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Mitochondria, traditionally recognized as cellular 'powerhouses' due to their pivotal role in energy production, have emerged as multifunctional organelles at the intersection of bioenergetics, metabolic signaling, and immunity. However, the understanding of their exact contributions to immunity and inflammation is still developing. This review first introduces the innovative concept of intracellular immunity, emphasizing how mitochondria serve as critical immune signaling hubs. They are instrumental in recognizing and responding to pathogen and danger signals, and in modulating immune responses. We also propose mitochondria as the leading immune organelles, drawing parallels with the broader immune system in their functions of antigen presentation, immune regulation, and immune response. Our comprehensive review explores mitochondrial immune signaling pathways, their therapeutic potential in managing inflammation and chronic diseases, and discusses cutting-edge methodologies for mitochondrial research. Targeting a broad readership of both experts in mitochondrial functions and newcomers to the field, this review sets forth new directions that could transform our understanding of intracellular immunity and the integrated immune functions of intracellular organelles.

IntroductionVascular smooth muscle cells (VSMCs) are the predominant cell type in the medial layer of the aorta, which plays a critical role in aortic diseases. Innate immunity is the main driving force for cardiovascular diseases. MethodsTo determine the roles of innate immunity in VSMC and aortic pathologies, we performed transcriptome analyses on aortas from ApoE–/– angiotensin II (Ang II)-induced aortic aneurysm (AAA) time course, and ApoE–/– atherosclerosis time course, as well as VSMCs stimulated with danger-associated molecular patterns (DAMPs).ResultsWe made significant findings: 1) 95% and 45% of the upregulated innate immune pathways (UIIPs, based on data of 1226 innate immune genes) in ApoE–/– Ang II-induced AAA at 7 days were different from that of 14 and 28 days, respectively; and AAA showed twin peaks of UIIPs with a major peak at 7 days and a minor peak at 28 days; 2) all the UIIPs in ApoE–/– atherosclerosis at 6 weeks were different from that of 32 and 78 weeks (two waves); 3) analyses of additional 12 lists of innate immune-related genes with 1325 cytokine and chemokine genes, 2022 plasma membrane protein genes, 373 clusters of differentiation (CD) marker genes, 280 nuclear membrane protein genes, 1425 nucleoli protein genes, 6750 nucleoplasm protein genes, 1496 transcription factors (TFs) including 15 pioneer TFs, 164 histone modification enzymes, 102 oxidative cell death genes, 68 necrotic cell death genes, and 47 efferocytosis genes confirmed two-wave inflammation in atherosclerosis and twin-peak inflammation in AAA; 4) DAMPs-stimulated VSMCs were innate immune cells as judged by the upregulation of innate immune genes and genes from 12 additional lists; 5) DAMPs-stimulated VSMCs increased trans-differentiation potential by upregulating not only some of 82 markers of 7 VSMC-plastic cell types, including fibroblast, osteogenic, myofibroblast, macrophage, adipocyte, foam cell, and mesenchymal cell, but also 18 new cell types (out of 79 human cell types with 8065 cell markers); 6) analysis of gene deficient transcriptomes indicated that the antioxidant transcription factor NRF2 suppresses, however, the other five inflammatory transcription factors and master regulators, including AHR, NF-KB, NOX (ROS enzyme), PERK, and SET7 promote the upregulation of twelve lists of innate immune genes in atherosclerosis, AAA, and DAMP-stimulated VSMCs; and 7) both SET7 and trained tolerance-promoting metabolite itaconate contributed to twin-peak upregulation of cytokines in AAA. DiscussionOur findings have provided novel insights on the roles of innate immune responses and nuclear stresses in the development of AAA, atherosclerosis, and VSMC immunology and provided novel therapeutic targets for treating those significant cardiovascular and cerebrovascular diseases.

To determine the roles of endoplasmic reticulum (ER) stress and trained immunity, we performed transcriptome analyses on the thoracic aorta (TA) and abdominal aorta (AA) from the angiotensin II (Ang II)-HFD-ApoE-KO aneurysm model and made significant findings: 1) Ang II bypassed HFD-induced metabolic reprogramming and induced stronger inflammation in AA than in TA; 2) Ang II and HFD upregulated 890 genes in AA versus TA and induced cytokine signaling; 3) Ang II AA and TA upregulated 73 and 68 cytokines, scRNA-Seq identified markers of macrophages and immune cells, cell death regulators, respectively; transdifferentiation markers of neuron, glial, and squamous epithelial cells were upregulated by Ang II-AA and TA; and pyroptosis signaling with IL-1β and caspase-4 were more upregulated in Ang II-AA than in TA; 4) Six upregulated transcriptomes in patients with AAA, Ang II AA, Ang II TA, additional aneurysm models, PPE-AAA and BAPN-Ang II-AAA, were partially overlapped with 10 lists of new ER stress gene sets including 3 interaction protein lists of ER stress regulators ATF6, PERK, and IRE1, HPA ER localization genes, KEGG signal genes, XBP1 transcription targets, ATF4 (PERK) targets, ATF6 targets, thapsigargin ER stress genes, tunicamycin-ER stress genes, respectively; 5) Ang II-AA and TA upregulated ROS regulators, MitoCarta genes, trained immunity genes, and glycolysis genes; and 6) Gene KO transcriptomes indicated that ATF6 and PERK played more significant roles than IRE1 in promoting AAA and trained immunity whereas antioxidant NRF2 inhibited them. Our unprecedented ER-focused transcriptomic analyses have provided novel insights on the roles of ER as an immune organelle in sensing various DAMPs and initiating ER stress that triggers Ang II-accelerated trained immunity and differs susceptibilities of thoracic and abdominal aortas to diseases.

To identify metabolomic reprogramming in early hyperlipidemia, unbiased metabolome was screened in four tissues from ApoE−/− mice fed with high fat diet (HFD) for 3 weeks. 30, 122, 67, and 97 metabolites in the aorta, heart, liver, and plasma, respectively, were upregulated. 9 upregulated metabolites were uremic toxins, and 13 metabolites, including palmitate, promoted a trained immunity with increased syntheses of acetyl-CoA and cholesterol, increased S-adenosylhomocysteine (SAH) and hypomethylation and decreased glycolysis. The cross-omics analysis found upregulation of 11 metabolite synthetases in ApoE‾/‾ aorta, which promote ROS, cholesterol biosynthesis, and inflammation. Statistical correlation of 12 upregulated metabolites with 37 gene upregulations in ApoE‾/‾ aorta indicated 9 upregulated new metabolites to be proatherogenic. Antioxidant transcription factor NRF2−/− transcriptome analysis indicated that NRF2 suppresses trained immunity-metabolomic reprogramming. Our results have provided novel insights on metabolomic reprogramming in multiple tissues in early hyperlipidemia oriented toward three co-existed new types of trained immunity.

