The Arabidopsis J-Protein AtDjC5 Facilitates Thermotolerance Likely by Aiding in the ER Stress Response.

AtDjC5 belongs to the J-protein family in Arabidopsis thaliana. Its biological functions remain unclear. In this study, we examined the roles of AtDjC5 in resisting heat stress using reverse genetic analysis. After the seedlings were exposed directly to 44 °C for 90 min, AtDjC5 knockout seedlings displayed decreases in the survival rate, membrane system stability, and cell vitality compared to WT seedlings, indicating that AtDjC5 is involved in plant basal thermotolerance. The AtDjC5 knockout seedlings pre-exposed to 37 °C for 30 min exhibited decreases in the survival rate and total chlorophyll contents and increased cell death when they were subsequently exposed to 45 °C compared to the WT seedlings, indicating that AtDjC5 plays an important role in plant acquired thermotolerance. AtDjC5 was found to localize to the endoplasmic reticulum. The expression of the AtDjC5 gene was induced by heat and TM (an ER stress inducer) treatment. Furthermore, we found that the knockout of AtDjC5 inhibited ER stress-induced autophagy and the expression of ER stress-related genes. Taken together, these results suggest that AtDjC5 facilitates thermotolerance, likely by aiding in the ER stress response.

Risk factors for renal failure and short-term prognosis in patients with spontaneous intracerebral haemorrhage complicated by acute kidney injury.

BACKGROUND: Although acute kidney injury (AKI) is a known risk factor for adverse clinical outcomes in patients with spontaneous intracerebral haemorrhage (SICH), little is known about the predisposing factors that contribute to renal failure and short-term prognosis in the setting of SICH already complicated by AKI. In this study, we aimed to identify the renal failure factors in SICH patents with AKI. METHODS: Five hundred forty-three patients with SICH complicated by differential severities of AKI who were admitted to the First Affiliated Hospital of Fujian Medical University from January 2016 to December 2018 were retrospectively studied. Logistic regression and receiver operator characteristic (ROC) curve analysis were performed to determine the best predictive and discriminative variables. Multivariate Cox regression analysis was performed to identify prognostic factors for renal recovery. RESULTS: In the multivariable adjusted model, we found that hypernatremia, metabolic acidosis, elevated serum creatine kinase, hyperuricaemia, proteinuria, and the use of colloids and diuretics were all independent risk factors for the occurrence of stage 3 AKI in SICH patients. The area under the curve analysis indicated that hypernatremia and hyperuricaemia were predictive factors for stage 3 AKI, and the combination of these two parameters increased their predictability for stage 3 AKI. Kaplan-Meier survival curves revealed that the renal recovery rate in SICH patients with stages 1 and 2 AKI was significantly higher than that in SICH patients with stage 3 AKI. Multivariate Cox regression analysis suggested that hypernatremia and the occurrence of stage 3 AKI are predictors for poor short-term renal recovery. CONCLUSIONS: These findings illustrate that hypernatremia and hyperuricaemia represent potential risk factors for the occurrence of stage 3 AKI in SICH patients. Those patients with hypernatremia and stage 3 AKI were associated with a poor short-term prognosis in renal recovery.

Knockout of AtDjB1, a J-domain protein from Arabidopsis thaliana, alters plant responses to osmotic stress and abscisic acid.

AtDjB1 is a member of the Arabidopsis thaliana J-protein family. AtDjB1 is targeted to the mitochondria and plays a crucial role in A. thaliana heat and oxidative stress resistance. Herein, the role of AtDjB1 in adapting to saline and drought stress was studied in A. thaliana. AtDjB1 expression was induced through salinity, dehydration and abscisic acid (ABA) in young seedlings. Reverse genetic analyses indicate that AtDjB1 is a negative regulator in plant osmotic stress tolerance. Further, AtDjB1 knockout mutant plants (atj1-1) exhibited greater ABA sensitivity compared with the wild-type (WT) plants and the mutant lines with a rescued AtDjB1 gene. AtDjB1 gene knockout also altered the expression of several ABA-responsive genes, which suggests that AtDjB1 is involved in osmotic stress tolerance through its effects on ABA signaling pathways. Moreover, atj1-1 plants exhibited higher glucose levels and greater glucose sensitivity in the post-germination development stage. Applying glucose promoted an ABA response in seedlings, and the promotion was more evident in atj1-1 than WT seedlings. Taken together, higher glucose levels in atj1-1 plants are likely responsible for the greater ABA sensitivity and increased osmotic stress tolerance.

