p38 MAPK inhibition alleviates experimental acute pancreatitis in mice.

BACKGROUND: The mitogen-activated protein kinases (MAPKs) signaling pathway is involved in inflammatory process. However, the mechanism is not clear. The present study was to investigate the role of p38 MAPK in acute pancreatitis in mice. METHODS: Mice were divided into 4 groups: saline control; acute pancreatitis induced with repeated injections of cerulein; control plus p38 MAPK inhibitor SB203580; and acute pancreatitis plus SB203580. The pancreatic histology, pancreatic enzymes, cytokines, myeloperoxidase activity, p38 MAPK and heat shock protein (HSP) 60 and 70 were evaluated. RESULTS: Repeated injections of cerulein resulted in acute pancreatitis in mice, accompanying with the activation of p38 MAPK and overexpression of HSP60 and HSP70 in the pancreatic tissues. Treatment with SB203580 significantly inhibited the activation of p38 MAPK, and furthermore, inhibited the expression of HSP60 and HSP70 in the pancreas, the inflammatory cytokines in the serum, and myeloperoxidase activity in the lung. CONCLUSION: The p38 MAPK signaling pathway is involved in the regulation of inflammatory response and the expression of HSP60 and HSP70 in acute pancreatitis.

The role of the CXCR6/CXCL16 axis in the pathogenesis of fibrotic disease.

CXC chemokine receptor 6 (CXCR6), a seven-transmembrane domain G-protein-coupled receptor, plays a pivotal regulatory role in inflammation and tissue damage through its interaction with CXC chemokine ligand 16 (CXCL16). This axis is implicated in the pathogenesis of various fibrotic diseases and correlates with clinical parameters that indicate disease severity, activity, and prognosis in organ fibrosis, including afflictions of the liver, kidney, lung, cardiovascular system, skin, and intestines. Soluble CXCL16 (sCXCL16) serves as a chemokine, facilitating the migration and recruitment of CXCR6-expressing cells, while membrane-bound CXCL16 (mCXCL16) functions as a transmembrane protein with adhesion properties, facilitating intercellular interactions by binding to CXCR6. The CXCR6/CXCL16 axis is established to regulate the cycle of damage and repair during chronic inflammation, either through modulating immune cell-mediated intercellular communication or by independently influencing fibroblast homing, proliferation, and activation, with each pathway potentially culminating in the onset and progression of fibrotic diseases. However, clinically exploiting the targeting of the CXCR6/CXCL16 axis requires further elucidation of the intricate chemokine interactions within fibrosis pathogenesis. This review explores the biology of CXCR6/CXCL16, its multifaceted effects contributing to fibrosis in various organs, and the prospective clinical implications of these insights.

Predicting short-term major postoperative complications in intestinal resection for Crohn's disease: A machine learning-based study.

BACKGROUND: Due to the complexity and numerous comorbidities associated with Crohn's disease (CD), the incidence of postoperative complications is high, significantly impacting the recovery and prognosis of patients. Consequently, additional studies are required to precisely predict short-term major complications following intestinal resection (IR), aiding surgical decision-making and optimizing patient care. AIM: To construct novel models based on machine learning (ML) to predict short-term major postoperative complications in patients with CD following IR. METHODS: A retrospective analysis was performed on clinical data derived from a patient cohort that underwent IR for CD from January 2017 to December 2022. The study participants were randomly allocated to either a training cohort or a validation cohort. The logistic regression and random forest (RF) were applied to construct models in the training cohort, with model discrimination evaluated using the area under the curves (AUC). The validation cohort assessed the performance of the constructed models. RESULTS: Out of the 259 patients encompassed in the study, 5.0% encountered major postoperative complications (Clavien-Dindo ≥ III) within 30 d following IR for CD. The AUC for the logistic model was 0.916, significantly lower than the AUC of 0.965 for the RF model. The logistic model incorporated a preoperative CD activity index (CDAI) of ≥ 220, a diminished preoperative serum albumin level, conversion to laparotomy surgery, and an extended operation time. A nomogram for the logistic model was plotted. Except for the surgical approach, the other three variables ranked among the top four important variables in the novel ML model. CONCLUSION: Both the nomogram and RF exhibited good performance in predicting short-term major postoperative complications in patients with CD, with the RF model showing more superiority. A preoperative CDAI of ≥ 220, a diminished preoperative serum albumin level, and an extended operation time might be the most crucial variables. The findings of this study can assist clinicians in identifying patients at a higher risk for complications and offering personalized perioperative management to enhance patient outcomes.