We determined whether gut microbiota-produced trimethylamine (TMA) is oxidized into trimethylamine N-oxide (TMAO) in nonliver tissues and whether TMAO promotes inflammation via trained immunity (TI). We found that endoplasmic reticulum (ER) stress genes were coupregulated with MitoCarta genes in chronic kidney diseases (CKD); TMAO upregulated 190 genes in human aortic endothelial cells (HAECs); TMAO synthesis enzyme flavin-containing monooxygenase 3 (FMO3) was expressed in human and mouse aortas; TMAO transdifferentiated HAECs into innate immune cells; TMAO phosphorylated 12 kinases in cytosol via its receptor PERK and CREB, and integrated with PERK pathways; and PERK inhibitors suppressed TMAO-induced ICAM-1. TMAO upregulated 3 mitochondrial genes, downregulated inflammation inhibitor DARS2, and induced mitoROS, and mitoTEMPO inhibited TMAO-induced ICAM-1. β-Glucan priming, followed by TMAO restimulation, upregulated TNF-α by inducing metabolic reprogramming, and glycolysis inhibitor suppressed TMAO-induced ICAM-1. Our results have provided potentially novel insights regarding TMAO roles in inducing EC activation and innate immune transdifferentiation and inducing metabolic reprogramming and TI for enhanced vascular inflammation, and they have provided new therapeutic targets for treating cardiovascular diseases (CVD), CKD-promoted CVD, inflammation, transplantation, aging, and cancer.

IntroductionNon-alcoholic fatty liver disease (NAFLD) has a global prevalence of 25% of the population and is a leading cause of cirrhosis and hepatocellular carcinoma. NAFLD ranges from simple steatosis (non-alcoholic fatty liver) to non-alcoholic steatohepatitis (NASH). Hepatic macrophages, specifically Kupffer cells (KCs) and monocyte-derived macrophages, act as key players in the progression of NAFLD. Caspases are a family of endoproteases that provide critical connections to cell regulatory networks that sense disease risk factors, control inflammation, and mediate inflammatory cell death (pyroptosis). Caspase-11 can cleave gasdermin D (GSDMD) to induce pyroptosis and specifically defends against bacterial pathogens that invade the cytosol. However, it’s still unknown whether high fat diet (HFD)-facilitated gut microbiota-generated cytoplasmic lipopolysaccharides (LPS) activate caspase-11 and promote NAFLD.MethodsTo examine this hypothesis, we performed liver pathological analysis, RNA-seq, FACS, Western blots, Seahorse mitochondrial stress analyses of macrophages and bone marrow transplantation on HFD-induced NAFLD in WT and Casp11–/– mice.Results and DiscussionOur results showed that 1) HFD increases body wight, liver wight, plasma cholesterol levels, liver fat deposition, and NAFLD activity score (NAS score) in wild-type (WT) mice; 2) HFD increases the expression of caspase-11, GSDMD, interleukin-1β, and guanylate-binding proteins in WT mice; 3) Caspase-11 deficiency decreases fat liver deposition and NAS score; 4) Caspase-11 deficiency decreases bone marrow monocyte-derived macrophage (MDM) pyroptosis (inflammatory cell death) and inflammatory monocyte (IM) surface GSDMD expression; 5) Caspase-11 deficiency re-programs liver transcriptomes and reduces HFD-induced NAFLD; 6) Caspase-11 deficiency decreases extracellular acidification rates (glycolysis) and oxidative phosphorylation (OXPHOS) in inflammatory fatty acid palmitic acid-stimulated macrophages, indicating that caspase-11 significantly contributes to maintain dual fuel bioenergetics—glycolysis and OXPHOS for promoting pyroptosis in macrophages. These results provide novel insights on the roles of the caspase-11-GSDMD pathway in promoting hepatic macrophage inflammation and pyroptosis and novel targets for future therapeutic interventions involving the transition of NAFLD to NASH, hyperlipidemia, type II diabetes, metabolic syndrome, metabolically healthy obesity, atherosclerotic cardiovascular diseases, autoimmune diseases, liver transplantation, and hepatic cancers.

Abstract Extra-virgin olive oil (EVOO) is a critical component of the Mediterranean diet, which has been found beneficial to human health. Bitterness is often positively associated with the presence of phenolic compounds in EVOO. There are twenty-five bitter taste receptors (TAS2Rs) in humans, each of which responds to specific bitter tastants. The identity of phenolic compounds and the bitter taste receptors they stimulate remain unknown. In this study, we isolated 12 phenolic and secoiridoid compounds from the olive fruit and the oil extracted from it, and tested their ability to stimulate bitter taste receptor activity, using a calcium mobilization functional assay. Our results showed that seven out of twelve studied compounds activated TAS2R8, and five of them activated TAS2R1, TAS2R8, and TAS2R14. The phenolic compounds oleuropein aglycon and ligstroside aglycon were the most potent bitter tastants in olive oil. TAS2R1 and TAS2R8 were the major bitter taste receptors activated most potently by these phenolic compounds. The results obtained here could be utilized to predict and control the bitterness of olive oil based on the concentration of specific bitter phenolics produced during the milling process of olives.

Most patients with end-stage renal disease (ESRD) and advanced chronic kidney disease (CKD) choose hemodialysis as their treatment of choice. Thus, upper-extremity veins provide a functioning arteriovenous access to reduce dependence on central venous catheters. However, it is unknown whether CKD reprograms the transcriptome of veins and primes them for arteriovenous fistula (AVF) failure. To examine this, we performed transcriptomic analyses of bulk RNA sequencing data of veins isolated from 48 CKD patients and 20 non-CKD controls and made the following findings: (1) CKD converts veins into immune organs by upregulating 13 cytokine and chemokine genes, and over 50 canonical and noncanonical secretome genes; (2) CKD increases innate immune responses by upregulating 12 innate immune response genes and 18 cell membrane protein genes for increased intercellular communication, such as CX3CR1 chemokine signaling; (3) CKD upregulates five endoplasmic reticulum protein-coding genes and three mitochondrial genes, impairing mitochondrial bioenergetics and inducing immunometabolic reprogramming; (4) CKD reprograms fibrogenic processes in veins by upregulating 20 fibroblast genes and 6 fibrogenic factors, priming the vein for AVF failure; (5) CKD reprograms numerous cell death and survival programs; (6) CKD reprograms protein kinase signal transduction pathways and upregulates SRPK3 and CHKB; and (7) CKD reprograms vein transcriptomes and upregulates MYCN, AP1, and 11 other transcription factors for embryonic organ development, positive regulation of developmental growth, and muscle structure development in veins. These results provide novel insights on the roles of veins as immune endocrine organs and the effect of CKD in upregulating secretomes and driving immune and vascular cell differentiation.

There is a high incidence of tobacco use among intravenous opioid drug users. It is well established that opioids and tobacco smoke induce a degree of immune activation, and recent work suggests that the combination of these drugs promotes further activation of the immune system. Our approach involved the treatment of wild-type mice with cigarette smoke (SM) for a period of eight weeks, and the chronic continuous administration of morphine (M) via mini-pumps for the final four weeks. In an effort to examine the responses of CD4+CD25highCD127low regulatory T (Treg) cells, the major immune suppressive cell type, to the combined chronic administration of SM and M, we determined the frequency of these cells in the spleen, lymph nodes and lungs. Flow cytometric analyses showed that SM and M individually, and the combination (SM + M) have differential effects on the numbers of Treg in the spleen, lymph node, and lung. Either SM or M alone increased Treg cell numbers in the spleen, but SM+M did not. Furthermore, SM + M decreased Treg cell numbers in the lymph node and lung. We then performed RNA-Seq on Treg cells from mice treated with SM, M, or SM + M, and we found that the S + M induced a number of significant changes in the transcriptome, that were not as apparent following treatment with either SM or M alone. This included an activation of TWEAK, PI3K/AKT and OXPHOS pathways and a shift to Th17 immunity. Our results have provided novel insights on tissue Treg cell changes, which we suggest are the result of transcriptomic reprogramming induced by SM, M, and SM + M, respectively. We believe these results may lead to the identification of novel therapeutic targets for suppressing smoke and opioid induced Treg cell impairment.

