMs had minimum cytotoxicity on Caco-2 cells, as demonstrated through lactose dehydrogenase release and 3-(4,5-dimethyliazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assays. Of the three CNMs, fSWCNT-COOH significantly reduced transepithelial electrical resistance and enhanced transport of Lucifer Yellow across the Caco-2 monolayer. Confocal fluorescence microscopy showed that fSWCNT-COOH treated cells had the highest perturbation in the distribution of ZO-1, a protein marker of tight junction, suggesting that fSWCNT-COOH could enhance paracellular permeability via disruption of tight junctions. This modulating effect of fSWCNT-COOH can be reversed over time. Furthermore, cellular accumulation of the P-gp substrate, rhodamine-123, was significantly increased in cells treated with fSWCNT-COOH, suggestive of P-gp inhibition. Of note, fSWCNT-PEG could increase rhodamine-123 accumulation without modifying the tight junction. Collectively, these results suggest that the functionalized CNMs could be useful as modulators for oral drug
Water, Jorrit Jeroen; Smart, Simon; Franzyk, Henrik;
high plectasin encapsulation efficiency (71-90%) and mediated release of the peptide over 24h. The antimicrobial efficacy of the peptide-loaded nanoparticles was investigated using bronchiolar epithelial Calu-3 cell monolayers infected with S. aureus. The plectasin-loaded nanoparticles displayed......A number of pathogenic bacterial strains, such as Staphylococcus aureus, are difficult to kill with conventional antibiotics due to intracellular persistence in host airway epithelium. Designing drug delivery systems to deliver potent antimicrobial peptides (AMPs) intracellularly to the airway...
Messerschmidt, Sylvia K E; Musyanovych, Anna; Altvater, Martin; Scheurich, Peter; Pfizenmaier, Klaus; Landfester, Katharina; Kontermann, Roland E
Polymeric nanoparticles displaying tumor necrosis factor on their surface (TNF nanocytes) are useful carrier systems capable of mimicking the bioactivity of membrane-bound TNF. Thus, TNF nanocytes are potent activators of TNF receptor 1 and 2 leading to a striking enhancement of apoptosis. However, in vivo applications are hampered by potential systemic toxicity. Here, using TNF nanocytes as a model system, we developed a procedure to generate targeted lipid-coated particles (TLP) in which TNF activity is shielded. The TLPs generated here are composed of an inner single-chain TNF (scTNF)-functionalized, polymeric nanoparticle core surrounded by a lipid coat endowed with polyethylene glycol (PEG) for sterical stabilization and a single-chain Fv (scFv) fragment for targeting. Using a scFv directed against the tumor stroma marker fibroblast activation protein (FAP) we show that TLP and scTNF-TLP specifically bind to FAP-expressing, but not to FAP-negative cells. Lipid coating strongly reduced nonspecific binding of particles and scTNF-mediated cytotoxicity towards FAP-negative cells. In contrast, an increased cytotoxicity of TLP was observed for FAP-positive cells. Thus, through liposome encapsulation, nanoparticles carrying bioactive molecules, which are subject to nonselective uptake and activity towards various cells and tissues, can be converted into target cell-specific composite particles exhibiting a selective activity towards antigen-positive target cells. Besides safe and targeted delivery of death ligands such as TNF, TLP should be suitable for various diagnostic and therapeutic applications, which benefit from a targeted delivery of reagents embedded into the particle core or displayed on the core particle surface. PMID:19306900
Roy, D G; Power, A T; Bourgeois-Daigneault, M C; Falls, T; Ferreira, L; Stern, A; Tanese de Souza, C; McCart, J A; Stojdl, D F; Lichty, B D; Atkins, H; Auer, R C; Bell, J C; Le Boeuf, F
Due to cancer's genetic complexity, significant advances in the treatment of metastatic disease will require sophisticated, multi-pronged therapeutic approaches. Here we demonstrate the utility of a Drosophila melanogaster cell platform for the production and in vivo delivery of multi-gene biotherapeutic systems. We show that cultured Drosophila S2 cell carriers can stably propagate oncolytic viral therapeutics that are highly cytotoxic for mammalian cancer cells without adverse effects on insect cell viability or gene expression. Drosophila cell carriers administered systemically to immunocompetent animals trafficked to tumors to deliver multiple biotherapeutics with little apparent off-target tissue homing or toxicity, resulting in a therapeutic effect. Cells of this Dipteran invertebrate provide a genetically tractable platform supporting the integration of complex, multi-gene biotherapies while avoiding many of the barriers to systemic administration of mammalian cell carriers. These transporters have immense therapeutic potential as they can be modified to express large banks of biotherapeutics with complementary activities that enhance anti-tumor activity. PMID:26482080
Aims: Delivery of siRNA to the lungs via inhalation offers a unique opportunity to develop a new treatment paradigm for a range of respiratory conditions. However, progress has been greatly hindered by safety and delivery issues. This study developed a high-throughput method for screening novel nanotechnologies for pulmonary siRNA delivery. Methodology: Following physicochemical analysis, the ability of PEI–PEG–siRNA nanoparticles to facilitate siRNA delivery was determined using high-content analysis (HCA) in Calu-3 cells. Results obtained from HCA were validated using confocal microscopy. Finally, cytotoxicity of the PEI–PEG–siRNA particles was analyzed by HCA using the Cellomics® multiparameter cytotoxicity assay. Conclusion: PEI–PEG–siRNA nanoparticles facilitated increased siRNA uptake and luciferase knockdown in Calu-3 cells compared with PEI–siRNA.
