The Effect Of Different Exclusive Enteral Nutrition Contents On Growth Factor Expression And The Proximal Tibial Epiphyseal Growth Plate Of IBD Rats

Objective We used IBD-specific formula, short peptides formula,ordinary formula and normal diet enteral nutrition respectively to intervene in IBD rats.Aimed to explore the role of four different components enteral nutrition on IGF-1, IGFBP3 expression and proliferation of proximal tibial epiphyseal cartilage at different periods of inflammatory bowel disease in rats. And which kind of them can obtain more sustained remission, even be able to significantly promote growth?Methods 4-5 weeks SD rats (80-110g) were randomly divided into 8 groups.Including 4 model groups(IBD-specific formula group 22 rats,short peptides formula group 21 rats,ordinary formula group 20 rats and normal diet group 20 rats) and 4 control groups(IBD-specific formula group,short peptides formula group,ordinary formula group and normal diet group had 16 rats respectively).Model groups underwent the infusing operation of trinitro benzene sulfonic (TNBS 100 mg/kg in ethanol) through anus with 2mm diameter rubber tube after fasting 24h. The control groups received the same dose 0.9% NaCl solution of required TNBS and ethanol mixture.4 model groups and 4 control groups were given IBD-specific formula,short peptides formula,ordinary formula and normal diet enteral nutrition respectively after operation.Each group was free diet and recorded the diet,weight and stool everyday.8-10 rats were executed at 7th day,14th day after operation and the specimens from blood, ileum, colon and right side of the tibia.To Measure the length of the tibia of each group rats.Colon and ileum samples were stored in 10% neutral formalin and pathological change was observed by section HE staining. Intercepted proximal epiphysis segment of tibial and stored in 10% neutral formalin and and observed proliferation of chondrocytes in growth plate by section HE staining and assayed GHR, IGF-1R expression with immunohistochemical methods. IGF-1 and IGFBP3 expression levels in the blood were tested by ELISA.Results 1. Diet:Food intake (g) of each model group rats was significantly less than the corresponding control group rats within 7 days after operation. The average daily food intake (g) of short peptide formula milk group was more than the other three groups among 4 different enteral nutrition model groups within 4 days and was more than the other two formula groups from 5 days to a week. The average daily food intake (g) of model groups were gradually returned to normal compared to the corresponding control groups.2. Weight:(1)Model groups:The average weight of each model group with 4 different enteral nutrition was significantly decreased within 4 days after operation. But the short peptide formula group's average weight loss (g) was significantly less than the special formula group (-0.61±13.5 vs -9.59±10.3, P<0.05) and ordinary formula group (-0.61±13.5 vs-5.61±12.8, P<0.05) at the 4th day after operation. There is no obvious difference between short peptides formula milk group and normal diet group. The average weight gain (g) of short peptide formula group was more than the special formula group (12.06±16.7 vs 0.14±17.7, P<0.05) and the ordinary formula group (12.06±16.7 vs 5.14±19.1, P<0.05). There is no obvious difference among 3 different formula groups at 14th day after operation,but the average weight gain(g) of normal diet group was significantly more than the other three different formula milk groups respectively(84.35±18.0 vs 25.35±31.8,35.26±27.2,23.36± 23.5, P<0.05). (2)Control groups:The average weight gain (g) of the ordinary formula group was significantly less than the other groups respectively (P<0.05).3. The length of the tibia:The average tibia length (cm) of short peptides formula group was apparently longer than IBD-specific formula group and normal diet group(3.21±0.10 vs 3.06±0.15,2.98±0.11, P<0.05) at the 7th day; But there was no significant superiority on tibia length of peptides formula milk group than the other three groups at 14th day. The growth rate of average tibial length was the greatest in normal diet group (13.1%), followed by IBD-specific formula group (8.8%), peptides formula group (3.1%) and ordinary formula group (2.6%) compared 14th day to 7th day among the 4 model groups. There was no significant difference of average tibial length among 4 different enteral nutrition control groups at 7th day. The average tibial length of ordinary formula group was the shortest than the other three groups at 14th day among the 4 different enteral nutrition control groups, and there was no apparent difference among peptides formula