Cholesterol Homeostasis and Infertility: The Liver X Receptor Connection
http://www.100md.com
内分泌学杂志 2005年第6期
Department of Biochemistry and Molecular Biology, Georgetown University Medical Center, Washington, D.C. 20057
Address all correspondence and requests for reprints to: Vassilios Papadopoulos, Department of Biochemistry and Molecular Biology, Georgetown University Medical Center, 3900 Reservoir Road Northwest, Washington, D.C. 20057. E-mail: papdopv@georgetown.edu.
Cholesterol is a basic component of all membranes and is the precursor of steroid hormones, bile acids, and vitamin D. Cholesterol and cholesteryl esters are insoluble in water and are transported in the blood from the tissues of origin, mainly liver, to the tissue of storage. Reverse cholesterol transport describes the pathway for removal of excess cholesterol from peripheral tissues via the liver into bile, and its subsequent excretion (1, 2). Although it generally has been assumed that high-density lipoprotein is the obligate transport vehicle for reverse cholesterol transport, new protein members of the nuclear receptor subfamilies and ATP-binding cassette (ABC) transporter were found to regulate cholesterol fluxes (3, 4). Experiments with transgenic animals have indicated that disruption of steps involved in reverse cholesterol transport results in atherosclerosis, whereas overexpression of key components of reverse cholesterol transport exerts atheroprotective effects (4).
The liver X receptor (LXR) belongs to the nuclear receptor family recently shown to act a sensor of cholesterol metabolism and lipid biosynthesis (5). The LXR subfamily consists of two members, LXR and LXR?. LXR is found mainly in liver, adipose tissue, small intestine, adrenal and macrophages, whereas LXR? is a ubiquitous protein. Both proteins are activated by oxysterols. After activation, LXRs heterodimerize with retinoid X receptors (RXR) and initiate the transcription of genes involved in cholesterol efflux, including ABC transporter A1 (ABCA1), sterol regulatory element binding protein, apolipoprotein E, and scavenger receptor class B, member I (5, 6). A physiological role for LXR in the maintenance of cholesterol homeostasis was indicated by the finding that LXR knockout mice, when fed a high-cholesterol diet, developed massive hepatic accumulation of cholesterol (7). Interestingly, LXR–/–, but not LXR?–/–, mice showed a significant increase in hepatic cholesterol content (8), and LXR–/–/LXR?–/– mice showed changes in blood lipid profiles (9).
In this issue of Endocrinology, Robertson et al. (10) report data demonstrating that LXR? plays a crucial role in maintaining cholesterol homeostasis in the testis. Importantly, detailed morphological analysis of the testis revealed that deletion of the LXR? gene, but not LXR, resulted in dramatic, time-dependent Sertoli cell cholesterol ester lipid accumulation and germ cell depletion that were correlated with infertility. The effects on germ cell loss were even more dramatic in LXR–/–?–/– mice. In contrast to the effects seen in Sertoli cells, there was no lipid accumulation in Leydig cells in LXR?–/–or in LXR–/–?–/– mice, although testosterone formation by these cells was reduced.
The Sertoli cell phagocytizes germ cells undergoing apoptosis and degrades residual bodies generated in the last steps of spermatogenesis (11, 12). This extensive phagocytotic activity would be expected to result in excess lipid accumulation in Sertoli cells, and an active lipid efflux process would be required to eliminate this material. Lipid accumulation in Sertoli cells was shown in this manuscript (10) to correlate with reduced numbers of germ cells and with infertility and has been shown in humans to correlate with azoospermia (13). Sertoli cell cholesterol could come from de novo synthesis, increased uptake of high-density lipoprotein cholesterol (14), or reduced efflux. Because hydroxymethylglutaryl-coenzyme A reductase or synthase, low-density lipoprotein (LDL) receptor and vLDL receptor gene expression were not affected by the deletion of LXR?, it can be concluded that the cholesterol efflux mechanism was impaired in LXR? null mice. Thus, the data presented in this paper suggest that a LXR?-target gene mediates cholesterol efflux in Sertoli cells. ABCA1 is a LXR? target gene. Indeed, recent data obtained in ABCA1–/– mice indicated a phenotype close to that seen in LXR?–/– mice, with increased lipid accumulation in Sertoli cells paralleling reduced fertility (15). In another recent paper, it was shown that ABCA1 levels were reduced in LXR?–/– mice, and that this led to Sertoli cell lipid accumulation (16). However, the authors failed to observe an effect on ABCA1 gene expression suggesting that other not-yet-identified LXR target genes might be involved in mediating cholesterol efflux in Sertoli cells.
