MicroRNA-26a alleviates tubulointerstitial fibrosis in diabetic kidney disease by targeting PAR4.

Our previous study found that miR-26a alleviates aldosterone-induced tubulointerstitial fibrosis (TIF). However, the effect of miR-26a on TIF in diabetic kidney disease (DKD) remains unclear. This study clarifies the role and possible mechanism of exogenous miR-26a in controlling the progression of TIF in DKD models. Firstly, we showed that miR-26a was markedly decreased in type 2 diabetic db/db mice and mouse tubular epithelial cells (mTECs) treated with high glucose (HG, 30 mM) using RT-qPCR. We then used adeno-associated virus carrying miR-26a and adenovirus miR-26a to enhance the expression of miR-26a in vivo and in vitro. Overexpressing miR-26a alleviated the TIF in db/db mice and the extracellular matrix (ECM) deposition in HG-stimulated mTECs. These protective effects were caused by reducing expression of protease-activated receptor 4 (PAR4), which involved in multiple pro-fibrotic pathways. The rescue of PAR4 expression reversed the anti-fibrosis activity of miR-26a. We conclude that miR-26a alleviates TIF in DKD models by directly targeting PAR4, which may provide a novel molecular strategy for DKD therapy.

High quality haplotype-resolved genome assemblies of Populus tomentosa Carr., a stabilized interspecific hybrid species widespread in Asia.

Populus has a wide ecogeographical range spanning the Northern Hemisphere, and interspecific hybrids are common. Populus tomentosa Carr. is widely distributed and cultivated in the eastern region of Asia, where it plays multiple important roles in forestry, agriculture, conservation, and urban horticulture. Reference genomes are available for several Populus species, however, our goals were to produce a very high quality de novo chromosome-level genome assembly in P. tomentosa genome that could serve as a reference for evolutionary and ecological studies of hybrid speciation throughout the genus. Here, combining long-read sequencing and Hi-C scaffolding, we present a high-quality, haplotype-resolved genome assembly. The genome size was 740.2 Mb, with a contig N50 size of 5.47 Mb and a scaffold N50 size of 46.68 Mb, consisting of 38 chromosomes, as expected with the known diploid chromosome number (2n = 2x = 38). A total of 59,124 protein-coding genes were identified. Phylogenomic analyses revealed that P. tomentosa is comprised of two distinct subgenomes, which we deomonstrate is likely to have resulted from hybridization between Populus adenopoda as the female parent and Populus alba var. pyramidalis as the male parent, with an origin of approximately 3.93 Ma. Although highly colinear, significant structural variation was found between the two subgenomes. Our study provides a valuable resource for ecological genetics and forest biotechnology.

Comparative transcriptomic analysis reveals the coordinated mechanisms of Populus × canadensis 'Neva' leaves in response to cadmium stress.

Cadmium (Cd), a heavy metal element has strong toxicity to living organisms. Excessive Cd accumulation directly affects the absorption of mineral elements, inhibits plant tissue development, and even induces mortality. Populus × canadensis 'Neva', the main afforestation variety planted widely in northern China, was a candidate variety for phytoremediation. However, the genes relieving Cd toxicity and increasing Cd tolerance of this species were still unclear. In this study, we employed transcriptome sequencing on two Cd-treated cuttings to identify the key genes involved in Cd stress responses of P. × canadensis 'Neva' induced by 0 (CK), 10 (C10), and 20 (C20) mg/L Cd(NO3)2 4H2O. We discovered a total of 2,656 (1,488 up-regulated and 1,168 down-regulated) and 2,816 DEGs (1,470 up-regulated and 1,346 down-regulated) differentially expressed genes (DEGs) between the CK vs C10 and CK vs C20, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses in response to the Cd stress indicated that many DEGs identified were involved in the catalytic activity, the oxidoreductase activity, the transferase activity, and the biosynthesis of secondary metabolites. Based on the enrichment results, potential candidate genes were identified related to the calcium ion signal transduction, transcription factors, the antioxidant defense system, and transporters and showed divergent expression patterns under the Cd stress. We also validated the reliability of transcriptome data with the real-time PCR. Our findings deeper the understanding of the molecular responsive mechanisms of P. × canadensis 'Neva' on Cd tolerance and further provide critical resources for phytoremediation applications.

[Bufalin inhibits PDGF-BB-induced mesangial cell proliferation via mediating gap junctional intercellular communication].