To determine whether aorta becomes immune organ in pathologies, we performed transcriptomic analyses of six types of secretomic genes (SGs) in aorta and vascular cells and made the following findings: 1) 53.7% out of 21,306 human protein genes are classified into six secretomes, namely, canonical, caspase 1, caspase 4, exosome, Weibel–Palade body, and autophagy; 2) Atherosclerosis (AS), chronic kidney disease (CKD) and abdominal aortic aneurysm (AAA) modulate six secretomes in aortas; and Middle East Respiratory Syndrome Coronavirus (MERS-CoV, COVID-19 homologous) infected endothelial cells (ECs) and angiotensin-II (Ang-II) treated vascular smooth muscle cells (VSMCs) modulate six secretomes; 3) AS aortas upregulate T and B cell immune SGs; CKD aortas upregulate SGs for cardiac hypertrophy, and hepatic fibrosis; and AAA aorta upregulate SGs for neuromuscular signaling and protein catabolism; 4) Ang-II induced AAA, canonical, caspase 4, and exosome SGs have two expression peaks of high (day 7)-low (day 14)-high (day 28) patterns; 5) Elastase induced AAA aortas have more inflammatory/immune pathways than that of Ang-II induced AAA aortas; 6) Most disease-upregulated cytokines in aorta may be secreted via canonical and exosome secretomes; 7) Canonical and caspase 1 SGs play roles at early MERS-CoV infected ECs whereas caspase 4 and exosome SGs play roles in late/chronic phases; and the early upregulated canonical and caspase 1 SGs may function as drivers for trained immunity (innate immune memory); 8) Venous ECs from arteriovenous fistula (AVF) upregulate SGs in five secretomes; and 9) Increased some of 101 trained immunity genes and decreased trained tolerance regulator IRG1 participate in upregulations of SGs in atherosclerotic, Ang-II induced AAA and CKD aortas, and MERS-CoV infected ECs, but less in SGs upregulated in AVF ECs. IL-1 family cytokines, HIF1α, SET7 and mTOR, ROS regulators NRF2 and NOX2 partially regulate trained immunity genes; and NRF2 plays roles in downregulating SGs more than that of NOX2 in upregulating SGs. These results provide novel insights on the roles of aorta as immune organ in upregulating secretomes and driving immune and vascular cell differentiations in COVID-19, cardiovascular diseases, inflammations, transplantations, autoimmune diseases and cancers.

We performed a database mining on 102 transcriptomic datasets for the expressions of 29 m6A-RNA methylation (epitranscriptomic) regulators (m6A-RMRs) in 41 diseases and cancers and made significant findings: (1) a few m6A-RMRs were upregulated; and most m6A-RMRs were downregulated in sepsis, acute respiratory distress syndrome, shock, and trauma; (2) half of 29 m6A-RMRs were downregulated in atherosclerosis; (3) inflammatory bowel disease and rheumatoid arthritis modulated m6A-RMRs more than lupus and psoriasis; (4) some organ failures shared eight upregulated m6A-RMRs; end-stage renal failure (ESRF) downregulated 85% of m6A-RMRs; (5) Middle-East respiratory syndrome coronavirus infections modulated m6A-RMRs the most among viral infections; (6) proinflammatory oxPAPC modulated m6A-RMRs more than DAMP stimulation including LPS and oxLDL; (7) upregulated m6A-RMRs were more than downregulated m6A-RMRs in cancer types; five types of cancers upregulated ≥10 m6A-RMRs; (8) proinflammatory M1 macrophages upregulated seven m6A-RMRs; (9) 86% of m6A-RMRs were differentially expressed in the six clusters of CD4+Foxp3+ immunosuppressive Treg, and 8 out of 12 Treg signatures regulated m6A-RMRs; (10) immune checkpoint receptors TIM3, TIGIT, PD-L2, and CTLA4 modulated m6A-RMRs, and inhibition of CD40 upregulated m6A-RMRs; (11) cytokines and interferons modulated m6A-RMRs; (12) NF-κB and JAK/STAT pathways upregulated more than downregulated m6A-RMRs whereas TP53, PTEN, and APC did the opposite; (13) methionine-homocysteine-methyl cycle enzyme Mthfd1 downregulated more than upregulated m6A-RMRs; (14) m6A writer RBM15 and one m6A eraser FTO, H3K4 methyltransferase MLL1, and DNA methyltransferase, DNMT1, regulated m6A-RMRs; and (15) 40 out of 165 ROS regulators were modulated by m6A eraser FTO and two m6A writers METTL3 and WTAP. Our findings shed new light on the functions of upregulated m6A-RMRs in 41 diseases and cancers, nine cellular and molecular mechanisms, novel therapeutic targets for inflammatory disorders, metabolic cardiovascular diseases, autoimmune diseases, organ failures, and cancers.

To determine the roles of nuclear localization of pro-caspase-1 in human aortic endothelial cells (HAECs) activated by proatherogenic lipid lysophosphatidylcholine (LPC), we examined cytosolic and nuclear localization of pro-caspase-1, identified nuclear export signal (NES) in pro-caspase-1 and sequenced RNAs. We made the following findings: 1) LPC increases nuclear localization of procaspase-1 in HAECs. 2) Nuclear pro-caspase-1 exports back to the cytosol, which is facilitated by a leptomycin B-inhibited mechanism. 3) Increased nuclear localization of pro-caspase-1 by a new NES peptide inhibitor upregulates inflammatory genes in oxidative stress and Th17 pathways; and SUMO activator N106 enhances nuclear localization of pro-caspase-1 and caspase-1 activation (p20) in the nucleus. 4) LPC plus caspase-1 enzymatic inhibitor upregulates inflammatory genes with hypercytokinemia/hyperchemokinemia and interferon pathways, suggesting a novel capsase-1 enzyme-independent inflammatory mechanism. 5) LPC in combination with NES inhibitor and caspase-1 inhibitor upregulate inflammatory gene expression that regulate Th17 activation, endotheli-1 signaling, p38-, and ERK- MAPK pathways. To examine two hallmarks of endothelial activation such as secretomes and membrane protein signaling, LPC plus NES inhibitor upregulate 57 canonical secretomic genes and 76 exosome secretomic genes, respectively, promoting four pathways including Th17, IL-17 promoted cytokines, interferon signaling and cholesterol biosynthesis. LPC with NES inhibitor also promote inflammation via upregulating ROS promoter CYP1B1 and 11 clusters of differentiation (CD) membrane protein pathways. Mechanistically, all the LPC plus NES inhibitor-induced genes are significantly downregulated in CYP1B1-deficient microarray, suggesting that nuclear caspase-1-induced CYP1B1 promotes strong inflammation. These transcriptomic results provide novel insights on the roles of nuclear caspase-1 in sensing DAMPs, inducing ROS promoter CYP1B1 and in regulating a large number of genes that mediate HAEC activation and inflammation. These findings will lead to future development of novel therapeutics for cardiovascular diseases (CVD), inflammations, infections, transplantation, autoimmune disease and cancers. (total words: 284).

