Protective effects of ghrelin on brain mitochondria after cardiac arrest and resuscitation.

Mitochondrial dysfunction plays a critical role in brain injury after cardiac arrest (CA) and cardiopulmonary resuscitation (CPR). Our recent study demonstrated that ghrelin protected against post-resuscitation brain injury with an elevated expression of mitochondrial uncoupling protein 2 (UCP2). However, the effects of ghrelin on mitochondrial dysfunction after CA are not clear. In the present study, the protective role of ghrelin was evaluated on mitochondrial dysfunction and the subsequent damage induced by CA in rats. In addition, mitochondrial unfolded protein response (UPRmt), an intrinsic cytoprotective pathway, was observed at the same time. Either vehicle (saline) or ghrelin (80 µg/kg) was injected blindly immediately after 6 min of CA and successful resuscitation. Neurological deficit was evaluated 6 h after CA and then cortex was collected for assessments. As a result, we found that ghrelin significantly improved the neurological deficit score in rats after CA. The functional analysis of isolated mitochondria revealed that ghrelin improved the mitochondrial ATP synthesis capacity and significantly reduced the reactive oxygen species (ROS) leakage after 6 h of CA. Concomitantly, we observed an increased ATP level and an attenuated oxidative stress in ghrelin treated animals. Moreover, ghrelin markedly improved the mitochondrial morphology compared with the vehicle animals. Further research revealed that ghrelin treatment significantly activated the UPRmt as demonstrated by the increased expression of heat shock protein 60 (HSP60), heat shock protein 10 (HSP10), caseinolytic protease 1 (CLPP1), and high-temperature requirement protein A2 (HTRA2). Our results suggest that ghrelin protected against cerebral mitochondria dysfunction after CA and the mechanism may involve a UPRmt pathway.

Inhibiting Succinate Dehydrogenase by Dimethyl Malonate Alleviates Brain Damage in a Rat Model of Cardiac Arrest.

Brain damage is a leading cause of death in patients with cardiac arrest (CA). The accumulation of succinate during ischemia by succinate dehydrogenase (SDH) is an important mechanism of ischemia-reperfusion injury. It was unclear whether inhibiting the oxidation of accumulated succinate could also mitigate brain damage after CA. In this study, rats were subjected to a 6 min of CA, and cardiopulmonary resuscitation (CPR) was performed with administration of normal saline or dimethyl malonate (DMM, a competitive inhibitor of SDH). After the return of spontaneous circulation, neurological function of the rats was assessed by a tape removal test for 3 days. The rats were then sacrificed, and their brains were used to assess neuronal apoptosis by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Hippocampal tissues were used for Western blotting analysis and biochemical detection. In addition, hippocampal mitochondria during CA and CPR were isolated. The relative mitochondrial membrane potential (MMP) and cytochrome C in the cytosol were detected. Our results show that DMM promoted ROSC and neurological performance in rats after CA. The TUNEL assay showed that DMM reduced neuronal apoptosis. Western blotting analysis showed that DMM inhibited the activation of caspase-3 and enhanced the expression of HIF-1α. Moreover, DMM inhibited excessive hyperpolarization of MMP after CPR, and prevented the release of cytochrome C. Therefore, inhibiting SDH by DMM alleviated brain damage after CA, and the main mechanisms included inhibiting the excessive hyperpolarization of MMP, reducing the generation of mtROS and stabilizing the structure of HIF-1α.

Design, synthesis, and biological evaluation of novel EF24 and EF31 analogs as potential IκB kinase ß inhibitors for the treatment of pancreatic cancer.

