甲状旁腺激素相关蛋白对去卵巢大鼠骨组织中OPG、RANKL基因表达的影响

目的观察不同剂量间歇性注射甲状旁腺激素相关蛋白氨基端片段1—34（PTHrP1—34）对去卵巢大鼠骨密度及股骨RANKL、OPG基因表达的影响。方法4月龄健康雌性未孕Wistar大鼠60只，随机分为6组，假手术组（Sham组）和卵巢切除＋安慰剂组（Placebo组）给予生理盐水；卵巢切除＋雌激素治疗组（E2组）给予苯甲酸雌二醇注射液；卵巢切除＋PIHrP治疗组分别用20、40、80ktg／kg剂量，每日1次注射PTHrP1—34。给药12周后，测定腰椎L3-6及股骨BMD，利用实时荧光定量RT-PCR方法检测大鼠股骨RANKL、OPG基因表达的水平。结果①BMD结果：PTHrP40组和PTHrPS0组大鼠股骨、腰椎BMD明显高于Placeb0组。②与Sham组相比，Placebo组OPGmRNA明显下降（P〈0．01），RANKLmRNA水平则显著升高（P〈0．01）。E2组OPGmRNA水平较Placebo组明显升高（P〈0．01），RANKLmRNA水平显著下降（P〈0．01）。与Placebo组相比，不同剂量胛HrP（1-34）治疗组OPG、RANKLmRNA水平无明显变化（P〉0．05）。结论PTHrPl—3440或80ug／kg每天皮下注射能提高去卵巢大鼠股骨、腰椎BMD，间歇性注射PTHrP（1-34）对去卵巢大鼠骨组织RANKL、OPGmRNA水平无明显影响。     

rhPTH（1-34）治疗OVX大鼠骨质疏松延后效应的初步研究

目的观察重组人甲状旁腺激素（1-34）[rhPTH（1-34）]对卵巢摘除（OVX）大鼠骨质疏松症的治疗作用及停药后效应.方法应用双侧卵巢摘除方法建立模拟绝经后骨质疏松大鼠模型;给予皮下注射20μg/kg/d rhPTH（1-34）治疗8周,观察其骨量、骨生物力学、骨小梁形态计量及骨代谢相关血、尿生化指标,综合评价PTH对模型大鼠的治疗效果;同时观察停药8周后上述指标的变化.结果外源性PTH（1-34）治疗能显著增加模型大鼠的骨量、骨力学性能,改善骨微结构、增加骨转换.用药组的骨密度、股骨三点弯曲与腰椎压缩最大载荷、腰椎骨小梁百分面积显著高于对照组（P＜0.05～0.001）;血ALP（P＜0.05～0.01）与尿Pyd（P＜0.05）保持高水平;PTH停药8周后大鼠股骨与腰椎骨密度、股骨三点弯曲与腰椎压缩最大载荷及腰椎骨小梁百分面积均较停药前显著降低（P＜0.05～0.001）,但仍显著高于OVX对照组（P＜0.05～0.001）.结论外源性PTH（1-34）可显著增加OVX大鼠的骨量,提高骨力学性能,改善骨微结构,对卵巢摘除诱导的大鼠骨质疏松具有明显治疗作用;停药后出现骨量的快速丢失,骨力学性能下降等变化,但仍显示出其对OVX大鼠骨骼的保护作用.     

去卵巢对大鼠骨密度的影响

目的：探讨去卵巢对大鼠骨密度（BMD）的影响。方法；20只3．5月龄SD雌性大鼠分别除双侧卵巢（OVX）或假性去卵巢（Sham)，术后14周处死，应用QDR－4500A型扇形束双能X射线吸收法（DXA）测量大鼠全身、离体股骨、胫骨、腰维及兴趣区的BMD。结果：①术后6周OVX组全身BMD显低Sham组（P＝0．048），术后14周两组无显性差异；②术后14周OVX组离体股骨BMD显低于Sham组（P＜0．01），股骨远侧干骺端平均降低11．6％（P＜0．001）；③术后14周右侧离体胫骨BMD两组间差异无显性，但OVX组胫骨的端干骺端BMD显低于Sham组（P＜0．001）；④术后14周OVX组腰椎（L4－L6）的BMD显低于Sham组（P＝0．014），第六腰椎降低明显，平均降低8．1％（P＝0．005）。结论：去卵巢所致骨丢失以松质骨含量丰富的兴趣区明显。     

