A nomogram model for predicting prognosis of obstructive colorectal cancer.

BACKGROUND: The prognosis of obstructive colorectal cancer (oCRC) is worse than that of nonobstructive colorectal cancer. However, no previous study has established an individualized prediction model for the prognosis of patients with oCRC. We aimed to screen the factors that affect the prognosis of oCRC and to use these findings to establish a nomogram model that predicts the individual prognosis of patients with oCRC. METHODS: This retrospective study collected data of 181 patients with oCRC from three medical hospitals between February 2012 and December 2017. Among them, 129 patients from one hospital were used as the training cohort. Univariate and multivariate analyses were used in this training cohort to select independent risk factors that affect the prognosis of oCRC, and a nomogram model was established. The other 52 patients from two additional hospitals were used as the validation cohort to verify the model. RESULTS: Multivariate analysis showed that carcinoembryonic antigen level (p = 0.037, hazard ratio [HR] = 2.872 [1.065-7.740]), N stage (N1 vs. N0, p = 0.028, HR = 3.187 [1.137-8.938]; N2 vs. N0, p = 0.010, HR = 4.098 [1.393-12.051]), and surgical procedures (p = 0.002, HR = 0.299 [0.139-0.643]) were independent prognostic factors of overall survival in patients with oCRC. These factors were used to construct the nomogram model, which showed good concordance and accuracy. CONCLUSION: Carcinoembryonic antigen, N stage, and surgical method are independent prognostic factors for overall survival in patients with oCRC, and the nomogram model can visually display these results.

BRAFV600E mutant colorectal cancer cells mediate local immunosuppressive microenvironment through exosomal long noncoding RNAs.

BACKGROUND: BRAFV600E mutated colorectal cancer (CRC) is prone to peritoneal and distant lymph node metastasis and this correlates with a poor prognosis. The BRAFV600E mutation is closely related to the formation of an immunosuppressive microenvironment. However, the correlation between BRAFV600E mutation and changes in local immune microenvironment of CRC is not clear. AIM: To explore the effect and mechanism of BRAFV600E mutant on the immune microenvironment of CRC. METHODS: Thirty patients with CRC were included in this study: 20 in a control group and 10 in a treatment group. The density of microvessels and microlymphatic vessels, and M2 subtype macrophages in tumor tissues were detected by immunohistochemistry. Screening and functional analysis of exosomal long noncoding RNAs (lncRNAs) were performed by transcriptomics. The proliferation and migration of human umbilical vein endothelial cells (HUVECs) and human lymphatic endothelial cells (HLECs) were detected by CCK-8 assay and scratch test, respectively. The tube-forming ability of endothelial cells was detected by tube formation assay. The macrophage subtypes were obtained by flow cytometry. The expression of vascular endothelial growth factor (VEGF)-A, basic fibroblast growth factor (bFGF), transforming growth factor (TGF)-ß1, VEGF-C, claudin-5, occludin, zonula occludens (ZO)-1, fibroblast activation protein, and α-smooth muscle actin was assessed by western blot analysis. The levels of cytokines interleukin (IL)-6, TGF-ß1, and VEGF were assessed by enzyme-linked immunosorbent assay. RESULTS: BRAFV600E mutation was positively correlated with the increase of preoperative serum carbohydrate antigen 19-9 (P < 0.05), and with poor tumor tissue differentiation in CRC (P < 0.01). Microvascular density and microlymphatic vessel density in BRAFV600E mutant CRC tissues were higher than those in BRAF wild-type CRC (P < 0.05). The number of CD163+ M2 macrophages in BRAFV600E mutant CRC tumor tissue was markedly increased (P < 0.05). Compared with exosomes from CRC cells with BRAF gene silencing, the expression of 13 lncRNAs and 192 mRNAs in the exosomes from BRAFV600E mutant CRC cells was upregulated, and the expression of 22 lncRNAs and 236 mRNAs was downregulated (P < 0.05). The biological functions and signaling pathways predicted by differential lncRNA target genes and differential mRNAs were closely related to angiogenesis, tumor cell proliferation, differentiation, metabolism, and changes in the microenvironment. The proliferation, migration, and tube formation ability of HUVECs and HLECs induced by exosomes in the 1627 cell group (HT29 cells with BRAF gene silencing) was greatly reduced compared with the HT29 cell group (P < 0.05). Compared with the HT29 cell group, the expression levels of VEGF-A, bFGF, TGF-ß1, and VEGF-C in the exosomes derived from 1627 cells were reduced. The expression of ZO-1 in HUVECs, and claudin-5, occludin, and ZO-1 in HLECs of the 1627 cell group was higher. Compared with the 1627 cell group, the exosomes of the HT29 cell group promoted the expression of CD163 in macrophages (P < 0.05). IL-6 secretion by macrophages in the HT29 cell group was markedly elevated (P < 0.05), whereas TGF-ß1 was decreased (P < 0.05). The levels of IL-6, TGF-ß1, and VEGF secreted by fibroblasts in the 1627 cell group decreased, compared with the HT29 cell group (P < 0.05). CONCLUSION: BRAFV600E mutant CRC cells can reach the tumor microenvironment by releasing exosomal lncRNAs, and induce the formation of an immunosuppressive microenvironment.

