Developmental expression of genes for the stereoidogenic enzymes P450scc (20,22-desmolase), P450c17 (17 alpha-hydroxylase/17,20-lyase), and P450c21 (21-hydroxylase) in the human fetus

Fetal adrenal steroidogenesis is required for the production of placental estrogen, and fetal testicular steroidogenesis is required for the development of male external genitalia. We studied the ontogeny and tissue specificity of expression of the genes for three steroidogenic enzymes: P450scc (the cholesterol side-chain cleavage enzyme), P450c17 (17 alpha-hydroxylase/17,20-lyase), and P450c21 (21-hydroxylase) in the human fetus. RNA from fetal tissues was probed with homologous human P450scc, P450c17, and P450c21 cDNAs cloned in our laboratory. At 20-21 weeks gestation, P450scc mRNA was most abundant in the adrenal, followed by testis, placenta, and ovary. P450c17 mRNA was also most abundant in the adrenal, followed by testis and ovary, but was undetectable in the placenta. P450c21 mRNA was detected only in the adrenal. None of these mRNAs was detected in kidney, liver, spleen, intestine, or muscle. Twenty-two fetal testis samples (13-25.8 weeks gestation) were studied. P450scc and P450c17 mRNAs were most abundant at 14-16 weeks and diminished to 35 and 19% of their peak values, respectively, by 20-25.8 weeks. Ovarian P450scc and P450c17 mRNAs were present, respectively, in only 6.2% and 1.8% of the maximum amount in the testis and did not vary detectably from 14.9 to 21.5 weeks gestation. The testicular and ovarian steroidogenic enzyme mRNA data correlate well with previously reported changes in gonadal steroidogenesis with gestational age. The presence of P450scc mRNA, but not P450c17 mRNA, in the placenta indicates that the placenta is able to initiate the synthesis of some steroid hormones, but is not able to synthesize estrogen de novo. Since P450c21 was found only in the adrenal, the extraadrenal 21-hydroxylation of progesterone to deoxycorticosterone, a common event in the fetus, is probably mediated by an enzyme(s) other than P450c21.     

Combined 17-hydroxylase and 17,20-desmolase deficiencies: evidence for synthesis of a defective cytochrome P450c17

We studied in vivo and in vitro steroidogenesis in six phenotypic female children with 17-hydroxylase deficiency. The diagnosis was suspected as a likely cause of familial low renin hypertension and was confirmed by findings of reduced basal and ACTH-stimulated serum and urinary levels of cortisol and other 17-hydroxysteroids, together with hypergonadotropic hypogonadism in both 46,XY and 46,XX patients, and abnormally increased secretion of 17-desoxysteroids, such as progesterone, 11-deoxycorticosterone, and corticosterone. ACTH stimulation testing demonstrated a lesser degree of 17-hydroxylase deficiency in the obligate heterozygous parents; one father had increased basal serum 17-hydroxyprogesterone values, unresponsive to ACTH, suggesting partial Leydig cell 17,20-desmolase deficiency. In vitro kinetic analysis of testicular microsomal enzymes in the affected 46,XY male pseudohermaphrodites confirmed that both 17-hydroxylase and 17,20-desmolase activities were less than 2% of those in age-matched normal subjects. However, in spite of this virtual absence of both enzymatic activities of cytochrome P450c17, Northern blot analysis demonstrated abundant amounts of RNA in these tests that hybridized to a cDNA specific for this P450 enzyme. Moreover, immunoblot analysis of sodium dodecyl sulfate-polyacrylamide gel electrophoresis-resolved testicular microsomes showed an apparently normal content of an immunoreactive protein with a mol wt similar to that of authentic P450c17. These results suggest that these patients have a point mutation in the gene for P450c17; the mutant gene is transcribed, but gives rise to a protein defective in normal 17-hydroxylase and 17,20-desmolase activities.     

