MiR-145 Alleviates Sepsis-Induced Inflammatory Responses and Organ Injury by Targeting ADAM17.

BACKGROUND: Current studies have demonstrated that disintegrin and metalloproteinase 17 (ADAM17) plays a critical role in the pathogenesis of sepsis. MicroRNA (miR)-145 is known to control immune responses as an anti-inflammatory modulatory molecule. However, a fundamental understanding of how miR-145 regulates ADAM17 and, more broadly, sepsis-induced inflammatory response remains unknown. METHODS: We used western blotting and quantitative real-time PCR (qRT-PCR) to measure expression levels of ADAM17 and miR-145. Enzyme-linked immunosorbent assays (ELISA) were performed to measure cytokine production. To determine if ADAM17 is a target gene of miR-145, bioinformatics analyses and luciferase reporter assays were conducted. The impacts of ADAM17 and miR-145 on sepsis-induced inflammatory responses were accessed in vitro using human umbilical endothelial cells (HUVECs) treated with lipopolysaccharide (LPS). Sepsis-induced inflammatory response was measured in vivo using a polymicrobial septic mouse model induced by cecal ligation and puncture (CLP) with pre-injection of a miR-145 agomir. RESULTS: In HUVECs treated with LPS, miR-145 expression was downregulated and miR-145 negatively regulated ADAM17 expression through direct binding to the ADAM17 transcript 3'-UTR. MiR-145 overexpression markedly reduced LPS-induced inflammatory cytokine production by targeting ADAM17 in HUVECs. In comparison to CLP-induced septic mice treated with a control agomir, treatment with a miR-145 agomir significantly reduced the expression of ADAM17, numerous downstream cytokines such as IL-6, TNF-α, IL-1ß and MCP-1, and the endothelial injury factors ICAM-1, VCAM-1. The miR-145 agomir also alleviated acute lung and kidney injury and improved the survival rate of septic mice. CONCLUSIONS: This study showed that miR-145, by specifically targeting ADAM17, negatively regulates sepsis-induced inflammatory responses and vascular endothelial injury, and ultimately improved organ injury and survival during sepsis. The underlying mechanism for the regulation of ADAM17 expression by miR-145 and sepsis-induced inflammatory reactions may offer sepsis patients a novel therapeutic option.

ADAM10-a "multitasker" in sepsis: focus on its posttranslational target.

BACKGROUND: Sepsis has a complex pathogenesis in which the uncontrolled systemic inflammatory response triggered by infection leads to vascular barrier disruption, microcirculation dysfunction and multiple organ dysfunction syndrome. Numerous recent studies reveal that a disintegrin and metalloproteinase 10 (ADAM10) acts as a "molecular scissor" playing a pivotal role in the inflammatory response during sepsis by regulating proteolysis by cleaving various membrane protein substrates, including proinflammatory cytokines, cadherins and Notch, which are involved in intercellular communication. ADAM10 can also act as the cellular receptor for Staphylococcus aureus α-toxin, leading to lethal sepsis. However, its substrate-specific modulation and precise targets in sepsis have not yet to be elucidated. METHODS: We performed a computer-based online search using PubMed and Google Scholar for published articles concerning ADAM10 and sepsis. CONCLUSIONS: In this review, we focus on the functions of ADAM10 in sepsis-related complex endothelium-immune cell interactions and microcirculation dysfunction through the diversity of its substrates and its enzymatic activity. In addition, we highlight the posttranslational mechanisms of ADAM10 at specific subcellular sites, or in multimolecular complexes, which will provide the insight to intervene in the pathophysiological process of sepsis caused by ADAM10 dysregulation.

Genetic Arg-304-His substitution in GRK5 protects against sepsis progression by alleviating NF-κB-mediated inflammation.

