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The present study aimed to investigate whether the drug nicorandil can improve cardiac remodeling after myocardial infarction (MI) and the underlying mechanisms.Mouse MI was established by the ligation of the left anterior descending coronary artery and H9C2 cells were cultured to investigate the underlying molecular mechanisms. The degree of myocardial collagen (Col) deposition was evaluated by Masson's staining. The expressions of nucleolin, autophagy and myocardial remodeling-associated genes were measured by Western blotting, qPCR, and immunofluorescence. The apoptosis of myocardial tissue cells and H9C2 cells were detected by TUNEL staining and flow cytometry, respectively. Autophagosomes were observed by transmission electron microscopy.Treatment with nicorandil mitigated left ventricular enlargement, improved the capacity of myocardial diastolic-contractility, decreased cardiomyocyte apoptosis, and inhibited myocardial fibrosis development post-MI. Nicorandil up-regulated the expression of nucleolin, promoted autophagic flux, and decreased the expressions of TGF-β1 and phosphorylated Smad2/3, while enhanced the expression of BMP-7 and phosphorylated Smad1 in myocardium. Nicorandil decreased apoptosis and promoted autophagic flux in HTreatment with nicorandil alleviated myocardial remodeling post-MI through up-regulating the expression of nucleolin, and subsequently promoting autophagy, followed by regulating TGF-β/Smad signaling pathway.

Sepsis with subsequent multiple-organ dysfunction is a distinct systemic inflammatory response to concealed or obvious infection, and it is a leading cause of death in intensive care units. Thus, one of the key goals in critical care medicine is to develop novel therapeutic strategies that will affect favorably on outcome of septic patients. In addition to systemic response to infection, apoptosis is implicated to be an important mechanism of the death of immune cells, including neutrophils, macrophages, T lymphocytes, and dendritic cells, and it is usually followed by the development of multiple-organ failure in sepsis. The implication of apoptosis of immune cells is now highlighted by multiple studies that demonstrate that prevention of cell apoptosis can improve survival in relevant animal models of severe sepsis. In this review, we focus on major apoptotic death pathways and molecular mechanisms that regulate apoptosis of different immune cells, and advances in these areas that may be translated into more promising therapies for the prevention and treatment of severe sepsis.

The present study aimed to investigate whether puerarin regulated tissue factor (TF) expression induced by oxidative low-density lipoprotein (ox-LDL), an independent risk factor for atherosclerosis, and its mechanisms.TF expression at the mRNA level was determined by reverse transcription-quantitative polymerase chain reaction, and its expression at the protein level, as well as other target proteins, was assessed by western blotting. Nitric oxide (NO) production was measured by a nitrate reduction method.Results demonstrated that treatment with ox-LDL (50mg/l) for 24h significantly increased (P0.01) TF expression at the mRNA and protein levels in human umbilical vein endothelial cells (HUVECs). Such an ox-LDL exposure also triggered the dephosphorylation of Akt, resulting in a reduction of NO production and activated the extracellular signal-regulated kinase (ERK)1/2 and nuclear factor (NF)-κB signaling pathways. Pre-treatment with puerarin (50-200μM) for 1h significantly attenuated the ox-LDL-induced TF expression, augmented the phosphorylation of Akt, with a resultant increase of the NO production, and inhibited the activation of ERK1/2 and NF-κB (P0.01). However, this beneficial effect of puerarin (100μM) was abolished by LY294002 (10μM), an inhibitor of phosphoinositide 3-kinase (PI3K), or NG-nitro-L-arginine methyl ester (100μM), an inhibitor of NO synthase.These results suggested that puerarin suppressed TF expression in HUVECs through activating the PI3K/Akt/endothelial nitric oxide synthase signaling pathway and inhibiting the activation of ERK1/2 and NF-κB. These findings suggested that puerarin possessed certain anticoagulation and may be a potential novel therapeutic drug for thrombosis in coronary artery disease.

Sepsis with subsequent multiple organ dysfunction is a pronounced systemic inflammatory response to concealed or known infection and is a leading cause of death in intensive care units. The survival rate of severe sepsis and septic shock has not markedly improved in recent decades despite a great number of receptors and molecules involved in its pathogenesis have been found and taken as therapeutic targets. It is essential to thoroughly understand the host cell-mediated immunity involved in the development of sepsis and sepsis-related organ injury. Recent studies indicate that innate immune cells (such as neutrophils, macrophages, dendritic cells, T lymphocytes, regulatory T cells, and natural killer T cells) play pivotal roles in the maintenance of peripheral homeostasis and regulation of immune responses during sepsis. Therefore, an understanding of the biological significance and pathophysiological roles of different cell populations might gain novel insights into the immunoregulatory mechanisms of sepsis. In this review, we focus on major immune cells that may play potential roles in the contribution of new therapeutic approaches for sepsis.