Despite the dramatic effect of the deletion of LXR genes on Sertoli cell lipid accumulation, it is unlikely that this was the sole factor involved in the arrest of spermatogenesis, and thus, infertility. As noted earlier, LXR heterodimerizes with RXR to induce changes in gene transcription. It recently was shown that although RXR?–/– mice are sterile, RXR?af20 males are fertile. RXR?af20 mice express RXR? carrying a mutation of its activation function-2 core responsible for the transcriptional function of the protein (16). Interestingly, testes from both RXR?–/– and RXR?af20 mice show increased lipid accumulation and same size lipid droplets in the Sertoli cells of the seminiferous epithelium.
The finding that Leydig cells in LXR?–/– and LXR–/– ?–/– mice produced less testosterone than the wild-type mice is of interest. Considering that LH levels have been reported to be unaffected or even increased in older animals, these results suggest the possibility of a direct effect of LXR deletion on Leydig cell function. There are other possibilities, however. For example, the observation that the lipid content of Leydig cells was not affected, whereas Sertoli cells were dramatically affected, led the authors to hypothesize that the effect on Leydig cells might be due to changes in paracrine regulation of Leydig cell androgen formation by Sertoli cell products (17). Indeed, data presented in this paper and by Mascrez et al. (16) suggest that Sertoli cell function is under the control of LXR, and there is solid evidence for paracrine regulation of Leydig cell function by the Sertoli cell (17). Another possibility is that reduced androgen formation might be due to the increased serum corticosterone levels seen in LXR–/–?–/– compared with wild-type mice (18). Indeed, increased glucocorticoid levels have been show to inhibit in vitro and in vivo Leydig cell function (19, 20), and in men, stress-induced increase in glucocorticoid levels resulted in inhibition of androgen formation, low sperm counts, and infertility (21).
Changes in Sertoli cell function, coupled with reduced testosterone in LXR? null mice, might also result in reduced estrogen formation by the testis. The testis forms approximately 25% of circulating estrogens in rodents (22, 23). Recent data suggest that estradiol may be a critical factor for normal reproduction, but also for various physiopathological processes, such as atherosclerosis (24, 25, 26).
In conclusion, the study of Robertson et al. (10) identifies LXR? as the key regulator of cholesterol homeostasis in the Sertoli cells and confirms recent findings on the critical nature of cholesterol efflux by Sertoli cells in spermatogenesis and fertility in rodents and humans. Identification of the LXR? target genes mediating this effect of LXR on Sertoli cell cholesterol efflux would elucidate the mechanism of cholesterol efflux by these cells. Importantly, the data showing that LXR–/–?–/– mice have impaired triglyceride metabolism and increased LDL (9), increased glucocorticoid (18), and reduced testosterone (10) levels, accumulated cholesterol in the arterial wall (9), and are infertile (10) could also be interpreted as an indication that LXR might be the link between risk factors (high cholesterol) for atherosclerosis and cardiovascular disease in general, the leading cause of global mortality (27), and reports of global decline in the numbers of spermatozoa produced by men (28, 29), an alarming prospect for the future of our species.
