摘要: Objectives Fibrotic cataract, including posterior capsule opacification (PCO) and anterior subcapsular cataract (ASC), renders millions of people visually impaired worldwide. However, the underlying mechanism remains poorly understood. Here, we report a miRNA-based regulatory pathway that controls pathological fibrosis of lens epithelium. Materials and methods Expression of miR-22-3p and histone deacetylase 6 (HDAC6) in normal and PCO patient samples were measured by qPCR. Human lens epithelial explants were treated with TGF-beta 2 in the presence or absence of miR-22-3p mimics or inhibitor. Cell proliferation was determined by MTS assay, and migration was tested by transwell assay. Expression of HDAC6 and EMT-related molecules were analysed by Western blot, qPCR and immunocytochemical experiments. Results We identify miR-22-3p as a downregulated miRNA targeting HDAC6 in LECs during lens fibrosis and TGF-beta 2 treatment. Mechanistically, gain- and loss-of-function experiments in human LECs and lens epithelial explants reveal that miR-22-3p prevents proliferation, migration and TGF-beta 2 induced EMT of LECs via targeting HDAC6 and thereby promoting alpha-tubulin acetylation. Moreover, pharmacological targeting of HDAC6 deacetylase with Tubacin prevents fibrotic opaque formation through increasing alpha-tubulin acetylation under TGF-beta 2 stimulated conditions in both human lens epithelial explants and the whole rat lenses. Conclusions These findings suggest that miR-22-3p prevents lens fibrotic progression by targeting HDAC6 thereby promoting alpha-tubulin acetylation. The 'miR-22-HDAC6-alpha-tubulin (de)acetylation' signalling axis may be therapeutic targets for the treatment of fibrotic cataract.

摘要: Background Hypoxia-inducible factors (HIFs) are thought to play important roles in the carcinogenesis and progression of VHL-deficient clear cell renal cell carcinoma (ccRCC). Methods The roles of HIF-1/2 alpha in VHL-deficient clear cell renal cell carcinoma were evaluated by bioinformatics analysis, immunohistochemistry staining and Kaplan-Meier survival analysis. The downstream genes that counteract the cancer-promoting effect of HIF were analysed by unbiased proteomics and verified by in vitro and in vivo assays. Results There was no correlation between the high protein level of HIF-1/2 alpha and the poor prognosis of ccRCC patients in our large set of clinical data. Furthermore, NDRG1 was found to be up-regulated by both HIF-1 alpha and -2 alpha at the cellular level and in ccRCC tissues. Intriguingly, the high NDRG1 expression was correlated with lower Furman grade, TNM stage and longer survival for ccRCC patients compared with the low NDRG1 expression. In addition, NDRG1 suppressed the expression of series oncogenes as well as the proliferation, metastasis and invasion of VHL-deficient ccRCC cells in vitro and vivo. Conclusions Our study demonstrated that HIF downstream gene of NDRG1 may counteract the cancer-promoting effect of HIF. These results provided evidence that NDRG1 may be a potential prognostic biomarker as well as a therapeutic target in ccRCC.

摘要: Objectives Most bone fracture heals through enchondral bone formation that relies on the involvement of periosteal progenitor cells. However, the identity of periosteal progenitor cells and the regulatory mechanism of their proliferation and differentiation remain unclear. The aim of this study was to investigate whetherGli1-Cre(ERT2)can identify a population of murine periosteal progenitor cells and the role of TGF-beta signalling in periosteal progenitor cells on fracture healing. Materials and methods Double heterozygous Gli1-CreER(T2);Rosa26-tdTomato(flox/wt) mice were sacrificed at different time points for tracing the fate of Gli1(+) cells in both intact and fracture bone.Gli1-CreER(T2)-mediatedTgfbr2knockout (Gli1-CreER(T2);Tgfbr2(flox/flox)) mice were subjected to fracture surgery. At 4, 7, 10, 14 and 21 days post-surgery, tibia samples were harvested for tissue analyses including mu CT, histology, real-time PCR and immunofluorescence staining. Results Through cell lineage-tracing experiments, we have revealed that Gli1-Cre(ERT2) can be used to identify a subpopulation of periosteal progenitor cells in vivo that persistently reside in periosteum and contribute to osteochondral elements during fracture repair. During the healing process, TGF-beta signalling is continually activated in the reparative Gli1(+)periosteal cells. Conditional knockout ofTgfbr2in these cells leads to a delayed and impaired enchondral bone formation, at least partially due to the reduced proliferation and chondrogenic and osteogenic differentiation of Gli1(+) periosteal cells. Conclusions TGF-beta signalling plays an essential role on fracture repair via regulating enchondral bone formation process of Gli1(+)periosteal cells.