group, IBD-specific formula group and normal diet group at 14th day.4. Ileum pathological:(1)The model groups:The intestinal wall was edema, the serosa, muscularis and mucosa layer boundaries is not clear and the intestinal villi was damaged and shedding in all of the 4 different enteral nutrition groups at 7th day. However, the intestinal pathological changes above of the short-peptide formula group were slight among 4 model groups. The pathological changes were significantly alleviated at 14th day in 4 groups. (2)The control groups:There was no significant pathological changes about the ileal tissues in 4 different enteral nutrition control groups at 7th and 14th days.5. The expression of IGF-1 and IGFBP3:(1) The expression of IGF-1:Average blood IGF-1 levels (ng/ml) of the peptides formula group was significantly higher than IBD-specific formula group, ordinary formula group and normal diet group(3.79±0.42 vs 2.93±0.89?2.69±0.49?2.58±0.50, P<0.05) at 7th day among 4 model groups. However, there was no significantly difference among 4 different enteral nutrition groups at 14th day. The highest level of IGF-1 appeared in the normal diet group just at 14th day in the 4 control groups. (2) The expression of IGFBP3:Average blood IGFBP3 levels (ng/ml) of the peptides formula group was significantly higher than normal diet group (21.28±4.52 vs 16.11±2.86, P<0.05). But it was only slightly higher than the IBD-specific formula group and gordinary formula group and there was no statistically significant.In 4 different enteral nutrition control groups, the highest level of blood IGFBP3 (ng/ml) which was higher than peptides formula group and ordinary formula group (21.50±4.81 vs 15.32±4.49?16.58± 4.72, P<0.05) appeared in normal diet group just at 14th day.6. Tibial pathology and immunohistochemistry:(1) Thickness of epiphyseal growth plate:In 4 different enteral nutrition model groups, the thickness of epiphyseal growth plate of peptides formula group was the thickest and the thinnest thickness of epiphyseal growth plate appeared in IBD-specific formula group at 7th day. At the 14th day, the thickness of epiphyseal growth plate was significantly restored in IBD-specific formula group and normal diet group.However,the thickness of epiphyseal growth plate of the ordinary formula group was thinner than that at 7th day. In 4 different enteral nutrition control groups, the epiphyseal growth plate of IBD-specific formula group and normal diet group was thicker than that of peptides formula group and ordinary formula group at 7th and 14th day. (2) Chondrocytes count of proliferation zone and hypertrophy zone in epiphyseal growth plate:In 4 different enteral nutrition model groups, the average chondrocytes count of proliferation zone and hypertrophy zone in epiphyseal growth plate of proximal tibial of peptides formula group was significantly more than that of IBD-specific formula group and normal diet group at 7th day (66.0±16.1 vs 47.9±6.3?51.2±6., P<0.05). Dramatic changes occurred at the 14th day, the average chondrocytes count of both of above two zones of peptides formula group was less than that of IBD-specific formula group and normal diet group, but there was no statistically significant. In 4 different enteral nutrition control groups, the average chondrocytes count of both of the two zones of IBD-specific formula group was significantly more than that of the other three groups at 7th day (P<0.05).And the average chondrocytes count of both of the two zones of peptides formula group was slightly less than that of normal diet group and IBD-specific formula group at 14th day, but there was no statistically significant. In 4 different enteral nutrition control groups, the cells count of the two zones of IBD-specific formula group was significantly more than that of the other three groups at 7th day(P<0.05). But there was no significantly difference among the peptides formula group, IBD-specific formula group and normal diet group at 14th day.(3)The expression of GHR and IGF-1R in the epiphyseal growth plate:GHR expressed in the entire epiphyseal growth plate area, but according to the immunohistochemical sections it mainly expressed in resting zone. The strong positive expression cells count of 4 different enteral nutrition model groups had no significant difference (P> 0.05) at 7th day and 14th day respectively.Conclusion The peptides formula enteral nutrition intervention can quickly relieve intestinal inflammation of IBD rats, promote the expression of IGF-1 and IGFBP3, and accelerate the growth of long bones within 7 days after operation. Compared with the normal diet group this advantage is more and more unobvious 7 days later after operation.