雌激素治疗去卵巢大鼠骨丢失的不同部位效应观察

目的探讨雌激素对去卵巢(OVX)大鼠骨丢失的作用敏感部位。方法8月龄SD大鼠去卵巢后,皮下注射17β-雌二醇(E2)(10、30μg.kg-1.d-1)3月,双能X线骨密度仪检测全身总骨密度(BMD),离体腰椎(L4-6)、股骨、胫骨BMD。实验分为假手术组、去卵巢模型组、雌激素小剂量组、雌激素大剂量组,每组10只。结果E2可逆转OVX所致总BMD降低,增加OVX后腰椎整体及3个兴趣区BMD。E2治疗可升高OVX后股骨BMD,其中股骨整体、股骨近端和股骨远端处改变最为明显(P<0.01),其余各兴趣区BMD改变相对稍弱,而股骨中段最不敏感。胫骨BMD指标中以胫骨近端最为敏感(P<0.01),而中远端几乎无改变。结论雌激素对去卵巢(OVX)大鼠的作用敏感部位与骨量丢失敏感部位具有一致性,雌激素增加骨密度主要在腰椎、股骨近端、股骨远端和胫骨近端。     

rhPTH(1-34)对骨质疏松大鼠的作用及对sclerostin的影响

[目的]探讨人重组甲状旁腺素1-34(rhPTH1-34)对骨质疏松的治疗作用以及与血钙、磷代谢和生长因子的关系。[方法]用摘除大鼠双侧卵巢的方式制备骨质疏松模型,实验动物分为3个组:模型对照组(OVX组,摘除大鼠双侧卵巢不作任何处理);rhPTH1-34治疗组(PTH组,摘除大鼠双侧卵巢12周后用rhPTH1-34治疗8周);假手术组(sham组,仅切除卵巢周围的脂肪组织约3 g,术后12周纳入实验)。应用第4代双能X线骨密度仪测量大鼠股骨上段骨密度值(BMD);用ELISA法测定血清硬化蛋白(sclerostin)水平及骨钙素(BGP)浓度;用自动生化仪测定血清碱性磷酸酶(ALP)。[结果]rhPTH1-34治疗组、sham组均较OVX组股骨上段骨密度增高,组间比较差异有显著性(P<0.01)。rhPTH1-34治疗组血清BGP浓度值升高及sclerostin值降低,与OVX组比较差异有显著性(P<0.01)。各组血清钙、磷含量无明显变化,与OVX组比较差异无显著性,ALP值治疗组与OVX组无明显差异。[结论]rhPTH1-34能够预防股骨上段骨密度丢失,并且血清BGP浓度值升高及sclerostin值降...     

糖皮质激素对成年大鼠骨量的影响

目的利用双能X线吸收法(DXA)探讨成年大鼠接受糖皮质激素后骨量变化的规律。方法 21只44周龄SD雌性大鼠分别假性去卵巢+未注射糖皮质激素(SHAM组)、摘除双侧卵巢(OVX组)或注射甲基强的松龙[2.5 mg/(kg·d)](PRED组),应用扇形束DXA(QDR-4500A)每4周测定一次全身骨密度(BMD)、骨矿含量(BMC)、骨骼面积(AREA);术后12周处死,测定离体腰椎、股骨、胫骨及其兴趣区的BMD、BMC、AREA。压缩试验测定第二腰椎最大载荷和弹性模量。结果 (1)术后8周开始OVX组体重显著重于同龄SHAM组(8周时,P0.05,12周时P0.01),术后4周开始PRED组体重显著轻于同龄SHAM组(P0.05);(2)术后12周OVX组整体BMC显著高于SHAM组(P0.05),术后8、12周OVX组整体BMC显著高于PRED组(P0.05);(3)术后12周OVX组离体第5、6腰椎BMD及第6腰椎BMC显著低于SHAM组和PRED组(P0.05),PRED组离体各腰椎BMD、BMC、AREA与SHAM组无明显差异;(4)术后12周与SHAM组比较,OVX组离体股骨(-7.42%)、股骨远端(-10.85%)和近端(-6.92%)、胫骨近端(-11.40%)BMD显著降低(P0.05),其中股骨、股骨远端、胫骨近端BMC也显著降低(P0.05);(5)术后12周与SHAM组比较,PRED组离体股骨及各区BMD、BMC、AREA无显著性差异,整体胫骨及各区BMD无显著性差异;(6)术后12周与SHAM组比较,OVX组及PRED组胫骨中远端骨量增加;(7)与SHAM组比较,OVX组最大载荷和弹性模量显著降低,PRED组最大载荷显著降低。结论成熟期大鼠接受甲基强的松龙后,皮质骨和松质骨骨量没有显著变化,DXA检查难以发现其骨丢失情况;力学性能改变提示糖皮质激素更多的是引起骨质量的改变而导致了力学性能的下降及骨折的发生。     

