Gasdermin D-mediated pyroptosis is regulated by AMPK-mediated phosphorylation in tumor cells.

Gasdermin D (GSDMD) is a critical mediator of pyroptosis, which consists of a N-terminal pore-forming domain and a C-terminal autoinhibitory domain. Its cytolytic activity is sequestered by the intramolecular autoinhibitory mechanism. Upon caspase-1/11 mediated cleavage of GSDMD, the N-terminal pore-forming domain (GD-NT) is released to mediate pyroptosis. However, it remains unclear how GD-NT is regulated once it is generated. In the current study, we developed a TetOn system in which GD-NT was selectively induced in tumor cells to explore how the cytolytic activity of GD-NT is regulated. We found that the cytolytic activity of GD-NT was negatively regulated by the AMP-activated protein kinase (AMPK) and AMPK activation rendered tumor cells resistant to GD-NT-mediated pyroptosis. Mechanistically, AMPK phosphorylated GD-NT at the serine 46 (pS46-GD), which altered GD-NT oligomerization and subsequently eliminated its pore-forming ability. In our in vivo tumor model, AMPK-mediated phosphorylation abolished GD-NT-induced anti-tumor activity and resulted in an aggressive tumor growth. Thus, our data demonstrate the critical role of AMPK in negatively regulating the cytolytic activity of GD-NT. Our data also highlight an unexpected link between GSDMD-mediated pyroptosis and the AMPK signaling pathway in certain tumor cells.

Abnormal mechanical stress on bicuspid aortic valve induces valvular calcification and inhibits Notch1/NICD/Runx2 signal.

Background: Bicuspid aortic valve (BAV) is a congenital cardiac deformity, increasing the risk of developing calcific aortic valve disease (CAVD). The disturbance of hemodynamics can induce valvular calcification, but the mechanism has not been fully identified. Methods: We constructed a finite element model (FEM) of the aortic valve based on the computed tomography angiography (CTA) data from BAV patients and tricuspid aortic valve (TAV) individuals. We analyzed the hemodynamic properties based on our model and investigated the characteristics of mechanical stimuli on BAV. Further, we detected the expression of Notch, NICD and Runx2 in valve samples and identified the association between mechanical stress and the Notch1 signaling pathway. Results: Finite element analysis showed that at diastole phase, the equivalent stress on the root of BAV was significantly higher than that on the TAV leaflet. Correspondingly, the expression of Notch1 and NICH decreased and the expression of Runx2 elevated significantly on large BAV leaflet belly, which is associated with equivalent stress on leaflet. Our findings indicated that the root of BAV suffered higher mechanical stress due to the abnormal hemodynamic environment, and the disturbance of the Notch1/NICD/Runx2 signaling pathway caused by mechanical stimuli contributed to valvular calcification.

Adenovirus-IL-10 relieves chronic rejection after mouse heart transplantation by inhibiting miR-155 and activating SOCS5.

Objective: Chronic rejection remains the main factor that influence long-term survival of patients after heart transplantation. Interleukin-10 (IL-10) play critical role in macrophages-mediated transplant immune responses. We investigated the mechanism of IL-10 in macrophage related chronic rejection after mouse heart transplantation. Methods: Mouse heart transplant chronic rejection model was established to evaluate pathological changes in the allograft. Myocardial interstitial fibrosis, apoptosis, and inflammatory factor levels were detected in ad-IL-10-treated mice. The positive iNOS+ and Arg-1+ expressions, macrophage subset changes, and the proportion of regulatory T-cells (Tregs) and TIGIT+ Tregs were quantified by flow. In in vitro experiments, ad-IL-10 was transfected into macrophages followed by detection of apoptosis, phagocytosis, and CD163, CD16/32, and CD206 expression. The expression and relationships between IL-10, miR-155, and SOCS5 were also detected and verified. A rescue experiment was performed to evaluate macrophage function through the combined treatment of ad-IL-10 and overexpression of miR-155. Results: Significantly decreased IL-10 expression in chronic rejection during mouse heart transplantation was observed. Ad-IL-10-treated mice showed decreased pathological injury, perivascular fibrosis, apoptosis, inflammation, and iNOS+ and CD16/32+ expression, and increased Treg/TIGIT+ Treg cell, Arg-1+ and CD206+ cell proportion. Ad-IL-10-treated macrophages in vitro showed reduced apoptosis, improved phagocytosis, and M2 polarization. Mechanically, IL-10 negatively regulated miR-155 to activate SOCS5. Overexpression of miR-155 reversed IL-10 mediated-positive regulation of macrophage function. Conclusion: IL-10 downregulated miR-155 and activated SOCS5, thereby promoting macrophage M2 polarization to relieve chronic rejection after heart transplantation.

