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Retinal Müller glial cells have the potential of neurogenic retinal progenitor cells, and could reprogram into retinal-specific cell types such as photoreceptor cells. How to promote the differentiation of Müller cells into photoreceptor cells represents a promising therapy strategy for retinal degeneration diseases. This study aimed to enhance the transdifferentiation of rat Müller cells-derived retinal stem cells (MC-RSCs) into photoreceptor-like cells and explore the signalling mechanism. We dedifferentiated rat Müller cells into MC-RSCs which were infected with Otx2 overexpression lentivirus or control. The positive rate of photoreceptor-like cells among MC-RSCs treated with Otx2 overexpression lentivirus was significantly higher compared to control. Furthermore, pre-treatment with Crx siRNA, Nrl siRNA, or GSK-3 inhibitor SB-216763 reduced the positive rate of photoreceptor-like cells among MC-RSCs treated with Otx2 overexpression lentivirus. Finally, Otx2 induced photoreceptor precursor cells were injected into subretinal space of N-methyl-N-nitrosourea induced rat model of retinal degeneration and partially recovered retinal degeneration in the rats. In conclusion, Otx2 enhances transdifferentiation of MC-RSCs into photoreceptor-like cells and this is associated with the inhibition of Wnt signalling. Otx2 is a potential target for gene therapy of retinal degenerative diseases.

Recently, we demonstrated that a fully differentiated tissue developed on a ventricular septal occluder that had been implanted due to infarct-related septum rupture. We suggested that this tissue originated from circulating stem cells. The aim of the present study was to evaluate this hypothesis and to investigate the physiological differentiation and transdifferentiation potential of circulating stem cells. We developed an animal model in which a freely floating membrane was inserted into each the left ventricle and the descending aorta. Membranes were removed after pre-specified intervals of 3 days, and 2, 6 and 12 weeks; the newly developed tissue was evaluated using quantitative RT-PCR, immunohistochemistry and in situ hybridization. The contribution of stem cells was directly evaluated in another group of animals that were by treated with granulocyte macrophage colony-stimulating factor (GM-CSF) early after implantation. We demonstrated the time-dependent generation of a fully differentiated tissue composed of fibroblasts, myofibroblasts, smooth muscle cells, endothelial cells and new blood vessels. Cells differentiated into early cardiomyocytes on membranes implanted in the left ventricles but not on those implanted in the aortas. Stem cell mobilization with GM-CSF led to more rapid tissue growth and differentiation. The GM-CSF effect on cell proliferation outlasted the treat ment period by several weeks. Circulating stem cells contributed to the development of a fully differentiated tissue on membranes placed within the left ventricle or descending aorta under physiological conditions. Early cardiomyocyte generation was identified only on membranes positioned within the left ventricle.

Liver fibrosis is the excessive accumulation of extracellular matrix proteins in response to the inflammatory response that accompanies tissue injury, which at an advanced stage can lead to cirrhosis and even liver failure. This study investigated the role of the CXC chemokine CXCL6 (GCP-2) in liver fibrosis. The expression of CXCL6 was found to be elevated in the serum and liver tissue of high stage liver fibrosis patients. Furthermore, treatment with CXCL6 (100 ng/mL) stimulated the phosphorylation of EGFR and the expression of TGF-β in cultured Kupffer cells (KCs). Although treatment with CXCL6 directly did not activate the hepatic stellate cell (HSC) line, HSC-T6, HSCs cultured with media taken from KCs treated with CXCL6 or TGF-β showed increased expression of α-SMA, a marker of HSC activation. CXCL6 was shown to function via the SMAD2/BRD4/C-MYC/EZH2 pathway by enhancing the SMAD3-BRD4 interaction and promoting direct binding of BRD4 to the C-MYC promoter and CMY-C to the EZH2 promoter, thereby inducing profibrogenic gene expression in HSCs, leading to activation and transdifferentiation into fibrogenic myofibroblasts. These findings were confirmed in a mouse model of CCl4 -induced chronic liver injury and fibrosis in which the levels of CXCL6 and TGF-β in serum and the expression of α-SMA, SMAD3, BRD4, C-MYC, and EZH2 in liver tissue were increased. Taken together, our results reveal that CXCL6 plays an important role in liver fibrosis through stimulating the release of TGF-β by KCs and thereby activating HSCs.