Given the important role that inhibitory kappa B (IκB) kinase ß (IKKß) plays in pancreatic cancer (PC) development and progression, inhibitors targeting IKKß are believed to be increasingly popular as novel anti-PC therapies. Two synthetic molecules, named EF24 and EF31, exhibited favorable potential in terms of inhibition of both IKKß activity and PC cell proliferation. Aiming to enhance their cellular efficacy and to analyze their structure-activity relationship, four series of EF24 and EF31 analogs were designed and synthesized. Through kinase activity and vitality screening of cancer cells, D6 displayed excellent inhibition of both IKKß activity and PC cell proliferation. Additionally, multiple biological evaluations showed that D6 was directly bound to IKKß and significantly suppressed the activation of the IKKß/nuclear factor κB pathway induced by tumor necrosis factor-α, as well as effectively inducing cancer cell apoptosis. Moreover, molecular docking and molecular dynamics simulation analysis indicated that the dominant force between D6 and IKKß comprised hydrophobic interactions. In conclusion, D6 may be a promising therapeutic agent for PC treatment and it also provides a structural lead for the design of novel IKKß inhibitors.

Sphingosine kinase 1: A novel independent prognosis biomarker in hepatocellular carcinoma.

Sphingosine kinase 1 (Sphk1) is an oncogenic kinase that is responsible for the phosphorylation of sphingosine to sphingosine-1-phosphate (S1P). Mounting evidence suggests that Sphk1 serves a crucial role in the proliferation and development of a variety of human cancer cells. However, the role of Sphk1 in hepatocellular carcinoma (HCC) has not been fully elucidated. Therefore, the expression of Sphk1 was examined in 127 formalin-fixed, paraffin-embedded HCC tissues using immunohistochemistry, and its clinical implications and prognostic significance were analyzed. As a result, the expression of Sphk1 in HCC tissue was revealed to be significantly higher than in normal tissue (P<0.01). In addition, Sphk1 expression was significantly associated with tumor size, tumor stage and histological differentiation (all P<0.05). The patients with low Sphk1 expression had higher overall survival and recurrence-free survival rates compared with patients with high Sphk1 expression. Furthermore, Sphk1-specific shRNA was used to downregulate the expression of Sphk1 in HCC cell lines, including hepatoblastoma G2 and HCC-9724. The CRISPR/Cas9 based transcription activation system was used to upregulate Sphk1 expression in the normal live cell, L02. Cell proliferation, mRNA expression and protein expression were measured using Cell Counting Kit-8, reverse transcription polymerase chain reaction and western blot analysis in the transfected cells. To the best of our knowledge, the present study provides the first evidence that Sphk1 promotes HCC cell proliferation and is involved in tumor progression. Notably, the data presented suggest that Sphk1 may be a potential independent prognosis biomarker for the treatment of HCC.

Schisantherin A suppresses interleukin-1ß-induced inflammation in human chondrocytes via inhibition of NF-κB and MAPKs activation.

Osteoarthritis is a degenerative joint disease that is characterized by the inflammation of synovium. Schisantherin A (SchA), a dibenzocyclooctadiene lignan isolated from the fruit of Schisandra sphenanthera, has been shown to have anti-inflammatory activity. The aim of this study was to investigate the anti-inflammatory effects of SchA on interleukin-1ß (IL-1ß)-stimulated human osteoarthritis chondrocytes. Human osteoarthritis chondrocytes were pretreated with SchA 1h before IL-1ß treatment. The effects of SchA on NO, PGE2, iNOS, COX-2, and TNF-α production were detected in this study. The production of MMP-1, MMP3, MMP13 were measured by ELISA. The expression of NF-κB and MAPKs were detected by western blotting. Our results showed that SchA inhibited IL-1ß-induced NO, PGE2, and TNF-α production in a dose-dependent manner. Moreover, IL-1ß-induced MMP1, MMP3, and MMP13 expression were significantly inhibited by treatment of SchA. In addition, SchA significantly inhibited IL-1ß-induced NF-κB and MAPKs activation. Taken together, these results suggest that SchA exhibits anti-inflammatory effects against IL-1ß-stimulated chondrocytes by blocking NF-κB and MAPKs signaling pathways.

Berberine modulates cisplatin sensitivity of human gastric cancer cells by upregulation of miR-203.

