Generation and transplantation of hepatocytes-like cells using human origin hepatogenic serum for acute liver injury treatment.

Liver failure is a critical disease for which regenerative therapies are still being explored. The major limitation in the use of a clinical grade, viable cell-based therapy approach is the scarce availability of sufficient number of in-vitro differentiated hepatocyte-like cells (HLC) that can induce regeneration and ameliorate liver injury. Here, we report for the first time an approach to engineer HLCs using sera of hyperbilirubin patients that act as a reservoir of differentiation factor. Utilizing our humanized approach, mesenchymal stem cells (hMSC) derived from umbilical cord tissue were transdifferentiated into HLC using patient-derived serum along with dimethyl sulfoxide (DMSO). We studied the effects of serum on the proliferation, cell cycle analysis, and apoptosis of hMSC by various differentiation combinations. We optimized the hepatic transdifferentiation ability of hMSC with hyperbilirubin serum treatment for a period of 7 days. Assessment of HLC functionalities was shown by quantifying the HLC spent medium for albumin and urea secretions. Transplantation of HLC in an acute liver injury (ALI) rat model showed an effective improvement in the liver function and histological changes in the liver. The results of this study suggest that hMSC-derived HLC using humanized hepatogenic serum holds a promising potential for cell transplantation, as an efficient therapy modality for liver failure in humans.

Recent Advances in Cardiac Tissue Engineering for the Management of Myocardium Infarction.

Myocardium Infarction (MI) is one of the foremost cardiovascular diseases (CVDs) causing death worldwide, and its case numbers are expected to continuously increase in the coming years. Pharmacological interventions have not been at the forefront in ameliorating MI-related morbidity and mortality. Stem cell-based tissue engineering approaches have been extensively explored for their regenerative potential in the infarcted myocardium. Recent studies on microfluidic devices employing stem cells under laboratory set-up have revealed meticulous events pertaining to the pathophysiology of MI occurring at the infarcted site. This discovery also underpins the appropriate conditions in the niche for differentiating stem cells into mature cardiomyocyte-like cells and leads to engineering of the scaffold via mimicking of native cardiac physiological conditions. However, the mode of stem cell-loaded engineered scaffolds delivered to the site of infarction is still a challenging mission, and yet to be translated to the clinical setting. In this review, we have elucidated the various strategies developed using a hydrogel-based system both as encapsulated stem cells and as biocompatible patches loaded with cells and applied at the site of infarction.

A critical appraisal of humanized alternatives to fetal bovine serum for clinical applications of umbilical cord derived mesenchymal stromal cells.

OBJECTIVE: The study is aimed to verify the possibility of using humanized alternatives to fetal bovine serum (FBS) such as umbilical cord blood plasma (CBP) and AB+ plasma to support the long-term growth of mesenchymal stromal cells (MSCs) derived from the umbilical cord. We hypothesized that umbilical CBP would be a potential substitute to FBS, especially for small scale autologous clinical transplantations. METHODS: The MSCs were cultured for six consecutive passages to evaluate xeno-free media's ability to support long-term growth. Cell proliferation rates, colony-forming-unit (CFU) efficiency and population doublings of expanded MSCs, were investigated. Ex vivo expanded MSCs were further characterized using flow cytometry and quantitative PCR. The impact of cryopreservation and composition of cryomedium on phenotype, viability of MSC was also assessed. RESULTS: Our results on cell proliferation, colony-forming unit efficiency suggested that the expansion of the cells was successfully carried out in media supplemented with humanized alternatives. MSCs showed lower CFU counts in FBS (~ 25) than humanized alternatives (~ 35). The gene expression analysis revealed that transcripts showed significant differential expression by two to three folds in the FBS group compared with MSCs grown in medium with humanized alternatives (p < 0.05). In addition, MSCs grown in a medium with FBS had more osteogenic activity, a signature of unwanted differentiation. The majority of ex vivo expanded MSCs at early and late passages expressed CD44+, CD73+, CD105+, CD90+, and CD166+ in all the experimental groups tested (~ 90%). In contrast to the other MSC surface markers, expression levels of STRO-1+ (~ 21-10%) and TNAP+ (~ 29-11%) decreased with the increase in passage number for MSCs cultured in a FBS-supplemented medium (p < 0.05). CONCLUSION: Our results established that CBP supported culture of umbilical cord tissue-derived MSCs and is a safer Xeno free replacement to FBS. The use of CBP also enables the storage of umbilical cord tissue derived MSCs in patient-specific conditions to minimize adverse events if cells are delivered directly to the patient.