Chang, Jui-Chih; Liu, Ko-Hung; Li, Yu-Chi; Kou, Shou-Jen; Wei, Yau-Huei; Chuang, Chieh-Sen; Hsieh, Mingli; Liu, Chin-San
We explored the feasibility of mitochondrial therapy using the cell-penetrating peptide Pep-1 to transfer mitochondrial DNA (mtDNA) between cells and rescue a cybrid cell model of the mitochondrial disease myoclonic epilepsy with ragged-red fibres (MERRF) syndrome. Pep-1-conjugated wild-type mitochondria isolated from parent cybrid cells incorporating a mitochondria-specific tag were used as donors for mitochondrial delivery into MERRF cybrid cells (MitoB2) and mtDNA-depleted Rho-zero cells (Mitoρ°). Forty-eight hours later, translocation of Pep-1-labelled mitochondria into the mitochondrial regions of MitoB2 and Mitoρ° host cells was observed (delivery efficiencies of 77.48 and 82.96%, respectively). These internalized mitochondria were maintained for at least 15 days in both cell types and were accompanied by mitochondrial function recovery and cell survival by preventing mitochondria-dependent cell death. Mitochondrial homeostasis analyses showed that peptide-mediated mitochondrial delivery (PMD) also increased mitochondrial biogenesis in both cell types, but through distinct regulatory pathways involving mitochondrial dynamics. Dramatic decreases in mitofusin-2 (MFN2) and dynamin-related protein 1/fission 1 were observed in MitoB2 cells, while Mitoρ° cells showed a significant increase in optic atrophy 1 and MFN2. These findings suggest that PMD can be used as a potential therapeutic intervention for mitochondrial disorders. PMID:23006856
Jiang, Ling; Xiong, Xiao-Peng; Hu, Chao-su; Ou, Zhou-Luo; Zhu, Guo-Pei; Ying,, Hong-Mei
Intensity-modulated radiation therapy, when used in the clinic, prolongs fraction delivery time. Here we investigated both the in vivoand in vitroradiobiological effects on the A549 cell line, including the effect of different delivery times with the same dose on A549 tumor growth in nude mice. The in vitroeffects were studied with clonogenic assays, using linear-quadratic and incomplete repair models to fit the dose-survival curves. Fractionated irradiation of different doses was given at on...
Zhao, Xiao; Yang, Keni; Zhao, Ruifang; Ji, Tianjiao; Wang, Xiuchao; Yang, Xiao; Zhang, Yinlong; Cheng, Keman; Liu, Shaoli; Hao, Jihui; Ren, He; Leong, Kam W; Nie, Guangjun
Immunogenic cell death (ICD) occurs when apoptotic tumor cell elicits a specific immune response, which may trigger an anti-tumor effect, via the release of immunostimulatory damage-associated molecular patterns (DAMPs). Hypothesizing that nanomedicines may impact ICD due to their proven advantages in delivery of chemotherapeutics, we encapsulated oxaliplatin (OXA) or gemcitabine (GEM), an ICD and a non-ICD inducer respectively, into the amphiphilic diblock copolymer nanoparticles. Neither GEM nor nanoparticle-encapsulated GEM (NP-GEM) induced ICD, while both OXA and nanoparticle-encapsulated OXA (NP-OXA) induced ICD. Interestingly, NP-OXA treated tumor cells released more DAMPs and induced stronger immune responses of dendritic cells and T lymphocytes than OXA treatment in vitro. Furthermore, OXA and NP-OXA exhibited stronger therapeutic effects in immunocompetent mice than in immunodeficient mice, and the enhancement of therapeutic efficacy was significantly higher in the NP-OXA group than the OXA group. Moreover, NP-OXA treatment induced a higher proportion of tumor infiltrating activated cytotoxic T-lymphocytes than OXA treatment. This general trend of enhanced ICD by nanoparticle delivery was corroborated in evaluating another pair of ICD inducer and non-ICD inducer, doxorubicin and 5-fluorouracil. In conclusion, although nanoparticle encapsulation did not endow a non-ICD inducer with ICD-mediated anti-tumor capacity, treatment with a nanoparticle-encapsulated ICD inducer led to significantly enhanced ICD and consequently improved anti-tumor effects than the free ICD inducer. The proposed nanomedicine approach may impact cancer immunotherapy via the novel cell death mechanism of ICD. PMID:27343466
Durymanov, Mikhail O; Yarutkin, Alexey V; Khramtsov, Yuri V; Rosenkranz, Andrey A; Sobolev, Alexander S
Utilizing nanoparticles made of cationic polymers as gene carriers is a promising approach in cancer gene therapy. One of the major requirements for successful gene delivery is DNA translocation into the nuclei of cancer cells. Nuclear envelope breakdown during mitosis has been considered as the most favorable opportunity for DNA translocation to the nucleus. Here, we aimed to study the influence of mitosis on polyplex-mediated gene delivery using time-lapse microscopy as a safe and accurate tool. Studying of reporter gene expression on a single cell level enabled to confirm the significance of cell division for gene delivery to Cloudman S91 melanoma cells, in spite of minor mitosis-independent transfection, and to discover some important details of polyplex delivery process. We have found that cell division can result in only one post-mitotic transfected cell of the two that could indicate non-uniform distribution of a very small number of intact plasmid DNA between daughter cells. According to our data, the shorter the time interval from polyplex addition to cell division, the longer time is required for the start of reporter gene expression after completed cytokinesis that presumably is a result of gradual polyplex dissociation in cell. Most probably, the development of new gene delivery carriers which would combine the strong ability to protect DNA and ability to release it during mitosis can provide an increase in intact DNA molecule number per cell, uniform DNA distribution between two post-mitotic cells, and fast reporter gene expression resulting in superior transfection of proliferating cells. PMID:26239430
Malecki, Maciej; PROCZKA, ROBERT; Chorostowska-Wynimko, Joanna; Swoboda, Paweł; DELBANI, ANNA; Pachecka, Jan
Peritoneal dissemination of cancer cells is characteristic of advanced stages of ovarian, breast and lung cancers, and is associated with poor patient survival. The presence of cancer cells in effusions complicates treatment protocols, while cell eradication is seriously limited. One of the novel options available is cancer gene therapy with recombinant adeno-associated viruses. This combination represents the most promising gene delivery vehicles to neoplasmatic cells within serosal cavities...