Chemotherapeutic resistance is the main reason of the failure in clinical treatment of gastric cancer. Berberine (BER) is the active compound of traditional Chinese medicine Huang Lian. The aim of this present study is to evaluate the effect of BER on cisplatin resistance in gastric cancer cells and to investigate its possible mechanism. Gastric cancer cell lines SGC-7901 and BGC-823 and their respective cisplatin-resistant variants SGC-7901/DDP and BGC-823/DDP were used in this study. We found that BER treatment significantly reversed cisplatin sensitivity and induced caspase-dependent apoptosis in SGC-7901/DDP and BGC-823/DDP cells; BER treatment induced miR-203 expression, and overexpression of miR-203 mimicked the cisplatin-sensitizing effect of BER. Importantly, we showed that miR-203 was able to target the 3'UTR of Bcl-w. Therefore, we conclude that BER treatment reduces cisplatin resistance of gastric cancer cells by modulating the miR-203/Bcl-w apoptotic axis. BER may be a novel agent to enhance chemotherapeutic responses in cisplatin-resistant gastric cancer patients.

Downregulated pseudogene CTNNAP1 promote tumor growth in human cancer by downregulating its cognate gene CTNNA1 expression.

Accumulating evidence indicates that deregulation of cancer-associated pseudogene is involved in the pathogenesis of cancer. In the study, we demonstrated that pseudogene CTNNAP1, for the CTNNA1 gene, was dysregulated in colorectal cancer and the degree of dysregulation was remarkably associated with tumor node metastasis (TNM) stage (P<0.05). The mechanistic experiments revealed that pseudogene CTNNAP1 played a pivotal role in the regulation of its cognate gene CTNNA1 by competition for microRNA-141. Moreover, gain-of-function approaches showed that overexpression of CTNNAP1 or CTNNA1 significantly inhibited cell proliferation and tumor growth in vitro and in vivo by inducing G0/G1 cell cycle arrest. Our findings add a new regulatory circuit via competing endogenous RNA (ceRNA) cross-talk between pseudogene CTNNAP1 and its cognate gene CTNNA1, and provide new insights into potential diagnostic biomarker for monitoring human colorectal cancer.

Pitavastatin suppressed liver cancer cells in vitro and in vivo.

Pitavastatin classically functions as a blood cholesterol-lowering drug. Previously, it was discovered with antiglioma stem cell properties through drug screening. However, whether it can be used for liver cancer cell therapy has never been reported. In this study, the cell viability and colony formation assay were utilized to analyze the cytotoxicity of pitavastatin on liver cancer cells. The cell cycle alteration was checked after pitavastatin treatment. Apoptosis-related protein expression and the effect of caspase inhibitor were also checked. The in vivo inhibitory effect of pitavastatin on the growth of liver tumor was also tested. It was found that pitavastatin inhibited growth and colony formation of liver cancer Huh-7 cells and SMMC7721 cells. It induced arrest of liver cancer cells at the G1 phase. Increased proportion of sub-G1 cells was observed after pitavastatin treatment. Pitavastatin promoted caspase-9 cleavage and caspase-3 cleavage in liver cancer cells. Caspase inhibitor Z-VAD-FMK reversed the cleavage of cytotoxic effect of pitavastatin. Moreover, pitavastatin decreased the tumor growth and improved the survival of tumor-bearing mice. This study suggested the antiliver cancer effect of the old drug pitavastatin. It may be developed as a drug for liver cancer therapy.

Effects of ghrelin on postresuscitation brain injury in a rat model of cardiac arrest.

