Diversity of Androgens; Comparison of Their Significance and Characteristics in Vertebrate Species.

Androgen(s) is one of the sex steroids that are involved in many physiological phenomena of vertebrate species. Although androgens were originally identified as male sex hormones, it is well known now that they are also essential in females. As in the case of other steroid hormones, androgen is produced from cholesterol through serial enzymatic reactions. Although testis is a major tissue to produce androgens in all species, androgens are also produced in ovary and adrenal (interrenal tissue). Testosterone is the most common and famous androgen. It represents a major androgen both in males and females of almost vertebrate species. In addition, testosterone is a precursor for producing significant androgens such as11-ketotestosterone, 5α-dihydrotestosterone, 11-ketodihydrotestosterones and 15α-hydroxytestosterone in a species- or sex-dependent manner for their homeostasis. In this article, we will review the significance and characteristics of these androgens, following a description of the history of testosterone discovery and its synthetic pathways.

Comparison of Placental HSD17B1 Expression and Its Regulation in Various Mammalian Species.

During mammalian gestation, large amounts of progesterone are produced by the placenta and circulate for the maintenance of pregnancy. In contrast, primary plasma estrogens are different between species. To account for this difference, we compared the expression of ovarian and placental steroidogenic genes in various mammalian species (mouse, guinea pig, porcine, ovine, bovine, and human). Consistent with the ability to synthesize progesterone, CYP11A1/Cyp11a1, and bi-functional HSD3B/Hsd3b genes were expressed in all species. CYP17A1/Cyp17a1 was expressed in the placenta of all species, excluding humans. CYP19A/Cyp19a1 was expressed in all placental estrogen-producing species, whereas estradiol-producing HSD17B1 was only strongly expressed in the human placenta. The promoter region of HSD17B1 in various species possesses a well-conserved SP1 site that was activated in human placental cell line JEG-3 cells. However, DNA methylation analyses in the ovine placenta showed that the SP1-site in the promoter region of HSD17B1 was completely methylated. These results indicate that epigenetic regulation of HSD17B1 expression is important for species-specific placental sex steroid production. Because human HSD17B1 showed strong activity for the conversion of androstenedione into testosterone, similar to HSD17B1/Hsd17b1 in other species, we also discuss the biological significance of human placental HSD17B1 based on the symptoms of aromatase-deficient patients.

Expression of Chrna9 is regulated by Tbx3 in undifferentiated pluripotent stem cells.

It was reported that nicotinic acetylcholine receptor (nAChR)-mediated signaling pathways affect the proliferation and differentiation of pluripotent stem cells. However, detail expression profiles of nAChR genes were unrevealed in these cells. In this study, we comprehensively investigated the gene expression of α subunit of nAChRs (Chrna) during differentiation and induction of pluripotent stem cells. Mouse embryonic stem (ES) cells expressed multiple Chrna genes (Chrna3-5, 7 and 9) in undifferentiated status. Among them, Chrna9 was markedly down-regulated upon the differentiation into mesenchymal cell lineage. In mouse tissues and cells, Chrna9 was mainly expressed in testes, ES cells and embryonal F9 teratocarcinoma stem cells. Expression of Chrna9 gene was acutely reduced during differentiation of ES and F9 cells within 24 h. In contrast, Chrna9 expression was increased in induced pluripotent stem cells established from mouse embryonic fibroblast. It was shown by the reporter assays that T element-like sequence in the promoter region of Chrna9 gene is important for its activities in ES cells. Chrna9 was markedly reduced by siRNA-mediated knockdown of Tbx3, a pluripotency-related transcription factor of the T-box gene family. These results indicate that Chrna9 is a nAChR gene that are transcriptionally regulated by Tbx3 in undifferentiated pluripotent cells.

Purification, characterization and cDNA cloning of a lectin from the brittle star Ophioplocus japonicus.

