Studies have highlighted the importance of histone deacetylase (HDAC)-mediated epigenetic processes in the development of diabetic complications. Inhibitors of HDAC are a novel class of therapeutic agents in diabetic nephropathy, but currently available inhibitors are mostly nonselective inhibit multiple HDACs, and different HDACs serve very distinct functions. Therefore, it is essential to determine the role of individual HDACs in diabetic nephropathy and develop HDAC inhibitors with improved specificity. First, we identified the expression patterns of HDACs and found that, among zinc-dependent HDACs, HDAC2/4/5 were upregulated in the kidney from streptozotocin-induced diabetic rats, diabetic db/db mice, and in kidney biopsies from diabetic patients. Podocytes treated with high glucose, advanced glycation end products, or transforming growth factor-β (common detrimental factors in diabetic nephropathy) selectively increased HDAC4 expression. The role of HDAC4 was evaluated by in vivo gene silencing by intrarenal lentiviral gene delivery and found to reduce renal injury in diabetic rats. Podocyte injury was associated with suppressing autophagy and exacerbating inflammation by HDAC4-STAT1 signaling in vitro. Thus, HDAC4 contributes to podocyte injury and is one of critical components of a signal transduction pathway that links renal injury to autophagy in diabetic nephropathy.

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Guangxi Huanjiang Xiang pig is a unique miniature pig strain that is originally from Huanjiang Maonan Autonomous County of Guangxi province, China, and shows great potential in agricultural and biomedical research. Although cloning and genetic modification of this pig would enhance its application value, cloning of this strain has not yet been reported. We sought to establish appropriate cloning procedures and produce transgenic embryos in Huanjiang Xiang pigs through the following methods. We isolated fibroblasts from tails of Huanjiang Xiang pig and genetically modified them using Xfect transfection. Fibroblasts, either in non-transgenic or transgenic forms, were used as donor cells for reconstructed embryos by somatic cell nuclear transfer (SCNT), and in vitro development was monitored after the reconstruction. We found no difference in blastocyst formation rate between non-transgenic and transgenic embryos (10.8% vs. 10.3%; P ≥ 0.05). In addition, we tested whether Scriptaid, a widely used histone deacetylase inhibitor, could enhance the in vitro development of Huanjiang Xiang pig cloned embryos. Treatment with 500 nM Scriptaid for 16 h post-activation significantly increased the blastocyst formation rate (26.1% vs. 10.8% for non-transgenic nuclear transfer groups with vs. without the Scriptaid treatment and 28.5% vs. 10.3% for transgenic nuclear transfer groups with vs. without the Scriptaid treatment; P < 0.05). This study provided a basis for further generation of cloned and transgenically cloned Huanjiang Xiang pigs used in agricultural and biomedical research.

Prolonged culture of metaphase II oocytes is an in vitro aging process that compromises oocyte quality. We tested whether melatonin preserves epigenetic modifications in oocytes after prolonged culture. The porcine oocytes were maturated in vitro for 44 h, and then metaphase II oocytes were continuously cultured in medium supplemented with or without melatonin for 24 h. We found that the parthenogenetic blastocyst formation rate of prolonged-culture oocytes was lower than in fresh oocytes. We further observed that methylation at H3K4me2 and H3K27me2 of oocytes enhanced after prolonged culture. However, 5mc fluorescence intensity was lower in prolonged-culture oocytes than in fresh oocytes. Moreover, the promoter of the imprinted gene NNAT exhibited a higher level of DNA methylation in prolonged-culture oocytes than in fresh oocytes, which was associated with a reduced expression level and glucose uptake capability. Conversely, melatonin improved blastocyst formation rate and preserved histone and DNA methylation modifications, as well as NNAT function in the oocytes after prolonged culture. Notably, DNA methyltransferase inhibitor 5-aza significantly attenuated the protective role of melatonin on genomic DNA methylation. In summary, our results revealed that epigenetic modifications are disrupted in oocytes after prolonged culture, but the changes are reversed by melatonin.