Poor neurological outcome remains a major problem in patients with cardiac arrest. Ghrelin has been shown to be neuroprotective in models of neurologic injury in vitro and in vivo. This study was performed to assess the effects of ghrelin on postresuscitation brain injury in a rat model of cardiac arrest. Sprague-Dawley rats were subjected to 6-min cardiac arrest and resuscitated successfully. Either vehicle (saline) or ghrelin (80 µg/kg) was injected blindly immediately after return of spontaneous circulation (ROSC). A tape removal test was performed to evaluate neurological function at 24, 48, and 72 h after ROSC. Then, brain tissues were harvested and coronal brain sections were analyzed by hematoxylin and eosin (HE) staining for neuronal viability and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining for apoptosis in hippocampal CA1 sectors. In additional groups, rats were sacrificed at 6 h after ROSC, and hippocampal tissues were collected for further analysis. We found that animals treated with ghrelin had improved neurological performances, reduced neuronal injury, and inhibited neuronal apoptosis compared with the vehicle group. Moreover, ghrelin treatment was associated with the following: (1) a decrease in caspase-3 up-regulation and an increased Bcl-2/Bax ratio, (2) a reduction in maleic dialdehyde content and an up-regulation in superoxide dismutase activity, and (3) an increase in uncoupling protein 2 (UCP-2) expression. Our results suggest that ghrelin treatment attenuated postresuscitation brain injury in rats, possibly via regulation of apoptosis, oxidative stress, and mitochondrial UCP-2 expression. Ghrelin may have therapeutic potential when administered after cardiac arrest and cardiopulmonary resuscitation.

Role of Endoplasmic Reticulum Stress in Brain Damage After Cardiopulmonary Resuscitation in Rats.

Postcardiac arrest syndrome yields poor neurological outcomes, but the mechanisms underlying this condition remain poorly understood. This study investigated whether endoplasmic reticulum (ER) stress-mediated apoptosis is induced in injured brain after resuscitation. Sprague-Dawley rats were subjected to 6 min of cardiac arrest (CA) and then resuscitated successfully. In the first experiment, animals were sacrificed 1, 3, 6, 12, or 24 h (n = 3 per group) after successful cardiopulmonary resuscitation. Brain tissues were analyzed by real-time polymerase chain reaction and Western blotting. In the second experiment, either dimethyl sulfoxide or salubrinal (Sal; 1 mg/kg), an ER stress inhibitor, was injected 30 min before the induction of CA (n = 10 per group). Neurological deficits were evaluated 24 h after CA. Brain specimens were analyzed using electron microscopy, terminal deoxynucleotidyl transferase dUTP nick end labeling assays and immunohistochemistry. We found that the messenger RNA and protein levels of glucose-regulated protein 78, X-box binding protein 1, C/EBP homologous protein, and caspase 12 were significantly elevated after resuscitation. We also observed that rats treated with Sal exhibited an improved neurological deficit score (32.3 ± 15.5 in the Sal group vs. 49.8 ± 20.9 in controls, P < 0.05). In addition, morphological improvements in the hippocampal ER were observed in the Sal group compared with the dimethyl sulfoxide group 24 h after reperfusion. Furthermore, in situ immunostaining revealed that markers of ER stress were significantly inhibited by Sal pretreatment. Our findings suggested that ER stress and the associated apoptotic pathways were activated in the hippocampus after resuscitation. Administration of Sal 30 min before cardiopulmonary resuscitation ameliorated neurological dysfunction 24 h after CA, possibly through the inhibition of ER stress after postresuscitation brain injury.

Duodenal-jejunal bypass for the treatment of type 2 diabetes in Chinese patients with an average body mass index<24 kg/m2.

BACKGROUND: It is frequently reported that bariatric surgery often leads to resolution of type 2 diabetes mellitus (T2 DM). Limited experience with duodenal-jejunal bypass (DJB) for the treatment of T2 DM has shown controversial results. We present the first study of DJB for T2 DM patients in China. The objective of this study was to evaluate the effects of DJB in nonobese Chinese patients with T2 DM. METHODS: From March 2009 to March 2011, a total of 10 T2 DM patients with an average body mass index (BMI) of 23.8 ± 1.2 kg/m(2) were enrolled in the study. DJB was performed in all patients. BMI and glycometabolic parameters were collected at baseline and 1, 3, 6, 12, and 24 months postoperatively. Remission of T2 DM was defined as a glycosylated hemoglobin (HbA1c) level of<7% without diabetic medication. RESULTS: Remission of T2 DM was observed in 1 (10%) of 10 T2 DM patients at 6 months. Without increasing antihyperglycemic agents, fasting plasma glucose (FPG), 2-hour postprandial plasma glucose, and HbA1c decreased significantly at each postoperative time point, compared with the preoperative baseline. BMI statistically decreased at 1 and 3 months, but did not reach statistical significance at 6, 12, and 24 months. CONCLUSIONS: DJB can improve glycemic control in nonobese T2 DM patients without significant weight loss but may not be effective enough to induce remission of T2 DM in nonobese Chinese patients. A larger sample size and more constrictive inclusion criteria may be required for better evaluation.