References
von Eckardstein A, Nofer JR, Assmann G 2001 High density lipoproteins and arteriosclerosis: role of cholesterol efflux and reverse cholesterol transport. Arterioscler Thromb Vasc Biol 21:13–27
Groen AK, Oude Elferink RP, Verkade HJ, Kuipers F 2004 The ins and outs of reverse cholesterol transport. Ann Med 36:135–145
Assmann G, Gotto Jr AM 2004 HDL cholesterol and protective factors in atherosclerosis. Circulation 109:III8-III14
von Eckardstein A, Nofer JR, Assmann G 2000 Acceleration of reverse cholesterol transport. Curr Opin Cardiol 15:348–354
Steffensen KR, Gustafsson JA 2004 Putative metabolic effects of the liver X receptor (LXR). Diabetes 53(Suppl 1):S36–S42
Francis GA, Fayard E, Picard F, Auwerx J 2003 Nuclear receptors and the control of metabolism. Annu Rev Physiol 65:261–311
Peet DJ, Turley SD, Ma W, Janowski BA, Lobaccaro JM, Hammer RE, Mangelsdorf DJ 1998 Cholesterol and bile acid metabolism are impaired in mice lacking the nuclear oxysterol receptor LXR . Cell 93:693–704
Alberti S, Schuster G, Parini P, Feltkamp D, Diczfalusy U, Rudling M, Angelin B, Bjorkhem I, Pettersson S, Gustafsson JA 2001 Hepatic cholesterol metabolism and resistance to dietary cholesterol in LXR?-deficient mice. J Clin Invest 107:565–573
Schuster GU, Parini P, Wang L, Alberti S, Steffensen KR, Hansson GK, Angelin B, Gustafsson JA 2002 Accumulation of foam cells in liver X receptor-deficient mice. Circulation 106:1147–1153
Robertson KM, Schuster GU, Steffensen KR, Hovatta O, Meaney S, Hultenby K, Johansson LC, Svechnikov K, Soder O, Gustafsson JA 2005 The liver X receptor-? is essential for maintaining cholesterol homeostasis in the testis. Endocrinology 146:2519–2530
Russell LD, Hikim APS, Ettlin RA, Clegg ED 1990 Histological and histopathological evaluation of the testis. Clearwater, FL: Cache River Press
Nakanishi Y, Shiratsuchi A 2004 Phagocytic removal of apoptotic spermatogenic cells by Sertoli cells: mechanisms and consequences. Biol Pharm Bull 27:13–16
Holstein AF, Roosen-Runge EC, Schirren C 1988 Illustrated pathology of human spermatogenesis. Berlin: Grosse Verlag
Fofana M, Travert C, Carreau S, Le Goff D 2000 Evaluation of cholesteryl ester transfer in the seminiferous tubule cells of immature rats in vitro and in vivo. J Reprod Fertil 118:79–83
Selva DM, Hirsch-Reinshagen V, Burgess B, Zhou S, Chan J, McIsaac S, Hayden MR, Hammond GL, Vogl AW, Wellington CL 2004 The ATP-binding cassette transporter 1 mediates lipid efflux from Sertoli cells and influences male fertility. J Lipid Res 45:1040–1050
Mascrez B, Ghyselinck NB, Watanabe M, Annicotte JS, Chambon P, Auwerx J, Mark M 2004 Ligand-dependent contribution of RXR? to cholesterol homeostasis in Sertoli cells. EMBO Rep 5:285–290
Habert R, Lejeune H, Saez JM 2001Origin, differentiation and regulation of fetal and adult Leydig cells. Mol Cell Endocrinol 179:47–74
Steffensen KR, Neo SY, Stulnig TM, Vega VB, Rahman SS, Schuster GU, Gustafsson JA 2004, Liu ET204 Genome-wide expression profiling; a panel of mouse tissues discloses novel biological functions of liver X receptors in adrenals. J Mol Endocrinol 33:609–622
Bambino TH, Hsueh AJ 1981 Direct inhibitory effect of glucocorticoids upon testicular luteinizing hormone receptor and steroidogenesis in vivo and in vitro. Endocrinology 108:2142–2148
Hardy MP, Ganjam VK 1997 Stress, 11?-HSD, and Leydig cell function. J Androl 18:475–479
Fenster L, Katz DF, Wyrobek AJ, Pieper C, Rempel DM, Oman D, Swan SH 1997 Effects of psychological stress on human semen quality. J Androl 18:194–202
Jong FH, Hey AH, Van Der Molen HJ 1973 Effect of gonadotropins on the secretion of oestradiol 17? and testosterone by the rat testis. J Endocr 57:277–284