Immune checkpoint inhibitors combined with paclitaxel-based chemotherapy versus chemotherapy alone as first-line treatment in HER2-negative advanced gastric cancer: result of a multicenter retrospective study.

Background: Platinum-based chemotherapy combined with immune checkpoint inhibitors (ICIs) is now becoming the standard first-line therapy for human epidermal growth factor receptor 2 (HER2)-negative advanced gastric cancer (AGC). In China, paclitaxel has shown good efficacy and tolerability in AGC as an alternative for first-line therapy. Combining ICIs with paclitaxel-based chemotherapy may lead to improved tumor immune microenvironment, but evidence in paclitaxel combing with ICIs as first-line regimen is lacking. This multicenter, retrospective research aims to compare effectiveness and tolerability of paclitaxel-based chemotherapy combined with ICIs versus chemotherapy alone as a first-line treatment of HER2-negative AGC in a real-world setting. Methods: Eighty-six patients with HER2-negative AGC were included from 2017 to 2022. Among them, 57 patients received paclitaxel-based chemotherapy plus ICIs, and 29 patients received paclitaxel-based chemotherapy alone. We compared the efficacy and incidence of adverse events between the two therapy options. Results: Significant improvements in median progression-free survival (PFS) (8.77 versus 7.47 months; P=0.04) and median overall survival (OS) (15.70 versus 14.33 months; P=0.04) were observed in the ICIs combined with paclitaxel-based chemotherapy group. The use of ICIs also significantly prolonged the duration of response (DOR) (7.47 versus 4.59 months; P=0.02). Meanwhile, the ICIs plus chemotherapy group demonstrated significantly improved objective response rate (ORR) (50.9% vs. 27.6%; P=0.03) and disease control rate (DCR) (98.3% vs. 82.8%; P=0.01), and the side effects were tolerable. Conclusions: In summary, for HER2-negative AGC, ICIs plus paclitaxel-based chemotherapy is effective with mild toxicities, which should be considered as an alternative first-line therapy regimen.

In silico epitope prediction and evolutionary analysis reveals capsid mutation patterns for enterovirus B.

Enterovirus B (EVB) is a common species of enterovirus, mainly consisting of Echovirus (Echo) and Coxsackievirus B (CVB). The population is generally susceptible to EVB, especially among children. Since the 21st century, EVB has been widely prevalent worldwide, and can cause serious diseases, such as viral meningitis, myocarditis, and neonatal sepsis. By using cryo-electron microscopy, the three-dimensional (3D) structures of EVB and their uncoating receptors (FcRn and CAR) have been determined, laying the foundation for the study of viral pathogenesis and therapeutic antibodies. A limited number of epitopes bound to neutralizing antibodies have also been determined. It is unclear whether additional epitopes are present or whether epitope mutations play a key role in molecular evolutionary history and epidemics, as in influenza and SARS-CoV-2. In the current study, the conformational epitopes of six representative EVB serotypes (E6, E11, E30, CVB1, CVB3 and CVB5) were systematically predicted by bioinformatics-based epitope prediction algorithm. We found that their epitopes were distributed into three clusters, where the VP1 BC loop, C-terminus and VP2 EF loop were the main regions of EVB epitopes. Among them, the VP1 BC loop and VP2 EF loop may be the key epitope regions that determined the use of the uncoating receptors. Further molecular evolution analysis based on the VP1 and genome sequences showed that the VP1 C-terminus and VP2 EF loop, as well as a potential "breathing epitope" VP1 N-terminus, were common mutation hotspot regions, suggesting that the emergence of evolutionary clades was driven by epitope mutations. Finally, footprints showed mutations were located on or near epitopes, while mutations on the receptor binding sites were rare. This suggested that EVB promotes viral epidemics by breaking the immune barrier through epitope mutations, but the mutations avoided the receptor binding sites. The bioinformatics study of EVB epitopes may provide important information for the monitoring and early warning of EVB epidemics and developing therapeutic antibodies.