Assignment of the gene for adrenal P450c17 (steroid 17 alpha-hydroxylase/17,20 lyase) to human chromosome 10

P450c17 is the single enzyme mediating both 17 alpha-hydroxylase and 17,20 lyase activities. We identified several human P450c17 cDNA clones in a human adrenal cDNA library we constructed in lambda gt10. A short clone containing the 3'-terminal 650 bases of the full-length sequence was used to examine Southern blots of DNA from normal persons and from a panel of mouse/human somatic cell hybrid lines. The pattern of hybridization of this cDNA to normal human DNA cut with 8 restriction endonucleases suggests the human genome has two (or more) P450c17 genes. The pattern of hybridization to the somatic cell hybrid cell lines, each containing a limited, known number of human chromosomes, indicates the human adrenal P450c17 gene lies on chromosome 10. The chromosomal locations of the other P450c17 genes could not be determined.     

绝经后妇女膝骨关节炎与骨密度、骨代谢血清标志物的临床相关性

目的探讨绝经后妇女膝骨关节炎(KOA)与骨密度(BMD)、骨代谢血清标志物临床相关性。方法 180例绝经后妇女KOA患者分别进行体重、体重指数计算、绝经时间、腰1～4、股骨颈BMD测定及双膝关节X线拍片进行Kellgren-Lawrence(KL)分级,根据KOA KL分级比较BMD变化及酶联免疫吸附试验检测骨代谢血清标志物各项指标的变化。结果以KL结果分组,各组间绝经年龄无显著统计学差异(P>0.05),体重指数、年龄差异有统计学意义(P<0.05),随着KL级别增加,腰1～4和股骨颈BMD呈升高趋势,Ⅰ型胶原N端肽和Ⅰ膝胶原C端肽呈降低趋势。结论随着KL分级越高,BMD值越高,血清标志物水平越低。     

Fractures and bone mineral density in adult women with 21-hydroxylase deficiency

CONTEXT: Patients with classical congenital adrenal hyperplasia (CAH) receive lifelong, often supraphysiological, glucocorticoid therapy. Pharmacological doses of glucocorticoids are an established risk factor for osteoporosis. OBJECTIVE: Our objective was to evaluate bone mineral density (BMD), fracture prevalence, and markers of bone metabolism in adult females with CAH. DESIGN: This was a cross-sectional observational study. SETTING: Tertiary care referral centers were used in this study. PARTICIPANTS: We studied 61 women, aged 18-63 yr, with genetically verified CAH due to 21-hydroxylase deficiency. They were patients with salt wasting (n = 27), simple virilizing (n = 28), and nonclassical 21-hydroxylase deficiency (n = 6). A total of 61 age-matched women were controls. MAIN OUTCOME MEASURES: History of fractures was recorded. Total body, lumbar spine, and femoral neck BMD were measured by dual-energy x-ray absorptiometry. The World Health Organization criteria for osteopenia and osteoporosis were used. Serum marker of bone resorption, beta-C telopeptide was studied. RESULTS: The mean glucocorticoid dose in hydrocortisone equivalents was 16.9 +/- 0.9 mg/m2. Patients had lower BMD than controls at all measured sites (P < 0.001). In patients younger than 30 yr old, 48% were osteopenic vs. 12% in controls (P < 0.009). In patients 30 yr or older, 73% were osteopenic or osteoporotic vs. 21% in controls (P < 0.001). BMD was similar in the two classical forms and had no obvious relationship to genotypes. beta-C-telopeptide was decreased in older patients. More fractures were reported in patients than controls (P < 0.001). The number of vertebrae and wrist fractures almost reached significance (P = 0.058). CONCLUSIONS: Women with CAH have low BMD and increased fracture risk. BMD should be monitored, adequate prophylaxis and treatment instituted, and glucocorticoid doses optimized from puberty.     

Age-related changes in bone mineral density and serum bone-related proteins in premenopausal and postmenopausal Japanese women

The purpose of this cross-sectional study was to characterize the age-related change in bone metabolism during the pre- and postmenopausal periods, and to define the standard levels of three serum markers of bone metabolism, pyridinoline cross-linked carboxyterminal telopeptide of type I collagen (ICTP), carboxyterminal propeptide of type I procollagen (PICP), and bone gla protein (BGP), in Japanese adult women. The bone mineral density (BMD) of the lumbar spine (L2-L4) and the serum levels of ICTP, PICP and BGP were determined in a total of 207 healthy Japanese women (108 premenopausal and 99 postmenopausal). The lumbar BMD decreased significantly with increasing age not only in postmenopausal women (P<0.001) but also in premenopausal women (P=0.014). There was a clear gap in the serum levels of ICTP, PICP and BGP between the premenopausal and postmenopausal group (P<0.001), but those were absolutely the same within each group except for ICTP in the postmenopausal women. These findings and the values of serum ICTP, PICP and BGP in pre- and postmenopausal women obtained in this study are expected to be very useful for treatment of postmenopausal osteoporosis.     