BACKGROUND: Previous studies have demonstrated that G protein-coupled receptor kinase 5 (GRK5) exerts a pivotal regulatory effect on the inflammation associated with sepsis. The present study aimed to investigate the clinical association of GRK5 genetic variants with sepsis and to further explore the underlying genetic mechanisms involved in regulating sepsis-induced inflammatory responses and the pathogenesis of sepsis. METHODS: This case-control study enrolled 1081 septic patients and 1147 matched controls for genotyping of GRK5 rs2230349 and rs2230345 polymorphisms. The effect of these genetic variants on GRK5-mediated inflammatory responses was analyzed in peripheral blood mononuclear cells (PBMCs) and THP-1 macrophages. A clinically relevant polymicrobial sepsis model was established by subjecting wild-type (WT) and GRK5-knockout mice to cecal ligation and puncture (CLP) to evaluate the role of GRK5 in sepsis. RESULTS: We identified significant differences in the genotype/allele distribution of rs2230349 G > A, but not rs2230345, between the sepsis subtype and septic shock subgroups (GA + AA vs. GG genotype, OR = 0.698, 95% CI = 0.547-0.893, P = 0.004; A vs. G allele, OR = 0.753, 95% CI = 0.620-0.919, P = 0.005) and between the survivor and nonsurvivor subgroups (GA + AA vs. GG genotype, OR = 0.702, 95% CI = 0.531-0.929, P = 0.015; A vs. G allele, OR = 0.753, 95% CI = 0.298-0.949, P = 0.017). PBMCs carrying the sepsis-associated protective A allele produced significantly lower levels of TNF-α and IL-1ß upon LPS stimulation. The results from the in vitro experiment showed that the Arg-304-His substitution caused by the rs2230349 G-to-A mutation in GRK5 significantly decreased the LPS-induced production of several proinflammatory cytokines, such as TNF-α, IL-6, IL-1ß and MCP-1, via the IκB-α/NF-κB signaling pathway in THP-1 macrophages. Furthermore, GRK5-knockout mice exhibited a significant decrease in IκB-α phosphorylation/degradation, the p-p65/p65 ratio, the p-p50/p50 ratio, p65 nuclear translocation and downstream cytokine (TNF-α, IL-6, IL-1ß and VCAM-1) production compared to WT mice after CLP surgery. A significant improvement in 7-day survival rate in GRK5-KO septic mice was observed in the presence of antibiotics. CONCLUSIONS: The Arg-304-His substitution caused by the rs2230349 G-to-A mutation in GRK5 might disrupt GRK5 function and alleviate IKB-α/NF-κB-mediated inflammatory responses, which ultimately conferred a genetic protective effect against susceptibility to sepsis progression and mortality. These results may, to some extent, explain the heterogeneity of the clinical prognoses of septic patients and provide novel opportunities for individualized approaches for sepsis treatment.

A comparative evaluation of lasers and photodynamic therapy in the nonsurgical treatment of peri-implant diseases: A Bayesian network meta-analysis.

PURPOSE: We conducted this Bayesian network meta-analysis (NMA) to evaluate the safety and efficacy of different lasers and PDT compared to conventional mechanical debridement (CMD) for peri-implant treatment. METHODS: The Web of Science, Cochrane Library and PubMed databases were searched for randomized clinical trials (RCTs) assessing the clinical effectiveness of adjunctive PDT, different lasers, and CMD until January 1st, 2022. Clinical outcomes were the changes in pocket probing depth (PPD), marginal bone loss (MBL), and clinical attachment level (CAL). RESULTS: Twenty-three studies, including 4 types of lasers, were included. Compared to that with CMD alone, PPD reduction was significantly more efficient in the diode laser (LD)+CMD groups (MD, 0.53; 95%CI, 0.13-0.93) and the PDT+CMD groups (MD, 0.83; 95%CI, 0.32-1.34) than in the CMD group in the follow-up period. Moreover, PDT+CMD treatment also showed a significantly better marginal bone level gain (MD, 0.32; 95%CI, 0.06-0.57). No significant effect on ΔCAL was observed among the different treatment strategies. Despite no differences in PPD reduction, MBL and CAL gains were found among the adjunctive laser treatment groups, PDT+CMD had the highest ranking probability of the most effective treatment in these clinical indices of periodontitis. The certainty of evidence for all outcomes was judged as very low to moderate. CONCLUSIONS: Within the limits of this NMA, we found that adjunctive PDT achieved a small additional benefit on PPD reduction and MBL gain compared with CMD alone and had the highest probability of being ranked first on the changes in PPD, MBL and CAL. PDT+CMD may represent an alternative method for peri­implant treatment. Further high-quality RCTs are needed to assess the influence of potential confounders on the efficacy of lasers and PDT.

