The CaMK Family Differentially Promotes Necroptosis and Mouse Cardiac Graft Injury and Rejection.

Organ transplantation is associated with various forms of programmed cell death which can accelerate transplant injury and rejection. Targeting cell death in donor organs may represent a novel strategy for preventing allograft injury. We have previously demonstrated that necroptosis plays a key role in promoting transplant injury. Recently, we have found that mitochondria function is linked to necroptosis. However, it remains unknown how necroptosis signaling pathways regulate mitochondrial function during necroptosis. In this study, we investigated the receptor-interacting protein kinase 3 (RIPK3) mediated mitochondrial dysfunction and necroptosis. We demonstrate that the calmodulin-dependent protein kinase (CaMK) family members CaMK1, 2, and 4 form a complex with RIPK3 in mouse cardiac endothelial cells, to promote trans-phosphorylation during necroptosis. CaMK1 and 4 directly activated the dynamin-related protein-1 (Drp1), while CaMK2 indirectly activated Drp1 via the phosphoglycerate mutase 5 (PGAM5). The inhibition of CaMKs restored mitochondrial function and effectively prevented endothelial cell death. CaMKs inhibition inhibited activation of CaMKs and Drp1, and cell death and heart tissue injury (n = 6/group, p < 0.01) in a murine model of cardiac transplantation. Importantly, the inhibition of CaMKs greatly prolonged heart graft survival (n = 8/group, p < 0.01). In conclusion, CaMK family members orchestrate cell death in two different pathways and may be potential therapeutic targets in preventing cell death and transplant injury.

Mitogenomic analysis and phylogenetic relationships of Agrilinae: Insights into the evolutionary patterns of a diverse buprestid subfamily.

Agrilinae is the largest subfamily in Buprestidae, which includes the four tribes, namely Coraebini, Agrilini, Aphanisticini, and Tracheini. However, there is a need to verify the evolutionary relationships among the taxa in Buprestidae. Thus, to explore the phylogenetic position of Aphanisticini, the mitochondrial genomes of Endelus continentalis and Cantonius szechuanensis were sequenced using next-generation sequencing technology. Three other mitogenomes of agriline beetles, Agrilus discalis, Sambus kanssuensis, and Habroloma sp., were also sequenced for the phylogenetic analyses. The divergence time of Buprestidae was estimated based on the mitogenomes. The general features of the known mitogenomes of Agrilinae were compared, analyzed, and summarized. Out of these five species, S. kanssuensis had the shortest mitogenome length (15,411), while Habroloma sp. had the longest (16,273). The gene arrangement of the five new sequences was identical to that of the reported buprestid mitogenomes. The Ka/Ks ratios of Meliboeus (0.79) and Endelus (0.78) were significantly larger than those of the other agriline genera. The results of the phylogeny indicated that Aphanisticini was more closely related to Tracheini and that the genus Sambus separated from the base of the Agrilinae clade at about 130 Ma. Moreover, Aphanisticini and Tracheini diverged at around 26 Ma.

First mitochondrial genome of subfamily Julodinae (Coleoptera, Buprestidae) with its phylogenetic implications.

Complete mitochondrial genomes of three species of the family Buprestidae were sequenced, annotated, and analyzed in this study. To explore the mitogenome features of the subfamily Julodinae and verify its phylogenetic position, the complete mitogenome of Julodisvariolaris was sequenced and annotated. The complete mitogenomes of Ptosimachinensis and Chalcophorajaponica were also provided for the phylogenetic analyses within Buprestidae. Compared to the known mitogenomes of Buprestidae species varied from 15,499 bp to 16,771 bp in length, three newly sequenced mitogenomes were medium length (15,759-16,227 bp). These mitogenomes were encoded 37 typical mitochondrial genes. Among the three studied mitogenomes, Leu2 (L2), Ser2 (S2), and Pro (P) were the three most frequently encoded amino acids. Within the Buprestidae, the heterogeneity in sequence divergences of Agrilinae was highest, whereas the sequence homogeneity of Chrysochroinae was highest. Moreover, phylogenetic analyses were performed based on nucleotide matrix (13 PCGs + 2 rRNAs) among the available sequenced species of Buprestidae using Bayesian Inference and Maximum Likelihood methods. The results showed that the Julodinae was closely related to the subfamily Polycestinae. Meanwhile, the genera Melanophila, Dicerca, and Coomaniella were included in Buprestinae, which was inconsistent with the current classification system of Buprestidae. These results could contribute to further studies on genetic diversity and phylogeny of Buprestidae.

