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On-site flexible sigmoidoscopy (FS) positively influences colorectal cancer screening practices by primary care providers (PCPS)
A676 AGA ABSTRACTS
• G2790 ON-SITE FLEXIBLE SIGMOIDOSCOPY (FS) POSITIVELY INFLUENCES COLORECTAL CANCER SCREENING PRACTICES BY PRIMARY CARE PROVIDERS (PCPS). Schrov PC, Prout M, Bliss CM, Bliss CM Jr, Pincus J, Wilson S, Herren T. Boston University Schools of Medicine and Public Health. Boston, MA. FS is a cost-effective screening strategy for colorectal cancer that is neither widely utilized nor recommended by PCPs. To address this issue, we have developed a model for primary care-directed community-based coloreetal cancer screening that incorporates an on-site practice-oriented provider educational program. The model commences with the implementation of onsite FS screening services initially provided by gastroenterologists (Phase 1) and culminates with a self-standing screening FS program staffed by PCPs (Phase 3). The aim of this study was to assess the impact of Phase 1 alone on PCP compliance with American Cancer Society (ACS) screening guidelines. Methods: A controlled trial was initiated at nine urban community health centers (CHCs). Four CHCs were selected to serve as intervention sites and 5 CHCs as control sites. The two groups were balanced with respect to annual adult patient visits, patient demographics, provider demographics, and on-site resources. Baseline data on provider attitudes and practice patterns were collected using a validated questionnaire just prior to a didactic educational seminar at all CHCs. Phase 1 was initiated within the ensuing 3 months at each of the intervention CHCs only. Outcome measures included a Year 1 follow-up survey of provider attitudes and quarterly review of sigmoidoscopy referrals to assess practice patterns. Results: Eighty percent (n=49) of the targeted providers at the intervention CHCs and 79% (n=21) of those at the control CHCs completed the baseline survey. No significant differences in self-reported screening behavior were observed between the two groups: overall, almost all providers claimed that they follow current ACS recommendations for digital rectal examinations (intervention, 98% vs. control, 95%) and fecal occult blood testing (94% vs 95%) in average risk individuals; in contrast, relatively few (22% vs 19%) recommended FS. Significant differences were noted at the Year 1 follow-up survey. Response rates were again excellent: 89% (n=51) at the intervention sites and 89% (n=24) at the control sites. Whereas compliance remained high and comparable for digital rectal examinations (98% vs 95%) and fecal occult blood testing (98% vs 95%), compliance rates for FS were significantly higher at the intervention sites (67% vs 32 %; P=.005). When stratified by site, compliance rates increased at each of the intervention sites but only one of the control sites. Importantly, concordance between self-reported compliance rates and referral patterns utilizing FS appointment logs was > 90%. Univariate analysis of Year 1 data identified concerns about cost, skills required, lack of proven efficacy, and low yield as significant factors for noncompliance. Conclusions: Implementation of on-site FS services significantly increases PCP compliance with ACS screening guidelines. We speculate that on-site FS provides a cue-to-action that supercedes PCP concerns about cost, skills required, lack of efficacy, and low yield. This research was supported by NIH/NCI grant KO7-CA68058 (PS). • G2791 EVIDENCE FOR THE PRESENCE OF 5-HT3 RECEPTOR ON THE HUMAN ADENOCARCINOMA TUMOUR CELL LINE COLO 320 DM AND FUNCTIONAL CONSEQUENCES OF ITS ACTIVATION. H. Schwtirer, D. Raddatz. G. Ramadori. Department of Internal Medicine, Division of Gastroenterologyand Endocrinology, University of G6ttingen, Germany There is evidence that serotonin (5-hydroxytryptamine, 5-HT) has growth stimulatory effects (proliferation and mitogenesis) in different cells. A positive autoregulation of 5-HT release from enterochromaffin cells of the intestinal mucosa by the activation of 5-HT3 receptors, an ion channel, was reported in guinea pig and rat. There are only few cell lines which contain 5-HT. The COLO 320 DM cell line was derived from a human colonic adenocarcinoma, which contains 5-HT. We investigated the presence of 5-HT3 receptors in COLO 320 DM cells by reverse transcription polymerase chain reaction (RT-PCR). Furthermore we studied the effect of 5-HT3 receptor activation on 5-HT release from COLO 320 DM cells as well as the effect of 5-HT 3 receptor activation on 5-bromo-2'-deoxy-uridin (BrdU) incorporation, a measure for DNA synthesis. The presence of 5-HT3 receptor-mRNA on COLO 320 DM cells could be confirmed by RT-PCR with 5-HT3 receptor specific primers. The 5-HT 3 receptor agonist 2-methyl-5-HT (1-100 gmol/1) increased the outflow of 5-HT by maximally 190%. This effect was antagonized by the 5-HT 3 receptor antagonist tropisetron (30 nmol/1). 2-methyl-5-HT increased the incorporation of BrdU by 50%. This effect was also antagonized by tropisetron. The present results clearly demonstrate in a human tumour cell line that activation of 5-HT3 receptors is coupled to an increase of 5-HT outflow from these cells as well as to an increase of DNA synthesis as indicated by the increased incorporation of BrdU labelling of the cells. In conclusion, the human colonic tumour cell line COLO 320 DM is a valuable tool for further research on these 5-HT receptors on human cells in vitro. Furthermore these results suggest that the biogenic amine 5-HT might be involved in autocrine growth mechanisms in colonic carcinoma. This would imply interesting therapeutic consequences e.g. treatment with 5-HT3 receptor antagonists.