Gegen Qinlian decoction enhances immunity and protects intestinal barrier function in colorectal cancer patients via gut microbiota.

BACKGROUND: We previously showed, using the Traditional Chinese Medicine System Pharmacology Database, that Gegen Qinlian decoction (GQD) had a direct antitumor effect, and was combined with programmed cell death protein (PD)-1 inhibitors to treat microsatellite stable (MSS) tumor-bearing mice. However, the effect of GQD on patients with colorectal cancer (CRC) is not clear. AIM: To determine the therapeutic mechanism of GQD in improving immune function, reducing inflammation and protecting intestinal barrier function. METHODS: Seventy patients with CRC were included in this study: 37 in the control group and 33 in the treatment group. The proportions of CD4+ T, CD8+ T, natural killer (NK), NKT and T regulatory cells were measured by flow cytometry. Levels of the cytokines tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-2, IL-6, IL-10 and serotonin (5-hydroxytryptamine; 5-HT) in serum were assessed by enzyme-linked immunosorbent assay (ELISA). The expression of zonula occludens (ZO)-1, occludin, nuclear factor (NF)-κB and TNF-α in tumor and normal tissues was measured by immunohistochemistry. The composition of gut microbiota from patients in the treatment group was assessed using 16S rDNA analysis. RESULTS: There were no adverse events in the treatment group. The proportion of CD4+ T cells and NKT cells in the post-treatment group was significantly higher than that in the pre-treatment and control groups (P < 0.05). The level of TNF-α in the post-treatment group was significantly lower than that in the pre-treatment and control groups (P < 0.05). The concentration of 5-HT in the post-treatment group was significantly lower than that in the pre-treatment group (P < 0.05). The expression of ZO-1 and occludin in tumor tissues in the treatment group was significantly higher than that in the control group (P < 0.05). The expression of ZO-1 in normal tissues of the treatment group was significantly higher than that in the control group (P = 0.010). Compared with the control group, expression of NF-κB and TNF-α in tumor tissues of the treatment group was significantly decreased (P < 0.05). Compared with the pre-treatment group, GQD decreased the relative abundance of Megamonas and Veillonella. In addition, GQD increased the relative abundance of Bacteroides, Akkermansia and Prevotella. CONCLUSION: GQD enhances immunity and protects intestinal barrier function in patients with CRC by regulating the composition of gut microbiota.

Sister Mary Joseph's nodule in endometrial carcinoma: A case report.

BACKGROUND: Sister Mary Joseph's nodule (SMJN) is rare and may occur independently or simultaneously with malignant tumors in the abdominal cavity. It has a poor prognosis. Endometrial carcinoma is the most common malignant tumor in the upper segment of the female reproductive tract; however, it rarely occurs with SMJN. We here report a case of endometrial carcinoma with SMJN. CASE SUMMARY: A 75-year-old woman was diagnosed with endometrial cancer > 2 years ago. After multiple cycles of chemotherapy, an obvious but painless umbilical mass was detected 2 wk before admission. The patient did not undergo surgical treatment but received chemotherapy, which was different from previous chemotherapy, for three consecutive cycles after discovery of umbilical metastases. Currently, the umbilical metastatic tumor has reduced, and the quality of life of the patient has significantly improved. CONCLUSION: Once the umbilical mass is found, the possibility of SMJN should be considered. We should also take into account the poor prognosis of endometrial carcinoma complicated with umbilical metastasis, especially in patients with advanced tumors, and it is important to choose an appropriate treatment.

