SOX4 maintains the stemness of cancer cells via transcriptionally enhancing HDAC1 revealed by comparative proteomics study.

BACKGROUND: Cancer stem cells (CSCs) are the root of human cancer development and the major cause of treatment failure. Aberrant elevation of SOX4, a member of SOX (SRY-related HMG-box) family transcription factors, has been identified in many types of human cancer and promotes cancer development. However, the role of SOX4 in CSCs, especially at a proteome-wide level, has remained elusive. The aim of this study is to investigate the effect of SOX4 on the stemness of CSCs and reveal the underlying mechanisms by identification of SOX4-induced proteome changes through proteomics study. RESULTS: Overexpression of SOX4 promotes sphere formation and self-renewal of colorectal cancer cells in vitro and in vivo and elevates the expression levels of CSCs markers. Through iTRAQ-based quantitative proteomics analysis, 215 differentially expressed proteins (128 upregulated, 87 downregulated) in SOX4-overexpressing HCT-116 spheres were identified. The bioinformatic analysis highlighted the importance of HDAC1 as the fundamental roles of its impacted pathways in stem cell maintenance, including Wnt, Notch, cell cycle, and transcriptional misregulation in cancer. The mechanistic study showed that SOX4 directly binds to the promoter of HDAC1, promotes HDAC1 transcription, thereby supporting the stemness of colorectal cancer cells. HDAC1 hallmarks colorectal cancer stem cells and depletion of HDAC1 abolished the stimulatory effect of SOX4. Furthermore, SOX4-HDAC1 axis is conserved in multiple types of cancer. CONCLUSIONS: The results of this study reveal SOX4-induced proteome changes in HCT-116 spheres and demonstrates that transcriptional activation of HDAC1 is the primary mechanism underlying SOX4 maintaining CSCs. This finding suggests that HDAC1 is a potential drug target for eradicating SOX4-driven human CSCs.

Tea polyphenols and Levofloxacin alleviate the lung injury of hepatopulmonary syndrome in common bile duct ligation rats through Endotoxin -TNF signaling.

BACKGROUND & AIMS: Hepatopulmonary syndrome (HPS) is characterized by pulmonary vasodilation and arterial blood oxygen desaturation in patients with chronic liver disease. Generally, common bile duct ligation (CBDL) rats are a suitable experimental model for studying hepatopulmonary syndrome. Our previous study demonstrated that endotoxin surges markedly, followed by bacterial translocation and the loss of liver immune function in all the stages of CBDL, thereby contributing to the pathogenesis of HPS. However, the mechanisms behind the increase of the endotoxin and how to alleviate it have not yet been elucidated. Pulmonary injury induced by increased bilirubin, endotoxin, and inflammatory mediators occurs in the early and later stages of CBDL. This study assessed the effects of Tea polyphenols (TP) and Levofloxacin on endotoxin reduction and suppression of lung injury in HPS rats in the long and short term, respectively. METHODS: Morphological change of pulmonary injury, HPS relative index, endotoxin concentration, and the activation extent of Malondialdehyde (MDA) and Myeloperoxidase (MPO) were evaluated in CBDL rats with or without TP and Levofloxacin for three weeks or six weeks. The inflammation factors of serum, lung tissue, and BALF were then compared at the same condition for the two time periods. This was followed by adoption of the network pharmacology approach, which was mainly composed of active component gathering, target prediction, HPS gene collection, network analysis, and gene enrichment analysis. Finally, the mRNA and protein levels of the inflammatory factors were studied and relative signaling expression was assessed using RT-PCR and Western blot analysis. RESULTS: The obtained results indicated that the pulmonary injury manifestation was perceived and endotoxin, MDA, and MPO activation were markedly increased in the early and later stages of CBDL. TP and Levofloxacin treatment alleviated endotoxin infection and inflammation factor expression three weeks and six weeks after CBDL. In addition, Levofloxacin displayed a short time anti-bacterial effect, while TP exerted a long period function. TP and Levofloxacin also reduced TNF-α, TGF-ß, IL-1ß, PDGF-BB, NO, ICAM-1, and ET-1 expression on the mRNA or protein expression. With regard to the pharmacological mechanism, the network analysis indicated that 12 targets might be the therapeutic targets of TP and Levofloxacin on HPS, namely ET-1, NOs3, VEGFa, CCl2, TNF, Ptgs2, Hmox1, Alb, Ace, Cav1, and Mmp9. The gene enrichment analysis implied that TP and Levofloxacin probably benefited patients with HPS by modulating pathways associated with the AGE-RAGE signaling pathway, the TNF signaling pathway, the HIF-1 signaling pathway, the VEGF signaling pathway, and the IL-17 signaling pathway, Rheumatoid arthritis, Fluid shear stress, and atherosclerosis. Finally, the TNF-α level was mainly diminished on the protein level following CBDL. CONCLUSIONS: TP and Levofloxacin could alleviate pulmonary injury for short and long period, respectively, while at the same time preventing endotoxin and the development of HPS in CBDL rats. These effects are possibly associated with the regulation of the Endotoxin -TNF-α pathways.