Şalva, Emine; Turan, Suna Özbaş; Ekentok, Ceyda; Akbuğa, Jülide
Establishing stable cell lines are useful tools to study the function of various genes and silence or induce the expression of a gene of interest. Nonviral gene transfer is generally preferred to generate stable cell lines in the manufacturing of recombinant proteins. In this study, we aimed to establish stable recombinant HEK-293 cell lines by transfection of chitosan complexes preparing with pDNA which contain LacZ and GFP genes. Chitosan which is a cationic polymer was used as gene delivery system. Stable HEK-293 cell lines were established by transfection of cells with complexes which were prepared with chitosan and pVitro-2 plasmid vector that contains neomycin drug resistance gene, beta gal and GFP genes. The transfection efficiency was shown with GFP expression in the cells using fluorescence microscopy. Beta gal protein expression in stable cells was examined by beta-galactosidase assay as enzymatically and X-gal staining method as histochemically. Full complexation was shown in the above of 1/1 ratio in the chitosan/pDNA complexes. The highest beta-galactosidase activity was obtained with transfection of chitosan complexes. Beta gal gene expression was 15.17 ng/ml in the stable cells generated by chitosan complexes. In addition, intensive blue color was observed depending on beta gal protein expression in the stable cell line with X-gal staining. We established a stable HEK-293 cell line that can be used for recombinant protein production or gene expression studies by transfecting the gene of interest. PMID:26134852
Norberg, Seth A.; Johnsen, Eric; Kushner, Mark J.
The use of atmospheric pressure plasma jets (APPJs) in plasma medicine have produced encouraging results in wound treatment, surface sterilization, deactivation of bacteria, and treatment of cancer cells. It is known that many of the reactive oxygen and nitrogen species produced by the APPJ are critical to these processes. Other key components to treatment include the ion and photon fluxes, and the electric fields produced in cells by the ionization wave of the APPJ striking in the vicinity of the cells. These relationships are often complicated by the cells being covered by a thin liquid layer—wet cells. In this paper, results from a computational investigation of the interaction of APPJs with tissue beneath a liquid layer are discussed. The emphasis of this study is the delivery of electric fields by an APPJ sustained in He/O2 = 99.8/0.2 flowing into humid air to cells lying beneath water with thickness of 200 μm. The water layer represents the biological fluid typically covering tissue during treatment. Three voltages were analyzed—two that produce a plasma effluent that touches the surface of the water layer and one that does not touch. The effect of the liquid layer thickness, 50 μm to 1 mm, was also examined. Comparisons were made of the predicted intracellular electric fields to those thresholds used in the field of bioelectronics.