Effects of PHA-665752 and Cetuximab Combination Treatment on In Vitro and Murine Xenograft Growth of Human Colorectal Cancer Cells with KRAS or BRAF Mutations.

BACKGROUND: It remains unknown whether blockade of c-Met signaling and epidermal growth factor receptor signaling is effective in suppressing the growth of human colorectal cancer (CRC) cells. In this study, we investigated the effects of the c-Met inhibitor PHA-665752 alone and in combination with cetuximab on the growth of human CRC cells in vitro and in mouse xenografts. METHODS: Human CRC cell lines (Caco2, HCT-116, and HT-29) and mice bearing HCT-116 xenografts were treated with cetuximab in the absence or presence of PHA-665752. Cell viability and apoptosis were examined using the MTT and TUNEL assays, respectively. Vimentin was measured by immunohistochemistry as a marker for epithelial-to-mesenchymal transition. Western blotting was used to determine signaling protein expression levels. RESULTS: The MTT assay showed that the growth of Caco2, HCT-116, and HT-29 cells was inhibited by PHA-665752 in a dose-dependent manner, but only Caco2 cell growth was suppressed by cetuximab. Combination treatment with PHA-665752 and cetuximab inhibited the proliferation of Caco2 cells and RAS mutant CRC cell lines. However, relative to the PHA-665752-alone treatment group, HT-29 cells with a BRAF mutation showed no noticeable effect. The mean tumor volume in mice treated with cetuximab in combination with PHA-665752 was significantly smaller than that in the mice treated with only cetuximab (P = 0.033) or PHA-665752 (P < 0.01). Similarly, the expression of vimentin in the mice treated with PHA-665752 in combination with cetuximab was significantly lower than that in the mice treated with cetuximab or PHA-665752 alone (P < 0.05 in each case). TUNEL assays revealed that treatment with PHA-665752 in combination with cetuximab markedly increased CRC cell apoptosis. Western blotting analysis of signaling protein expression showed that PHA- 665752 inhibited Met phosphorylation (P < 0.05). In addition, treatment with cetuximab alone or in combination with PHA-665752 effectively inhibited EGFR phosphorylation (P < 0.05). CONCLUSION: Combination treatment with PHA-665752 and cetuximab suppressed in vitro and in vivo CRC cell growth more than treatment with either agent alone did.

Absence of NF-κB subunit p50 ameliorates cold immobilization stress-induced gastric ulcers.

Stress ulcers are a common complication in critically ill patients, but the underlying mechanism is little known. This study characterized the function of the p50 subunit of NF-κB in an experimental model of cold immobilization stress-induced gastric ulcers. Stress-induced gastric mucosal inflammation and gastric injury were examined in wild-type and NF-κB p50-deficient mice. When subjected to cold immobilization stress, NF-κB was rapidly activated in the gastric mucosa in WT mice whereas the majority of κB DNA-binding activity was abrogated from p50(-/-) mice. Deficiency of p50 ameliorated stress-induced expression of TNF-α, MIP-2, and ICAM-1, resulting in reduced mucosal accumulation of neutrophils and gastric injury. These data indicated a critical role for the p50 in the gastric mucosal inflammatory response to cold restraint stress.

[Role of angiotensin II type 1 receptor in activation of nuclear factor-kappaB and activator protein-1 in lung of mice with acute lung injury].