The larval, pupal and mitogenomic characteristics of Agrilusadelphinus Kerremans, 1895 (Coleoptera, Buprestidae) from China.

In this study, the larva and pupa of Agrilusadelphinus are described and illustrated. DNA barcoding (COI gene) was used to associate the larval and pupal stages with adults based on the maximum-likelihood method. In the resulting phylogenetic tree, species from the same species-group were found to be clustered on a branch with high support value. To better understand A.adelphinus, the complete mitochondrial genome of this species was also sequenced and annotated. Comparing this genome to the known mitogenomes of Agrilus species, the newly sequenced genome is shorter, with 15,732 bp. However, its whole mitogenome composition and gene orientation were consistent with that of most species of Buprestidae. In the mitogenome of A.adelphinus, the ATGATAG sequence was observed between ATP8 and ATP6, which is ATGATAA in other insect mitogenomes. Leu2, Phe, Ile, Gly, and Ser2 were the five most frequently encoded amino acids. The results further prove that DNA barcoding can remove the limitation of traditional taxonomy which cannot identify to species all developmental stages. This study also provides valuable molecular and morphological data for species identification and phylogenetic analyses of the genus Agrilus.

Constructing and Validating a Pyroptosis-Related Genes Prognostic Signature for Stomach Adenocarcinoma and Immune Infiltration: Potential Biomarkers for Predicting the Overall Survival.

Background: Stomach adenocarcinoma (STAD) is a kind of cancer that begins in the stomach cells and has a poor overall survival rate. Following resection surgery, chemotherapy has been suggested as a curative method for stomach cancer. However, it is ineffective. Pyroptosis, a kind of inflammatory programmed cell death, has been shown to play a significant role in the development and progression of STAD. However, whether pyroptosis-related genes (PRGs) can be utilized to predict the diagnosis and prognosis of gastric cancer remains unknown. Method: The research measured at predictive PRGs in STAD samples from TCGA and GEO. Lasso regression was used to build the prediction model. Coexpression analysis revealed that gene expression was linked to pyroptosis. PRGs were found to be overexpressed in high-risk individuals, implying that they could be used in a model to predict STAD prognosis. Result: Immunological and tumor-related pathways were discovered using GSEA. In STAD patients, the genes GPX3, PDGFRL, RGS2, and SERPINE1 may be connected to the cancer process. The levels of expression also differed between the two risk groups. Conclusion: The purpose of this study is to identify and verify STAD-associated PRGs that can effectively guide prognosis and the immunological milieu in STAD patients as well as offer evidence for the development of pyroptosis-related molecularly targeted therapeutics. Therefore, PRGs and the link between immunological and PRGs in STAD may be therapeutic targets.

Potential biomarkers for predicting the overall survival outcome of kidney renal papillary cell carcinoma: an analysis of ferroptosis-related LNCRNAs.

