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ABSTRACT: To clarify the role of mast cells and neuropeptides substance P (SP), somatostatin (SS), and vasoactive intestinal peptide (VIP) in dextran sulfate sodium (DSS)-induced colitis in rats. Methods Experimental colitis was induced in Sprague-Dawley rats (180-200 g, n=20) by oral ingestion of 4% (w/v) DSS in drinking water for 7 days. Control rats (n=5) drank water and were sacrificed on day 0. Mast cell number, histamine levels in whole blood and tissue, tissue levels of SP, SS and, VIP in the distal colon of the rats were measured on day 8, day 13, and day 18 of experimentation. Results Oral administration of 4% DSS solution for 7 days resulted in surface epithelial loss and crypt loss in the distal colon. Mast cell count increased on day 8 (1.75±1.09/mm vs. 0.38±0.24/mm, P<0.05) and day 13 (1.55±1.01/mm vs. 0.38±0.24/mm, P<0.05) after DSS treatment. Whole blood histamine levels were increased on day 8 (266.93±35.62 ng/mL vs. 76.87±32.28 ng/mL, P<0.01) and gradually decreased by day 13 and day 18 after DSS treatment. Histamine levels in the distal colon were decreased on day 8 (1.77±0.65 ng/mg vs. 3.06±0.87 ng/mg, P<0.05) and recovered to control levels by day 13 after DSS treatment. SP level in the distal colon gradually increased and were raised significantly by day 13 (8777.14±3056.14 pg/mL vs. 4739.66±3299.81 pg/mL, P<0.05) after DSS treatment. SS and VIP levels in the distal colon were not changed. Conclusions Mast cell degranulation followed by histamine release may play an important role in the pathogenesis of colitis induced by DSS. SP may be a significant substance in the progression of inflammation and the recovery process of DSS-induced colitis.
Chinese Medical Sciences Journal 03/2013; 28(1):28-33.
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ABSTRACT: BACKGROUND: Cyclooxygenase-2(COX-2) promotes carcinogenesis, tumor proliferation, angiogenesis, prevention of apoptosis, and immunosuppression. Meanwhile, COX-2 over-expression has been associated with tumor behavior and prognosis in several cancers. This study investigated the antitumor effects of the selective COX-2 inhibitor, Celecoxib, on breast cancer in vitro and in vivo. METHODS: Human breast cancer MCF-7 and MDA-MB-231 cells were cultured with different concentration (10, 20, 40mumol/L) of celecoxib after 0-96 hours in vitro. MTT assay was used to determine the growth inhibition of breast cancer cells in vitro. The expression of COX-2 on mRNA was measured by real-time quantitive PCR analysis. Flow cytometry was performed to analyze the cell cycle of MCF-7 cells. Levels of PGE2 were measured by ELISA method. The in vivo therapeutic effects of celecoxib were determined using rat breast cancer chemically induced by 7,12-dimethylben anthracene (DMBA). RESULTS: The inhibition of proliferation of both MCF-7 and MDA-MB-231 cells in vitro by celecoxib was observerd in time and dose dependent manner. Celecoxib effectively down-regulated the expression of COX-2. The cell cycle was arrested at G0/G1, and rate of cells in S phase was obviously decreased. Levels of PGE2 were inhibited by Celecoxib. The tumor incidence rate of the celecoxib group was lower than that of the control group. In addition, the tumor latency period of the celecoxib group was longer than that of the control group. CONCLUSIONS: Celecoxib inhibited the proliferation of breast cancer cell lines in vitro, and prevented the occurrence of rat breast cancer chemically induced by DMBA. Therefore, celecoxib exhibits an antitumor activity and seems to be effective in anti-tumor therapy.
Cancer Cell International 12/2012; 12(1):53. · 1.97 Impact Factor
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ABSTRACT: The exact mechanism of the effects of hypoxia on the proliferation and apoptosis in carcinoma cells is still conflicting. This study investigated the variation of hypoxia-inducible factor-1α(HIF-1α) expression and the apoptosis effect of hypoxia stimulated by cobalt chloride (CoCl(2)) in pancreatic cancer PC-2 cells.
