Effects of angiotensin II type 1 receptor blocker on bones in mice with type 1 diabetes induced by streptozotocin.

INTRODUCTION: This study was performed to address the pathological roles of the skeletal renin-angiotensin system (RAS) in type 1 diabetes-induced osteoporosis and the effects of the angiotensin II type 1 receptor blocker losartan on bones in diabetic mice. MATERIALS AND METHODS: Bone histomorphology was detected by H&E staining, Safranin O staining and X-ray radiography. Micro-CT was performed for the analysis of bone parameters. Gene and protein expression were determined by RT-PCR and immunoblotting. RESULTS: Type 1 diabetic mice displayed osteopenia phenotype, and losartan treatment had no osteoprotective effects on diabetic mice as shown by the reduction of bone mineral density and microarchitectural parameters at the proximal metaphysis of the tibia. The mRNA expression of AGT, renin receptor and ACE, and protein expression of renin and AT1R were markedly up-regulated in the bones of vehicle-treated diabetic mice compared to those of non-diabetic mice. The treatment with losartan further significantly increased the expression of AGT, renin, angiotensin II and AT1R, and reduced the expression of AT2R receptor as compared to those of diabetic mice. CONCLUSION: Local bone RAS functionally played a role in the development of type 1 diabetic osteoporosis, and losartan had no bone-sparing function in diabetes mice because of enhance skeletal RAS activity.

Effects of angiotensin-converting enzyme inhibitor, captopril, on bone of mice with streptozotocin-induced type 1 diabetes.

There are contradictory results about the effect of angiotensin-converting enzyme inhibitors (ACEIs) on bone. This study was performed to address the skeletal renin-angiotensin system (RAS) activity and the effects of the ACEI, captopril, on the bone of streptozotocin-induced type 1 diabetic mice. Histochemical assessment on bone paraffin sections was conducted by Safranin O staining and tartrate-resistant acid phosphatase staining. Micro-computed tomography was performed to analyze bone biological parameters. Gene and protein expression were determined by real-time polymerase chain reaction and immunoblotting, respectively. Type 1 diabetic mice displayed osteopenia phenotype and captopril treatment showed no osteoprotective effects in diabetic mice as shown by the reduction of bone mineral density, trabecular thickness and bone volume/total volume. The mRNA expression of ACE and renin receptor, and the protein expression of renin and angiotensin II were markedly up-regulated in the bone of vehicle-treated diabetic mice compared to those of non-diabetic mice, and these molecular changes of skeletal RAS components were effectively inhibited by treatment with captopril. However, treatment with captopril significantly elevated serum tartrate-resistant acid phosphatase 5b levels, reduced the ratio of osteoprotegerin/receptor activator of nuclear factor-κB ligand expression, increased carbonic anhydrase II mRNA expression and the number of matured osteoclasts and decreased transforming growth factor-ß and osteocalcin mRNA expression in the tibia compared to those of diabetic mice. The present study demonstrated that the use of the ACEI, captopril, has no beneficial effect on the skeletal biological properties of diabetic mice. However, this could be attributed, at least partially, to its suppression of osteogenesis and stimulation of osteoclastogenesis, even though it could effectively inhibit high activity of local RAS in the bone of diabetic mice.

Trabecular bone deterioration at the greater trochanter of mice with unilateral obstructive nephropathy.

Our previous study showed the early molecular responses of bone in response to obstructive nephropathy in a unilateral ureteral obstruction (UUO) mouse model. Here, we addressed the changes in trabecular bone properties at greater trochanter, the proximal and the distal metaphysis of femur in UUO mice. The male mice were subjected to UUO (n=10) or sham operation (n=10). All mice were killed on day 7 after the surgical operation. The micro-computed tomography (micro-CT) analysis for different femoral trabecular bone sites demonstrated pathological alterations of trabecular bone mass and micro-networks at greater trochanter as shown by decreases in bone mineral density/bone volume (P<0.05) and trabecular number (P<0.05) and increases in trabecular separation (P<0.01) and bone surface/bone volume (P<0.05) in UUO mice. The present study demonstrates that UUO-induced unilateral obstructive nephropathy has markedly detrimental effects on the trabecular trochanter of the femur.

