A Two-Stage Process for Differentiation of Wharton's Jelly-Derived Mesenchymal Stem Cells into Neuronal-like Cells.

With no permanent cure for neurodegenerative diseases, the symptoms reappear shortly after the withdrawal of medicines. A better treatment outcome can be expected if the damaged neurons are partly replaced by functional neurons and/or they are repaired using trophic factors. In this regard, safe cell therapy has been considered as a potential alternative to conventional treatment. Here, we have described a two-stage culture process to differentiate Wharton Jelly mesenchymal stem cells (WJ-MSCs) into neuronal-like cells in the presence of various cues involved in neurogenesis. The fate of cells at the end of each stage was analyzed at the morphometric, transcriptional, and translational levels. In the first stage of priming, constitutively, wingless-activated WJ-MSCs crossed the lineage boundary in favor of neuroectodermal lineage, identified by the loss of mesenchymal genes with concomitant expression of neuron-specific markers, like SOX1, PAX6, NTRK1, and NEUROD2. Neuronal-like cells formed in the second stage expressed many mature neuronal proteins like Map2, neurofilament, and Tuj1 and possessed axon hillock-like structures. In conclusion, the differentiation of a large number of neuronal-like cells from nontumorigenic and trophic factors secreting WJ-MSCs promises the development of a therapeutic strategy to treat neurodegenerative diseases.

Aggressive serous epithelial ovarian cancer is potentially propagated by EpCAM+CD45+ phenotype.

Epithelial ovarian carcinoma (EOC) patients often acquire resistance against common chemotherapeutic drugs like paclitaxel and cisplatin. The mechanism responsible for the same is ambiguous. We have identified a putative drug-resistant tumour cell phenotype (EpCAM+CD45+) in the ascitic fluid of EOC patients, which appears to originate from the primary tumour. These cells represent the major tumour burden and are more drug resistant compared to EpCAM+ tumour cells due to the over-expression of SIRT1, ABCA1 and BCL2 genes. We have found that the entire EpCAM+CD45+ population is highly invasive with signature mesenchymal gene expression and also consists of subpopulations of ovarian cancer stem cells (CD133+ and CD117+CD44+). Additionally, we demonstrate that the EpCAM+CD45+ tumour cells over-express major histocompatibility complex class I antigen, which enable them to evade the natural killer cell-mediated immune surveillance. Preliminary evidence obtained in OVCAR-5 cells suggests that exosomes, secreted by non-tumour cells of the ascitic fluid, play an important role in rendering drug resistance and invasive properties to the cancer cells. Identification of such aggressive tumour cells and deciphering their origin is important for designing better drug targets for EOC.

JMJD3 aids in reprogramming of bone marrow progenitor cells to hepatic phenotype through epigenetic activation of hepatic transcription factors.

The strictly regulated unidirectional differentiation program in some somatic stem/progenitor cells has been found to be modified in the ectopic site (tissue) undergoing regeneration. In these cases, the lineage barrier is crossed by either heterotypic cell fusion or direct differentiation. Though studies have shown the role of coordinated genetic and epigenetic mechanisms in cellular development and differentiation, how the lineage fate of adult bone marrow progenitor cells (BMPCs) is reprogrammed during liver regeneration and whether this lineage switch is stably maintained are not clearly understood. In the present study, we wanted to decipher genetic and epigenetic mechanisms that involve in lineage reprogramming of BMPCs into hepatocyte-like cells. Here we report dynamic transcriptional change during cellular reprogramming of BMPCs to hepatocytes and dissect the epigenetic switch mechanism of BM cell-mediated liver regeneration after acute injury. Genome-wide gene expression analysis in BM-derived hepatocytes, isolated after 1 month and 5 months of transplantation, showed induction of hepatic transcriptional program and diminishing of donor signatures over the time. The transcriptional reprogramming of BM-derived cells was found to be the result of enrichment of activating marks (H3K4me3 and H3K9Ac) and loss of repressive marks (H3K27me3 and H3K9me3) at the promoters of hepatic transcription factors (HTFs). Further analyses showed that BMPCs possess bivalent histone marks (H3K4me3 and H3K27me3) at the promoters of crucial HTFs. H3K27 methylation dynamics at the HTFs was antagonistically regulated by EZH2 and JMJD3. Preliminary evidence suggests a role of JMJD3 in removal of H3K27me3 mark from promoters of HTFs, thus activating epigenetically poised hepatic genes in BMPCs prior to partial nuclear reprogramming. The importance of JMJD3 in reprogramming of BMPCs to hepatic phenotype was confirmed by inhibiting catalytic function of the enzyme using small molecule GSK-J4. Our results propose a potential role of JMJD3 in lineage conversion of BM cells into hepatic lineage.