SKP1 promotes YAP-mediated colorectal cancer stemness via suppressing RASSF1.

BACKGROUND: Cancer stem cells (CSCs) have been recognized as an important drug target, however, the underlying mechanisms have not been fully understood. SKP1 is a traditional drug target for cancer therapy, while, whether SKP1 promotes colorectal cancer (CRC) stem cells (CRC-SCs) and the underlying mechanisms have remained elusive. METHODS: Human CRC cell lines and primary human CRC cells were used in this study. Gene manipulation was performed by lentivirus system. The mRNA and protein levels of target genes were examined by qRT-PCR and western blot. The sphere-forming and in vitro migration capacities were determined by sphere formation and transwell assay. The self-renewal was determined by limiting dilution assay. The tumorigenicity and metastasis of cancer cells were examined by xenograft model. The promoter activity was examined by luciferase reporter assay. Nuclear run-on and Chromatin immunoprecipitation-PCR (ChIP-PCR) assay were employed to examine the transcription and protein-DNA interaction. Co-immunoprecipitation assay was used to test protein-protein interaction. The relationship between gene expression and survival was analyzed by Kaplan-meier analysis. The correlation between two genes was analyzed by Spearman analysis. Data are represented as mean ± SD and the significance was determined by Student's t test. RESULTS: SKP1 was upregulated in CRC-SCs and predicted poor prognosis of colon cancer patients. Overexpression of SKP1 promoted the stemness of CRC cells reflected by increased sphere-forming, migration and self-renewal capacities as well as the expression of CSCs markers. In contrast, SKP1 depletion produced the opposite effects. SKP1 strengthened YAP activity and knockdown of YAP abolished the effect of SKP1 on the stemness of CRC cells. SKP1 suppressed RASSF1 at both mRNA and protein level. Overexpression of RASSF1 abolished the effect of SKP1 on YAP activity and CRC stemness. CONCLUSION: Our results demonstrated that SKP1 suppresses RASSF1 at both mRNA and protein level, attenuates Hippo signaling, activates YAP, and thereby promoting the stemness of CRC cells.

A Novel Polysaccharide Depolymerase Encoded by the Phage SH-KP152226 Confers Specific Activity Against Multidrug-Resistant Klebsiella pneumoniae via Biofilm Degradation.