Lectins are carbohydrate-binding proteins that possess specific sugar-binding properties and are involved in various biological activities in different organisms. In this study, purification, characterization, and cDNA cloning of a brittle star lectin, designated as Ophioplocus japonicus agglutinin (OJA), were conducted. OJA was isolated from the brittle star O. japonicus by affinity chromatography on a Sephadex G-25 column, followed by ion-exchange chromatography on a Resource Q column. This lectin yielded distinct bands at approximately 176 or 17 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) under non-reducing or reducing conditions, respectively. It also exhibited Ca2+-dependent hemagglutination activity, which, however, was not affected by other metal cations, such as Ba2+, Co2+, Cu2+, Zn2+, Fe2+, Mg2+, and Mn2+. The OJA activity was strongly inhibited by glucose and xylose among the monosaccharides tested, and by bovine thyroglobulin among the glycoproteins tested. Cloning of the OJA cDNA revealed that its primary structure contained the C-type lectin domain (CTLD). The results of this study showed that OJA is an echinoderm-derived glucose/xylose-specific lectin that belongs to the C-type lectin superfamily.

Facilitation of colonic T cell immune responses is associated with an exacerbation of dextran sodium sulfate-induced colitis in mice lacking microsomal prostaglandin E synthase-1.

BACKGROUND: Microsomal prostaglandin E synthase-1 (mPGES-1) is a key enzyme that acts downstream of cyclooxygenase and plays a major role in inflammation by converting prostaglandin (PG) H2 to PGE2. The present study investigated the effect of genetic deletion of mPGES-1 on the development of immunologic responses to experimental colitis induced by dextran sodium sulfate (DSS), a well-established model of inflammatory bowel disease (IBD). METHODS: Colitis was induced in mice lacking mPGES-1 (mPGES-1-/- mice) and wild-type (WT) mice by administering DSS for 7 days. Colitis was assessed by body weight loss, diarrhea, fecal bleeding, and histological features. The colonic expression of mPGES-1 was determined by real-time PCR, western blotting, and immunohistochemistry. The impact of mPGES-1 deficiency on T cell immunity was determined by flow cytometry and T cell depletion in vivo. RESULTS: After administration of DSS, mPGES-1-/- mice exhibited more severe weight loss, diarrhea, and fecal bleeding than WT mice. Histological analysis further showed significant exacerbation of colonic inflammation in mPGES-1-/- mice. In WT mice, the colonic expression of mPGES-1 was highly induced on both mRNA and protein levels and colonic PGE2 increased significantly after DSS administration. Additionally, mPGES-1 protein was localized in the colonic mucosal epithelium and infiltrated inflammatory cells in underlying connective tissues and the lamina propria. The abnormalities consistent with colitis in mPGES-1-/- mice were associated with higher expression of colonic T-helper (Th)17 and Th1 cytokines, including interleukin 17A and interferon-γ. Furthermore, lack of mPGES-1 increased the numbers of Th17 and Th1 cells in the lamina propria mononuclear cells within the colon, even though the number of suppressive regulatory T cells also increased. CD4+ T cell depletion effectively reduced symptoms of colitis as well as colonic expression of Th17 and Th1 cytokines in mPGES-1-/- mice, suggesting the requirement of CD4+ T cells in the exacerbation of DSS-induced colitis under mPGES-1 deficiency. CONCLUSIONS: These results demonstrate that mPGES-1 is the main enzyme responsible for colonic PGE2 production and deficiency of mPGES-1 facilitates the development of colitis by affecting the development of colonic T cell-mediated immunity. mPGES-1 might therefore impact both the intestinal inflammation and T cell-mediated immunity associated with IBD.

Analyses of Molecular Characteristics and Enzymatic Activities of Ovine HSD17B3.

