Zhang Hui, Ameen Nadia, Melvin James E, Vidyasagar Sadasivan
Digestive Diseases, Department of Medicine, Center for Oral Biology, University of Rochester School of Medicine, 601 Elmwood Ave, Box 646, Rochester, NY 14642, USA.
J Physiol. 2007 Jun 15;581(Pt 3):1221-33. doi: 10.1113/jphysiol.2007.129262. Epub 2007 Mar 29.
T-cell mediated acute inflammation of the ileum may occur during Crohn's disease exacerbations. During ileal inflammation, absorption of nutrients and electrolytes by villus cells is decreased with a concomitant increase in crypt and/or villus fluid secretion. These alterations lead to fluid accumulation and the subsequent diarrhoea. Net intestinal fluid secretion consists of HCO3--rich plasma-like fluid. However, the regulation and mechanisms of HCO3- secretion in normal and acutely inflamed ileum are not clearly understood. To study this phenomenon, anti-CD3 monoclonal antibody (mAb)- induced in vivo ileal inflammatory mouse models was used for in vitro functional studies with Ussing chamber and pH stat techniques. Three hours after anti-CD3 mAb injection, ileal mucosa stripped of muscular and serosal layers showed a significant increase in short circuit current (Isc) (0.58+/-0.07 microEq h(-1) cm2 versus 1.63+/-0.14 microEq h(-1) cm2). The cAMP-stimulated Isc component was sensitive to glibenclamide but not to DIDS, suggesting that a cystic fibrosis transmembrane conductance regulator (Cftr)-mediated anion conductance was responsible. Basal Cl--dependent HCO3- secretion, measured using a pH stat technique, was decreased significantly in anti-CD3-injected mice, with a simultaneous increase in Cl--independent HCO3- secretion that was also inhibited by glibenclamide. Experiments using Cftr-/- mice showed neither an increase in Isc nor an increase in HCO3- secretion, confirming the role for Cftr protein in stimulating anion secretion following anti-CD3 treatment. Western blot analysis indicated that Cftr protein levels were unaltered by anti-CD3 treatment, at least acutely. Finally, an immunoassay for cAMP showed significant increases in intracellular cAMP in villus cells, but not in crypt cells. These studies therefore suggest a shift from a predominantly electroneutral Cl-HCO3- exchange in normal mice, to a predominantly electrogenic anion secretion including HCO3- that occurs via functional Cftr during anti-CD3-mediated acute inflammation.
在克罗恩病病情加重期间,可能会发生T细胞介导的回肠急性炎症。在回肠炎症期间,绒毛细胞对营养物质和电解质的吸收减少,同时隐窝和/或绒毛液分泌增加。这些改变导致液体蓄积及随后的腹泻。肠道净液体分泌由富含HCO3-的血浆样液体组成。然而,正常和急性炎症回肠中HCO3-分泌的调节及机制尚不清楚。为研究此现象,采用抗CD3单克隆抗体(mAb)诱导的体内回肠炎症小鼠模型,运用尤斯灌流小室和pH计技术进行体外功能研究。注射抗CD3 mAb三小时后,去除肌层和浆膜层的回肠黏膜短路电流(Isc)显著增加(0.58±0.07微当量·小时-1·厘米-2 对比1.63±0.14微当量·小时-1·厘米-2)。cAMP刺激的Isc成分对格列本脲敏感,但对二异丙基氟磷酸(DIDS)不敏感,提示囊性纤维化跨膜传导调节因子(Cftr)介导的阴离子传导起作用。采用pH计技术测量,注射抗CD3的小鼠中,基础Cl-依赖性HCO3-分泌显著减少,同时Cl-非依赖性HCO3-分泌增加,且也被格列本脲抑制。使用Cftr基因敲除小鼠的实验显示,Isc和HCO3-分泌均未增加,证实了Cftr蛋白在抗CD3治疗后刺激阴离子分泌中的作用。蛋白质免疫印迹分析表明,抗CD3治疗至少在急性期未改变Cftr蛋白水平。最后,一项cAMP免疫测定显示,绒毛细胞内的细胞内cAMP显著增加,但隐窝细胞中未增加。因此,这些研究表明,在抗CD3介导的急性炎症过程中,正常小鼠中主要的电中性Cl-HCO3-交换转变为主要通过功能性Cftr发生的包括HCO3-的电生性阴离子分泌。