Efficacy and safety of nafamostat mesilate anticoagulation in blood purification treatment of critically ill patients: a systematic review and meta-analysis.

BACKGROUND: Nafamostat mesilate (NM), a broad-spectrum and potent serine protease inhibitor, can be used as an anticoagulant during extracorporeal circulation, as well as a promising drug effective against coronavirus disease 2019 (COVID-19). We conducted a systematic meta-analysis to evaluate the safety and efficacy of NM administration in critically ill patients who underwent blood purification therapy (BPT). METHODS: The Cochrane Library, Web of Science and PubMed were comprehensively searched from inception to August 20, 2021, for potential studies. RESULTS: Four randomized controlled trials (RCTs) and seven observational studies with 2723 patients met the inclusion criteria. The meta-analysis demonstrated that conventional therapy (CT) significantly increased hospital mortality compared with NM administration (RR = 1.25, p = 0.0007). In subgroup analyses, the in-hospital mortality of the NM group was significantly lower than that of the anticoagulant-free (NA) group (RR = 1.31, p = 0.002). The CT interventions markedly elevated the risk ratio of bleeding complications by 45% (RR = 1.45, p = 0.010) compared with NM interventions. In another subgroup analysis, NM used exhibited a significantly lower risk of bleeding complications than those of the low-molecular-weight heparin (LMWH) used (RR = 4.58, p = 0.020). The filter lifespan was decreased significantly (MD = -10.59, p < 0.0001) in the NA groups compared with the NM groups. Due to the poor quality of the included RCTs, these results should be interpreted with caution. CONCLUSION: Given the better survival outcomes, lower risk of bleeding, NM anticoagulation seems to be a safe and efficient approach for BPT patients and could yield a favorable filter lifespan. More multi-center RCTs with large samples are required for further validation of this study.

The -172 A-to-G variation in ADAM17 gene promoter region affects EGR1/ADAM17 pathway and confers susceptibility to septic mortality with sepsis-3.0 criteria.

BACKGROUND: A disintegrin and metalloproteinase 17 (ADAM17) is a proteolytic cleaving protein with a crucial function in the inflammatory responses, especially sepsis. But the clear role of ADAM17 in sepsis and the underlying mechanism remained unknown. In this study, we aim to determine the clinical association of ADAM17 -172A > G (rs12692386) promoter polymorphism with sepsis and to further explore the effect and mechanism of the early growth response 1 (EGR1)/ADAM17 pathway in inflammatory process during sepsis. METHODS: A total of 477 sepsis patients and 750 controls were enrolled in this study to determine the association of ADAM17 -172A > G polymorphism with sepsis. The transcription factor binding to the promoter region of ADAM17 gene was predicted by bioinformatics analysis and verified by Chromatin Immunoprecipitation (ChIP) and luciferase assays. Quantitative real-time PCR and Western blot were performed to detect EGR1 and ADAM17 expression. Cytokine production was detected by enzyme-linked immunosorbent assay. The effect of EGR1/ADAM17 pathway on sepsis-induced inflammatory responses was evaluated in EGR1-silenced cells and endotoxemia mouse model. RESULTS: The frequencies of non-survivors among the sepsis patients with the -172AG/GG genotypes and G allele were distinctly higher than those among patients with the AA genotype (53.9% vs. 39.7%, OR = 1.779, 95% CI = 1.119-2.829, P = 0.0142) and A allele (30.9% vs. 22.2%, OR = 1.570, 95% CI = 1.095-2.251, P = 0.0136). The Kaplan-Meier survival analysis indicated that the 28-day survival in septic patients with -172AG/GG genotypes of this functional ADAM17 promoter polymorphism was much worse than in the AA genotype carriers (log-rank = 5.358, P = 0.021). The results of in vitro lipopolysaccharide-stimulated and luciferase assays indicated that the -172 A-to-G variation could functionally upregulate promoter activity and transcription of ADAM17 gene via enhancing the binding affinity of its promoter region with the EGR1. The ChIP assay identified the direct interaction. Further studies demonstrated that inhibition of EGR1 significantly decreased ADAM17 expression and the pro-inflammatory cytokine secretion in vitro, and improved the survival and inflammatory response of sepsis mouse model. CONCLUSIONS: These results provided evidence that the ADAM17 -172A > G polymorphism functionally promoted ADAM17 expression and enhanced sepsis-induced inflammatory responses via the EGR1/ADAM17 pathway, which ultimately conferred susceptibility to sepsis mortality and poor prognosis.