17ß-hydroxysteroid dehydrogenase type 3 (HSD17B3) converts androstenedione (A4) into testosterone (T), which regulates sex steroid production. Because various mutations of the HSD17B3 gene cause disorder of sex differentiation (DSD) in multiple mammalian species, it is very important to reveal the molecular characteristics of this gene in various species. Here, we revealed the open reading frame of the ovine HSD17B3 gene. Enzymatic activities of ovine HSD17B3 and HSD17B1 for converting A4 to T were detected using ovine androgen receptor-mediated transactivation in reporter assays. Although HSD17B3 also converted estrone to estradiol, this activity was much weaker than those of HSD17B1. Although ovine HSD17B3 has an amino acid sequence that is conserved compared with other mammalian species, it possesses two amino acid substitutions that are consistent with the reported variants of human HSD17B3. Substitutions of these amino acids in ovine HSD17B3 for those in human did not affect the enzymatic activities. However, enzymatic activities declined upon missense mutations of the HSD17B3 gene associated with 46,XY DSD, affecting amino acids that are conserved between these two species. The present study provides basic information and tools to investigate the molecular mechanisms behind DSD not only in ovine, but also in various mammalian species.

Profiles of 5α-Reduced Androgens in Humans and Eels: 5α-Dihydrotestosterone and 11-Ketodihydrotestosterone Are Active Androgens Produced in Eel Gonads.

Although 11-ketotestosterone (11KT) and testosterone (T) are major androgens in both teleosts and humans, their 5α-reduced derivatives produced by steroid 5α-reductase (SRD5A/srd5a), i.e., 11-ketodihydrotestosterone (11KDHT) and 5α-dihydrotestosterone (DHT), remains poorly characterized, especially in teleosts. In this study, we compared the presence and production of DHT and 11KDHT in Japanese eels and humans. Plasma 11KT concentrations were similar in both male and female eels, whereas T levels were much higher in females. In accordance with the levels of their precursors, 11KDHT levels did not show sexual dimorphism, whereas DHT levels were much higher in females. It is noteworthy that plasma DHT levels in female eels were higher than those in men. In addition, plasma 11KDHT was undetectable in both sexes in humans, despite the presence of 11KT. Three srd5a genes (srd5a1, srd5a2a and srd5a2b) were cloned from eel gonads. All three srd5a genes were expressed in the ovary, whereas only both srd5a2 genes were expressed in the testis. Human SRD5A1 was expressed in testis, ovary and adrenal, whereas SRD5A2 was expressed only in testis. Human SRD5A1, SRD5A2 and both eel srd5a2 isoforms catalyzed the conversion of T and 11KT into DHT and 11KDHT, respectively, whereas only eel srd5a1 converted T into DHT. DHT and 11KDHT activated eel androgen receptor (ar)α-mediated transactivation as similar fashion to T and 11KT. In contrast, human AR and eel arß were activated by DHT and11KDHT more strongly than T and 11KT. These results indicate that in teleosts, DHT and 11KDHT may be important 5α-reduced androgens produced in the gonads. In contrast, DHT is the only major 5α-reduced androgens in healthy humans.

11-Ketotestosterone is a major androgen produced in porcine adrenal glands and testes.

Porcine steroid hormone profiles have some unique characteristics. We previously studied human and murine steroidogenesis using steroidogenic cells-derived from mesenchymal stem cells (MSCs). To investigate porcine steroidogenesis, we induced steroidogenic cells from porcine subcutaneous preadipocytes (PSPA cells), which originate from MSCs. Using cAMP, adenovirus-mediated introduction of steroidogenic factor-1 (SF-1)/adrenal 4-binding protein (Ad4BP) induced the differentiation of PSPA cells into sex steroid-producing cells. Introducing SF-1/Ad4BP also induced the aldo-keto reductase 1C1 (AKR1C1) gene. Porcine AKR1C1 had 17ß-hydroxysteroid dehydrogenase activity, which converts androstenedione and 11-ketoandrostenedione into testosterone (T) and 11-ketotestosteorne (11KT). Furthermore, differentiated cells expressed hydroxysteroid 11ß-dehydrogenase 2 (HSD11B2) and produced 11KT. HSD11B2 was expressed in testicular Leydig cells and the adrenal cortex. 11KT was present in the plasma of both immature male and female pigs, with slightly higher levels in the male pigs. T levels were much higher in the male pigs. It is noteworthy that in the female pigs, the 11KT levels were >10-fold higher than the T levels. However, castration altered the 11KT and T plasma profiles in the male pigs to near those of the females. 11KT induced endothelial nitric oxide synthase (eNOS) in porcine vascular endothelial cells. These results indicate that 11KT is produced in porcine adrenal glands and testes, and may regulate cardiovascular functions through eNOS expression.