GASTROENTEROLOGY Vol. 114, No. 4
• G2792 REDUCED APOPTOSIS IN H. PYLORI-ASSOCIATED GASTRIC INTESTINAL METAPLASIA: A FACTOR IN GASTRIC CARCINOGENESIS? IA Seetiniotis 1,4, T Rokkas 2, EE Furth3, JW Plotkin4, B Rigas 4, SJ Shill~ lDivision of Gastroenterology and 3Department of Pathology, Hospital of the University of Pennsylvania, Philadelphia, PA, 2401 General Military Hospital, Athens, Greece; 4The Rockefeller University, New York, NY. BACKGROUND: The mechanisms whereby H. pylori (Hp) infection induces intestinal metaplasia, a harbinger of gastric dysplasia and adenocarcinoma, are unknown. Apoptosis is important in GI epithelial cell turnover and carcinogenesis. We investigated the role of Hp in gastric carcinogenesis by comparing epithelial cell apoptosis in the gastric pits in antral biopsies from: a) normals (n=l 1); b) those with Hp-associated gastritis (n=22); and c) those with Hp-associated gastritis and fuci of intestinal metaplasia (n=9). METHODS: Antral biopsies from 42 patients (31 M, ages 18-78; 11 F, ages 28-59), examined endoscopically for dyspeptic symptoms, were reviewed by one pathologist. Acute and chronic inflammation was graded 1-3 according to the Updated Sydney Scale. A cumulative inflammatory score, graded 1-6, was the sum of the two: mild gastritis=l-2; moderate=3-4; and marked=5-6. The Hp status of each patient was determined by CLO testing and thiazine staining. Apoptosis was assessed by TUNEL staining. 1000 epithelial cells were counted from gastric pits in each patient and the apoptotic index (AI), the percentage of epithelial cells with TUNEL-positive nuclei, was determined. In biopsies with intestinal metaplasia, separate measurements were obtained from glands with and without metaplasia. RESULTS: A) Hp status: normal biopsies, all (-); gastritis ( _+ metaplasia), all (+). B) AI: a) normals (nl) (mean _+SEM) =1.5 x 10-3 _+0.3 x 10-3; b) mild gastritis (n=10) 6.6 x 10-3 + 1.8 x 10-3; c) moderate gastritis (n=16) = 14.3 x 10.3 __.1.7 x 10-3; d) marked gastritis (n=5) 16.6 x 10"3 - 1.6 x 10-3, e) all gastritis= 12.2 x 10.3 _ 1.3 x 10-3; f) intestinal metaplasia =1.96 x 10-3 _+0.6 x 10-3; g) gastritis with intestinal metaplasia (n=9) = 13.4 x 10.3 -+ 2.3 x 10"3. AI was greater in gastritis specimens (8-fold nl vs. all gastritis, P<0.0001; 4-fold nl vs. mild gastritis, P<0.009; 11-fold nl vs. marked gastritis, P<0.0001). The AI increases with the severity of lip-associated gastritis (mild gastritis vs. moderate and mild vs. marked, P<0.05; moderate vs. marked, P=0.05). In gastric metaplasia, the AI was 6-fold less than in areas of gastritis (f vs. e and f vs. g, P<0.002) and approached that seen in normal mueosa (P=0.43). CONCLUSIONS: In Hp-infected antral biopsy samples from dyspeptic subjects: 1) gastric epithelial apoptosis increases in parallel to the degree of gastric mucosal inflammation; 2) significantly less epithelial apoptosis is seen in Hp-associated intestinal metaplasia than in lip-associated gastritis, regardless of severity. It is plausible that changes in apoptosis in the gastric mucosa may contribute to in the pathogenesis of gastritis (increased apoptosis) or gastric intestinal metaplasia (decreased apoptosis) and ultimately to Hp-associated gastric cancer. Research is funded by the Arthur & Rochelle Belfer Foundation. • G2793 INTERLEUKIN-10 (IL-10) RESULTS IN LOSS OF NEURAL CELL ADHESION MOLECULE (NCAM) EXPRESSION IN COLON CARCINOMA. J. Sebastian, S. Huerta, T. Blinman, EH Livingston. Surgical Molecular Biology Research Laboratory, VAMC West Los Angeles/UCLA School of Medicine Division of General Surgery. Los Angeles, California. Objective: To determine what role cytokines may play in the progression of colon cancer cell phenotype. Using semi-quantitative RT-PCR, we previously screened two colon cancer cell lines obtained from the same patient (SW 480 and SW620) for cytokine expression. Although both cell lines expressed IL-6 and IL-8 in near equal amounts, there was a dramatic difference in IL-10 expression, with much greater expression in the SW 620 cell line. This cell line is known clinically to have possessed a more aggressive phenotype, with metastatic behavior. Furthermore, we known from prior work that this cell line