Department of Surgery, College of Human Medicine, Michigan State University, East Lansing2Research Service, John D. Dingell Veterans Affairs Medical Center, Detroit, Michigan3Department of Anesthesiology, Wayne State University, Detroit, Michigan.
JAMA Surg. 2013 Nov;148(11):1037-42. doi: 10.1001/jamasurg.2013.3731.
Short bowel syndrome occurs when a shortened intestine cannot absorb sufficient nutrients or fluids. Teduglutide is a recombinant analogue of human glucagonlike peptide 2 that reduces dependence on parenteral nutrition in patients with short bowel syndrome by promoting enterocytic proliferation, increasing the absorptive surface area. However, enterocyte function depends not only on the number of cells that are present but also on differentiated features that facilitate nutrient absorption and digestion.
To test the hypothesis that teduglutide impairs human intestinal epithelial differentiation.
We investigated the effects of teduglutide in the modulation of proliferation and differentiation in human Caco-2 intestinal epithelial cells at a basic science laboratory. This was an in vitro study using Caco-2 cells, a human-derived intestinal epithelial cell line commonly used to model enterocytic biology.
Cells were exposed to teduglutide or vehicle control.
We analyzed the cell cycle by bromodeoxyuridine incorporation or propidium iodide staining and flow cytometry and measured cell proliferation by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay. We used quantitative reverse transcription-polymerase chain reaction to assay the expression of the enterocytic differentiation markers villin, sucrase-isomaltase, glucose transporter 2 (GLUT2), and dipeptidyl peptidase 4 (DPP-4), as well as that of the putative differentiation signals schlafen 12 (SLFN12) and caudal-related homeobox intestine-specific transcription factor (Cdx2). Villin promoter activity was measured by a luciferase-based assay.
The MTS assay demonstrated that teduglutide increased cell numbers by a mean (SD) of 10% (2%) over untreated controls at a maximal 500 nM (n = 6, P < .05). Teduglutide increased bromodeoxyuridine-positive cells vs untreated controls by a mean (SD) of 19.4% (2.3%) vs 12.0% (0.8%) (n = 6, P < .05) and increased the S-phase fraction by flow cytometric analysis. Teduglutide reduced the mean (SD) expression of villin by 29% (6%), Cdx2 by 31% (10%), DPP-4 by 15% (6%), GLUT2 by 40% (11%), SLFN12 by 61% (14%), and sucrase-isomaltase by 28% (8%) (n = 6, P < .05 for all).
Teduglutide increased Caco-2 proliferation but tended to inhibit intestinal epithelial differentiation. The effects of mitogenic stimulation with teduglutide in patients with short bowel syndrome might be greater if the more numerous teduglutide-treated cells could be stimulated toward a more fully differentiated phenotype.
当肠缩短导致无法吸收足够的营养或液体时，就会发生短肠综合征。特杜格鲁肽是一种人胰高血糖素样肽 2 的重组类似物，通过促进肠细胞增殖，增加吸收表面积，减少短肠综合征患者对肠外营养的依赖。然而，肠细胞的功能不仅取决于存在的细胞数量，还取决于促进营养吸收和消化的分化特征。
检验特杜格鲁肽损害人类肠道上皮细胞分化的假说。
我们在基础科学实验室研究了特杜格鲁肽在调节人 Caco-2 肠上皮细胞增殖和分化中的作用。这是一项在体外研究，使用 Caco-2 细胞，一种常用于模拟肠细胞生物学的人源肠上皮细胞系。
细胞暴露于特杜格鲁肽或载体对照。
我们通过溴脱氧尿苷掺入或碘化丙啶染色和流式细胞术分析细胞周期，并通过 3-(4,5-二甲基噻唑-2-基)-5-(3-羧甲氧基苯基)-2-(4-磺苯基)-2H-四唑 (MTS) 测定法测量细胞增殖。我们使用定量逆转录聚合酶链反应来检测肠细胞分化标志物绒毛蛋白、蔗糖酶-异麦芽糖酶、葡萄糖转运蛋白 2 (GLUT2) 和二肽基肽酶 4 (DPP-4) 的表达，以及假定的分化信号 Schlafen 12 (SLFN12) 和尾相关同源盒肠特异性转录因子 (Cdx2) 的表达。绒毛蛋白启动子活性通过基于荧光素酶的测定来测量。
MTS 测定表明，特杜格鲁肽使未经处理的对照细胞数量增加了 10%（2%），最大浓度为 500 nM（n = 6，P <.05）。与未经处理的对照相比，特杜格鲁肽使溴脱氧尿苷阳性细胞增加了 19.4%（2.3%）（n = 6，P <.05），通过流式细胞术分析增加了 S 期分数。特杜格鲁肽使绒毛蛋白的平均（SD）表达降低了 29%（6%），Cdx2 降低了 31%（10%），DPP-4 降低了 15%（6%），GLUT2 降低了 40%（11%），SLFN12 降低了 61%（14%），蔗糖酶-异麦芽糖酶降低了 28%（8%）（n = 6，所有 P <.05）。
特杜格鲁肽增加了 Caco-2 的增殖，但倾向于抑制肠上皮细胞的分化。在短肠综合征患者中，特杜格鲁肽的有丝分裂刺激作用可能更大，如果更多的特杜格鲁肽处理的细胞能够被刺激向更完全分化的表型发展。