Yao, Kevin; Ricardo, Sharon D
MicroRNAs (miRNAs) are short single strands of RNA responsible for post-transcriptional regulation of gene expression and have been implicated in the pathogenesis of chronic kidney disease (CKD). Emerging evidence reports that miRNAs can reduce kidney fibrosis through regulation of targets associated with collagen and extracellular matrix accumulation. However, the development of miRNA therapies has been hampered by the lack of targeted and sustainable methods of systemic miRNA delivery. Mesenchymal stem cells (MSCs) provide a promising miRNA delivery platform to overcome toxicity, the potential for insertional mutations and the low efficiency of previous methods. MSCs are endogenously immunoprivileged and home to sites of inflammation. They also release trophic growth factors to modulate the immune system, alter the polarization of macrophages and provide renal protection and repair. The potential to engineer MSCs to express or overexpress miRNAs, released by exosomes, may enhance their natural functions. Clinical studies are already being conducted individually for the use of miRNAs in cancer and MSCs in diseases associated with CKD. Hence, the combination of miRNAs and MSCs may provide an unparalleled cell-based therapy for treating CKD. PMID:26437381
Anirudhan, Thayyath Sreenivasan; Binusreejayan
Curcumin (Cur), a poly phenolic yellow colored compound present in Indian spice turmeric, has a wide variety of biological properties. Bioavailability of Cur is limited by its low water solubility, rapid metabolism and low stability. In the present study, we mainly focus on synthesis and characterization of dextran based nano-sized drug carrier (GHDx) for the delivery of Cur. A liver targeting moiety is incorporated in GHDx so as to improve the therapeutic efficiency and decrease adverse effects of conventional cancer therapy. The effect of different parameters on grafting variables was studied. GHDx was characterised by FTIR, (1)H NMR XRD, TG/DTG, TEM, SEM, AFM, DLS and zeta potential analyses. Adsorption experiments were carried out for drug loading. Swelling of GHDx was studied as a function of pH and temperature. Three step release of Cur from GHDx was confirmed by analyzing in vitro release data in simulated intracellular pH using different kinetic models. In vitro cytotoxicity analysis on L929 and Hep G2 cells shows that GHDx is safe carrier while Cur loaded GHDx exhibits high toxicity with slow drug release towards hepatic cells. The results show that the GHDx can be customized as a stimuli sensitive potential carrier for the delivery of drugs. PMID:27012895
Microinjection of extracellular molecules into a single animal cell was performed by an amplified femtosecond laser irradiation. When a single-shot laser pulse was focused on the plasma membrane of a single fibroblast from the mouse cell line NIH3T3 with a high-numerical aperture objective lens, a transient hole with a diameter of 1 μm was formed. The delivery process of extracellular molecules immediately after the hole formation was monitored by a fluorescence staining with fluoresceinisothiocyanate-dextran (FITC-dextran). Then the gene expression was confirmed using a DNA plasmid of an enhanced green fluorescent protein (EGFP). The gene expression was observed when the laser pulse was focused first on the cellular membrane and then on the nuclear membrane, while the gene was not expressed when the laser was focused only on the cellular membrane. On the basis of these results, the efficiency of gene delivery by the femtosecond laser microinjection and the subsequent gene expression were clarified.
Gharat, L; Taneja, R; Weerapreeyakul, N; Rege, B; Polli, J; Chikhale, P J
We demonstrate transport across, intracellular accumulation and bioreductive activation of a conformationally constrained, anticancer drug delivery system (the CH(3)-TDDS) using Caco-2 cell monolayers (CCMs) as an in vitro model of the human intestinal mucosa. Reverse-phase High Performance Liquid Chromatography (HPLC) coupled with UV detection was used to detect CH(3)-TDDS, the bioreduction product (lactone) and the released drug (melphalan methyl ester; MME). Upon incubation of the CH(3)-TDDS with the apical (AP) surface of 21-day-old CCM, we observed rapid decrease in the AP concentration of the CH(3)-TDDS (60%/hr) as a result of cellular uptake. Rapid intracellular accumulation of the CH(3)-TDDS was followed by bioreductive activation to deplete the cellular levels of CH(3)-TDDS. The drug part (MME) and lactone, as well as CH(3)-TDDS, were detected in the basolateral (BL) chamber. Intracellular Caco-2 levels of TDDS and lactone were also detectable. Bioreductive activation of the CH(3)-TDDS was additionally confirmed by formation of lactone after incubation of the CH(3)-TDDS in the presence of freshly prepared Caco-2 cell homogenates. During transport studies of melphalan or MME alone (as control), the intact drug was not detected in the intracellular compartment or in the BL chamber. These observations demonstrate that CH(3)-TDDS has potential for improving intestinal delivery of MME. TDDS could be useful in facilitating oral absorption of MME as well as the oral delivery of other agents. PMID:11337161
Huang, Yuan-Pin; Lin, I.-Jou; Chen, Chih-Chen; Hsu, Yi-Chiang; Chang, Chi-Chang; Lee, Mon-Juan