The increasing prevalence of infections caused by multidrug-resistant Klebsiella pneumoniae necessitates the development of alternative therapies. Here, we isolated, characterized, and sequenced a K. pneumoniae bacteriophage (SH-KP152226) that specifically infects and lyses K. pneumoniae capsular type K47. The phage SH-KP152226 contains a genome of 41,420 bp that encodes 48 predicted proteins. Among these proteins, Dep42, the gene product of ORF42, is a putative tail fiber protein and hypothetically possesses depolymerase activity. We demonstrated that recombinant Dep42 showed specific enzymatic activities in the depolymerization of the K47 capsule of K. pneumoniae and was able to significantly inhibit biofilm formation and/or degrade formed biofilms. We also showed that Dep42 could enhance polymyxin activity against K. pneumoniae biofilms when used in combination with antibiotics. These results suggest that combination of the identified novel depolymerase Dep42, encoded by the phage SH-KP152226, with antibiotics may represent a promising strategy to combat infections caused by drug-resistant and biofilm-forming K. pneumoniae.

PTPRU, As A Tumor Suppressor, Inhibits Cancer Stemness By Attenuating Hippo/YAP Signaling Pathway.

BACKGROUND: PTPRU is an important signaling molecule that regulates a variety of cellular processes; however, the role of PTPRU in cancer development has remained elusive. Here, we report that PTPRU serves as a tumor suppressor that inhibits cancer stemness by attenuating Hippo/YAP signaling pathway. METHODS: Primary cancer cells and cell line cells were used in the study. The gene expression data were downloaded from R2 analysis and visualization platform and Kaplan-Meier analysis was performed to study the relationship between survival and PTPRU expression. qRT-PCR and Western blot were employed to study the expression of target genes in tissues and cells. Sphere and colony formation, proliferation, migration activities and the expression of stem cell and EMT markers were employed for characterizing the stemness. Gene manipulation was achieved by lentivirus-mediated gene delivery system. Luciferase reporter gene assay was used to study the transcriptional activity of the promoter, and ChIP-qPCR was employed to study the target binding sequence of the protein. Spearman correlation analysis was performed to study the correlation between two genes. Student's t-test was used for determination of the significance between two experimental groups. RESULTS: PTPRU is downregulated in colorectal and gastric cancer tissues and cancer stem cells. High expression of PTPRU predicts poor prognosis. Overexpression of PTPRU attenuates the stemness of gastric cancer stem cells and knockdown of PTRPU improves the maintenance of the stemness of cancer stem cells. Mechanistic analysis showed that PTPRU inhibits Hippo/YAP signaling by suppressing the expression of YAP in a transcriptional level. Overexpression of YAP restored PTPRU-induced inhibited stemness of gastric cancer stem cells. CONCLUSION: PTPRU serves as a tumor suppressor that inhibits the stemness of cancer stem cell by inhibiting Hippo/YAP signaling pathway.

[Surgical management of locally advanced digestive tract neuroendocrine neoplasm with or without liver metastasis].

Neuroendocrine tumors in the digestive tract are rare, however their incidences increased obviously for the past few years. The purpose of this paper is to elucidate the surgical management of locally advanced digestive tract neuroendocrine neoplasms(NENs) with or without liver metastasis and to discuss the present classification of gastrointestinal NENs in order to provide reference for clinicians. WHO re-classified the gastroenteropancreatic NENs in 2010, but this classification remains many questions and needs further clinical trials to answer. Up to now, radical resection of the lesions is the only cure for the gastrointestinal NENs. For resectable locally advanced gastrointestinal NENs, standard radical or extended resection should be performed according to gastrointestinal cancer. For patients who can not receive radical procedure because of unresectable primary lesions or diffuse metastases, cytoreductive operation should be considered when endocrine symptoms exist. Palliative surgery is beneficial to the improvement of bleeding or obstruction by tumor. For unresectable liver metastatic lesion and resectable primary lesion, the primary lesion should be resected. For tolerable patients with resectable liver metastatic lesion, one-stage resection involving the primary and the liver metastatic lesions should be performed. For unresectable liver metastasis, hepatic arterial chemoembolization, systematic chemotherapy, biotherapy, targeted therapy or radio frequency ablation (RFA) should be considered to control symptoms and prevent the tumor progression.

MiR-146a suppresses hepatocellular carcinoma by downregulating TRAF6.