To investigate the release and intracellular localization of high mobility group box chromosomal protein 1 (HMGB1) in the peripheral blood monocytes of rheumatoid arthritis (RA) patients and the inhibitive effect of thalidomide.19 RA patients and 20 healthy controls were included in the study. Monocytes were separated from peripheral blood with Ficoll density gradient centrifugation. Monocytes were treated with 100 ng/ml tumor necrosis factor alpha (TNFalpha) or 100 ng/ml TNFalpha plus 40 microg/ml thalidomide and grown in an incubator at 37 degrees C with 5% CO2 for 24 hours. The culture supernatants of the monocytes were collected. HMGB1 level in the culture medium was detected with Western blot. In addition, the intracellular localization of HMGB1 in the monocytes was investigated with immunocytochemical analysis.Without stimulation, the release of HMGB1 protein was significantly increased in the culture supernatants of peripheral blood monocytes from RA patients as compared with that from healthy controls (P0.05). TNFalpha (100 ng/ml) did not further increase the release of HMGB1 in the monocytes from the patients with RA. Thalidomide (40 microg/ml) could inhibit the release of HMGB1 in the monocytes from RA patients stimulated with TNFalpha (P0.05). In the monocytes from RA patients, HMGB1 was mainly localized in the nucleus. Treatment with TNFalpha (100 ng/ml) for 24 hours resulted in a cytoplasmic translocation of HMGB1, which was inhibited significantly by thalidomide.TNFalpha induces the release and cytoplasmic translocation of HMGB1 in the peripheral blood monocytes of RA patients and thalidomide inhibits the release and translocation of HMGB1.

To observe the effect of Krüppel-like factor 4 (KLF4) overexpression on heat stress-induced apoptosis of Raw264.7 macrophages.The fragment containing full length mouse KLF4 cDNA coding sequence was inserted into the pcDNA3.1 vector and Raw264.7 macrophages were transfected with pcDNA3.1-KLF4 plasmids using lipofectamine.The expression of KLF4 was examined by Western blot in the Raw264.7 macrophages stably transfected with pcDNA3.1- KLF4 plasmids. Raw264.7 cells transfected with pcDNA3.1 and pcDNA3.1-KLF4 were exposed to heat stress (42 degrees C) for 1h and recovered at 37 degrees C for 12h. Flow cytometry, Hoechest 33258 staining assay, and DNA ladder assays were performed to assess the apoptosis.The KLF4 overexpressed Raw264.7 macrophages were established. After the heat stress, flow cytometry showed that apoptotic cells increased significantly in KLF4 overexpressed cells compared with the vector control; Hoechest 33258 staining was characterized with classical changes including apoptotic body, and nuclear condensation. Agarose gel electrophoresis showed that "DNA ladder" could be observed clearly.KLF4 overexpression can increase heat stress-induced apoptosis of Raw264.7 macrophages.

To investigate the mRNA and protein expression of high mobility group box chromosomal protein 1 (HMGB1) in peripheral blood mononuclear cells (PBMCs) and serum.Levels of HMGB1mRNA were detected with reverse transcription polymerase chain reaction (RT-PCR) in PBMC and levels of HMGB1 protein in PBMC and plasm were measured with Western blot in 38 patients with active rheumatoid arthritis (RA), 24 with inactive RA and 20 healthy controls. Ficoll density gradient centrifugation was used to separate PBMCs from peripheral blood. The correlation between the levels of HMGB1 in serum and the index of disease activity in RA was analyzed.There was no statistically significant difference of the mRNA expression of HMGB1 in PBMCs from active RA patients as compared with those from inactive RA group and healthy controls (F = 1.23, P0.05). The protein expression of HMGB1 was significantly lower in PBMCs from active RA patients (F = 70.91, P0.01), while the protein expression of HMGB1 was higher in plasma from active RA patients (P0.001) as compared with that from inactive RA patients and healthy controls. However, there was no statistically significant difference between inactive RA patients and healthy controls (P0.05). The level of HMGB1 protein in plasma of RA patients was correlated with erythrocyte sedimentation rate (ESR) (r(s) = 0.478, P0.001) and C- reactive protein (CRP) (r(s) = 0.574, P0.05). It was not correlated with age, the number of tender joints and swollen joints, radiographic scores and therapeutic effect.The protein expression of HMGB1 was significantly decreased in PBMCs from active RA patients, while it was increased in serum from active RA patients. HMGB1 plays a pivotal role in the pathogenesis of RA and may be a target of therapy as a novel cytokine.