OBJECTIVE: To explore the role of angiotensin II type 1 (AT1) receptor in activation of nuclear factor-kappaB (NF-kappaB) and activator protein-1 (AP-1) in lung of mice with LPS-induced acute lung injury (ALI). METHODS: Eighty-eight BABL/c mice were divided into control group (n = 8), LPS group (n = 40), and LPS + AT1 receptor antagonist ZD7155 group (n = 40) according to the random number table. Puncture of trachea was done in all mice. Mice in LPS + ZD7155 group were intraperitoneally injected with 10 mg/kg ZD7155. Mice in LPS and control groups were intraperitoneally injected with normal saline in the same volume as that of ZD7155. Thirty minutes later, 1 mg/mL LPS was dripped into trachea of mice in LPS and LPS + ZD7155 groups (2 mg/kg). Normal saline in the same volume as that of LPS was dripped into trachea of mice in control group. Lung tissue samples of mice in LPS and LPS + ZD7155 groups were harvested at post dripping hour (PDH) 1, 3, 6, 12, and 24. Lung tissue sample of mice in control group was harvested at PDH 24. Expression of AT1 receptor was determined with Western blot. AP-1 and NF-kappaB activity in lung tissue was detected with electrophoretic mobility shift assay. Data were processed with one-way analysis of variance. RESULTS: The relative expression amount of AT1 receptor protein in lung tissue of mice in LPS group at each time point was increased obviously as compared with that of mice in control group (0.69 +/- 0.28, F = 9.356, with P values all below 0.01), and it peaked at PDH 6 (3.44 +/- 0.90), while that of mice in LPS + ZD7155 group was less than that in LPS group at each time point (F = 9.356, with P values all below 0.01). NF-kappaB activity in mice lung was markedly increased in LPS group at each time point as compared with mice in control group (5.47 +/- 0.08, F = 26.443, with P values all below 0.05), and its peak value in LPS group was found at PDH 3 (52.33 +/- 3.25). While NF-kappaB activity in mice of LPS + ZD7155 group was obviously lower than that in LPS group at each time point (F = 26.443, with P values all below 0.05). AP-1 activity in lung was enhanced significantly in LPS group at each time point as compared with that in control group (2.5 +/- 0.4, F = 34.685, with P values all below 0.05), and the activity peaked at PDH 6 (73.3 +/- 9.5) in LPS group. The activity was obviously weaker in mice in LPS + ZD7155 group as compared with that in LPS group at each time point (F = 34.685, with P values all below 0.05). CONCLUSIONS: AT1 receptor contributes to LPS-induced ALI through activating NF-kappaB and AP-1 in lung tissue.

Notoginsenoside R1 attenuates renal ischemia-reperfusion injury in rats.

Ischemia-reperfusion (I/R) injury of the kidney is a complex pathophysiological process and a major cause of acute renal failure. It has been shown that I/R injury is related to inflammatory responses and activation of apoptotic pathways. Inhibition of certain elements of inflammatory responses and apoptotic pathway seemed to ameliorate renal I/R injury. As an effective element of Panax notoginseng, NR1 has antioxidant, anti-inflammatory, antiapoptotic, and immune-stimulatory activities. Therefore, we speculate that NR1 can attenuate renal I/R injury. Ischemia-reperfusion injury was induced by renal pedicle ligation followed by reperfusion along with a contralateral nephrectomy. Male Sprague-Dawley rats were randomized to four groups: sham group, I/R control group, NR1-1 group (rats treated with NR1, 20 mg.kg.d) and NR1-2 group (rats treated with NR1, 40 mg.kg.d). All animals were killed 72 h after I/R induction. Blood and renal tissues were collected. Renal dysfunction was observed by the level of serum creatinine and histological evaluation. Apoptosis and inflammatory response in the tissue of kidney were detected mainly with molecular biological methods. NR1 attenuated I/R-induced renal dysfunction as indicated by the level of serum creatinine and histological evaluation. It prevented the I/R-induced increases in the levels of proinflammatory cytokine TNF-alpha, myeloperoxidase activity, phosphorylation of p38, and activation of nuclear factor kappaB with cell apoptosis in the kidney and enhanced expression of antiapoptosis cytokine bcl-2. Treatment with NR1 improves renal function after I/R associated with a significant reduction in cell apoptosis and inflammatory responses, which may be related to p38 and nuclear factor kappaB inhibition.

Angiotensin II type-1 receptor antagonist attenuates LPS-induced acute lung injury.