Protective and biogenesis effects of sodium hydrosulfide on brain mitochondria after cardiac arrest and resuscitation.

Mitochondrial dysfunction plays a critical role in brain injury after cardiac arrest and cardiopulmonary resuscitation (CPR). Recent studies demonstrated that hydrogen sulfide (H2S) donor compounds preserve mitochondrial morphology and function during ischemia-reperfusion injury. In this study, we sought to explore the effects of sodium hydrosulfide (NaHS) on brain mitochondria 24h after cardiac arrest and resuscitation. Male Sprague-Dawley rats were subjected to 6min cardiac arrest and then resuscitated successfully. Rats received NaHS (0.5mg/kg) or vehicle (0.9% NaCl, 1.67ml/kg) 1min before the start of CPR intravenously, followed by a continuous infusion of NaHS (1.5mg/kg/h) or vehicle (5ml/kg/h) for 3h. Neurological deficit was evaluated 24h after resuscitation and then cortex was collected for assessments. As a result, we found that rats treated with NaHS revealed an improved neurological outcome and cortex mitochondrial morphology 24h after resuscitation. We also observed that NaHS therapy reduced intracellular reactive oxygen species generation and calcium overload, inhibited mitochondrial permeability transition pores, preserved mitochondrial membrane potential, elevated ATP level and ameliorated the cytochrome c abnormal distribution. Further studies indicated that NaHS administration increased mitochondrial biogenesis in cortex at the same time. Our findings suggested that administration of NaHS 1min prior CPR and followed by a continuous infusion ameliorated neurological dysfunction 24h after resuscitation, possibly through mitochondria preservation as well as by promoting mitochondrial biogenesis.

Effects of sodium hydrosulfide on intestinal mucosal injury in a rat model of cardiac arrest and cardiopulmonary resuscitation.

AIMS: Cardiac arrest and cardiopulmonary resuscitation (CPR) can lead to intestinal ischemia/reperfusion (I/R) injury. Increasing studies have indicated that hydrogen sulfide (H2S) is in favor of a variety of tissue I/R injury. The purpose of this study was to explore whether sodium hydrosulfide (NaHS), a H2S donor, can protect intestinal mucosa after CPR and its potential mechanisms. MAIN METHODS: Male Sprague-Dawley rats were subjected to 6min cardiac arrest induced by transcutaneous electrical epicardium stimulation and then resuscitated successfully. A bolus of either NaHS (0.5mg/kg) or placebo (NaCl 0.9%) was blindly injected 1min before the start of CPR intravenously, followed by a continuous injection of NaHS (2mg/kg/h) or placebo for 3h. Intestinal and plasma samples were collected for assessments 24h after CPR. KEY FINDINGS: We found that NaHS can markedly alleviate cardiac arrest induced intestinal mucosal injury. Rats treated with NaHS showed a lower malondialdehyde content, higher superoxide dismutase activity and glutathione content in intestine after CPR. Increased intestinal myeloperoxidase activity was significantly decreased by NaHS after CPR. Moreover, a reduced intestinal apoptotic cells after CPR were evident when pretreated with NaHS. Further studies indicated that NaHS enhances the expression of hypoxia-inducible factor-1α (HIF-1α) in intestine after CPR. SIGNIFICANCE: Our data demonstrated that NaHS treatment before CPR induces intestinal mucosal protection 24h post-resuscitation. The protective effects may be through oxidative stress reduction, inflammation alleviation, apoptosis inhibition and HIF-1α activation.