Cytokines and irritable bowel syndrome: where do we stand?

Irritable bowel syndrome (IBS) is a functional gastrointestinal disorder, which presents with one or more gastrointestinal symptoms without any structural or organic abnormality. The etiology and pathophysiological mechanisms of IBS remain uncertain. Residual or reactivated inflammation at the molecular level is considered the underlying mechanism of post-infectious IBS. On the other hand, genetic variations in the immunological components of the body, including cytokine gene polymorphisms, are proposed as a potential mechanism of IBS even in patients without previous gastrointestinal infection. Several studies have suggested imbalanced cytokine signaling as an etiology for IBS. In this review, recent findings on cytokine profiles and cytokine gene polymorphisms in patients with IBS are described and the role of cytokines in animal models of IBS is discussed.

Timing of individualized surgical intervention in Crohn's disease.

Crohn's disease (CD) is a chronic inflammatory disorder of the gastrointestinal tract with an increasing incidence worldwide. Comprehensive therapy for CD focuses on symptom control and healing the intestinal mucosa to improve the quality of life and prevent complications. Surgical intervention plays a vital role in comprehensive therapy. However, deciding the optimal timing for surgical intervention has long been a focus of controversy. This review provides insights into the timing of surgery for CD and guides clinicians in daily treatment.

Laparoscopy for Crohn's disease: A comprehensive exploration of minimally invasive surgical techniques.

BACKGROUND: Along with the unceasing progress of medicine, Crohn's disease (CD), especially complex CD, is no longer a taboo for minimally invasive surgery. However, considering its special disease characteristics, more clinical trials are needed to confirm the safety and feasibility of laparoscopic surgery for CD. AIM: To investigate the safety and feasibility of laparoscopic enterectomy for CD, assess the advantages of laparoscopy over laparotomy in patients with CD, and discuss comprehensive minimally invasive surgical techniques in complex CD. METHODS: This study prospectively collected clinical data from patients with CD who underwent enterectomy from January 2017 to January 2020. It was registered in the Chinese clinical trial database with the registration number ChiCTR-INR-16009321. Patients were divided into a laparoscopy group and a traditional laparotomy group according to the surgical method. The baseline characteristics, operation time, intraoperative blood loss, temporary stoma, levels of abdominal adhesion, pathological characteristics, days to flatus and soft diet, postoperative complications, hospitalization time, readmission rate within 30 d, and hospitalization cost were compared between the two groups. RESULTS: A total of 120 eligible patients were enrolled into the pre-standardized groups, including 100 in the laparoscopy group and 20 in the laparotomy group. Compared with the laparotomy group, the postoperative hospitalization time in the laparoscopy group was shorter (9.1 ± 3.9 d vs 11.0 ± 1.6 d, P < 0.05), the days to flatus were fewer (2.8 ± 0.8 d vs 3.5 ± 0.7 d, P < 0.05), the days to soft diet were fewer (4.2 ± 2.4 d vs 6.2 ± 2.0 d, P < 0.05) and the intraoperative blood loss was less (103.3 ± 80.42 mL vs 169.5 ± 100.42 mL, P < 0.05). There were no statistically significant differences between the two groups in preoperative clinical data, operation time (149.0 ± 43.8 min vs 159.2 ± 40.0 min), stoma rate, levels of abdominal adhesion, total cost of hospitalization, incidence of postoperative complications [8.0% (8/100) vs 15.0% (3/20)], or readmission rate within 30 days [1.0% (1/100) vs 0.00 (0/20)]. CONCLUSION: Compared with laparotomy, laparoscopic enterectomy promotes the recovery of gastrointestinal function, shortens the postoperative hospitalization time, and does not increase the incidence of postoperative complications. Laparoscopic enterectomy combined with varieties of minimally invasive surgical techniques is a safe and acceptable therapeutic method for CD patients with enteric fistulas.

Minimally invasive surgery for glycogen storage disease combined with inflammatory bowel disease: A case report.