DXA测量活体大鼠骨的精密性及骨丢失的检测

目的 了解QDR－4500A型双能X射线吸收法（DXA）测量活体大鼠的精密性和探测去卵巢后大鼠骨丢失的能力。方法 测量15只体重为202－311g的SD大鼠全身、股骨及腰椎的骨密度（BMD）,每只大鼠测量3次,可得变异系数（CV）,15只大鼠CV的平均值为该指标的批内CV。结果 ①全身、股骨、腰椎BMD的批内CV分别为0．71％、2．02％和2．44％。全身BMD的批内CV显著低于股骨和腰椎（P＜0．05）;②全身BMD的批间CV为0．99％,股骨整体为2．81％,腰椎总体（L3－L6）为3．42％;③术后4周去卵巢组全身、股骨、腰椎BMD与假手术组比较无显著性变化,而股骨远侧干骺端BMD低于假手术组（P＜0．05）;④去卵巢后14周腰椎总体（L4－L6）的BMD低于假手术组（P＜0．05）。结论 QDR－4500A型DXA测量大鼠全身、股骨和腰椎BMD有较好的精密性,全身优于局部骨骼;其精密性能满足检出去卵巢后骨丢失。     

激素与去卵巢诱导大鼠骨质疏松症的差异及其分子机制探讨

目的 比较激素及去卵巢对大鼠骨量、骨转换指标、雌激素水平、骨髓细胞成脂分化水平的影响,并探讨两种模型可能的分子机制。方法 3月龄雌性SD大鼠36只,随机分为空白组、甲强龙组（MP组）、去卵巢组（OVX组）。3组大鼠于8w后处死,测量体重及子宫、肾上腺重量;双能X线骨密度仪测量左股骨BMD、BMC、BA;ELISA法测量血清PINP、β-CTX及雌激素水平;油红“O”染色观察右股骨骨髓间充质干细胞（BMSCs）成脂情况。RT-PCR检测左胫骨Runx2、Col1α1、MMP9 mRNA表达。结果 术后8w,MP组体重显著低于空白组及OVX组,OVX组体重明显高于空白组（P<0.05）。MP组BMC、BMD、BA及OVX组BMC、BMD均较空白组明显降低,且MP组BMC、BMD、BA明显低于OVX组（P<0.05）。两组血清PINP、β-CTX均较空白组显著升高,且MP组明显高于OVX组,两组雌激素水平均显著低于空白组（P<0.05）。两组骨髓脂滴均较空白组增多,骨组织Runx2 mRNA表达均较空白组显著下调（P<0.05）,Col1α1、MMP9 mRNA表达较空白组高,但比较差异无统计学意义。结论 激素及去卵巢均可诱导大鼠高转换型骨质疏松症,以激素导致的骨量丢失更严重,且两者均可下调大鼠血清雌激素水平、促进骨髓细胞成脂分化,其机制可能均与调控骨组织Runx2 mRNA表达水平密切相关。     

甲状旁腺激素促进系膜细胞合成分泌转化生长因子β1

目的：研究甲状旁腺激素（PTH）对大鼠系膜细胞合成与分泌转化生长因子β1（TGF－β1）的影响。方法：（1）分别以10^-12,10^-11,10^-10,10^-9,10^-8mol/l的hPTH1-34刺激大鼠系膜细胞6，12，24，48h后，ELISA方法测定上清中TGF－β1的浓度；（2）分别以10^-12,10^-11,10^-10,10^-9,10^-8mol/L的hPTH1-34刺激大鼠系细胞48h,采用半定量RT－PCR方法检测细胞TGF－β1 mRNA的表达；（3）以10^-8mol/L的hPTH1-34分别刺激大鼠系膜细胞6，12，24，48h，采用半定量RT－PCR方法检测细胞TGF－β1 mRNA的表达，结果：（1）ELISA结果显示，hPTH1-34促进大鼠系膜细胞合成与分泌TGF－β1分泌作用达高峰（P＜0．01），（2）半定量RT－PCR方法结果显示，hPTH1-34促进大鼠系膜细胞TGF－β1 mRNA的表达，并具有浓度依赖和时间依赖性特点（各组与对照组间P＜0．05），结论：hPTH1-34从蛋白和基因水平显著促进系膜细胞TGF－β1的合成与分泌，且呈浓度依赖和时间依赖性特点。     

