The GABAergic System Regulates The Proliferation And Differentiation Of Intestinal Epithelial Stem Cell And Prevents Intestine From Injury

Objectives:Cancer research has been rightly and successfully focused on prevention, early detection, and identification of specific molecular targets that distinguish the malignant cells from the neighboring benign cells. However, reducing lethal tissue injury caused by intensive chemotherapy during treatment of late-stage metastatic cancers remains a key clinical challenge. Gamma aminobutyric acid (GABA) can inhibit neural crest stem cells proliferation, we found that GABAergic signaling molecules were expressed in the intestinal epithelium and GABA involved in regulating intestinal stem cells proliferation and differentiation. So here we tested whether induction of adult stem cells through inhibition of GABA receptors could repair chemotherapy-induced intestinal injury.Methods:we performed immunohistochemisty staining to analyze the expression of GABAergic system molecule in the mouse and human small intestine.We administered muscimol, bicuculline, baclofen and CGP54646 in the adult mice for 2 weeks and then collected the jejunum to do HE staining for morphological analysis and immunohistochemisty staining for transit amplifying cells(TACs), BrdU+ cells, Goblet cells and apoptotic cells. Next we administered therapeutic dose of 5-Fluorouracil for 5 days or high dose of 5-Fluorouracil once in adult mice to establish intestinal epithelium injury model and in the same time we injected bicuculline or CGP54626 to the mice. Thereafter, we collected the jejunum to do HE staining for morphological analysis and immunohistochemisty staining for transit amplifying cells, BrdU+ cells, apoptosis cells and yH2AX+ cells, performed βcatenin immunohistochemisty staining to confirm if it could affect Wnt signaling, and did QPCR analysis of the relative mRNA expression of intestinal stem cells markers lgr5 and olfm5,reserve stem cells markers mTert and bmi1,and DNA damage related gene.Results:1.GAD65,GAD67,GAT3,GABAARα1,β1/2/3,GABABR1 and GABABR2 were expressed in the intestinal epithelium and were high expressed in the crypts. In addition, GABAaRπ, q5,r, e2 and GAT2 were also expressed in the intestinal epithelium, but not ABAT.2.Intraperatoneal injection of muscimol decreased the villus height and the numbers of TACs and goblet cells in mice, and increased the number of apoptotic cells; injection of bicuculline increased the villus height and the number of goblet cells; injection of baclofen decreased the TACs number while CGP54626 increased TACs number.3.By comparison with NS group, administration of bicuculline and CGP54626 prevented intestinal epithelium from 5-Fluorouracil induced injury, increased TACs,and decreased yH2AX+cells, higher expression of olfm4 and lgr5 mRNA while lower expression of bmi1 and mTert mRNA in the chemotherapy induced intestinal injury model.Moreover, injection of CGP54626 could decrease apoptotic intestinal stem cells, decreased the expression of gtf2h2 and prkdc mRNA in mice injury model.Conclusions:1.There is GABAergic signaling system in intestinal epithelium. 2.GABA regulates intestinal stem cells proliferation,differentiation and apoptosis through its A receptors.3. GABA regulates intestinal stem cells proliferation through its B receptors.4.1nhibition of GABAa R and GABAb R can protect intestinal epithelium from chemotherapy induced injury.