Papadopoulos V, Carreau S, Szerman-Joly E, Drosdowsky MA, Dehennin L, Scholler R 1986 Rat testis 17?-estradiol: identification by gas chromatography-mass spectrometry and age related cellular distribution. J Steroid Biochem 24:1211–1216
Sharpe RM, Skakkebaek NE 1993 Are oestrogens involved in falling sperm counts and disorders of the male reproductive tract? Lancet 341:1392–1395
Carreau S 2003 Estrogens—male hormones? Folia Histochem Cytobiol 41:107–111
von Eckardstein A, Wu FCW 2003 Testosterone and atherosclerosis. Growth Horm IGF Res 13:S72–S84
Smith Jr SC, Jackson R, Pearson TA, Fuster V, Yusuf S, Faergeman O, Wood DA, Alderman M, Horgan J, Home P, Hunn M, Grundy SM 2004 Principles for national and regional guidelines on cardiovascular disease prevention: a scientific statement from the World Heart and Stroke Forum. Circulation 109:3112–3121
Skakkebaek NE, Rajpert-De Meyts E, Main KM 2001 Testicular dysgenesis syndrome: an increasingly common developmental disorder with environmental aspects. Hum Reprod 16:972–978
Jouannet P, Wang C, Eustache F, Kold-Jensen T, Auger J 2001 Semen quality and male reproductive health: the controversy about human sperm concentration decline. APMIS 109:333–344(Vassilios Papadopoulos)
Address all correspondence and requests for reprints to: Vassilios Papadopoulos, Department of Biochemistry and Molecular Biology, Georgetown University Medical Center, 3900 Reservoir Road Northwest, Washington, D.C. 20057. E-mail: papdopv@georgetown.edu.
Cholesterol is a basic component of all membranes and is the precursor of steroid hormones, bile acids, and vitamin D. Cholesterol and cholesteryl esters are insoluble in water and are transported in the blood from the tissues of origin, mainly liver, to the tissue of storage. Reverse cholesterol transport describes the pathway for removal of excess cholesterol from peripheral tissues via the liver into bile, and its subsequent excretion (1, 2). Although it generally has been assumed that high-density lipoprotein is the obligate transport vehicle for reverse cholesterol transport, new protein members of the nuclear receptor subfamilies and ATP-binding cassette (ABC) transporter were found to regulate cholesterol fluxes (3, 4). Experiments with transgenic animals have indicated that disruption of steps involved in reverse cholesterol transport results in atherosclerosis, whereas overexpression of key components of reverse cholesterol transport exerts atheroprotective effects (4).
The liver X receptor (LXR) belongs to the nuclear receptor family recently shown to act a sensor of cholesterol metabolism and lipid biosynthesis (5). The LXR subfamily consists of two members, LXR and LXR?. LXR is found mainly in liver, adipose tissue, small intestine, adrenal and macrophages, whereas LXR? is a ubiquitous protein. Both proteins are activated by oxysterols. After activation, LXRs heterodimerize with retinoid X receptors (RXR) and initiate the transcription of genes involved in cholesterol efflux, including ABC transporter A1 (ABCA1), sterol regulatory element binding protein, apolipoprotein E, and scavenger receptor class B, member I (5, 6). A physiological role for LXR in the maintenance of cholesterol homeostasis was indicated by the finding that LXR knockout mice, when fed a high-cholesterol diet, developed massive hepatic accumulation of cholesterol (7). Interestingly, LXR–/–, but not LXR?–/–, mice showed a significant increase in hepatic cholesterol content (8), and LXR–/–/LXR?–/– mice showed changes in blood lipid profiles (9).