BACKGROUND: Kidney renal papillary cell carcinoma (KIRP) is a dangerous cancer, which accounts for 15-20% of all kidney malignancies. Ferroptosis is a rare kind of cell death that overcomes medication resistance. Ferroptosis-related long non-coding RNAs (LNCRNAs) in KIRP, remain unknown. METHOD: We wanted to express how ferroptosis-related LNCRNAs interact with immune cell infiltration in KIRP. Gene set enrichment analysis in the GO and KEGG databases were used to explore gene expression enrichment. The prognostic model was constructed using Lasso regression. In addition, we also analyzed the modifications in the tumor microenvironment (TME) and immunological association. RESULT: The expression of LNCRNA was closely connected to the ferroptosis, according to co-expression analyses. CASC19, AC090197.1, AC099850.3, AL033397.2, LINC00462, and B3GALT1-AS1 were found to be significantly increased in the high-risk group, indicating that all of these markers implicates the malignancy processes for KIRP patients and may be cancer-promoting variables. LNCTAM34A and AC024022.1 were shown to be significantly elevated in the low-risk group; these might represent as the KIRP tumor suppressor genes. According to the TCGA, CCR, and inflammation-promoting genes were considered to be significantly different between the low-risk and high-risk groups. The expression of CD160, TNFSF4, CD80, BTLA, and TNFRSF9 was different in the two risk groups. CONCLUSION: LNCRNAs associated with ferroptosis were linked to the occurrence and progression of KIRP. Ferroptosis-related LNCRNAs and immune cell infiltration in the TME may be potential biomarkers in KIRP that should be further investigated.

First Report of Complete Mitochondrial Genome in the Tribes Coomaniellini and Dicercini (Coleoptera: Buprestidae) and Phylogenetic Implications.

The complete mitochondrial genomes (mitogenomes) of the tribes Coomaniellini and Dicercini were sequenced and described in this study, including Coomaniella copipes (16,196 bp), Coomaniella dentata (16,179 bp), and Dicerca corrugata (16,276 bp). These complete mitogenomes are very similar in length and encoded 37 typical mitochondrial genes, including 22 transfer RNA genes (tRNAs), 2 ribosomal RNA genes (rRNAs) and 13 protein-coding genes (PCGs). Most of PCGs had typical ATN start codons and terminated with TAR. Among these mitogenomes, Leu2 (L2), Ile (I), Ser2 (S2), and Phe (F) were the four most frequently encoded amino acids. Moreover, phylogenetic analyses were performed based on three kinds of nucleotide matrixes (13 PCGs, 2 rRNAs, and 13 PCGs + 2 rRNAs) among the available sequenced species of the family Buprestidae using Bayesian inference and Maximum-likelihood methods. The results showed that a Chrysochroninae species interspersed in Buprestinae, and Coomaniellini is more closely related to Dicercini than Melanophilini. Moreover, the clade of Buprestidae was well separated from outgroups and the monophyly of Agrilinae is confirmed again. Our whole mitogenome phylogenetic results support that the genus Dicerca can be transferred from Chrysochroinae to Buprestinae; whether Dicercini can be completely transferred remains to be further verified after enriching samples. Our results have produced new complete mitogenomic data, which will provide information for future phylogenetic and taxonomic research.

S100A8 as a Promising Biomarker and Oncogenic Immune Protein in the Tumor Microenvironment: An Integrative Pancancer Analysis.

Background: S100 Calcium Binding Protein A8 (S100A8) is beneficial for cancer immunotherapy. However, the processes underlying its therapeutic potential have not been completely studied. Methods: The Cancer Genome Atlas provides raw data on 33 different cancer types. GEO made available GSE67501, GSE78220, and IMvigor210. We investigated S100A8's genetic changes, expression patterns, and survival studies. The linkages between S100A8 and TME, as well as its association with immunological processes/elements and the major histocompatibility complex, were explored to effectively understand the role of S100A8 in cancer immunotherapy. Three distinct immunotherapeutic cohorts were employed to examine the relationship between S100A8 and immunotherapeutic response. Results: S100A8 expression was high in tumor tissue. The overexpression of S100A8 is associated with poor clinical outcome in patients with overall survival. S100A8 is associated with immune cell infiltration, immunological modulators, and immunotherapeutic indicators. S100A8 overexpression is connected to immune-related pathways. However, no statistically significant connection between S100A8 and immunotherapeutic response was identified. Conclusions: S100A8 may be a reliable biomarker for tumor prognosis and a viable prospective therapeutic target for human cancer immunotherapy (e.g., GBM, KIRC, LGG, and LIHC).

Disaggregation of Ploidy, Gender, and Genotype Effects on Wood and Fiber Traits in a Diploid and Triploid Hybrid Poplar Family.