Since the findings of Murry and Currie et al. that ischemic preconditioning (IPC) and heat shock response (HSR) could protect evidently myocardium against ischemia-reperfusion injury in the middle of 1980s, endogenous myocardial protection has drawn widespread attentions. A great quantity of studies completed during the past 25 years made much progress in endogenous myocardial protection. Abundant research experiences have been accumulated and a basic theoretical framework has been established in this field. However, there are still many questions need to be solved. In this review, we focused on clarifying some hot questions and important future directions in IPC, heat shock proteins (HSPs), research models and strategies in endogenous myocardial protection.

Cytokine mediated cell immunity is the main mode of anti-tumor immunity in organism, and the disequilibrium of cytokine network is the main cause of tumor cells escaping immunologic surveillance. High mobility group box 1 (HMGB1), a nuclear protein, has recently been identified as an important mediator of local and systemic inflammatory diseases when released into the extracellular milieu. In the present study, the investigators explored the clinical significance of alteration in the serum levels of HMGB1 in childhood acute lymphocytic leukemia (ALL) and the mechanism of HMGB1-induced tumor necrosis factor (TNF)-alpha secretion in leukemic cells.The serum levels of HMGB1 in healthy children and childhood ALL were assayed by Western blotting. K562 leukemic cells were stimulated with recombinant HMGB1 protein in vitro, and the secretion of TNF-alpha was determined by using ELISA. The effects of HMGB1 on activation of p38, c-Jun amino-terminal kinase (JNK), and extracellular-signal regulated protein kinase (ERK) and mitogen-activated protein kinase (MAPK) in K562 cells were assayed by using Western blotting. The effects of inhibitors specific for the MAPK on HMGB1-induced TNF-alpha secretion were assayed by using ELISA.The serum levels of HMGB1 were significantly higher in ALL initial treatment group (n = 15, 43.78 +/- 4.62 microg/ml) than those in healthy control group (n = 15, 0.60 +/- 0.48 microg/ml, P0.01) and ALL complete remission group (n = 15, 0.89 +/- 0.62 microg/ml, P0.01). No significant difference was found between the healthy control group and ALL complete remission group in HMGB1 levels (P0.05). TNF-alpha started to become detectable at 2 h and was still increasing at 16 h after HMGB1 (1 microg/ml) treatment in K562 cell culture. TNF-alpha was also secreted from K562 cells in a dose-dependent manner after HMGB1 (1 ng/ml-1 microg/ml) exposure. HMGB1 induced the phosphorylation of p38, JNK and ERK in k562 cells. Inhibitors specific for the JNK (SP600125), MEK (PD98059), and p38 MAPK (SB203580), abrogated HMGB1-induced TNF-alpha secretion.The measurement of serum HMGB1 is helpful to evaluate the prognosis of the childhood ALL. HMGB1 stimulates leukemic cells to secrete TNF-alpha through a MAPK-dependent mechanism.

To investigate the diagnostic value of heat shock protein 70 (HSP70) in central nervous system infection (CNSI) in children.The level of HSP70 in the cerebrospinal fluid (CSF) was determined in children with CNSI of different etiology. The concentration of HSP70 was determined in the CSF of 104 children, among them 13 patients had purulent meningitis (PM), 38 patients had acute viral meningitis (VM), 7 patients had tuberculous meningitis (TM), and 46 with no CNSI to serve as controls. The concentration of HSP70 was determined by Western blotting assay. The CSF specimens were also analyzed for the total cellular score (TCS), white blood cell count (WBC), lactate dehydrogenase (LDH), protein content (PC), adenosine deaminase (ADD), glucose, chloride content (Cl(í)), and pressure.The CSF level of HSP70 was significantly higher in the PM, TM and VM groups [76.61+/-27.69, 65.85+/-33.16, 33.65+/-16.93] compared with the control group (23.28+/-19.77) (P0.05 or P0.01). The HSP70 concentration was markedly higher in the CSF of patients with PM and TM than patients with VM (both P0.01). No significant difference was found between PM group and TM group in HSP70 level in CSF (P0.05). The concentration of HSP70 in the CSF was positively correlated to TCS (r=0.298, P=0.002), WBC (r=0.274, P=0.005), LDH (r=0.322, P=0.001), PC (r=0.629, P0.001), ADD (r=0.363, P0.001), and negatively correlated to the glucose (r=-0.443, P0.001) in CSF. The HSP70 concentration was not correlated to the Cl(í) (r=0.148, P=0.133) and pressure (r=0.001, P=0.993) of CSF.HSP70 is increased in the CSF of patients with CNSI. It may be one of the pathophysiological mechanisms of acute infection of the central nervous system. The level of HSP70 in CSF may be a valuable index in the differential diagnosis of CNSI, and it may be helpful in differentiating PM and TM from VM.

