WRN loss accelerates abnormal adipocyte metabolism in Werner syndrome.

BACKGROUND: Metabolic dysfunction is one of the main symptoms of Werner syndrome (WS); however, the underlying mechanisms remain unclear. Here, we report that loss of WRN accelerates adipogenesis at an early stage both in vitro (stem cells) and in vivo (zebrafish). Moreover, WRN depletion causes a transient upregulation of late-stage of adipocyte-specific genes at an early stage. METHODS: In an in vivo study, we generated wrn-/- mutant zebrafish and performed histological stain and Oil Red O staining to assess the fat metabolism. In an in vitro study, we used RNA-seq and ATAC-seq to profile the transcriptional features and chromatin accessibility in WRN depleted adipocytes. Moreover, we performed ChIP-seq to further study the regulatory mechanisms of metabolic dysfunction in WS. RESULTS: Our findings show that mechanistically WRN deficiency causes SMARCA5 upregulation. SMARCA5 is crucial in chromatin remodeling and gene regulation. Additionally, rescuing WRN could normalize SMARCA5 expression and adipocyte differentiation. Moreover, we find that nicotinamide riboside (NR) supplementation restores adipocyte metabolism in both stem cells and zebrafish models. CONCLUSIONS: Our findings unravel a new mechanism for the influence of WRN in the early stage of adipogenesis and provide a possible treatment for metabolic dysfunction in WS. These data provide promising insights into potential therapeutics for ageing and ageing-related diseases.

WRN promotes bone development and growth by unwinding SHOX-G-quadruplexes via its helicase activity in Werner Syndrome.

Werner Syndrome (WS) is an autosomal recessive disorder characterized by premature aging due to mutations of the WRN gene. A classical sign in WS patients is short stature, but the underlying mechanisms are not well understood. Here we report that WRN is indispensable for chondrogenesis, which is the engine driving the elongation of bones and determines height. Zebrafish lacking wrn exhibit impairment of bone growth and have shorter body stature. We pinpoint the function of WRN to its helicase domain. We identify short-stature homeobox (SHOX) as a crucial and direct target of WRN and find that the WRN helicase core regulates the transcriptional expression of SHOX via unwinding G-quadruplexes. Consistent with this, shox-/- zebrafish exhibit impaired bone growth, while genetic overexpression of SHOX or shox expression rescues the bone developmental deficiency induced in WRN/wrn-null mutants both in vitro and in vivo. Collectively, we have identified a previously unknown function of WRN in regulating bone development and growth through the transcriptional regulation of SHOX via the WRN helicase domain, thus illuminating a possible approach for new therapeutic strategies.

The relationship between online learning and eye strain in college students during the COVID-19 outbreak / 中国学校卫生

Objective@#To describe online learning and eye strain situation of college students during the COVID-19 outbreak, to provide a scientific basis for guiding students eye health.@*Methods@#A self-filled electronic questionnaire survey through questionnaire star was administered to college students across China. Information about online learning and eye strain of 1 046 college students during the epidemic was collected in Hefei, Anhui Province from March 16 to 20, 2020. The univariate and multivariate Logistic regression analysis were performed to analyze the association between online learning and eye strain of college students.@*Results@#The rate of eye strain during online learning was 72.1%, totally of 68.4% in 421 male students and 74.6% in 625 female students. Boys with online learning time <6 h/d, slow internet access，difficulty in understanding online class reported higher rate of eye strain than girls( χ 2=17.36,8.72,7.02, P ＜0.05). Freshmen reported the highest rate of slow internet access occasionally and active online class( χ 2=15.26,16.11, P ＜0.05), junior students reported highest rate of online learning time <6 h/d, and easy understandable online class( χ 2=15.33,32.59, P ＜0.05), medical college students reported higher rate of slow internet access, inactive online class than non-medical college students( χ 2=11.79,11.03, P ＜0.05). Multivariate Logistic regression analysis showed that odds ratio( OR ) of eye strain in females was 1.40 (1.06-1.87), compared with males; the OR of eye strain were 1.43 (1.01-2.03) and 1.54 (1.10-2.15) in the groups with online learning time 6-<8 h/d and ≥8 h/d, respectively, compared with the group with online learning time <6 h/d, the OR of eye strain in the groups with slow internet access was 2.28 (1.25-4.14), compared with students without slow internet access, the OR of eye strain in the capable to understand and difficult to understand group were 2.54 (1.73-3.74) and 5.40 (2.70-10.80) respectively, compared with the easy to understand group.@*Conclusion@#Female students, online learing time ≥ 8 h/d, slow internet access, difficult to understand class content were positively related with college students eye strain. Attention should be paid to the eye health of college students to reduce the adverse effects of online learning on vision.during the COVID-19 epidemic.

Genetic correction of Werner syndrome gene reveals impaired pro-angiogenic function and HGF insufficiency in mesenchymal stem cells.

WRN mutation causes a premature aging disease called Werner syndrome (WS). However, the mechanism by which WRN loss leads to progeroid features evident with impaired tissue repair and regeneration remains unclear. To determine this mechanism, we performed gene editing in reprogrammed induced pluripotent stem cells (iPSCs) derived from WS fibroblasts. Gene correction restored the expression of WRN. WRN+/+ mesenchymal stem cells (MSCs) exhibited improved pro-angiogenesis. An analysis of paracrine factors revealed that hepatocyte growth factor (HGF) was downregulated in WRN-/- MSCs. HGF insufficiency resulted in poor angiogenesis and cutaneous wound healing. Furthermore, HGF was partially regulated by PI3K/AKT signaling, which was desensitized in WRN-/- MSCs. Consistently, the inhibition of the PI3K/AKT pathway in WRN+/+ MSC resulted in reduced angiogenesis and poor wound healing. Our findings indicate that the impairment in the pro-angiogenic function of WS-MSCs is due to HGF insufficiency and PI3K/AKT dysregulation, suggesting trophic disruption between stromal and epithelial cells as a mechanism for WS pathogenesis.