It was shown that endothelial progenitor cells (EPCs) have bidirectional differentiation potential and thus perform different biological functions. The purpose of this study was to investigate the effects of slight up-regulation of the Kir2.1 channel on EPC transdifferentiation and the potential mechanism on cell function and transformed cell type. First, we found that the slight up-regulation of Kir2.1 expression promoted the expression of the stem cell stemness factors ZFX and NS and inhibited the expression of senescence-associated β-galactosidase. Further studies showed the slightly increased expression of Kir2.1 could also improve the expression of pericyte molecular markers NG2, PDGFRβ and Desmin. Moreover, adenovirus-mediated Kir2.1 overexpression had an enhanced contractile response to norepinephrine of EPCs. These results suggest that the up-regulated expression of the Kir2.1 channel promotes EPC transdifferentiation into a pericyte phenotype. Furthermore, the mechanism of EPC transdifferentiation to mesenchymal cells (pericytes) was found to be closely related to the channel functional activity of Kir2.1 and revealed that this channel could promote EPC EndoMT by activating the Akt/mTOR/Snail signalling pathway. Overall, this study suggested that in the early stage of inflammatory response, regulating the Kir2.1 channel expression affects the biological function of EPCs, thereby determining the maturation and stability of neovascularization.

Prevention of infarct scar thinning and dilatation and stimulation of scar contracture can prevent progressive heart failure. Since microRNA 145 (miR-145) plays an important role in cardiac fibroblast response to wound healing and cardiac repair after an myocardial infarction (MI), using a miR-145 knock-out (KO) mouse model, we evaluated contribution of down-regulation of miR-145 to cardiac fibroblast and myofibroblast function during adverse cardiac remodelling. Cardiac function decreased more and the infarct size was larger in miR-145 KO than that in WT mice after MI and this phenomenon was accompanied by a decrease in cardiac fibroblast-to-myofibroblast differentiation. Quantification of collagen I and α-SMA protein levels as well as wound contraction revealed that transdifferentiation of cardiac fibroblasts into myofibroblasts was lower in KO than WT mice. In vitro restoration of miR-145 induced more differentiation of fibroblasts to myofibroblasts and this effect involved the target genes Klf4 and myocardin. MiR-145 contributes to infarct scar contraction in the heart and the absence of miR-145 contributes to dysfunction of cardiac fibroblast, resulting in greater infarct thinning and dilatation. Augmentation of miR-145 could be an attractive target to prevent adverse cardiac remodelling after MI by enhancing the phenotypic switch of cardiac fibroblasts to myofibroblasts.

Acute respiratory distress syndrome/acute lung injury (ARDS/ALI) is histologically characterized by extensive alveolar barrier disruption and excessive fibroproliferation responses. Protectin DX (PDX) displays anti-inflammatory and potent inflammation pro-resolving actions. We sought to investigate whether PDX attenuates LPS (lipopolysaccharide)-induced lung injury via modulating epithelial cell injury repair, apoptosis and fibroblasts activation. In vivo, PDX was administered intraperitoneally (IP) with 200 ng/per mouse after intratracheal injection of LPS, which remarkedly stimulated proliferation of type II alveolar epithelial cells (AT II cells), reduced the apoptosis of AT II cells, which attenuated lung injury induced by LPS. Moreover, primary type II alveolar cells were isolated and cultured to assess the effects of PDX on wound repair, apoptosis, proliferation and transdifferentiation in vitro. We also investigated the effects of PDX on primary rat lung fibroblast proliferation and myofibroblast differentiation. Our result suggests PDX promotes primary AT II cells wound closure by inducing the proliferation of AT II cells and reducing the apoptosis of AT II cells induced by LPS, and promotes AT II cells transdifferentiation. Furthermore, PDX inhibits transforming growth factor-β1 (TGF-β1 ) induced fibroproliferation, fibroblast collagen production and myofibroblast transformation. Furthermore, the effects of PDX on epithelial wound healing and proliferation, fibroblast proliferation and activation partly via the ALX/ PI3K signalling pathway. These data present identify a new mechanism of PDX which targets the airway epithelial cell and fibroproliferation are potential for treatment of ARDS/ALI.

