Dihydroartemisinin inhibits the activation and proliferation of hepatic stellate cells by regulating miR­29b­3p.

Liver fibrosis is an early pathological feature of hepatic diseases. Hepatic stellate cell (HSC) activation and disordered proliferation are associated with liver fibrosis. The present study identified significant differences in the expression levels of microRNA (miRNA/miR)­29b­3p in clinical samples and multiple miRNA databases. Subsequently, the specific antifibrotic mechanism of miR­29b­3p was further elucidated. Reverse transcription­quantitative PCR, western blot, ELISA and immunofluorescence were used to detect the expression levels of target genes and proteins. Oil red O, Nile red and trypan blue staining were used to evaluate HSC activation and cell viability. A luciferase assay was used to detect the relationship between miR­29b­3p and VEGFA. Adhesion, wound healing, apoptosis double staining and JC­1 assays were used to detect the effects of VEGFR1 and VEGFR2 knockdown on HSCs. Immunoprecipitation and fluorescence colocalization were used to identify interactions between the proteins. Furthermore, a rat fibrosis model was constructed to investigate the effects of dihydroartemisinin (DHA) and miR­29b­3p in vivo and in vitro. The results indicated that miR­29b­3p both inhibited the activation of HSCs and limited the proliferation of activated HSCs via lipid droplet recovery and VEGF pathway regulation. VEGFA was identified as a direct target of miR­29b­3p, and knockdown of VEGFA induced cell apoptosis and autophagy. Notably, VEGFR1 and VEGFR2 knockdown both promoted apoptosis; however, VEGFR1 knockdown inhibited autophagy, whereas VEGFR2 knockdown induced autophagy. Furthermore, it was revealed that VEGFR2 regulated autophagy by mediating the PI3K/AKT/mTOR/ULK1 pathway. VEGFR2 knockdown also led to ubiquitination of heat shock protein 60, ultimately inducing mitochondrial apoptosis. Finally, DHA was identified as a natural agonist of miR­29­3p that effectively prevented liver fibrosis in vivo and in vitro. Overall, the present study determined the molecular mechanism by which DHA inhibited HSC activation and prevented liver fibrosis.

HpaP divergently regulates the expression of hrp genes in Xanthomonas oryzae pathovars oryzae and oryzicola.

The bacterial pathogens Xanthomonas oryzae pathovars oryzae (Xoo) and oryzicola (Xoc) cause leaf blight and leaf streak diseases on rice, respectively. Pathogenesis is largely defined by the virulence genes harboured in the pathogen genome. Recently, we demonstrated that the protein HpaP of the crucifer pathogen Xanthomonas campestris pv. campestris is an enzyme with both ATPase and phosphatase activities, and is involved in regulating the synthesis of virulence factors and the induction of the hypersensitive response (HR). In this study, we investigated the role of HpaP homologues in Xoo and Xoc. We showed that HpaP is required for full virulence of Xoo and Xoc. Deletion of hpaP in Xoo and Xoc led to a reduction in virulence and alteration in the production of virulence factors, including extracellular polysaccharide and cell motility. Comparative transcriptomics and reverse transcription-quantitative PCR assays revealed that in XVM2 medium, a mimic medium of the plant environment, the expression levels of hrp genes (for HR and pathogenicity) were enhanced in the Xoo hpaP deletion mutant compared to the wild type. By contrast, in the same growth conditions, hrp gene expression was decreased in the Xoc hpaP deletion mutant compared to the wild type. However, an opposite expression pattern was observed when the pathogens grew in planta, where the expression of hrp genes was reduced in the Xoo hpaP mutant but increased in the Xoc hpaP mutant. These findings indicate that HpaP plays a divergent role in Xoo and Xoc, which may lead to the different infection strategies employed by these two pathogens.

Stroke volume variation for predicting responsiveness to fluid therapy in patients undergoing cardiac and thoracic surgery: a systematic review and meta-analysis.