Triploid breeding based on unilateral sexual polyploidization is an effective approach for genetic improvement of Populus, which can integrate heterosis and ploidy vigor in an elite variety. However, the phenotypic divergence of unselected allotriploids with the same cross-combination remains poorly understood, and the contributions of ploidy, gender, and genotype effects on phenotypic variation are still unclear. In this study, wood and fiber traits, including basic density (BD), lignin content (LC), fiber length (FL), fiber width (FW), and fiber length/width (FL/W), were measured based on a 10-year-old clonal trial, including full-sib diploid and triploid hybrids of (Populus pseudo-simonii × P. nigra 'Zheyin3#') × P. × beijingensis, and contributions of ploidy, gender, and genotype effects on the variation of these traits, were disaggregated to enhance our understanding of triploid breeding. We found a significant phenotypic variation for all measured traits among genotypes. All the wood and fiber traits studied here underwent strong clonal responses with high repeatabilities (0.55-0.76). The Pearson's correlation analyses based on the best linear unbiased predictors (BLUPs) revealed that BD was significantly positively correlated with FL (r = 0.65, p = 0.030), suggesting that BD could be improved together with FL during triploid breeding. The FL of the triploids was significantly larger than that of the diploids (p < 0.001), suggesting that ploidy strongly affected the variation of FL traits. The difference between females and males was not significant for any measured trait, implying that gender might not be a major factor for variation in these traits. Further analyses of variance components showed that genotype dominantly contributed to the variation of BD, LC, and FW traits (with 54, 62, and 53% contributions, respectively) and ploidy contributed strongly to variation in FL and FL/W (77 and 50%, respectively). The genetic coefficient of variation (CVG) of triploids for each trait was low, suggesting that it is necessary to produce many triploids for selection or to use different Populus species as parents. Our findings provide new insights into the genetic effects of ploidy, gender, and genotype on wood and fiber traits within a full-sib poplar family, enhancing the understanding of the triploid breeding program of Populus.

In the Tumor Microenvironment, ETS1 Is an Oncogenic Immune Protein: An Integrative Pancancer Analysis.

Background: Previous research suggested that ETS1 (ETS proto-oncogene 1, transcription factor) could be useful for cancer immunotherapy. The processes underlying its therapeutic potential, on the other hand, have yet to be thoroughly investigated. The purpose of this study was to look into the relationship between ETS1 expression and immunity. Methods: TCGA and GEO provide raw data on 33 different cancers as well as GSE67501, GSE78220, and IMvigor210. In addition, we looked at ETS1's genetic changes, expression patterns, and survival studies. The linkages between ETS1 and TME, as well as its association with immunological processes/elements and the major histocompatibility complex, were explored to effectively understand the role of ETS1 in cancer immunotherapy. Three distinct immunotherapeutic cohorts were employed to examine the relationship between ETS1 and immunotherapeutic response. Results: ETS1 expression was shown to be high in tumor tissue. ETS1 overexpression is linked to a worse clinical outcome in individuals with overall survival. Immune cell infiltration, immunological modulators, and immunotherapeutic signs are all linked to ETS1. Overexpression of ETS1 is linked to immune-related pathways. However, no statistically significant link was found between ETS1 and immunotherapeutic response. Conclusions: ETS1 may be a reliable biomarker for tumor prognosis and a viable prospective therapeutic target for human cancer immunotherapy (e.g., KIRP, MESO, BLCA, KIRC, and THYM).

Effective biomarkers and therapeutic targets of nerve-immunity interaction in the treatment of depression: an integrated investigation of the miRNA-mRNA regulatory networks.