Investigation of Association of Complement 5 Genetic Polymorphisms with Sepsis and Sepsis-Induced Inflammatory Responses.

BACKGROUND: Complement 5 (C5) and C5a production play a pivotal role in the pathophysiology of sepsis. Strong evidence demonstrates an association of C5 gene polymorphisms with various inflammatory diseases. However, no current studies have explored the clinical relevance of C5 polymorphisms in sepsis. METHODS: Two C5 gene polymorphisms, rs17611 and rs2269067, were identified by genotyping in 636 sepsis patients and 753 controls in a Han Chinese population. C5 gene expression was detected via quantitative real-time PCR. C5a and proinflammatory cytokine production was measured by enzyme-linked immunosorbent assay. An Annexin V apoptosis assay was performed to assess cell apoptosis. RESULTS: Our results showed significantly lower frequencies of rs2269067 GC/CC genotypes or C allele in sepsis patients than healthy controls. The frequencies of rs17611 CC/CT genotypes or C allele were significantly overrepresented in both the septic shock and non-survivor subgroups. Patients with this sepsis-associated high-risk rs17611 C allele exhibited a significant increase in C5a, TNF-α and IL-6 production. However, no significant difference in C5a and downstream proinflammatory cytokine production was observed among patients with different rs2269067 genotypes. In addition, in vitro experiments showed an effect of recombinant C5a on enhancing LPS-stimulated IL-1ß, IL-6 and TNF-α production and cell apoptosis in THP-1 monocytes. CONCLUSION: The rs2269067 polymorphism conferred protection against sepsis susceptibility. The rs17611 polymorphism was associated with increased C5a production, which ultimately potentiated the secretion of downstream proinflammatory cytokines and conferred susceptibility to sepsis progression and poor prognosis.

A gain-of-function NLRP3 3'-UTR polymorphism causes miR-146a-mediated suppression of NLRP3 expression and confers protection against sepsis progression.

Nucleotide-binding domain and leucine-rich repeat (LRR)-containing family protein 3 (NLRP3) regulated the maturation of inflammation-related cytokines by forming NLRP3 inflammasome, which plays pivotal roles in sepsis pathogenesis. In this study, we evaluated the genetic association of NLRP3 polymorphisms with sepsis (640 patients and 769 controls) and characterized the impact of NLRP3 polymorphisms on NLRP3 expression and inflammatory responses. No significant differences were observed in genotype/allelic frequencies of NLRP3 29940G>C between sepsis cases and controls. The G allele was significantly overrepresented in patients with septic shock than those in sepsis subgroup, and the GC/GG genetypes were related to the 28-day mortality of sepsis. Lipopolysaccharide challenge to peripheral blood mononuclear cells showed a significant suppression of NLRP3 mRNA expression and release of IL-1ß and TNF-α in CC compared with the GC/GG genotype category. Functional experiments with luciferase reporter vectors containing the NLRP3 3'-UTR with the 29940 G-to-C variation in HUVECs and THP-1 cells showed a potential suppressive effect of miR-146a on NLRP3 transcription in the presence of the C allele. Taken together, these results demonstrated that the 29940 G-to-C mutation within the NLRP3 3'-UTR was a gain-of-function alteration that caused the suppression of NLRP3 expression and downstream inflammatory cytokine production via binding with miR-146a, which ultimately protected patients against susceptibility to sepsis progression and poor clinical outcome.