五味子甲素对去卵巢骨质疏松大鼠SDF-1/CXCR4信号通路的影响

目的 探究五味子甲素对去卵巢骨质疏松症大鼠的治疗作用以及对基质细胞衍生因子-1（SDF-1）/CXC趋化因子受体4（CXCR4）信号通路的影响。方法 采用去卵巢法构建骨质疏松症大鼠模型，将建模成功的60只大鼠随机分为模型组、五味子甲素低（20 mg/kg）、高（40 mg/kg）剂量组、阿仑膦酸钠（1 mg/kg）组、五味子甲素高剂量+SDF-1/CXCR4通路抑制剂（AMD3100）（40 mg/kg+5 mg/kg）组，每组12只。另取12只大鼠作为假手术组（手术过程中仅暴露卵巢不切除）。建模结束后，各组给予对应药物干预，每天1次，连续12周。采用双能 X 射线骨密度仪检测大鼠左、右股骨骨密度（BMD）；酶联免疫吸附法检测大鼠血清中Ⅰ型胶原交联羧基末端肽（CTX-Ⅰ）、骨钙素（OC）水平；HE染色观察大鼠股骨组织病理学变化；荧光定量PCR法检测大鼠股骨组织中SDF-1、CXCR4信使RNA（mRNA）水平；蛋白印迹法检测大鼠股骨组织中SDF-1、CXCR4蛋白水平。结果 假手术组大鼠股骨组织骨小梁结构正常，排列规则；与假手术组相比，模型组大鼠股骨组织骨小梁稀疏断裂，数量减少，排列紊乱，左、右股骨BMD、血清中OC、CTX-Ⅰ水平、股骨组织中SDF-1、CXCR4 mRNA和蛋白水平显著降低（P<0.05）；与模型组相比，五味子甲素低、高剂量组大鼠股骨组织病变程度依次减轻，左、右股骨BMD、血清中OC、CTX-Ⅰ水平、股骨组织中SDF-1、CXCR4 mRNA和蛋白水平依次升高（P<0.05）；阿仑膦酸钠组和五味子甲素高剂量组大鼠股骨组织病理学变化及各项指标差异比较均无统计学意义（P>0.05）；AMD3100可逆转高剂量五味子甲素对骨质疏松症大鼠上述指标的改善效果（P<0.05）。结论 五味子甲素可抑制骨质疏松症大鼠BMD的下降，减轻大鼠股骨组织病变，改善大鼠骨质疏松，其机制可能与激活SDF-1/CXCR4信号通路有关。     

Human parathyroid hormone (1-34) increases mass and structure of the cortical shell,with resultant increase in lumbar bone strength,in ovariectomized rats

Estrogen deficiency causes reduction of bone mass and abnormal bone microarchitecture, consequently reducing bone strength. Human parathyroid hormone (hPTH) (1-34) increases bone mass and strength. To clarify the factors that determine the recovery of bone strength in the lumbar vertebrae of ovariectomized rats by intermittent hPTH administration, we analyzed the relationship between skeletal measurements and bone strength. Human PTH (1-34) administration resulted in recovery of cortical bone mineral content (BMC) and cortical bone area to sham the levels, but in resulted in a less pronounced recovery of trabecular BMC and no increase in the total cross-sectional area of the vertebral body. Of the three-dimensional (3D) trabecular bone parameters, hPTH (1-34) increased trabecular thickness (Tb.Th). The cortical shell area of L4, determined by histomorphometry, was also increased. In hPTH-treated rats, the only determinant of the compressive load of L5 was the cortical shell BMC, in the early recovery period (days 42–84). Our data suggest that increased cortical bone mass contributes more than trabecular bone mass and structure to the recovery of bone strength in response to hPTH therapy in the rat lumbar vertebral body after ovariectomy.     

Maintenance of increased bone mass after recombinant human parathyroid hormone (1-84) with sequential zoledronate treatment in ovariectomized rats.