Carbon nanotubes are capable of penetrating the cell membrane and are widely considered as potential carriers for gene or drug delivery. Because the C-C and C=C bonds in carbon nanotubes are nonpolar, functionalization is required for carbon nanotubes to interact with genes or drugs as well as to improve their biocompatibility. In this study, polyethylenimine (PEI)-functionalized single-wall (PEI-NH-SWNTs) and multiwall carbon nanotubes (PEI-NH-MWNTs) were produced by direct amination method. PEI functionalization increased the positive charge on the surface of SWNTs and MWNTs, allowing carbon nanotubes to interact electrostatically with the negatively charged small interfering RNAs (siRNAs) and to serve as nonviral gene delivery reagents. PEI-NH-MWNTs and PEI-NH-SWNTs had a better solubility in water than pristine carbon nanotubes, and further removal of large aggregates by centrifugation produced a stable suspension of reduced particle size and improved homogeneity and dispersity. The amount of grafted PEI estimated by thermogravimetric analysis was 5.08% ( w/ w) and 5.28% ( w/ w) for PEI-NH-SWNTs and PEI-NH-MWNTs, respectively. For the assessment of cytotoxicity, various concentrations of PEI-NH-SWNTs and PEI-NH-MWNTs were incubated with human cervical cancer cells, HeLa-S3, for 48 h. PEI-NH-SWNTs and PEI-NH-MWNTs induced cell deaths in a dose-dependent manner but were less cytotoxic compared to pure PEI. As determined by electrophoretic mobility shift assay, siRNAs directed against glyceraldehyde-3-phosphate dehydrogenase (siGAPDH) were completely associated with PEI-NH-SWNTs or PEI-NH-MWNTs at a PEI-NH-SWNT/siGAPDH or PEI-NH-MWNT/siGAPDH mass ratio of 80:1 or 160:1, respectively. Furthermore, PEI-NH-SWNTs and PEI-NH-MWNTs successfully delivered siGAPDH into HeLa-S3 cells at PEI-NH-SWNT/siGAPDH and PEI-NH-MWNT/siGAPDH mass ratios of 1:1 to 20:1, resulting in suppression of the mRNA level of GAPDH to an extent similar to that of DharmaFECT, a common transfection
Hörner, Sebastian; Fabritz, Sebastian; Herce, Henry D; Avrutina, Olga; Dietz, Christian; Stark, Robert W; Cardoso, M Cristina; Kolmar, Harald
Cube octameric silsesquioxanes (COSS) are among the smallest nanoparticles known to date with a diameter of only 0.7 nm. We describe a COSS-based delivery system which allows for the drug targeting in human cells. It comprises a siloxane core with seven pendant aminopropyl groups and a fluorescently labeled peptidic ligand attached to one cage corner via a reversible disulfide bond to ensure its intracellular release. Bimodal amplitude-modulated atomic force microscopy (AFM) experiments revealed the formation of dendritic COSS structures by a self-assembly of single particles on negatively charged surfaces. Nuclear targeting was demonstrated in HeLa cells by selective binding of released p21(Cip1/Waf1)-derived cargo peptide to PCNA, a protein involved in DNA replication and repair. PMID:23250285
Hong, Tu; Guo, Honglian; Xu, Yaqiong
Small interfering RNA (siRNA) is potentially a promising tool in influencing gene expression with a high degree of target specificity. However, its poor intracellular uptake, instability in vivo, and non-specific immune stimulations impeded its effect in clinical applications. In this study, carbon nanotubes (CNTs) functionalized with two types of phospholipid-polyethylene glycol (PEG) have shown capabilities to stabilize siRNA in cell culture medium during the transfection and efficiently deliver siRNA into neuroblastoma and breast cancer cells. Moreover, the intrinsic optical properties of CNTs have been investigated through absorption and fluorescence measurements. We have found that the directly-functionalized groups play an important role on the fluorescence imaging of functionalized CNTs. The unique fluorescence imaging and high delivery efficiency make CNTs a promising material to deliver drugs and evaluate the treatment effect simultaneously.
Regional cerebral blood flow (rCBF) was determined in 77 normal females and 53 normal males of different ages and in 26 men and 45 women with multiple sclerosis by the inhalation of radioactive Xe133 method. In the normal subjects the CBF was relatively high in the teens and fell, at first rapidly and then slowly in both sexes with age. During adult life the flow in females was significantly higher than in males. The delivery of packed red cells (RCD) was determined by multiplying the CBF by the percentage concentration of red cells (HCT). The RCD for both sexes was nearly the same. In the patients with multiple sclerosis there occurred a progressive generalized decrease in CBF and in RCD with age which was significantly greater than observed in normal subjects. The rate of decrease in CBF and RCD correlated directly with the rate of progress of the disease
Full Text Available Abstract Background Mesenchymal stem cells (MSCs have been considered to be the attractive vehicles for delivering therapeutic agents toward various tumor diseases. This study was to explore the distribution pattern, kinetic delivery of adenovirus, and therapeutic efficacy of the MSC loading of E1A mutant conditionally replicative adenovirus Adv-Stat3(- which selectively replicated and expressed high levels of anti-sense Stat3 complementary DNA in breast cancer and melanoma cells. Methods We assessed the release ability of conditionally replicative adenovirus (CRAd from MSC using crystal violet staining, TCID50 assay, and quantitative PCR. In vitro killing competence of MSCs carrying Adv-Stat3(- toward breast cancer and melanoma was performed using co-culture system of transwell plates. We examined tumor tropism of MSC by Prussian blue staining and immunofluorescence. In vivo killing competence of MSCs carrying Adv-Stat3(- toward breast tumor was analyzed by comparison of tumor volumes and survival periods. Results Adv-Stat3(- amplified in MSCs and were released 4 days after infection. MSCs carrying Adv-Stat3(- caused viral amplification, depletion of Stat3 and its downstream proteins, and led to significant apoptosis in breast cancer and melanoma cell lines. In vivo experiments confirmed the preferential localization of MSCs in the tumor periphery 24 hours after tail vein injection, and this localization was mainly detected in the tumor parenchyma after 72 hours. Intravenous injection of MSCs carrying Adv-Stat3(- suppressed the Stat3 pathway, down-regulated Ki67 expression, and recruited CD11b-positive cells in the local tumor, inhibiting tumor growth and increasing the survival of tumor-bearing mice. Conclusions These results indicate that MSCs migrate to the tumor site in a time-dependent manner and could be an effective platform for the targeted delivery of CRAd and the amplification of tumor killing effects.