OBJECTIVE: To study the role and possible mechanisms of gap junctional intercellular communication (GJIC) involved in mesangial cell (MC) proliferation which could be inhibited by bufalin. METHODS: Rat mesangial cells were cultured in vitro. The effect of bufalin on platelet-derived growth factor-BB (PDGF-BB)-induced MC proliferation was evaluated by MTT assay. The function of GJIC was detected by Lucifer Yellow scrape loading and dye transfer (SLDT). mRNA levels of Cx43, Cx45 and Cx40 were measured by RT-PCR. Intracellular calcium concentrations ([Ca(2+)]i) were examined in laser scanning confocal microscopy after loading by Fura-3/AM. RESULTS: MTT indicated that bufalin could inhibited PDGF-BB-induced MC proliferation (P<0.01). Compared with the hormal control group, PDGF-BB inhibited GJIC function, increased the expression of Cx45 and Cx40 (P<0.01) without altering the Cx43 (P>0.05) in gene level and also increased [Ca(2+)]i. However, bufalin treatment enhanced GJIC function, decreased Cx45 mRNA and Cx40 mRNA expression (P<0.01), and reduced [Ca(2+)]i (P<0.01). CONCLUSIONS: Bufalin inhibits PDGF-BB-induced MC proliferation, and its possible mechanisms may be related to regulation of Cx45 and Cx40 expression in the gene level, reduction of [Ca(2+)]i and enhancement of GJIC function.

Attenuation of glomerular filtration barrier damage in adriamycin-induced nephropathic rats with bufalin: an antiproteinuric agent.

Proteinuria is an important risk factor for the progression and prognosis of chronic kidney disease. Bufalin, a cardiotonic steroid, has been shown to posses a variety of biological activities including cardiotonic, anaesthetic and antineoplastic activities, and regulate the immune response. This study investigated the effects of bufalin against proteinuria and glomerular filtration barrier damage in rats with adriamycin (ADR)-induced nephropathy. We compared the blood and urine biochemical indices and the histologic and ultrastructure of the glomerulus in ADR rats with and without the intervention of bufalin or prednisone. The transcription, expression and distribution of the podocyte-associated molecules were compared utilising RT-PCR, western blotting and immunohistochemical staining. We found that bufalin reduced the urinary protein excretion and optimised the lipidaemia of the ADR rats. Bufalin alleviated the removal of podocyte foot processes and attenuated the changes in nephrin, podocin and integrin-linked kinase (ILK) stainings in the glomerulus of the ADR rats. Bufalin notably decreased the expression of nephrin and ILK but inhibited the down-regulation of podocin in protein levels on the renal cortex of the ADR rats. Additionally, bufalin inhibited the up-regulation of podocin and ILK in mRNA levels but did not affect nephrin mRNA levels. These results suggest that bufalin could alleviate ADR-induced proteinuria by protecting the glomerular filtration barrier and may be a novel potential therapeutic agent for proteinuria-associated kidney disease.

Bufalin inhibits platelet-derived growth factor-BB-induced mesangial cell proliferation through mediating cell cycle progression.

Bufalin, a traditional Chinese medicine, has been reported as a protective factor in many tumors. We therefore investigated the effect of bufalin on platelet-derived growth factor (PDGF)-BB-induced proliferation of cultured rat mesangial cells. The effect of bufalin on cell proliferation and its underlying mechanisms were investigated in cultured rat mesangial cells (MCs) by the methylthiazoletetrazolium (MTT) assay, flow cytometry, reverse transcription-polymerase chain reaction (RT-PCR), Western blotting, and cyclin-dependent kinases (CDK)2 and CDK4 kinase assays. Bufalin inhibited 20 ng/ml PDGF-BB-induced MC proliferation in a dose-dependent manner. Similar results were observed in different concentrations of bufalin, which blocked PDGF-BB-induced progression through G0/G1 to S phase of the cell cycle. Furthermore, bufalin not only inhibited upregulation of cyclin D1 and CDK4, but also downregulation of p21 in both mRNA and protein levels. Although bufalin did not affect p27 and CDK2 mRNA expression, it reversed downregulation of p27 and upregulation of CDK2 in protein level. Activity of CDK2 and CDK4 was also inhibited by bufalin. However, both bufalin and PDGF-BB did not affect cyclin E mRNA or protein expression. These results suggest that bufalin could inhibit MC proliferation by modulating cell cycle progress, indicating that bufalin could be a potential therapeutic agent for the prevention of mesangial proliferative glomerulonephritis.