To investigate the killing effects of VP(3) on nasopharyngeal carcinoma cell line CNE-2.Plasmid expression vector pcDNA3.1(-) CMV.VP(3)-His was constructed and identified by Kpn I/EcoR I endonuclease analysis, and then sequenced to verify successful insertion in the sense direction of VP(3) gene. pcDNA3.1(-) CMV.VP(3)-His and pcDNA3.1(-)-His expression plasmid was transiently transfected into nasopharyngeal carcinoma cell line CNE-2 . VP(3) protein expression was detected by Western blotting. MTT assay was used to determine the killing effects of VP(3) gene on nasopharyngeal carcinoma cell line CNE-2.Endonuclease analysis and sequencing confirmed the recombinant plasmid contained the complete VP(3) CDS sequence. Western blotting detected a 14.03 kD protein expression from the transfected cells, which was the expecting band of VP(3) gene. The growth of CNE-2 cells that expressed VP(3) gene was inhibited,while the growth of CNE-2 cells that did not express VP(3) gene was not inhibited.VP(3) gene can kill nasopharyngeal carcinoma cell CNE-2.

To observe the proliferation of SW480 cells exposed to different concentrations of CoCl2, and to examine the expression of hypoxiainducible factor-1 alpha (HIF-1alpha) and heme oxygenase-1 (HO-1) during hypoxia to explore the chemotherapy resistance effect and role of HIF-1alpha and HO-1.Methyl thiazolyl tetrazolium (MTF) method was used to detect the proliferation of SW480 cells in the presence of fluorouracil (FU). RT-PCR was applied to examine the expression of HIF-1alpha and HO-1 mRNA in hypoxia.SW480 cells were proliferated at a slow rate, and had a strong resistance to FU with the increase of CoCl2. RT-PCR showed that the up-regulated expression of HIF-1alpha and HO-1 mRNA was consistent with the dose-effect curve and time-effect curve.The hypoxia induced by CoCl2 can inhibit the proliferation of SW480, and it can also decrease the sensitivity of the cell to FU. The mechanism is probably related to the up-regulated expression of HIF-1alpha and HO-1 mRNA.

To observe the effect of heat shock factor 1 (HSF1) on heat stress-induced apoptosis in Raw264.7 macrophages.Raw264.7 cells transfected with pcDNA3.1 and pcDNA3.1-HSF1 were exposed to heat stress (42.5 degrees C +/- 0.5 degrees C) for 1 h and recovered at 37 degrees C for 6, 9, 12, and 24 h respectively. Flow cytometry (FCM), Hoechst 33258 staining and DNA ladder assays were performed to assess the apoptosis.After heat stress, FCM showed that apoptotic cells were increased significantly and reached the peak at 9 h in Raw 264.7 cells transfected with pcDNA3.1, and were characterized with classical morphologic changes including apoptotic body and nuclear condensation. Agarose gel electrophoresis showed that "DNA ladder" could be observed clearly at 6, 9, and 12 h after the heat stress. But the overexpression of HSF1 could reduce the number of apoptotic cells and inhibit DNA fragmentation.HSF1 can inhibit heat stress-induced apoptosis in Raw264.7 macrophages.