Schuster, Martin; Martin-Urdiroz, Magdalena; Higuchi, Yujiro; Hacker, Christian; Kilaru, Sreedhar; Gurr, Sarah J; Steinberg, Gero
Fungal cells are surrounded by an extracellular cell wall. This complex matrix of proteins and polysaccharides protects against adverse stresses and determines the shape of fungal cells. The polysaccharides of the fungal wall include 1,3-β-glucan and chitin, which are synthesized by membrane-bound synthases at the growing cell tip. A hallmark of filamentous fungi is the class V chitin synthase, which carries a myosin-motor domain. In the corn smut fungus Ustilago maydis, the myosin-chitin synthase Mcs1 moves to the plasma membrane in secretory vesicles, being delivered by kinesin-1 and myosin-5. The myosin domain of Mcs1 enhances polar secretion by tethering vesicles at the site of exocytosis. It remains elusive, however, how other cell-wall-forming enzymes are delivered and how their activity is coordinated post secretion. Here, we show that the U. maydis class VII chitin synthase and 1,3-β-glucan synthase travel in Mcs1-containing vesicles, and that their apical secretion depends on Mcs1. Once in the plasma membrane, anchorage requires enzyme activity, which suggests co-synthesis of chitin and 1,3-β-glucan polysaccharides at sites of exocytosis. Thus, delivery of cell-wall-forming enzymes in Mcs1 vesicles ensures local foci of fungal cell wall formation. PMID:27563844
Geninatti Crich, S.; Cadenazzi, M.; Lanzardo, S.; Conti, L.; Ruiu, R.; Alberti, D.; Cavallo, F.; Cutrin, J. C.; Aime, S.
In this work the selective uptake of native horse spleen ferritin and apoferritin loaded with MRI contrast agents has been assessed in human breast cancer cells (MCF-7 and MDA-MB-231). The higher expression of L-ferritin receptors (SCARA5) led to an enhanced uptake in MCF-7 as shown in T2 and T1 weighted MR images, respectively. The high efficiency of ferritin internalization in MCF-7 has been exploited for the simultaneous delivery of curcumin, a natural therapeutic molecule endowed with antineoplastic and anti-inflammatory action, and the MRI contrast agent Gd-HPDO3A. This theranostic system is able to treat selectively breast cancer cells over-expressing ferritin receptors. By entrapping in apoferritin both Gd-HPDO3A and curcumin, it was possible to deliver a therapeutic dose of 167 μg ml-1 (as calculated by MRI) of this natural drug to MCF-7 cells, thus obtaining a significant reduction of cell proliferation.In this work the selective uptake of native horse spleen ferritin and apoferritin loaded with MRI contrast agents has been assessed in human breast cancer cells (MCF-7 and MDA-MB-231). The higher expression of L-ferritin receptors (SCARA5) led to an enhanced uptake in MCF-7 as shown in T2 and T1 weighted MR images, respectively. The high efficiency of ferritin internalization in MCF-7 has been exploited for the simultaneous delivery of curcumin, a natural therapeutic molecule endowed with antineoplastic and anti-inflammatory action, and the MRI contrast agent Gd-HPDO3A. This theranostic system is able to treat selectively breast cancer cells over-expressing ferritin receptors. By entrapping in apoferritin both Gd-HPDO3A and curcumin, it was possible to deliver a therapeutic dose of 167 μg ml-1 (as calculated by MRI) of this natural drug to MCF-7 cells, thus obtaining a significant reduction of cell proliferation. Electronic supplementary information (ESI) available: Competition studies with free apoferritin, Fig. S1; APO-FITC intracellular distribution by
Genova M. P.
Full Text Available The aim of the present study was to assess β-cell function using homeostasis model (HOMA-B and disposition index (DI in pregnant women with/without gestational diabetes, and after delivery. A total of 102 pregnant women between 24-28 gestational weeks (53 with gestational diabetes mellitus (GDM and 49 with normal glucose tolerance (NGT and 22 GDM postpartum women (8-12 weeks after delivery were included in the study. All postpartum women had a history of GDM. HOMA indexes (insulin resistance - HOMA-IR and HOMA-B for assessing β-cell function were calculated from fasting glucose and insulin concentrations. To estimate insulin secretion independent of insulin sensitivity, DI was calculated using glucose and insulin levels at 0 and 60 min during the course of a 2 h 75g oral glucose tolerance test (OGTT. In GDM pregnant women HOMA-B was significantly lower compared to NGT women (p = 0.017, but there was no significant difference compared to women after birth (NS. There was difference between NGT and postpartum women (p < 0.05. DI was significantly lower for GDM pregnant women in comparison to NGT and postpartum women (p < 0.0001; p = 0.011, between NGT and women after birth (p < 0.04. In our study, comparison of НОМА-В in NGT and GDM pregnant women demonstrated that the OR of developing GDM was 0.989 (95% CI, 0.980-0.998, P = 0.013, and comparison of DI in healthy pregnant and GDM showed that the OR of developing GDM was 0.967 (95% CI, 0.947-0.988, P = 0.002. Therefore, HOMA-B and DI appear to be protective factors in the risk of developing GDM. According to our results, assessment of β-cell function, using HOMA-B and DI, showed that they are lower in GDM than NGT group and postpartum women. It is important to note that HOMA-B did not show significant difference between GDM pregnant and women after delivery with a history for GDM. We assume that pregnant women with GDM have a pancreatic β-cell defect that remains after birth. These women
Stem cell transplantation is a promising therapy for a myriad of debilitating diseases and injuries; however, current delivery protocols are inadequate. Transplantation by direct injection, which is clinically preferred for its minimal invasiveness, commonly results in less than 5% cell viability, greatly inhibiting clinical outcomes. We demonstrate that mechanical membrane disruption results in significant acute loss of viability at clinically relevant injection rates. As a strategy to protect cells from these damaging forces, we show that cell encapsulation within hydrogels of specific mechanical properties will significantly improve viability. Building on these fundamental studies, we have designed a reproducible, bio-resorbable, customizable hydrogel using protein-engineering technology. In our Mixing-Induced Two-Component Hydrogel (MITCH), network assembly is driven by specific and stoichiometric peptide-peptide binding interactions. By integrating protein science methodologies with simple polymer physics models, we manipulate the polypeptide chain interactions and demonstrate the direct ability to tune the network crosslinking density, sol-gel phase behavior, and gel mechanics. This is in contrast to many other physical hydrogels, where predictable tuning of bulk mechanics from the molecular level remains elusive due to the reliance on non-specific and non-stoichiometric chain interactions for network formation. Furthermore, the hydrogel network can be easily modified to deliver a variety of bioactive payloads including growth factors, peptide drugs, and hydroxyapatite nanoparticles. Through a series of in vitro and in vivo studies, we demonstrate that these materials may significantly improve transplanted stem cell retention and function.