Rapamycin efficiently promotes cardiac differentiation of mouse embryonic stem cells.

To investigate the effects of rapamycin on cardiac differentiation, murine embryonic stem cells (ESCs) were induced into cardiomyocytes by 10-4 M ascorbic acid (AA), 20 nM rapamycin alone or 0.01% solvent DMSO. We found that rapamycin alone was insufficient to initiate cardiomyogenesis. Then, the ESCs were treated with AA and rapamycin (20 nM) or AA and DMSO (0.01%) as a control. Compared with control, mouse ESCs (mESCs) treated with rapamycin (20 nM) and AA yielded a significantly higher percentage of cardiomyocytes, as confirmed by the percentage of beating embryonic bodies (EBs), the immunofluorescence and FACS analysis. Rapamycin significantly increased the expression of a panel of cardiac markers including Gata4, α-Mhc, ß-Mhc, and Tnnt2. Additionally, rapamycin enhanced the expression of mesodermal and cardiac transcription factors such as Mesp1, Brachyury T, Eomes, Isl1, Gata4, Nkx2.5, Tbx5, and Mef2c. Mechanistic studies showed that rapamycin inhibits Wnt/ß-catenin and Notch signaling but promotes the expression of fibroblast growth factor (Fgf8), Fgf10, and Nodal at early stage, and bone morphogenetic protein 2 (Bmp 2) at later stages. Sequential treatment of rapamycin showed that rapamycin promotes cardiac differentiation at the early and later stages. Interestingly, another mammalian target of rapamycin (mTOR) inhibitor Ku0063794 (1 µM) had similar effects on cardiomyogenesis. In conclusion, our results highlight a practical approach to generate cardiomyocytes from mESCs by rapamycin.

Cross-talk of SFRP4, integrin α1ß1, and Notch1 inhibits cardiac differentiation of P19CL6 cells.

Signaling pathways play an important role in cardiogenesis. Secreted frizzled-related protein 4 (SFRP4), a member of the Wnt family, contributes to adipogenesis and tumorigenesis. However, how SFRP4 participates in cardiogenesis and the detailed molecular mechanisms involved have not been elucidated. The aim of this work was to determine cross-talk between SFRP4, integrin α1ß1, and Notch1 during cardiac differentiation of P19CL6 cells. Using a well-established in vitro P19CL6 cell cardiomyocyte differentiation system, we found that SFRP4 inhibited P19CL6 cell cardiac differentiation via SFRP4 overexpression or knockdown. In addition, the SFRP4 overexpression augmented Notch1 and HES1 production. Further investigation demonstrated that SFRP4 bound to integrin α1ß1 to activate the focal adhesion kinase (FAK) pathway and that phosphorylated FAK Y397 (p-FAK Y397) aided Notch intracellular domain 1 (NICD1) nuclear translocation to form a p-FAK Y397-NICD1 complex that activated the Hes1 promoter. Taken together, the cross-talk between SFRP4, integrin α1ß1, and Notch1 suppresses the cardiac differentiation of P19CL6 cells.

SOX6 and PDCD4 enhance cardiomyocyte apoptosis through LPS-induced miR-499 inhibition.

Sepsis-induced cardiac apoptosis is one of the major pathogenic factors in myocardial dysfunction. As it enhances numerous proinflammatory factors, lipopolysaccharide (LPS) is considered the principal mediator in this pathological process. However, the detailed mechanisms involved are unclear. In this study, we attempted to explore the mechanisms involved in LPS-induced cardiomyocyte apoptosis. We found that LPS stimulation inhibited microRNA (miR)-499 expression and thereby upregulated the expression of SOX6 and PDCD4 in neonatal rat cardiomyocytes. We demonstrate that SOX6 and PDCD4 are target genes of miR-499, and they enhance LPS-induced cardiomyocyte apoptosis by activating the BCL-2 family pathway. The apoptosis process enhanced by overexpression of SOX6 or PDCD4, was rescued by the cardiac-abundant miR-499. Overexpression of miR-499 protected the cardiomyocytes against LPS-induced apoptosis. In brief, our results demonstrate the existence of a miR-499-SOX6/PDCD4-BCL-2 family pathway in cardiomyocytes in response to LPS stimulation.

Autophagy eliminates cytoplasmic ß-catenin and NICD to promote the cardiac differentiation of P19CL6 cells.

Autophagy plays important roles in adipogenesis and neuron development. However, how autophagy contributes to cardiac development is not well understood. The main aim of our study was to determine the association between autophagy and myocardial differentiation and its roles in this process. Using a well-established in vitro cardiomyocyte differentiation system, P19CL6 cells, we found that autophagy occurred from the early stage of cardiac differentiation. Blocking autophagy by knocking-down of autophagy-related gene Atg7 or Atg5 inhibited the cardiac differentiation of P19CL6 cells. Further investigation demonstrated that LC3 and P62 could form a complex with ß-catenin and NICD, respectively, and promoted the degradation of ß-catenin and NICD. Enhancing autophagy promoted the formation of complex, whereas blocking autophagy attenuated the degradation of ß-catenin and NICD. Taken together, autophagy could facilitate P19CL6 cells to complete the cardiac differentiation process through blocking Wnt and Notch signaling pathways.