绝经后妇女腰椎骨质疏松的定量CT研究

目的：研究绝经后妇女不同年龄组腰椎骨密度（ＢＭＤ）与骨质疏松关系及临床表现。方法采用ＣＴ机横断扫描及自制固体标准模测量９９例绝经后妇女第２－４腰椎上１／３处骨密度（ＢＭＤ）,并对其临床症状进行观察。结果９９例绝经后妇女有７６例被诊断为骨质疏松,占７７％,并显示腰椎骨密度（ＢＭＤ）随增龄逐渐减低,以大于７０岁年龄密度最低,骨质疏松最严重,平均值６０．２ｍｇ／ｃｍ＾３,方差为１８．９,而且１００％有不     

亚临床甲状腺功能减退症与绝经后女性骨代谢指标的相关性分析

目的探讨亚临床甲状腺功能减退症(SCH)与绝经后女性骨密度及骨代谢相关指标的关系。方法选取2015年1月至2018年6月在空军军医大学西京医院老年病科就诊的138例绝经后女性临床资料,根据患者是否患SCH分为SCH组(68例)和正常对照组(70例)。检测并比较2组患者骨密度相关指标[碱性磷酸酶(ALP)、Ca~(2+)、骨化三醇、骨密度T值(T -1.0为骨密度异常)]以及甲状腺功能相关指标[甲状旁腺激素(PTH)、促甲状腺激素(TSH)、游离三碘甲状腺原氨酸(FT3)、游离甲状腺素(FT4)、甲状腺过氧化物酶抗体(TPOAb~+)比例]。采用SPSS 18.0统计软件对数据进行分析。相关性采用Spearman相关分析。结果 SCH组及对照组患者骨密度异常率分别为50.0%(34/68)和25.7%(18/70),2组比较差异有统计学意义(P=0.003)。与对照组比较,SCH组患者TSH水平和TPOAb~+比例显著升高(P0.05),但FT3、FT4、PTH及骨代谢相关指标比较,差异无统计学意义(P0.05)。Spearman相关分析显示,TSH、TPOAb~+与ALP、骨化三醇、骨密度T值呈负相关,其中TSH与T值呈高度负相关(r=-0.804,P0.01)。结论 SCH可能引起绝经后女性骨量异常和骨密度测定值降低,这可能与血清TSH水平升高和TPOAb呈阳性有关。     

Effects of denosumab on bone mineral density and bone turnover in postmenopausal women

CONTEXT: Denosumab is an investigational fully human monoclonal antibody against receptor activator of nuclear factor-kappaB ligand, a mediator of osteoclastogenesis and osteoclast survival. OBJECTIVE: This study evaluated the ability of denosumab to increase bone mineral density (BMD) and decrease bone turnover markers (BTMs) in early and later postmenopausal women with low BMD. DESIGN AND SETTING: This 2-yr randomized, double-blind, placebo-controlled study was conducted in North America. PARTICIPANTS: Subjects included 332 postmenopausal women with lumbar spine BMD T-scores between -1.0 and -2.5. Interventions: SUBJECTS were randomly assigned to receive denosumab sc, 60 mg every 6 months, or placebo. Randomization was stratified by time since onset of menopause (< or =5 yr or > 5 yr). MAIN OUTCOME MEASURES: The primary end point was the percent change in lumbar spine BMD by dual-energy x-ray absorptiometry at 24 months. Additional end points were percent change in volumetric BMD of the distal radius by quantitative computed tomography; percent change in BMD by dual-energy x-ray absorptiometry for the total hip, one-third radius, and total body; hip structural analysis; percent change in BTMs; and safety. RESULTS: Denosumab significantly increased lumbar spine BMD, compared with placebo at 24 months (6.5 vs. -0.6%; P<0.0001) with similar results for both strata. Denosumab also produced significant increases in BMD at the total hip, one-third radius, and total body (P < 0.0001 vs. placebo); increased distal radius volumetric BMD (P < 0.01); improved hip structural analysis parameters; and significantly suppressed serum C-telopeptide, tartrate-resistant acid phosphatase-5b, and intact N-terminal propeptide of type 1 procollagen. The overall incidence of adverse events was similar between both study groups. CONCLUSIONS: Twice-yearly denosumab increased BMD and decreased BTMs in early and later postmenopausal women.     

