LncRNA GAS5 suppresses TGF-ß1-induced transformation of pulmonary pericytes into myofibroblasts by recruiting KDM5B and promoting H3K4me2/3 demethylation of the PDGFRα/ß promoter.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a condition that may cause persistent pulmonary damage. The transformation of pericytes into myofibroblasts has been recognized as a key player during IPF progression. This study aimed to investigate the functions of lncRNA growth arrest-specific transcript 5 (GAS5) in myofibroblast transformation during IPF progression. METHODS: We created a mouse model of pulmonary fibrosis (PF) via intratracheal administration of bleomycin. Pericytes were challenged with exogenous transforming growth factor-ß1 (TGF-ß1). To determine the expression of target molecules, we employed quantitative reverse transcription-polymerase chain reaction, Western blotting, and immunohistochemical and immunofluorescence staining. The pathological changes in the lungs were evaluated via H&E and Masson staining. Furthermore, the subcellular distribution of GAS5 was examined using FISH. Dual-luciferase reporter assay, ChIP, RNA pull-down, and RIP experiments were conducted to determine the molecular interaction. RESULTS: GAS5 expression decreased whereas PDGFRα/ß expression increased in the lungs of IPF patients and mice with bleomycin-induced PF. The in vitro overexpression of GAS5 or silencing of PDGFRα/ß inhibited the TGF-ß1-induced differentiation of pericytes to myofibroblasts, as evidenced by the upregulation of pericyte markers NG2 and desmin as well as downregulation of myofibroblast markers α-SMA and collagen I. Further mechanistic analysis revealed that GAS5 recruited KDM5B to promote H3K4me2/3 demethylation, thereby suppressing PDGFRα/ß expression. In addition, KDM5B overexpression inhibited pericyte-myofibroblast transformation and counteracted the promotional effect of GAS5 knockdown on pericyte-myofibroblast transformation. Lung fibrosis in mice was attenuated by GAS5 overexpression but promoted by GAS5 deficiency. CONCLUSION: GAS5 represses pericyte-myofibroblast transformation by inhibiting PDGFRα/ß expression via KDM5B-mediated H3K4me2/3 demethylation in IPF, identifying GAS5 as an intervention target for IPF.

AUF1 protects against ferroptosis to alleviate sepsis-induced acute lung injury by regulating NRF2 and ATF3.

BACKGROUND: The AU-rich element (ARE)-binding factor 1 (AUF1) acts as a switch for septic shock, although its underlying mechanisms remain largely unknown. In this study, we examined the biological significance and potential molecular mechanism of AUF1 in regulating ferroptosis in sepsis-induced acute lung injury (ALI). METHODS: Alveolar epithelial cells (AECs) challenged with ferroptosis-inducing compounds and cecum ligation and puncture (CLP)-induced ALI were used as the in vitro and in vivo model, respectively. The stability of AUF1 and its degradation by ubiquitin-proteasome pathway were examined by cycloheximide chase analysis and co-immunoprecipitation assay. The regulation of AUF1 on nuclear factor E2-related factor 2 (NRF2) and activation transcription factor 3 (ATF3) was explored by RNA immunoprecipitation (RIP), RNA pull-down, and mRNA stability assays. Functionally, the effects of altering AUF1, NRF2 or ATF3 on ferroptosis in AECs or ALI mice were evaluated by measuring cell viability, lipid peroxidation, iron accumulation, and total glutathione level. RESULTS: AUF1 was down-regulated in AECs challenged with ferroptosis-inducing compounds, both on mRNA and protein levels. The E3 ubiquitin ligase FBXW7 was responsible for protein degradation of AUF1 during ferroptosis. By up-regulating NRF2 and down-regulating ATF3, AUF1 antagonized ferroptosis in AECs in vitro. In the CLP-induced ALI model, the survival rate of AUF1 knockout mice was significantly reduced and the lung injuries were aggravated, which were related to the enhancement of lung ferroptosis. CONCLUSIONS: FBXW7 mediates the ubiquitination and degradation of AUF1 in ferroptosis. AUF1 antagonizes ferroptosis by regulating NRF2 and ATF3 oppositely. Activating AUF1 pathway may be beneficial to the treatment of sepsis-induced ALI.