PC-2 cells were cultured with different concentration (50-200 μmol/L) of CoCl(2) after 24-120 hours to simulate hypoxia in vitro. The proliferation of PC-2 cells was examined by MTT assay. The cellular morphology of PC-2 cells were observed by light inverted microscope and transmission electron microscope(EM). The expression of HIF-1α on mRNA and protein level was measured by semi-quantitative RT-PCR and Western blot analysis. Apoptosis of PC-2 cells were demonstrated by flow cytometry with Annexin V-FITC/PI double staining.
MTT assay showed that the proliferation of PC-2 cells were stimulated in the first 72 h, while after treated over 72 h, a dose- dependent inhibition of cell growth could be observed. By using transmission electron microscope, swollen chondrosomes, accumulated chromatin under the nuclear membrane and apoptosis bodies were observed. Flow cytometer(FCM) analysis showed the apoptosis rate was correlated with the dosage of CoCl(2). RT-PCR and Western blot analysis indicated that hypoxia could up-regulate the expression of HIF-1α on both mRNA and protein levels.
Hypoxic microenvironment stimulated by CoCl(2) could effectively induce apoptosis and influence cell proliferation in PC-2 cells, the mechanism could be related to up-expression of HIF-1α.
Journal of Experimental & Clinical Cancer Research 03/2012; 31:28. · 2.15 Impact Factor
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ABSTRACT: In the present study, we investigated the in vitro and in vivo antitumor effects of crude extract of Scutellaria Barbate (CE-SB) on mouse hepatoma H22 cells. The MTT assay was used to determine the growth inhibition of H22 cells in vitro. The in vivo therapeutic effects of CE-SB were determined using H22 tumor bearing mice. Besides, the body weight, tumor weight, thymus index and spleen index of H22 bearing mice were also measured. The tumor inhibitory rate (IR) was calculated according to the mean weight of tumor (MWT). The phagocytotic function of macrophages was examined by observing peritoneal macrophages phagocytize chicken RBC. The results showed that CE-SB could inhibit the growth of hepatoma H22 Cells in vitro and in vivo. Furthermore, CE-SB could improve immune function of H22 tumor bearing mice. Together these results indicate that CE-SB has antitumor activity and seems to be safe and effective for the use of anti-tumor therapy.
Molecules 01/2011; 16(6):4389-400. · 2.39 Impact Factor
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ABSTRACT: To investigate the role of mast cells and gut hormones and their interactions in TNBS-induced ulcerative colitis.
Rat models of ulcerative colitis were established by a single intracolonic injection of 100 mg/kg TNBS (in 0.3 ml 50% ethanol). At 0, 6, 11, 16, 21 days after TNBS injection, the rats were sacrificed to determine the count of the mast cells. Histamine level in the whole blood, and the levels of histamine, substance P (SP), vasoactive intestinal peptide (VIP), and somatostatin (SS) in the distal colons were measured by fluoremetry or radioimmunal assay. Immunofluorescence double staining was used to observe the relationship of the mast cells with SP, VIP, and SS positive nerve fibers.
On day 6 after TNBS injection, obvious ulcers occurred in the distal colon of the rats with significantly increased histamine level in the whole blood (P<0.05) but significantly decreased colonic histamine levels (P<0.05). The histamine levels in the whole blood and distal colon gradually recovered the normal levels. The mast cells significantly increased on day 16 (P<0.05) and maintained the high level till day 21. The distribution of mast cells was altered after TNBS injection, and the cells were found to aggregate in the myenteric region. SP levels in the distal colon significantly increased on day 11 (P<0.05) and maintained the high level till day 21. Immunofluorescence double staining revealed numerous mast cells close to the SP- and VIP-positive nerve fibers at different time points after TNBS injection. VIP positivivity and the number of VIP-positive nerve fibers in the myenteric region were markedly increased, but no mast cells were observed in association with SP- and VIP-positive nerve fibers. The distribution of MC was not found to associate with the SS-positive nerve fibers.
The mast cells and histamine released by them, as well as parasecretion of SP and VIP, participate in tissue damage by TNBS-induced colitis. Bidirectional neuroimmunomodulation of the mast cells, SP and VIP have important effect on the development of TNBS-induced colitis.