Recent trends in biodegradable polyester nanomaterials for cancer therapy.

Biodegradable polyester nanomaterials-based drug delivery vehicles (DDVs) have been largely used in most of the cancer treatments due to its high biological performance and wider applications. In several previous studies, various biodegradable and biocompatible polyester backbones were used which are poly(lactic acid) (PLA), poly(ε-caprolactone) (PCL), poly(propylene fumarate) (PPF), poly(lactic-co-glycolic acid) (PLGA), poly(propylene carbonate) (PPC), polyhydroxyalkanoates (PHA), and poly(butylene succinate) (PBS). These polyesters were fabricated into therapeutic nanoparticles that carry drug molecules to the target site during the cancer disease treatment. In this review, we elaborately discussed the chemical synthesis of different synthetic polyesters and their use as nanodrug carriers (NCs) in cancer treatment. Further, we highlighted in brief the recent developments of metal-free semi-aromatic polyester nanomaterials along with its role as cancer drug delivery vehicles.

Low frequency magnetic force augments hepatic differentiation of mesenchymal stem cells on a biomagnetic nanofibrous scaffold.

Liver tissue engineering using polymeric nanofibrous scaffold and stem cells holds great promises for treating end-stage liver failures. The aim of this study was to evaluate hepatic trans-differentiation potential of human mesenchymal stem cells (hMSCs) on a biomagnetic electrospun nanofibrous scaffold fabricated from a blend of poly-L-lactide (PLLA), collagen and fibrin-rich blood clot, under the influence of a low frequency magnetic field. The scaffold was characterized for surface properties, biochemical and biomechanical parameters and bio-magnetic behaviour. Cell proliferation assay revealed that the scaffold was suitable for hMSCs adhesion and proliferation. Hepatic trans-differentiation potential of hMSCs was augmented on nanofibrous scaffold in magnetic field exposure group compared to control groups, as evident by strong expression of hepatocyte specific markers, albumin release, urea synthesis and presence of an inducible cytochrome P450 system. Our results conclude that biomagnetic scaffold of PLLA/collagen/blood clot augments hepatic trans-differentiation of hMSCs under magnetic field influence.

Nanofibers coated on acellular tissue-engineered bovine pericardium supports differentiation of mesenchymal stem cells into endothelial cells for tissue engineering.

AIM: This study aimed to develop biodegradable, polymer-based nanofibers coated on acellular tissue-engineered bovine pericardium (ATEBP) for cell interfaces, enabling more exquisite functionality, such as mesenchymal stem cell (MSC) adhesion, proliferation and differentiation into endothelial cells for tissue engineering. MATERIALS & METHODS: ATEBP coated with nanofibers of poly(L-lactic acid)-co-poly(ε-caprolactone) (PLACL) and a blend of PLACL and gelatin were analyzed for human bone marrow-derived MSC adhesion, proliferation and differentiation into endothelial cells. RESULTS: The cell culture-based approach showed an increase in human bone marrow-derived MSC adhesion, proliferation and differentiation into endothelial cells on ATEBP coated with PLACL/gelatin nanofibers compared with ATEBP and PLACL nanofibers coated on ATEBP. CONCLUSION: ATEBP coated with PLACL/gelatin nanofibrous scaffolds, along with human bone marrow-derived MSCs differentiated into endothelial cells, might improve the scaffolds' functionality for tissue engineering.