Evaluation of 17ß-hydroxysteroid dehydrogenase activity using androgen receptor-mediated transactivation.

17ß-Hydroxysteroid dehydrogenases (17ß-HSDs) catalyze the reduction of 17-ketosteroids and the oxidation of 17ß-hydroxysteroids to regulate the production of androgens and estrogens. Among them, 17ß-HSD type 3 (HSD17B3) is expressed almost exclusively in testicular Leydig cells and contributes to development of male sexual characteristics by converting androstenedione (A4) to testosterone (T). Mutations of HSD17B3 genes cause a 46,XY disorder of sexual development (46,XY DSD) as a result of low T production. Therefore, the evaluation of 17ß-HSD3 enzymatic activity is important for understanding and diagnosing this disorder. We adapted a method that easily evaluates enzymatic activity of 17ß-HSD3 by quantifying the conversion from A4 to T using androgen receptor (AR)-mediated transactivation. HEK293 cells were transduced to express human HSD17B3, and incubated medium containing A4. Depending on the incubation time with HSD17B3-expressing cells, the culture media progressively increased luciferase activities in CV-1 cells, transfected with the AR expression vector and androgen-responsive reporter. Culture medium from HSD17B1 and HSD17B5-expressing cells also increased the luciferase activities. This system is also applicable to detect the conversion of 11-ketoandrostenedione to 11-ketotestosterone by HSD17B3. Establishment of HEK293 cells expressing various missense mutations in the HSD17B3 gene associated with 46,XY DSD revealed that this system is effective to evaluate the enzymatic activities of mutant proteins.

Prostaglandin F2α Facilitates Platelet Activation by Acting on Prostaglandin E2 Receptor Subtype EP3 and Thromboxane A2 Receptor TP in Mice.

Platelets play an important role in both physiological hemostasis and pathological thrombosis. Thromboxane (TX) A2 and prostaglandin (PG) I2 are well known as a potent stimulator and an inhibitor of platelet function, respectively. Recently, PGE2 has also been reported to regulate platelet function via PGE2 receptor subtypes. However, the effect of PGF2α on platelet function remains to be determined. The aim of the present study was to clarify the effect of PGF2α on murine platelet function both in vitro and in vivo. Platelets prepared from wild-type mice (WT platelets) expressed several types of prostanoid receptors, including the PGE2 receptor subtype EP3 and the TXA2 receptor TP, while expression of the PGF2α receptor FP was not detected. In WT platelets, PGF2α potentiated adenosine diphosphate-induced aggregation in a concentration-dependent manner, while PGF2α alone did not induce aggregation. In platelets prepared from mice lacking FP, however, PGF2α-induced potentiation was not significantly different from that in WT platelets. Interestingly, the potentiation was significantly blunted in platelets lacking EP3 or TP and disappeared completely in platelets lacking both EP3 and TP. Accordingly, PGF2α decreased the cyclic adenosine monophosphate level via EP3 and increased the inositol triphosphate level via TP in WT platelets. Intravenously administered PGF2α significantly shortened the bleeding time and aggravated arachidonic acid-induced acute thromboembolism in WT mice, suggesting that PGF2α works as a platelet stimulator also in vivo. In conclusion, PGF2α potentiates platelet aggregation in vitro via EP3 and TP but not FP. Accordingly, PGF2α facilitates hemostasis and thromboembolism in vivo.