Exosomal miR-107 antagonizes profibrotic phenotypes of pericytes by targeting a pathway involving HIF-1α/Notch1/PDGFRß/YAP1/Twist1 axis in vitro.

Microvascular pericytes have been demonstrated as an origin for myofibroblasts that produce excessive extracellular matrix (ECM) proteins such as α-smooth muscle actin (α-SMA) and type I collagen (ColIA1) and contribute to pulmonary fibrosis (PF). However, the signaling mechanism responsible for ECM production within pericytes is poorly understood. In this study, we examined exosomal miR-107 in the fibrotic phenotypes of pericytes and the pathogenesis of PF. Using RT-qPCR, MiR-107 level was compared between clinical or bleomycin-induced PF and normal pulmonary tissues. Exosomes were isolated from cultured microvascular endothelial cells (ECs) derived from either normal or PF tissues, characterized using dynamic light scattering, transmission electron microscopy, flow cytometry, Western blot, and immunofluorescence, and then applied to pericytes. The effects of exosomes or different fibrosis-related signaling molecules were examined by Western blot, and the potential regulations between the signaling molecules were identified using bioinformatic analysis and assessed by electrophoretic mobility shift assay, chromatin immunoprecipitation, luciferase assay, and RNA binding protein immunoprecipitation. MiR-107 was downregulated in clinical or experimental PF tissues and also in exosomes from PF-derived ECs. EC-derived exosomal miR-107 essentially controlled the miR-107 level and inhibited α-SMA and ColIA1 expression in pericytes. The antifibrosis effect of miR-107 was mediated through the suppression of a pathway involving HIF-1α/Notch1/PDGFRß/YAP1/Twist1, where miR-107 directly targeted HIF-1α mRNA, whereas the latter directly activated the transcriptions of both Notch1 and PDGFRß. Functionally, targeting miR-107 promoted and targeting HIF-1α abolished the fibrotic phenotypes of pericytes. Exosomal miR-107 produced by pulmonary vascular ECs may alleviate pericyte-induced fibrosis by inhibiting a signaling pathway involving HIF-1α/Notch1/PDGFRß/YAP1/Twist1.NEW & NOTEWORTHY This work reveals a novel mechanism by which pulmonary vascular endothelial cells, via regulating the transdifferentiation of microvascular pericytes into myofibroblasts, contribute to the pathogenesis of pulmonary fibrosis. Since targeting the formation of myofibroblasts may prevent the development and benefit the treatment of pulmonary fibrosis, this study provides not only mechanistic understanding but also promising therapeutic targets for pulmonary fibrosis.

Low let-7d exosomes from pulmonary vascular endothelial cells drive lung pericyte fibrosis through the TGFßRI/FoxM1/Smad/ß-catenin pathway.

The pathogenesis of pulmonary fibrosis (PF) was mediated by the progressive deposition of excessive extracellular matrix, but little is known about the regulatory mechanisms of fibrogenesis by lung pericytes. The mouse PF model was established by treatment with bleomycin, followed by isolation of exosomes from mouse broncho-alveolar lavage fluids by the centrifuge method. Relative mRNA/microRNA levels and protein expression were assessed by qRT-PCR and Western blotting, respectively. The binding of let-7d with gene promoter was validated by dual-luciferase reporter assay. Protein interactions were verified via GST pull-down and co-immunoprecipitation. Nuclear retention of Smad3 was analysed by extraction of cytoplasmic and nuclear fraction of pericytes followed by Western blotting. Association of FoxM1 with gene promoter was detected by EMSA and ChIP-PCR methods. FoxM1 expression is significantly elevated in human lung fibroblasts of PF patients and mouse PF model. The expression of let-7d is repressed in exosomes derived from broncho-alveolar lavage fluids of PF mice. Let-7d or FoxM1 knockdown suppressed the expression of FoxM1, Smad3, ß-catenin, Col1A and α-SMA expression in mouse lung pericytes under TGF-ß1 treatment. FoxM1 overexpression elevated above gene expression in mouse lung pericytes under TGF-ß1 treatment. Let-7d directly targets TGFßRI to regulate FoxM1 and downstream gene expression in mouse lung pericytes. FoxM1 directly interacts with Smad3 proteins to promote Smad3 nuclear retention and binds with ß-catenin promoter sequence to promote fibrogenesis. Exosomes with low let-7d from pulmonary vascular endothelial cells drive lung pericyte fibrosis through activating the TGFßRI/FoxM1/Smad/ß-catenin signalling pathway.

