Autophagy, a type II programmed cell death, is essential for cell survival under stress, e.g. lung injury, and bone marrow-derived mesenchymal stem cells (BM-MSCs) have g... More
Autophagy, a type II programmed cell death, is essential for cell survival under stress, e.g. lung injury, and bone marrow-derived mesenchymal stem cells (BM-MSCs) have great potential for cell therapy. However, the mechanisms underlying the BM-MSC activation of autophagy to provide a therapeutic effect in ischaemia/reperfusion-induced lung injury (IRI) remain unclear. Thus, we investigate the activation of autophagy in IRI following transplantation with BM-MSCs. Seventy mice were pre-treated with BM-MSCs before they underwent lung IRI surgery in vivo. Human pulmonary micro-vascular endothelial cells (HPMVECs) were pre-conditioned with BM-MSCs by oxygen-glucose deprivation/reoxygenation (OGD) in vitro. Expression markers for autophagy and the phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) signalling pathway were analysed. In IRI-treated mice, administration of BM-MSCs significantly attenuated lung injury and inflammation, and increased the level of autophagy. In OGD-treated HPMVECs, co-culture with BM-MSCs attenuated endothelial permeability by decreasing the level of cell death and enhanced autophagic activation. Moreover, administration of BM-MSCs decreased the level of PI3K class I and p-Akt while the expression of PI3K class III was increased. Finally, BM-MSCs-induced autophagic activity was prevented using the inhibitor LY294002. Administration of BM-MSCs attenuated lung injury by improving the autophagy level via the PI3K/Akt signalling pathway. These findings provide further understanding of the mechanisms related to BM-MSCs and will help to develop new cell-based therapeutic strategies in lung injury. Less
Background: To determine the effect of different statins on the induction of diabetes mellitus. Materials and methods: Four statins (atorvastatin, pravastatin, rosuvastat... More
Background: To determine the effect of different statins on the induction of diabetes mellitus. Materials and methods: Four statins (atorvastatin, pravastatin, rosuvastatin, and pitavastatin) were used. Cytotoxicity, insulin secretion, glucose-stimulated insulin secretion, and G0/G1 phase cell cycle arrest were investigated in human pancreas islet β cells, and glucose uptake and signaling were studied in human skeletal muscle cells (HSkMCs). Results: Human pancreas islet β cells treated with 100 nM atorvastatin, pravastatin, rosuvastatin, and pitavastatin had reduced cell viability (32.12%, 41.09%, 33.96%, and 29.19%, respectively) compared to controls. Such cytotoxic effect was significantly attenuated by decreasing the dose to 10 and 1 nM, ranged from 1.46% to 17.28%. Cells treated with 100 nM atorvastatin, pravastatin, rosuvastatin, and pitavastatin had a reduction in the rate of insulin secretion rate by 34.07%, 30.06%, 26.78%, and 19.22%, respectively. The inhibitory effect was slightly attenuated by decreasing the dose to 10 and 1 nM, ranging from 10.84% to 29.60%. Insulin secretion stimulated by a high concentration of glucose (28 mmol/L) was significantly higher than a physiologic concentration of glucose (5.6 mmol/L) in all treatment groups. The glucose uptake rates at a concentration of 100 nM were as follows: atorvastatin (58.76%) < pravastatin (60.21%) < rosuvastatin (72.54%) < pitavastatin (89.96%). We also found that atorvastatin and pravastatin decreased glucose transporter (GLUT)-2 expression and induced p-p38 MAPK levels in human pancreas islet β cells. Atorvastatin, pravastatin, and rosuvastatin inhibited GLUT-4, p-AKT, p-GSK-3β, and p-p38 MAPK levels in HSkMCs. Conclusion: Statins similar but different degree of effects on pancreas islet β cells damage and induce insulin resistance in HSkMC. Keywords: glucose; human pancreas islet β cell; human skeletal muscle cells; insulin; statins. Less
Introduction
Type 1 interferon (IFN)-inducible genes and their inducible products are upregulated in dermatomyositis muscle. Of these, IFN-stimulated gene 15 (ISG15) is o... More
Introduction
Type 1 interferon (IFN)-inducible genes and their inducible products are upregulated in dermatomyositis muscle. Of these, IFN-stimulated gene 15 (ISG15) is one of the most upregulated, suggesting its possible involvement in the pathogenesis of this disease. To test this postulate, we developed a model of type 1 IFN mediated myotube toxicity and assessed whether or not downregulation of ISG15 expression prevents this toxicity.