Cyclooxygenase-2 is acutely induced by CCAAT/enhancer-binding protein ß to produce prostaglandin E 2 and F 2α following gonadotropin stimulation in Leydig cells.

Cyclooxygenase 2 (COX-2) is an inducible rate-limiting enzyme for prostanoid production. Because COX-2 represents one of the inducible genes in mouse mesenchymal stem cells upon differentiation into Leydig cells, we investigated COX-2 expression and production of prostaglandin (PG) in Leydig cells. Although COX-2 was undetectable in mouse testis, it was transiently induced in Leydig cells by human chorionic gonadotropin (hCG) administration. Consistent with the finding that Leydig cells expressed aldo-keto reductase 1B7 (PGF synthase) and PGE synthase 2, induction of COX-2 by hCG caused a marked increase in testicular PGF 2α and PGE 2 levels. Using mouse Leydig cell tumor-derived MA-10 cells as a model, it was indicated by reporter assays and electron mobility shift assays that transcription of the COX-2 gene was activated by CCAAT/enhancer-binding protein ß (C/EBPß) with cAMP-stimulation. C/EBPß expression was induced by cAMP-stimulation, whereas expression of C/EBP homolog protein (CHOP) was robustly downregulated. Transfection of CHOP expression plasmid inhibited cAMP-induced COX-2 promoter activity. In addition, CHOP reduced constitutive COX-2 expression in other mouse Leydig cell tumor-derived TM3 cells. These results indicate that COX-2 is induced in Leydig cells by activation of C/EBPß via reduction of CHOP expression upon gonadotropin-stimulation to produce PGF 2α and PGE 2 .

Transcriptional Regulation of Ovarian Steroidogenic Genes: Recent Findings Obtained from Stem Cell-Derived Steroidogenic Cells.

Ovaries represent one of the primary steroidogenic organs, producing estrogen and progesterone under the regulation of gonadotropins during the estrous cycle. Gonadotropins fluctuate the expression of various steroidogenesis-related genes, such as those encoding steroidogenic enzymes, cholesterol deliverer, and electronic transporter. Steroidogenic factor-1 (SF-1)/adrenal 4-binding protein (Ad4BP)/NR5A1 and liver receptor homolog-1 (LRH-1) play important roles in these phenomena via transcriptional regulation. With the aid of cAMP, SF-1/Ad4BP and LRH-1 can induce the differentiation of stem cells into steroidogenic cells. This model is a useful tool for studying the molecular mechanisms of steroidogenesis. In this article, we will provide insight into the transcriptional regulation of steroidogenesis-related genes in ovaries that are revealed from stem cell-derived steroidogenic cells. Using the cells derived from the model, novel SF-1/Ad4BP- and LRH-1-regulated genes were identified by combined DNA microarray and promoter tiling array analyses. The interaction of SF-1/Ad4BP and LRH-1 with transcriptional regulators in the regulation of ovarian steroidogenesis was also revealed.

Angiotensin II promotes pulmonary metastasis of melanoma through the activation of adhesion molecules in vascular endothelial cells.