BACKGROUND: Inflammatory bowel disease (IBD) is rare in patients with glycogen storage disease (GSD). In GSD patients, a decrease in the number of neutrophils leads to prolonged intestinal infection, leading to the formation of chronic inflammation and eventually the development of IBD. Minimally invasive surgery for patients with IBD has been proven to reduce inflammatory responses and postoperative risks and ultimately promote rapid recovery. Herein we discuss minimally invasive surgery and the perioperative management in a patient with GSD and IBD. CASE SUMMARY: A 23-year-old male had GSD Ib associated with IBD-like disease for 10 years. Despite standard treatments, such as mesalazine, prednisone and adalimumab, the patient eventually developed colonic stenosis with incomplete ileus. After adequate assessment, the patient was treated with minimally invasive surgery and discharged in stable condition. CONCLUSION: Minimally invasive surgery for patients with IBD and GSD is safe, feasible and effective.

Laparoscopic vs open surgery in ileostomy reversal in Crohn's disease: A retrospective study.

BACKGROUND: Although minimally invasive surgery is becoming more commonly applied for ileostomy reversal (IR), there have been relatively few studies of IR for patients with Crohn's disease (CD). It is therefore important to evaluate the potential benefits and risks of laparoscopy for patients with CD. AIM: To compare the safety, feasibility, and short-term and long-term outcomes of laparoscopic IR (LIR) vs open IR (OIR) for the treatment of CD. METHODS: The baseline characteristics, operative data, and short-term (30-d) and long-term outcomes of patients with CD who underwent LIR and OIR at our institution between January 2017 and January 2020 were retrieved from an electronic database and retrospectively reviewed. RESULTS: Of the 60 patients enrolled in this study, LIR was performed for 48 and OIR for 12. There were no statistically significant differences in baseline characteristics, operation time, intraoperative blood loss, days to flatus and soft diet, postoperative complications, hospitalization time, readmission rate within 30 d, length of hospitalization, hospitalization costs, or reoperation rate after IR between the two groups. However, patients in the LIR group more frequently required lysis of adhesions as compared to those in the OIR group (87.5% vs 41.7%, respectively, P < 0.05). Notably, following exclusion of patients who underwent enterectomy plus IR, OIR was more advantageous in terms of postoperative recovery of gastrointestinal function and hospitalization costs. CONCLUSION: The safety and feasibility of LIR for the treatment of CD are comparable to those of OIR with no increase in intraoperative or postoperative complications.

Mast Cell Tryptase Promotes Inflammatory Bowel Disease-Induced Intestinal Fibrosis.

BACKGROUND: Intestinal fibrosis is the final pathological outcome of chronic intestinal inflammation without specific therapeutic drugs, which leads to ileus and surgical intervention. Intestinal fibrosis is characterized by excessive deposition of extracellular matrix (ECM). The role of mast cells (MCs), which are members of the sentinel immune cell population, is unknown in intestinal fibrosis. METHODS: In this study, we analyzed changes in MCs, tryptase proteins, and ECM components in human fibrotic and control patient intestines. We constructed dextran sodium sulfate-induced intestinal fibrosis models using wild-type mice, MC-reconstituted mice, and MC-deficient mice to explore the role of MCs and tryptase in intestinal fibrosis. The roles and mechanisms of MCs and tryptase on fibroblasts were evaluated using human MCs (HMC-1 and LAD-2), commercial tryptase proteins, human colon fibroblasts (CCD-18Co fibroblasts), the tryptase inhibitor APC366, and the protease-activated receptor-2 (PAR-2) antagonist ENMD-1068. RESULTS: Regardless of whether the colon was a human colon or a mouse colon, the fibrotic intestinal tissue had increased MC infiltration and a higher expression of ECM proteins or genes than that of the control group. The dextran sodium sulfate-induced intestinal fibrosis in MC-deficient mice was alleviated compared with that in wild-type mice. After MC reconstruction in MC-deficient mice, the alleviating effect disappeared. Tryptase, as a content stored in MC granules, was released into fibrotic intestinal tissues in the form of degranulation, resulting in an increased expression of tryptase. Compared with the control group, the tryptase inhibition group (the APC366 group) had reduced intestinal fibrosis. The CCD-18Co fibroblasts, when cocultured with MCs or treated with tryptase proteins, were activated to differentiate into myofibroblasts and secrete more ECM proteins (such as collagen and fibronectin). The underlying mechanism of fibroblast activation by tryptase was the activation of the PAR-2/Akt/mTOR pathway. CONCLUSIONS: We found that MC tryptase promotes inflammatory bowel disease-induced intestinal fibrosis. The underlying mechanism is that tryptase promotes the differentiation of fibroblasts into fibrotic-phenotype myofibroblasts by activating the PAR-2/Akt/ mTOR pathway of fibroblasts.

