Salutary effects of androstenediol on cardiac function and splanchnic perfusion after trauma-hemorrhage

Recent studies have shown that dehydroepiandrosterone (DHEA) administration after trauma-hemorrhage (T-H) improves cardiovascular function and decreases cytokine production in male animals. Although androstenediol, one of the metabolites of DHEA, is reported to have estrogen-like activity, it remains unknown whether androstenediol per se has any salutary effects on cytokines and cardiovascular function after T-H. To examine this effect, male Sprague-Dawley rats underwent laparotomy and were bled to and maintained at a mean arterial blood pressure of 35-40 mmHg for approximately 90 min. The animals were resuscitated with four times the volume of maximal bleedout volume in the form of Ringer lactate. Androstenediol (1 mg/kg body wt i.v.) or vehicle was administered at the end of resuscitation. Twenty-four hours after resuscitation, cardiac function and organ blood flow were measured by using (85)Sr-microspheres. Circulating levels of nitrate/nitrite and IL-6 were also determined. Cardiovascular function and organ blood flow were significantly depressed after T-H. However, these parameters were restored by androstenediol treatment. The elevated plasma IL-6 levels after T-H were also lowered by androstenediol treatment. In contrast, plasma levels of nitrate/nitrite were the highest in the androstenediol-treated T-H animals. Because androstenediol administration after T-H decreases cytokine production and improves cardiovascular function, this agent appears to be a novel and useful adjunct for restoring the depressed cardiovascular function and for cytokine production in males after adverse circulatory conditions.     

Mechanism of salutary effects of estradiol on organ function after trauma-hemorrhage: upregulation of heme oxygenase

A growing body of evidence indicates that heme degradation products may counteract the deleterious consequences of hypoxia and/or ischemia-reperfusion injury. Because heme oxygenase (HO)-1 induction after adverse circulatory conditions is known to be protective, and because females in the proestrus cycle (with high estrogen) have better hepatic function and less hepatic damage than males after trauma-hemorrhage, we hypothesized that estrogen administration in males after trauma-hemorrhage will upregulate HO activity and protect the organs against dysfunction and injury. To test this hypothesis, male Sprague-Dawley rats underwent 5-cm laparotomy and hemorrhagic shock (35-40 mmHg for 93 +/- 2 min), followed by resuscitation with four times the shed blood volume in the form of Ringer lactate. 17beta-Estradiol and/or the specific HO enzyme inhibitor chromium mesoporphyrin (CrMP) were administered at the end of resuscitation, and the animals were killed 24 h thereafter. Trauma-hemorrhage reduced cardiac output, myocardial contractility, and serum albumin levels. Portal pressure and serum alanine aminotransferase levels were markedly increased under those conditions. These parameters were significantly improved in the 17beta-estradiol-treated rats. Estradiol treatment also induced increased HO-1 mRNA expression, HO-1 protein levels, and HO enzymatic activity in cardiac and hepatic tissue compared with vehicle-treated trauma-hemorrhage rats. Administration of the HO inhibitor CrMP prevented the estradiol-induced attenuation of shock-induced organ dysfunction and damage. Thus the salutary effects of estradiol administration on organ function after trauma-hemorrhage are mediated in part via upregulation of HO-1 expression and activity.     

Maintenance of lung myeloperoxidase activity in proestrus females after trauma-hemorrhage: upregulation of heme oxygenase-1

Previous studies showed that females in the proestrus stage of the reproductive cycle maintain organ functions after trauma-hemorrhage. However, it remains unknown whether the female reproductive cycle is an important variable in the regulation of lung injury after trauma-hemorrhage and, if so, whether the effect is mediated via upregulation of heme oxygenase (HO)-1. To examine this, female Sprague-Dawley rats during diestrus, proestrus, estrus, and metestrus phases of the reproductive cycle or 14 days after ovariectomy underwent soft tissue trauma and then hemorrhage (mean blood pressure 40 mmHg for 90 min followed by fluid resuscitation). At 2 h after trauma-hemorrhage or sham operation, lung myeloperoxidase (MPO) activity and intercellular adhesion molecule (ICAM)-1, cytokine-induced neutrophil chemoattractant (CINC)-1, CINC-3, and HO-1 protein levels were measured. Plasma 17beta-estradiol concentration was also determined. The results indicated that trauma-hemorrhage increased lung MPO activity and ICAM-1, CINC-1, and CINC-3 levels in ovariectomized females. These parameters were found to be similar to sham-operated animals in proestrus female rats subjected to trauma-hemorrhage. Lung HO-1 protein level in proestrus females was increased significantly compared with female rats subjected to trauma-hemorrhage during diestrus, estrus, and metestrus phases of the reproductive cycle and ovariectomized rats. Furthermore, plasma 17beta-estradiol level was highest in proestrus females. Administration of the HO inhibitor chromium mesoporphyrin prevented the attenuation of shock-induced lung damage in proestrus females. Thus these findings suggest that the female reproductive cycle is an important variable in the regulation of lung injury following trauma-hemorrhage and that the protective effect in proestrus females is likely mediated via upregulation of HO-1.     

