| Aims The aims of this study were to investigate ICC changes in the stomach of diabetic mice after being treated with EA.Methods A total of fifty male six to eight weeks old C57BL/6 mice were used in this study and were randomly divided into five groups:the control group,the diabetic group(DM),a group of diabetic mice with sham EA(DM+SEA),a group of diabetic mice treated with low-frequency EA group(DM+LEA),a group of diabetic mice treated with high-frequency EA group(DM+HEA).To make the diabetic mouse model,a single intraperitoneal injection of streptozotocin(STZ)was applied at a dose of 150 mg/Kg.The location of acupuncture was the Zusanli(ST-36),which was located at the posterolateral knee of bilateral hind limbs.The parameters for each group:the DM+SEA group(only acupuncture without electric current);the DM+LEA group(10Hz.1-3mA,continuous wave);the DM+HEA group(100Hz,1-3mA,continuous wave).The chronic stimulation was as long as eight weeks and 30 minutes per day.The glucose and body weight changes were recorded in mice before modeling 1 week,after 2 weeks,4 weeks,6 weeks and 8 weeks,respectively.After 8 weeks,the tissues of antrum and corpus for each group were obtained and the whole-mount preparations were made to label ICC by double immunofluorescence staining.Results(1)There were no significant difference for the glucose level among five groups before modeling.Compared with the control group,the glucose levels in other four groups were obviously elevated after modeling 2 weeks,4 weeks,6 weeks and 8 weeks,respectively.But no difference were found among the four groups after modeling.(2)There were no obvious difference for the body weight among five groups before modeling.Compared to the control group,the body weight for other four groups were markedly decreased after modeling 2 weeks,4 weeks,6 weeks and 8 weeks.Compared with the DM group,the body weight in the DM+LEA group was significantly increase after modeling 4 weeks and the body weight in the DM+HEA group was dramaticlly increase after modeling 8 weeks.(3)There were abundant and bright Anol positive cells and c-Kit positive cells in the stomach of the control group and Anol positive cells were almost completely merged with c-Kit positive cells.Compared with the control group.Anol+ cells and c-Kit+ cells were obvious decreased in the stomach of the DM group,but no marked changes were found in the DM+LEA group and the DM+HEA group.In the DM group and the DM+SEA group,c-Kit positive cells were decreased obviously than Ano1 positive cells and a few Anol+c-Kitlow and Anol+c-Kit-cells were found.Conclusion(1)There were no effects for the variations of glucose in diabetic mice under the condition of EA;(2)After treated with EA,loss of body weight in diabetic mice were decreased,especially in the DM+LEA group;(3)Compared with c-Kit,Anol was more sensitive and specific to label ICC in diabetic mice;(4)Low-and high-frequency EA can protect the networks of ICC in the stomach of diabetic mice.Objective To investigate whether the mSCF/Kit-ETV1 signaling participates the protective effects for the ICC networks in the diabetic mice after treated with EA.Methods The expression of mSCF,c-Kit and ETV1 protein were detected by Western blot;RT-PCR was employed to detected the mRNA of mSCF,c-Kit and ETV1 expression;The technique of double immunofluorescence was used to evaluate the location and expression of c-Kit and ETV1.Results(1)Western blot showed that compared to the control group,the expression of mSCF,c-Kit and ETV1 protein in gastric antrum and corpus of the DM group were significantly decreased,but in the DM+LEA and the DM+HEA,the protein expression of mSCF,c-Kit and ETV1 in gastric antrum and corpus were markedly elevated when compared with the DM group;(2)RT-PCR showed that when compared with the control group,the expression of mSCF,c-Kit and ETV1 mRNA in the antrum and corpus of the DM group were obviously reduced,but the expression of mSCF.c-Kit and ETV1 mRNA in the antrum and corpus of the DM+LEA group and the DM+HEA group were significantly increased when compared to the DM group;(3)The result of double immunofluorescent staining revealed that,in the control group,plenty of ETV1 expressed in the myenteric plexus of antrum and corpus,connected each other to form networks.The ETV1 distribution was closely associated with the networks of ICC,even parts of ETV1 and ICC were overlapped.In intramuscular of antrum and corpus,the expression of ETV1 was relative decreased and the distribution of ETV1 was along the smooth muscle.Most parts of ETV1 were accompanied with ICC and a few merged parts were also observed.However,in the antrum and corpus of the DM group,accompanied with the disrupted ICC networks,the expression of ETV1 was significantly decreased.Compared with the DM group,the expression of ETV1 in the DM+LEA group and the DM+HEA group formed abundant and bright networks along the networks of ICC,which were similar to the control group.Conclusion(1)In diabetes,the decreased mSCF and c-Kit in the tissues of stomach may lead to ETV1 lost and further make the ICC networks to be disrupted;(2)The m SCF/Kit-ETV1 signaling can be normally activated to sustain the expression of ETV1 after treated with EA,further protect ICC networks from diabetes.Objective To investigate whether the HO-1 positive M2 macrophages participated the protective effects of EA for the ICC networks in the stomach tissues of diabetic mice.Methods Sixty male C57BL/6 mice were randomized into five groups:the control group,the diabetic group(DM group),the group of diabetic mice plus sham EA(DM+SEA group),the group of diabetic mice plus low-frequency EA group(DM+LEA group),the group of diabetic mice plus high-frequency EA group(DM+HEA group).EA was started to carry out after diabetic model had been made successfully.Eight weeks later,the tissues of stomach and serum were obtained.The technique of double immunofluorescence was used to observed HO-1 positive macrophages expression;Western blot and RT-PCR were applied to investigate the expression of HO-1 and IL-10 in the stomach tissues;the serum MDA level was detected by a commercial kit;the M1 macrophage marker-iNOS and the M2 macrophage marker-Arg-1 and CD163 expression were evaluated by RT-PCR.Results(1)Compared with the control group,the amounts of F4/80 positive macrophages were obviously increased,but F4/80 positive macrophages were significantly decreased in the DM+LEA group and the DM+HEA group;(2)There were nearly no HO-1 positive macrophages in the control group,the DM group and the DM+SEA group,but many HO-1 positive macrophages were observed in the DM+LEA group and the DM+HEA group;(3)The results of western blot revealed that the expression of HO-1 and IL-10 protein was very low in the control group,the DM group and the DM+SEA group,but was rich in the DM+LEA group and the DM+HEA group;(4)RT-PCR also showed that the levels of HO-1 and IL-10 mRNA was very low in the control group,the DM group and the DM+SEA group,but increased in the DM+LEA group and the DM+HEA group;(5)Compared to the control group,the serum MDA was markedly increased in the DM group,but it was decreased in the DM+LEA group and the DM+HEA group when compared with the DM group;(6)The results of RT-PCR also exhibited that the mRNA expression of M2 macrophage marker-Arg-1 and CD 163 was relative low in the DM group,but significantly upregulated in the DM+LEA group and the DM+HEA group.However,the situation of the Ml macrophage marker iNOS was reversed as the M1 macrophage makers.Conclusion EA stimulation can promote the expression of HO-1 positive M2 macrophage expression,enhance the secretion of anti-inflammatory molecule IL-10,decrease oxidative stress and inflammatory response and further protect the ICC networks of stomach in diabetes. |
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