The Mechanism Of Bone Formation And BMP/Smad Pathway Inhibited By AlCl₃

Two hundred 4-week-old healthy male Wistar rats were randomly divided into control group and treatment group. The rats of treatment group were orally exposed Al³⁺ 64 mg/kg body weight in drink water. The rats of control group were given distilled water. A standard laboratory diet was fed to rats with free access to food and water. All the appliances are not the use of aluminum containing products.Twenty five rats in each group were sacrificed under light ether anesthesia on day 30, 60, 90 and 120 respectively. The blood was collected to obtain serum. The femora were separated and stored at-80°C. The Al contents of serum and bone were measured by graphite furnace atomic absorption spectrophotometry; The bone mineral density（BMD） of the distal and proximal femoral metaphysis were measured by dual-energy X-ray absorptiometry（DXA）; The mRNA expressions of BMP-2、BMPRI/II、Smad4、Runx2、Osterix、ALP、ColI and BGP were detected by real-time fluorescence quantitative PCR（q RT-PCR）; The protein expressions of BMP-2、p-BMPRIA、p-Smad1/5/8、Runx2 and Osterix were detected by Western blost; The p-Smad1/5/8/4 protein expression in nucleus was detected by co-immunoprecipitation. The results showed as follows:（1）The Al contents of serum and bone in Al-treated groups were significantly higher than those in the control ones（p<0.01）. It indicated that Al can accumulate in bone of rats.（2）The BMD of the distal and proximal femoral metaphysis were lower in the treatment group than those in control group on day 120（P<0.05）. It indicated that AlCl₃ exposure induced bone lose and inhibited bone formation.（3）The m RNA expressions of ALP and Col I in Al-treated rats were significantly lower than those in the control ones from day 60（p<0.05, p<0.01）; The m RNA expressions of BGP in Al-treated rats were significantly lower than those in the control ones from day 30（p<0.05, p<0.01）. It indicated that AlCl₃ can inhibit the bone formation and the mRNA expressions of BGP can be used as an early diagnostic indicator for the inhibition of bone formation induced by Al.（4）The mRNA expressions of BMPRI/IIand Smad4 in Al-treated rats were significantly lower than those in the control ones from day 60 and day 30（p<0.05, p<0.01）; The protein expressions of BMP-2、p-Smad1/5/8、p-BMPR-IAin Al-treated rats were significantly lower than those in the control ones from day 90, 90, 60, respectively（p<0.05, p<0.01）;The p-Smad1/5/8/4 protein expression in nucleus of Al-treated rats were significantly lower than those in the control ones from day 60（p<0.05, p<0.01）. It indicated that AlCl₃ can inhibit BMP/Smad singal pathway.（5）The m RNA and protein expressions of Runx2 and Osterix in Al-treated rats were significantly lower than those in the control ones from day 30（ p<0.01）. It indicated that AlCl₃ inhibited the activity of nuclear transcription factors in bone.The above results showed that AlCl₃ can down-regulate the BMP/Smad pathway and inhibit the activity of nuclear transcription factors in bone, then decrease the ALP 、Col I and BGP mRNA expressions, inhibiting bone formation finally. Moreover, the mRNA expressions of BGP can be used as an early diagnostic indicator for the inhibition of bone formation induced by Al.