Norcantharidin inhibits growth of human gallbladder carcinoma xenografted tumors in nude mice by inducing apoptosis and blocking the cell cycle in vivo.

BACKGROUND: Gallbladder carcinoma, a lethal malignant neoplasm with poor prognosis, has dismal results of surgical resection and chemoradiotherapy. We previously reported that norcantharidin (NCTD) is useful against growth, proliferation, and invasion of human gallbladder carcinoma GBC-SD cells in vitro. In this study, we further studied the inhibitory effect of NCTD on the growth of xenografted tumors of human gallbladder carcinoma in nude mice in vivo and the underlying mechanisms. METHODS: The tumor xenograft model of human gallbladder carcinoma in nude mice in vivo was established with subcutaneous GBC-SD cells. The experimental mice were randomly divided into control, 5-FU, NCTD, and NCTD+5-FU groups which were given different treatments. Tumor growth in terms of size, growth curve, and inhibitory rate was evaluated. Cell cycle, apoptosis, and morphological changes of the xenografted tumors were assessed by flow cytometry and light/electron microscopy. The expression of the cell cycle-related proteins cyclin-D1 and p27 as well as the apoptosis-related proteins Bcl-2, Bax, and survivin were determined by the streptavidin-biotin complex (SABC) method and RT-PCR. RESULTS: NCTD inhibited the growth of the xenografted tumors in a dose- and time-dependent manner. Tumor volume decreased (5.61+/-0.39 vs. 9.78+/-0.61 cm3, P=0.000) with an increased tumor inhibitory rate (42.63% vs. 0%, P=0.012) in the NTCD group compared with the control group. The apoptosis rate increased (15.08+/-1.49% vs. 5.49+/-0.59%, P=0.0001) along with a decreased percentage of cells in S phase (43.47+/-2.83% vs. 69.85+/-1.96%, P=0.0001) in the NTCD group compared with the control group. The morphological changes of apoptosis such as nuclear shrinkage, chromatin aggregation, chromosome condensation, and typical apoptosis bodies in the xenografted tumor cells induced by NCTD were observed by light and electron microscopy. The expression of cyclin-D1, Bcl-2 and survivin proteins/mRNAs decreased significantly, with increased expression of p27 and Bax proteins/mRNAs in the NCTD group compared with the control group. CONCLUSION: NCTD inhibits the growth of xenografted tumors of human gallbladder carcinoma in nude mice by inducing apoptosis and blocking the cell cycle in vivo.

Prognostic factors of retroperitoneal soft tissue sarcomas: analysis of 132 cases.

BACKGROUND: Retroperitoneal sarcoma is a rare disease with poor prognosis. The aim of this study was to investigate the prognostic factors of the disease. METHODS: Between January 1988 and December 2003, 132 patients with retroperitoneal sarcoma were surgically treated in our hospital, 79 of them were followed up for 1 - 122 months (median: 19 months). Their clinicopathological data including tumor size, histological subtype, grade and resection margin status, were studied. The Kaplan-Meier method and log-rank test were used to analyze the disease-specific survival rates after the resection. RESULTS: Among the 132 patients, 98 (74.2%) received macroscopic complete resection, 29 (22.0%) incomplete resection, and 5 (3.8%) surgical biopsy. In the 79 patients who were followed up, macroscopic clear resection of retroperitoneal sarcoma (n = 49) was associated with a significantly higher survival rate compared with unclear resection (n = 30, P < 0.001). The median survival period was 31 months (95% CI, 20.09 - 41.91; actuarial 1-year survival, 85.7%) in the patients with the tumor completely resected and 11 months (95% CI, 6.71 - 15.29; actuarial 1-year survival, 46.7%) in those with incomplete resection. Patients with high-grade sarcomas had a significantly shorter survival time (n = 39; median: 24, 95% CI: 5.71 - 42.29) than those with low-grade sarcomas (n = 40; median: 15; 95% CI: 8.80 - 21.20; P < 0.01). Moreover, compared with the patients with the tumor sized < 15 cm in diameter (n = 53), the survival rate was lower in those with a sarcoma sized > 15 cm (n = 26). (Median: 12, 95% CI: 8.26 - 15.74 vs median: 24, 95% CI: 17.25 - 30.75; P < 0.05). Furthermore, the survival of the patients with liposarcomas (n = 29, median: 29, 95% CI: 12.84 - 45.16), leiomyosarcomas (n = 14, median: 11, 95% CI: 6.11 - 15.89), and others (n = 36, median: 22, 95% CI: 14.95 - 29.05) varied significantly (P < 0.05). CONCLUSION: Completeness of resection, tumor volume, grade, and subtype are prognostic factors of retroperitoneal soft tissue sarcomas.