To establish immortalized embryonic fibroblast lines in heat shock transcription factor 1 (HSF1) HSF1-/- and HSF1+/+ mice and to provide experimental models to study the function of HSF1.A mammalian expression vector (pSV3neo) containing the SV40 large T antigen was used to transfect the HSF1-/- and HSF1+/+ mouse embryonic fibroblast using Lipofectamine 2000. Colonies were screened by G418 and expanded to immortalized cell lines. PCR was used to detect the integration of the large T antigen with genome in the mouse embryonic fibroblast. Expression of SV40 large T antigen gene in expanded cells was identified by RT-PCR. HSP70 expression was examined by Western blot in the embryonic fibroblast lines.The stable growth and serial propagation were observed in the HSF1-/- and HSF1+/+ cell lines for six months. The mRNA of SV40 T antigen gene expressed in the two cell lines. HSP70 expression could not be induced in the heat-treated HSF1-/- mouse embryo fibroblasts.The immortalized cells of HSF1+/+ and HSF1-/- mouse embryo fibroblasts are successfully established.

To explore the protective effect of HSP72 on the acute injury of cardiomyocyte induced by oxidative stress.Cardiomyocytes of neonatal rats treated with heat shock (42 degrees C, 30 min, recovery for 6 h) to induce the expression of HSP72 and HSP72 antisense oligonucleotide was transformed to block the expression of HSP72. 0.5 mmol/L (final concentration) H2O2 was added into the culture medium to mimic oxidative stress, and to induce the acute injury of neonatal cardiomyocytes. The release of LDH and the total protein synthesis were applied to evaluate the injury of cardiomyocyte of neonatal rats.Oxidative stress could significantly increase the release of LDH, and inhibit the total protein synthesis. By inducing the expression of HSPs, heat shock pretreatment significantly reduced the release of LDH and relieved the oxidative stress-mediated inhibition of total protein synthesis. Moreover, HSP7-2 anti-sense oligonucleotide could remarkably block the protective effect of heat shock pretreatment on the cellular injuries induced by H2O2.HSP72 plays a most important role in the acute injury of cardiomyocyte mediated by oxidative stress.

To screen the inflammatory mediators genes regulated by HSF1, and explore the mechanism of downstream genes regulated by HSF1.HSF-/- and HSF1+/+ mice were injected with 15 mg/kg LPS intraperitoneally (ip), respectively, and were treated as previous after HSR. The total RNA of lung tissues were extracted and filtrated by SuperArray gene Microarry. The promoter of candidate genes were analyzed by transcription element search software to search for heat shock element (HSE). Select the suppressor of cytokine signaling 3 (SOCS3) with HSE. Macrophage cells were stimulated with 400 ng/mL LPS, and were treated as previous after HSR, then the total RNA was extracted respectively. RT-PCR and northern blot assay were performed to detect the expression levels of SOCS3 mRNA.Fifteen genes were repressed by HSF1, including 9 genes with complete HSE. Eleven genes were accelerated by HSF1 possibly, including 8 genes with complete HSE. The promoter of SOCS3 gene contained one complete HSE. LPS stimulation obviously increased the levels of SOCS3 mRNA in macrophages of RAW264.7 mice, which was inhibited by HSR and over-expression of HSF1.HSR or HSF1 inhibits LPS induced expression of SOCS3 mRNA; HSF1 might inhibit LPS-induced expression of SOCS3 mRNA by binding to HSE in the promoter of SOCS3 gene.

To determine the characteristics of a novel gene Mip5 (GenBank accession number AY553870) and its expression under physiological and pathological conditions.The characteristics of Mip5 were analyzed by bioinformatic programs including BLAST, spidey, psort, ClustalW and so on. RT-PCR was performed to detect Mip5 expression.Bioinformatic analysis showed that Mip5 gene lied in the 13th chromosome and contained 8 exons and 7 introns, its open reading frame contained 909 bp and its protein production was 302 amino acid residues including 6 kelth domains. Under normal conditions, MIP5 expressed abundantly in the heart, brain and kidney, but its expression could not be detected in the liver and muscle. Expression of Mip5 gene was increased significantly after ischemia-reperfusion compared with the sham groups, and reached its peak at 3 h and recovered at 12 h after the reperfusion. Conclusion Mip5 gene is a novel gene containing a putative open reading frame of 302 amino acids residues and may play an important role in rat cardiomyocytes suffering ischemia processing.

