Repairing Effect And Mechanism Of Chronic Intermittent Hypobaric Hypoxia On Degenerative Intervertebral Disc

Part 1 Reparative effects of chronic intermittent hypobaric hypoxia pre-treatment on intervertebral disc degeneration in ratsObjective:The study aimed to explore the repair mechanisms of chronic intermittent hypobaric hypoxia(CIHH)on intervertebral disc degeneration(IDD)in rats.Methods:In the experiment,48 adult Sprague–Dawley rats were selected and randomly divided into experimental group(CIHH-IDD),degenerative group(IDD)and control group(CON).The CIHH-IDD group of rats(n=16)were treated with CIHH(simulated 3000m altitude,5 hours per day,28 days,PO~2=108.8 mm Hg)before surgery for disc degeneration.The IDD group of rats(n=16)were treated with tail-vertebrae intervertebral disc surgery to make a model of intervertebral disc degeneration.The CON group of rats(n=16)did not receive any treatment.After surgery,we calculated Disc height index(DHI)with X-rays of rat tail vertebrae,observed the degeneration process and evaluated the repair effect by chemical staining of the degenerative intervertebral disc tissue slice.We measured the expressions of b FGF,TGFβ1,Collagen I and Collagen II in intervertebral disc tissue using Western blot(WB)technology,and determined the expressions of b FGF,TGFβ1 and HIF-1αin rat serum using Enzyme-Linked Immunosorbent Assay(ELISA).Results:(1)The degree of intervertebral disc height degeneration in CIHH-IDD rats was significantly lower compared with that in IDD rats(P<0.05);(2)the expression levels of b FGF,TGFβ1 and HIF-1αwere higher in CIHH-IDD rat serum compared with those in IDD rat serum(P<0.05);(3)optical microscopy revealed that the degree of disc degeneration was relatively mild in CIHH-IDD rats;(4)the protein expression levels of b FGF,TGFβ1 and collagen II were increased in CIHH-IDD rat intervertebral disc tissues compared with those of IDD rats,while the overexpression of collagen I protein was inhibited.Conclusions:after CIHH pre-treatment,the expression levels of b FGF and TGFβ1 were up-regulated,which play notable roles in repairing degenerative intervertebral discs in rats.Part 2 Preventive effect and mechanism of chronic intermittent hypobaric hypoxia(CIHH)post-treatment on degenerative intervertebral disc in ratsObjective:The purpose of this study was to investigate the prophylactic effect of chronic intermittent hypobaric hypoxia(CIHH)post-treatment on the degenerative intervertebral disc in rats with acupuncture.Methods: Forty-eight SD rats were randomly divided into experimental group CIHH group,IDD degeneration group and CON control group.CIHH group rats(n=16)were treated with CIHH(simulated height 3000 m,5 h/day,28 days,PO2=108.8 mm Hg)after needling intervertebral disc degeneration model.The model of intervertebral disc degeneration was made by acupuncture in IDD group(n=16).Rats in CON group(n=16)were used as blank control.The disc height index was calculated using X-ray analysis of rat tail vertebrae in 1week,2weeks,4weeks and 8weeks after surgery.At each time point,the degeneration and changes of intervertebral disc and nucleus pulposus were observed and evaluated by HE staining,saffron solid green staining and Masson staining.Immunohistochemistry(IHC)was used to detect b FGF,TGFβ1,Collagen Ⅰ and Collagen Ⅱ tissues,and then the optical density was measured and the expression of factors was recorded.The expressions of b FGF,TGFβ1 and HIF-1α in rat serum were determined by Enzyme-Linked Immunosorbent Assay(ELISA).Results:(1)there was no statistically significant difference in the degenerative value of intervertebral disc height between the CIHH group and the IDD group(P>0.05).(2)The expression of b FGF in serum of rats was high in IDD group,and increased first and then decreased to normal level in CIHH group.TGFβ1 was highly expressed in CIHH and IDD groups,while the expression of TGFβ1 was decreased in CIHH and IDD groups(P<0.05).The expression of HIF-1α was irregular and high in IDD group(P<0.05),while in CIHH group,the expression of HIF-1α first increased and then decreased to normal level.(3)Under light microscope,there was no significant improvement in the degree of disc degeneration in CIHH and IDD groups.(4)In intervertebral disc tissue,compared with IDD rats,the expression of b FGF factor and TGFβ1 factor in CIHH group was significantly inhibited.And Collagen Ⅰ and II expressions increased in CIHH(P<0.05).Conclusions: CIHH post-treatment can inhibit the expression of b FGF and TGFβ1,but has no obvious repair effect on degenerative intervertebral disc.Part 3 Effect of chronic intermittent hypobaric hypoxia(CIHH)pre-treatment and post-treatment on the relative height change of degenerative discs in ratsObjective: In this study,we observed the changes in the level of the degenerative intervertebral disc after acupuncture in rats with chronic intermittent hypoxia(CIHH)pretreatment and post-treatment to speculate the influence of different time of CIHH treatment on the degenerative intervertebral disc.Methods: Sixty-four SD rats were randomly divided into CIHH pretreatment group(CIHH-IDD),CIHH post-treatment group(CIHH),degeneration group(IDD)and blank control group(CON).CIHH preconditioning group(CIHH-IDD)rats(n=16)were treated with CIHH(simulated altitude 3000 m,5 hours a day for 28 consecutive days,PO2=108.8 mm Hg),and then THE IDD model was established.CIHH post-treatment group(CIHH)rats were first established with IDD model and then immediately treated with CIHH(simulated altitude 3000 m,5 hours a day for 56 consecutive days,PO2= 108.8mm Hg).Degeneration group(IDD)rats were given IDD model without CIHH treatment.The blank control group(CON)maintained normally without IDD and CIHH treatment.Since the IDD model was established,the time was calculated,and at 1week,2weeks,4weeks,and 8weeks after surgery,4 rats were selected for X-ray imaging in each group,the height value of the intervertebral disc and its adjacent vertebral body was measured,the intervertebral disc height index was calculated,and the change in height was represented by the rate of change of DHI.Results:(1)Intervertebral disc height degeneration value in CIHH-IDD group was significantly higher than that in CON group at 8weeks(P<0.05).(2)Intervertebral disc height degeneration of rats in CIHH and IDD groups was significantly different from that in CON group from 2weeks(P<0.05),and significant statistical difference was observed after 4weeks(P<0.01).(3)Compared with the CIHH and IDD groups,the CIHH-IDD group showed statistical significance from the 4weeks(P<0.05),and the CIHH-IDD group showed significant difference from the other two groups at the 8weeks(P<0.01).(4)There was no significant statistical significance in postoperative disc height degeneration in CIHH group and IDD group(P>0.05).Conclusions: CIHH pretreatment can inhibit the degeneration degree of degenerative intervertebral disc in rats.CIHH posttreatment had no significant protective effect on degenerative intervertebral disc in rats.