OBJECTIVE: To evaluate the reliability of stroke volume variation (SVV) for predicting responsiveness to fluid therapy in patients undergoing cardiac and thoracic surgery. DESIGN: Systematic review and meta-analysis. DATA SOURCES: PubMed, EMBASE, Cochrane Library, Web of Science up to 9 August 2020. METHODS: Quality of included studies were assessed with the Quality Assessment of Diagnostic Accuracy Studies-2 tool. We conducted subgroup analysis according to different anaesthesia and surgical methods with Stata V.14.0, Review Manager V.5.3 and R V.3.6.3. We used random-effects model to pool sensitivity, specificity and diagnostic odds ratio with 95% CI. The area under the curve (AUC) of receiver operating characteristic was calculated. RESULTS: Among the 20 relevant studies, 7 were conducted during thoracic surgery, 8 were conducted during cardiac surgery and the remaining 5 were conducted in intensive critical unit (ICU) after cardiac surgery. Data from 854 patients accepting mechanical ventilation were included in our systematic review. The pooled sensitivity and specificity were 0.73 (95％ CI: 0.59 to 0.83) and 0.62 (95％ CI: 0.46 to 0.76) in the thoracic surgery group, 0.71 (95％ CI: 0.65 to 0.77) and 0.76 (95％ CI: 0.69 to 0.82) in the cardiac surgery group, 0.85 (95％ CI: 0.60 to 0.96) and 0.85 (95％ CI: 0.74 to 0.92) in cardiac ICU group. The AUC was 0.73 (95% CI: 0.69 to 0.77), 0.80 (95% CI: 0.77 to 0.83) and 0.88 (95% CI: 0.86 to 0.92), respectively. Results of subgroup of FloTrac/Vigileo system (AUC=0.80, Youden index=0.38) and large tidal volume (AUC=0.81, Youden index=0.48) in thoracic surgery, colloid (AUC=0.85, Youden index=0.55) and postoperation (AUC=0.85, Youden index=0.63) in cardiac surgery, passive leg raising (AUC=0.90, Youden index=0.72) in cardiac ICU were reliable. CONCLUSION: SVV had good predictive performance in cardiac surgery or ICU after cardiac surgery and had moderate predictive performance in thoracic surgery. Nevertheless, technical and clinical variables may affect the predictive value potentially.

McvR, a single domain response regulator regulates motility and virulence in the plant pathogen Xanthomonas campestris.

Signal transduction pathways mediated by sensor histidine kinases and cognate response regulators control a variety of physiological processes in response to environmental conditions in most bacteria. Comparatively little is known about the mechanism(s) by which single-domain response regulators (SD-RRs), which lack a dedicated output domain but harbour a phosphoryl receiver domain, exert their various regulatory effects in bacteria. Here we have examined the role of the SD-RR proteins encoded by the phytopathogen Xanthomonas campestris pv. campestris (Xcc). We describe the identification and characterization of a SD-RR protein named McvR (motility, chemotaxis, and virulence-related response regulator) that is required for virulence and motility regulation in Xcc. Deletion of the mcvR open reading frame caused reduced motility, chemotactic movement, and virulence in Xcc. Global transcriptome analyses revealed the McvR had a broad regulatory role and that most motility and pathogenicity genes were down-regulated in the mcvR mutant. Bacterial two-hybrid and protein pull-down assays revealed that McvR did not physically interact with components of the bacterial flagellum but interacts with other SD-RR proteins (like CheY) and the subset of DNA-binding proteins involved in gene regulation. Site-directed mutagenesis and phosphor-transfer experiments revealed that the aspartyl residue at position 55 of the receiver domain is important for phosphorylation and the regulatory activity of McvR protein. Taken together, the findings describe a previously unrecognized class of SD-RR protein that contributes to the regulation of motility and virulence in Xcc.

Identification and Functional Analysis of the Pheromone Response Factor Gene of Sporisorium scitamineum.

The sugarcane smut fungus Sporisorium scitamineum is bipolar and produces sporidia of two different mating types. During infection, haploid cells of opposite mating types can fuse to form dikaryotic hyphae that can colonize plant tissue. Mating and filamentation are therefore essential for S. scitamineum pathogenesis. In this study, we obtained one T-DNA insertion mutant disrupted in the gene encoding the pheromone response factor (Prf1), hereinafter named SsPRF1, of S. scitamineum, via Agrobacterium tumefaciens-mediated transformation (ATMT) mutagenesis. Targeted deletion of SsPRF1 resulted in mutants with phenotypes similar to the T-DNA insertion mutant, including failure to mate with a compatible wild-type partner strain and being non-pathogenic on its host sugarcane. qRT-PCR analyses showed that SsPRF1 was essential for the transcription of pheromone-responsive mating type genes of the a1 locus. These results show that SsPRF1 is involved in mating and pathogenicity and plays a key role in pheromone signaling and filamentous growth in S. scitamineum.

The mating-type locus b of the sugarcane smut Sporisorium scitamineum is essential for mating, filamentous growth and pathogenicity.

Sporisorium scitamineum is the causal agent of sugarcane smut, which is one of the most serious constraints to global sugarcane production. S. scitamineum and Ustilago maydis are two closely related smut fungi, that are predicted to harbor similar sexual mating processes/system. To elucidate the molecular basis of sexual mating in S. scitamineum, we identified and deleted the ortholog of mating-specific U. maydis locus b, in S. scitamineum. The resultant b-deletion mutant was defective in mating and pathogenicity in S. scitamineum. Furthermore, a functional b locus heterodimer could trigger filamentous growth without mating in S. scitamineum, and functionally replace the b locus in U. maydis in terms of triggering aerial filament production and forming solopathogenic strains, which do not require sexual mating prior to pathogenicity on the host plants.