The concept of lose, restore, maintain (LRM) for reversing existing osteoporosis was tested in rats. The withdrawal of PTH results in the loss of the acquired bone mass, but sequential therapy with zoledronate quite effectively maintained the PTH(1-84)-acquired bone quantity and quality. INTRODUCTION: Because antiresorptive agents against osteoporosis are presently quite limited, strong anabolic agents such as human parathyroid hormone (hPTH) are quite helpful. However, because hPTH(1-34) is available only through injection and has a critical side effect of causing bone tumors during life-long administration in the rat, it would be practical to use PTH for the shortest possible duration to obtain the maximal effect. To determine the effectiveness of the osteoporosis-reversing concept of lose, restore, and maintain (LRM), recombinant hPTH(1-84) [rhPTH(1-84)] and the respective antiresorptive agents were sequentially studied. MATERIALS AND METHODS: Thirty-six, 20-week-old Sprague-Dawley rats were used. Treatment started at the 25th week after ovariectomy, which was performed at 20 weeks of age, with 5 weeks of rhPTH(1-84) 100 microg/kg/day, 5 days/week, followed by the respective sequential therapies for 5 weeks as follows: (1) ovariectomized rats (OVX; n = 6), (2) sham-operated rats (SHAM; n = 6), (3) OVX rats with PTH maintenance (PTH-M; n = 6), (4) OVX rats treated with PTH and then PTH was withdrawn (PTH-W; n = 6), (5) PTH-treated OVX rats treated with 17beta-estradiol (PTH-E; 10 microg/day SC, 5 days/week; n = 6), and (6) PTH-treated OVX rats treated with zoledronate (PTH-Z; 12.5 microg/kg SC weekly; n = 6). BMD of the right femora was measured by DXA. microCT was used to measure the structural parameters of the second lumbar vertebrae. Three-point bending test of the femora and compressive tests of vertebrae were also performed. RESULTS: Bone quantity data showed that the BMD and most of the microstructural parameters were significantly higher in the PTH-M and PTH-Z groups than in the OVX and PTH-W groups (p < 0.05). Measurement of the cortical thickness revealed that only the PTH-M group showed a significant increase (p = 0.001). The ultimate force (Fu) at the midshaft of the femora was similar in the treated groups and stronger than in the OVX group (p < 0.05). However, in the vertebrae, the Fu of the PTH-M and PTH-Z groups was significantly higher, by approximately 44-47%, than in the OVX and PTH-E groups and showed a higher tendency than in the PTH-W group. CONCLUSION: PTH withdrawal resulted in the loss of acquired BMD, and sequential therapy with antiresorptives prevented further loss (17beta-estradiol versus zoledronate). The zoledronate after rhPTH(1-84) as a sequential regimen was quite consistently effective.     

Effect of parathyroid hormone (hPTH[1-84]) treatment on bone mass and strength in ovariectomized rats.

Skeletal fragility in osteoporotic patients is a prominent underlying cause of low-trauma fractures of most bone sites in humans. Clinical research is now focused on developing treatment strategies, including anabolic agents such as parathyroid hormone (PTH), to recover osteoporosis-related bone loss. Female Sprague-Dawley rats (4.5 mo old) were allowed to become osteopenic for 10 wk postovariectomy. Eight rats were killed at the time of ovariectomy (-10 wk) as a baseline control; sham and ovariectomized (OVX) groups were killed at wk 0. Eight rats per group (sham, OVX + vehicle, OVX + hPTH [5 d/wk], and OVX + hPTH [3 d/wk]) were killed after 4, 8, 14, and 20 wk of treatment with 50 microg/kg of human parathyroid hormone (hPTH[1-84]). Bone mineral content and density were measured only in the vertebral body. Bone strength was evaluated in the vertebral body, femoral diaphysis, femoral neck, and distal femur. Significant, lasting osteopenia developed in the vertebral body of OVX rats by 10 wk postovariectomy. Bone mineral density of the vertebral body partially recovered by 8 wk and fully recovered to that seen in sham animals only by 20 wk posttreatment with either a 5 or 3 d/wk dosing schedule of PTH[1-84]. Therefore, hPTH[1-84] (50 microg/kg) given either 3 or 5 d/wk fully restores vertebral and femoral bone strength in osteopenic OVX rats.     

Lactam formation increases receptor binding, adenylyl cyclase stimulation and bone growth stimulation by human parathyroid hormone (hPTH)(1-28)NH2.

