CETP inhibition enhances monocyte activation and bacterial clearance and reduces streptococcus pneumonia-associated mortality in mice.

Sepsis is a leading cause of mortality worldwide, and pneumonia is the most common cause of sepsis in humans. Low levels of high-density lipoprotein cholesterol (HDL-C) levels are associated with an increased risk of death from sepsis, and increasing levels of HDL-C by inhibition of cholesteryl ester transfer protein (CETP) decreases mortality from intraabdominal polymicrobial sepsis in APOE*3-Leiden.CETP mice. Here, we show that treatment with the CETP inhibitor (CETPi) anacetrapib reduced mortality from Streptococcus pneumoniae-induced sepsis in APOE*3-Leiden.CETP and APOA1.CETP mice. Mechanistically, CETP inhibition reduced the host proinflammatory response via attenuation of proinflammatory cytokine transcription and release. This effect was dependent on the presence of HDL, leading to attenuation of immune-mediated organ damage. In addition, CETP inhibition promoted monocyte activation in the blood prior to the onset of sepsis, resulting in accelerated macrophage recruitment to the lung and liver. In vitro experiments demonstrated that CETP inhibition significantly promoted the activation of proinflammatory signaling in peripheral blood mononuclear cells and THP1 cells in the absence of HDL; this may represent a mechanism responsible for improved bacterial clearance during sepsis. These findings provide evidence that CETP inhibition represents a potential approach to reduce mortality from pneumosepsis.

Identification of hub genes and potential ceRNA networks of diabetic cardiomyopathy.

Diabetic cardiomyopathy (DCM), a common complication of diabetes, is defined as ventricular dysfunction in the absence of underlying heart disease. Noncoding RNAs (ncRNAs), including long noncoding RNAs (lncRNAs) and microRNAs (miRNAs), play a crucial role in the development of DCM. Weighted Gene Co-Expression Network Analysis (WGCNA) was used to identify key modules in DCM-related pathways. DCM-related miRNA-mRNA network and DCM-related ceRNA network were constructed by miRNA-seq to identify hub genes in these modules. We identified five hub genes that are associated with the onset of DCM, including Troponin C1 (Tnnc1), Phospholamban (Pln), Fatty acid binding proteins 3 (Fabp3), Popeye domain containing 2 (Popdc2), and Tripartite Motif-containing Protein 63 (Trim63). miRNAs that target the hub genes were mainly involved in TGF-ß and Wnt signaling pathways. GO BP enrichment analysis found these miRNAs were involved in the signaling of TGF-ß and glucose homeostasis. Q-PCR results found the gene expressions of Pln, Fabp3, Trim63, Tnnc1, and Popdc2 were significantly increased in DCM. Our study identified five hub genes (Tnnc1, Pln, Fabp3, Popdc2, Trim63) whose associated ceRNA networks are responsible for the onset of DCM.

MicroRNA-126 affects rheumatoid arthritis synovial fibroblast proliferation and apoptosis by targeting PIK3R2 and regulating PI3K-AKT signal pathway.

Rheumatoid arthritis (RA) is a chronic autoimmune disease that causes inflammation and destruction of the joints as well as an increased risk of cardiovascular disease. RA synovial fibroblasts (RASFs) are involved in the progression of RA and release pro-inflammatory cytokines. On the other hand, microRNAs (miRs) may help control the inflammatory response of immune and non-immune cells. Therefore, our study used lentiviral expression vectors to test the effects of miR-126 overexpression on RASF proliferation and apoptosis. Luciferase experiments verified the targeting relationship between miR-126 and PIK3R2 gene. The co-transfection of anti-miR-126 and PIK3R2 siRNA to RASFs were used to identify whether PIK3R2 was directly involved in proliferation and apoptosis of miR-126-induced RASFs. Real-time polymerase chain reaction (PCR) was used to detect miR-126 and PIK3R2 expressions. MTT assay was used to detect cell proliferation. Flow cytometry was used to detect cell apoptosis and cell cycle. Western blotting was used to detect PIK3R2, PI3K, AKT and p-AKT proteins. After Lv-miR-126 infected RASFs, the relative expression of miR-126 was significantly enhanced. MiR-126 promoted RASF proliferation and inhibited apoptosis. Levels of PIK3R2 decreased while total PI3K and p-AKT levels increased in RASFs overexpressing miR-126. Co-transfection of anti-miR-126 and PIK3R2 siRNA also increased PI3K and p-AKT levels as well as RASF proliferation and reduced apoptosis, as compared to anti-miR-126 treatment alone. Finally, luciferase reporter assays showed that miR-126 targeted PIK3R2. Our data indicate that miR-126 overexpression in RASFs inhibits PIK3R2 expression and promotes proliferation while inhibiting apoptosis. This suggests inhibiting miR-126 may yield therapeutic benefits in the treatment of RA.

Apoptosis in cardiac myocytes during the early stage after severe burn.