Angiotensin II is able to trigger inflammatory responses through an angiotensin II type 1 (AT1) receptor. The role of AT1 receptor in acute lung injury (ALI) is poorly understood. Mice were randomly divided into three groups (n=40 each groups): NS group; LPS group (2mg/kg LPS intratracheally); and LPS+ZD 7155 group, 10mg/kg ZD 7155 (an AT1 receptor antagonist) intraperitoneally 30 min prior to LPS exposure. Samples from the lung were isolated and assayed for histopathology analyses or proinflammatory gene expressions, angiotensin II receptors expressions and nuclear factors activities. LPS exposure resulted in severe ALI, elevated levels of TNF-alpha and IL-1 beta mRNA expressions, and increased activities of NF-kappaB and activated protein (AP)-1. Upregulation of AT1 receptor and down-regulation of AT2 receptor were also observed after LPS challenge. Pretreatment with ZD 7155 significantly inhibited the increase of AT1 receptor expression and upregulated AT2 receptor expression. ZD 7155 also reduced the mRNA expression of TNF-alpha and IL-1 beta, inhibited the activation of NF-kappaB and AP-1, and improved lung histopathology. These findings suggest that antagonism of AT1 receptor inhibits the activation of NF-kappaB and AP-1 in the lung, which may mediate the release of TNF-alpha and IL-1 beta and contribute to LPS-induced ALI.

[Endostar reduces the growth and metastasis by inhibiting angiogenesis and lymphangiogenesis in nude mouse models of human cervical cancer].

OBJECTIVE: To observe the influence of endostar alone or in combination with cisplatin on tumor growth and metastasis, as well as the inhibition of angiogenesis and lymphangiogenesis in nude mouse models of human cervical cancer. METHODS: HeLa cells were inoculated subcutaneously into the hind flank region of female nu/mice to establish xenograft models. The nude mice were randomly divided into 5 groups: (1) sodium chloride (as control); (2) cisplatin alone; (3) endostar alone; (4) cisplatin plus endostar (10 mg/kg); (5) cisplatin plus endostar (20 mg/kg). The course of all the treatments lasted for 4 weeks. The tumor growth and lymph node metastasis were observed. Immunohistochemical staining was employed to detect the angiogenesis and lymphangiogenesis. RESULTS: (1) Either endostar alone or endostar with cisplatin inhibited the tumor growth significantly than cisplatin and NS (P < 0.05). (2) The rates of lymph node metastasis in the endostar (20 mg/kg) with cisplatin, the endostar (10 mg/kg) with cisplatin, the endostar, the cisplatin and the NS groups were 0 (0/8), 12.5% (1/8), 12.5% (1/8), 62.5% (5/8) and 75.0% (6/8) (P = 0.002), respectively. (3) The MVD of tumor tissue in these five groups were 10.88 +/- 1.38, 10.25 +/- 1.22, 10.83 +/- 2.29, 15.58 +/- 2.31 and 22.08 +/- 1.93, respectively (P < 0.05). The MLD were 5.00 +/- 0.63, 5.17 +/- 0.75, 6.00 +/- 0.63, 14.33 +/- 1.63 and 13.67 +/- 1.21, respectively (P < 0.05). CONCLUSION: Endostar can reduce the tumor growth and metastasis by inhibiting angiogenesis and lymphangiogenesis in nude mouse model of human cervical cancer.

Tec kinase mediating IL-8 transcription in monocytes stimulated with LPS.

The purpose of present study was to explore the possibility of Tec kinase as a mediator for IL-8 transcription in monocytes stimulated with LPS. Plasmids of mouse Tec kinase IV or Tec kinase IV with inactivating point mutations generated with QuikChange site-directed mutagenesis were co-transfected with IL-8 promoter driven luciferase construct into RAW264.7 cells, then luciferase activity was measured with a luminometer. The results shown Tec kinase could significantly enhance IL-8 transcription. Furthermore, point inactivating mutation in SH2, PH or PTK domain almost completely abolish the effects of Tec kinase on the transcription of IL-8. In the transfection experiment, PD98059, a MEK1 inhibitor, decreased the transcription of IL-8 in a dose dependent pattern. When siRNA for Tec kinase was transfected into THP-1 cells, it could efficiently block the production of IL-8 from THP-1 cells (p < 0.01) stimulated with LPS. In conclusion, Tec kinase may mediate the transcription of IL-8 in monocyte stimulated with LPS.