摘要: Objectives Genetic engineering of human-induced pluripotent stem cell-derived neural stem cells (hiPSC-NSC) may increase the risk of genomic aberrations. Therefore, we asked whether genetic modification of hiPSC-NSCs exacerbates chromosomal abnormalities that may occur during passaging and whether they may cause any functional perturbations in NSCs in vitro and in vivo. Materials and Methods The transgenic cassette was inserted into the AAVS1 locus, and the genetic integrity of zinc-finger nuclease (ZFN)-modified hiPSC-NSCs was assessed by the SNP-based karyotyping. The hiPSC-NSC proliferation was assessed in vitro by the EdU incorporation assay and in vivo by staining of brain slices with Ki-67 antibody at 2 and 8 weeks after transplantation of ZFN-NSCs with and without chromosomal aberration into the striatum of immunodeficient rats. Results During early passages, no chromosomal abnormalities were detected in unmodified or ZFN-modified hiPSC-NSCs. However, at higher passages both cell populations acquired duplication of the entire long arm of chromosome 1, dup(1)q. ZNF-NSCs carrying dup(1)q exhibited higher proliferation rate than karyotypically intact cells, which was partly mediated by increased expression ofAKT3located on Chr1q. Compared to karyotypically normal ZNF-NSCs, cells with dup(1)q also exhibited increased proliferation in vivo 2 weeks, but not 2 months, after transplantation. Conclusions These results demonstrate that, independently of ZFN-editing, hiPSC-NSCs have a propensity for acquiring dup(1)q and this aberration results in increased proliferation which might compromise downstream hiPSC-NSC applications.

摘要: Objectives SIRT1 is an antioxidative factor, but its mechanism in methamphetamine (MA)-induced lung injury remains unclear. The purpose of this study is to determine whether MA can disrupt the integrity of alveolar epithelial barrier, whether SIRT1 is involved in MA-induced chronic lung injury and whether Resveratrol (Res) can protect the integrity of alveolar epithelial cells by regulating ROS to activate SIRT1/PTEN/p-Akt pathway. Materials and methods The rats were randomly divided into control group and MA group. Extracted lungs were detected by Western blot, HE staining and immunohistochemistry. The alveolar epithelial cells were treated with MA or/and Res, following by Western blot, LDH leakage assay and flow cytometry. MOE is used for bio-informatics prediction. Results Chronic exposure to MA can cause slower growth ratio of weight, increased RVI and induced lung injury including the reduced number of alveolar sacs and the thickened alveolar walls. MA-induced apoptosis was associated with SIRT1-related oxidative stress. Res suppressed ROS levels, activated SIRT1, negatively regulated PTEN, phosphorylated Akt, reduced LDH leakage, increased the expression of ZO-1 and E-cadherin and inhibited the apoptosis of alveolar epithelial cells to attenuate MA-induced higher permeability of alveolar epithelium. Conclusions MA disrupted the integrity of alveolar epithelial barrier. Res inhibited oxidative stress and reversed MA-induced higher permeability and apoptosis of alveolar epithelium by the activation of SIRT1/PTEN/p-Akt pathway.