High Hydrostatic Pressure Exacerbates Bladder Fibrosis through Activating Piezo1.

OBJECTIVE: Bladder outlet obstruction (BOO) results in significant fibrosis in the chronic stage and elevated bladder pressure. Piezo1 is a type of mechanosensitive (MS) channel that directly responds to mechanical stimuli. To identify new targets for intervention in the treatment of BOO-induced fibrosis, this study investigated the impact of high hydrostatic pressure (HHP) on Piezo1 activity and the progression of bladder fibrosis. METHODS: Immunofluorescence staining was conducted to assess the protein abundance of Piezo1 in fibroblasts from obstructed rat bladders. Bladder fibroblasts were cultured under normal atmospheric conditions (0 cmH2O) or exposed to HHP (50 cmH2O or 100 cmH2O). Agonists or inhibitors of Piezo1, YAP1, and ROCK1 were used to determine the underlying mechanism. RESULTS: The Piezo1 protein levels in fibroblasts from the obstructed bladder exhibited an elevation compared to the control group. HHP significantly promoted the expression of various pro-fibrotic factors and induced proliferation of fibroblasts. Additionally, the protein expression levels of Piezo1, YAP1, ROCK1 were elevated, and calcium influx was increased as the pressure increased. These effects were attenuated by the Piezo1 inhibitor Dooku1. The Piezo1 activator Yoda1 induced the expression of pro-fibrotic factors and the proliferation of fibroblasts, and elevated the protein levels of YAP1 and ROCK1 under normal atmospheric conditions in vitro. However, these effects could be partially inhibited by YAP1 or ROCK inhibitors. CONCLUSION: The study suggests that HHP may exacerbate bladder fibrosis through activating Piezo1.

Rhizobium mesosinicum sp. nov., isolated from root nodules of three different legumes.

Thirteen novel strains were isolated from root nodules of three different leguminous plants of the genera Albizia, Kummerowia and Dalbergia grown in China. Cells of these strains were Gram-negative, strictly aerobic, non-spore-forming, motile rods. Phylogenetic analysis of 16S rRNA gene sequences revealed that they belong to the genus Rhizobium. A representative strain, CCBAU 25010T, showed 98.8% 16S rRNA gene sequence similarity to its closest phylogenetic relative, Rhizobium sullae IS123T. The 16S-23S intergenic spacer (ITS) sequence of CCBAU 25010T shared 91.5 and 87.2% sequence similarity, respectively, with those of Rhizobium etli CFN 42T and Rhizobium leguminosarum LMG 14904T. Analysis of the sequences of the housekeeping genes atpD and recA was in agreement with the results of ITS sequence analysis. The nodC gene sequence of CCBAU 25010T was identical to that of Rhizobium tropici CFN 299. DNA-DNA hybridization values for strain CCBAU 25010T ranged from 20.7% (with Rhizobium mongolense USDA 1844T) to 34.4% (with R. leguminosarum USDA 2370T). Cell protein SDS-PAGE, BOX-PCR and several phenotypic characteristics, such as use of sole carbon sources and antibiotic resistance, could differentiate the novel strains from defined Rhizobium species. We therefore propose that the novel strains reported in this study form a novel species, Rhizobium mesosinicum sp. nov., with the type strain CCBAU 25010T (=LMG 24135T =JCM 14777T).

An ABC transporter is required for alkaline stress and potassium transport regulation in Sinorhizobium meliloti.

To understand the mechanisms of high-pH-induced protection in Sinorhizobium meliloti, a cDNA-amplified fragment length polymorphism analysis of S. meliloti cells grown in minimal medium under alkali stress was undertaken. This revealed that the first four genes of a seven-gene cluster encode the characteristic components of a putative sugar ATP-binding cassette (ABC) transporter. A functional study suggested that this putative sugar ABC transporter might play a role in potassium transport regulation, which we therefore designated supABCD. The transcription of three potassium uptake genes, trkH, kdpA and kup1, in S. meliloti is significantly attenuated in the supA mutant in the presence of potassium. The supA mutant was unable to grow at elevated levels of potassium. The expression of supA, as determined by beta-galactosidase activity, was shown to be induced by potassium but not by sodium.

Shinella kummerowiae sp. nov., a symbiotic bacterium isolated from root nodules of the herbal legume Kummerowia stipulacea.

Bacterial strain CCBAU 25048(T) was isolated from root nodules of Kummerowia stipulacea grown in Shandong province of China. Cells of the strain were Gram-negative, strictly aerobic, non-spore-forming, motile short rods. Phylogeny of 16S rRNA gene sequences revealed that the strain belonged to the genus Shinella, a member of family Rhizobiaceae. Its closest phylogenetic relatives were Shinella granuli Ch06(T) and Shinella zoogloeoides IAM 12669(T), respectively showing 98.3 and 98.9 % 16S rRNA gene sequence similarity. Strain CCBAU 25048(T) had DNA-DNA relatedness of 43.5 and 34.8 %, respectively, with S. zoogloeoides JCM 20728(T) and S. granuli JCM 13254(T). In addition, in TP-RAPD analysis, different patterns were obtained for these three strains and some rhizobial strains. The nifH, nodC and nodD sequences of CCBAU 25048(T) were identical or very similar to those of bean-nodulating Rhizobium tropici strains. Several phenotypic characteristics, including the use of citrate and d-ribose as carbon sources and growth at pH 11.0, as well as the fatty acid composition, could differentiate CCBAU 25048(T) from the two defined Shinella species. Therefore, a novel species Shinella kummerowiae sp. nov. is proposed, with strain CCBAU 25048(T) (=JCM 14778(T) =LMG 24136(T)) as the type strain.

Diverse rhizobia that nodulate two species of Kummerowia in China.

A total of 63 bacterial strains were isolated from root nodules of Kummerowia striata and K. stipulacea grown in different geographic regions of China. These bacteria could be divided into fast-growing (FG) rhizobia and slow-growing (SG) rhizobia according to their growth rate. Genetic diversity and taxonomic relationships among these rhizobia were revealed by PCR-based 16 S rDNA RFLP and sequencing, 16 S-IGS RFLP, SDS-PAGE of whole cell soluble proteins, BOX-PCR and symbiotic gene (nifH/nodC) analyses. The symbiotic FG strains were mainly isolated from temperate regions and they were identified as four genomic species in Rhizobium and Sinorhizobium meliloti based on the consensus of grouping results. The SG strains were classified as five genomic species within Bradyrhizobium and they were mainly isolated fron the subtropic and tropical regions. The phylogenetic analyses of nifH and nodC genes showed relationships similar to that of 16 S rDNA but the symbiotic genes of Bradyrhizobium strains isolated from Kummerowia were distinct from those isolated from Arachis and soybean. These results offered evidence for rhizobial biogeography and demonstrated that the Kummerowia-nodulating ability might have evolved independently in different regions in association with distinctive genomic species of rhizobia.