BACKGROUND: Major depressive disorder (MDD) is an emotional condition that interferes with sufferers' work and daily life. Numerous studies have found that miRNAs play a significant role in the development of MDD and can be utilized as a biomarker for its diagnosis and therapy. However, there have been few studies on nerve-immunity interaction treatment for the brains of MMD patients. METHODS: The work is performed on microarray data. We analyzed the differences of miRNAs (GSE58105, GSE81152, GSE152267, and GSE182194) and mRNA (GSE19738, GSE32280, GSE44593, GSE53987, and GSE98793) in MDD and healthy samples from GEO datasets. FunRich was used to predict the transcription factors and target genes of the miRNAs, and TF and GO enrichment analyses were performed. Then, by comparing the differential expression of the anticipated target genes and five mRNAs, intersecting mRNAs were discovered. The intersecting genes were submitted to GO and KEGG analyses to determine their functions. These intersecting potential genes and pathways that linked to MDD in neurological and immunological aspects have been identified for future investigation. RESULTS: We discovered five hub genes: KCND2, MYT1L, GJA1, CHL1, and SNAP25, which were all up-regulated genes. However, in MMD, the equivalent miRNAs, hsa-miR-206 and hsa-miR-338-3p, were both down-regulated. These miRNAs can activate or inhibit the T cell receptor signal pathway, JAK-STAT and other signal pathways, govern immune-inflammatory response, neuronal remodeling, and mediate the onset and development of MMD Conclusions: The results of a thorough bioinformatics investigation of miRNAs and mRNAs in MDD showed that miR-338-3P and miR-206 might be effective biomarkers and possible therapeutic targets for the treatment of MDD via nerve-immunity interaction.

Novel necroptosis-related gene signature for predicting the prognosis of pancreatic adenocarcinoma.

Pancreatic adenocarcinoma (PAAD) is a deadly digestive system tumor with a poor prognosis. Recently, necroptosis has been considered as a type of inflammatory programmed cell death. However, the expression of necroptosis-related genes (NRGs) in PAAD and their associations with prognosis remain unclear. NRGs' prediction potential in PAAD samples from The TCGA and GEO datasets was investigated. The prediction model was constructed using Lasso regression. Co-expression analysis showed that gene expression was closely related to necroptosis. NRGs were shown to be somewhat overexpressed in high-risk people even when no other clinical symptoms were present, indicating that they may be utilized in a model to predict PAAD prognosis. GSEA showed immunological and tumor-related pathways in the high-risk group. Based on the findings, immune function and m6A genes differ significantly between the low-risk and high-risk groups. MET, AM25C, MROH9, MYEOV, FAM111B, Y6D, and PPP2R3A might be related to the oncology process for PAAD patients. Moreover, CASKIN2, TLE2, USP20, SPRN, ARSG, MIR106B, and MIR98 might be associated with low-risk patients with PAAD. NRGs and the relationship of the immune function, immune checkpoints, and m6A gene expression with NRGs in PAAD may be considered as potential therapeutic targets that should be further studied.

Cyclophilin D Regulates the Nuclear Translocation of AIF, Cardiac Endothelial Cell Necroptosis and Murine Cardiac Transplant Injury.

Ischemia-reperfusion injury (IRI) is an inevitable consequence of organ transplant procedure and associated with acute and chronic organ rejection in transplantation. IRI leads to various forms of programmed cell death, which worsens tissue damage and accelerates transplant rejection. We recently demonstrated that necroptosis participates in murine cardiac microvascular endothelial cell (MVEC) death and murine cardiac transplant rejection. However, MVEC death under a more complex IRI model has not been studied. In this study, we found that simulating IRI conditions in vitro by hypoxia, reoxygenation and treatment with inflammatory cytokines induced necroptosis in MVECs. Interestingly, the apoptosis-inducing factor (AIF) translocated to the nucleus during MVEC necroptosis, which is regulated by the mitochondrial permeability molecule cyclophilin D (CypD). Furthermore, CypD deficiency in donor cardiac grafts inhibited AIF translocation and mitigated graft IRI and rejection (n = 7; p = 0.002). Our studies indicate that CypD and AIF play significant roles in MVEC necroptosis and cardiac transplant rejection following IRI. Targeting CypD and its downstream AIF may be a plausible approach to inhibit IRI-caused cardiac damage and improve transplant survival.

Comparative efficacy and dysmenorrhea score of 6 object-separated moxibustions for the treatment of Chinese patients with dysmenorrhea: A systematic review and network meta-analysis.