Zheng, Lixia; Wu, Shao; Tan, Li; Tan, Huo; Yu, Baodan
Delivery of amphiphobic drugs (insoluble in both water and oil) has been a great challenge in drug delivery. SNX-2112, a novel inhibitor of Hsp90, is a promising drug candidate for treating various types of cancers; however, the insolubility greatly limits its clinical application. This study aimed to build a new type of drug delivery system using single-walled carbon nanotubes (SWNTs) for controllable release of SNX-2112; chitosan (CHI) was non-covalently added to SWNTs to improve their biocompatibility. SWNTs-CHI demonstrated high drug-loading capability; the release of SNX-2112 was pH triggered and time related. The intracellular reactive oxygen species of SWNTs-CHI increased, compared with that of SWNTs, leading to higher mitogen-activated protein kinase and cell apoptosis. The results of western-blotting, lactate dehydrogenase (LDH) release assay, and cell viability assay analyses indicated that apoptosis-related proteins were abundantly expressed in K562 cells and that the drug delivery system significantly inhibited K562 cells. Thus, SWNT-CHI/SNX-2112 shows great potential as a drug delivery system for cancer therapy. PMID:27231263
Wu, Fang; Wuensch, Sherry A; Azadniv, Mitra; Ebrahimkhani, Mohammad R; Crispe, I Nicholas
We aim to define the role of Kupffer cells in intrahepatic antigen presentation, using the selective delivery of antigen to Kupffer cells rather than other populations of liver antigen-presenting cells. To achieve this we developed a novel antigen delivery system that can target antigens to macrophages, based on a galactosylated low-density lipoprotein nanoscale platform. Antigen was delivered via the galactose particle receptor (GPr), internalized, degraded and presented to T cells. The conjugation of fluoresceinated ovalbumin (FLUO-OVA) and lactobionic acid with LDL resulted in a substantially increased uptake of FLUO-OVA by murine macrophage-like ANA1 cells in preference to NIH3T3 cells, and by primary peritoneal macrophages in preference to primary hepatic stellate cells. Such preferential uptake led to enhanced proliferation of OVA specific T cells, showing that the galactosylated LDL nanoscale platform is a successful antigen carrier, targeting antigen to macrophages but not to all categories of antigen presenting cells. This system will allow targeted delivery of antigen to macrophages in the liver and elsewhere, addressing the question of the role of Kupffer cells in liver immunology. It may also be an effective way of delivering drugs or vaccines directly at macrophages. PMID:19637876
Bartlett, Rush Lloyd, II
Inflammatory diseases such as osteoarthritis and rheumatoid arthritis cause $127.8 billion in US healthcare expenditures each year and are the cause of disability for 27% of disabled persons in the United States. Current treatment options rarely halt disease progression and often result in significant unwanted and debilitating side effects. Our laboratory has previously developed a family of cell penetrating peptides (CPPs) which inhibit the activity of mitogen activated protein kinase activate protein kinase 2 (MK2). MK2 mediates the inflammatory response by activating Tristetraprline (TTP). Once activated, TTP rapidly stabilizes AU rich regions of pro-inflammatory cytokine mRNA which allows translation of pro-inflammatory cytokines to occur. Blocking MK2 with our labs CPPs yields a decrease in inflammatory activity but CPPs by are highly non specific and prone to rapid enzymatic degradation in vivo.. In order to increase the potency of MK2 inhibiting CPPs we have developed a novel nanoparticle drug carrier composed of poly(N-isopropylacrylamide-co-2-acrylamido-2-methyl-1-propanesulfonic acid). This drug carrier has been shown to have preliminary efficacy in vitro and ex vivo for suppressing pro-inflammatory cytokine production when releasing CPPs. This thesis will present progress made on three aims: Specific Aim 1) Create and validate a NIPAm based drug delivery system that mimics the binding and release previously observed between cell penetrating peptides and glycosaminoglycans. Specific Aim 2) Engineer degradability into poly(NIPAm-AMPS) nanoparticles to enable more drug to be released and qualify that system in vitro. Specific Aim 3) Validate poly(NIPAm-AMPS) nanoparticles for targeted drug delivery in an ex vivo inflammatory model. Overall we have developed a novel anionic nanoparticle system that is biocompatible and efficient at loading and releasing cell penetrating peptides to inflamed tissue. Once loaded with a CPP the nanoparticle drug complex is
Full Text Available "nBackground and the purpose of the study: Of the gene delivery systems, non-viral polycationic gene delivery nanosystems have been alternatively exploited as a relatively safe delivery reagents compared to viral vectors. However, little is known about the genomic impacts of these delivery systems in target cells/tissues. In this study, the toxicogenomics and genotoxicity potential of some selected polycationic lipid/polymer based nanostructures (i.e., Oligofectamine® (OF, starburst polyamidoamine Polyfect® (PF and diaminobutane (DAB dendrimers were investigated in human alveolar epithelial A549 cells. "nMethods: To study the nature and the ontology of the gene expression changes in A549 cells upon treatment with polycationic nanostructures, MTT assay and microarray gene expression profiling methodology were employed. For microarray analysis, cyanine (Cy3/Cy5 labeled cDNA samples from treated and untreated cells were hybridized on target arrays housing 200 genes. "nResults and major conclusions: The polycationic nanosystems induced significant gene expression changes belonging to different genomic ontologies such as cell defence and apoptosis pathways. These data suggest that polycationic nanosystems can elicit multiple gene expression changes in A549 cells upon their chemical structures and interactions with cellular/subcellular components. Such impacts may interfere with the main goals of the desired genemedicine.