BACKGROUND: Cardiac dysfunction after severe burn is associated with postburn myocardial injury. We hypothesize that myocyte apoptosis is triggered and presented as the pathologic basis of postburn myocardial injury during the early stage after severe burn, and that apoptosis may be related to inflammatory responses in the postburn myocardium. METHODS: Rats with 40% total body surface area full-thickness burn were used. The following functions were measured at several time points after the burn injury: myocyte apoptosis (TUNEL staining, DNA ladder, and caspase-3 activity assay); mRNA levels of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (reverse transcriptase-polymerase chain reaction [RT-PCR]); activities of myeloperoxidase and p38 mitogen activated protein (MAP) kinase (Western blots); and left cardiac function. RESULTS: TUNEL positive myocytes appeared as early as 6-hour and their numbers showed further increases at 12-hour and 24-hour postburn; DNA fragmentation was clearly observed, and caspase-3 activity was significantly increased in the myocardium after burn. Infiltration of neutrophils, evidenced by the levels of myeloperoxidase activity, expression of TNF-alpha, and p38 MAP kinase activity in the heart, were all significantly increased within 24-hour after burn. Cardiac function was decreased after burn, which approximately paralleled the increased amount of cardiac apoptosis. CONCLUSION: These results demonstrate that cardiomyocyte apoptosis progressively develops during the early stage after severe burn, which may in part contribute to burn-induced cardiac dysfunction. Myocardial inflammatory responses, evidenced by the increased infiltration of neutrophils, as well as production of TNF-alpha probably because of the activation of p38 MAP kinase, may be involved in burn-induced cardiomyocyte apoptosis.

Involvement of p38 MAP kinase in burn-induced degradation of membrane phospholipids and upregulation of cPLA2 in cardiac myocytes.

This study was aimed to evaluate the role of p38 mitogen-activated protein (MAP) kinase in the degradation of membrane phospholipids and the regulation of cytosolic phospholipase A2 (cPLA2) in cardiac myocytes after burn trauma. In an in vivo study, rats were randomized into four groups: (1) sham-burn group, (2) burn group (40% total body surface area full-thickness burn), (3) burn + SB203580 group, and (4) burn + vehicle group. The rats from each group were killed at varying times after burn to examine the p38 MAP kinase activation (by means of Western blot analysis and immunohistochemical assay), the expression of cPLA2 (by means of reverse transcriptase polymerase chain reaction), the level of cardiac membrane phospholipids, and the level of the remaining creatine kinase-MB (CK-MB) isoenzyme in the heart. These studies showed that burn resulted in a significant decrease in the level of cardiac membrane phospholipids from 3 to 24 h after burn, which was paralleled with a persistent activation of p38 MAP kinase and an increased expression of cPLA2 in the heart. SB203580, a selective inhibitor of p38 MAP kinase, inhibited the activation of cardiac p38 MAP kinase, suppressed the burn-induced upregulation of cPLA2 and the increased PLA2 activity, and prevented burn-induced decrease in the levels of the cardiac membrane phospholipids and the remaining creatine kinase-MB isoenzyme. In addition, the in vitro treatment of cardiac myocytes with SB203580 also abolished the upregulation of cPLA2 and the disturbance of phospholipid homeostasis elicited by hypoxia and burn serum challenge. Taken together, these results have demonstrated for the first time that p38 MAP kinase is involved in burn-induced membrane phospholipids degradation in cardiac myocytes, at least in part through the regulation of cPLA2.

Calcium induced the damage of myocardial mitochondrial respiratory function in the early stage after severe burns.

OBJECTIVE: To explore the role of Ca(2+) in the damage to myocardial mitochondrial respiratory function in the early stage after severe burns. METHODS: An experimental model of 30%TBSA full-thickness skin scalding was reproduced in rats. Myocardial mitochondria were isolated from control and burned rats in the 1st, 3rd, 6th, 12th and 24th hour post-burn. The mitochondrial respiratory function, contents of mitochondrial calcium ([Ca(2+)](m)), activities of mtPLA(2), mtNOS, F(0)F(1)-ATPase and cytochrome c oxidase were determined. RESULTS: (1) At the 1st hour post-burn, [Ca(2+)](m) was increased significantly and the myocardial mitochondrial respiratory function was significantly reinforced. At the same time, mitochondrial respiratory control rate (RCR) was elevated and positively correlated with [Ca(2+)](m) (r=0.8415, P<0.01). At the 3rd, 6th, 12th and 24th hour post-burn, [Ca(2+)](m) increased further to a higher level, however, the mitochondrial respiratory function was decreased from the peak value at 6h, and RCR was negatively correlated with [Ca(2+)](m). (2) The activities of mtNOS and mtPLA(2) were higher significantly at the 3rd, 6th, 12th and 24th hour post-burn than that of the control. After severe burns, mtNOS and mtPLA(2) activities were both positively correlated with [Ca(2+)](m) (r=0.8945, P<0.05; r=0.9271, P<0.01, respectively). (3) The F(0)F(1)-ATPase synthetic activity increased at the 1st hour post-burn, but it decreased to 51.4, 44.9, 77.6 and 87.4% of that of the control at the 3rd, 6th, 12th and 24th hour post-burn respectively. The F(0)F(1)-ATPase hydrolytic activity decreased at the 1st hour post-burn and increased at the 3rd, however, it decreased again at the 6th, 12th and 24th hour post-burn. The activity of cytochrome c oxidase at the 3rd, 6th, 12th and 24th hour was low compared to the control. CONCLUSIONS: The changes of [Ca(2+)](m) were involved in damage to or regulation of mitochondrial respiratory function after severe burns. Appropriate increase of [Ca(2+)](m) reinforced the mitochondrial respiration at 1st hour after of burn injury, but Ca(2+) severe overload impairing F(0)F(1)-ATPase and cytochrome c oxidase directly, or, indirectly by activation of mtPLA(2) and mtNOS, might play an important role in damage to myocardial mitochondrial respiratory function at later stages after severe burns.