iPSCs ameliorate hypoxia-induced autophagy and atrophy in C2C12 myotubes via the AMPK/ULK1 pathway

BACKGROUND: Duchenne muscular dystrophy (DMD) is an X-linked lethal genetic disorder for which there is no effective treatment. Previous studies have shown that stem cell transplantation into mdx mice can promote muscle regeneration and improve muscle function, however, the specific molecular mechanisms remain unclear. DMD suffers varying degrees of hypoxic damage during disease progression. This study aimed to investigate whether induced pluripotent stem cells (iPSCs) have protective effects against hypoxia-induced skeletal muscle injury. RESULTS: In this study, we co-cultured iPSCs with C2C12 myoblasts using a Transwell nested system and placed them in a DG250 anaerobic workstation for oxygen deprivation for 24 h. We found that iPSCs reduced the levels of lactate dehydrogenase and reactive oxygen species and downregulated the mRNA and protein levels of BAX/BCL2 and LC3II/ LC3I in hypoxia-induced C2C12 myoblasts. Meanwhile, iPSCs decreased the mRNA and protein levels of atrogin-1 and MuRF-1 and increased myotube width. Furthermore, iPSCs downregulated the phosphorylation of AMPKA and ULK1 in C2C12 myotubes exposed to hypoxic damage. CONCLUSIONS: Our study showed that iPSCs enhanced the resistance of C2C12 myoblasts to hypoxia and inhibited apoptosis and autophagy in the presence of oxidative stress. Further, iPSCs improved hypoxia-induced autophagy and atrophy of C2C12 myotubes through the AMPK/ULK1 pathway. This study may provide a new theoretical basis for the treatment of muscular dystrophy in stem cells.

MALAT1 is involved in type I IFNs-mediated systemic lupus erythematosus by up-regulating OAS2, OAS3, and OASL

Systemic lupus erythematosus (SLE) is an autoimmune disease associated with an aberrant activation of immune cells partly due to the dysfunction of cytokines such as type I interferons (IFNs). Long non-coding RNA MALAT1 has been found to play a pathogenic role in SLE; however, the underlying mechanisms are still poorly understood. Bioinformatics analysis showed the up-regulation of type I IFN downstream effectors OAS2, OAS3, and OASL (OAS-like) in CD4+ T cells, CD19+ B cells, and CD33+ myeloid cells in patients with active SLE compared to healthy participants. In this study, peripheral blood mononuclear cells (PBMCs), CD19+ B, and CD4+ T cells were isolated from active SLE patients and healthy participants. PCR was performed to quantify MALAT1, OAS2, OAS3, and OASL expression in immune cells. MALAT1, OAS2, OAS3, and OASL were knocked down in CD4+ T cells to investigate the regulatory effect of MALAT1 on the effectors and their involvement in type I IFNs-mediated inflammation. Results showed higher OAS2, OAS3, and OASL expression in active SLE patients. MALAT1 expression was positively correlated to OAS2, OAS3, and OASL expression in CD19+ B or CD4+ T cells. MALAT1 knockdown decreased OAS2, OAS3, and OASL expression. Treatment with IFN-α-2a increased the expression of TNF-α, IL-1β, and IFN-α in CD4+ T cells. However, knockdown of MALAT1, OAS2, OAS3, and OASL alone inhibited the effect of IFN-α-2a on TNF-α and IL-1β. This study suggested the involvement of MALAT1 in type I IFNs-mediated SLE by up-regulating OAS2, OAS3, and OASL.

A SNP in 5′ untranslated region of CD40 gene is associated with an increased risk of ischemic stroke in a Chinese population: a case-control study