The role of conformational epitopes in the evolutionary divergence of enterovirus D68 clades: A bioinformatics-based study.

Enterovirus D68 (EV-D68), as one of the major pathogens of paediatric respiratory disease, has been widely spread in the population in recent years. As the basis of virus antigenicity, antigenic epitopes are essential to monitoring the transformation of virus antigenicity. However, there is a lack of systematic studies on the antigenic epitopes of EV-D68. In this study, a bioinformatics-based prediction algorithm for human enteroviruses was used to predict the conformational epitopes of EV-D68. The prediction results showed that the conformational epitopes of EV-D68 were clustered into three sites: site 1, site 2, and site 3. Site 1 was located in the "north rim" region of the canyon near the fivefold axis; site 2 was located in the "puff" region near the twofold axis; and site 3 consisted of two parts, one in the "knob" region on the south rim of the canyon and the other in the threefold axis region. The predicted epitopes overlapped highly with the binding regions of four reported monoclonal antibodies (mAbs), indicating that the predictions were highly reliable. Phylogenetic analysis showed that amino acid mutations in the epitopes of the VP1 BC loop, DE loop, C-terminus, and VP2 EF loop played a crucial role in the evolutionary divergence of EV-D68 clades/subclades and epidemics. This finding indicated that the VP1 BC loop, DE loop, C-terminus, and VP2 EF loop were the most important epitopes of EV-D68. Research on the epitopes of EV-D68 will contribute to outbreak surveillance and to the development of diagnostic reagents and recombinant vaccines.

Bioinformatics-based prediction of conformational epitopes for Enterovirus A71 and Coxsackievirus A16.

Enterovirus A71 (EV-A71), Coxsackievirus A16 (CV-A16) and CV-A10 are the major causative agents of hand, foot and mouth disease (HFMD). The conformational epitopes play a vital role in monitoring the antigenic evolution, predicting dominant strains and preparing vaccines. In this study, we employed a Bioinformatics-based algorithm to predict the conformational epitopes of EV-A71 and CV-A16 and compared with that of CV-A10. Prediction results revealed that the distribution patterns of conformational epitopes of EV-A71 and CV-A16 were similar to that of CV-A10 and their epitopes likewise consisted of three sites: site 1 (on the "north rim" of the canyon around the fivefold vertex), site 2 (on the "puff") and site 3 (one part was in the "knob" and the other was near the threefold vertex). The reported epitopes highly overlapped with our predicted epitopes indicating the predicted results were reliable. These data suggested that three-site distribution pattern may be the basic distribution role of epitopes on the enteroviruses capsids. Our prediction results of EV-A71 and CV-A16 can provide essential information for monitoring the antigenic evolution of enterovirus.

Bioinformatics-based prediction of conformational epitopes for human parechovirus.

