Geranylgeranylacetone (GGA), an isoprenoid compound widely utilized as an antiulcer agent in Asia, confers protection against ischemia, anoxia, and oxidative stress by rapidly enhancing the expression of HSP70. Nevertheless, the impact of GGA on sepsis-associated intestinal injury remains unexplored. Thus, this study is crafted to elucidate the protective efficacy and underlying mechanisms of GGA against septic intestinal damage. Our findings revealed that GGA significantly extended the survival duration of septic mice, and mitigated lipopolysaccharide (LPS)-induced alterations in intestinal permeability and tissue damage. Furthermore, GGA effectively suppressed LPS-induced cytokine release, attenuated levels of reactive oxygen species (ROS) and malondialdehyde, and bolstered antioxidant-related parameters within the intestinal tissue of LPS-stimulated mice. Mechanistically, GGA significantly increased HSP70 expression and promoted E3 ubiquitin ligase CHIP to play the role in ubiquitination and degradation of karyopherin-α2 (KPNA2), resulting in inhibition of nuclear translocation of NF-κB and reduced NOX1, NOX2 and NOX4 expression. The inhibitory action of GGA on cytokine release and ROS generation was abolished by CHIP knockdown in IEC-6 cells treated with LPS. Simultaneously, the downregulation of CHIP reversed the suppressive role of GGA in the LPS-induced NF-κB activation and the expression of NOX1, NOX2 and NOX4 in IEC-6 cells. The effects of GGA on mitigating intestinal damage, inflammation and oxidative stress caused by LPS were eliminated in CHIP knockout mice. Our results demonstrate that the protective effect of GGA against LPS-caused intestinal injury of mice is dependent on CHIP activation, which promotes KPNA2 degradation and restrains translocation of NF-κB into nucleus, leading to suppressing LPS-induced inflammatory response and oxidative stress.

Sepsis is a severe inflammatory disorder that can lead to life-threatening multiple organ injury. Lipopolysaccharide (LPS)-induced inflammation is the leading cause of multiple organ failure in sepsis. This study aimed to explore the effect of a novel agent, 2-(4-hydroxy-3-methoxyphenyl)-benzothiazole (YL-109), on LPS-induced multiple organ injury and the molecular mechanisms underlying these processes. The results showed that YL-109 protected against LPS-induced high mortality, cardiac dysfunction, pulmonary and intestinal injury through inhibiting the proinflammatory response, NLRP3 expression and pyroptosis-associated indicators in mouse tissues. YL-109 suppressed LPS-initiated cytokine release, pyroptosis and pyroptosis-related protein expression in HL-1, IEC-6 and MLE-12 cells, which was consistent with the results of the in vivo experiments. Mechanistically, YL-109 reduces phosphorylated ERK (extracellular signal-regulated kinase) levels and NF-κB activation, which are achieved through upregulating CHIP (carboxy terminus of Hsc70-interacting protein) expression, thereby inhibiting c-Jun and c-Fos activation as well as NLRP3 expression. As an E3 ligase, CHIP overexpression obviously promoted the degradation of phosphorylated ERK and inhibited the expression of NF-κB-mediated NLRP3 in cells stimulated with LPS. The protective effects of YL-109 against cardiac, pulmonary and intestinal damage, inflammation and pyroptosis caused by LPS were eliminated in CHIP knockout mice. Our results not only reveal the protective effect and molecular mechanism of YL-109 against LPS-mediated organs damage but also provide additional insights into the effect of CHIP on negatively regulating pyroptosis and inflammatory pathways.