also lacks Neural Cell Adhesion Molecule (NCAM) expression. Although it is known that IL-10 plays an immunosuppressive role in the progression of certain cancer types, we hypothesize that IL-10 also influences colon cancer cell phenotype, namely that of NCAM expression. Methods: The SW 480 (CCL 228) human colon cancer cell line was obtained from the American Type Culture Collection (ATCC). We have previously demonstrated that the NCAM-180 isoform is expressed in this cell line. Cells were seeded in flasks and cell cycle synchronization was achieved with serum deprivation for a period of 12 hours. Cells were then treated with increasing doses of hIL-10 for a period of 48 hours (n=3). Protein extraction was performed for Western blot analysis, looking for the expression of the NCAM - 180 isoform. Results: Untreated cells exhibited strong expression of the NCAM-180 isoform, whereas, cells treated with IL-10 appeared to have progressive loss of the protein in a dose-dependant fashion. At the highest concentration of hlL-10, there appeared to be complete loss of NCAM expression. Conclusion: Although IL-10 is known to possess important properties allowing tumor ceils to escape immunosurveillance, our results suggest an additional mechanism whereby this cytokine may facilitate the progression of colon carcinoma by down-regulating expression of NCAM. We have previously shown that NCAM has important tumor suppressor activity, and its
• G2790 ON-SITE FLEXIBLE SIGMOIDOSCOPY (FS) POSITIVELY INFLUENCES COLORECTAL CANCER SCREENING PRACTICES BY PRIMARY CARE PROVIDERS (PCPS). Schrov PC, Prout M, Bliss CM, Bliss CM Jr, Pincus J, Wilson S, Herren T. Boston University Schools of Medicine and Public Health. Boston, MA. FS is a cost-effective screening strategy for colorectal cancer that is neither widely utilized nor recommended by PCPs. To address this issue, we have developed a model for primary care-directed community-based coloreetal cancer screening that incorporates an on-site practice-oriented provider educational program. The model commences with the implementation of onsite FS screening services initially provided by gastroenterologists (Phase 1) and culminates with a self-standing screening FS program staffed by PCPs (Phase 3). The aim of this study was to assess the impact of Phase 1 alone on PCP compliance with American Cancer Society (ACS) screening guidelines. Methods: A controlled trial was initiated at nine urban community health centers (CHCs). Four CHCs were selected to serve as intervention sites and 5 CHCs as control sites. The two groups were balanced with respect to annual adult patient visits, patient demographics, provider demographics, and on-site resources. Baseline data on provider attitudes and practice patterns were collected using a validated questionnaire just prior to a didactic educational seminar at all CHCs. Phase 1 was initiated within the ensuing 3 months at each of the intervention CHCs only. Outcome measures included a Year 1 follow-up survey of provider attitudes and quarterly review of sigmoidoscopy referrals to assess practice patterns. Results: Eighty percent (n=49) of the targeted providers at the intervention CHCs and 79% (n=21) of those at the control CHCs completed the baseline survey. No significant differences in self-reported screening behavior were observed between the two groups: overall, almost all providers claimed that they follow current ACS recommendations for digital rectal examinations (intervention, 98% vs. control, 95%) and fecal occult blood testing (94% vs 95%) in average risk individuals; in contrast, relatively few (22% vs 19%) recommended FS. Significant differences were noted at the Year 1 follow-up survey. Response rates were again excellent: 89% (n=51) at the intervention sites and 89% (n=24) at the control sites. Whereas compliance remained high and comparable for digital rectal examinations (98% vs 95%) and fecal occult blood testing (98% vs 95%), compliance rates for FS were significantly higher at the intervention sites (67% vs 32 %; P=.005). When stratified by site, compliance rates increased at each of the intervention sites but only one of the control sites. Importantly, concordance between self-reported compliance