To determine whether pro-inflammatory lipid lysophosphatidylinositols (LPIs) upregulate the expressions of membrane proteins for adhesion/signaling and secretory proteins in human aortic endothelial cell (HAEC) activation, we developed an EC biology knowledge-based transcriptomic formula to profile RNA-Seq data panoramically. We made the following primary findings: first, G protein-coupled receptor 55 (GPR55), the LPI receptor, is expressed in the endothelium of both human and mouse aortas, and is significantly upregulated in hyperlipidemia; second, LPIs upregulate 43 clusters of differentiation (CD) in HAECs, promoting EC activation, innate immune trans-differentiation, and immune/inflammatory responses; 72.1% of LPI-upregulated CDs are not induced in influenza virus-, MERS-CoV virus- and herpes virus-infected human endothelial cells, which hinted the specificity of LPIs in HAEC activation; third, LPIs upregulate six types of 640 secretomic genes (SGs), namely, 216 canonical SGs, 60 caspase-1-gasdermin D (GSDMD) SGs, 117 caspase-4/11-GSDMD SGs, 40 exosome SGs, 179 Human Protein Atlas (HPA)-cytokines, and 28 HPA-chemokines, which make HAECs a large secretory organ for inflammation/immune responses and other functions; fourth, LPIs activate transcriptomic remodeling by upregulating 172 transcription factors (TFs), namely, pro-inflammatory factors NR4A3, FOS, KLF3, and HIF1A; fifth, LPIs upregulate 152 nuclear DNA-encoded mitochondrial (mitoCarta) genes, which alter mitochondrial mechanisms and functions, such as mitochondrial organization, respiration, translation, and transport; sixth, LPIs activate reactive oxygen species (ROS) mechanism by upregulating 18 ROS regulators; finally, utilizing the Cytoscape software, we found that three mechanisms, namely, LPI-upregulated TFs, mitoCarta genes, and ROS regulators, are integrated to promote HAEC activation. Our results provide novel insights into aortic EC activation, formulate an EC biology knowledge-based transcriptomic profile strategy, and identify new targets for the development of therapeutics for cardiovascular diseases, inflammatory conditions, immune diseases, organ transplantation, aging, and cancers.

Tregs play vital roles in suppressing atherogenesis. Pathological conditions reshape Tregs and increase Treg-weakening plasticity. It remains unclear how Tregs preserve their function and how Tregs switch into alternative phenotypes in the environment of atherosclerosis. In this study, we observed a great induction of CD4+Foxp3+ Tregs in the spleen and aorta of ApoE–/– mice, accompanied by a significant increase of plasma IL-35 levels. To determine if IL-35 devotes its role in the rise of Tregs, we generated IL-35 subunit P35–deficient (IL-35P35–deficient) mice on an ApoE–/– background and found Treg reduction in the spleen and aorta compared with ApoE–/– controls. In addition, our RNA sequencing data show the elevation of a set of chemokine receptor transcripts in the ApoE–/– Tregs, and we have validated higher CCR5 expression in ApoE–/– Tregs in the presence of IL-35 than in the absence of IL-35. Furthermore, we observed that CCR5+ Tregs in ApoE–/– have lower Treg-weakening AKT-mTOR signaling, higher expression of inhibitory checkpoint receptors TIGIT and PD-1, and higher expression of IL-10 compared with WT CCR5+ Tregs. In conclusion, IL-35 counteracts hyperlipidemia in maintaining Treg-suppressive function by increasing 3 CCR5-amplified mechanisms, including Treg migration, inhibition of Treg weakening AKT-mTOR signaling, and promotion of TIGIT and PD-1 signaling.

CD4+ regulatory T cells (Tregs) respond to environmental cues to permit or suppress inflammation, and atherosclerosis weakens Treg suppression and promotes plasticity. However, the effects of smoking plus morphine (SM + M) on Treg plasticity remain unknown. To determine whether SM + M promotes Treg plasticity to T helper 17 (Th17) cells, we analyzed the RNA sequencing data from SM, M, and SM + M treated Tregs and performed knowledge-based and IPA analysis. We demonstrated that (1) SM + M, M, and SM upregulated the transcripts of cytokines, chemokines, and clusters of differentiation (CDs) and modulated the transcripts of kinases and phosphatases in Tregs; (2) SM + M, M, and SM upregulated the transcripts of immunometabolism genes, trained immunity genes, and histone modification enzymes; (3) SM + M increased the transcripts of Th17 transcription factor (TF) RORC and Tfh factor CXCR5 in Tregs; M increased the transcripts of T helper cell 1 (Th1) TF RUNX3 and Th1-Th9 receptor CXCR3; and SM inhibited Treg TGIF1 transcript; (4) six genes upregulated in SM + M Tregs were matched with the top-ranked Th17 pathogenic genes; and 57, 39 genes upregulated in SM + M Tregs were matched with groups II and group III Th17 pathogenic genes, respectively; (5) SM + M upregulated the transcripts of 70 IPA-TFs, 11 iTregs-specific TFs, and 4 iTregs-Th17 shared TFs; and (6) SM + M, M, and SM downregulated Treg suppression TF Rel (c-Rel); and 35 SM + M downregulated genes were overlapped with Rel−/− Treg downregulated genes. These results provide novel insights on the roles of SM + M in reprogramming Treg transcriptomes and Treg plasticity to Th17 cells and novel targets for future therapeutic interventions involving immunosuppression in atherosclerotic cardiovascular diseases, autoimmune diseases, transplantation, and cancers.

To examine whether the expressions of 260 organelle crosstalk regulators (OCRGs) in 16 functional groups are modulated in 23 diseases and 28 tumors, we performed extensive -omics data mining analyses and made a set of significant findings: (1) the ratios of upregulated vs. downregulated OCRGs are 1:2.8 in acute inflammations, 1:1 in metabolic diseases, 1:1.2 in autoimmune diseases, and 1:3.8 in organ failures; (2) sepsis and trauma-upregulated OCRG groups such as vesicle, mitochondrial (MT) fission, and mitophagy but not others, are termed as the cell crisis-handling OCRGs. Similarly, sepsis and trauma plus organ failures upregulated seven OCRG groups including vesicle, MT fission, mitophagy, sarcoplasmic reticulum–MT, MT fusion, autophagosome–lysosome fusion, and autophagosome/endosome–lysosome fusion, classified as the cell failure-handling OCRGs; (3) suppression of autophagosome–lysosome fusion in endothelial and epithelial cells is required for viral replications, which classify this decreased group as the viral replication-suppressed OCRGs; (4) pro-atherogenic damage-associated molecular patterns (DAMPs) such as oxidized low-density lipoprotein (oxLDL), lipopolysaccharide (LPS), oxidized-1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (oxPAPC), and interferons (IFNs) totally upregulated 33 OCRGs in endothelial cells (ECs) including vesicle, MT fission, mitophagy, MT fusion, endoplasmic reticulum (ER)–MT contact, ER– plasma membrane (PM) junction, autophagosome/endosome–lysosome fusion, sarcoplasmic reticulum–MT, autophagosome–endosome/lysosome fusion, and ER–Golgi complex (GC) interaction as the 10 EC-activation/inflammation-promoting OCRG groups; (5) the expression of OCRGs is upregulated more than downregulated in regulatory T cells (Tregs) from the lymph nodes, spleen, peripheral blood, intestine, and brown adipose tissue in comparison with that of CD4+CD25− T effector controls; (6) toll-like receptors (TLRs), reactive oxygen species (ROS) regulator nuclear factor erythroid 2-related factor 2 (Nrf2), and inflammasome-activated regulator caspase-1 regulated the expressions of OCRGs in diseases, virus-infected cells, and pro-atherogenic DAMP-treated ECs; (7) OCRG expressions are significantly modulated in all the 28 cancer datasets, and the upregulated OCRGs are correlated with tumor immune infiltrates in some tumors; (8) tumor promoter factor IKK2 and tumor suppressor Tp53 significantly modulate the expressions of OCRGs. Our findings provide novel insights on the roles of upregulated OCRGs in the pathogenesis of inflammatory diseases and cancers, and novel pathways for the future therapeutic interventions for inflammations, sepsis, trauma, organ failures, autoimmune diseases, metabolic cardiovascular diseases (CVDs), and cancers.