Human parathyroid hormone (1-28)NH2 [hPTH(1-28)NH2] is the smallest of the PTH fragments that can fully stimulate adenylyl cyclase in ROS 17/2 rat osteoblast-like osteosarcoma cells. This fragment has an IC50 of 110 nM for displacing 125I-[Nle8,18,Tyr34]bovine PTH(1-34)NH2 from HKRK B7 porcine kidney cells, which stably express 950,000 human type 1 PTH/PTH-related protein (PTHrP) receptors (PTH1Rs) per cell. It also has an EC50 of 23.9 nM for stimulating adenylyl cyclase in ROS 17/2 cells. Increasing the amphiphilicity of the alpha-helix in the residue 17-28 region by replacing Lys27 with Leu and stabilizing the helix by forming a lactam between Glu22 and Lys26 to produce the [Leu27]cyclo(Glu22-Lys26)hPTH(1-28)NH2 analog dramatically reduced the IC50 for displacing 125I-[Nle8,18,Tyr34]bPTH(1-34)NH2 from hPTH1Rs from 110 to 6 nM and dropped the EC50 for adenylyl cyclase stimulation in ROS 17/2 cells from 23.9 to 9.6 nM. These modifications also increased the osteogenic potency of hPTH(1-28)NH2. Thus, hPTH(1-28)NH2 did not significantly stimulate either femoral or vertebral trabecular bone growth in rats when injected daily at a dose of 5 nmol/100 g body weight for 6 weeks, beginning 2 weeks after ovariectomy (OVX), but it strongly stimulated the growth of trabeculae in the cancellous bone of the distal femurs and L5 vertebrae when injected at 25 nmol/100 g body weight. By contrast [Leu27]cyclo(Glu22-Lys26)hPTH(1-28)NH2 significantly stimulated trabecular bone growth when injected at 5 nmol/100 g of body weight. Thus, these modifications have brought the bone anabolic potency of hPTH(1-28)NH2 considerably closer to the potencies of the larger PTH peptides and analogs.     

BEMF对去卵巢骨质疏松大鼠骨组织bFGF表达的影响及意义

目的 观察BEMF对去卵巢骨质疏松大鼠骨组织bFGF蛋白表达的影响,探讨BEMF治疗骨质疏松的机理.方法 6月龄雌性未孕Wistar大鼠40只,按体质量随机分为模型组（OVX）、假手术组（Sham）、仿生脉冲电磁场治疗组（BEMF＋OVX,EM）、雌激素治疗组（Estrogen＋OVX,E）.OVX、EM、E组行双侧卵巢切除术,Sham组行假手术.术后第9 W开始治疗：E组行苯甲酸雌二醇肌肉注射,0.5mg/kg,1次/2 W.EM组大鼠暴露于仿生脉冲电磁场治疗,1 h/1次/d,OVX、Sham组不予以任何处理,作为对照组.治疗10 W后处死各组实验动物,测量腰椎骨密度、椎体的最大载荷、扫描电镜观察椎体骨结构的变化.采用免疫组化检测椎体骨组织中bFGF蛋白表达情况并以图像分析进行组间比较.结果 治疗10 W后EM组大鼠骨密度、椎体的最大载荷较OVX组显著性增高（P＜0.05）,骨结构明显改善.bFGF的表达主要位于成骨细胞、骨细胞,EM组椎体骨组织中bFGF表达显著性高于OVX组（P＜0.01）.结论 提高骨组织中bFGF的表达是BEMF治疗骨质疏松的重要机理之一.     

Maintenance of cortical bone in human parathyroid hormone(1-84)-treated ovariectomized rats

The purpose of this cross-sectional study was to evaluate the effects of human parathyroid hormone(1-84) (hPTH) followed by maintenance treatment with 17beta-estradiol (E(2)), risedronate (Ris), or a reduced dose of hPTH (LowPTH) on cortical bone in the ovariectomized (ovx) rat. Eight groups of ovx and one group of intact female rats (3.5 months) were left untreated for 11 weeks. For the following 12 weeks, four groups received subcutaneous injections of hPTH (75 microg/kg per day on 3 days/week) and four groups received vehicle. Treatments were then changed to E(2) (10 microg/kg per day on 2 days/week), Ris (3 microg/kg per day on 3 days/week), LowPTH (25 microg/kg per day on 3 days/week), or vehicle. Bone tissue was collected at weeks -11 (baseline), 0 (ovx effect), 12 (hPTH effect), 24, 36, and 48 (maintenance effect). Bone mineral density (BMD) and bone mineral content (BMC) of the diaphyseal femur and total cross-sectional area (Tt.Ar), marrow area (Ma.Ar), cortical area (Ct.Ar), and periosteal and endocortical bone formation of the tibia were measured. Ovariectomy resulted in lower BMD (weeks 0-48), unaffected BMC, and greater Tt.Ar (weeks 12 and 36), Ma.Ar (week 48), and Ct.Ar (weeks 0 and 12) compared with intact rats. Endocortical and periosteal bone formation were greater in the ovx than in the intact rats up to 23 weeks postovariectomy. Treatment of ovx rats with hPTH for 12 weeks resulted in greater cortical BMD, BMC, and endocortical bone formation than in intact or ovx controls. In ovx rats pretreated with hPTH and then treated with Ris for 36 weeks, BMD and BMC were greater and Ma.Ar was smaller than in ovx controls. In ovx rats pretreated with hPTH and then treated with LowPTH, BMD, BMC, Ct.Ar, and endocortical bone formation were greater and Ma.Ar was smaller than in ovx controls. Treatment of hPTH-pretreated rats with E(2) for 36 weeks did not affect cortical BMD, BMC, and Ct.Ar, although periosteal bone formation was lower in the E(2) group compared with the ovx group. Thus, in ovariectomized rats, cortical bone gained by 12 weeks of hPTH treatment was maintained for up to 36 weeks by treatment with risedronate or low-dose hPTH, but not with 17beta-estradiol.     

