The Effects Of The Related Factors Of Bone And Kidney In The Pathological Mechanism Of The Type2Diabetes And Osteoporosis

Objective: Osteoporosis (OP) is a kind of whole body bone diseasecharacterized by bone loss and bone microstructure destroy, which lead toincreased bone fragile and high risk of fracture. Type2diabetes is a metabolicdisorder that is characterized by high blood glucose in the context of insulinresistance and relative insulin deficiency, with the lifestyle and environmentchanging, the incidence of the two diseases has increased year by year.Though traditionally these two diseases are entirely independent, more andmore research evidenced that type2diabetes and osteoporosis influence eachother, and between them may be had common pathophysiologic mechanisms,but the connect signaling pathway or mechanisms is unclear. In recent years,the relevant research showed that the bone and kidney are two important targetorgans which can both affect osteoporosis and type2diabetes, but whethercertain mechanisms or common factors through the kidney and bone toinfluence osteoporosis and type2diabetes is no definite report.Insulin signaling pathway plays an important role in the occurrence of thetype2diabetes, especially the central factors: insulin growth factor-1(IGF-1),insulin receptor substrate-1(IRS-1) and insulin receptor substrate-2(IRS-2).When the pathway inhibited, insulin resistance or the type2diabetes mayhappened. The genes research found that knock out IGF-1, IRS-1and IRS-2ordecreased expression levels could induce bone mass reduced and bonestructure abnormality. It has been reported that the bone and kidney mayexpress IGF-1, IRS-1or IRS-2, but how they expressed in the bone and kidneyof type2diabetes and osteoporosis together is not clear. Vitamin D is animportant hormone that influence bone growth and mineralization, thesynthesis of its active form1,25(OH)2D3depends on the structure and function of bone, kidney and intestinal tract complete well,1,25(OH)2D3plays the roleneed combine with vitamin D receptor(VDR). We have known that1,25(OH)2D3insufficiency can reduce bone mass and result to the occurrenceof osteoporosis, and recently many studies had found1,25(OH)2D3insufficiency was associate with type2diabetes. The fibroblast growth factor23(FGF23) and bone morphogenetic protein7(BMP7) are two new importantfactors which can express in both the bone and kidney, and found them have aregulatory function to1,25(OH)2D3and also can affect VDR expression, buthow VDR, FGF23and BMP7express in the bone and kidney of type2diabetes and osteoporosis is no definite report. Type2diabetes affectosteoporosis may be through influence bone formation, fibroblast growthfactor2(FGF2) and bone morphogenetic protein2(BMP2) are the twoimportant factors that play the role in the process of osteoblast bone formation.The expression levels of them decreasing may inhibit the effect of osteoblastin the process of bone formation, papers showed two factors can express in thebone and kidney, but how they express in the type2diabetes and osteoporosisis not clear.This study on the basis of set up type2diabetes and osteoporosis ratmodel, to analysis the relevant factors which can express in both the bone andkidney but has different mechanisms,to clear these factors expression in thebone and kidney of type2diabetes and osteoporosis rats, to provide newthoughts for the pathological mechanism of the type2diabetes andosteoporosis.Methods:Part1:After ovariectomized4,8and12weeks, each5rats choserandomly to separate the bone and renal tissues, the paraffin-embedded renaltissue sections were stained with HE to observe the pathological changes oftissue and cell structure.Used immunohistochemistry to observe the renaltissue express sites of IGF-1, IRS-1and IRS-2in each group, used RT-PCR toclear the IGF-1, IRS-1and IRS-2mRNA expression levels of the bone andkidney in each group, used western blot to measure the IGF-1, IRS-1and IRS-2protein expression levels of the kidney in12weeks each group.Part2:After ovariectomized4,8and12weeks, each5rats choserandomly to separate the bone and renal tissues. Used immunohistochemistryto observe the renal tissue express sites of VDR, FGF23and BMP7in eachgroup, used RT-PCR to clear the VDR, FGF23and BMP7mRNA expressionlevels of the bone and kidney in each group, used western blot to measure theVDR, FGF23and BMP7protein expression levels of the kidney in12weekseach group.Part3: After ovariectomized4,8and12weeks, each5rats choserandomly to separate the bone and renal tissues. Used immunohistochemistryto observe the renal tissue express sites of FGF2and BMP2in each group,used RT-PCR to clear the FGF2and BMP2mRNA expression levels of thebone and kidney in each group, used western blot to measure the FGF2andBMP2protein expression levels of the kidney in12weeks each group.Results:Part1: Compared to the non-diabetic groups(NS and NOVX groups), thediabetic groups(DS and DOVX groups) showed increased glomerular volume,expanded medullary loop cavity.IGF-1, IRS-1and IRS-2expressed mainly inthe cytoplasm of renal tubular epithelial cells in each group renal tissues.IHSgrade result showed that the positive expression of IGF-1, IRS-1and