MicroRNAs have been proven to play important roles in many biological processes such as cellular growth and differentiation, apoptosis, and modulation of host response to viral infection. In the present study, we find that the expression of miR-146a was decreased in hepatocellular carcinoma (HCC) tissues compared with corresponding adjacent tissues, and the expression level in HCC cell lines was lower than in a normal liver cell. Over-expression suppressed the proliferation and invasion of HCC cells. In addition, luciferase reporter assays and western blotting confirmed that miR-146a directly target TRAF6 which attenuated the effect of miR-146a on cell proliferation and invasion in HepG2 and SMMC7721 cells. Meanwhile, lentivirus-mediated increased expression of miR146a repressed tumor formation in nude mice. Taken together, our findings demonstrate that miR-146a suppresses HCC by down-regulating TRAF6. We also discovered that miR-146a may represent a novel potential candidate of the HCC carcinoma diagnostic marker in the long term.

[Application of enhanced recovery after surgery for patients with laparoscopic radical gastrectomy].

OBJECTIVE: To investigate the clinical application effects of enhanced recovery after surgery (ERAS) during perioperative period on patients undergoing laparoscopic radical gastrectomy. METHODS: A total of 127 patients with gastric cancer receiving laparoscopic radical gastrectomy in our department from July 2013 to February 2015 were prospectively enrolled. Patients were randomly divided into ERAS group (n=67) treated with ERAS method and conventional group (n=60) treated with the traditional perioperative management. The perioperative efficacy and postoperative short-term outcomes were compared between the two groups. RESULTS: All the patients were recovery with no mortality during perioperative period. Compared with conventional group, patients in ERAS group showed shorter time of nasogastric tube placement[(1.4±0.6) days vs. (2.2±0.6) days, P=0.000], shorter time to first flatus [(3.2±0.9) days vs. (3.8±1.1) days, P=0.004], shorter time to first ambulation [(2.3±1.1) days vs. (4.2±6.4) days, P=0.026], shorter time to initiate oral intake (1.9±0.7) days vs. (4.0±1.1) days, P=0.000] and shorter postoperative hospital stay [(13.9±4.0) days vs. (18.7±9.1) days, P=0.000]. Visual pain scores were all significantly lower in the ERAS group as compared to the traditional group 2 hours after anesthesia awakening, 12, 24, 48 hours after operation and at the first ambulation respectively(all P<0.05). The overall complication rate was significantly lower in ERAS group than that in conventional group[14.9%(10/67) vs. 30.0%(18/60), P = 0.041]. Patients in the ERAS group had shorter postoperative hospital stay [(13.9±4.0) days vs. (18.7±9.1) days, P=0.000]. CONCLUSION: ERAS during perioperative period in laparoscopic radical gastrectomy is safe and effective, which is associated with quicker postoperative rehabilitation, better pain control and shorter hospital stay.

Safety and efficacy of laparoscopy-assisted gastrectomy for advanced gastric cancer in the elderly.

To evaluate safety and efficacy of laparoscopy-assisted radical gastrectomy (LARG) for advanced gastric cancer patients aged 70 years or older. Clinical data were retrospectively collected from patients with IIA-IIIC gastric cancer who underwent LARG (n = 30) and open radical gastrectomy (ORG, n = 34) in Department of Gastrointestinal Surgery in the Ningbo First Hospital from January 2012 to December 2013. The mean operative time was longer in the LARG group than in the ORG group but there was no statistical difference between the two groups. The intraoperative blood loss (120 ± 52.7 ml vs 227.3 ± 146.9 ml), incidence of postoperative complication (23.0% vs 47%) were lower in the LARG group than those in the ORG group. In addition, the time to first flatus (2.9 ± 0.8 d vs 4.6 ± 1.2 d), time to first ambulation (1.2 ± 0.4 vs 4.1 ± 1.0 d), time of nasogastric intubation (2.5 ± 1.0 d vs 3.5 ± 1.4 d), and postoperative hospital stay (13.0 ± 4.2 d vs 16.9 ± 4.1 d) were significantly shorter in the LARG group than in the ORG group, respectively. No statistical difference in the number of harvested lymph nodes was noted between the two groups (30.2 ± 12.0 vs 28.1 ± 11.8, P > 0.05). LARG is safer, more effective and less invasive for the elderly patients with advanced gastric cancer.