Angiotensin II induces type I collagen gene expression in human dermal fibroblasts through an AP-1/TGF-beta1-dependent pathway.

Angiotensin II is critically involved in skin wound healing, but the underlying mechanism remains unclear. This study investigated the effect of angiotensin II on type I collagen gene activation in human dermal fibroblasts and the possible mechanism involved. Angiotensin II stimulated the mRNA and protein expression of type I collagen and TGF-beta1. Effects were abolished by the angiotensin AT1 receptor antagonist ZD7155 but not by the AT2 blocker PD123319. Blockade of TGF-beta1 markedly inhibited angiotensin II-induced type I collagen gene expression. Activator protein-1 (AP-1) decoy ODNs transfection suppressed angiotensin II-induced TGF-beta1 expression, and also, diminished type I collagen expression. These data indicated that angiotensin II induces collagen gene activation in human dermal fibroblasts through an AT1-mediated AP-1/TGF-beta1 signaling pathway.

Activation of p38 MAPK by reactive oxygen species is essential in a rat model of stress-induced gastric mucosal injury.

Stress ulceration is a common complication in critically ill patients and can result in significant upper gastrointestinal bleeding associated with a high morbidity and mortality. At present, little is known of the molecular mechanisms underlying the incidence of this type of gastric damage. In the present study, we investigated the temporal activation of the redox-sensitive p38 signaling transduction cascade and its roles in a well-defined experimental model of cold immobilization stress-induced gastric ulceration. Exposure of Sprague-Dawley rats to 6 h of cold immobilization stress led to a rapid activation of p38 in the gastric mucosa at as early as 15 min after stress, and this activation was maximal after 1.5 h of stress and still persisted until the end of stress. Selectively blocking p38 by pretreatment with SB 239063, a potent and selective p38 inhibitor, suppressed the stress-promoted TNF-alpha, IL-1beta, and CINC-1 production and then prevented the subsequent neutrophil infiltration, gastric mucosal epithelial necrosis and apoptosis, and the ulcerative lesions formation. Prior administration of the free radical scavengers, tempol and N-acetyl-L-cysteine, abolished the stress induction of p38 activation and the resulting mucosal inflammation and gastric injury. These results demonstrate that reactive oxygen species-mediated p38 activation plays an essential role in the pathogenesis of stress-induced gastric inflammatory damage in the rat model of cold immobilization stress. Our findings suggested that inhibition of p38 activation might be a potential strategy for the prophylaxis and treatment of stress ulceration.

Inhibition of p38 mitogen-activated protein kinase attenuates experimental autoimmune hepatitis: involvement of nuclear factor kappa B.

AIM: To investigate the role of p38 mitogen-activated protein kinase (p38MAPK) in murine experimental autoimmune hepatitis (EAH). METHODS: To induce EAH, the syngeneic S-100 antigen emulsified in complete Freud's adjuvant was injected intraperitoneally into adult male C57Bl/6 mice. Liver injury was assessed by serum ALT and liver histology. The expression and activity of p38 MAPK were measured by Western blot and kinase activity assays. In addition, DNA binding activities of nuclear factor kappa B (NF-kappaB) were analyzed by electrophoretic mobility shift assay. The effects of SB203580, a specific p38 MAPK inhibitor, on liver injuries and expression of proinflammatory cytokines (interferon-gamma, IL-12, IL-1beta and TNF-alpha) were observed. RESULTS: The activity of p38 MAPK and NF-kappaB was increased and reached its peak 14 or 21 d after the first syngeneic S-100 administration. Inhibition of p38 MAPK activation by SB203580 decreased the activation of NF-kappaB and the expression of proinflammatory cytokines. Moreover, hepatic injuries were improved significantly after SB203580 administration. CONCLUSION: p38 MAPK and NF-kappaB play an important role in an animal model of autoimmune hepatitis (AIH) induced by autoantigens.

Sustained activation of nuclear factor-kappaB by reactive oxygen species is involved in the pathogenesis of stress-induced gastric damage in rats.