Association of serum sex steroid receptor bioactivity and sex steroid hormones with breast cancer risk in postmenopausal women

Postmenopausal women with elevated serum sex steroids have an increased risk of breast cancer. Most of this risk is believed to be exerted through binding of the sex steroids to their receptors. For the first time, we investigate the association of estrogen receptor (ER) and androgen receptor (AR) serum bioactivity (SB) in addition to hormone levels in samples from women with breast cancer collected before diagnosis. Two hundred postmenopausal women participating in the UK Collaborative Trial of Ovarian Cancer Screening who developed ER-positive breast cancer 0.6-5 years after sample donation were identified and matched to 400 controls. ER and AR bioassays were used to measure ERα, ERβ, and AR SB. Androgen and estrogen levels were measured with immunoassays. Subjects were classified according to quintiles of the respective marker among controls and the associations between SB and hormones with breast cancer risk were determined by logistic regression analysis. ERα and ERβ SB were significantly higher before diagnosis compared with controls, while estrogens showed no difference. Women had a twofold increased breast cancer risk if ERα SB (odds ratio (OR), 2.114; 95% confidence interval (CI), 1.050-4.425; P=0.040) was in the top quintile >2 years before diagnosis or estrone (OR, 2.205; 95% CI, 1.104-4.586; P=0.029) was in the top quintile <2 years before diagnosis. AR showed no significant association with breast cancer while androstenedione (OR, 3.187; 95% CI, 1.738-6.044; P=0.0003) and testosterone (OR, 2.145; 95% CI, 1.256-3.712; P=0.006) were significantly higher compared with controls and showed a strong association with an almost threefold increased breast cancer risk independent of time to diagnosis. This study provides further evidence on the association of androgens and estrogens with breast cancer. In addition, it reports that high ER but not AR SB is associated with increased breast risk >2 years before diagnosis.     

Growth hormone and sex steroid effects on bone metabolism and bone mineral density in healthy aged women and men.

BACKGROUND: Aging is associated with concomitant declines in activity of the growth hormone (GH) and gonadal steroid axes, and in bone mineral density (BMD), in both sexes. Long-term estrogen replacement slows bone loss and prevents fractures in postmenopausal women, whereas the effects of supplementation of GH or testosterone on bone metabolism and BMD in aged individuals remains uncertain. METHODS: Using a randomized, placebo-controlled, double-blind study design, we investigated the separate and interactive effects of 6 months of administration of recombinant human GH and/or gonadal steroids on bone biochemical markers and BMD in 125 healthy, older (>65 years) women (n = 53) and men (n = 72) with age-related reductions in GH and gonadal steroids. RESULTS: In women, administration of GH, but not GH + hormone replacement therapy (HRT), increased serum levels of osteocalcin and procollagen peptide (PICP) and increased urinary excretion of deoxypyridinoline (DPD) crosslinks. Urinary calcium excretion decreased after HRT. In men, GH, and to a greater extent GH + T, increased osteocalcin. GH increased serum PICP, and GH + T increased urinary DPD. Urinary calcium excretion was unaffected by hormone treatment in men. In women, administration of HRT and GH + HRT, but not GH, increased BMD at the lumbar spine, femoral neck, and distal radius. In men, GH + T led to a small decrease in BMD at the proximal radius; there were no other significant effects of hormone administration on BMD. CONCLUSIONS: These data suggest that short-term administration of HRT exerts beneficial effects on bone metabolism and BMD in postmenopausal women, which are not significantly altered by the coadministration of GH. In andropausal men, T administration to achieve physiologic levels did not result in significant effects on bone metabolism or BMD, whereas GH + T increased one marker of bone formation and decreased one marker of bone resorption. Given the known biphasic actions of GH on bone and the apparent favorable biochemical effects of GH + T in men, the longer-term effects of GH + T on BMD in aged men remain to be clarified.     