Inhibitory effect of norcantharidin on the growth of human gallbladder carcinoma GBC-SD cells in vitro.

BACKGROUND: Gallbladder carcinoma is a lethal malignant neoplasm with dismal surgical results. Unfortunately, the adjuvant therapies for gallbladder carcinoma such as chemotherapy and radiotherapy are also disappointing. We reported that norcantharidin (NCTD), a demethylated form of cantharidin, which is an active ingredient of the Chinese medicine Mylabris, was used against human gallbladder carcinoma GBC-SD cells. In the present study, we further studied the mechanism underlying the inhibitory effect of NCTD on growth of human gallbladder carcinoma GBC-SD cells in vitro. METHODS: Human gallbladder carcinoma GBC-SD cells were grown in cell culture and divided into a NCTD group and a control group. The inhibitory effect of NCTD on growth of GBC-SD cells was investigated by evaluation of proliferation, cell cycle, apoptosis and morphological changes of the cells. Cell proliferation was assessed by tetrazolium-based colorimetric assay. The induction of cell cycle arrest and apoptosis was measured by flow cytometry. The morphological changes of the cells were observed by light- and electron-microscopy. To elucidate the anticancer mechanism of NCTD, expression of the proliferation-related gene proteins PCNA, Ki-67, cyclin-D1 and p27 and the apoptosis-related gene proteins Bcl-2, Bax and Survivin were determined by the streptavidin-biotin complex method and RT-PCR. RESULTS: NCTD inhibited the proliferation of GBC-SD cells in a dose- and time-dependent manner, with an IC50 of 56.18 microg/ml at 48 hours. The flow cytometric profiles revealed that NCTD (at the IC50 for 48 hours) significantly increased the proportion of cells in G2/M phase and significantly decreased the proportion of cells in S phase, with a significantly increased rate of cell apoptosis. After treatment with the 48-hour IC50 dose of NCTD, cell shrinkage, vacuolar cytoplasm, membrane budding, karyorrhexis, karyolysis, chromosome condensation and chromatin aggregation in some GBC-SD cells were observed by light-microscopy; decreased microvilli, Golgiosome atrophy, mitochondrial swelling, nuclear shrinkage, chromosome condensation and typical apoptosis bodies were seen by electron-microscopy, and the morphological changes of apoptosis occurred in GBC-SD cells. The expression of PCNA, Ki-67 and Bcl-2 proteins decreased significantly; the Pix or relative levels of PCNA mRNA, cyclin-D1 mRNA, Bcl-2 mRNA and Survivin mRNA decreased significantly, whereas the Pix or relative levels of p27 mRNA and Bax mRNA increased significantly. CONCLUSIONS: NCTD inhibits the growth of human gallbladder carcinoma GBC-SD cells in vitro. Its anticancer mechanism may correlate with inhibition of cell proliferation, arrest of the cell cycle, blockage of DNA synthesis, influence on cell metabolism, induction of cell apoptosis and influence on expression of the proliferation-related genes PCNA, Ki-67, cyclin-D1 and p27, and the apoptosis-related genes Bcl-2, Bax and Survivin in human gallbladder carcinoma GBC-SD cells.

[Primary diffuse large B-cell non-Hodgkin's lymphoma of the small intestine: clinicopathologic features, management, and prognosis in 24 patients].