To characterize transcriptomic changes in endothelial cells (ECs) infected by coronaviruses, and stimulated by DAMPs, the expressions of 1311 innate immune regulatomic genes (IGs) were examined in 28 EC microarray datasets with 7 monocyte datasets as controls. We made the following findings: The majority of IGs are upregulated in the first 12 hours post-infection (PI), and maintained until 48 hours PI in human microvascular EC infected by middle east respiratory syndrome-coronavirus (MERS-CoV) (an EC model for COVID-19). The expressions of IGs are modulated in 21 human EC transcriptomic datasets by various PAMPs/DAMPs, including LPS, LPC, shear stress, hyperlipidemia and oxLDL. Upregulation of many IGs such as nucleic acid sensors are shared between ECs infected by MERS-CoV and those stimulated by PAMPs and DAMPs. Human heart EC and mouse aortic EC express all four types of coronavirus receptors such as ANPEP, CEACAM1, ACE2, DPP4 and virus entry facilitator TMPRSS2 (heart EC); most of coronavirus replication-transcription protein complexes are expressed in HMEC, which contribute to viremia, thromboembolism, and cardiovascular comorbidities of COVID-19. ECs have novel trained immunity (TI), in which subsequent inflammation is enhanced. Upregulated proinflammatory cytokines such as TNFα, IL6, CSF1 and CSF3 and TI marker IL-32 as well as TI metabolic enzymes and epigenetic enzymes indicate TI function in HMEC infected by MERS-CoV, which may drive cytokine storms. Upregulated CSF1 and CSF3 demonstrate a novel function of ECs in promoting myelopoiesis. Mechanistically, the ER stress and ROS, together with decreased mitochondrial OXPHOS complexes, facilitate a proinflammatory response and TI. Additionally, an increase of the regulators of mitotic catastrophe cell death, apoptosis, ferroptosis, inflammasomes-driven pyroptosis in ECs infected with MERS-CoV and the upregulation of pro-thrombogenic factors increase thromboembolism potential. Finally, NRF2-suppressed ROS regulate innate immune responses, TI, thrombosis, EC inflammation and death. These transcriptomic results provide novel insights on the roles of ECs in coronavirus infections such as COVID-19, cardiovascular diseases (CVD), inflammation, transplantation, autoimmune disease and cancers.

We performed a transcriptomic analyses using the strategies we pioneered and made the following findings: 1) Normal lymphoid Tregs, diseased kidney Tregs, splenic Tregs from mice with injured muscle have 3, 17 and 3 specific (S-) pathways, respectively; 2) Tumor splenic Tregs share 12 pathways with tumor Tregs; tumor splenic Tregs and tumor Tregs have 11 and 8 S-pathways, respectively; 3) Normal and non-tumor disease Tregs upregulate some of novel 2641 canonical secretomic genes (SGs) with 24 pathways, and tumor Tregs upregulate canonical secretomes with 17 pathways; 4) Normal and non-tumor disease tissue Tregs upregulate some of novel 6560 exosome SGs with 56 exosome SG pathways (ESP), tumor Treg ESP are more focused than other Tregs; 5) Normal, non-tumor diseased Treg and tumor Tregs upregulate some of novel 961 innate immune caspase-1 SGs and 1223 innate immune caspase-4 SGs to fulfill their tissue/SG-specific and shared functions; 6) Most tissue Treg transcriptomes are controlled by Foxp3; and Tumor Tregs had increased Foxp3 non-collaboration genes with ROS and 17 other pathways; 7) Immune checkpoint receptor PD-1 does, but CTLA-4 does not, play significant roles in promoting Treg upregulated genes in normal and non-tumor disease tissue Tregs; and tumor splenic and tumor Tregs have certain CTLA-4-, and PD-1-, non-collaboration transcriptomic changes with innate immune dominant pathways; 8) Tumor Tregs downregulate more immunometabolic and innate immune memory (trained immunity) genes than Tregs from other groups; and 11) ROS significantly regulate Treg transcriptomes; and ROS-suppressed genes are downregulated more in tumor Tregs than Tregs from other groups. Our results have provided novel insights on the roles of Tregs in normal, injuries, regeneration, tumor conditions and some of canonical and innate immune non-canonical secretomes via ROS-regulatory mechanisms and new therapeutic targets for immunosuppression, tissue repair, cardiovascular diseases, chronic kidney disease, autoimmune diseases, transplantation, and cancers.

We used functional -omics angles and examined transcriptomic heterogeneity in CD4+Foxp3+ regulatory T cells (Treg) from spleen (s-Treg), lymph nodes (LN-Treg), intestine (int-Treg), and visceral adipose tissue (VAT-Treg), and made significant findings: 1) Five new shared Treg genes including NIBAN, TNFRSF1b, DUSP4,VAV2, and KLRG1, and 68 new signatures are identified. Among 27 signaling pathways shared in four tissue Treg, 22 pathways are innate immune pathways (81.5%); 2) s-Treg, LN-Treg, int-Treg, and VAT-Treg have zero, 49, 45, and 116 upregulated pathways, respectively; 3) 12, 7, and 15 out of 373 CD markers are identified as specific for LN-Treg, int-Treg, and VAT-Treg, respectively, which may initiate innate immune signaling; 4) 7, 49, 44, and 79 increased cytokines out of 1176 cytokines are identified for four Treg, respectively, suggesting that Treg have much more secretory proteins/cytokines than IL-10, TGF-β, and IL-35; 5) LN-Treg, int-Treg, and VAT-Treg have 13 additional secretory functions more than s-Treg, found by analyzing 1,706 secretomic genes; 6) 2, 20, 25, and 43 increased transcription factors (TFs) out of 1,496 TFs are identified four Treg, respectively; 7) LN-Treg and int-Treg have increased pyroptosis regulators but VAT-Treg have increased apoptosis regulators; 8) 1, 15, 19, and 31 increased kinases out of 661 kinome are identified for s-Treg, LN-Treg, int-Treg, and VAT-Treg, respectively; 9) comparing with that of s-Treg, LN-Treg, int-Treg, and VAT-Treg increase activated cluster (clusters 1–3) markers; and decrease resting cluster (clusters 4–6) markers; and 10) Treg promote tissue repair by sharing secretomes and TFs AHR, ETV5, EGR1, and KLF4 with stem cells, which partially promote upregulation of all the groups of Treg genes. These results suggest that stem cell-shared master genes make tissue Treg as the first T cell type using a Treg niche to maintain their Treg-ness with 80% innate immune pathways, and triple functions of immunosuppression, tissue repair, and homeostasis maintenance. Our results have provided novel insights on the roles of innate immune pathways on Treg heterogeneity and new therapeutic targets for immunosuppression, tissue repair, cardiovascular diseases, chronic kidney disease, autoimmune diseases, transplantation, and cancers.