Maintenance of cancellous bone in ovariectomized, human parathyroid hormone [hPTH(1-84)]-treated rats by estrogen, risedronate, or reduced hPTH

This study compares effects of maintenance doses of human parathyroid hormone [hPTH(1-84)], 17beta-estradiol (E2), and risedronate on distal femur bone mineral density and proximal tibia cancellous bone histomorphometry in ovariectomized (ovx), osteopenic rats previously administered a higher dose of hPTH. Nine groups (n = 8) of 3.5-month-old ovx or intact Sprague-Dawley rats were left untreated for 11 weeks to allow for the development of cancellous osteopenia in the ovx groups. Next, the ovx rats received subcutaneous injections of hPTH (75 microg/kg per day, three times per week) or vehicle for 12 weeks. Treatments were then changed to E2 (10 microg/kg per day, two times per week), risedronate (Ris; 3 microg/kg per day, three times per week), low-dose hPTH(1-84) (LowPTH; 25 microg/kg per day, three times per week), or vehicle, and administered for 36 weeks. The intact control group remained untreated for the duration of study. Femora and tibiae were collected at weeks -11 (baseline); 0 (ovx effect); 12 (hPTH effect), and 24, 36, and 48 (maintenance effects). Endpoints evaluated included distal femur bone mineral density (BMD) and proximal tibia cancellous bone volume (BV/TV), osteoclast surface (Oc.S), mineralizing surface (MS), mineral apposition rate (MAR), and bone formation rate (BFR). Ovariectomy had a negative effect on distal femur BMD and proximal tibia BV/TV. Treatment of ovx rats with hPTH for 12 weeks resulted in higher BMD in comparison to intact controls, and higher cancellous BV/TV in comparison to ovx controls. Discontinuation of hPTH resulted in loss of gained BMD within 24 weeks and loss of gained BV/TV within 12 weeks. Treatment of ovx rats with hPTH for 12 weeks followed by E2 treatment left BMD and BV/TV similar to vehicle-treated ovx rats by week 48 (36 weeks after commencement of the E2 maintenance treatment). Maintenance treatment with risedronate resulted in BMD and BV/TV similar to that of intact controls. Maintenance treatment with low-dose hPTH resulted in greater BMD and similar BV/TV in comparison to intact controls. MS and BFR were highest after low-dose hPTH administration. MS and BFR were lowest after E2 or risedronate, whereas Oc.S was lowest after risedronate administration. Thus, in osteopenic rats, the increment in distal femur BMD and proximal tibia BV/TV gained by 12 weeks of hPTH treatment was lost within 24 and 12 weeks of treatment termination, respectively. Low-dose hPTH maintained BMD and BV/TV after hPTH treatment by stimulating bone formation, whereas risedronate maintained BMD and BV/TV by reducing bone resorption. E2 in a maintenance dose failed to maintain BMD and BV/TV after withdrawal of hPTH treatment.     

Both hPTH(1-34) and bFGF increase trabecular bone mass in osteopenic rats but they have different effects on trabecular bone architecture.