Nan fang yi ke da xue xue bao = Journal of Southern Medical University 08/2009; 29(7):1359-63.
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ABSTRACT: Matrine, an alkaloid purified from the chinese herb Sophora flavescens Ait, is well known to possess activities including anti-inflammation, anti-fibrotic and anticancer. In this study, the mechanism of matrine inducing the apoptosis of gastric carcinoma cells was investigated.
Proliferation of SGC-7901 cells was examined by MTT assay. Cellular morphology was observed under transmission electron microscope. Flow cytometry (FCM) was used to observe the apoptosis of SGC-7901 cells by staining with annexinV-FITC/PI. The expression levels of Fas/FasL in SGC-7901 cells were monitored by FCM analysis using an indirect immunofluorescence method. Activity of caspase-3 enzyme was measured by spectrofluorometry.
MTT assay showed that matrine inhibited SGC-7901 cells proliferation in a dose-dependent and time-dependent manner. Apoptosis induction was demonstrated by morphological changes under electron microscope and FCM analysis. Fluorescence intensity levels of Fas and FasL were found to be equally up-regulated after matrine treatment, which were both correlated with apoptosis rate. The activity of caspase-3 enzyme increased in matrine groups, positively correlated with apoptosis rate.
Matrine could inhibit cell proliferation and induce apoptosis of SGC-7901 cells in vitro. The apoptosis induction appears to proceed by up-regulating Fas/FasL expression and activating caspase-3 enzyme.
Journal of ethnopharmacology 06/2009; 123(1):91-6. · 2.32 Impact Factor
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ABSTRACT: To study the growth inhibitory and apoptotic effects of Scutellaria barbata D.Don (S. barbata) and to determine the underlying mechanism of its antitumor activity in mouse liver cancer cell line H22.
Proliferation of H22 cells was examined by MTT assay. Cellular morphology of PC-2 cells was observed under fluorescence microscope and transmission electron microscope (EM). Mitochondrial transmembrane potential was determined under laser scanning confocal microscope (LSCM) with rhodamine 123 staining. Flow cytometry was performed to analyze the cell cycle of H22 cells with propidium iodide staining. Protein level of cytochrome C and caspase-3 was measured by semi-quantitive RT-PCR and Western blot analysis. Activity of caspase-3 enzyme was measured by spectrofluorometry.
MTT assay showed that extracts from S. barbata (ESB) could inhibit the proliferation of H22 cells in a time-dependent manner. Among the various phases of cell cycle, the percentage of cells in S phase was significantly decreased, while the percentage of cells in G(1) phase was increased. Flow cytometry assay also showed that ESB had a positive effect on apoptosis. Typical apoptotic morphologies such as condensation and fragmentation of nuclei and blebbing membrane of apoptotic cells could be observed under transmission electron microscope and fluorescence microscope. To further investige the molecular mechanism behind ESB-induced apoptosis, ESB-treated cells rapidly lost their mitochondrial transmembrane potential, released mitochondrial cytochrome C into cytosol, and induced caspase-3 activity in a dose-dependent manner.
ESB can effectively inhibit the proliferation and induce apoptosis of H22 cells involving loss of mitochondrial transmembrane potential, release of cytochrome C, and activation of caspase-3.
World Journal of Gastroenterology 01/2009; 14(48):7321-8. · 2.47 Impact Factor
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ABSTRACT: To investigate the effects of serum containing Scutellaria Barbata extract (ESB) on apoptosis rate and mitochondrial transmembrane potential (MTP) of liver cancer cell line H22 from mice in vitro.
H22 cells were cultured in vitro and divided into 5 groups: blank control group, low-dose ESB group, medium-dose ESB group, high-dose ESB group and fluorouracil (5-Fu) group. Methyl thiazolyl tetrazolium assay was utilized to determine the proliferation rates of H22 cells. Cellular morphology was observed under a transmission electron microscope (EM). The rhodamine 123 was used as a fluorescence probe to label the H22 cells, and the fluorescence intensities were observed with a laser scanning confocal microscope. The fluorescence intensity of H22 cells indicated the MTP of H22 cells.