Metabolically healthy obesity (MHO) accounts for roughly 35% of all obese patients. There is no clear consensus that has been reached on whether MHO is a stable condition or merely a transitory period between metabolically healthy lean and metabolically unhealthy obesity (MUO). Additionally, the mechanisms underlying MHO and any transition to MUO are not clear. Macrophages are the most common immune cells in adipose tissues and have a significant presence in atherosclerosis. Fas (or CD95), which is highly expressed on macrophages, is classically recognized as a pro-apoptotic cell surface receptor. However, Fas also plays a significant role as a pro-inflammatory molecule. Previously, we established a mouse model (ApoE-/-/miR155-/-; DKO mouse) of MHO, based on the criteria of not having metabolic syndrome (MetS) and insulin resistance (IR). In our current study, we hypothesized that MHO is a transition phase toward MUO, and that inflammation driven by our newly classified CD95+CD86- macrophages is a novel mechanism for this transition. We found that, with extended (24 weeks) high-fat diet feeding (HFD), MHO mice became MUO, shown by increased atherosclerosis. Mechanistically, we found the following: 1) at the MHO stage, DKO mice exhibited increased pro-inflammatory markers in adipose tissue, including CD95, and serum; 2) total adipose tissue macrophages (ATMs) increased; 3) CD95+CD86- subset of ATMs also increased; and 4) human aortic endothelial cells (HAECs) were activated (as determined by upregulated ICAM1 expression) when incubated with conditioned media from CD95+-containing DKO ATMs and human peripheral blood mononuclear cells-derived macrophages in comparison to respective controls. These results suggest that extended HFD in MHO mice promotes vascular inflammation and atherosclerosis via increasing CD95+ pro-inflammatory ATMs. In conclusion, we have identified a novel molecular mechanism underlying MHO transition to MUO with HFD. We have also found a previously unappreciated role of CD95+ macrophages as a potentially novel subset that may be utilized to assess pro-inflammatory characteristics of macrophages, specifically in adipose tissue in the absence of pro-inflammatory miR-155. These findings have provided novel insights on MHO transition to MUO and new therapeutic targets for the future treatment of MUO, MetS, other obese diseases, and type II diabetes.

Background. The immune mechanisms underlying low-intensity ultrasound- (LIUS-) mediated suppression of inflammation and tumorigenesis remain poorly determined. Methods. We used microarray datasets from the NCBI GEO DataSet repository and conducted comprehensive data-mining analyses, where we examined the gene expression of 1376 innate immune regulators (innatome genes (IGs) in cells treated with LIUS. Results. We made the following findings: (1) LIUS upregulates proinflammatory IGs and downregulates metastasis genes in cancer cells, and LIUS upregulates adaptive immunity pathways but inhibits danger-sensing and inflammation pathways and promote tolerogenic differentiation in bone marrow (BM) cells. (2) LIUS upregulates IGs encoded for proteins localized in the cytoplasm, extracellular space, and others, but downregulates IG proteins localized in nuclear and plasma membranes, and LIUS downregulates phosphatases. (3) LIUS-modulated IGs act partially via several important pathways of reactive oxygen species (ROS), reverse signaling of immune checkpoint receptors B7-H4 and BTNL2, inflammatory cytokines, and static or oscillatory shear stress and heat generation, among which ROS is a dominant mechanism. (4) LIUS upregulates trained immunity enzymes in lymphoma cells and downregulates trained immunity enzymes and presumably establishes trained tolerance in BM cells. (5) LIUS modulates chromatin long-range interactions to differentially regulate IGs expression in cancer cells and noncancer cells. Conclusions. Our analysis suggests novel molecular mechanisms that are utilized by LIUS to induce tumor suppression and inflammation inhibition. Our findings may lead to development of new treatment protocols for cancers and chronic inflammation.

We performed a panoramic analysis on both human nonalcoholic steatohepatitis (NASH) microarray data and microarray/RNA-seq data from various mouse models of nonalcoholic fatty liver disease NASH/NAFLD with total 4249 genes examined and made the following findings: (i) human NASH and NAFLD mouse models upregulate both cytokines and chemokines; (ii) pathway analysis indicated that human NASH can be classified into metabolic and immune NASH; methionine- and choline-deficient (MCD)+high-fat diet (HFD), glycine N-methyltransferase deficient (GNMT-KO), methionine adenosyltransferase 1A deficient (MAT1A-KO), and HFCD (high-fat-cholesterol diet) can be classified into inflammatory, SAM accumulation, cholesterol/mevalonate, and LXR/RXR-fatty acid β-oxidation NAFLD, respectively; (iii) canonical and noncanonical inflammasomes play differential roles in the pathogenesis of NASH/NAFLD; (iv) trained immunity (TI) enzymes are significantly upregulated in NASH/NAFLD; HFCD upregulates TI enzymes more than cytokines, chemokines, and inflammasome regulators; (v) the MCD+HFD is a model with the upregulation of proinflammatory cytokines and canonical and noncanonical inflammasomes; however, the HFCD is a model with upregulation of TI enzymes and lipid peroxidation enzymes; and (vi) caspase-11 and caspase-1 act as upstream master regulators, which partially upregulate the expressions of cytokines, chemokines, canonical and noncanonical inflammasome pathway regulators, TI enzymes, and lipid peroxidation enzymes. Our findings provide novel insights on the synergies between hyperlipidemia and hypomethylation in establishing TI and promoting inflammation in NASH and NAFLD progression and novel targets for future therapeutic interventions for NASH and NAFLD, metabolic diseases, transplantation, and cancers.

Reactive oxygen species (ROS) are critical for the progression of cardiovascular diseases, inflammations and tumors. However, the mechanisms of how ROS sense metabolic stress, regulate metabolic pathways and initiate proliferation, inflammation and cell death responses remain poorly characterized. In this analytic review, we concluded that: 1) Based on different features and functions, eleven types of ROS can be classified into seven functional groups: metabolic stress-sensing, chemical connecting, organelle communication, stress branch-out, inflammasome-activating, dual functions and triple functions ROS. 2) Among the ROS generation systems, mitochondria consume the most amount of oxygen; and nine types of ROS are generated; thus, mitochondrial ROS systems serve as the central hub for connecting ROS with inflammasome activation, trained immunity and immunometabolic pathways. 3) Increased nuclear ROS production significantly promotes cell death in comparison to that in other organelles. Nuclear ROS systems serve as a convergent hub and decision-makers to connect unbearable and alarming metabolic stresses to inflammation and cell death. 4) Balanced ROS levels indicate physiological homeostasis of various metabolic processes in subcellular organelles and cytosol, while imbalanced ROS levels present alarms for pathological organelle stresses in metabolic processes. Based on these analyses, we propose a working model that ROS systems are a new integrated network for sensing homeostasis and alarming stress in metabolic processes in various subcellular organelles. Our model provides novel insights on the roles of the ROS systems in bridging metabolic stress to inflammation, cell death and tumorigenesis; and provide novel therapeutic targets for treating those diseases. (Word count: 246).