rates and referral patterns utilizing FS appointment logs was > 90%. Univariate analysis of Year 1 data identified concerns about cost, skills required, lack of proven efficacy, and low yield as significant factors for noncompliance. Conclusions: Implementation of on-site FS services significantly increases PCP compliance with ACS screening guidelines. We speculate that on-site FS provides a cue-to-action that supercedes PCP concerns about cost, skills required, lack of efficacy, and low yield. This research was supported by NIH/NCI grant KO7-CA68058 (PS). • G2791 EVIDENCE FOR THE PRESENCE OF 5-HT3 RECEPTOR ON THE HUMAN ADENOCARCINOMA TUMOUR CELL LINE COLO 320 DM AND FUNCTIONAL CONSEQUENCES OF ITS ACTIVATION. H. Schwtirer, D. Raddatz. G. Ramadori. Department of Internal Medicine, Division of Gastroenterologyand Endocrinology, University of G6ttingen, Germany There is evidence that serotonin (5-hydroxytryptamine, 5-HT) has growth stimulatory effects (proliferation and mitogenesis) in different cells. A positive autoregulation of 5-HT release from enterochromaffin cells of the intestinal mucosa by the activation of 5-HT3 receptors, an ion channel, was reported in guinea pig and rat. There are only few cell lines which contain 5-HT. The COLO 320 DM cell line was derived from a human colonic adenocarcinoma, which contains 5-HT. We investigated the presence of 5-HT3 receptors in COLO 320 DM cells by reverse transcription polymerase chain reaction (RT-PCR). Furthermore we studied the effect of 5-HT3 receptor activation on 5-HT release from COLO 320 DM cells as well as the effect of 5-HT 3 receptor activation on 5-bromo-2'-deoxy-uridin (BrdU) incorporation, a measure for DNA synthesis. The presence of 5-HT3 receptor-mRNA on COLO 320 DM cells could be confirmed by RT-PCR with 5-HT3 receptor specific primers. The 5-HT 3 receptor agonist 2-methyl-5-HT (1-100 gmol/1) increased the outflow of 5-HT by maximally 190%. This effect was antagonized by the 5-HT 3 receptor antagonist tropisetron (30 nmol/1). 2-methyl-5-HT increased the incorporation of BrdU by 50%. This effect was also antagonized by tropisetron. The present results clearly demonstrate in a human tumour cell line that activation of 5-HT3 receptors is coupled to an increase of 5-HT outflow from these cells as well as to an increase of DNA synthesis as indicated by the increased incorporation of BrdU labelling of the cells. In conclusion, the human colonic tumour cell line COLO 320 DM is a valuable tool for further research on these 5-HT receptors on human cells in vitro. Furthermore these results suggest that the biogenic amine 5-HT might be involved in autocrine growth mechanisms in colonic carcinoma. This would imply interesting therapeutic consequences e.g. treatment with 5-HT3 receptor antagonists.
GASTROENTEROLOGY Vol. 114, No. 4
• G2792 REDUCED APOPTOSIS IN H. PYLORI-ASSOCIATED GASTRIC INTESTINAL METAPLASIA: A FACTOR IN GASTRIC CARCINOGENESIS? IA Seetiniotis 1,4, T Rokkas 2, EE Furth3, JW Plotkin4, B Rigas 4, SJ Shill~ lDivision of Gastroenterology and 3Department of Pathology, Hospital of the University of Pennsylvania, Philadelphia, PA, 2401 General Military Hospital, Athens, Greece; 4The Rockefeller University, New York, NY. BACKGROUND: The mechanisms whereby H. pylori (Hp) infection induces intestinal metaplasia, a harbinger of gastric dysplasia and adenocarcinoma, are unknown. Apoptosis is important in GI epithelial cell turnover and carcinogenesis. We investigated the role of Hp in gastric carcinogenesis by comparing epithelial cell apoptosis in the gastric pits in antral biopsies from: a) normals (n=l 1); b) those with Hp-associated gastritis (n=22); and c) those with Hp-associated gastritis and fuci of intestinal metaplasia (n=9). METHODS: Antral biopsies from 42 patients (31 M, ages 18-78; 11 F, ages 28-59), examined endoscopically for dyspeptic symptoms, were reviewed by one pathologist. Acute and chronic inflammation was graded 1-3 according to the Updated Sydney Scale. A cumulative inflammatory score, graded 1-6, was the sum of the two: mild gastritis=l-2; moderate=3-4; and marked=5-6. The Hp status of each patient was determined by CLO testing and thiazine staining. Apoptosis was assessed by TUNEL staining. 