Early molecular responses of bone to obstructive nephropathy induced by unilateral ureteral obstruction in mice.

AIM: This study was performed to address the bone injury and the early molecular responses of bone to obstructive nephropathy induced by unilateral ureteral obstruction in mice. METHODS: The male mice were subjected to unilateral ureteral obstruction (UUO, n = 10) or sham operation (n = 10). All mice were killed on day 7 after the surgical operation. Hematoxylin and eosin and tartate-resistant acid phosphatase staining were performed on paraffin-embedded bone sections. Expression of genes and proteins was analyzed by reverse transcription-polymerase chain reaction, and Western blotting and immunohistochemistry staining, respectively. RESULTS: The serum calcium level was significantly reduced in UUO mice compared with that of Sham mice. The proximal tibia of UUO mice exhibited the increased expansion of chondrocytes zone, the reduction of osteoid content, and the increased separation and disconnection of woven bones. Reverse transcription-polymerase chain reaction results showed the downregulation of Cbfa1 and Col mRNA expression and the upregulation of Tgf-ß, CtsK, CaII, Opg and Rankl mRNA expression in tibia of UUO mice compared to those of Sham mice. The ratio of Opg and Rankl was unchanged between Sham and the UUO group. Local protein expression of angiotensin II and its type 2 receptor was dramatically upregulated in tibia of UUO mice. CONCLUSION: Together, it is concluded that the obstructive nephropathy has defective effects on bone, and the underlying mechanisms are the reduction of bone formation and the increase of bone resorption, which is mediated, at least partially through local angiotensin II signalling.

Involvement of the skeletal renin-angiotensin system in age-related osteoporosis of ageing mice.

The local tissue-specific renin-angiotensin system (RAS) was identified. The aim of this study was to investigate the role of local bone RAS in the osteoporosis of aging mice. Twelve-month-old and two-month-old male mice were respectively assigned to the ageing and young groups. The tibias and femurs were collected for an analysis of histomorphology, bone mass, and gene and protein expression. H&E staining and micro-CT measurement showed a loss of the trabecular bone network and decrease of bone mineral density in the proximal tibial metaphysis of the aged mice. The PCR results indicated the significant up-regulation of renin and angiotensinogen (AGT) mRNA expression in both the tibia and femur of the ageing mice. Western blotting data showed that the tibial angiotensin II protein expression was significantly increased in the ageing group. The enhancement of renin and AGT expression in the bone tissue resulted in the increased production of angiotensin II which plays an important role in the pathology of age-related osteoporosis.

Changes of renal vitamin D metabolic enzyme expression and calcium transporter abundance in obstructive nephropathy.

The effects of urinary-tract obstruction on renal function have been clarified. However, there is little known about the change of renal vitamin D metabolic enzyme expression and vitamin D-dependent calcium transporting proteins expression in obstructive nephropathy. The male mice were subjected to unilateral ureteral obstruction (n = 10) or sham operation (n = 10). All mice were killed on day 7 after the surgical operation. Kidney sections were stained with Masson's trichrome and gene expression was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and real-time PCR. The obstructed kidney exhibited interstitial fibrosis as shown by the strong collagen deposition in the interstitium. Quantitative PCR results showed the increase of 1-OHase (P < 0.001) mRNA expression and the decrease of 24-OHase (P < 0.01), CaBP-9k (P < 0.01) and CaBP-28k (P < 0.01) mRNA expression in obstructed kidney as compared to that of the Sham group. In addition, the mRNA expression of 1-OHase and CaBP-9k was significantly increased and decreased, respectively, in obstructed kidney as compared to that of the contra-lateral kidney in unilateral ureteral obstruction (UUO) mice. Together, the present finding supports the hypothesis that the ureteral obstruction leads to the alteration of renal vitamin D metabolic enzyme expression and calcium transporter abundance, which may secondarily induce the abnormality of vitamin D endocrine system and bone health.