Knockdown of protein tyrosine phosphatase receptor U inhibits growth and motility of gastric cancer cells.

Protein tyrosine phosphatase receptor U (PTPRU) has been shown to be a tumor suppressor in colon cancer by dephosphorylating ß-catenin and reducing the activation of ß-catenin signaling. Here, we investigate the expression of PTPRU protein in gastric cancer cell lines, gastric cancer tissues and respective adjacent non-cancer tissues and find that the 130 kDa nuclear-localized PTPRU fragment is the main PTPRU isoform in gastric cancer cells, whereas the full-length PTPRU is relatively lowly expressed. The level of the 130 kDa PTPRU is higher in gastric cancer tissues than in adjacent non-cancer tissues. Knockdown of endogenous PTPRU in gastric cancer cells using lentivirus-delivered specific shRNA results in the attenuation of cell growth, migration, invasion and adhesion. Knockdown of PTPRU also inhibits tyrosine phosphorylation and transcriptional activity of ß-catenin as well as levels of focal adhesion proteins and lysine methylation of histone H3. These results indicate that PTPRU is required for gastric cancer progression and may serve as a potential therapeutic target.

High fever as an initial symptom of primary gastric inflammatory myofibroblastic tumor in an adult woman.

Inflammatory myofibroblastic tumor, also known as inflammatory pseudotumor, plasma cell granuloma or inflammatory myofibroblastoma, is characterized histopathologically by myofibroblastic spindle cells with inflammatory cell infiltrates composed of plasma cells, lymphocytes and eosinophils. Inflammatory myofibroblastic tumor is typically seen in children or young adults and is most commonly localized to the lungs, but it can occur anywhere in the body. To date, however, only a few cases involving the stomach have been reported. Herein, we present a case of gastric inflammatory myofibroblastic tumor in an adult woman with an initial symptom of high fever.

Adult Hirschsprung's disease: report of four cases.

Adult Hirschsprung's disease (HD) is a rare motor disorder of the gut that is frequently misdiagnosed as refractory constipation. The primary pathogenic defect in adult HD is identical to that seen in infancy or childhood, and is characterized by the total absence of intramural ganglion cells of the submucosal (Meissner) and myenteric (Auerbach) neural plexuses in the affected segment of the bowel. Ninety-four percent of HD cases are diagnosed before the patient reaches 5 years of age, however, on rare occasion, mild cases of HD may go undiagnosed until he or she reaches adulthood. In this study, we describe four cases of adult HD with a history of longstanding recurrent constipation, relieved by laxatives, and presenting to the Department of Gastrointestinal Surgery with progressive abdominal distention, colicky pain or acute intestinal obstruction. Barium enema or computed tomography revealed a grossly distended proximal large colon with fecal retention. Intraoperative frozen section biopsy was performed in all cases and showed aganglionosis of the stenotic segment and a normal distal rectum. In all cases, patient symptoms were completely resolved and there were no complications arising immediately post-surgery or at one-year follow-up. Adult HD should be considered in the differential diagnosis of cases where adult patients present with chronic constipation or even acute intestinal obstruction. The modified one-stage Martin-Duhamel or Rehbein's procedure is a feasible surgical option for treating cases of adult HD involving a segment or the entire bowel.

Lentivirus-mediated RNAi knockdown of VEGFA in RKO colorectal cancer cells decreases tumor formation and growth in vitro and in vivo.