Airway epithelial MyD88 restores control of Pseudomonas aeruginosa murine infection via an IL-1-dependent pathway

The opportunistic human pathogen Pseudomonas aeruginosa causes rapidly progressive and tissue-destructive infections, such as hospital-acquired and ventilator-associated pneumonias. Innate immune responses are critical in controlling P. aeruginosa in the mammalian lung, as demonstrated by the increased susceptibility of MyD88(-/-) mice to this pathogen. Experiments conducted using bone marrow chimeric mice demonstrated that radio-resistant cells participated in initiating MyD88-dependent innate immune responses to P. aeruginosa. In this study we used a novel transgenic mouse model to demonstrate that MyD88 expression by epithelial cells is sufficient to generate a rapid and protective innate immune response following intranasal infection with P. aeruginosa. MyD88 functions as an adaptor for many TLRs. However, mice in which multiple TLR pathways (e.g., TLR2/TLR4/TLR5) are blocked are not as compromised in their response to P. aeruginosa as mice lacking MyD88. We demonstrate that IL-1R signaling is an essential element of MyD88-dependent epithelial cell responses to P. aeruginosa infection.     

Cold-induced PGC-1alpha expression modulates muscle glucose uptake through an insulin receptor/Akt-independent, AMPK-dependent pathway

Peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) participates in control of expression of genes involved in adaptive thermogenesis, muscle fiber type differentiation, and fuel homeostasis. The objective of the present study was to evaluate the participation of cold-induced PGC-1alpha expression in muscle fiber type-specific activity of proteins that belong to the insulin-signaling pathway. Rats were exposed to 4 degrees C for 4 days and acutely treated with insulin in the presence or absence of an antisense oligonucleotide to PGC-1alpha. Cold exposure promoted a significant increase of PGC-1alpha and uncoupling protein-3 protein expression in type I and type II fibers of gastrocnemius muscle. In addition, cold exposure led to higher glucose uptake during a hyperinsulinemic clamp, which was accompanied by higher expression and membrane localization of GLUT4 in both muscle fiber types. Cold exposure promoted significantly lower insulin-induced tyrosine phosphorylation of the insulin receptor (IR) and Ser473 phosphorylation of acute transforming retrovirus thymoma (Akt) and an insulin-independent increase of Thr172 phosphorylation of adenosine 5'-monophosphate-activated protein kinase (AMPK). Inhibition of PGC-1alpha expression in cold-exposed rats by antisense oligonucleotide treatment diminished glucose clearance rates during a hyperinsulinemic clamp and reduced expression and membrane localization of GLUT4. Reduction of PGC-1alpha expression resulted in no modification of insulin-induced tyrosine phosphorylation of the IR and Ser473 phosphorylation of Akt. Finally, reduction of PGC-1alpha resulted in lower Thr172 phosphorylation of AMPK. Thus cold-induced hyperexpression of PGC-1alpha participates in control of skeletal muscle glucose uptake through a mechanism that controls GLUT4 expression and subcellular localization independent of the IR and Akt activities but dependent on AMPK.     

Estradiol improves cardiac and hepatic function after trauma-hemorrhage: role of enhanced heat shock protein expression

Although studies indicate that 17beta-estradiol administration after trauma-hemorrhage (T-H) improves cardiac and hepatic functions, the underlying mechanisms remain unclear. Because the induction of heat shock proteins (HSPs) can protect cardiac and hepatic functions, we hypothesized that these proteins contribute to the salutary effects of estradiol after T-H. To test this hypothesis, male Sprague-Dawley rats ( approximately 300 g) underwent laparotomy and hemorrhagic shock (35-40 mmHg for approximately 90 min) followed by resuscitation with four times the shed blood volume in the form of Ringer lactate. 17beta-estradiol (1 mg/kg body wt) was administered at the end of the resuscitation. Five hours after T-H and resuscitation there was a significant decrease in cardiac output, positive and negative maximal rate of left ventricular pressure. Liver function as determined by bile production and indocyanine green clearance was also compromised after T-H and resuscitation. This was accompanied by an increase in plasma alanine aminotransferase (ALT) levels and liver perfusate lactic dehydrogenase levels. Furthermore, circulating levels of TNF-alpha, IL-6, and IL-10 were also increased. In addition to decreased cardiac and hepatic function, there was an increase in cardiac HSP32 expression and a reduction in HSP60 expression after T-H. In the liver, HSP32 and HSP70 were increased after T-H. There was no change in heart HSP70 and liver HSP60 after T-H and resuscitation. Estradiol administration at the end of T-H and resuscitation increased heart/liver HSPs expression, ameliorated the impairment of heart/liver functions, and significantly prevented the increase in plasma levels of ALT, TNF-alpha, and IL-6. The ability of estradiol to induce HSPs expression in the heart and the liver suggests that HSPs, in part, mediate the salutary effects of 17beta-estradiol on organ functions after T-H.     