Osteoporosis is a syndrome of excessive skeletal fragility that results from both the loss of trabecular bone mass and trabecular bone connectivity. Recently, bFGF has been found to increase trabecular bone mass in osteoporotic rats. The purpose of this study was to compare how trabecular bone architecture, bone cell activity, and strength are altered by two different bone anabolic agents, bFGF and hPTH(1-34), in an osteopenic rat model. MATERIALS AND METHODS: Six-month-old female Sprague-Dawley rats (n = 74) were ovariectomized (OVX) or sham-operated (sham) and maintained untreated for 2 months. Then OVX rats were subcutaneously injected with basic fibroblast factor (bFGF; 1 mg/kg, 5 days/week), human parathyroid hormone [hPTH(1-34); 40 microg/kg, 5 days/week], or vehicle for 60 days (days 60-120). Sham-operated and one group of OVX animals were injected with vehicle. Biochemical markers of bone turnover (urinary deoxypyridinoline cross-links; Quidel Corp., San Diego, CA, USA) and serum osteocalcin (Biomedical Technologies, Stroughton, MA, USA) were obtained at study days 0, 60, 90, and 120 and analyzed by ELISA. At death, the right proximal tibial metaphysis was removed, and microcomputed tomography was performed for trabecular bone structure and processed for histomorphometry to assess bone cell activity. The left proximal tibia was used for nanoindentation/mechanical testing of individual trabeculae. The data were analyzed with Kruskal Wallis and post hoc testing as needed. RESULTS: Ovariectomy at day 60 resulted in about a 50% loss of trabecular bone volume compared with sham-treated animals. By day 120 post-OVX, OVX + vehicle treated animals had decreased trabecular bone volume, connectivity, number, and high bone turnover compared with sham-operated animals [p < 0.05 from sham-, hPTH(1-34)-, and bFGF-treated groups]. Treatment of OVX animals with bFGF and hPTH(1-34) both increased trabecular bone mass, but hPTH(1-34) increased trabecular thickness and bFGF increased trabecular number and connectivity. Histomorphometry revealed increased mineralizing surface and bone formation rate in both bFGF and hPTH(1-34) animals. However, osteoid volume was greater in bFGF-treated animals compared with both the hPTH(1-34) and OVX + vehicle animals (p < 0.05). Nanoindentation by atomic force microscope was performed on approximately 20 individual trabeculae per animal (three animals per group) and demonstrated that elastic modulus and hardness of the trabeculae in bFGF-treated animals were similar to that of the hPTH-treated and sham + vehicle-treated animals. CONCLUSION: Both hPTH(1-34) and bFGF are anabolic agents in the osteopenic female rat. However, hPTH(1-34) increases trabecular bone volume primarily by thickening existing trabeculae, whereas bFGF adds trabecular bone mass through increasing trabecular number and trabecular connectivity. These results suggest the possibility of sequential treatment paradigms for severe osteoporosis.     

The addition of a raloxifene analog (LY117018) allows for reduced PTH(1-34) dosing during reversal of osteopenia in ovariectomized rats.

To test the hypothesis that an antiresorptive agent might reduce the dosing requirement for an anabolic drug during reversal of osteopenia due to estrogen deficiency, the following experiment was conducted in 6-month-old female rats. Ovariectomy or sham surgery was performed and the following six experimental groups were studied. Untreated (SHAM) or ovariectomized (OVX) animals served as control groups. Four weeks post-OVX, osteopenic rats (now 7 months old), were treated in one of four experimental protocols: human parathyroid hormone (hPTH(1-34)), 80 microg/kg/day, given by subcutaneous injection 5 days/week; a selective estrogen receptor modulator (SERM), raloxifene analog LY117018 (RA), 3 mg/kg/day, given by gavage 5 days/week; and two combinations of LY117018 at the same dose and frequency with hPTH(1-34) (same dose, 5 times/week) and a reduced dosing interval of hPTH(1-34) (same dose, 2 times/week). After 12 weeks of treatment, the four experimental groups were sacrificed at age 10 months. SHAM and OVX controls were also studied at 7 and 10 months of age. Bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry at four skeletal sites: two mixed cortical/trabecular sites (femur and tibia) and two predominantly trabecular sites (lumbar spine and pelvis). The differences in BMD were consistent at all four sites. RA alone maintained BMD at all skeletal sites, but the results were not significantly improved over OVX controls, at age 10 months. hPTH(1-34) injections given 5 days/week resulted in BMD increments significantly higher than in either OVX or SHAM controls (p < 0.001). While the RA did not enhance the anabolic effects of full doses of hPTH(1-34), the addition of RA treatment to twice-weekly hPTH(1-34) dosing resulted in BMD increments at all four skeletal sites that were similar to the more intensive anabolic regimen of hPTH(1-34) therapy given 5 times/week. Therefore, an antiresorptive agent such as SERMs may potentially reduce the pharmacologic doses of PTH needed to reverse estrogen deficiency-induced osteopenia.