OBJECTIVE: To investigate the clinicopathological features of primary diffuse large B-cell lymphomas (DLBCLs) of the small intestine, CD10 expression, and their relationship to prognosis. METHODS: Twenty-four cases of small intestinal DLBCLs were studied clinically and pathologically. All cases were staged according to the Ann Arbor classification of lymphoma. RESULTS: Fifteen cases (62.5%) were at stages I and II, and nine cases (37.5%) at stages III and IV. The Karnofsky performance status ranged from 40% to 100% (mean 75.5%). Twenty cases (83.3%) received surgical resection, sixteen cases (66.7%) received chemotherapy, and no patient received radiotherapy. Seven of 19 cases (36.8%) were CD10+. Although there was no statistically significant difference(P = 0.28) in therapy result between the CD10+ and CDO1--groups, patients with CD10+ lymphoma more frequently presented with stages I compared with those with CD10 - lymphoma (P = 0.013). Follow-up information was available in 19 cases ranging from 1 to 111 months (mean 32.7 months). Five cases died of the disease. The mortality rate was 26.3%. The analysis of survival rate showed a longer overall survival duration in the stage I and II group compared with that of the stage III and IV group ( P = 0.0197 ) , but there was no significant difference between CD10+ and CD1- groups. CONCLUSION: The primary small intestnal diffuse large B cell lymphoma patients at stage I and II respond better to therapy including surgical resection and chemotherapy than those at stage III and IV. CD10+ expression is more common in stage I lymphomas.

[Research of rat small intestinal mesentery lymphoid tissue stimulating allograft mixed lymphocyte reaction].

OBJECTIVE: To evaluate the effect of the small intestinal mesenteric lymphoid tissues stimulating mixed lymphocyte reaction with dendritic cells (DC) and peripheral blood monocyte cells (PBMC), and observe the changes of the MHC molecular expression on DC. METHODS: DC, PBMC and mixed lymphocyte were separated to culture from SD rats. Lymphoid tissue suspension was adopted from small intestinal mesentery of Wistar rats. In the mixed lymphocyte reaction (MLR), the cellular proliferation of small intestinal mesenteric lymphoid tissue antigen act on DC and PBMC was detected with cell counting of CCK-8 assay, the same assay used in small intestinal mesenteric lymphoid tissue antigen and ovalbumin (OVA) acting on DC. FACS analysis was performed after lymphoid tissue suspension stimulating DC to observe the MHC molecular expression. RESULTS: In the lymphoid tissue suspension, 91% of the cells was lymphocyte, others including granulocyte, plasmocyte, epithelium. The effect of stimulating mixed lymphocyte proliferation were higher in DC groups than in PBMC groups with the small intestinal mesenteric lymphoid tissue (P < 0.05). In the proportion of DC and mixed lymphocyte >or= 1:100 groups, the mixed lymphocyte proliferation were higher in the small intestinal mesenteric lymphoid tissues groups than in the OVA groups (P < 0.05). After stimulated by the small intestinal mesenteric lymphoid tissue, DC expressed higher MHC-I and -II molecules than control groups. CONCLUSIONS: The small intestinal mesenteric lymphoid tissue has high antigenicity; the antigen presenting ability of DC was much stronger than granulocytes; DC expresses high MHC-I and MHC-II molecules after stimulated by mixed lymphoid tissue suspension.

[Clinical analysis of infectious complications following abdominal cluster transplantation].

OBJECTIVE: To investigate the characteristic and management of postoperative infection in abdominal cluster transplantation. METHODS: Preliminary experience of two cases of abdominal cluster transplantation including small intestine was reviewed. RESULTS: Combination of five immunosuppressive agents based on tacrolimus was used. Severe Gram-negative bacillus infections occurred. The majority of invasive fungal infections was due to Candida species. Cytomegalovirus (CMV) infection increased monocytes and caused eosinopenia and an inversion of the CD4(+) to CD8(+) cell ratio in recipient I, and human CMV matrix proteins pp71 (CMV-pp71) was detected and identified in bile by PCR. Microabscesses in liver transplant biopsies were presented. CONCLUSIONS: Infectious complications after cluster transplantation were complicated. Strategies to optimize the immunity suppression protocol and early diagnosis and treatment will be important to reduce infection after abdominal cluster transplantation.

[Effects of norcantharidin on angiogenesis of human gallbladder carcinoma and its anti-angiogenic mechanisms].