丙型肝炎病毒核心蛋白通过FOXO1/PGC-1α途径上调磷酸烯醇式丙酮酸羧基酶的转录

【目的】分析丙型肝炎病毒(HCV)核心蛋白(CORE)稳定表达对磷酸烯醇式丙酮酸羧基酶(PCK1)转录水平的影响,并分析HCV CORE调控PCK1转录的分子机制,为进一步阐明HCV感染致2型糖尿病机理的探讨提供新的思路。【方法】利用反转录病毒表达系统构建稳定表达HCV CORE的Huh7-lunet-core细胞系。采用Real-time PCR和萤光素酶报告基因技术检测Huh7-lunet-core细胞系中PCK1、FOXO1以及PGC-1α转录水平变化,并结合Western blot分析FOXO1的活性变化。【结果】HCV CORE的稳定表达显著增强PCK1的转录水平,HCV CORE不影响FOXO1的转录和表达水平,但降低FOXO1的磷酸化水平,激活了FOXO1的转录活性,并增强PGC-1α的mRNA表达水平。【结论】HCV CORE在Huh7-lunet细胞中的稳定表达激活FOXO1的转录活性,并与PGC-1α协同作用,上调PCK1的转录,从而导致肝糖异生过度发生,对HCV CORE调控PCK1转录的分子机制的揭示可能为HCV感染相关的糖尿病的治疗提供新的靶点。     

Pomalidomide restores immune recognition of primary effusion lymphoma through upregulation of ICAM-1 and B7-2

Pomalidomide (Pom) is an immunomodulatory drug that has efficacy against Kaposi’s sarcoma, a tumor caused by Kaposi’s sarcoma-associated herpesvirus (KSHV). Pom also induces direct cytotoxicity in primary effusion lymphoma (PEL), a B-cell malignancy caused by KSHV, in part through downregulation of IRF4, cMyc, and CK1α as a result of its interaction with cereblon, a cellular E3 ubiquitin ligase. Additionally, Pom can reverse KSHV-induced downregulation of MHCI and co-stimulatory immune surface molecules ICAM-1 and B7-2 on PELs. Here, we show for the first time that Pom-induced increases in ICAM-1 and B7-2 on PEL cells lead to an increase in both T-cell activation and NK-mediated cytotoxicity against PEL. The increase in T-cell activation can be prevented by blocking ICAM-1 and/or B7-2 on the PEL cell surface, suggesting that both ICAM-1 and B7-2 are important for T-cell co-stimulation by PELs. To gain mechanistic insights into Pom’s effects on surface markers, we generated Pom-resistant (PomR) PEL cells, which showed about 90% reduction in cereblon protein level and only minimal changes in IRF4 and cMyc upon Pom treatment. Pom no longer upregulated ICAM-1 and B7-2 on the surface of PomR cells, nor did it increase T-cell and NK-cell activation. Cereblon-knockout cells behaved similarly to the pomR cells upon Pom-treatment, suggesting that Pom’s interaction with cereblon is necessary for these effects. Further mechanistic studies revealed PI3K signaling pathway as being important for Pom-induced increases in these molecules. These observations provide a rationale for the study of Pom as therapy in treating PEL and other KSHV-associated tumors.     