OBJECTIVE: Stress ulceration is a common complication in critically ill patients, but the mechanisms involved are poorly understood. In this study we investigated the temporal activation of the redox-sensitive transcription factor nuclear factor-kappaB and its roles in an experimental model of cold immobilization stress-induced gastric mucosal lesions. DESIGN: Prospective, controlled, and randomized animal study. SETTING: University research laboratory. SUBJECTS: Male Sprague-Dawley rats. INTERVENTIONS: The rats were subjected to cold immobilization stress for a total of 6 hrs. The temporal profiles of nuclear factor-kappaB activation and expression of tumor necrosis factor-alpha, interleukin-1beta, cytokine-induced neutrophil chemoattractant-1 (CINC-1), intercellular adhesion molecule-1 (ICAM-1), and inducible nitric oxide synthase (iNOS) were determined in the gastric corpus mucosa of stressed rats. To study the roles of nuclear factor-kappaB activation, rats received an intravenous bolus of a specific nuclear factor-kappaB inhibitor Bay 11-7082 (20 mg/kg) 1 hr before stress. For antioxidant administration, rats were treated with intravenous injection of a free radical scavenger pyrrolidine dithiocarbamate (50, 100, and 200 mg/kg) 1 hr before stress. MEASUREMENTS AND MAIN RESULTS: Exposure of rats to 6 hrs of stress led to a rapid and persistent activation of nuclear factor-kappaB, which was associated with transient degradation of inhibitory protein IkappaBalpha and slower but sustained degradation of IkappaBbeta. Nuclear factor-kappaB activation preceded the induction of tumor necrosis factor-alpha, interleukin-1beta, CINC-1, ICAM-1, and iNOS messenger RNAs, all of which were linearly increased with the duration of stress. Bay 11-7082 selectively blocked the stress-induced nuclear factor-kappaB activation and up-regulation of tumor necrosis factor-alpha, interleukin-1beta, CINC-1, ICAM-1, and iNOS messenger RNAs. Inhibition of expression of these proinflammatory genes prevented the increases in myeloperoxidase activity (an indicator of neutrophil infiltration) in gastric mucosa and the development of gastric damage. Pyrrolidine dithiocarbamate dose-dependently inhibited the stress-induced nuclear factor-kappaB pathway activation and consequential proinflammatory gene expression, neutrophil infiltration, and gastric damage, suggesting the involvement of reactive oxygen species in these processes. CONCLUSIONS: Sustained activation of nuclear factor-kappaB by reactive oxygen species is an important in vivo mechanism mediating stress-induced gastric inflammatory damage in rats.

[Activation of nuclear factor-kappaB signaling pathway in rat gastric mucosa during stress ulceration].

OBJECTIVE: To investigate the time course of nuclear factor-kappaB (NF-kappaB) activation in the process of stress ulcer formation. METHODS: Model of stress ulcer was reproduced by subjecting male Sprague-Dawley rats to water-immersion restraint (WIR) stress. At indicated time after the beginning of WIR stress, animals were sacrificed and cytoplasmic and nuclear protein and total RNA were prepared from gastric corpus mucosal tissues. DNA-binding activity of NF-KB was assessed as an index of NF-kappaB activation with electrophoretic mobility shift assay. Degradation of IkappaBalpha and IkappaBbeta, the inhibitory proteins of NF-kappaB, was analyzed by Western blot analysis. Expression of NF-kappaB dependent genes including tumor necrosis factor-alpha (TNF-alpha) , interleukin-1beta (IL-1beta), cytokine-inducible neutrophil chemoattractant-1 ( CINC-1), intercellular adhesion molecule-1 (ICAM-1), and inducible nitric oxide synthase (iNOS) was detected with Northern blot analysis. RESULTS: WIR stress induced a rapid biphasic activation of gastric mucosal NF-kappaB within 15 min of the beginning of stress, peaking at 45 min and 360 min. Compared with baseline, NF-kappaB activation by stress was increased (10.6 +/- 1.3) and (8.9 +/- 1.2) fold at 45 min and 360 min, respectively (P < 0.01). Antibody supershift assays revealed that p50/p65 heterodimer was the major active component of mucosal NF-kappaB. Western blot analysis showed that degradation of IkappaBalpha and IkappaBbeta occurred at first and second wave of NF-kappaB activation. Corresponding with the rapid and persistent activation of NF-kappaB, the levels of TNF-alpha, IL-1beta, CINC-1 and ICAM-1 mRNA in gastric mucosa were markedly increased 15 to 30 min after stress, respectively. Up-regulation of iNOS mRNAs was observed 30 to 90 min after stress, and the expression of all of these genes was increased consistently until the end of stress. CONCLUSION: NF-kappaB activation is an early event and may play an important role in proinflammatory gene over-expression in rat gastric mucosa during WIR stress.