1000 epithelial cells were counted from gastric pits in each patient and the apoptotic index (AI), the percentage of epithelial cells with TUNEL-positive nuclei, was determined. In biopsies with intestinal metaplasia, separate measurements were obtained from glands with and without metaplasia. RESULTS: A) Hp status: normal biopsies, all (-); gastritis ( _+ metaplasia), all (+). B) AI: a) normals (nl) (mean _+SEM) =1.5 x 10-3 _+0.3 x 10-3; b) mild gastritis (n=10) 6.6 x 10-3 + 1.8 x 10-3; c) moderate gastritis (n=16) = 14.3 x 10.3 __.1.7 x 10-3; d) marked gastritis (n=5) 16.6 x 10"3 - 1.6 x 10-3, e) all gastritis= 12.2 x 10.3 _ 1.3 x 10-3; f) intestinal metaplasia =1.96 x 10-3 _+0.6 x 10-3; g) gastritis with intestinal metaplasia (n=9) = 13.4 x 10.3 -+ 2.3 x 10"3. AI was greater in gastritis specimens (8-fold nl vs. all gastritis, P<0.0001; 4-fold nl vs. mild gastritis, P<0.009; 11-fold nl vs. marked gastritis, P<0.0001). The AI increases with the severity of lip-associated gastritis (mild gastritis vs. moderate and mild vs. marked, P<0.05; moderate vs. marked, P=0.05). In gastric metaplasia, the AI was 6-fold less than in areas of gastritis (f vs. e and f vs. g, P<0.002) and approached that seen in normal mueosa (P=0.43). CONCLUSIONS: In Hp-infected antral biopsy samples from dyspeptic subjects: 1) gastric epithelial apoptosis increases in parallel to the degree of gastric mucosal inflammation; 2) significantly less epithelial apoptosis is seen in Hp-associated intestinal metaplasia than in lip-associated gastritis, regardless of severity. It is plausible that changes in apoptosis in the gastric mucosa may contribute to in the pathogenesis of gastritis (increased apoptosis) or gastric intestinal metaplasia (decreased apoptosis) and ultimately to Hp-associated gastric cancer. Research is funded by the Arthur & Rochelle Belfer Foundation. • G2793 INTERLEUKIN-10 (IL-10) RESULTS IN LOSS OF NEURAL CELL ADHESION MOLECULE (NCAM) EXPRESSION IN COLON CARCINOMA. J. Sebastian, S. Huerta, T. Blinman, EH Livingston. Surgical Molecular Biology Research Laboratory, VAMC West Los Angeles/UCLA School of Medicine Division of General Surgery. Los Angeles, California. Objective: To determine what role cytokines may play in the progression of colon cancer cell phenotype. Using semi-quantitative RT-PCR, we previously screened two colon cancer cell lines obtained from the same patient (SW 480 and SW620) for cytokine expression. Although both cell lines expressed IL-6 and IL-8 in near equal amounts, there was a dramatic difference in IL-10 expression, with much greater expression in the SW 620 cell line. This cell line is known clinically to have possessed a more aggressive phenotype, with metastatic behavior. Furthermore, we known from prior work that this cell line also lacks Neural Cell Adhesion Molecule (NCAM) expression. Although it is known that IL-10 plays an immunosuppressive role in the progression of certain cancer types, we hypothesize that IL-10 also influences colon cancer cell phenotype, namely that of NCAM expression. Methods: The SW 480 (CCL 228) human colon cancer cell line was obtained from the American Type Culture Collection (ATCC). We have previously demonstrated that the NCAM-180 isoform is expressed in this cell line. Cells were seeded in flasks and cell cycle synchronization was achieved with serum deprivation for a period of 12 hours. Cells were then treated with increasing doses of hIL-10 for a period of 48 hours (n=3). Protein extraction was performed for Western blot analysis, looking for the expression of the NCAM - 180 isoform. Results: Untreated cells exhibited strong expression of the NCAM-180 isoform, whereas, cells treated with IL-10 appeared to have progressive loss of the protein in a dose-dependant fashion. At the highest concentration of hlL-10, there appeared to be complete loss of NCAM expression. Conclusion: Although IL-10 is known to possess important properties allowing tumor ceils to escape immunosurveillance, our results suggest an additional mechanism whereby this cytokine may facilitate the progression of colon carcinoma by down-regulating expression of NCAM. We have previously shown that NCAM has important tumor suppressor activity, and its