Abstract Cluster of differentiation 40 (CD40), the receptor for CD154, is a member of the tumor necrosis factor (TNF) receptor superfamily. Several studies have been conducted to investigate the effect of the CD40 rs1883832 polymorphism on atherosclerotic disease in different population; however, inconsistent results were obtained. In this study, we investigated the association of four polymorphisms (rs1883832, rs13040307, rs752118 and rs3765459) of CD40 gene and their effect on CD40 expression with the risk of ischemic stroke (IS) in a Chinese population. Three hundred and eighty patients with IS and 450 control subjects were included in the study. The CD40 polymorphisms were discriminated by Snapshot SNP genotyping assay. Serum soluble CD40 (sCD40) levels were detected by ELISA. We found that the rs1883832CT and rs1883832TT genotypes were associated with an increased risk of IS compared with the rs1883832CC genotype (OR = 1.42, 95% CI: 1.03-1.95, p = 0.030 and OR = 1.91, 95% CI: 1.29-2.82, P = 0.001, respectively), and the rs1883832T allele was associated with a significantly increased risk of IS compared with rs1883832C allele (OR = 1.40, 95% CI: 1.15-1.70, P = 0.001). Elevated serum sCD40 levels were observed in patients with IS compared with the control gropu (P < 0.01). Individuals carrying the rs1883832TT or rs1883832CT genotypes showed significantly higher sCD40 levels compared with the rs1883832CC genotype in the IS group [(64.8 ± 25.4 pg/mL, TT = 94); (63.9 ± 24.3 pg/mL, CT = 185) vs (53.3 ± 22.5 pg/mL, CC = 101), P < 0.01]. The TCCA haplotype was associated with an increased risk of IS compared with the control group (OR = 2.10, 95% CI: 1.23-3.58, p = 0.005). However, we did not find a significant association between the other three polymorphisms and IS risk. In conclusion, after a comprehensive comparison with other studies, we confirmed that the rs1883832T allele but not the rs1883832C allele is associated with an increased risk of IS. The rs1883832 polymorphism may exert influences on abnormal CD40 expression in IS patients among the Chinese population.

MiR-20 Regulates Myocardiac Ischemia by Targeting KATP Subunit Kir6.1 / 华中科技大学学报（医学）（英德文版）

This study aimed to examine the functional role of microRNA-20 (miR-20) and its potential target,Kir6.1,in ischemic myocardiocytes.The expression of miR-20 was detected by real-time PCR.Myocardiocytes were stained with terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) reagent for apoptosis evaluation.Western blotting was used to detect the Kit6.1 protein in ischemic myocardiocytes transfected with miR-20 mimics or inhibitors.Luciferase reporter gene assay was performed to confirm the targeting effect of miR-20 on KCNJ8.The results showed that miR-20 was remarkably down-regulated,while the KATP subunit Kir6.1 was significantly up-regulated,during myocardial ischemia.The miR-20 overexpression promoted the apoptosis of ischemic myocardiocytes,but showed no such effect on normal cells.Under ischemic condition,myocardiocytes transfected with miR-20 mimics expressed less Kir6.1.On the contrary,inhibiting miR-20 increased the expression of Kir6.1 in the cells.Co-transfection of miR-20 mimics with the KCNJ8 3’-UTR plasmid into HEK293 cells consistently produced less luciferase activity than transfection of the plasmid alone.It was concluded that miR-20 may regulate myocardiac ischemia by targeting KATP subunit Kir6.1 to accelerate the cell apoptosis.Therefore miR-20 may serve as a therapeutic target for myocardial ischemic disease.

Genetic analysis and gene fine mapping of aroma in rice (Oryza sativa L. Cyperales, Poaceae)

We investigated inheritance and carried out gene fine mapping of aroma in crosses between the aromatic elite hybrid rice Oryza sativa indica variety Chuanxiang-29B (Ch-29B) and the non-aromatic rice O. sativa indica variety R2 and O. sativa japonica Lemont (Le). The F1 grains and leaves were non-aromatic while the F2 non-aroma to aroma segregation pattern was 3:1. The F3 segregation ratio was consistent with the expected 1:2:1 for a single recessive aroma gene in Ch-29B. Linkage analysis between simple sequence repeat (SSR) markers and the aroma locus for the aromatic F2 plants mapped the Ch-29B aroma gene to a chromosome 8 region flanked by SSR markers RM23120 at 0.52 cM and RM3459 at 1.23 cM, a replicate F2 population confirming these results. Three bacterial artificial chromosome (BAC) clones cover chromosome 8 markers RM23120 and RM3459. Our molecular mapping data from the two populations indicated that the aroma locus occurs in a 142.85 kb interval on BAC clones AP005301 or AP005537, implying that it might be the same gene reported by Bradbury et al (2005a; Plant Biotec J. 3:363-370). The flanking markers Aro7, RM23120 and RM3459 identified by us could greatly accelerate the efficiency and precision of aromatic rice breeding programs.