Doxorubicin (DOX)-related cardiotoxicity has been recognized as a serious complication of cancer chemotherapy. Effective targeted strategies for myocardial protection in addition to DOX treatment are urgently needed. The purpose of this paper was to determine the therapeutic effect of berberine (Ber) on DOX-triggered cardiomyopathy and explore the underlying mechanism. Our data showed that Ber markedly prevented cardiac diastolic dysfunction and fibrosis, reduced cardiac malondialdehyde (MDA) level and increased antioxidant superoxide dismutase (SOD) activity in DOX-treated rats. Moreover, Ber effectively rescued the DOX-induced production of reactive oxygen species (ROS) and MDA, mitochondrial morphological damage and membrane potential loss in neonatal rat cardiac myocytes and fibroblasts. This effect was mediated by increases in the nuclear accumulation of nuclear erythroid factor 2-related factor 2 (Nrf2) and levels of heme oxygenase-1 (HO-1) and mitochondrial transcription factor A (TFAM). We also found that Ber suppressed the differentiation of cardiac fibroblasts (CFs) into myofibroblasts, as indicated by decreased expression of α-smooth muscle actin (α-SMA), collagen I and collagen III in DOX-treated CFs. Pretreatment with Ber inhibited ROS and MDA production and increased SOD activity and the mitochondrial membrane potential in DOX-challenged CFs. Further investigation indicated that the Nrf2 inhibitor trigonelline reversed the protective effect of Ber on both cardiomyocytes and CFs after DOX stimulation. Taken together, these findings demonstrated that Ber effectively alleviated DOX-induced oxidative stress and mitochondrial damage by activating the Nrf2-mediated pathway, thereby leading to the prevention of myocardial injury and fibrosis. The current study suggests that Ber is a potential therapeutic agent for DOX-induced cardiotoxicity that exerts its effects by activating Nrf2.

Sepsis is a dysregulated systemic inflammatory response caused by infection that leads to multiple organ injury and high mortality without effective treatment. Corilagin, a natural polyphenol extracted from traditional Chinese herbs, exhibits strong anti-inflammatory properties. However, the role for Corilagin in lipopolysaccharide (LPS)-induced sepsis and the molecular mechanisms underlying this process have not been completely explored. Here we determine the effect of Corilagin on LPS-treated mice and use a screening approach integrating surface plasmon resonance with liquid chromatography-tandem mass spectrometry (SPR-LC-MS/MS) to further explore the therapeutic target of Corilagin. We discovered that Corilagin significantly prolonged the survival time of septic mice, attenuated the multi-organ injury and the expression of pyroptosis-related proteins in tissues of LPS-treated mice. In vitro studies revealed that Corilagin inhibited pyroptosis and NLRP3 inflammasome activation in LPS-treated macrophages followed with ATP stimulation, as reflected by decreased levels of GSDMD-NT and activated caspase-1, and reduced ASC specks formation. Mechanistically, Corilagin alleviated the formation of ASC specks and blocked the interaction of ASC and pro-caspase1 by competitively binding with the caspase recruitment domain (CARD) of ASC. Additionally, Corilagin interrupted the TLR4-MyD88 interaction through targeting TIR domain of MyD88, leading to the inhibition of NF-κB activation and NLRP3 production. In addition, Corilagin downregulated genes associated with several inflammatory responses and inflammasome-related signaling pathways in LPS-stimulated macrophages. Overall, our results indicate that the inhibitory effect of Corilagin on pyroptosis through targeting TIR domain of MyD88 and binding the CARD domain of ASC in macrophages plays an essential role in protection against LPS-induced sepsis.

ABSTRACT:Pulmonary fibrosis is an important factor affecting the prognosis of severe septic patients with acute lung injury. The objective of this study was to explore the effect of norepinephrine (NE) and α 2 -adrenoreceptor (AR) on sepsis-associated pulmonary fibrosis and the mechanism underlying these effects. We found pulmonary fibrotic changes, and increased NE production and α 2A -AR expression in the pulmonary tissue of mice subjected to cecal ligation and puncture surgery. Reserpine and yohimbine alleviated pulmonary fibrosis in mice with sepsis by exhausting NE derived from the lung's adrenergic nerve and blocking α 2 -AR, respectively. There was no significant difference in the expression of the three α 1 -AR subtypes. The effect of NE on promoting pulmonary fibroblast differentiation in vitro was suppressed by yohimbine. Both the protein and mRNA expression levels of α 2A -AR were increased in pulmonary fibroblasts treated with LPS. Clonidine, a selective α 2 -AR agonist, enhanced LPS-induced differentiation in pulmonary fibroblasts, as indicated by the increase in α-smooth muscle actin and collagen I/III, which was mitigated by inhibiting PKC and p38. Further in vivo results indicated that yohimbine alleviated pulmonary fibrosis and inhibited the phosphorylation of PKC, p38, and Smad2/3 in lung tissue of mice exposed to LPS for 4 weeks. Clonidine showed the opposite effect to yohimbine, which aggravated LPS-induced pulmonary fibrosis. These findings demonstrated that the sepsis-induced increase in NE promoted fibroblast differentiation via activating α 2 -AR. Blockage of α 2 -AR effectively ameliorated sepsis-associated pulmonary fibrosis by abolishing NE-induced lung fibroblast differentiation and inhibiting the PKC-p38-Smad2/3 pathway.

