| ObjectiveTo investigate the influence of transforming growth factor betat1 (TGF-β1) and interleukin 1 beta (IL-1β) on human cartilage endplate and in vitro cultured cervical cartilage endplate. To preliminarily probe the action mechanism of TGF-β1 and IL-1βin the occurrence and evolution of intervertebral disc degeneration and to provide novel pathway for preventing and curing intervertebral disc degeneration.Methods1.①Monitor the expression of TGF-β1 and IL-1βof centrum cartilage endplate gathered from November 2009 to June 2010 in our hospital by immunohistochemistry (IHC). The lesion group consists of 20 centrum cartilage endplates resectionlied in the anterior surgery of cervical spondylosis and the control group is comprised of 20 centrum cartilage endplates gathered in the anterior surgery of cervical fracture.②Monitor the expression of TGF-β1 and IL-1βin both lesion and control group by means of western blot.2. Adopt cervical cartilage endplate to carry out the primary culture experiment, observe the morphology of cells, draw the growth curve and choose the cells from passage 2-4 as samples.①Add TGF-β1 and IL-1βinto sample cartilage cells cultured in serum-free nutrient solution, with the final concentration of 0.5pg/ml, 5pg/ml , 10pg/ml, 50pg/ml. Detect the proliferation of cells after culturing for 24h by the colorimetric method using 4-methyl thiazolyl tetrazolium (MTT). Results1.①The positive rate of TGF-β1 and IL-1βin the lesion group was 32.9076%and 51.5207%,respectively; while the positive rate of TGF-β1 and IL-1βin the control groupwas 42.4816% and 33.9185%,respectively. The overall positive expression of TGF-β1 in the control group was significantly higher than that in the lesion group, whereas the overall positive expression of IL-1βin the lesion group was significantly higher than that in the control group, and both of the differences were distinct enough to have a statistical implication(P <0.01).②In the western blot experiment, the expression of TGF-β1 in the control group and IL-1βin the lesion group were both significantly higher than their counterparts2. The MTT method showed that TGF-β1 promoted the proliferation of cartilage cells at concentration of 0.5, 5 and 10 pg/mL and inhibited it at 50 pg/mL. The difference between the experimental and control group had a statistical implication (P<0.05 or P<0.01). The effect of TGF-β1 was also dependent on its concentration, and the optimal promotion occurs when cartilage cells were exposed to 10 pg/mL TGF-β1 for 24 h(P<0.01.) Obvious inhibition of proliferation was observed when the concentration of IL-1βin the experimental group is 0.5, 5, 10 and 50 pg/mL.Conclusion1. The expression TGF-β1 and IL-1βexisted in degenerated human cartilage endplate, and the expression of TGF-βand IL-1βin the lesion group was lower and higher than their counterparts in the control group, respectively. The positive stain of TGF-β1 and IL-1βwere mainly located in the cytoplasm and cytomembrane of cartilage cells, and also appeared in the nucleus. The qualitative and quantitative results of expression of TGF-β1 and IL-1βobtained by IHC and western blot are in good accordance, thus providing reference for research and clinical work.2. MTT results suggested that TGF-β1 in primary cervical cartilage cells could promote the proliferation of cartilage cells at limited concetrations and inhibited it at a concentration of 50pg/ml, while IL-1βinhibited it at all concentrations. The qualitative and quantitative results of expression of TGF-β1 and IL-1βin cells from passage 2 and 4 obtained by IHC and western blot are in good accordance, , thus also providing reference for research and clinical work. |
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