Vascular calcification (VC) is a pathological process underpinning major cardiovascular conditions and has attracted public attention due to its high morbidity and mortality. Chronic kidney disease (CKD) is a common disease related to VC. Ginsenoside Rb1 (Rb1) has been reported to protect the cardiovascular system against vascular diseases, yet its role in VC and the underlying mechanisms remain unclear. In this study, we established a CKD-associated VC rat model and a β-glycerophosphate (β-GP)-induced vascular smooth muscle cell (VSMC) calcification model to investigate the effects of Rb1 on VC. Our results demonstrated that Rb1 ameliorated calcium deposition and VSMC osteogenic transdifferentiation both in vivo and in vitro. Rb1 treatment inhibited the Wnt/β-catenin pathway by activating peroxisome proliferator-activated receptor-γ (PPAR-γ), and confocal microscopy was used to show that Rb1 inhibited β-catenin nuclear translocation in VSMCs. Furthermore, SKL2001, an agonist of the Wnt/β-catenin pathway, compromised the vascular protective effect of Rb1. GW9662, a PPAR-γ antagonist, reversed Rb1's inhibitory effect on β-catenin. These results indicate that Rb1 exerted anticalcific properties through PPAR-γ/Wnt/β-catenin axis, which provides new insights into the potential theraputics of VC.

Transforming growth factor-β (TGF-β) becomes rapidly activated in the infarcted heart. Hence, TGF-β-mediated persistent activation of cardiac fibroblasts (CFs) and exaggerated fibrotic responses may result in adverse cardiac remodelling and heart failure. Additionally, peptidylarginine deiminase 4 (PAD4) was described to be implicated in organ fibrosis. Here, we investigated the impact of PAD4 on CF function and myofibroblast transdifferentiation in vitro. The expression of fibrosis-related genes was largely similar in cultured WT and PAD4-/- CFs of passage 3, although collagen III was reduced in PAD4-/- CFs. Exposure to TGF-β inhibited proliferation and increased contractile activity and migration of WT CFs, but not of PAD4-/- CFs. However, under baseline conditions, PAD4-/- CFs showed comparable functional characteristics as TGF-β-stimulated WT CFs. Although the SMAD-dependent TGF-β pathway was not disturbed in PAD4-/- CFs, TGF-β failed to activate protein kinase B (Akt) and signal transducer and activator of transcription 3 (STAT3) in these cells. Similar results were obtained in WT CFs treated with the PAD4 inhibitor Cl-amidine. Abrogated Akt activation was associated with diminished levels of phosphorylated, inactive glycogen synthase kinase-3β (GSK-3β). Consequently, PAD4-/- CFs did not upregulate collagen I and α-smooth muscle actin (α-SMA) expression after TGF-β treatment. Thus, PAD4 is substantially involved in the regulation of non-canonical TGF-β signalling and may represent a therapeutic target for the treatment of adverse cardiac remodelling.

MafA is a pancreatic transcriptional factor that controls β-cell-specific transcription of the insulin gene. However, the role of MafA in the regulation of pancreatic transdifferentiation and reprogramming in human stem cells is still unclear. In this study, we investigate the role of MafA in placenta-derived multipotent stem cells (PDMSCs) that constitutively expressed Oct-4 and Nanog. PDMSCs were isolated and transfected with MafA using a lentivector. Our results showed that overexpression of MafA in PDMSCs significantly up-regulated the expression of pancreatic development-related genes (Sox17, Foxa2, Pdx1 and Ngn3). Microarray analysis suggested that the gene expression profile of MafA-overexpressing PDMSCs was similar to that of pancreas and islet tissues. MafA increased the expression levels of the mRNAs of NKx2.2, Glut2, insulin, glucagons and somatostatin, and further facilitated the differentiation of PDMSCs into insulin(+) cells. The glucose-stimulated responses to insulin and c-peptide production in MafA-overexpressing PDMSCs were significantly higher than in PDMSCs with vector control. Our results indicated that MafA-overexpressing PDMSCs were more resistant to oxidative damage and oxidative damage-induced apoptosis than PDMSCs carrying the vector control were. Importantly, the expression of MafA in PDMSCs xenotransplanted into immunocompromised mice improved the restoration of blood insulin levels to control values and greatly prolonged the survival of graft cells in immunocompromised mice with STZ-induced diabetes. In summary, these data suggest that MafA plays a novel role in the reprogramming of stem cells into pancreatic β-progenitors, promotes the islet-like characteristics of PDMSCs, as well as functionally enhances insulin production to restore the regulation of blood glucose levels in transplanted grafts.