The inhibition of serum containing ESB on the proliferation of H22 cells in vitro was observed in a time-dependent manner. The typical morphological changes of apoptosis were observed after incubation with ESB-containing serum in high dose for 48h. The apoptosis rates of blank control group, 5-Fu group, low-dose ESB group, medium-dose ESB group and high-dose ESB group were (0.51+/-0.32)%, (11.26+/-2.97)%, (1.07+/-0.46)%, (3.15+/-1.12)%, (7.83+/-2.25)% respectively. ESB could reduce the MTP of H22 cells from mice as compared with the untreated group. The MTPs of the blank control group, 5-Fu group, and low-, medium- and high-dose ESB groups were (245.45+/-67.37), (127.42+/-41.35), (213.68+/-65.52), (186.34+/-56.37) and (142.65+/-39.44) respectively, which were negatively correlated with the apoptosis rates.
ESB-containing serum effectively induces apoptosis, which may be related to the decrease of MTP in H22 cells.
Journal of Chinese Integrative Medicine 09/2008; 6(8):821-6.
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ABSTRACT: To investigate the mechanism of gastric carcinoma cells apoptosis induced by matrine injection in vitro.
Effects of 24, 48, 72, 96 h incubation with different concentrations (0.25-1.5 g/L) of matrine injection on proliferation of SGC-7901 cells were evaluated using 3-4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT) assay. The cellular morphology of SGC-7901 cells was observed by transmission electron microscope (EM). Flow cytometry was used to analyze the apoptosis of SGC-7901 cells by staining with annexin V-FITC/PI. The expression of Fas/FasL was examined by flow cytometry using specific antibody. The activity of caspase-3 was measured by spectrofluorometry.
Matrine injection could inhibit the proliferation of SGC-7901 cells in a dose- and time-dependent manner. The typical morphological changes of apoptosis were observed after incubation with 1.0 g/L matrine injections for 48 h. The apoptosis rates of 0.5 g/L, 1.0 g/L and 1.5 g/L groups were 39.80%, 58.11% and 79.00% respectively. The apoptotic cells in matrine injection group were mainly early apoptotic cells, and those in 5-FU group were mainly late apoptotic cells and necrotic cells. Spectrofluorometry revealed FI levels of Fas and FasL were equal, which were both correlated with apoptosis rate. The activity of caspase-3 increased with the elevation of matrine concentration, and was correlated with the apoptosis rate.
Matrine injection can induce apoptosis of SGC-7901 cells through the up-regulation of Fas/FasL expression and activation of caspase-3.
Zhonghua wei chang wai ke za zhi = Chinese journal of gastrointestinal surgery 06/2008; 11(3):261-5.
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ABSTRACT: To investigate the effects of Scutellaria barbate extract (ESB) on suppressing proliferation and inducing apoptosis of mouse hepatoma H22 cells.
H22 cells cultured in vitro were divided into 5 groups: blank control group, ESB in high, medium, low dose groups and 5-Fu group. H22 cells were cultured in media with serum containing different concentrations of ESB and blank serum. The proliferation of H22 cells was determined by microculture tetrazolium (MTT) assay. Fluorescence microscopy was utilized to observe the apoptosis of H22 cells by staining with Hoechst 33258. The cell cycle and apoptosis were analyzed by flow cytometry (FCM).
The inhibition of serum containing ESB on the proliferation of H22 cells in vitro was observerd in a dose and time dependent manner. The typical morphological changes of apoptosis were observed after incubation with ESB-containing serum in high dose for 48 hours. Among the various phases of cell cycle, the percentage of cells in S phase decreased significantly, while the percentage of cells in G1 phase increased. Drug-containing serum showed positive effect on cell apoptosis. The apoptosis rate of blank control group, ESB in low, medium, high dose groups and 5-Fu group were 0.51%, 1.07%, 3.15%, 7.83%, 11.26%, respectively.
ESB containing serum can inhibit proliferation and induce apoptosis of H22 cells in vitro.
Zhong yao cai = Zhongyaocai = Journal of Chinese medicinal materials 04/2008; 31(4):550-3.