Analysis of volume management by comparing between critical care ultrasound examination and pulse indicator cardiac output in patients with septic shock.

OBJECTIVE: To investigate volume management by comparing between critical care ultrasound examination and pulse indicator cardiac output (PICCO) in patient with septic shock. METHODS: Patients with septic shock during July 2017 and June 2018 were included. Inferior Vena Cava (IVC), total end-diastolic volume index (GEDI), central venous pressure (CVP), lactic acid and oxygenation index were measured by ultrasound. First, the accuracy difference of IVC, GEDI and CVP estimation capacity was compared. According to the changes of IVCmin, IVCmax, and GEDI, they were divided into 5 groups to compare the differences of lactic acid and oxygenation index between the groups and the correlation of lactate and Oxygenation index (PaO2/FiO2) between IVC and GEDI was analyzed. The correlation of lactate and PaO2/FiO2 between B lines and extravascular pulmonary water index (ELWI) was noted. RESULTS: The accuracy of IVC and GEDI in volume estimation was greater than 75%, significantly higher than that of CVP (53.3%) (P<0.05). The correlation results showed that GEDI was significantly correlated with IVCmax and IVCmin (P<0.05), while there was a significant correlation between b-line area and oxygenation index, ELWI and lactic acid, ELWI and oxygenation index (P<0.05). IVCmin, IVCmax and GEDI were respectively divided into 5 groups for comparing the difference between lactic acid and oxygenation. It was found that there were significant differences between the two indicators of IVCmin in different groups (P>0.05). The oxygenation index of the group ≤IVCmax was significantly lower than that of the group 0.5 ≤IVCmax < 1.0cm (P<0.05). The oxgenation indexes of groups 500≤GEDI < 600mL/m2; 600≤GEDI < 700mL/m2. 700≤GEDI < 800mL/m2 were significantly higher than that of group 0 < GEDI < 500mL/m2 (P<0.05). CONCLUSIONS: Critical care ultrasound examination and PICCO are better methods than in volume management, but PICCO is more individualized, and PICCO in patients with valvular heart disease is not recommended.

Statins Reduce Mortality After Non-severe but Not After Severe Pneumonia: A Systematic Review and Meta-Analysis.

PURPOSE: The objective of this study was to perform a systematic review and meta-analysis of the effects of statins on mortality for patients with non-severe pneumonia or severe pneumonia. METHODS: PubMed, EMBASE, Cochrane Database of Systematic Reviews, Cochrane central register of controlled trials and Clinicaltrials.gov were searched for the association between statins and non-severe/severe pneumonia. Eligible articles were analyzed in Stata 12.0. RESULTS: The database search yielded a total of 566 potential publications, 24 studies involving 312,309 patients met the eligibility criteria. Pooled unadjusted data showed that statin use was associated with lower mortality after non-severe pneumonia (odds ratio [OR] 0.70, 95% confidence interval [CI], 0.66-0.73), but not severe pneumonia (OR 1.05; 95% CI, 0.86-1.28). However, this protective effect of statins was weakened using adjusted estimates (OR 0.78, 95% CI, 0.75-0.82). Besides, protective effect of statins was attenuated by confounders in a subgroup analysis, especially when accounting for pneumonia severity indicators (OR 0.88; 95% CI, 0.80-0.96). CONCLUSIONS: Statin use was associated with reduced mortality after non-severe pneumonia but not severe pneumonia and this protective effect was weakened in subgroups.