Matthias, Nadine; Hunt, Samuel D; Wu, Jianbo; Darabi, Radbod
Muscular dystrophies are among major inherited muscle disorders characterized by progressive muscle damage and fibrosis with no definitive cure. Recently, gene or cell based therapies have been developed to restore the missing gene expression or replace the damaged tissues. In order to test the efficiency of these therapies in mice models of muscular dystrophies, the arterial route of delivery is very advantageous as it provides uniform muscle exposure to the therapeutic agents or cells. Although there are few reports of arterial delivery of the therapeutic agents or cells in mice, there is no in-depth description and evaluation of its efficacy in perfusion of downstream muscles. This study is aimed to develop a practical method for intra-femoral artery perfusion in mice and to evaluate perfusion efficiency using near-infrared-fluorescence (NIRF) imaging as well as histology following stem cell delivery. Our results provide a practical guide to perform this delicate method in mice. By using a sensitive fluorescent dye, different muscle groups of the hindlimb have been evaluated for proper perfusion. As the final step, we have validated the efficiency of arterial cell delivery into muscles using human iPS-derived myogenic cells in an immunodeficient mouse model for Duchenne muscular dystrophy (NSG-mdx(4cv)). PMID:26341268
Flanary, Suzanne; Hoffman, Allan S.; Stayton, Patrick S.
While many infectious diseases are controlled by vaccine strategies, important limitations continue to motivate the development of better antigen delivery systems. This study focuses on the use of a pH-sensitive polymeric carrier based on poly(propylacrylic acid) (PPAA) to address the need for more potent CD8 cytotoxic T-cell (CTL) responses. An MHC-1/CD8 CTL cell model system with ovalbumin as the protein antigen was used to test whether PPAA could enhance the delivery of ovalbumin into the ...
Qiu, Liping; Chen, Tao; Öçsoy, Ismail; Yasun, Emir; Wu, Cuichen; Zhu, Guizhi; You, Mingxu; Han, Da; Jiang, Jianhui; Yu, Ruqin; Tan, Weihong
The development of multidrug resistance (MDR) has become an increasingly serious problem in cancer therapy. The cell-membrane overexpression of P-glycoprotein (P-gp), which can actively efflux various anticancer drugs from the cell, is a major mechanism of MDR. Nuclear-uptake nanodrug delivery systems, which enable intranuclear release of anticancer drugs, are expected to address this challenge by bypassing P-gp. However, before entering the nucleus, the nanocarrier must pass through the cell...
Gaspar, V. M.; Marques, J. G.; Sousa, F.; Louro, R. O.; Queiroz, J. A.; Correia, I. J.
Bridging the gap between nanoparticulate delivery systems and translational gene therapy is a long sought after requirement in nanomedicine-based applications. However, recent developments regarding nanoparticle functionalization have brought forward the ability to synthesize materials with biofunctional moieties that mimic the evolved features of viral particles. Herein we report the versatile conjugation of both cell penetrating arginine and pH-responsive histidine moieties into the chitosan polymeric backbone, to improve the physicochemical characteristics of the native material. Amino acid coupling was confirmed by 2D TOCSY NMR and Fourier transform infrared spectroscopy. The synthesized chitosan-histidine-arginine (CH-H-R) polymer complexed plasmid DNA biopharmaceuticals, and spontaneously assembled into stable 105 nm nanoparticles with spherical morphology and positive surface charge. The functionalized delivery systems were efficiently internalized into the intracellular compartment, and exhibited remarkably higher transfection efficiency than unmodified chitosan without causing any cytotoxic effect. Additional findings regarding intracellular trafficking events reveal their preferential escape from degradative lysosomal pathways and nuclear localization. Overall, this assembly of nanocarriers with bioinspired moieties provides the foundations for the design of efficient and customizable materials for cancer gene therapy.