Hypertension is considered as one of the cancer progressive factors, and often found comorbidity in cancer patients. Renin-angiotensin system (RAS) plays an important role in the regulation of blood pressure, and angiotensin II (Ang II) is well known pressor peptide associated with RAS. Ang II has been reported to accelerate progression and metastasis of cancer cells. However, its precise mechanisms have not been fully understood. In this study, we sought to elucidate the mechanisms by which Ang II exacerbates hematogenous metastasis in mouse melanoma cells, focusing the adhesion pathway in vascular endothelial cells. For this purpose, B16/F10 mouse melanoma cells, which do not express the Ang II type 1 receptor (AT1R), were intravenously injected into C57BL/6 mice. Two weeks after cell injection, the number of lung metastatic colonies was significantly higher in the Ang II-treated group (1 µg/kg/min) than in the vehicle-treated group. The AT1R blocker valsartan (40 mg/kg/day), but not the calcium channel blocker amlodipine (5 or 10 mg/kg/day), significantly suppressed the effect of Ang II. In endothelium-specific Agtr1a knockout mice, Ang II-mediated acceleration of lung metastases of melanoma cells was significantly diminished. Ang II treatment significantly increased E-selectin mRNA expression in vascular endothelial cells collected from lung tissues, and thus promoted adherence of melanoma cells to the vascular endothelium. Ang II-accelerated lung metastases of melanoma cells were also suppressed by treatment with anti-E-selectin antibody (20 mg/kg). Taken together, Ang II-treatment exacerbates hematogenous cancer metastasis by promoting E-selectin-mediated adhesion of cancer cells to vascular endothelial cells.

Prostaglandin I2 suppresses the development of diet-induced nonalcoholic steatohepatitis in mice.

Nonalcoholic steatohepatitis (NASH) is a hepatic manifestation of metabolic syndrome. Although the prostaglandin (PG)I2 receptor IP is expressed broadly in the liver, the role of PGI2-IP signaling in the development of NASH remains to be determined. Here, we investigated the role of the PGI2-IP system in the development of steatohepatitis using mice lacking the PGI2 receptor IP [IP-knockout (IP-KO) mice] and beraprost (BPS), a specific IP agonist. IP-KO and wild-type (WT) mice were fed a methionine- and choline-deficient diet (MCDD) for 2, 5, or 10 wk. BPS was administered orally to mice every day during the experimental periods. The effect of BPS on the expression of chemokine and inflammatory cytokines was examined also in cultured Kupffer cells. WT mice fed MCDD developed steatohepatitis at 10 wk. IP-KO mice developed steatohepatitis at 5 wk with augmented histologic derangements accompanied by increased hepatic monocyte chemoattractant protein-1 (MCP-1) and TNF-α concentrations. After 10 wk of MCDD, IP-KO mice had greater hepatic iron deposition with prominent oxidative stress, resulting in hepatocyte damage. In WT mice, BPS improved histologic and biochemical parameters of steatohepatitis, accompanied by reduced hepatic concentration of MCP-1 and TNF-α. Accordingly, BPS suppressed the LPS-stimulated Mcp-1 and Tnf-α mRNA expression in cultured Kupffer cells prepared from WT mice. PGI2-IP signaling plays a crucial role in the development and progression of steatohepatitis by modulating the inflammatory response, leading to augmented oxidative stress. We suggest that the PGI2-IP system is an attractive therapeutic target for treating patients with NASH.-Kumei, S., Yuhki, K.-I., Kojima, F., Kashiwagi, H., Imamichi, Y., Okumura, T., Narumiya, S., Ushikubi, F. Prostaglandin I2 suppresses the development of diet-induced nonalcoholic steatohepatitis in mice.

Diethylstilbestrol administration inhibits theca cell androgen and granulosa cell estrogen production in immature rat ovary.

Diethylstilbestrol (DES), a strong estrogenic compound, is well-known to affect the reproductive system. In this study, we investigated the effects of DES administration on gonadotropin levels and ovarian steroidogenesis in prepubertal rats. DES treatment acutely reduced serum LH levels, followed by a reduction in the expression of various steroidogenesis-related genes in theca cells. Serum FSH levels were almost unaffected by DES-treatment, even though Cyp19a1 expression was markedly reduced. Serum progesterone, testosterone and estradiol levels were also declined at this time. LH levels recovered from 12 h after DES-treatment and gradually increased until 96 h with a reduction of ERα expression observed in the pituitary. Steroidogenesis-related genes were also up-regulated during this time, except for Cyp17a1 and Cyp19a1. Consistent with observed gene expression pattern, serum testosterone and estradiol concentrations were maintained at lower levels, even though progesterone levels recovered. DES-treatment induced the inducible nitric oxide synthase (iNOS) in granulosa cells, and a nitric oxide generator markedly repressed Cyp19a1 expression in cultured granulosa cells. These results indicate that DES inhibits thecal androgen production via suppression of pituitary LH secretion and ovarian Cyp17a1 expression. In addition, DES represses Cyp19a1 expression by inducing iNOS gene expression for continuous inhibition of estrogen production in granulosa cells.