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ABSTRACT: To construct an expression vector of small interfering RNA (siRNA) against survivin and observe its effects on survivin expression and proliferation of human pancreatic cancer cell line PC-2 and breast cancer cell line MCF-7.
Constructed an expression vector of siRNA against survivin and transfected it into PC-2 and MCF-7 cells using lipofectamine 2000. The expression of survivin was detected by semi-quantitive RT-PCR and immunohistochemistry, and its effects on proliferation of PC-2 and MCF-7 cells were detected by MTT assay.
The introduction of sequence-specific siRNA could efficiently suppress survivin expression at both mRNA and protein levels in the two cancer cell lines. In PC-2 cells, the expression inhibition rates were 81.25% at mRNA level and 74.24% at protein level. In MCF-7 cells, the expression inhibition rates were 64.91% at mRNA level and 79.72% at protein level. The proliferation of PC-2 and MCF-7 cells was also suppressed, and 24 and 48 hours after the cells were reseeded, the proliferation inhibition rates of PC-2 cells were 28.00% and 33.38%, and that of MCF-7 cells were 31.58% and 33.02%, respectively.
The expression vector of siRNA against survivin can block survivin expression in PC-2 and MCF-7 cells efficiently and specifically. Down regulation of survivin expression can suppress proliferation of PC-2 and MCF-7 cells. Survivin RNAi may have potential value in gene therapy of human cancers.
Chinese Medical Sciences Journal 07/2006; 21(2):115-9.
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ABSTRACT: Blocking the expression of survivin with RNA interference techniques, the effects of suppressing proliferation and inducing apoptosis of breast cancer MCF-7 cells were investigated.
A siRNA eukaryotic expression vector against survivin was constructed and transfected into breast cancer MCF-7 cells with lipofectamine 2000. The changes of survivin expression were detected by semi-quantitive RT-PCR and immunohistochemistry. The effect of suppressing proliferation of MCF-7 cell was detected by MTT assay. The effect of inducing MCF-7 cell apoptosis was detected by TUNEL assay.
The sequence-specific siRNA can efficiently block the expression of survivin both at mRNA and protein levels. The expression inhibition rate was 64.9% at mRNA level detected by semi-quantitive RT-PCR and 79.7% at protein level detected by immunohistochemistry. Blocking the expression of survivin can suppress proliferation of MCF-7 cells significantly. At 24 and 48 h after the cells were reseeded, the proliferation inhibition rates were 31.6% and 33.0%, respectively. At 24 h after transfection, apoptosis was induced in 12.9% of the cells as detected by TUNEL assay.
Blocking the expression of survivin with RNA interference technology can significantly suppress proliferation of MCF-7 cells and induce apoptosis to a certain degree. RNAi targeted to survivin has a potential value in gene therapy of breast cancer.
Zhonghua zhong liu za zhi [Chinese journal of oncology] 06/2006; 28(5):326-30.
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ABSTRACT: To investigate the inhibitory effect of small interfering RNA (siRNA) on the expression of survivin in pancreatic cancer cell line PC-2 and the role of siRNA in inducing PC-2 cell apoptosis and enhancing its radiosensitivity.
A siRNA plasmid expression vector against survivin was constructed and transfected into PC-2 cells with Lipofectamine 2000. The down regulation of survivin expression was detected by semi-quantitative RT-PCR and immunohistochemical SP method and the role of siRNA in inducing PC-2 cell apoptosis and enhancing its radiosensitivity was detected by flow cytometry.
The sequence-specific siRNA efficiently and specifically down-regulated the expression of survivin at both mRNA and protein levels. The expression inhibition ratio was 81.25% at mRNA level detected by semi-quantitative RT-PCR and 74.24% at protein level detected by immunohistochemical method. Forty-eight hours after transfection,apoptosis was induced in 7.03% cells by siRNA and in 14.58% cells by siRNA combined with radiation.
The siRNA plasmid expression vector against survivin can inhibit the expression of survivin in PC-2 cells efficiently and specifically. Inhibiting the expression of survivin can induce apoptosis of PC-2 cells and enhance its radiosensitivity significantly. RNAi against survivin is of potential value in gene therapy of pancreatic cancer.