Bone marrow stem cell therapy partially ameliorates pathological consequences in livers of mice expressing mutant human α1-antitrypsin.

Alpha-1-antitrypsin (AAT) deficiency (AATD) is a genetic disease, caused by mutation of the AAT gene. Accumulation of mutated AAT protein aggregates in hepatocytes leads to endoplasmic reticulum stress, resulting in impairment of liver functions and, in some cases, hepatocellular carcinoma, whereas decline of AAT levels in sera is responsible for pulmonary emphysema. In advanced liver disease, the only option for treatment is liver transplantation, whereas AAT replacement therapy is therapeutic for emphysema. Given that hepatocytes are the primary affected cells in AATD, we investigated whether transplantation of bone marrow (BM)-derived stem cells in transgenic mice expressing human AATZ (the Z variant of AAT) confers any competitive advantages compared to host cells that could lead to pathological improvement. Mouse BM progenitors and human mesenchymal stem cells (MSCs) appeared to contribute in replacement of 40% and 13% host hepatocytes, respectively. Transplantation of cells resulted in decline of globule-containing hepatocytes, improvement in proliferation of globule-devoid hepatocytes from the host-derived hepatocytes, and apparently, donor-derived cells. Further analyses revealed that transplantation partially improves liver pathology as reflected by inflammatory response, fibrosis, and apoptotic death of hepatocytes. Cell therapy was also found to improve liver glycogen storage and sera glucose level in mice expressing human AATZ mice. These overall improvements in liver pathology were not restricted to transplantation of mouse BM cells. Preliminary results also showed that following transplantation of human BM-derived MSCs, globule-containing hepatocytes declined and donor-derived cells expressed human AAT protein. CONCLUSION: These results suggest that BM stem cell transplantation may be a promising therapy for AATD-related liver disease. (Hepatology 2017;65:1319-1335).

Genetic screening of THAP1 in primary dystonia patients of India.

BACKGROUND: Primary Dystonia is a common movement disorder manifested by dystonic symptoms only. DYT6, a major genetic factor, plays a significant role in primary pure dystonia pathogenesis. In this study we analyzed THAP1 (DYT 6) gene in primary pure dystonia patients, which has been widely studied in other populations but not in Indians. METHODS: The study cohort contained 227 index primary pure dystonia patients with the involvement of cervical region and 254 neurologically control individuals collected from East Indian population. All three exons of THAP1 and their flanking sequences, including exon-intron boundaries, were screened by PCR, DNA sequencing and/or RFLP analysis. RESULTS: A total of three nucleotide variants were detected, which include a reported missense mutation (c.427 A>G; p.Met143Val) in a juvenile onset generalized dystonia patient, a novel frameshift deletion mutation (c.208-209 ΔAA; p.K70VfsX15) in a juvenile onset cervical dystonia patient and a rare variant in 3' UTR of THAP1 (c.*157 T>C) in an adult-onset blepharospasm patient. In addition, two SNPs (rs71521601 and rs111989331) were detected both in the patients and controls with the major allele of the latter being significantly over represented in the patients. CONCLUSIONS: Our study suggests that the THAP1 is likely to have a causative role in the pathogenesis of Indian primary pure dystonia patients. Though the phenotypic spectrum is extensively diverse, the cervical involvement with dystonic tremor and speech problem is common amongst the patients harboring mutations.