To investigate the effect of oxidative stress on the accumulation of heat shock protein 70 (HSP70) within C2C12 myogenic cells.Heat shock response (42 degrees C for 1 h and recovery for 12 h at 37 degrees C) was used to induce the expression of heat shock protein 70. We constructed a recombinant plasmid of HSP70 with enhanced green fluorescent protein (EGFP). After being transfected transiently into C2C12 cells, immunoblotting was used to detect the expression of HSP70 induced by heat shock response and transfection. Immunocytochemistry, fluorescent microscopy and immunoblotting were used to detect the translocation of HSP70.Immunoblotting showed that the overexpression of HSP70 was induced by heat shock response and transient transfenction. HSP70 localized within the cytoplasm of the normal cells, but HSP70 translocated from the cytoplasm to the nucleus and the nucleolus at 1 h after the treatment of oxidative stress (0.5 mmol/L H2O2) by using immunocytochemistry, fluorescent microscopy and immunoblotting for cellular partial proteins.Oxidative stress may induce the accumulation of heat shock protein 70 within the nucleolus.

To observe the cleavage of nucleolin (C23) during apoptosis induced by oxidative stress and to clarify the effect of heat shock response (HSR) on the cleavage of nucleolin and its possible molecular mechanism.We added 0.5 mmol/L peroxide hydrogen (H2O2 ) into cultured cells to mimic oxidative stress. Apoptosis and cleavage of C23 were detected using caspase-3 colorimetric assay and Western blotting respectively. HSR was performed to observe the effect of HSR on cleavage of C23 induced by oxidative stress, and over-expressions of HSP70 and HSP25 were detected by Western blotting.Activity of caspase-3 increased significantly after 2 hours of 0.5 mmol/L H2O2 treatment, and reached the peak after 12 hours. The cleavage of C23 appeared 30 minutes to 1 hour after the treatment of H2O2 as indicated by a cleaved fragmentation of 80 kD, which was significantly inhibited by HSR. Moreover, HSR could induce HSP70 and HSP25 over-expressions.Oxidative stress can induce the activation of caspase-3, cleavage of C23, and apoptosis. HSR can significantly inhibit the cleavage of C23 induced by oxidative stress, which is related to the over-expressions of HSP70, HSP25, and other stress proteins.

To explore the mechanisms of myocardial apoptosis during myocardial ischemia-reperfusion injury and to further clarify the molecular mechanisms by which heat shock pretreatment in inhibiting myocardial apoptosis induced by ischemia-reperfusion injury.Myocardial ischemia-reperfusion injury was induced by the occlusion of left anterior descending branch of the coronary artery. Apoptosis was evaluated by DNA laddering assay and the activities of caspase 3, 8, or 9 was measured with Caspase Colorimetric Assay Kit. Expression of heat shock proteins was detected by Western blotting analysis. To explore the effect of heat shock pretreatment on myocardium against apoptosis, mice were pretreated with whole body hyperthermia before the myocardial ischemia-reperfusion injury.Ischemia-reperfusion injury induced myocardial apoptosis and activation of caspase-3,8,9. Heat shock pretreatment induced the expression of several family members of heat shock proteins and inhibited myocardial apoptosis and activation of the above caspases.Mitochondria and death receptor signaling pathways play important roles in myocardial apoptosis induced by ischemia-reperfusion injury. Heat shock pretreatment may increase the expression of several HSP, and inhibit the activation of both mitochondria and death receptor signaling pathways and apoptosis in cardiomyocytes induced by myocardial ischemia-reperfusion injury.

To analyze the pattern of gene expression programs during rat myocardial ischemia/reperfusion at multiple time points.Rat model of myocardial ischemia/reperfusion was established by repeating the occlusion and relaxation of left coronary artery. cDNA microarray was used to analyze the pattern of gene expression programs in rat myocardium at 1, 3, 6, 12, 24 hours after reperfusion. SOM cluster analysis was used to identify different cluster of genes in which each cluster had similar expression pattern.Altogether 75, 779, 205, 155, and 166 genes were differentially expressed at 1, 3, 6, 12, 24 hours after the reperfusion respectively. Clusters analysis identified 12 clusters of genes in which each cluster had similar expression pattern.Analysis of gene expression pattern revealed sequential induction of subsets of genes that characterize each response.