Somatic cloning, also known as somatic cell nuclear transfer (SCNT), is a promising technology which has been expected to rapidly extend the population of elaborately selected breeding boars with superior production performance. Chinese Guike No. 1 pig breed is a novel swine specialized strain incorporated with the pedigree background of Duroc and Chinese Luchuan pig breeds, thus inherits an excellent production performance. The present study was conducted to establish somatic cloning procedures of adult breeding boars from the Chinese Guike No. 1 specialized strain. Ear skin fibroblasts were first isolated from a three-year-old Chinese Guike No. 1 breeding boar, and following that, used as donor cell to produce nuclear transfer embryos. Such cloned embryos showed full in vitro development and with the blastocyst formation rate of 18.4 % (37/201, three independent replicates). Finally, after transferring of 1187 nuclear transfer derived embryos to four surrogate recipients, six live piglets with normal health and development were produced. The overall cloning efficiency was 0.5 % and the clonal provenance of such SCNT derived piglets was confirmed by DNA microsatellite analysis. All of the cloned piglets were clinically healthy and had a normal weight at 1 month of age. Collectively, the first successful cloning of an adult Chinese Guike No. 1 breeding boar may lay the foundation for future improving the pig production industry.

It is widely accepted that infusion of mesenchymal stem cells (MSCs) ameliorates hyperglycemia by alleviating insulin resistance in rats with type 2 diabetes mellitus (T2D). However, the detailed underlying mechanisms are not clearly defined. Mitsugumin 53 (MG53) is an E3 ligase that has recently been implicated in the aggravation of insulin resistance by promoting the ubiquitinoylation of insulin receptor substrate‑1 (IRS‑1) in skeletal muscles. It was therefore hypothesized that MG53 may be involved in MSC‑mediated therapeutic effects on insulin resistance. To test this hypothesis, in the present study, T2D rat models were induced by a high‑fat diet combined with streptozotocin administration and MSC infusion was performed four times (once every 2 weeks for 8 weeks). The therapeutic effects of MSC infusion on insulin resistance were evaluated and the effect on the expression of MG53 and insulin receptor signaling elements in skeletal muscle was also investigated by immunofluorescence staining and western blotting. The results demonstrated that MSC infusion ameliorated hyperglycemia and insulin resistance in T2D rats. Furthermore, MSC infusion inhibited MG53 elevation and reversed the decreases in glucose transporter type 4, insulin receptor, IRS‑1 and phosphorylated‑AKT levels in the skeletal muscle of T2D rats. These results indicated that MSC infusion has therapeutic effects in rats and that MG53 in skeletal muscle may be a promising novel therapeutic target protein for MSC‑mediated amelioration of insulin resistance in T2D.

As a natural plant‐derived antitoxin, resveratrol possesses several pharmacological activities. This study aimed to evaluate the effects of resveratrol addition on nuclear maturation, oocyte quality during in vitro maturation (IVM) of porcine oocytes and subsequent early embryonic development following somatic cell nuclear transfer (SCNT). Our experiments showed that the treatment of porcine oocytes with 5 µM resveratrol during IVM resulted in the highest rate of the first polar body extrusion. Treatment of oocytes with resveratrol had no influence on cytoskeletal dynamics, whereas it significantly increased glucose uptake ability compared to the control oocytes. Oocytes matured with 5 μM resveratrol displayed significantly lower intracellular reactive oxygen species (ROS) levels and higher relative mRNA expression levels of the genes encoding such antioxidant enzymes as catalase (CAT) and superoxide dismutase 1 (SOD1). In addition, resveratrol also prevented onset and progression of programmed cell death in porcine oocytes, which was confirmed by significant upregulation of the anti‐apoptotic B‐cell lymphoma 2 (BCL‐2) gene and significant downregulation of the pro‐apoptotic BCL2‐associated X (BAX) gene. Furthermore, the blastocyst rates and the blastocyst cell numbers in cloned embryos derived from the oocytes that had matured in the presence of 5 μM resveratrol were significantly increased. In conclusion, supplementation of IVM medium with 5 μM resveratrol improves the quality of porcine oocytes by protecting them from oxidative damage and apoptosis, which leads to the production of meiotically matured oocytes exhibiting enhanced developmental potential following SCNT.