OBJECTIVE: To investigate the effects of norcantharidin (NCTD) on angiogenesis of human gallbladder carcinoma and its anti-angiogenic mechanisms. METHODS: Human gallbladder carcinoma cells of the line GBC-SD were cultured. BALB/c nude mice were inoculated subcutaneously with the GBC-SD cells and then randomly divided into 6 groups: NCTD group, injected intraperitoneally with 1/5 of the LD(50) of NCTD twice a week for 6 weeks; 5-fluorouracil (5-FU) group, injected intraperitoneally with 1/5 of the LD(50) of 5-FU twice a week for 6 weeks; endostatin (ES) group, intraperitoneally with ES; NCTD + 5-FU group, injected intraperitoneally with 1/5 of the LD(50) of NCTD and 1/5 of the LD(50) of 5-FU twice a week for 6 weeks; NCTD + ES group, injected intraperitoneally with 1/5 of the LD(50) of NCTD and ES twice a week for 6 weeks; and normal saline (NS) group (control group), injected with NS. The mice were killed in the 7th week. The tumors were taken out to measure their volumes and undergo microscopy. SABC method of immunohistochemistry was used to measure the microvessel density (MVD) and the protein expression of the angiogenesis-related factors: proliferating cell nuclear antigen (PCNA), vascular endothelial growth factor (VEGF), angiopoietin (Ang)-2, thrombospondin (TSP), and tissue inhibitor of metalloprotease (TIMP)(2). Suspension of single tumor cell was prepared to examine the cell apoptosis by flow cytometry. RT-PCR was used to examine the mRNA expression of PCNA, VEGF, Ang-2, TSP, and TIMP2. RESULTS: (1) The MVD of the NCTD group was 4.12 +/- 1.4, significantly lower than those of the 5-FU group (15.8 +/- 5.9) and control group (17.6 +/- 3.2) (both P < 0.01), but not significantly different from those of the NCTD + 5-FU group (3.8 +/- 1.7), ES group (4.5 +/- 2.1), and NCTD + ES group (2.9 +/- 1.5) (all P > 0.05). The mice treated with NCTD showed significantly smaller tumor volume, lower PCNA protein expression, higher apoptotic rate, and higher PCNA/apoptosis ratio (P < 0.05 or P < 0.01), and significant correlation between MVD and tumor volume and between MVD and PCNA/apoptosis ratio (both P < 0.05). (2) The protein expression of VEGF and of Ang-2 of the NCTD group were both significantly lower than those of the control and 5-FU groups (all P < 0.01), however, not significantly different from those of the ES, NCTD + 5-FU, and NCTD + ES groups; and the protein expression of TSP and of TIMP2 of the NCTD group were both significantly higher than those of the control and 5-FU groups (all P < 0.01), however, not significantly different from those of the ES, NCTD + 5-FU, and NCTD + ES groups. MVD was positively correlated with VEGF and Ang-2 expression and negatively correlated with the expression of TSP and TIMP2 (all P < 0.05). (3) In comparison with the control group, the mRNA expression of VEGF and of Ang-2 of the tumor cells of the NCTD group were both significantly lower and the mRNA expression of TIMP2 was significantly higher. CONCLUSION: NCTD down-regulates the expression of the angiogenic factors, such as VEF|GF and Ang-2, and up-regulates the expression of the anti-angiogenic factors, such as TDP and TIMP2, thus inhibiting the angiogenesis in tumor, such as human gallbladder carcinoma, and further inhibiting the growth of tumor.

[Anti-tumor mechanism of norcantharidin for the implanted tumors of human gallbladder carcinoma in nude mice in vivo].