Identification and Verification of Tumor Immune Microenvironment-Related Prognostic Genes in Kidney Renal Clear Cell Carcinoma.

BACKGROUND: The tumor immune microenvironment is vital to kidney renal clear cell carcinoma (KIRC) progression, and immunotherapies have been shown to be effective in the management of KIRC. However, the prognostic genes associated with the tumor immune microenvironment in KIRC have not been fully identified. We obtained the KIRC RNA sequencing data and the clinical characteristics from The Cancer Genome Atlas (TCGA) and the International Cancer Genome Consortium (ICGC) database. We screened the gene modules associated with the tumor immune microenvironment based on the ESTIMATE algorithm and weighted gene coexpression network analysis (WGCNA). Univariate Cox analysis and the LASSO method were used to construct a prognostic model. Receiver Operating Characteristic (ROC) curve analysis was performed to evaluate the accuracy of our risk model. TIMER and Single-Sample Gene Set Enrichment Analysis (ssGSEA) were used to explore the correlation between prognostic genes and immune cell infiltration. RESULTS: Fifty-four genes in modules were significantly associated with the overall survival (OS) time of patients with KIRC. Furthermore, 12 hub genes were selected to construct the prognostic model. The prognostic model showed superior accuracy in both TCGA and ICGC cohorts using ROC curve analysis. Systematic analysis of immune cell infiltration revealed that nine genes were significantly correlated with levels of tumor-infiltrating immune cells. CONCLUSIONS: Our findings indicated that the tumor immune microenvironment was an important determinant of KIRC outcomes and revealed potential biomarkers for predicting patient OS and for targeted immunotherapies.

Erythropoietin and its derivatives: from tissue protection to immune regulation.

Erythropoietin (EPO) is an evolutionarily conserved hormone well documented for its erythropoietic role via binding the homodimeric EPO receptor (EPOR)2. In past decades, evidence has proved that EPO acts far beyond erythropoiesis. By binding the tissue-protective receptor (TPR), EPO suppresses proinflammatory cytokines, protects cells from apoptosis and promotes wound healing. Very recently, new data revealed that TPR is widely expressed on a variety of immune cells, and EPO could directly modulate their activation, differentiation and function. Notably, nonerythropoietic EPO derivatives, which mimic the structure of helix B within EPO, specifically bind TPR and show great potency in tissue protection and immune regulation. These small peptides prevent the cardiovascular side effects of EPO and are promising as clinical drugs. This review briefly introduces the receptors and tissue-protective effects of EPO and its derivatives and highlights their immunomodulatory functions and application prospects.

Adaptive features of innate immune cells and their relevance to graft rejection.

PURPOSE OF REVIEW: Allograft rejection involves both innate and adaptive immune cells, and the adaptive immune cells have dominated transplant studies for decades. Recent studies have identified surprising new features for the innate immune cells, including memory recall responses, which may have significant implications in further improvement of transplant outcomes. RECENT FINDINGS: Transplant survival is excellent in the short-term, but the long-term graft outcomes are not so, and most grafts are continuously lost to chronic rejection in the clinic. In both animal models and clinical settings, graft loss to chronic rejection is often dominated by innate immune cells, especially macrophages and natural killer (NK) cells in the grafts. Recent studies suggest that innate immune cells can acquire features of adaptive cells in that they either directly sense allogeneic nonself or become 'trained' in the allogeneic milieu, where they show features of memory recall responses. In certain models, targeting the adaptive features of such innate immune cells can promote long-term allograft survival. These findings may open new therapeutic opportunities in promoting transplant survival in the clinic. SUMMARY: The discovery of donor specificity and memory recall responses of certain innate immune cells, which are prominently featured in chronic allograft rejection, may open novel therapeutic opportunities in transplantation, as well as in treatment of cancers and autoimmune diseases.