[Role of p38 mitogen-activated protein kinase in the pathogenesis of stress ulcer].

OBJECTIVE: To determine whether the activation of p38 mitogen-activated protein kinase (MAPK) is involved in the pathogenesis of stress ulcer. METHODS: Model of stress ulcer was established with the treatment of rats with water-immersion restraint (WIR) stress. Ulcer index (UI) was macroscopically evaluated as a parameter of gastric mucosal lesions. Expression of phospho- and pan-p38 in gastric mucosa was detected using Western blot analysis. Tumor necrosis factor-alpha (TNF-alpha) and Interleukin 1beta (IL-1beta) gene expressions were analyzed by Northern blot analysis. As indicated in some experiments, rats were pretreated with intravenous injection of the specific p38 MAPK inhibitor CNI-1493 prior to WIR stress and then the changes of UI and TNF-alpha and IL-1beta mRNA expression were examined. RESULTS: The p38 MAPK was persistently activated in the gastric mucosa of rats with WIR stress, with maximal activation after 1 h of stress [(6.8 +/- 3.2) fold of baseline levels, P < 0.01]. Inhibition of p38 MAPK activation with CNI-1493 led to a marked decrease in UI in WIR stress rats. Similarly, the increased gene expression of proinflammatory cytokines TNF-alpha and IL-1beta in gastric mucosa induced by WIR stress were significantly diminished by p38 MAPK inhibition. CONCLUSION: p38 MAPK might have an important role in the pathogenesis of stress ulcer.

Dynamic changes of mitogen-activated protein kinase signal transduction in rats with severe acute pancreatitis.

OBJECTIVE: To investigate the dynamic changes of mitogen-activated protein kinase (MAPK) signal transduction in rats with severe acute pancreatitis (SAP). METHODS: The SAP model was induced by infusing the bilio-pancreatic duct of 56 Sprague-Dawley rats with 5% sterile sodium taurocholate solution. The rats were randomly divided into seven groups: control group, 0.5 h postoperative group, 1 h group, 3 h group, 6 h group, 12 h group and 24 h group. Western blot analysis was used to determine the activities of p38 MAPK and c-Jun N-terminal kinase (JNK) in the pancreas and lungs. RESULTS: In the rats of the control group, basal p38MAPK activity could be detected but not that of JNK. After SAP was induced, the p38MAPK activity in the pancreas increased markedly and peaked at 3 h, but in the lung it peaked at 6 h. The p38MAPK activity in the pancreas and lungs was significantly higher than the basal activity at the 24 h time point. The activity of INK was only increased at the 12 h point and was not detectable at 24 h. CONCLUSION: The MAPK signal transduction pathway, in particular p38MAPK, plays an important role in the pathogenesis of SAP.

Expression of vascular endothelial growth factor-C and the relationship between lymphangiogenesis and lymphatic metastasis in colorectal cancer.

AIM: To investigate the expression of vascular endothelial growth factor-C (VEGF-C) and the relationship between VEGF-C and lymphangiogenesis, lymph node metastasis in colorectal cancer. METHODS: Fifty six cases of colorectal cancer were selected randomly. Expression of VEGF-C was detected by immunohistochemistry, and lymphatic vessels were stained by enzyme histochemical method. RESULTS: VEGF-C expression was found in 66.7% (37/56) patients. In VEGF-C positive and negative patients, the lymphatic vessel density was 25.16+/-7.52 and 17.14+/-7.22, respectively (P<0.05). The rate of lymph node metastasis in VEGF-C positive patients (81.1%) was significantly higher than that in the negative group (42.1%). CONCLUSION: VEGF-C expression may induce lymphangiogenesis in colorectal cancer, as a result, tumor cells can entry the lymphatic vessels easily. VEGF-C may serve as a useful prognotic factor in colorectal carcinoma.