Cigarette Smoke Extract Inhibits Platelet Aggregation by Suppressing Cyclooxygenase Activity.

The results of studies that were performed to determine whether cigarette smoking affects platelet function have been controversial, and the effects of nicotine- and tar-free cigarette smoke extract (CSE) on platelet function remain to be determined. The aim of this study was to determine the effect of CSE on platelet aggregation and to clarify the mechanism by which CSE affects platelet function. CSE inhibited murine platelet aggregation induced by 9,11-dideoxy-9α,11α-methanoepoxy-prosta-5Z,13E-dien-1-oic acid (U-46619), a thromboxane (TX) A 2 receptor agonist, and that induced by collagen with respective IC 50 values of 1.05 ± 0.14% and 1.34 ± 0.19%. A similar inhibitory action of CSE was also observed in human platelets. CSE inhibited arachidonic acid-induced TXA 2 production in murine platelets with an IC 50 value of 7.32 ± 2.00%. Accordingly, the inhibitory effect of CSE on collagen-induced aggregation was significantly blunted in platelets lacking the TXA 2 receptor compared with the inhibitory effect in control platelets. In contrast, the antiplatelet effects of CSE in platelets lacking each inhibitory prostanoid receptor, prostaglandin (PG) I 2 receptor and PGE 2 receptor subtypes EP 2 and EP 4 , were not significantly different from the effects in respective control platelets. Among the enzymes responsible for TXA 2 production in platelets, the activity of cyclooxygenase (COX)-1 was inhibited by CSE with an IC 50 value of 1.07 ± 0.15% in an uncompetitive manner. In contrast, the activity of TX synthase was enhanced by CSE. The results indicate that CSE inhibits COX-1 activity and thereby decreases TXA 2 production in platelets, leading to inhibition of platelet aggregation.

Induction of steroidogenic cells from adult stem cells and pluripotent stem cells [Review].

Steroid hormones are mainly produced in adrenal glands and gonads. Because steroid hormones play vital roles in various physiological processes, replacement of deficient steroid hormones by hormone replacement therapy (HRT) is necessary for patients with adrenal and gonadal failure. In addition to HRT, tissue regeneration using stem cells is predicted to provide novel therapy. Among various stem cell types, mesenchymal stem cells can be differentiated into steroidogenic cells following ectopic expression of nuclear receptor (NR) 5A subfamily proteins, steroidogenic factor-1 (also known as adrenal 4 binding protein) and liver receptor homolog-1, with the aid of cAMP signaling. Conversely, these approaches cannot be applied to pluripotent stem cells, such as embryonic stem cells and induced pluripotent stem cells, because of poor survival following cytotoxic expression of NR5A subfamily proteins. However, if pluripotent stem cells are first differentiated through mesenchymal lineage, they can also be differentiated into steroidogenic cells via NR5A subfamily protein expression. This approach offers a potential suitable cells for future regenerative medicine and gene therapy for diseases caused by steroidogenesis deficiencies. It represents a powerful tool to investigate the molecular mechanisms involved in steroidogenesis. This article highlights our own and current research on the induction of steroidogenic cells from various stem cells. We also discuss the future direction of their clinical application.

11-Ketotestosterone Is a Major Androgen Produced in Human Gonads.