IRS-2inDOVX group were lower than other three groups.The result of bone RT-PCRshowed that at4,8and12weeks, the IGF-1, IRS-1and IRS-2mRNAexpression levels in DS and DOVX groups has different degree decreasing,compared with control groups, the IGF-1, IRS-1and IRS-2mRNA expressionlevels of NOVX group were lower than DS group at12weeks (P<0.05),andat12weeks the IGF-1, IRS-1and IRS-2mRNA expression levels of DOVXgroup were lowest. The result of kidney RT-PCR showed that at4,8and12weeks, the IRS-1and IRS-2mRNA expression levels in NOVX group werelower than NS group, the IGF-1mRNA expression level in DS group waslower than NOVX group at8and12weeks(P<0.05).The results of12weekskidney Western blot showed that the IGF-1protein expression level in DS and DOVX groups was lower than NS and NOVX groups, DS group was lowerthan NOVX group(P<0.05); the IRS-1and IRS-2protein expression levels inDS and DOVX groups were lower than NS group(P<0.05), but no significantdifference between NOVX and NS groups(P>0.05).Part2: Immunohistochemistry results showed VDR, FGF23and BMP7expressed mainly in the cytoplasm of renal tubular epithelial cells in eachgroup renal tissues. IHS grade result showed that the positive expression ofVDR in DOVX and NOVX groups were lower than NS group, DS group waslower than NS group at12weeks (P<0.05), the positive expression of FGF23in DOVX and DS groups were higher than NOVX and NS groups at8and12weeks, and the number in DOVX was highest (P<0.05); the positiveexpression of BMP7in DOVX and DS groups were lower than NS group,andthe number in DOVX was lowest (P<0.05). The result of bone RT-PCRshowed that at4,8and12weeks, the VDR, FGF23and BMP7mRNAexpression levels in DS and DOVX groups has different degree decreasing,compared with control groups, and the VDR and BMP7mRNA expressionlevels of NOVX group were lower than NS group at12weeks (P<0.05), theVDR, FGF23and BMP7mRNA expression levels of DOVX group werelowest. The result of kidney RT-PCR showed that at4,8and12weeks, theVDR and BMP7mRNA expression levels in DS and DOVX groups werelower than NS and NOVX groups, and the BMP7mRNA expression level inDS group was lower than NOVX group (P<0.05), but the FGF23mRNAexpression levels in DS and DOVX groups were higher than NS group at8and12weeks (P<0.05), the expression of DOVX group was highest. Theresults of12weeks kidney Western blot showed that the VDR proteinexpression level in NOVX and DOVX groups were lower than NS and DSgroups, NOVX group was lower than DS group(P<0.05); the BMP7proteinexpression level in DS and DOVX groups were lower than NS group, and DSgroup was lower than NOVX group(P<0.05); the FGF23protein expressionlevel in DS and DOVX groups were higher than NS group,and DOVX washigher than NOVX group(P<0.05). Part3: Immunohistochemistry results showed FGF2and BMP2expressed mainly in the cytoplasm of renal tubular epithelial cells in eachgroup renal tissues. IHS grade result showed that the positive expression ofFGF2in DS group was higher than NS and NOVX groups (P<0.05)ï¼›thepositive expression of BMP2in DOVX and DS groups were lower thanNOVX and NS groups at8and12weeks,the number in DOVX was lowest(P<0.05). The result of bone RT-PCR showed that at4,8and12weeks, theFGF2and BMP2mRNA expression levels in DS and DOVX groups hasdifferent degree decreasing, compared with NS group, and the FGF2andBMP2mRNA expression levels of NOVX group were lower than DS group at12weeks (P<0.05), the mRNA expression levels of DOVX group were lowest.The result of kidney RT-PCR showed that in4weeks, no significantdifference between each group, the FGF2and BMP2mRNA expression levelsin NOVX and DOVX groups were lower than NS group, but the FGF2mRNAexpression level in DS group was higher than NS group at8and12weeks(P<0.05), the FGF2expression of NOVX group was lowes at12weeks, nodifference of BMP2between DS and DOVX. The results of12weeks kidneyWestern blot showed that the FGF2protein expression level in DS group werehigher than other three groups, the BMP2protein expression level in NOVXand DS groups was lower than NS group (P<0.05); no difference of BMP2between DS and NOVX groups, the BMP2protein expression level in DOVXwas lowest.Conclusions:1The IGF-1, IRS-1and IRS-2expression levels of the bone and kidneyhas different degree reduction in type2diabetes and osteoporosis rats, theconnect changes of insulin signaling pathway in both bone and kidney may beprovide new thoughts for the pathological mechanism of type2diabetes andosteoporosis;2FGF23expressed different from VDR and BMP7in the bone andkidney of type2diabetes and osteoporosis, it is probably provide new thoughtfor research the effect of Vitamin D in the pathological mechanism of type2 diabetes and osteoporosis;3The FGF2and BMP2expression levels decreased in the bone of type2diabetes and osteoporosis rats, and the BMP2expressed reduction in thekidney of type2diabetes and osteoporosis rats, the FGF2expression in simpletype2diabetes group was higher than control rats, but this change has nodirect relationship with the occurrence of osteoporosis, this provide new ideafor study the effect of bone formation factors in the pathological mechanism oftype2diabetes and osteoporosis.