| Objective:To confirm the exact effect of GuBiZhiTong (GBZT) solution and discuss the mechanism of action of GBZT solution on the New Zealand white rabbits model of knee osteoarthritis, this project detects the express of transforming, growth factor-β (TGF-β) and insulin-like growth factor-1 (IGF-1) in different period influenced by different doses of GBZT solution, through polymerase chain reaction (PCR). To provide the experimental support and theoretical basis of the treatment of GBZT solution on O A in clinic applicationsMethods:Fifty male or female healthy New Zealand white rabbits of 8 to 14 months and 2.5 kg to 3 kg weight were randomLy divided into 6 groups. There were low, medium, high dose groups of GBZT solution, glucosamine hydrochloride (GH) group and model group respectively. And 12 rabbits were included in each group. Hulth method was refered to establish the model of osteoarthritis. After mo ding, different dose of GBZT solution, glucosamine hydrochloride and normal saling (NS) were intragastric administration for each group continuously for eight weeks. Rabbits in each group were put into death and sampled articular cartilage specimen after modling for four weeks and eight weeks respectively. Naked-eye observation was performed for morphology changes and pathological sections of hematoxylin-eosin (HE) staining were observed by microscope.Real-time polymerase chain reaction (RT-PCR) method was taken to detect the expression of TGF-β and IGF-1 in rabbit knee cartilage articularis. The statistical analysis was performed by SPSS 19.0 softwareResults:1. Macroscopic observation:Normal knee joint surface should be smooth and has transparent color. After 4 weeks’ moding, the model group articular cartilage faded dull, joint surface was not smooth, partly was beige, no obvious defects and the synovium was thicken. The transparency of cartilage in medium dose group, high dose group and GH group was higher than that in the model group and without defects and new creatures. After 8 weeks, the articular cartilage became yellow in the model group and the transparency of cartilage was significantly reduced. There were also osteophyte significantly formed and the cartilage even hecame erosion. But there no obviously changes in the GBZT solution groups and GH group compared with the previous groups, significant And involving the bone pain liquid group and glucosamine hydrochloride group, no obvious changes earlier.2. HE staining microscopic observation:normal cartilage surface is relatively smooth, with the tiny pores of symmetry, covered by the dotted white spherical gel-like substance. The cartilage surface of model set changed to thin, rough, uneven, and even some disappeared after 4 weeks, Some areas appeared crack appearance change, presenting villiform. Depigmentation of surface of the serious parts and bareness of the collagen fiber underneath were observed, and cartilage surface cracked, and collagen fiber fracture.Clear cell structure, cell membrane integrity and uniform distribution of chromatin in the nucleus were observed in normal cartilage, and the outline is clear, crisscrossed visible extracellular collagen fiber. The model set cell membrane dissolved, which is seen that density distribution inhomogeneity. There was cell clustering phenomenon exists and no significant cartilage cells arranged in order. Most of the tidal lines were not complete, The cartilage damages in the low dose, medium dose, high dose GBZT solution groups and the GH group were better than that in the model group. There were still some areas of cartilage cells arranged in irregular phenomenon, but obviously better than that in the model group with less clear boundary layers, microscopic phenomenon of individual regions had family collection. The majority of the tidal lines were imcomplete but individual integrity. Structural damage of the cartilage and the formation of the osteophyte were observed in the model group after 8 weeks. There were no different in each treatment group compared with before.3. RT-PCR:Compared with the model group after 8 weeks, the expression of IL-1 mRMA and TNF-α mRNA in the cartilage specimens of the white rabbits were significant lower in the low dose of GBZT solution group (P<0.05). The expression of IL-1 mRNA and TNF-α mRNA were significant lower in the medium dose, high dose and the GH groups compared with the model group (P<0.01). The expression of IL-1 mRNA and TNF-α mRNA were significant lower in the medium dose, high dose of GBZT solution groups and the GH group compared with the low dose of GBZT solution group after 8 weeks (P<0.05). Compared with the GH group after 8 weeks, there were no significant different of expression of IL-1 mRNA and TNF-α mRNA in the medium dose, high dose of GBZT solution groups (P>0.05). The expression of IL-1 mRNA and TNF-α mRNA in the low dose, medium dose, high dose of GBZT solution groups and the GH group after 8 weeks were significantly lower than that st alter 4 weeks.Conclusion:This experiment evaluated the histological damage index (HI) by HE staining to observe histological changes, and detected the express of transforming growth factor-β (TGF-β) and insulin-like growth factor-1 (IGF-1) in different period influenced by different doses of GBZT solution, through polymerase chain reaction {PCR). The results show that the cartilage surface of model set changed to thin, rough, uneven, and even some disappeared after 4 weeks, Some areas appeared crack appearance change, presenting villiform. Depig mentation of surface of the serious parts and bareness of the collagen fiber underneath were observed. This showed that the articular cartilage layer changed in model set. proved that building models successfully. The expression of articular cartilage facter in model set was higher than in the treatment group, showing that the GBZT solution can play the therapeutic effect on experimental rabbit knee osteoarthritis, and long-term usage can inhibit articular cartilage degeneration and inhibit the expression of inflammatory mediators. Glucosamine hydro chloride can inhibit the same cytokines (TGF-β, IGF-1), and showed no difference on the data statistics involving the bone pain liquid high dose group. We can draw a conclusion that the GBZT solution has comparative advantages in prevention and control of QA. and it’s necessary to carry out basic and clinical usage furtherly. The GBZT solution showed a better inhibition of the expression of TGF-β and IGF-1 factors in the cartilage cells, proving the clinic effects of the GBZT solution furtherly. |
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