Vascular endothelial growth factors (VEGF) play important roles in angiogenesis, vasculogenesis and endothelial cell growth. In endothelial cancers, secreted VEGF proteins induce endothelial cell proliferation, promote cell migration, inhibit apoptosis and induce blood vessel permeabilization. VEGFA is frequently overexpressed in human colorectal cancers (CRC) and its expression correlates with tumor progression and invasiveness. In this study we examine the effect of knocking down VEGFA expression by infecting RKO colorectal cancer cells with lentiviral particles containing VEGFA-targeting RNAi constructs. We found that suppressing VEGFA dramatically decreased RKO cell proliferation, colony formation, invasion, migration and tumor growth. Furthermore, VEGFA knock-down reduced MAPK pathway signaling and Smac/DIABLO expression. These results suggest that lentivirus-mediated RNAi knock-down of VEGFA could be an effective therapy for the treatment of CRC.

[RNA Interference targeting GRP75 decreases cisplatin resistance in human lung adenocarcinoma cell].

BACKGROUND AND OBJECTIVE: GRP75, a member of HSPs which is overexpressed in some resistant cancer cells, is a molecular chaperone mainly located in mitochondrial membrane. The aim of this study is to investigate the role of GRP75 on the resistant mechanisms of cancer cells by downregulating GRP75 expreesion via RNAi approach. METHODS: Cisplatin-resistant cell A549/CDDP was established from their parental human lung adenocarcinoma cell line A549 by combining gradually increasing concentrations of cisplatin with high dosage impact. The shRNA for GRP75 was transfected into A549 and A549/CDDP cells by lentivirus. Western blot and methyl thiazolyl tetrazolium (MTT) assay were applied to detect the influence of silencing GRP75 expression on sensitivity of the cells to cisplatin. RESULTS: The infection rate of six groups were all over 90%. After infection, the level of expression of GRP75 in both A549 and A549/CDDP were down-regulated (P < 0.05); the level of expression of p53 in A549/CDDP was up-regulated (P < 0.05) and the level of expression of bcl-2 of A549/CDDP was down-regulated (P < 0.05). The resistance index of A549/CDDP before and after infection were 21.52 and 4.14 respectively. CONCLUSIONS: Cisplatin resistance of lung cancer cells is associated with overexpression of GRP75 gene, which could regulate the expressions of p53 and bcl-2.

[Diagnosis, treatment, and prognosis of primary appendiceal tumors: analysis of 37 cases].

OBJECTIVE: To investigate the pathology, diagnosis,treatment, and prognosis of primary appendiceal tumors. METHODS: The clinical data of 37 patients with primary tumors of the appendix, 16 males and 21 females, aged 56 +/- 13 (35-87) hospitalized Jan. 1977 to Feb. 2007 were analyzed retrospectively. RESULTS: Appendicitis and abdominal mass were the major clinical manifestations. All 37 cases received surgical operation with the diagnosis confirmed by pathology. The pathological types included carcinoid tumor (n=12), mucinous tumor (n=17), and adenocarcinoma (n=8). Only 4 cases were diagnosed pathologically by biopsy before operation, 28 cases were diagnosed by intra-operative frozen section, and 5 cases were diagnosed after operation. Single appendectomy were performed in 17 cases, ileocecalectomy in 9 cases, right hemicolectomy in 8 cases, and other operation patterns in 3 cases. The 1, 3, and 5-year survival rates of the primary appendix carcinoid tumor, mucinous tumor, and adenocarcinoma were 100.0%, 100.0%, and 91.7%, 100.0%, 86.7%, and 71.5%, and 75.0%, 50.0%, and 50.0% respectively. CONCLUSION: A rare disease, appendiceal tumors lack specific clinical features. Intra-operative exploration and frozen section are very important for diagnosis and operation choice. The prognosis of primary appendix carcinoid tumors is better.

Liposome-mediated gene transfer of endothelial nitric oxide synthase to cirrhotic rat liver decreases intrahepatic vascular resistance.