Human parechoviruses (HPeVs) are human pathogens that usually cause diseases ranging from rash to neonatal sepsis in young children. HPeV1 and HPeV3 are the most frequently reported genotypes and their three-dimensional structures have been determined. However, there is a lack of systematic research on the antigenic epitopes of HPeVs, which are useful for understanding virus-receptor interactions, developing antiviral agents or molecular diagnostic tools, and monitoring antigenic evolution. Thus, we systematically predicted and compared the conformational epitopes of HPeV1 and HPeV3 using bioinformatics methods in the study. The results showed that both epitopes clustered into three sites (sites 1, 2 and 3). Site 1 was located on the "northern rim" near the fivefold vertex; site 2 was on the "puff"; and site 3 was divided into two parts, of which one was located on the "knob" and the other was close to the threefold vertex. The predicted epitopes highly overlapped with the reported antigenic epitopes, which indicated that the prediction results were accurate. Although the distribution positions of the epitopes of HPeV1 and HPeV3 were highly consistent, the residues varied largely and determined the genotypes. Three amino acid residues, VP3-91N, -92H and VP0-257S, were the key residues for monoclonal antibody (mAb) AM28 binding to HPeV1 and were also of great significance in distinguishing HPeV1 and HPeV3. We also found that two residues, VP1-85N and -87D, might affect the capability of mAb AT12-015 to bind to HPeV3.

RIPK3-MLKL-mediated necroinflammation contributes to AKI progression to CKD.

Necroptosis predominates functionally over apoptosis in the pathophysiology of renal ischemia-reperfusion injury (IRI). Inhibition of the core components of the necroptotic pathway-receptor-interacting protein kinase 1 (RIPK1), RIPK3 or mixed lineage kinase domain-like protein (MLKL) reduced renal injury after ischemia/reperfusion (IR). Necrosis can initiate inflammation, which enhances necrosis in a positive feedback loop, subsequently leading to triggering more inflammation, termed as necroinflammation. However, the mechanisms underlying necroinflammation driven by renal tubular cell necroptosis in progression of AKI to CKD are still largely unknown. Here we showed that the upregulated expression and interactions between RIPK3 and MLKL induced necroptosis of renal proximal tubular cells and contributed to NLRP3 inflammasome activation under the conditions of IRI. Gene deletion of Ripk3 or Mlkl ameliorated renal tubular cell necroptosis, macrophage infiltration and NLRP3 inflammasome activation with a reduction in caspase-1 activation and maturation of IL-1ß, and then finally reduced interstitial fibrogenesis in the long term after IRI. Bone marrow chimeras confirmed that RIPK3-MLKL-dependent necroptosis is responsible for the initiation of the early renal injury after IRI, and then necroptosis triggered NLRP3 inflammasome activation, which subsequently accelerates necroptosis and triggers more inflammation in an auto-amplification loop. These data indicate that necroinflammation driven by RIPK3-MLKL-dependent necroptosis plays a crucial role in the progression of IRI to CKD.

Cisplatin-induced necroptosis in TNFα dependent and independent pathways.

Cisplatin is a chemotherapeutic drug for treatment of many solid tumors. It has been shown to induce apoptosis and/or necrosis in different types of cancer cells. However, the underlying mechanisms remain elusive. In this study, we provide evidences that cisplatin induces necroptosis in receptor-interacting protein 3 (RIP3)-expressing cell lines, but not in cell lines lacking RIP3 protein expression. Deficiency of core components of necroptotic pathway, RIP1, RIP3, or mixed lineage kinase domain-like protein (MLKL) blocked cisplatin-induced cell death in L929 cells. This phenomenon is dependent on RIP1/RIP3/MLKL necrosome formation and translocation to mitochondria-associated membrane (MAM), but only partially via autocrine production of tumor necrosis factor α (TNFα). Moreover, we demonstrate that the mitochondrial permeability transition pore opening (mPTP) opening and reactive oxygen species (ROS) generation is a critical downstream event of the formation of necrosome in cisplatin-induced necroptosis, which is TNFα independent. Deficiency of cyclophilin-D (CypD) partially reduced cisplatin-induced cell death, indicating CypD mediated-mPTP opening plays an important role during cisplatin-induced necroptosis. Both deletion of CypD and TNFα completely blocked cisplatin-induced cell death, suggesting that cisplatin could induce necroptosis through TNFα dependent and independent pathway. These findings provide new insight into the molecular mechanisms underlying cisplatin-induced necroptosis.