Effects of alendronate combined with "Shen gu an" on the endplate structure and intervertebral disc degeneration in ovariectomized mice

English Keywords:Intervertebral disc degeneration  Ovariectomy  Endplate  Alendronate  Herb  Mice

English Abstract:

  【Abstract】 Objectives: To observe the effects of alendronate(ALN) combined with tonifying-kidney herbs (herbs, HB) on the endplate leision and intervertebral disc degeneration(IVDD) in ovariectomized mice, and to investigate the mechanism of ALN+HB in the treatment of IVDD. Methods: Thirty-two 12-week-old healthy female C57BL/6J mice were randomly divided into four groups as follows: control group(CT, n=8), the ovariectomized group(OVX, n=8), the alendronate treated group(ALN, n=8) and the group treated with alendronate combining with tonifying-kidney herbs(ALN+HB, n=8). The mice in CT group underwent sham operation, and the mice in the other groups were bilateral ovariectomized. Animals were dosed with medicine three days after surgery and sacrificed at two months later. HE and safranine O-fast green staining were used to detect the morphological changes of intervertebral disc, and the pathological score was recorded. The u-CT scan was used to examine the surface porosity of endplate and the osteophyte formation nearby. The expression of collagen-type Ⅱ(Col2) in the intervertebral disc was detected by immunohistochemistry. Results: No mice died during the experiment. The structure and morphology of intervertebral disc in CT group showed normal in Hematoxylin-Eosin and Safranine O-Fast green staining. In OVX group, severe ossification occurred on the endplate which was more obvious in the caudal side. Besides, the border between the endplate and the nucleus pulposus was disordered with matrix degradation and crack formation. However, the ossification remodeling of the endplate and structural morphology of the nucleus pulposus in ALN group were obviously improved when compared with OVX group, although small ossification lesions in endplates were still existed. In ALN+HB group, there was hardly no ossification remodeling in endplates, and safranin O staining showed deep dying in cartilagous endplate. The IVDD score, endplate surface porosity(%) and Col2 integral optical density(IOD) were 1.20±0.84, 28.60±4.04 and 27764.7±1958.2 in CT group, 4.00±1.58, 56.80±9.39 and 17213.6±1021.3 in OVX group, 2.40±0.89, 36.20±3.42 and 18921.4±888.1 in ALN group, and 2.00±0.71, 29.60±6.19 and 23420.1±2439.6 in ALN+HB group, respectively. Both the pathological score of IVDD and the porosity of the endplate were significantly higher in OVX group than in tCT group, ALN group and ALN+HB group(P<0.05). There was no significant difference among ALN group, ALN+HB group and CT group(P>0.05). The Col2 IOD in OVX, ALN and ALN+HB groups was significantly lower than that in CT group(P<0.05), but had no significant difference between ALN group and OVX group(P>0.05). However, the Col2 IOD in ALN+HB group was significantly higher than that in OVX group and ALN group(P<0.05). Conclusions: Estrogen deficiency in OVX mice can induce ossification remodeling and increasing permeability of endplate, which affects the disc metabolism and results in IVDD. Meanwhile, ALN and ALN+HB can inhibit the ossification remodeling, reduce the porosity, maintain the structural and the functional integrity of endplate, which delays the menopausal related IVDD. Futhermore, ALN＋HB has a better effect on IVDD.