The Study Of Two Kinds Of Collagen And The Effects On Rheumatoid Arthritis Of Mice

In this paper,the type I collagen?CI?and the type II collagen?CII?were extracted from porcine tendon and chicken sternal cartilage,respectively.Then the CI and CII were characterized by several methods,such as UV,FTIR,CD and SDS-PAGE and so on.Finally,we studied the effect of CI and CII on rheumatoid arthritis?RA?of C57BL/6 J mice.First of all,acid soluble collagen?ASC?and pepsin soluble collagen?PSC?from porcine tendon were isolated and characterized,respectively.The yields of ASC and PSC were12.10%and 17.39%,based on the dry weight of porcine tendon,respectively,with the accumulated yield of 29.49%.Ultraviolet spectral analysis showed that the characteristic absorption wavelength of ASC and PSC were 224 nm and 209 nm,respectively,suggesting that both of them are typical collagen.Sodium dodecyl sulfate-polyacrylamide-gel electrophoresis?SDS-PAGE?suggested that both ASC and PSC comprising two different?-chains??₁ and?₂?were characterized as type I collagen.Fourier transform infrared spectrometry?FTIR?of both ASC and PSC were almost similar and pepsin hydrolysis had no marked effect on the triple-helical structure of collagen.The thermal denaturation temperature?Td?of ASC and PSC were 39?and 32?,respectively.The results suggested that the collagen of porcine tendon were type I collagen and had a triple helical structure.Secondly,the pepsin soluble collagen from chicken sternal cartilage was isolated and characterized.The results was that the yields of collagen was 18.51%,based on the dry weight of chicken sternal cartilage.Ultraviolet spectral analysis showed that the characteristic absorption wavelength of collagen was 217 nm.Analysis results of SDS-PAGE showed that there was a heavy?-chain.Amino acid components showed that it was characterized as type II collagen.The thermal denaturation temperature?Td?of PSC was 38?.The results suggested that the collagen of chicken sternal cartilage was type II collagen and the extracted method did not damage the original native conformational structure of the type II collagen.The method of classical type II collagen-induced arthritis as RA animal model was adopted.C57BL/6 J mice were selected with similar weight and age,and randomly divided into seven groups averagely:control group,model group,CI group,CI+Na₂SeO₃ group,CII group,CII+Na₂SeO₃ group and Na₂SeO₃ group.Every group was determined the weight and monitored arthritic scores every three days after inflammation.We analyzed the content of IL-1??IL-6?IL-17?TNF-?and I?B in the blood serum on the 35^thh after inflammation.In addition,the toe joint section of mice was observed.Then the total RNA of joint was extracted and detected the relative expression of IL-1??IL-6 and TNF-?.Finally the micro CT of joint of the right rear foot was detected.The results were as follows:?1?The trend of weight in groups was that the control group increased slowly all the way;the other increased slowly after first immunization,and then decreased after boost immunization and remained stable eventually.?2?Compared with model group,the inflammation in the right rear foot of mice of model group,CI group was aggravating,but the other were reduced in various degrees,especially in C?+Na₂SeO₃ group.?3?Compared with model group,the content of inflammatory cytokines,such as,IL-1?,IL-6,IL-17,TNF-?and I?B in mice serum,was no signification in CI group?p>0.05?,and CII groups were the opposite?p<0.05?.?4?Histopathological analyses of the toe joints showed that in the H&E images,compared with model group,both CII and Na₂SeO₃ could maintain the smooth and integrity of articular cartilage,CI group could not suppress of synovium inflammation and protect from articular cartilage erosion.?5?The results of micro CT analysis showed that the degree of erosion of the ankle was not reduced in CI group,in contrast to the other groups.?6?The results of RT-PCR showed that compared with model group,the relative mRNA expression of inflammatory cytokines,contain IL-1?,IL-6 and TNF-?in joints,were no signification in CI group?p>0.05?,CII group groups were the opposite?p<0.05?.