To investigate the role of nuclear factor-kappaB (NF-kappaB) in heat shock pretreatment to abate cardiomyocyte injury induced by hydrogen peroxide (H(2)O(2)).The primary generation of cultured neonatal rat cardiomyocytes were injured by exposure to 1 mmol/L H(2)O(2) for different durations. The total antioxidant in cardiomyocytes was detected. The changes in heat shock protein 70 (HSP70), alphaB-crystallin, inhibitor of NF-kappaB (I-kappaB) were assayed by Western-blotting. The translocation of NF-kappaB and HSP70 from cytoplasm to nucleus was observed by immunohistochemical analysis.(1)Compared with H(2)O(2) (1 mmol/L, 3 h) treated cells, cells subjected to heat shock pretreatment showed significant increase in total antioxidant capability (all P0.01). (2)Western blot analysis demonstrated that heat shock pretreatment could induce expression of HSP70, alphaB-crystallin and I-kappaB. (3)Heat shock pretreatment inhibited H(2)O(2)-mediated I-kappaB degradation. (4)Immunohistochemical analysis showed that heat shock pretreatment could abate HSP70 and NF-kappaB translocation from cytoplasm to nucleus.Heat shock pretreatment could protect cardiomyocytes against H(2)O(2)-induced injury, and its mechanism might involve expression of HSP70, alphaB-crystallin and I-kappaB, which could inhibit H(2)O(2) -mediated NF-kappaB activation.

To clarify the effect of nucleolin on the proliferation and apoptosis in C2C12 cells.After inhibiting the expression of nucleolin using antisense oligonucleotides, the cellular proliferation was determined by MTT, and the apoptosis was detected by flow cytometry (FCM) assays and DNA ladder assays.After being transfected with antisense oligonucleotides for 24 hours, Western blotting showed that the expression of nucleolin was repressed significantly. In cells treated with antisense oligonucleotides, the cellular proliferation was obviously inhibited; the apoptotic cell increased significantly; and the "DNA ladder" was clearly observed. But the sense and random oligonucleotides had no effect on the cellular proliferation and apoptosis.The down-regulation of nucleolin can inhibit the cellular proliferation and initiate the apoptosis in C2C12cells.

To compare the effects of dobutamine under different fluids resuscitation for shock induced by intestinal ischemia/reperfusion (I/R) injury in rabbits.Thirty-two anesthetized rabbits were randomized into four groups of eight animals each. The groups were followed as: (1) lactated Ringer's solution (LRS) resuscitation; (2) LRS+hydroxyethyl starch solution (HES) resuscitation; (3) LRS resuscitation+dobutamine treatment; (4) LRS+HES resuscitation+dobutamine treatment. All these rabbits underwent the intestinal I/R injury developed by occluding superior mesenteric artery (SMA) with a non-crushing vascular clamp for 60 minutes and then loosing the clamp for 300 minutes. The fluid resuscitation and drug treatment began at the same time of reperfusion. Hemodynamic parameters including mean artery pressure (MAP), heart rate (HR), aortic velocity (AoV, as cardiac output) and SMA blood flow (Qsma) were measured. Tissue oxygenation was assessed indirectly by measuring the tonometric parameters of the gut, including difference between partial pressure of carbon dioxide in intestinal intramucosal and partial pressure of carbon dioxide in arterial blood (Pt-a CO2 gap), intestinal intramucosal pH (pHi), arterial blood lactate acid concentration and oxygen delivery (DO2).HR, AoV and Qsma as measured in two dobutamine groups were significantly higher in values than LRS and LRS+HES groups (all P0.05). But MAP as measured in two dobutamine groups were significantly higher in values than only LRS (P0.05), and in LRS+HES resuscitation+dobutamine treatment group was also significantly higher in values than LRS resuscitation+dobutamine treatment group (P0.05). Dobutamine in LRS resuscitation+dobutamine treatment and LRS+HES resuscitation+dobutamine treatment group could greatly decrease lactate and Pt-aCO2 gap, significantly improve pHi and DO2 compared with other two resuscitation groups (all P0.05). Dobutamine in LRS+HES resuscitation+dobutamine treatment group could also greatly decrease lactate and Pt-aCO2 gap, significantly improve pHi compared with LRS resuscitation+dobutamine treatment group (all P0.05).Dobutamine could improve hemodynamic parameters and tissue oxygenation in shock induced by intestinal I/R injury in rabbits, being better used under the LRS+HES resuscitation.