World Journal of Gastroenterology 06/2006; 12(18):2901-7. · 2.47 Impact Factor
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ABSTRACT: To use the sequence-specific siRNA knocking down the expressions of Survivin gene and inducing breast cancer MCF-7 cell line to apoptosis, and to couple the siRNA with Survivin for investigating the effects of MCF-7 cell induced to apoptosis and the chemotherapy sensitivity of breast cancer cell treated to epirubicin.
The molecular cloning technique was applied to construct the eukaryotic expression vector of siRNA against Survivin, and lipofectamine 2000 was used to transfect MCF-7 cell. Survivin expressions were detected by semi-quantitive RT-PCR and immunohistochemical SABC methods. The effects of inducing MCF-7 cell apoptosis and enhanced chemotherapy sensitivity to epirubicin were assessed by TUNEL method.
The sequence-specific siRNA can, effectively and specifically, knock the expressions of Survivin gene down at both mRNA and protein levels, in which the expression inhibition rates were 64.91 and 79.72% respectively. After 48 h, 8.75% cells transfected with siRNA expression vector were induced to apoptosis; Coupling siRNA against Survivin with epirubicin can induce the cell apoptosis rate up to 24.21%.
In the study, the siRNA against Survivin can, effectively and specifically, decrease the expressions of Survivin gene in MCF-7 cell; blocking the expressions of Survivin can, in certain degree, induce MCF-7 cell to apoptosis and enhance cell chemotherapy sensitivity to epirubicin significantly; Survivin RNAi has a great potential value in the gene therapy of breast cancer.
Sichuan da xue xue bao. Yi xue ban = Journal of Sichuan University. Medical science edition 04/2006; 37(2):221-5.
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ABSTRACT: To investigate the effect of a sequence-specific small interfering RNA (siRNA) in suppressing survivin expression and cell proliferation and inducing apoptosis of PC-2 cells.
The plasmid expression vector of siRNA targeted against survivin was constructed and transfected into PC-2 cells with Lipofectamine 2000. The changes of survivin expression were detected by semi-quantitative RT-PCR and immunohistochemical SP methods. The effect of siRNA in suppressing the proliferation of PC-2 cells was detected by MTT assay, and its role in inducing PC-2 cell apoptosis evaluated by flow cytometry.
The sequence-specific siRNA effectively suppressed survivin expression at both mRNA and protein levels with inhibition rate of 81.25% at mRNA level and 74.24% at protein level. Survivin expression suppression significantly inhibited the proliferation of PC-2 cells, and at 24 and 48 h after cell seeding, the proliferation inhibition rate was 28.00% and 33.38% respectively; 24, 48 h after the transfection, apoptosis occurred in 8.46% and 7.53% of the cells, respectively.
The plasmid expression vector for the siRNA against survivin constructed in the study can effectively and specifically suppress survivin expression in PC-2 cells, and blocking survivin expression suppresses PC-2 cell proliferation and induces cell apoptosis. siRNA targeted against survivin has a potential value in gene therapy for pancreatic cancer.
Nan fang yi ke da xue xue bao = Journal of Southern Medical University 03/2006; 26(2):169-73.
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ABSTRACT: Objective: To study the relationship between cyclooxygenase-2 (COX-2) expression and tumor angiogenesis in human breast cancer.
Methods: Archival primary breast carcinomas (n=62), adjacent ductal carcinomain situ (DCIS, n=13) and DCIS alone (n=5) were analyzed for COX-2 and VEGF expression by immunohistochemistry using specific monoclonal
antibodies. Microvessel density (MVD) was also examined the using CD34 staining. Results: A significant correlation was found
between COX-2 and VEGF expression (P<0.01). Both COX-2 and VEGF were significantly correlated with MVD (P<0.05) andP<0.01, respectively). COX-2 and VEGF genes were overexpressed in tumor specimens as compared with normal epithelia. Conclusion:
COX-2 is related to tumor angiogenesis in breast cancer. It is likely that VEGF is one of the most important mediators of
the COX-2 angiogenic pathway.
Chinese Journal of Cancer Research 05/2003; 15(2):149-151. · 0.18 Impact Factor