Interleukin (IL) 35 is a novel immunosuppressive heterodimeric cytokine in IL-12 family. Whether and how IL-35 regulates ischemia-induced angiogenesis in peripheral artery diseases are unrevealed. To fill this important knowledge gap, we used loss-of-function, gain-of-function, omics data analysis, RNA-Seq, in vivo and in vitro experiments, and we have made the following significant findings: i) IL-35 and its receptor subunit IL-12RB2, but not IL-6ST, are induced in the muscle after hindlimb ischemia (HLI); ii) HLI-induced angiogenesis is improved in Il12rb2−/− mice, in ApoE−/−/Il12rb2−/− mice compared to WT and ApoE−/− controls, respectively, where hyperlipidemia inhibits angiogenesis in vivo and in vitro; iii) IL-35 cytokine injection as a gain-of-function approach delays blood perfusion recovery at day 14 after HLI; iv) IL-35 spares regenerative angiogenesis at the late phase of HLI recovery after day 14 of HLI; v) Transcriptome analysis of endothelial cells (ECs) at 14 days post-HLI reveals a disturbed extracellular matrix re-organization in IL-35-injected mice; vi) IL-35 downregulates three reactive oxygen species (ROS) promoters and upregulates one ROS attenuator, which may functionally mediate IL-35 upregulation of anti-angiogenic extracellular matrix proteins in ECs; and vii) IL-35 inhibits human microvascular EC migration and tube formation in vitro mainly through upregulating anti-angiogenic extracellular matrix-remodeling proteins. These findings provide a novel insight on the future therapeutic potential of IL-35 in suppressing ischemia/inflammation-triggered inflammatory angiogenesis at early phase but sparing regenerative angiogenesis at late phase.

The mechanisms that underlie various inflammation paradoxes, metabolically healthy obesity, and increased inflammations after inflammatory cytokine blockades and deficiencies remain poorly determined. We performed an extensive –omics database mining, determined the expressions of 1367 innate immune regulators in 18 microarrays after deficiencies of 15 proinflammatory cytokines/regulators and eight microarray datasets of patients receiving Mab therapies, and made a set of significant findings: 1) proinflammatory cytokines/regulators suppress the expressions of innate immune regulators; 2) upregulations of innate immune regulators in the deficiencies of IFNγ/IFNγR1, IL-17A, STAT3 and miR155 are more than that after deficiencies of TNFα, IL-1β, IL-6, IL-18, STAT1, NF-kB, and miR221; 3) IFNγ, IFNγR and IL-17RA inhibit 10, 59 and 39 proinflammatory cytokine/regulator pathways, respectively; in contrast, TNFα, IL-6 and IL-18 each inhibits only four to five pathways; 4) The IFNγ-promoted and -suppressed innate immune regulators have four shared pathways; the IFNγR1-promoted and -suppressed innate immune regulators have 11 shared pathways; and the miR155-promoted and -suppressed innate immune regulators have 13 shared pathways, suggesting negative-feedback mechanisms in their conserved regulatory pathways for innate immune regulators; 5) Deficiencies of proinflammatory cytokine/regulator-suppressed, promoted programs share signaling pathways and increase the likelihood of developing 11 diseases including cardiovascular disease; 6) There are the shared innate immune regulators and pathways between deficiency of TNFα in mice and anti-TNF therapy in clinical patients; 7) Mechanistically, up-regulated reactive oxygen species regulators such as myeloperoxidase caused by suppression of proinflammatory cytokines/regulators can drive the upregulation of suppressed innate immune regulators. Our findings have provided novel insights on various inflammation paradoxes and proinflammatory cytokines regulation of innate immune regulators; and may re-shape new therapeutic strategies for cardiovascular disease and other inflammatory diseases.

Background: The molecular mechanisms underlying chronic kidney disease (CKD) transition to end-stage renal disease (ESRD) and CKD acceleration of cardiovascular and other tissue inflammations remain poorly determined. Methods: We conducted a comprehensive data analyses on 7 microarray datasets in peripheral blood mononuclear cells (PBMCs) from patients with CKD and ESRD from NCBI-GEO databases, where we examined the expressions of 2641 secretome genes (SG). Results: 1) 86.7% middle class (molecular weight >500 Daltons) uremic toxins (UTs) were encoded by SGs; 2) Upregulation of SGs in PBMCs in patients with ESRD (121 SGs) were significantly higher than that of CKD (44 SGs); 3) Transcriptomic analyses of PBMC secretome had advantages to identify more comprehensive secretome than conventional secretomic analyses; 4) ESRD-induced SGs had strong proinflammatory pathways; 5) Proinflammatory cytokines-based UTs such as IL-1β and IL-18 promoted ESRD modulation of SGs; 6) ESRD-upregulated co-stimulation receptors CD48 and CD58 increased secretomic upregulation in the PBMCs, which were magnified enormously in tissues; 7) M1-, and M2-macrophage polarization signals contributed to ESRD- and CKD-upregulated SGs; 8) ESRD- and CKD-upregulated SGs contained senescence-promoting regulators by upregulating proinflammatory IGFBP7 and downregulating anti-inflammatory TGF-β1 and telomere stabilizer SERPINE1/PAI-1; 9) ROS pathways played bigger roles in mediating ESRD-upregulated SGs (11.6%) than that in CKD-upregulated SGs (6.8%), and half of ESRD-upregulated SGs were ROS-independent. Conclusions: Our analysis suggests novel secretomic upregulation in PBMCs of patients with CKD and ESRD, act synergistically with uremic toxins, to promote inflammation and potential disease progression. Our findings have provided novel insights on PBMC secretome upregulation to promote disease progression and may lead to the identification of new therapeutic targets for novel regimens for CKD, ESRD and their accelerated cardiovascular disease, other inflammations and cancers. (Total words: 279).

The mechanisms underlying pathophysiological regulation of tissue macrophage (Mφ) subsets remain poorly understood. From the expression of 207 Mφ genes comprising 31 markers for 10 subsets, 45 transcription factors (TFs), 56 immunometabolism enzymes, 23 trained immunity (innate immune memory) enzymes, and 52 other genes in microarray data, we made the following findings. (1) When 34 inflammation diseases and tumor types were grouped into eight categories, there was differential expression of the 31 Mφ markers and 45 Mφ TFs, highlighted by 12 shared and 20 group-specific disease pathways. (2) Mφ in lung, liver, spleen, and intestine (LLSI-Mφ) express higher M1 Mφ markers than lean adipose tissue Mφ (ATMφ) physiologically. (3) Pro-adipogenic TFs C/EBPα and PPARγ and proinflammatory adipokine leptin upregulate the expression of M1 Mφ markers. (4) Among 10 immune checkpoint receptors (ICRs), LLSI-Mφ and bone marrow (BM) Mφ express higher levels of CD274 (PDL-1) than ATMφ, presumably to counteract the M1 dominant status via its reverse signaling behavior. (5) Among 24 intercellular communication exosome mediators, LLSI- and BM- Mφ prefer to use RAB27A and STX3 than RAB31 and YKT6, suggesting new inflammatory exosome mediators for propagating inflammation. (6) Mφ in peritoneal tissue and LLSI-Mφ upregulate higher levels of immunometabolism enzymes than does ATMφ. (7) Mφ from peritoneum and LLSI-Mφ upregulate more trained immunity enzyme genes than does ATMφ. Our results suggest that multiple new mechanisms including the cell surface, intracellular immunometabolism, trained immunity, and TFs may be responsible for disease group-specific and shared pathways. Our findings have provided novel insights on the pathophysiological regulation of tissue Mφ, the disease group-specific and shared pathways of Mφ, and novel therapeutic targets for cancers and inflammations.