To compare the effects of high and low doses of 6% hydroxyethyl starch solution (HES) on resuscitation for shock induced by ischemia/reperfusion injury of the intestine in rabbits.Thirty-two anesthetized rabbits were randomized into four groups of eight animals each. The animals in the control group received no fluid resuscitation, animals in group 2 received lactated Ringer's solution (LRS, 20 ml x kg(-1) x h(-1)), those in group 3 received LRS+HES (LRS 18 ml x kg(-1) x h(1)+HES 2 ml x kg(-1) x h(-1), low dosage of HES), and those in group 4 received HES only in high dosage of HES (20 ml x kg(-1) x h(-1)). All these rabbits underwent intestinal ischemia/reperfusion injury developed by occluding superior mesenteric artery (SMA) with a non-crushing vascular clamp for 60 minutes and then the clamp was loosened for 240 minutes. The fluid resuscitation began at the same time of reperfusion. Hemodynamic parameters including mean artery pressure, heart rate, aortic velocity (as cardiac output), and SMA blood flow were measured. Tissue oxygenation was assessed indirectly by measuring the tonometric parameters of the gut, including difference between partial pressure of carbon dioxide in intestinal intramucosal and partial pressure of carbon dioxide in arterial blood (Pt-aCO(2) gap), intestinal intramucosal pH (pHi), arterial blood lactate acid concentration and oxygen delivery. Mortality of the rabbits was counted at the end of the experiment.Hemodynamic parameters as measured in low and high doses of HES groups were significantly higher in values than LRS group (LRS) and control (all P0.05). Low dose of HES was better in restoring hemodynamic parameters than high dose of HES (all P0.05). Low dose of HES could greatly decrease lactate and Pt-aCO(2) gap, significantly improve pHi compared with other three groups (all P0.05), but high dose of HES did not do so, and oral and nasal bleeding even death of some animals were seen. Low dose and high dose of HES significantly improved oxygen delivery while LRS did not (all P0.05).Low dose of HES together with LRS is more effective than only high dose of HES or LRS in the resuscitation for shock induced by ischemia/reperfusion injury of the intestine in rabbits, resulting in better hemodynamic parameters and tissue oxygenation.

Members of the heat shock factor (HSF) family are evolutionarily conserved regulators that share a highly homologous DNA-binding domain. In mammals, HSF1 is the main factor controlling the stress-inducible expression of Hsp genes while the functions of HSF2 and HSF4 are less clear. Based on its developmental profile of expression, it was hypothesized that HSF2 may play an essential role in brain and heart development, spermatogenesis, and erythroid differentiation. To directly assess this hypothesis and better understand the underlying mechanisms that require HSF2, we generated Hsf2 knockout mice. Here, we report that Hsf2(-/-) mice are viable and fertile and exhibit normal life span and behavioral functions. We conclude that HSF2, most probably because its physiological roles are integrated into a redundant network of gene regulation and function, is dispensable for normal development, fertility, and postnatal psychomotor function.

Cardiomyocytes can resist ischemia/reperfusion (I/R) injury through ischemic postconditioning (IPoC) which is repetitive ischemia induced during the onset of reperfusion. Myocardial ischemic preconditioning up-regulated protein 2 (MIP2) is a member of the WD-40 family proteins, we previously showed that MIP2 was up-regulated during ischemic preconditioning (IPC). As IPC and IPoC engaged similar molecular mechanisms in cardioprotection, this study aimed to elucidate whether MIP2 was up-regulated during IPoC and contributed to IPoC-mediated protection against I/R injury. The experiment was conducted on two models, an in vivo open chest rat coronary artery occlusion model and an in vitro model with H9c2 myogenic cells. In both models, 3 groups were constituted and randomly designated as the sham, I/R and IPoC/hypoxia postconditioning (HPoC) groups. In the IPoC group, after 45 min of ischemia, hearts were allowed three cycles of reperfusion/ischemia phases (each of 30 s duration) followed by reperfusion. In the HPoC group, after 6 h of hypoxia, H9c2 cells were subjected to three cycles of 10 minute reoxygenation and 10 minute hypoxia followed by reoxygenation. IPoC significantly reduced the infarct size, plasma level of Lactate dehydrogenase and creatine kinase MB in rats. 12 h after the reperfusion, MIP2 mRNA levels in the IPoC group were 10 folds that of the sham group and 1.4 folds that of the I/R group. Increased expression of MIP2 mRNA and attenuation of apoptosis were similarly observed in the HPoC group in the in vitro model. These effects were blunted by transfection with MIP2 siRNA in the H9c2 cells. This study demonstrated that IPoC induced protection was associated with increased expression of MIP2. Both MIP2 overexpression and MIP2 suppression can influence the IPoC induced protection.