BACKGROUND AND AIMS: Nitric oxide (NO) production by endothelial nitric oxide synthase (eNOS) in sinusoidal endothelial cells is reduced in the injured liver and leads to intrahepatic portal hypertension. The present study evaluates the effects of liposome-mediated gene transfer of eNOS on the intrahepatic vascular resistance and portal venous pressure (PVP) in cirrhotic rats. METHODS: Hepatic cirrhosis was induced in male Sprague-Dawley rats by intraperitoneal injection of carbon tetrachloride (CCl(4)), whereas the control normal rats were given the same dose of peanut oil. Plasmid eukaryotic expression vector (liposome-pcDNA3/eNOS) was injected into the portal vein of CCl(4) cirrhotic rats, whereas cirrhotic controls received the same dose of naked plasmid (liposome-pcDNA3) or Tris buffer, and control normal rats received the same dose of Tris buffer. Five days after gene transfer, the levels of eNOS mRNA and protein, NO production, PVP and the changes of hepatic intrahepatic vascular resistance were investigated. RESULTS: Five days after eNOS gene transfer, the levels of eNOS mRNA, eNOS protein and NO production in cirrhotic rats increased remarkably, while hepatic vascular resistance and PVP decreased significantly in cirrhotic rats. CONCLUSION: Liposome-mediated eNOS gene transfer via intraportal injection is feasible and the increase of intrahepatic eNOS leads to a marked decrease in introhepatic vascular resistance and PVP. These data indicate that intrahepatic eNOS plays an important role in the pathogenesis of portal hypertension and gene transfer of eNOS is a potential and novel therapy for portal hypertension.

Cystamine ameliorates liver fibrosis induced by carbon tetrachloride via inhibition of tissue transglutaminase.

AIM: To investigate the anti-fibrosis effect of the tissue transglutaminase (tTG) specific inhibitor cystamine on liver fibrosis. METHODS: Sixty-eight male Sprague Dawley rats were divided into three groups: normal control, liver fibrosis control and cystamine-treated group. Liver fibrosis was induced by intraperitoneal injection of carbon tetrachloride (CCl(4)), and Cystamine was administrated by intraperitoneal injection starting 2 d before the first administration of CCl(4). Animals in each group were further divided into 2 subgroups according to two time points of 4 wk and 8 wk after treatment. Hepatic function, pathological evaluation (semi-quantitative scoring system, SSS) and liver hydroxyproline (Hyp) content were examined. Real-time PCR was used to detect the expression of tTG, smooth muscle alpha actin (alpha-SMA), tissue inhibitor of metalloproteinase 1 (TIMP-1) and collagen-1 mRNA. The expressions of tTG and alpha-SMA protein were detected by Western Blotting. RESULTS: Eight weeks after treatment, the SSS score of liver was significantly less in the cystamine group than that in the fibrosis control group (P < 0.01). The levels of alanine aminotransferase (ALT) and total bile acid (TBA) at the 4 wk and 8 wk time points were decreased in the cystamine group compared with those in fibrosis controls (P < 0.01). Liver hydroxyproline content at the 4 wk and 8 wk time points showed a substantial reduction in the cystamine group compared to fibrosis controls (P < 0.01). The expression of tTG, alpha-SMA, collagen-1, TIMP-1 mRNA and tTG, as well as alpha-SMA protein was downregulated in the cystamine group compared to fibrosis controls. CONCLUSION: Cystamine can ameliorate CCl(4) induced liver fibrosis and protect hepatic function. The possible mechanism is related to the reduced synthesis of the extracellular matrix (ECM) caused by the inhibition of hepatic stellate cell activation and decreased expression of TIMP-1.

[Protective effects of vasoactive intestinal peptide on septic shock rats].

