RESUMO
Objective To investigate the effects of Lactobacillus rhamnosus GG (LGG) colonization in early life on intestinal barrier and intestinal development in offspring mice and its possible mechanism.Methods Six C57BL/6 pregnant mice with the same conception time of 6 weeks were selected and randomly divided into experiment group given 108 cfu/ml LGG live bacteria and control group given LGG inactivated bacteria by gavage from the 18th day of pregnancy until natural birth.The progeny mice in the two groups were continued to be gavaged with 107 cfu/ml of LGG live bacteria or LGG inactivated bacteria on days 1-5 of birth.The body weight changes of 3 week'progeny mice were recorded.The colonization of LGG bacteria in offspring mice was detected at 2nd and 3rd weeks.The mRNA of intestinal proinflammatory cytokines and tight junction molecules were evaluated by real-time PCR method.HE,immunohistochemistry,immunofluorescence staining and enzyme-linked immunosorbent assay were used to evaluate the intestinal barrier of 3-week old off spring mice.Results Compared with the control group,the progeny mice of the experiment group showed no significant difference in body weight at the first week,and the body weight increased at the second week and the third week [2ndweek:(3.790±0.240) g vs.(4.326±0.140) g,t=3.707,P=0.006;3rd week:(7.295±0.326) g vs.(8.040±0.370) g,t=3.130,P=0.011].LGG colonization can be detected only in the feces of progeny mice in the experiment group.Intestinal colonization can promote the growth of small intestine villi and colon crypt depth [jejunum:(320.000±22.514) μm vs.(265.100±15.611) μm,t=8.258,P＜0.001;ileum:(150.500±13.099) μm vs.(111.000±11.308) μm,t=9.958,P＜0.001;colon:(295.000±15.209) μm vs.(233.100±6.678) μm,t=9.129,P＜0.001].Compared with the control group,the number of goblet cells in the colonic crypt of the experiment group increased (11.62 ± 0.780 vs.35.24 ±1.370,t=15.000,P＜0.001),and the relative mRNA expression levels of pro-inflammatory factors as IFN-γ (1.280±0.232 vs.0.512±0.206,t=4.970,P=0.001),IL-6 (1.364±0.271 vs.0.941±0.215,t=2.452,P=0.040),IL-10 (1.341±0.320vs.0.744±0.294,t=2.762,P=0.025)andTNF-α (3.702±0.150 vs.2.581±0.500,t=2.553,P=0.034) in the experiment group decreased;the expression levels of the intimate tight junction molecules (Claudin3) (1.283±0.152 vs.1.881±0.172,t=4.932,P=0.001) and the atresia protein molecule (Occludin) (1.164±0.342 vs.0.812±0.224,t=3.67,P=0.016) significantly increased.Conclusion Early life LGG colonization protects the intestinal barrier by inhibiting lowgrade intestinal inflammation.This study will lay the experimental foundation for the supplementation of probiotics in early life so as to prevent intestinal diseases.
RESUMO
Objective To observe the effects of maternal high fat diet (MHFD) during pregnancy and lactation on intestinal barrier function in offspring mice.Methods C57BL/6 pregnant mice were divided into high fat diet (MHFD) group and normal diet group (MND) randomly and were given high fat diet and normal diet during pregnancy (3 weeks) and lactation (3 weeks) respectively.Both groups of offspring mice were naturally given and bodyweight of pups was monitored at birth and weekly.After weaning,the intestinal permeability of offspring mice was detected by fluorescein isothiocyanate conjugated-dextran method (FITC-D).Immunofluorescence was used to detect the expression of ZO-1 in intestinal tissues.HE staining was used to assess the villus length and crypt depth.The intestinal cell proliferation (expression of Ki-67) and Mucin 2 (MUC2) were assessed by immunohistochemistry.PAS staining was used to evaluate the goblet cells.The expression of inflammatory cytokines including IL-1β,IL-6,and TNF-α in intestinal tissue were measured by real-time PCR.Results At the age of 2 and 3 weeks,the offspring in MHFD group were significantly heavier than those in MND group.HE staining showed no obvious microscopic inflammation in both groups of 3 weeks old offspring mice,however,the relative expression levels of IL-1β (1.95±0.53 vs.1.13±0.15;t =3.65,P=0.005),IL-6 (1.40±0.71 vs.0.73±0.17;t=2.72,P=0.04),and TNF-α (1.63±0.53 vs.1.04±0.12;t=2.64,P=0.02) mRNA were significantly higher in the MHFD group.Compared with the 3 weeks old offspring mice in MND group,MHFD significantly increased the permeability of intestine and decreased the expression of ZO-1 in membrane.The number of Ki-67 positive cells (18.00±4.74 vs.24.60±4.17;t =3.31,P=0.004) in each villus,goblet cells (14.70±2.91 vs.28.10±4.95;t =7.38,P＜0.001) and MUC2 positive cells (20.60± 3.13 vs.30.00±3.33;t=6.50,P＜0.001) in each crypt were significantly lower than those in MND group.Conclusion Maternal high fat diet in early life of offspring mice can induce intestinal low grade inflammation and lead to the disruption of intestinal mucosal barrier in offspring mice,which may be involved in the progeny diseases.