In vitro hepatic trans-differentiation of human mesenchymal stem cells using sera from congestive/ischemic liver during cardiac failure.

Cellular therapy for end-stage liver failures using human mesenchymal stem cells (hMSCs)-derived hepatocytes is a potential alternative to liver transplantation. Hepatic trans-differentiation of hMSCs is routinely accomplished by induction with commercially available recombinant growth factors, which is of limited clinical applications. In the present study, we have evaluated the potential of sera from cardiac-failure-associated congestive/ischemic liver patients for hepatic trans-differentiation of hMSCs. Results from such experiments were confirmed through morphological changes and expression of hepatocyte-specific markers at molecular and cellular level. Furthermore, the process of mesenchymal-to-epithelial transition during hepatic trans-differentiation of hMSCs was confirmed by elevated expression of E-Cadherin and down-regulation of Snail. The functionality of hMSCs-derived hepatocytes was validated by various liver function tests such as albumin synthesis, urea release, glycogen accumulation and presence of a drug inducible cytochrome P450 system. Based on these findings, we conclude that sera from congestive/ischemic liver during cardiac failure support a liver specific microenvironment for effective hepatic trans-differentiation of hMSCs in vitro.

Nanofiber-reinforced myocardial tissue-construct as ventricular assist device.

OBJECTIVES: This study aimed to create a myocardial tissue construct by tissue engineering to repair, replace, and regenerate damaged cardiac tissue. METHODS AND RESULTS: Human cardiac muscles harvested from a homograft heart retrieval system were decellularized followed by coating with electrospun nanofibers to make them amenable to scaffolding. These processed cardiac tissues were nourished in modified media having ischemic cardiac tissue conditioned media in 6 separate experimental variants, and cord blood mononuclear cells were injected into 4 of them. On the 17th day of culture, the nanofiber-coated scaffolds injected with mononuclear cells and/or reinforced by electrical and mechanical forces, started contracting spontaneously at varying rates, while the control remain noncontractile. Histological staining confirmed the pre-culture acellularity as well as post-culture stem cell viability, and revealed expression of troponin I and cardiac myosin. The acellular processed scaffold when implanted into sheep ischemic myocardial apex revealed transformation into sheep myocardium after 4 months of implantation. CONCLUSION: These results provide direct evidence for the re-cellularization of decellularized cardiac tissue grafts reinforced with a polymer nanofiber coating, by human mononuclear cells injection, leading to generation of a tissue-engineered myocardial construct.

A multiplex PCR technique to characterize human bone marrow derived mesenchymal stem cells.

Human mesenchymal stem cells (MSCs), with capacity to differentiate into adipocytes, osteoblasts and chondrocytes, offer potential for the development of novel treatments. A critical question in MSCs biology is whether this cell population possesses a relatively uniform differentiation capability or is comprised of distinct subsets of progenitors committed to differentiate in particular pathways. To quantify the changes during growth of MSCs, we analyzed the mesenchymal phenotype and differentiation ability using a multi-marker PCR with six primer sets specific for CD73, CD90, CD105, CD166, CD45 and beta-actin allowing a gel-based differential detection of the PCR products. To determine degree of variability of MSCs populations in terms of proliferation, cell proliferation assays were performed on expanded MSCs up to the sixth passage. At each passage, the osteogenic and adipogenic differentiation potentials of MSCs were verified by culture in inductive media. RT-PCR and cytochemical analysis revealed that, despite the loss of multipotentiality during expansion, certain markers remain expressed, indicating that these markers are unlikely to be reflective of the MSC's true 'stem cell' nature. Our results suggest that decrease in the expression of MSCs specific markers correlates with down-regulation of proliferation ability and differentiation efficiency of MSCs.