Cytochrome P450c17 (steroid 17 alpha-hydroxylase/17,20 lyase): cloning of human adrenal and testis cDNAs indicates the same gene is expressed in both tissues.

P450c17 is the single enzyme mediating both 17 alpha-hydroxylase (steroid 17 alpha-monooxygenase, EC 1.14.99.9) and 17,20 lyase activities in the synthesis of steroid hormones. It has been suggested that different P450c17 isozymes mediate these activities in the adrenal gland and testis. We sequenced 423 of the 509 amino acids (83%) of the porcine adrenal enzyme; based on this partial sequence, a 128-fold degenerate 17-mer was synthesized and used to screen a porcine adrenal cDNA library. This yielded a 380-base cloned cDNA, which in turn was used to isolate several human adrenal cDNAs. The longest of these, lambda hac17-2, is 1754 base pairs long and includes the full-length coding region, the complete 3'-untranslated region, and 41 bases of the 5'-untranslated region. This cDNA encodes a protein of 508 amino acids having a predicted molecular weight of 57,379.82. High-stringency screening of a human testicular cDNA library yielded a partial clone containing 1303 identical bases. RNA gel blots and nuclease S1-protection experiments confirm that the adrenal and testicular P450c17 mRNAs are indistinguishable. These data indicate that the testis possesses a P450c17 identical to that in the adrenal. The human amino acid sequence is 66.7% homologous to the corresponding regions of the porcine sequence, and the human cDNA and amino acid sequences are 80.1 and 70.3% homologous, respectively, to bovine adrenal P450c17 cDNA. Both comparisons indicate that a central region comprising amino acid residues 160-268 is hypervariable among these species of P450c17. Comparison of the amino acid sequence of P450c17 with two other human steroidogenic cytochromes P450 show much greater homology with P450c21 (28.9%), another microsomal enzyme, than with P450scc (12.3%), a mitochondrial enzyme.     

Hormonal regulation of messenger ribonucleic acids for P450scc (cholesterol side-chain cleavage enzyme) and P450c17 (17 alpha-hydroxylase/17,20-lyase) in cultured human fetal adrenal cells

ACTH has acute and long term effects on adrenal steroidogenesis by week 14 of fetal life. We used human fetal adrenal cells to investigate the long term effect of physiological doses of ACTH on mRNAs for P450scc (the cholesterol side-chain cleavage enzyme) and P450c17 (17 alpha-hydroxylase/17,20-lyase). Monolayer cultures of 18- to 24-week gestation fetal zone adrenal cells were maintained in the presence and absence of 10(-9) or 10(-8) M ACTH for up to 12 days. As assessed by RNA dot blots probed with cloned homologous human cDNAs, ACTH increased P450scc and P450c17 mRNAs 4- and 9-fold, respectively, over control values on day 7 of culture. ACTH-mediated stimulation was slightly less on day 12 of culture. The ACTH-mediated accumulation of those mRNAs were time dependent. When cells were exposed to a single 10(-8)-M dose of ACTH, the amount of P450scc and P450c17 mRNA was increased by 24 h, reaching a maximum at 48 h and diminishing by 72 h. When cells were maintained in 10(-8) M ACTH continuously, mRNA for both enzymes accumulated in a similar pattern, reaching a peak at 48 h but remaining at nearly maximal values thereafter, up to 96 h. Dibutyryl cAMP (10(-3) M) mimicked these stimulatory actions of ACTH, although its effect was greater at 24 h and more stable up to 96 h. Angiotensin II (1-100 ng/mL) and hCG (1-100 ng/mL) had no effect on accumulation of P450scc and P450c17 mRNAs. The production of both dehydroepiandrosterone sulfate and cortisol also was stimulated by ACTH, suggesting that the increased mRNAs were translated into active enzymes. These results indicate that ACTH induces human fetal adrenal cells to accumulate mRNAs for both P450scc and P450c17; this effect of ACTH is probably mediated by cAMP. Chronic 96-h stimulation of human fetal adrenal cells did not diminish their responsiveness to ACTH. Together with our earlier studies of the human fetal adrenal, these data indicate that fetal adrenal tissue does not exhibit the desensitization to trophic hormone stimulation characteristic of adult tissue.     