Cardiac dysfunction has been recognized as a major contributor to mortality in sepsis, which is closely associated with inflammatory reactions. The carboxy terminus of Hsc70-interacting protein (CHIP), a U-box E3 ubiquitin ligase, defends against cardiac injury caused by other factors, but its role in sepsis-induced cardiac dysfunction has yet to be determined. The present study was designed to investigate the effects of CHIP on cardiac dysfunction caused by sepsis and the molecular mechanisms underlying these processes. We discovered that the CHIP level decreased gradually in the heart at different time points after septic model construction. The decline in CHIP expression of lipopolysaccharide (LPS)-stimulated cardiomyocytes was related to c-Jun activation that inhibited the transcription of CHIP. Functional biology experiments indicated that CHIP bound directly to karyopherin-α 2 (KPNA2) and promoted its degradation through polyubiquitination in cardiomyocytes. CHIP overexpression in cardiomyocytes obviously inhibited LPS-initiated release of TNF-α and IL-6 by promoting KPNA2 degradation, reducing NF-κB translocation into the nucleus. Consistent with the in vitro results, data obtained from animal experiments indicated that septic transgenic mice with heart-specific CHIP overexpression showed a weaker proinflammatory response and reduced cardiac dysfunction than septic control mice. Furthermore, we found that the therapeutic effect of compound YL-109 on cardiac dysfunction in septic mice was due to the upregulation of myocardial CHIP expression. These findings demonstrated that sepsis-initiated the activation of c-Jun suppressed CHIP transcription. CHIP directly promoted ubiquitin-mediated degradation of KPNA2, which reduced the production of proinflammatory cytokines by inhibiting the translocation of NF-κB from the cytoplasm into the nucleus in myocardium, thereby attenuating sepsis-induced cardiac dysfunction.

Dexmedetomidine (DEX), a selective α2 adrenergic receptor (AR) agonist, is commonly used as a sedative drug during critical illness. In the present study, we explored a novel accelerative effect of DEX on cardiac fibroblast (CF) differentiation mediated by LPS and clarified its potential mechanism. LPS apparently increased the expression of α-SMA and collagen I/III and the phosphorylation of p38 and Smad-3 in the CFs of mice. These effects were significantly enhanced by DEX through increasing α2A-AR expression in CFs after LPS stimulation. The CFs from α2A-AR knockout mice were markedly less sensitive to DEX treatment than those of wild-type mice. Inhibition of protein kinase C (PKC) abolished the enhanced effects of DEX on LPS-induced differentiation of CFs. We also found that the α-SMA level in the second-passage CFs was much higher than that in the nonpassage and first-passage CFs. However, after LPS stimulation, the TNF-α released from the nonpassage CFs was much higher than that in the first- and second-passage CFs. DEX had no effect on LPS-induced release of TNF-α and IL-6 from CFs. Further investigation indicated that DEX promoted cardiac fibrosis and collagen I/III synthesis in mice exposed to LPS for four weeks. Our results demonstrated that DEX effectively accelerated LPS-induced differentiation of CFs to myofibroblasts through the PKC-p38-Smad2/3 signaling pathway by activating α2A-AR.