MARCH5 restores endothelial cell function against ischaemic/hypoxia injury via Akt/eNOS pathway

MARCH5 is a critical regulator of mitochondrial dynamics, apoptosis and mitophagy. However, its role in cardiovascular system remains poorly understood. This study aimed to investigate the role of MARCH5 in endothelial cell (ECs) injury and the involvement of the Akt/eNOS signalling pathway in this process. Rat models of myocardial infarction (MI) and human cardiac microvascular endothelial cells (HCMECs) exposed to hypoxia (1% O₂) were used in this study. MARCH5 expression was significantly reduced in ECs of MI hearts and ECs exposed to hypoxia. Hypoxia inhibited the proliferation, migration and tube formation of ECs, and these effects were aggravated by knockdown of MARCH5 but antagonized by overexpressed MARCH5. Overexpression of MARCH5 increased nitric oxide (NO) content, p-eNOS and p-Akt, while MARCH5 knockdown exerted the opposite effects. The protective effects mediated by MARCH5 overexpression on ECs could be inhibited by eNOS inhibitor L-NAME and Akt inhibitor LY294002. In conclusion, these results indicated that MARCH5 acts as a protective factor in ischaemia/hypoxia-induced ECs injury partially through Akt/eNOS pathway.     

Mechanism of estrogen-mediated intestinal protection following trauma-hemorrhage: p38 MAPK-dependent upregulation of HO-1

p38 MAPK has been reported to regulate the inflammatory response in various cell types via extracellular stimuli. p38 MAPK activation also results in the induction of heme oxygenase (HO)-1, which exerts potent anti-inflammatory effects. Although studies have shown that 17beta-estradiol (E(2)) prevented organ dysfunction following trauma-hemorrhage, it remains unknown whether p38 MAPK/HO-1 plays any role in E(2)-mediated attenuation of intestinal injury under those conditions. To study this, male rats underwent trauma-hemorrhage (mean blood pressure approximately 40 mmHg for 90 min) followed by fluid resuscitation. At the onset of resuscitation, rats were treated with vehicle, E(2) (1 mg/kg body wt), the p38 MAPK inhibitor SB-203580 (2 mg/kg body wt) or E(2) plus SB-203580. Two hours thereafter, intestinal myeloperoxidase (MPO) activity and lactate, TNF-alpha, IL-6, ICAM-1, cytokine-induced neutrophil chemoattractant (CINC)-1, and macrophage inflammatory protein (MIP)-2 levels were measured. Intestinal p38 MAPK and HO-1 protein levels were also determined. Trauma-hemorrhage led to an increase in intestinal MPO activity and lactate, TNF-alpha, IL-6, ICAM-1, CINC-1, and MIP-2 levels. This was accompanied with a decrease in intestinal p38 MAPK activity and increase in HO-1 expression. Administration of E(2) normalized all the above parameters except HO-1, which was further increased following trauma-hemorrhage. Administration of SB-203580 with E(2) abolished the E(2)-mediated restoration of the above parameters as well as the increase in intestinal HO-1 expression following trauma-hemorrhage. These results suggest that the p38 MAPK/HO-1 pathway plays a critical role in mediating the salutary effects of E(2) on shock-induced intestinal injury.     

Mechanism of cardiac depression after trauma-hemorrhage: increased cardiomyocyte IL-6 and effect of sex steroids on IL-6 regulation and cardiac function

A prolonged depression of cardiovascular function occurs in males after trauma-hemorrhagic shock (T-H). Although a correlation between increased circulatory IL-6 levels and poor outcome has been reported after T-H, it remains unknown whether T-H increases IL-6 levels locally in cardiomyocytes and whether there is a correlation between altered cardiac function and local IL-6 production after T-H. T-H was induced in normal, castrated (2 wk before T-H), and 17beta-estradiol (E2)-treated (0.5 mg sc, 1 wk before T-H) adult male rats. At 2 h after T-H or sham operation, cardiac output, heart rate, mean arterial pressure, positive and negative first derivative of pressure (+/-dP/dt), stroke volume, and total peripheral resistance were determined. Cardiomyocytes were isolated and divided into two parts: one was used for measurements of intracellular IL-6 levels using fluorescein-activated cell sorting, and the other was used to isolate RNA to determine IL-6 gene expression by quantitative real-time PCR. In addition, cardiac IL-6 protein levels were measured in freshly isolated hearts by Western blotting. Cardiac output, stroke volume, +dP/dt, -dP/dt, and total peripheral resistance were markedly altered after T-H. These parameters, except -dP/dt, improved significantly in the castrated group; however, all these parameters were restored in E2-treated males. Cardiomyocyte IL-6 mRNA expression and intracellular IL-6 production increased after T-H. Cardiac IL-6 protein levels increased after T-H in freshly isolated heart. Castration and E2 treatment attenuated cardiomyocyte intracellular IL-6 levels and cardiac IL-6 protein levels after T-H; however, only E2 treatment attenuated cardiomyocyte IL-6 gene expression. Thus there is an inverse correlation between cardiomyocyte IL-6 levels and cardiac function after T-H. The salutary effects of E2 on cardiac function after T-H may be due in part to decreased IL-6 synthesis in cardiomyocytes.