Methods
Mouse myoblasts (C2C12 cell line) were cultured in the presence of type 1 or type 2 IFNs and ISG15 expression assessed by microarray analysis. The morphology of newly formed myotubes was assessed by measuring their length, diameter, and area on micrographs using imaging software. ISG15 expression was silenced through transfection with small interference RNA.
Results
Type 1 IFNs, especially IFN-beta, increased ISG15 expression in C2C12 cells and impaired myotube formation. Silencing of ISG15 resulted in knockdown of ISG15 protein, but without phenotypic rescue of myotube formation.
Discussion
IFN-beta affects myoblast differentiation ability and myotube morphology in vitro.These studies provide evidence that ISG15, which is highly upregulated in dermatomyositis muscle, does not appear to play a key role in IFN-beta-mediated C2C12 myoblast cell fusion. Less
Aim: Non-thyroidal illness syndrome (NTIS) is related to changes in thyroid hormone (TH) physiology. Skeletal muscle (SM) plays a major role in metabolism, and TH regulat... More
Aim: Non-thyroidal illness syndrome (NTIS) is related to changes in thyroid hormone (TH) physiology. Skeletal muscle (SM) plays a major role in metabolism, and TH regulates SM phenotype and metabolism. We aimed to characterize the SM of non-septic shock NTIS patients in terms of: i) expression of genes and proteins involved in TH metabolism and actions; and ii) NFKB's pathway activation, a responsible factor for some of the phenotypic changes in NTIS. We also investigated whether the patient's serum can induce in vitro the effects observed in vivo. Methods: Serum samples and SM biopsies from 14 patients with non-septic shock NTIS and 11 controls. Gene and protein expression and NFKB1 activation were analyzed by quantitative PCR and immunoblotting. Human SM cell (HSkMC) cultures to investigate the effects of patient's serum on TH action mediators. Results: Patients with non-septic shock NTIS showed higher levels of pro-inflammatory cytokines than controls. Expression of TRβ (THRB), TRα1 (THRA), and retinoid X receptor γ (RXRG) was decreased in NTIS patients. RXRA gene expression was higher, but its protein was lower in NTIS than controls, suggesting the existence of a post-transcriptional mechanism that down-regulates protein levels. NFKB1 pathway activation was not different between NTIS and control patients. HSkMC incubated with patient's serum increased TH receptor and RXRG gene expression after 48 h. Conclusions: Patients with non-septic shock NTIS showed decreased expression of TH receptors and RXRs, which were not related to increased activation of the NFKB1 pathway. These findings could not be replicated in cultures of HSkMCs incubated in the patient's serum. Less
Objective: We investigated interferon-stimulated gene 15 (ISG15), a poorly understood ubiquitin-like modifier, and its enzymatic pathway in dermatomyositis (DM), an autoi... More
Objective: We investigated interferon-stimulated gene 15 (ISG15), a poorly understood ubiquitin-like modifier, and its enzymatic pathway in dermatomyositis (DM), an autoimmune disease primarily involving muscle and skin. Methods: We generated microarray data measuring transcript abundance for approximately 18,000 genes in each of 113 human muscle biopsy specimens, and studied biopsy specimens and cultured skeletal muscle using immunohistochemistry, immunoblotting proteomics, real-time quantitative polymerase chain reaction, and laser-capture microdissection. Results: Transcripts encoding ISG15-conjugation pathway proteins were markedly upregulated in DM with perifascicular atrophy (DM-PFA) muscle (ISG15 339-fold, HERC5 62-fold, and USP18 68-fold) compared with 99 non-DM samples. Combined analysis with publicly available microarray datasets showed that >50-fold ISG15 transcript elevation had 100% sensitivity and specificity for 28 biopsies from adult DM-PFA and juvenile DM patients compared with 199 muscle samples from other muscle diseases. Free ISG15 and ISG15-conjugated proteins were only found on immunoblots from DM-PFA muscle. Cultured human skeletal muscle exposed to type 1 interferons produced similar transcripts and ISG15 protein and conjugates. Laser-capture microdissection followed by proteomic analysis showed deficiency of titin in DM perifascicular atrophic myofibers. Interpretation: A large-scale microarray study of muscle samples demonstrated that among a diverse group of muscle diseases DM was uniquely associated with upregulation of the ISG15 conjugation pathway. Exposure of human skeletal muscle cell culture to type 1 interferons produced a molecular picture highly similar to that seen in human DM muscle. Perifascicular atrophic myofibers in DM were deficient in a number of skeletal muscle proteins including titin. Less
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