Skeletal and mandibular bone mineral density in dentate and edentulous postmenopausal women

Tooth loss is one of risk factors for osteoporosis. Reversibly osteoporotic condition is one of putative risk factors for tooth loss. The balance of bone remodeling in jawbone shift to absorption in systemically osteoporotic individuals and result in enhancement of tooth loss. The presence of tooth and skeletal and mandibular bone metabolism may relate to one another. We investigated the relationship between the presence of the tooth and skeletal and mandibular bone mineral density (BMD) in postmenopausal women. Periodontally healthy and edentulous subjects participated in this study. BMD of the lumbar spine and mandibular bone was determined. The results suggest that the presence of tooth contribute to maintain the BMD of mandibular cortical bone in postmenopausal women. Direct masticatory forces via natural teeth may influence the mandibular cortical bone metabolism and if tooth loss occurs, the bone metabolism may be regulated similar to other skeletal bone.     

绝经后妇女脊椎压缩性骨折与骨密度的关系

目的探讨绝经后妇女脊椎压缩性骨折与骨密度（BMD）的关系。方法为病例一对照研究，入选250例有脊椎压缩性骨折的绝经后妇女，另有250名无脊椎压缩性骨折的绝经后妇女作为对照组。两组均有胸腰椎正侧位X线摄片，并应用双能X线吸收仪检测腰椎1～4和左股骨近端各部位BMD。结果脊椎压缩性骨折组身高、体重、腰椎2～4和股骨近端各部位BMD值均显著低于对照组（均P〈0．01）。腰椎2～4BMD是发生脊柱骨折的预报因子（r=-0．416，P〈0．01）。身高和全髋部BMD与骨折次数和骨折椎体数目呈负相关（均P〈0．01）。按股骨颈和全髋部BMD值，骨折组骨质疏松检出率各为50．8％和50．4％；另外剔除在腰椎2～4发生椎体骨折53例，按腰椎2～4BMD检出骨质疏松占64．5％。同时，腰椎2～4、股骨颈或全髋部BMD值低于-2．5s者发生脊柱压缩性骨折的风险分别是BMD正常者的4．76、2．36和3．52倍。结论腰椎呈低骨量是发生脊椎压缩性骨折的重要危险因素。身高的下降和全髋部低BMD值是骨折发生次数和受累椎体数目的危险因子；对绝经后妇女在重视BMD测量的同时，应重视脊柱X线正侧位检查。     

Serum uric acid level and bone mineral density in Iraqi postmenopausal women

BackgroundThe role of uric acid (UA) in bone mineral density (BMD) has been investigated with diverse results.Aim of the workTo study the relation between serum UA and BMD in Iraqi postmenapausal women.Patients and methodsThe study involved 151 Iraqi postmenapausal women recruited from Baghdad Medical City. Serum UA was measured on two occasions and subjects were categorized into four quartiles according to the serum concentrations. BMD was measured by dual energy x-ray absorptiometry (DXA) and T-score calculated at the lumbar spine (L1-L4) and right femoral neck.ResultsThe mean age of the subjects was 53 ± 9.1 years, body mass index was 31 ± 3.68 and the menopause duration was 8.13 ± 5.86 years. Their mean serum UA level was 4.72 ± 1.35 mg/dl. 56 (37.1%) subjects were osteopenic and 34 (22.5%) were osteoporotic. The mean BMD increased significantly across the quartiles; the highest was in the fourth UA quartile (highest) for both the lumbar spine and right femoral neck. The percentage of women with osteoporosis and osteopenia were lowest in the fourth UA quartile. UA was significantly associated with BMD at L1-L4 spine (p = 0.04) and right femoral neck (p = 0.004) and with the corresponding T-scores (p = 0.008 and p = 0.01 respectively). After adjusting for confounding factors for UA on BMD, only the association of UA with L1-L4 BMD (β = 0.03, p = 0.01) and T-value (β = 0.32, p = 0.009) was still significant.ConclusionHigher serum UA levels were associated with higher BMDs at the lumbar spine suggesting that it may have a beneficial effect on the bone density.