In this issue of Endocrinology, Robertson et al. (10) report data demonstrating that LXR? plays a crucial role in maintaining cholesterol homeostasis in the testis. Importantly, detailed morphological analysis of the testis revealed that deletion of the LXR? gene, but not LXR, resulted in dramatic, time-dependent Sertoli cell cholesterol ester lipid accumulation and germ cell depletion that were correlated with infertility. The effects on germ cell loss were even more dramatic in LXR–/–?–/– mice. In contrast to the effects seen in Sertoli cells, there was no lipid accumulation in Leydig cells in LXR?–/–or in LXR–/–?–/– mice, although testosterone formation by these cells was reduced.
The Sertoli cell phagocytizes germ cells undergoing apoptosis and degrades residual bodies generated in the last steps of spermatogenesis (11, 12). This extensive phagocytotic activity would be expected to result in excess lipid accumulation in Sertoli cells, and an active lipid efflux process would be required to eliminate this material. Lipid accumulation in Sertoli cells was shown in this manuscript (10) to correlate with reduced numbers of germ cells and with infertility and has been shown in humans to correlate with azoospermia (13). Sertoli cell cholesterol could come from de novo synthesis, increased uptake of high-density lipoprotein cholesterol (14), or reduced efflux. Because hydroxymethylglutaryl-coenzyme A reductase or synthase, low-density lipoprotein (LDL) receptor and vLDL receptor gene expression were not affected by the deletion of LXR?, it can be concluded that the cholesterol efflux mechanism was impaired in LXR? null mice. Thus, the data presented in this paper suggest that a LXR?-target gene mediates cholesterol efflux in Sertoli cells. ABCA1 is a LXR? target gene. Indeed, recent data obtained in ABCA1–/– mice indicated a phenotype close to that seen in LXR?–/– mice, with increased lipid accumulation in Sertoli cells paralleling reduced fertility (15). In another recent paper, it was shown that ABCA1 levels were reduced in LXR?–/– mice, and that this led to Sertoli cell lipid accumulation (16). However, the authors failed to observe an effect on ABCA1 gene expression suggesting that other not-yet-identified LXR target genes might be involved in mediating cholesterol efflux in Sertoli cells.
Despite the dramatic effect of the deletion of LXR genes on Sertoli cell lipid accumulation, it is unlikely that this was the sole factor involved in the arrest of spermatogenesis, and thus, infertility. As noted earlier, LXR heterodimerizes with RXR to induce changes in gene transcription. It recently was shown that although RXR?–/– mice are sterile, RXR?af20 males are fertile. RXR?af20 mice express RXR? carrying a mutation of its activation function-2 core responsible for the transcriptional function of the protein (16). Interestingly, testes from both RXR?–/– and RXR?af20 mice show increased lipid accumulation and same size lipid droplets in the Sertoli cells of the seminiferous epithelium.
The finding that Leydig cells in LXR?–/– and LXR–/– ?–/– mice produced less testosterone than the wild-type mice is of interest. Considering that LH levels have been reported to be unaffected or even increased in older animals, these results suggest the possibility of a direct effect of LXR deletion on Leydig cell function. There are other possibilities, however. For example, the observation that the lipid content of Leydig cells was not affected, whereas Sertoli cells were dramatically affected, led the authors to hypothesize that the effect on Leydig cells might be due to changes in paracrine regulation of Leydig cell androgen formation by Sertoli cell products (17). Indeed, data presented in this paper and by Mascrez et al. (16) suggest that Sertoli cell function is under the control of LXR, and there is solid evidence for paracrine regulation of Leydig cell function by the Sertoli cell (17). Another possibility is that reduced androgen formation might be due to the increased serum corticosterone levels seen in LXR–/–?–/– compared with wild-type mice (18). Indeed, increased glucocorticoid levels have been show to inhibit in vitro and in vivo Leydig cell function (19, 20), and in men, stress-induced increase in glucocorticoid levels resulted in inhibition of androgen formation, low sperm counts, and infertility (21).
