Spinal Nitric Oxide Synthase Type II Increases Neurosteroid-metabolizing Cytochrome P450c17 Expression in a Rodent Model of Neuropathic Pain

We have previously demonstrated that the neurosteroid dehydroepiandrosterone sulfate (DHEAS) induces functional potentiation of N-methyl-D-aspartate (NMDA) receptors via increases in phosphorylation of NMDA receptor GluN1 subunit (pGluN1). However, the modulatory mechanisms responsible for the expression of the DHEA-synthesizing enzyme, cytochrome P450c17 following peripheral nerve injury have yet to be examined. Here we determined whether oxidative stress induced by the spinal activation of nitric oxide synthase type II (NOS-II) modulates the expression of P450c17 and whether this process contributes to the development of neuropathic pain in rats. Chronic constriction injury (CCI) of the sciatic nerve induced a significant increase in the expression of NOS-II in microglial cells and NO levels in the lumbar spinal cord dorsal horn at postoperative day 5. Intrathecal administration of the NOS-II inhibitor, L-NIL during the induction phase of neuropathic pain (postoperative days 0~5) significantly reduced the CCI-induced development of mechanical allodynia and thermal hyperalgesia. Sciatic nerve injury increased the expression of PKC- and PKA-dependent pGluN1 as well as the mRNA and protein levels of P450c17 in the spinal cord at postoperative day 5, and these increases were suppressed by repeated administration of L-NIL. Co-administration of DHEAS together with L-NIL restored the development of neuropathic pain and pGluN1 that were originally inhibited by L-NIL administration alone. Collectively these results provide strong support for the hypothesis that activation of NOS-II increases the mRNA and protein levels of P450c17 in the spinal cord, ultimately leading to the development of central sensitization and neuropathic pain induced by peripheral nerve injury.

Effect of Wi-Fi microwave irradiation on testosterone synthesis in Leydig cells of male workers / 中国职业医学

OBJECTIVE: To explore the effect of occupational wireless fidelity(Wi-Fi) microwave radiation on testosterone synthesis in male workers. METHODS: A total of 51 male workers exposed to microwave radiation in Wi-Fi test station of a mobile phone manufacturer were selected as exposure group by judgment sampling method. They were divided into <2.0 years subgroup and ≥2.0 years subgroup according to the length of work years. At the same time, 30 male workers who were not exposed to occupational hazards in the same factory were selected as the control group. Serum total cholesterol level was detected by colorimetry. Serum testosterone, cyclic adenosine monophosphate(cAMP), cytochrome P450 17 A1(P450 c17), cytochrome P450 cholesterol side chain cleavage enzyme(P450 scc), levels were detected by enzyme-linked immunosorbent assay. The relative expression of P450 scc and P450 c17 mRNA in whole blood was measured by real-time quantitative polymerase chain reaction. RESULTS: The levels of serum testosterone, P450 c17 and the relative expression of P450 c17 mRNA in workers of the exposure group were lower than that in the control group(P≤0.05), and the above indexes in the sub-exposure group with work age ≥2.0 years was lower than that in the control group(P<0.05). There was no significant difference in levels of serum total cholesterol, cAMP, P450 scc and relative expression of P450 scc in whole blood among the exposed group,two subgroups and the control group(P>0.05). CONCLUSION: Long-term exposure to Wi-Fi microwave radiation can inhibit the expression of P450 c17 mRNA and the synthesis of P450 c17 protein, both are key enzymes for testosterone synthesis in male workers, thereby affecting the synthesis and secretion of testosterone.

Identification of steroid biosynthetic defects in genotype-proven heterozygote individuals with 17α-hydroxylase/17,20-lyase deficiency / 中华内分泌代谢杂志

Objective To investigate the adrenal steroidogenic function in genotype-proven heterozygotes carrying mutations in CYP17A1 gene in vivo. Methods Eight patients and 14 family members from 5 families with 17-hydroxylase/17,20-lyase deficiency (17OHD) were recruited. The mutations of the CYP17A1 gene in these individuals were screened by direct sequencing of PCR products. The hormonal response to ACTH was evaluated in the 14 genotype-proven carriers and 45 age- and sex-matched normal subjects. Results Three mutations were found in 5 unrelated families. 14 carriers with CYP17A1 mutation were identified, including 7 heterozygotes with D487_F489del, 6 with Y329fs, and 1 for H373L. Compared to the normal subjects, the carriers exhibited lower basal and ACTH-stimulated cortisol levels, but higher ACTH-stimulated corticosterone level. The ratios of corticosterone to cortisol in the genotype-proven heterozygotes were higher than those of normal individuals at baseline and following ACTH-stimulation. Similarly, progesterone level and ratios of progesterone to 17-hydroxyprogesterone in the male heterozygotes were also higher than that of normal individuals before and after stimulation. No significant differences were observed in the hormone levels between two genotypes (D487_F489del vs Y329fs). Conclusions Genotype-proven carriers of 17OHD without apparent clinical symptoms exhibit decreased enzyme activity,analogous to mildly impaired adrenal 21-hydroxylase activity in the carriers of CYP21 A2 gene mutation.