BACKGROUND: Primary dysmenorrhea (PD), one of the most common diseases in women, is known to be effective with object-separated moxibustion. However, because there is no large sample size for comparison, it is difficult to choose the best method for the clinical treatment of these different treatments. Therefore, our aim was to compare and rank different moxibustion methods to determine the most effective treatment method for PD. MATERIALS AND METHODS: A systematic search was carried out in PubMed, Cochrane Central Register of Controlled Trials, China National Knowledge Infrastructure, Wanfang Database, and Chinese Biomedical Literature, to identify the randomized controlled trials (RCTs) investigated the object-separated moxibustion is associated with dysmenorrhea, as well as we also manually checked the bibliographies of eligible studies and topic-related reviews, RCTs from their inception to May 1, 2020. Three investigators read the citations and excluded quasi-randomized trials and trials that were incomplete. We extracted data following a predefined hierarchy. We assessed the studies' risk of bias in accordance with the Cochrane Handbook for Systematic Reviews of Interventions and certainty of evidence using the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) framework. The primary outcomes were efficacy (response rate) and dysmenorrhea scores. We estimated the summary odds ratio (OR) and mean difference (MD) using pairwise and network meta-analyses with random effects. STATA software version 16.0, ADDIS software version 1.16.5, and R software version 3.6.1 were used to statistically analyze all data. RESULTS: Fifty-six RCTs with 5550 patients were included, comparing 6 object-separated moxibustion therapies with acupuncture or oral medicine. All moxibustions were more effective than ibuprofen, with OR ranging between 6.75 (95%CI: 3.58 to 13.22) for moxibustion at the navel. For relieving pain which uses dysmenorrhea score to evaluate, mild moxibustion (MD = -1.42, -4.24 to 0.85) was more effective than others. A total of 24 (42.8%) of 56 trials were rated as having a high risk of bias, 31(55.4%) as moderate, and 1(1.8%) as low, and the certainty of the evidence was moderate. CONCLUSIONS: Mild moxibustion cannot only effectively treat PD but also relieve pain in comparison with ibuprofen. Although GRADE evidence indicate low to moderate for most comparisons, mild moxibustion seems to be an advisable option for PD treatment to relieve symptoms.

Toll-like receptor 3 is an endogenous sensor of cell death and a potential target for induction of long-term cardiac transplant survival.

Inflammation posttransplant is directly linked to cell death programs including apoptosis and necrosis. Cell death leads to the release of cellular contents which can promote inflammation. Targeting of these pathways should be an effective strategy to prevent transplant rejection. Toll-like receptor 3 (TLR3) is emerging as a major endogenous sensor of inflammation. In this study, we assessed the role of TLR3 on cell death and transplant rejection. We showed that TLR3 is highly expressed on mouse microvascular endothelial cell (ECs) and the endothelium of cardiac grafts. We demonstrated that TLR3 interacting with dsRNA or self-RNA triggered apoptosis and necroptosis in ECs. Interestingly, TLR3-induced necroptosis led mitochondrial damage. Inhibition of the mitochondrial membrane permeability molecule Cyclophilin D prevented necroptosis in ECs. In vivo, endothelium damage and activities of caspase-3 and mixed lineage kinase domain-like protein were inhibited in TLR3-/- cardiac grafts compared with C57BL/6 grafts posttransplant (n = 5, p < .001). Importantly, TLR3-/- cardiac grafts had prolonged survival in allogeneic BALB/c mice (mean survival = 121 ± 67 vs. 31 ± 6 days of C57BL/6 grafts, n = 7, p = .002). In summary, our study suggests that TLR3 is an important cell death inducer in ECs and cardiac grafts and thus a potential therapeutic target in preventing cardiac transplant rejection.

Development and Validation of a Prognostic Index Based on Genes Participating in Autophagy in Patients With Lung Adenocarcinoma.