Changes in Sertoli cell function, coupled with reduced testosterone in LXR? null mice, might also result in reduced estrogen formation by the testis. The testis forms approximately 25% of circulating estrogens in rodents (22, 23). Recent data suggest that estradiol may be a critical factor for normal reproduction, but also for various physiopathological processes, such as atherosclerosis (24, 25, 26).
In conclusion, the study of Robertson et al. (10) identifies LXR? as the key regulator of cholesterol homeostasis in the Sertoli cells and confirms recent findings on the critical nature of cholesterol efflux by Sertoli cells in spermatogenesis and fertility in rodents and humans. Identification of the LXR? target genes mediating this effect of LXR on Sertoli cell cholesterol efflux would elucidate the mechanism of cholesterol efflux by these cells. Importantly, the data showing that LXR–/–?–/– mice have impaired triglyceride metabolism and increased LDL (9), increased glucocorticoid (18), and reduced testosterone (10) levels, accumulated cholesterol in the arterial wall (9), and are infertile (10) could also be interpreted as an indication that LXR might be the link between risk factors (high cholesterol) for atherosclerosis and cardiovascular disease in general, the leading cause of global mortality (27), and reports of global decline in the numbers of spermatozoa produced by men (28, 29), an alarming prospect for the future of our species.
References
von Eckardstein A, Nofer JR, Assmann G 2001 High density lipoproteins and arteriosclerosis: role of cholesterol efflux and reverse cholesterol transport. Arterioscler Thromb Vasc Biol 21:13–27
Groen AK, Oude Elferink RP, Verkade HJ, Kuipers F 2004 The ins and outs of reverse cholesterol transport. Ann Med 36:135–145
Assmann G, Gotto Jr AM 2004 HDL cholesterol and protective factors in atherosclerosis. Circulation 109:III8-III14
von Eckardstein A, Nofer JR, Assmann G 2000 Acceleration of reverse cholesterol transport. Curr Opin Cardiol 15:348–354
Steffensen KR, Gustafsson JA 2004 Putative metabolic effects of the liver X receptor (LXR). Diabetes 53(Suppl 1):S36–S42
Francis GA, Fayard E, Picard F, Auwerx J 2003 Nuclear receptors and the control of metabolism. Annu Rev Physiol 65:261–311
Peet DJ, Turley SD, Ma W, Janowski BA, Lobaccaro JM, Hammer RE, Mangelsdorf DJ 1998 Cholesterol and bile acid metabolism are impaired in mice lacking the nuclear oxysterol receptor LXR . Cell 93:693–704
Alberti S, Schuster G, Parini P, Feltkamp D, Diczfalusy U, Rudling M, Angelin B, Bjorkhem I, Pettersson S, Gustafsson JA 2001 Hepatic cholesterol metabolism and resistance to dietary cholesterol in LXR?-deficient mice. J Clin Invest 107:565–573
Schuster GU, Parini P, Wang L, Alberti S, Steffensen KR, Hansson GK, Angelin B, Gustafsson JA 2002 Accumulation of foam cells in liver X receptor-deficient mice. Circulation 106:1147–1153
Robertson KM, Schuster GU, Steffensen KR, Hovatta O, Meaney S, Hultenby K, Johansson LC, Svechnikov K, Soder O, Gustafsson JA 2005 The liver X receptor-? is essential for maintaining cholesterol homeostasis in the testis. Endocrinology 146:2519–2530