Combined 17 alpha-hydroxylase/17, 20-lyase deficiency due to a homozygous 25 BP duplication (NT 4157-4181) at exon 5 in the CYP17 resulting in a premature stop codon predicted by molecular modeling / Deficiência combinada de 17 alfa-hidroxilase/17, 20 liase devido a duplicação de 25 PB (NT 4157-4181) no éxon 5 do CYP17 resultando em parada prematura de leitura predita por modelagem molecular

Combined 17alpha-hydroxylase/17,20-lyase deficiency is a rare, autosomal recessive form of congenital adrenal hyperplasia characterized by the coexistence of hypertension, caused by the hyperproduction of mineralocorticoid precursors and DSD in males and sexual infantilism in females, due to impaired production of sex hormones. Several CYP17 mutations resulting in 17alpha-hydroxylase/17,20-lyase deficiency have been reported previously. In the present study, we described a novel CYP17 mutation in two Brazilian sisters with primary amenorrhea, 46,XY karyotype, high basal levels of progesterone (3.4-4.9 ng/mL) and hypokalemic hypertension born to consanguineous parents. After PCR and automatic sequencing of CYP17 coding region, 25 bp duplication at exon 5 was found in the patients. This duplication started at codon 318 resulting in a premature stop codon at position 320 resulting in an ineffective and truncated protein and in accordance with the molecular modeling of P450c17. Therefore we expanded the repertoire of CYP17 mutations describing the largest duplication found in this gene in both sisters, with a clinical phenotype of combined 17alpha-hydroxylase/17,20-lyase deficiency and emphasizes the importance of the P450c 17 molecular modeling to predict the functional effect of these mutations.

A deficiência combinada de 17 alfa-hidroxilase/17,20 liase é uma doença rara, de herança autossômica recessiva, causa de hiperplasia adrenal congênita caracterizada pela presença de hipertensão resultante do acúmulo de precursores mineralocorticóides, distúrbio da diferenciação sexual em homens e infantilismo sexual em mulheres devido à falha na produção de esteróides sexuais. Várias mutações no gene CYP17 resultando em deficiência de 17 alfa-hidroxilase/17,20-liase têm sido descritas. No presente estudo, descrevemos uma nova mutação no CYP17 em duas irmãs, nascidas de pais consangüíneos, com quadro de amenorréia primária, cariótipo 46,XY, dosagens basais elevadas de progesterona (3,4-4,9 ng/mL) e hipertensão hipocalêmica. Após PCR e seqüenciamento automático da região codificadora do CYP17, uma duplicação de 25 pb no exon 5 foi identificada nas pacientes. Esta duplicação inicia-se no códon 318 resultando em parada prematura de leitura no códon 320 gerando uma proteína truncada e inativa conforme predito pela modelagem molecular do P450c17. Com este achado, ampliamos o repertório de mutações do CYP17 descrevendo a maior duplicação descrita até então neste gene em duas irmãs com fenótipo de deficiência combinada de 17 alfa-hidroxilase/17,20-liase e enfatizamos a importância da modelagem molecular do P450c 17 em predizer o efeito funcional destas mutações.

A Case of 17a-Hydroxylase Deficiency in 17-Year-Old Girl / 대한내분비학회지

The single enzyme P-450c17 hydroxylase catalyzes the 17a-hydroxylation of both pregnenolone and progesterone and the side-chain cleavage of 17a-hydroxypregnenolone and 17a-hydroxypro- gesterone to dehydroepiandrosterone and androstenedione. This enzyme is located in the endoplasmic reticulum and consists of a P-450c17 and a specific flavoprotein NADPH-cytochrome P-450 reductase. The clinical picture and hormonal pattern in 17a-hydroxylase deficiency have been consistent in both genotypic sexes with hypergonadotropic hypogonadism in whom the virtual absence of gonadal steroids results in a female phenotype with primary amenorrhea and pseudohermaphro- ditism in the male and underdeveloped secondary sex characteristics and hypermineralocorticoidism with hypertension, hypokalemia, suppressed renin-angiotensin system and extremely reduced aldo-sterone production. A 17-year-old girl visited endocrine clinic because of amenorrhea, absence of pubic and axillary hair, and hypertension. she had elevated levels of serum corticosterone, deoxycorticosterone(DOC), 18-hydroxycorticosterone(18-OHB). Stumulation with ACTH effected minimal increase in the elevated steroids and the ACTH-stimulated 18-OHB to aldosterone ratio was more than 280. These hormonal patterns appear to be homozygote in 17a-hydroxylase deficiency.

THE REGULATORY EFFECT OF hCG ON STEROIDOGENIC ENZYMES IN ADULT MOUSE LEYDIG CELLS / 解剖学报

Objective To study the regulation role on molecular level of hCG on P450scc,P450c17 and 3?HSD in adult mouse Leydig cells. Methods Leydig cells of adult mice were cultured in vitro for 1 and 8 hours with or without hCG,then P450scc,P450c17 and I,VI total 3?HSD mRNA levels were measured respectively by the semi quantitative RT PCR as well as Scal restriction endonuclease incised methods.Meanwhile,the testosterone contents were measured in two groups. Results 1.In stimulated group Leydig cells cultured for 1 and 8 hours with hCG,the testosterone contents were higher significantly than those in control groups(1 hour P 0.05),but the ratio of type VI/I 3?HSD mRNA gradually increased. Conclusion hCG can stimulate the transcription of P450scc,P450c17 and type VI 3?HSD in adult mouse Leydig cells.