BACKGROUND: Lung adenocarcinoma (LUAD) is a deadly respiratory system malignancy with poor prognosis. Autophagy is essential for the beginning, development, and therapy resistance of cancer. However, the expression of genes participating in autophagy in LUAD and their associations with prognosis remain unclear. METHODS: Predictive genes participating in autophagy in LUAD samples from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) datasets were investigated. TCGA and GEO cohorts were divided into two risk groups, while the low-risk group having a longer overall survival (OS) time. This article aims to point out the interaction between genes participating in autophagy and immune function, immune checkpoints, and m6a in LUAD. The prediction model was designed for exploring least absolute shrinkage and selection operator (LASSO) regression. It has been revealed that gene expression and autophagy are inextricably connected. RESULTS: Genes participating in autophagy were shown to be somewhat overexpressed in the high-risk group even though no different clinical symptoms were present, indicating that they might be used in a model to predict LUAD prognosis. The majority of genes participating in autophagy prognostic signatures controlled immunological and tumor-related pathways, according to gene set enrichment analysis (GSEA). KRT6A, KYNU, IGFBP1, DKK1, PKP2, PLEK2, GAPDH, FLNC, and NTSR1 might be related to the oncology process for LUAD patients. CERS4, CMAHP, and PLEKHB1 have been identified as being associated with low risk in patients with LUAD. Furthermore, the immune function and m6a gene expression differed significantly between the two groups. CONCLUSIONS: Genes participating in autophagy are connected to the development and progression of LUAD. LUAD patients' prognoses are often foreseen utilizing matched prognostic models. Genes participating in autophagy in LUAD may be therapeutic targets that ought to be investigated more.

Mitochondrial permeability regulates cardiac endothelial cell necroptosis and cardiac allograft rejection.

Transplantation is invariably associated with programmed cell death including apoptosis and necrosis, resulting in delayed graft function and organ rejection. We have demonstrated the contribution of necroptosis to mouse microvascular endothelial cell (MVEC) death and transplant rejection. Organ injury results in the opening of mitochondrial permeability transition pores (mPTPs), which can trigger apoptotic molecules release that ultimately results in cell death. The effect of mPTPs in the necroptotic pathway remains controversial; importantly, their role in transplant rejection is not clear. In this study, tumor necrosis factor-α triggered MVECs to undergo receptor-interacting protein kinase family (RIPK1/3)-dependent necroptosis. Interestingly, inhibition of mPTP opening could also inhibit necroptotic cell death. Cyclophilin-D (Cyp-D) is a key regulator of the mPTPs. Both inhibition and deficiency of Cyp-D protected MVECs from necroptosis (n = 3, P < .00001). Additionally, inhibition of Cyp-D attenuated RIPK3-downstream mixed-lineage kinase domain-like protein phosphorylation. In vivo, Cyp-D-deficient cardiac grafts showed prolonged survival in allogeneic BALB/c mice posttransplant compared with wild-type grafts (n = 7, P < .0001). Our study results suggest that the mPTPs may be important mechanistic mediators of necroptosis in cardiac grafts. There is therapeutic potential in targeting cell death via inhibition of the mPTP-regulating molecule Cyp-D to prevent cardiac graft rejection.

Intracellular pH Regulates TRAIL-Induced Apoptosis and Necroptosis in Endothelial Cells.

During ischemia or inflammation of organs, intracellular pH can decrease if acid production exceeds buffering capacity. Thus, the microenvironment can expose parenchymal cells to a reduced extracellular pH which can alter pH-dependent intracellular functions. We have previously shown that while silencing caspase-8 in an in vivo ischemia reperfusion injury (IRI) model results in improved organ function and survival, removal of caspase-8 function in a donor organ can paradoxically result in enhanced receptor-interacting protein kinase 1/3- (RIPK1/3-) regulated necroptosis and accelerated graft loss following transplantation. In our current study, TRAIL- (TNF-related apoptosis-inducing ligand-) induced cell death in vitro at neutral pH and caspase-8 inhibition-enhanced RIPK1-dependent necroptotic death were confirmed. In contrast, both caspase-8 inhibition and RIPK1 inhibition attenuated cell death at a cell pH of 6.7. Cell death was attenuated with mixed lineage kinase domain-like (MLKL) silencing, indicating that MLKL membrane rupture, a distinctive feature of necroptosis, occurs regardless of pH. In summary, there is a distinct regulatory control of apoptosis and necroptosis in endothelial cells at different intracellular pH. These results highlight the complexity of modulating cell death and therapeutic strategies that may need to consider different consequences on cell death dependent on the model.