OBJECTIVE: To investigate the protective effects of vasoactive intestinal peptide (VIP) on septic shock rats and explore its possible mechanism. METHODS: Cecal ligation and puncture (CLP) was performed to produce septic shock rat model. Thirty adult Sprague-dawley rats were randomly divided into 3 groups with 10 animals in each group: sham operation group, CLP group and VIP group. The rats in VIP group were given intravenous injection of VIP (5 nmol per rat) instantly after the CLP operation. Then the mean arterial pressure (MAP) was monitored consistently and survival rate was observed. Blood samples were obtained from femoral artery for measuring the serum concentrations of tumor necrosis factor-alpha (TNF-alpha) and interleukin-10 (IL-10) by using enzyme-linked immunosorbent assays (ELISA). Organs (lung, kidney and intestine) were harvested for pathological examination. RESULTS: At each time point after 8 h, the MAP of VIP rats was significantly higher than that in CLP rats (P<0.05). In VIP group rats, the serum TNF-alpha concentration was decreased meanwhile IL-10 level was increased with markedly alleviated organic pathological injuries and the survival rate was obviously raised. CONCLUSIONS: VIP exerts protective effects on septic shock rats through inhibiting production of proinflammatory factors and stimulating the production of anti-inflammatory cytokines.

Influence of nitric oxide synthase and cyclooxygenase blockade on expression of cyclooxygenase and hemodynamics in rats with portal hypertension.

BACKGROUND: The importance of nitric oxide (NO) in the pathogenesis of portal hypertension (PHT) has been extensively studied, but whether or not prostacyclin (PGI(2)) plays a role in formation and development of hyperdynamic circutatory state in PHT has not been verified. The present study was undertaken to investigate the possible interaction between prostacyclin (PGI(2)) and nitric oxide (NO) in the hyperdynamic circulatory state of rats with chronic portal hypertension (PHT), by measuring the hemodynamic changes and expression of cyclooxygenase (COX) mRNA in vessels and small intestine after administration of N(omega)-nitro-L-arginine (L-NNA) or indomethacin (INDO) either in the short-term (7 days) or long-term (15 days). METHODS: Ninety-seven male Sprague-Dawley rats were divided into three groups: intrahepatic portal hypertension (IHPH) induced by injection of CCl(4), prehepatic portal hypertension (PHPH) induced by partial stenosis of the portal vein, and sham-operated controls (SO). Animals of each group received L-NNA or INDO either for 7 or 15 days, with saline as control. Splanchnic hemodynamics was measured by the radioactive microsphere technique. The concentration of NO in serum was determined as the nitrate; nitrite ratio (NO(2)(-)/NO(3)(-), micromol/L) by a colorometric method, and that of PGI(2) was measured by specific radioimmunoassay for its stable hydrolysis product 6-keto-PGF(1alpha) (pg/ml). The reverse transcription-polymerase chain reaction measured the levels of COX-1 mRNA in the superior mesenteric artery, thoracic aorta, and small intestine of these rats. RESULTS: Compared with SO rats, COX-1 mRNA expression and the concentrations of plasma 6-keto-PGF(1alpha) and serum NO(2)(-)/NO(3)(-) were enhanced in both IHPH and PHPH rats; splanchnic vascular resistance (SVR) decreased, but portal venous inflow (PVI) markedly increased (P<0.05). Seven or 15 days of L-NNA treatment reduced COX-1 mRNA expression in these vessels and the small intestine, concomitant with a significant decrease in the concentration of plasma PGI(2) and serum NO in IHPH and PHPH rats (P<0.05). At the same time, PVI decreased but SVR increased significantly (P<0.05). In both IHPH and PHPH rats, the COX-1 mRNA expression and the concentration of plasma PGI(2) after No synthase (NOS) blockade for 15 days were higher than those for 7 days, whereas the hyperdynamic circulatory state was improved after NOS blockade for 15 days compared with 7 days. The concentration of PGI(2) treated by INDO for 15 days was not significantly different from that after 7-day COX blockade, and hemodynamics restored hyperdynamic circulatory state. CONCLUSIONS: The hyperdynamic circulatory state in rats with PHT is correlated with the concentration of serum NO. There is a possible interaction between PGI(2) and NO in the hyperhemodynamics of PHT. PGI(2) is probably not the mediator in the formation and development of the hyperdynamic circulatory state in rats with chronic PHT.