Russell LD, Hikim APS, Ettlin RA, Clegg ED 1990 Histological and histopathological evaluation of the testis. Clearwater, FL: Cache River Press
Nakanishi Y, Shiratsuchi A 2004 Phagocytic removal of apoptotic spermatogenic cells by Sertoli cells: mechanisms and consequences. Biol Pharm Bull 27:13–16
Holstein AF, Roosen-Runge EC, Schirren C 1988 Illustrated pathology of human spermatogenesis. Berlin: Grosse Verlag
Fofana M, Travert C, Carreau S, Le Goff D 2000 Evaluation of cholesteryl ester transfer in the seminiferous tubule cells of immature rats in vitro and in vivo. J Reprod Fertil 118:79–83
Selva DM, Hirsch-Reinshagen V, Burgess B, Zhou S, Chan J, McIsaac S, Hayden MR, Hammond GL, Vogl AW, Wellington CL 2004 The ATP-binding cassette transporter 1 mediates lipid efflux from Sertoli cells and influences male fertility. J Lipid Res 45:1040–1050
Mascrez B, Ghyselinck NB, Watanabe M, Annicotte JS, Chambon P, Auwerx J, Mark M 2004 Ligand-dependent contribution of RXR? to cholesterol homeostasis in Sertoli cells. EMBO Rep 5:285–290
Habert R, Lejeune H, Saez JM 2001Origin, differentiation and regulation of fetal and adult Leydig cells. Mol Cell Endocrinol 179:47–74
Steffensen KR, Neo SY, Stulnig TM, Vega VB, Rahman SS, Schuster GU, Gustafsson JA 2004, Liu ET204 Genome-wide expression profiling; a panel of mouse tissues discloses novel biological functions of liver X receptors in adrenals. J Mol Endocrinol 33:609–622
Bambino TH, Hsueh AJ 1981 Direct inhibitory effect of glucocorticoids upon testicular luteinizing hormone receptor and steroidogenesis in vivo and in vitro. Endocrinology 108:2142–2148
Hardy MP, Ganjam VK 1997 Stress, 11?-HSD, and Leydig cell function. J Androl 18:475–479
Fenster L, Katz DF, Wyrobek AJ, Pieper C, Rempel DM, Oman D, Swan SH 1997 Effects of psychological stress on human semen quality. J Androl 18:194–202
Jong FH, Hey AH, Van Der Molen HJ 1973 Effect of gonadotropins on the secretion of oestradiol 17? and testosterone by the rat testis. J Endocr 57:277–284
Papadopoulos V, Carreau S, Szerman-Joly E, Drosdowsky MA, Dehennin L, Scholler R 1986 Rat testis 17?-estradiol: identification by gas chromatography-mass spectrometry and age related cellular distribution. J Steroid Biochem 24:1211–1216
Sharpe RM, Skakkebaek NE 1993 Are oestrogens involved in falling sperm counts and disorders of the male reproductive tract? Lancet 341:1392–1395
Carreau S 2003 Estrogens—male hormones? Folia Histochem Cytobiol 41:107–111
von Eckardstein A, Wu FCW 2003 Testosterone and atherosclerosis. Growth Horm IGF Res 13:S72–S84
Smith Jr SC, Jackson R, Pearson TA, Fuster V, Yusuf S, Faergeman O, Wood DA, Alderman M, Horgan J, Home P, Hunn M, Grundy SM 2004 Principles for national and regional guidelines on cardiovascular disease prevention: a scientific statement from the World Heart and Stroke Forum. Circulation 109:3112–3121
Skakkebaek NE, Rajpert-De Meyts E, Main KM 2001 Testicular dysgenesis syndrome: an increasingly common developmental disorder with environmental aspects. Hum Reprod 16:972–978
Jouannet P, Wang C, Eustache F, Kold-Jensen T, Auger J 2001 Semen quality and male reproductive health: the controversy about human sperm concentration decline. APMIS 109:333–344(Vassilios Papadopoulos)