Necroptosis Is Involved in CD4+ T Cell-Mediated Microvascular Endothelial Cell Death and Chronic Cardiac Allograft Rejection.

BACKGROUND: Despite advances in immunosuppressive therapies, the rate of chronic transplant loss remains substantial. Organ injury involves various forms of cell death including apoptosis and necrosis. We now recognize that early injury of cardiac transplants involves a newly described form of programmed necrotic cell death, termed necroptosis. Because this involves receptor-interacting protein (RIP) kinase 1/3, this study aimed to establish the role of RIP3 in chronic cardiac allograft rejection. METHODS: We used major histocompatibility complex class II mismatched C57BL/6N (H-2; B6) or B6.RIP3 (H-2; RIP3) mice to B6.C-H-2 (H2-Ab1; bm12) mouse cardiac transplantation. Microvascular endothelial cells (MVEC) were developed from B6 and RIP3 cardiac grafts. RESULT: CD4 T cell-mediated cardiac graft rejection is inhibited using RIP3 deficient donor grafts, with reduced cellular infiltration and vasculopathy compared with wild type cardiac grafts. Alloreactive CD4 T cell-mediated MVEC death involves TNFα, Fas ligand (FasL) and granzyme B. Although necroptosis and release of danger molecule high-mobility group box 1 are eliminated by the absence of RIP3, CD4 T cells had attenuated MVEC death through granzyme B and FasL. CONCLUSIONS: CD4 T cell-mediated MVEC death involves in TNFα, FasL and granzyme B. Necroptotic cell death and release of the danger molecule may promote inflammatory responses and transplant rejection. Although loss of RIP3 does not eliminate alloimmune responses, chronic graft injury is reduced. RIP3 is an important therapeutic target but additional granzyme and caspases inhibition is required for sufficiently improving long-term graft survival.

Natural Killer Cells Mediate Long-term Kidney Allograft Injury.

BACKGROUND: Chronic allograft injury remains the leading cause of late kidney graft loss despite improvements in immunosuppressive drugs and a reduction in acute T cell-mediated rejection. We have recently demonstrated that natural killer (NK) cells are cytotoxic to tubular epithelial cells and contribute to acute kidney ischemia-reperfusion injury. The role of NK cells in kidney allograft rejection has not been studied. METHODS: A "parent to F1" kidney transplant model was used to study NK cell-mediated transplant rejection. RESULTS: The C57BL/6 kidneys were transplanted into fully nephrectomized CB6F1 (C57BL/6 x BALB/c) mice. Serum creatinine levels increased from baseline (18.8 ± 5.0 µmol/L to 37.2 ± 5.9 µmol/L, P < 0.001) at 60 days after transplantation. B6Rag-to-CB6F1Rag (B6RagxBALB/cRag) recipients, which lack T and B cells but retain NK cells, showed similar levels of kidney dysfunction 65 days after transplantation (creatinine, 33.8 ± 7.9 µmol/L vs 17.5 ± 5.1 µmol/L in nontransplant Rag mice, P < 0.05). Importantly, depletion of NK cells in Rag1 recipients inhibited kidney injury (24.6 ± 5.5 µmol/L, P < 0.05). Osteopontin, which can activate NK cells to mediate tubular epithelial cell death in vitro, was highly expressed in 60 days kidney grafts. Osteopontin null kidney grafts had reduced injury after transplantation into CB6F1 mice (17.7 ± 3.1 µmol/L, P < 0.001). CONCLUSIONS: Collectively, these data demonstrate for the first time that independent of T and B cells, NK cells have a critical role in mediating long-term transplant kidney injury. Specific therapeutic strategies that target NK cells in addition to conventional immunosuppression may be required to attenuate chronic kidney transplant injury.