Abstract Background ETS2 is a downstream effector of the RAS/RAF/ERK pathway, which plays a critical role in the development of malignant tumor. However, the clinical impact of ETS2 expression in AML remains unknown. Methods In this study, we evaluated the prognostic significance of ETS2 expression using two relatively large cohorts of AML patients. Results In the first cohort, compared to low expression of ETS2 (ETS2 low), high expression of ETS2 (ETS2 high) showed significant shorter OS, EFS and RFS in the current treatments including the allogeneic HCT group (n = 72) and the chemotherapy group (n = 100). Notably, among ETS2 high patients, those received allogeneic HCT had longer OS, EFS and RFS than those with chemotherapy alone (allogeneic HCT, n = 39 vs. chemotherapy, n = 47), but treatment modules play insignificant role in the survival of ETS2 low patients (allogeneic HCT, n = 33 vs. chemotherapy, n = 53). Moreover, gene/microRNA expression data provides insights into the biological changes associated with varying ETS2 expression levels in AML. The prognostic value of ETS2 was further validated in the second AML cohort (n = 329). Conclusions Our results indicate that ETS2 high is a poor prognostic factor in AML and may guide treatment decisions towards allogeneic HCT.

Ischemia reperfusion injury (IRI) during liver transplantation increases morbidity and contributes to allograft dysfunction. There are no therapeutic strategies to mitigate IRI. We examined a novel hypothesis: caspase 1 and caspase 11 serve as danger-associated molecular pattern (DAMPs) sensors in IRI. By performing microarray analysis and using caspase 1/caspase 11 double-knockout (Casp DKO) mice, we show that the canonical and non-canonical inflammasome regulators are upregulated in mouse liver IRI. Ischemic pre (IPC)- and post-conditioning (IPO) induce upregulation of the canonical and non-canonical inflammasome regulators. Trained immunity (TI) regulators are upregulated in IPC and IPO. Furthermore, caspase 1 is activated during liver IRI, and Casp DKO attenuates liver IRI. Casp DKO maintained normal liver histology via decreased DNA damage. Finally, the decreased TUNEL assay-detected DNA damage is the underlying histopathological and molecular mechanisms of attenuated liver pyroptosis and IRI. In summary, liver IRI induces the upregulation of canonical and non-canonical inflammasomes and TI enzyme pathways. Casp DKO attenuate liver IRI. Development of novel therapeutics targeting caspase 1/caspase 11 and TI may help mitigate injury secondary to IRI. Our findings have provided novel insights on the roles of caspase 1, caspase 11, and inflammasome in sensing IRI derived DAMPs and TI-promoted IRI-induced liver injury.

We conducted an experimental database analysis to determine the expression of 61 CD4+ Th subset regulators in human and murine tissues, cells, and in T-regulatory cells (Treg) in physiological and pathological conditions. We made the following significant findings: (1) adipose tissues of diabetic patients with insulin resistance upregulated various Th effector subset regulators; (2) in skin biopsy from patients with psoriasis, and in blood cells from patients with lupus, effector Th subset regulators were more upregulated than downregulated; (3) in rosiglitazone induced failing hearts in ApoE-deficient (KO) mice, various Th subset regulators were upregulated rather than downregulated; (4) aortic endothelial cells activated by proatherogenic stimuli secrete several Th subset-promoting cytokines; (5) in Treg from follicular Th (Tfh)-transcription factor (TF) Bcl6 KO mice, various Th subset regulators were upregulated; whereas in Treg from Th2-TF GATA3 KO mice and HDAC6 KO mice, various Th subset regulators were downregulated, suggesting that Bcl6 inhibits, GATA3 and HDAC6 promote, Treg plasticity; and (6) GATA3 KO, and Bcl6 KO Treg upregulated MHC II molecules and T cell co-stimulation receptors, suggesting that GATA3 and BCL6 inhibit Treg from becoming novel APC-Treg. Our data implies that while HDAC6 and Bcl6 are important regulators of Treg plasticity, GATA3 determine the fate of plastic Tregby controlling whether it will convert in to either Th1-Treg or APC-T-reg. Our results have provided novel insights on Treg plasticity into APC-Treg and Th1-Treg, and new therapeutic targets in metabolic diseases, autoimmune diseases, and inflammatory disorders.

Introduction: Non-alcoholic fatty liver disease (NAFLD) has a global prevalence of 25% of the population and is a leading cause of cirrhosis and hepatocellular carcinoma. NAFLD ranges from simple steatosis (non-alcoholic fatty liver) to non-alcoholic steatohepatitis (NASH). Hepatic macrophages, specifically Kupffer cells (KCs) and monocyte-derived macrophages, act as key players in the progression of NAFLD. Caspases are a family of endoproteases that provide critical connections to cell regulatory networks that sense disease risk factors, control inflammation, and mediate inflammatory cell death (pyroptosis). Caspase-11 can cleave gasdermin D (GSDMD) to induce pyroptosis and specifically defends against bacterial pathogens that invade the cytosol. However, it’s still unknown whether high fat diet (HFD)-facilitated gut microbiota-generated cytoplasmic lipopolysaccharides (LPS) activate caspase-11 and promote NAFLD. Methods: To examine this hypothesis, we performed liver pathological analysis, RNA-seq, FACS, Western blots, Seahorse mitochondrial stress analyses of macrophages and bone marrow transplantation on HFD-induced NAFLD in WT and Casp11–/– mice. Results and Discussion: Our results showed that 1) HFD increases body wight, liver wight, plasma cholesterol levels, liver fat deposition, and NAFLD activity score (NAS score) in wild-type (WT) mice; 2) HFD increases the expression of caspase-11, GSDMD, interleukin-1b, and guanylate-binding proteins in WT mice; 3) Caspase-11 deficiency decreases fat liver deposition and NAS score; 4) Caspase-11 deficiency decreases bone marrow monocyte-derived macrophage (MDM) pyroptosis (inflammatory cell death) and inflammatory monocyte (IM) surface GSDMD expression; 5) Caspase-11 deficiency re-programs liver transcriptomes and reduces HFD-induced NAFLD; 6) Caspase-11 deficiency decreases extracellular acidification rates (glycolysis) and oxidative phosphorylation (OXPHOS) in inflammatory fatty acid palmitic acid-stimulated macrophages, indicating that caspase-11 significantly contributes to maintain dual fuel bioenergetics— glycolysis and OXPHOS for promoting pyroptosis in macrophages. These results provide novel insights on the roles of the caspase-11-GSDMD pathway in promoting hepatic macrophage inflammation and pyroptosis and novel targets for future therapeutic interventions involving the transition of NAFLD to NASH, hyperlipidemia, type II diabetes, metabolic syndrome, metabolically healthy obesity, atherosclerotic cardiovascular diseases, autoimmune diseases, liver transplantation, and hepatic cancers. [ABSTRACT FROM AUTHOR]