CONTEXT: 11-ketotestosterone (11-KT) is a novel class of active androgen. However, the detail of its synthesis remains unknown for humans. OBJECTIVE: The objective of this study was to clarify the production and properties of 11-KT in human. Design, Participants, and Methods: Expression of cytochrome P450 and 11ß-hydroxysteroid dehydrogenase types 1 and 2 (key enzymes involved in the synthesis of 11-KT) were investigated in human gonads. The production of 11-KT was investigated in Leydig cells. Plasma concentrations of testosterone and 11-KT were measured in 10 women and 10 men of reproductive age. Investigation of its properties was performed using breast cancer-derived MCF-7 cells. RESULTS: Cytochrome P450 and 11ß-hydroxysteroid dehydrogenase types 1 and 2 were detected in Leydig cells and theca cells. Leydig cells produced 11-KT, and relatively high levels of plasma 11-KT were measured in both men and women. There was no sexual dimorphism in the plasma levels of 11-KT, even though testosterone levels were more than 20 times higher in men than in women. It is noteworthy that the levels of testosterone and 11-KT were similar in women. In a luciferase reporter system, 11-KT activated human androgen receptor-mediated transactivation. Conversely, 11-KT did not activate estrogen receptor-mediated transactivation in aromatase-expressed MCF-7 cells, whereas testosterone did following conversion to estrogen. 11-KT did not affect the estrogen/estrogen receptor -mediated cell proliferation of MCF-7 cells. Furthermore, it significantly inhibited cell proliferation when androgen receptor was transfected into MCF-7 cells. CONCLUSIONS: The current study indicates that 11-KT is produced in the gonads and represents a major androgen in human. It can potentially serve as a nonaromatizable androgen.

Regulation of Steroidogenesis, Development, and Cell Differentiation by Steroidogenic Factor-1 and Liver Receptor Homolog-1.

Steroidogenic factor-1 (SF-1) and liver receptor homolog-1 (LRH-1) belong to the nuclear receptor superfamily and are categorized as orphan receptors. In addition to other nuclear receptors, these play roles in various physiological phenomena by regulating the transcription of target genes. Both factors share very similar structures and exhibit common functions. Of these, the roles of SF-1 and LRH-1 in steroidogenesis are the most important, especially that of SF-1, which was originally discovered and named to reflect such roles. SF-1 and LRH-1 are essential for steroid hormone production in gonads and adrenal glands through the regulation of various steroidogenesis-related genes. As SF-1 is also necessary for the development of gonads and adrenal glands, it is also considered a master regulator of steroidogenesis. Recent studies have clearly demonstrated that LRH-1 also represents another master regulator of steroidogenesis, which similarly to SF-1, can induce differentiation of non-steroidogenic stem cells into steroidogenic cells. Here, we review the functions of both factors in these steroidogenesis-related phenomena.

Establishment and characterization of a novel orthotopic mouse model for human uterine sarcoma with different metastatic potentials.

Uterine sarcomas are rare and aggressive gynecologic tumors with a poor prognosis because of recurrence and metastasis. However, the mechanisms of uterine sarcoma metastasis are largely unknown. To investigate this mechanism, we developed a novel uterine sarcoma tissue-derived orthotopic and metastatic model in KSN nude mice using a green fluorescent protein stably expressed uterine sarcoma cell line, MES-SA. Histological analysis showed that all orthotopic primary tumors were undifferentiated sarcoma. Primary tumors were characterized by high (18)F-fluorodeoxyglucose uptake with a positive correlation to the number of pulmonary metastases. In addition, we generated uterine sarcoma cell sublines with high or low metastatic potentials by serial in vivo selection. Microarray analysis between orthotopic tumors with high and low metastatic potentials revealed differential expression of genes related to cell proliferation and migration (TNNT1, COL1A2, and ZIC1). Our model would be useful to compensate for the limited clinical cases of uterine sarcoma and to investigate the molecular mechanisms of metastatic uterine sarcoma.