OBJECTIVE: To explore the anti-tumor mechanism of norcantharidin (NCTD) for the implanted tumors of human gallbladder carcinoma in nude mice in vivo. METHODS: Animal model of implanted tumors of human gallbladder carcinoma in nude mice was established. Mice were randomly divided into control, 5-FU, NCTD and NCTD + 5-FU groups and were taken different treatment. The expressions of PCNA, Ki-67, cyclin D1, p27, Bcl-2, Bax, Survivin, nm23/nm23-H1, MMP2 and TIMP2 proteins or genes in each tissue section of every group were determined by immunohistochemistry and RT-PCR. RESULTS: (1) On proliferation-related gene proteins, the expression of PCNA, Ki-67, cyclin D1 was significantly decreased, with significantly increased expression of p27 protein, in paraffin sections of NCTD group when compared with control group (P < 0.05); The expression of PCNA mRNA, cyclin D1 mRNA was decreased, with significantly increased expression of p27 mRNA in NCTD group. (2) On apoptosis-related gene proteins, the expression of Bcl-2 was significantly decreased in paraffin sections of NCTD group when compared with control group (P < 0.05); The expression of Bcl-2 mRNA, Survivin mRNA was significantly decreased, with significantly increased expression of Bax mRNA in NCTD group. (3) There was significant difference on invasion around tumor and lung metastasis in NCTD group when compared with control group (P < 0.01). On metastasis-related gene proteins, the expression of nm23 and TIMP2 was significantly increased, with significantly decreased expression of MMP2 in paraffin sections of NCTD group when compared with control group (P < 0.05); The expression of nm23-H1 mRNA, TIMP2 mRNA was significantly increased, with significantly decreased expression of MMP2 mRNA in NCTD group. CONCLUSIONS: The anti-tumor mechanism of NCTD for human gallbladder carcinoma in nude mice might correlated with inhibition of cell proliferation, blockage of cell cycle, induction of cell apoptosis, reducing of cell motility and invasive capability, alteration of the expression of proliferation-, apoptosis- and metastasis-related gene proteins such as PCNA, Ki-67, cyclin D1, p27, Bcl-2, Bax, Survivin, nm23, MMP2 and TIMP2.

Protective effect of sodium nitroprusside on the rat small intestine transplanted mucosa.

The intestinal mucosal epithelium is extremely susceptible to even brief periods of ischemia. Mucosal barrier damage, which is associated with ischemia/reperfusion (I/R) injury and consequently bacterial translocation, remains a major obstacle for clinically successful small bowel transplantation (SBT). Previous studies have demonstrated a protective effect of nitric oxide (NO) on other transplanted organs and NO mediated intestinal protection has also been reported in vitro. The aim of this study was to evaluate the effect of sodium nitroprusside (SNP), NO donor, on graft mucosal histology and molecular markers of function after SBT in rats. We used SNP in different period of heterotopic SBT rats. The groups consisted of SBT, pre-SNP group, and post-SNP group. Interestingly, the pre-SNP graft samples exhibited less damage compared to the SBT and post-SNP samples. In addition, mucosal samples from the pre-SNP group showed higher Na(+)-K(+)-ATPase activity and higher levels of laminin expression compared to the SBT and post-SNP samples. The findings of the present study reveal that SNP given before graft ischemia/reperfusion injury has a protective effect on mucosal histology and molecular markers of function in the transplanted small intestine.

[The in vitro effect of norcantharidin on proliferation and invasion of human gallbladder carcinoma GBC-SD cells and its mechanism].

OBJECTIVE: To study the effect and mechanism of action of norcantharidin on proliferation and invasion of GBC-SD cells. METHODS: GBC-SD cells of human gallbladder carcinoma were cultured by cell culture technique. The tetrazolium-based colorimetric assay was used to evaluate cell growth. The Matrigel experiment and the crossing-river test were used to examine the invasiveness of GBC-SD cells. Expression of MMP(2), TIMP(2), PCNA and Ki-67 proteins of GBC-SD cells was determined by streptavidin-biotin complex method. RESULTS: Norcantharidin inhibited the growth and proliferation of GBC-SD cells in a dose and time dependent manner, with an IC(50) value of 56.18 micro g/ml at 48 h. The Matrigel experiment showed that norcantharidin began to inhibit the in vitro invasion of GBC-SD cells at the concentration of 5 micro g/ml. At 40 micro g/ml, the invasive action of GBC-SD cells was inhibited completely and their crossing-river time was prolonged significantly. After treatment with norcantharidin, the expression of PCNA, Ki-67, MMP(2) was significantly decreased. With the increase in TIMP(2) expression, the MMP(2) to TIMP(2) ratio was decreased significantly (P < 0.05). CONCLUSION: Norcantharidin inhibits the in vitro proliferation and growth of human gallbladder carcinoma cells at relatively low concentrations by inhibiting PCNA and Ki-67 expression. Its anti-invasive activity may be the results of decrease in MMP(2) to TIMP(2) ratio and reduced cell motility.