Differential gene expression of organic anion transporters in male and female rats.

Sex-related differential gene expression of organic anion transporters (rOAT1, rOAT2, and rOAT3) in rat brain, liver, and kidney was investigated. There were no sex differences in the expression of rOAT1 mRNA. rOAT2 mRNA was abundant in the liver and weakly expressed in the kidney of male rats; however, the OAT2 gene was strongly expressed in both organs of females. The abundance of rOAT2 mRNA markedly increased in castrated male rat kidney; however, treatment of castrated male rats with testosterone led to a decrease of rOAT2 mRNA. Expression of rOAT3 mRNA in intact female rats was found in the kidney and brain, whereas in males rOAT3 mRNA was also found in the liver. rOAT3 mRNA markedly decreased in the liver of castrated male rats but increased in testosterone-treated castrated male rats. Moreover, rOAT3 mRNA increased in the hypophysectomized female rat liver, indicating that rOAT3 is an inducible isoform. The present findings suggest that sex steroids play an important role in the expression and maintenance of OAT2/3 isoforms in the rat liver and kidney. Our results provide information on the differential gene expression of OAT isoforms with sex hormone dependency.     

Sex‐biased transcriptome divergence along a latitudinal gradient

Sex‐dependent gene expression is likely an important genomic mechanism that allows sex‐specific adaptation to environmental changes. Among Drosophila species, sex‐biased genes display remarkably consistent evolutionary patterns; male‐biased genes evolve faster than unbiased genes in both coding sequence and expression level, suggesting sex differences in selection through time. However, comparatively little is known of the evolutionary process shaping sex‐biased expression within species. Latitudinal clines offer an opportunity to examine how changes in key ecological parameters also influence sex‐specific selection and the evolution of sex‐biased gene expression. We assayed male and female gene expression in Drosophila serrata along a latitudinal gradient in eastern Australia spanning most of its endemic distribution. Analysis of 11 631 genes across eight populations revealed strong sex differences in the frequency, mode and strength of divergence. Divergence was far stronger in males than females and while latitudinal clines were evident in both sexes, male divergence was often population specific, suggesting responses to localized selection pressures that do not covary predictably with latitude. While divergence was enriched for male‐biased genes, there was no overrepresentation of X‐linked genes in males. By contrast, X‐linked divergence was elevated in females, especially for female‐biased genes. Many genes that diverged in D. serrata have homologs also showing latitudinal divergence in Drosophila simulans and Drosophila melanogaster on other continents, likely indicating parallel adaptation in these distantly related species. Our results suggest that sex differences in selection play an important role in shaping the evolution of gene expression over macro‐ and micro‐ecological spatial scales.     

Sex differences and the effects of ovariectomy on the β-adrenergic contractile response

The presence of sex differences in myocardial β-adrenergic responsiveness is controversial, and limited studies have addressed the mechanism underlying these differences. Studies were performed using isolated perfused hearts from male, intact female and ovariectomized female mice to investigate sex differences and the effects of ovarian hormone withdrawal on β-adrenergic receptor function. Female hearts exhibited blunted contractile responses to the β-adrenergic receptor agonist isoproterenol (ISO) compared with males but not ovariectomized females. There were no sex differences in β(1)-adrenergic receptor gene or protein expression. To investigate the role of adenylyl cyclase, phosphodiesterase, and the cAMP-signaling cascade in generating sex differences in the β-adrenergic contractile response, dose-response studies were performed in isolated perfused male and female hearts using forskolin, 3-isobutyl-1-methylxanthine (IBMX), and 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate (CPT-cAMP). Males showed a modestly enhanced contractile response to forskolin at 300 nM and 5 μM compared with females, but there were no sex differences in the response to IBMX or CPT-cAMP. The role of the A(1) adenosine receptor (A(1)AR) in antagonizing the β-adrenergic contractile response was investigated using both the A(1)AR agonist 2-chloro-N(6)-cyclopentyl-adenosine and A(1)AR knockout (KO) mice. Intact females showed an enhanced A(1)AR anti-adrenergic effect compared with males and ovariectomized females. The β-adrenergic contractile response was potentiated in both male and female A(1)ARKO hearts, with sex differences no longer present above 1 nM ISO. The β-adrenergic contractile response is greater in male hearts than females, and minor differences in the action of adenylyl cyclase or the A(1)AR may contribute to these sex differences.     

Sex-specific effects of spironolactone on blood pressure in gonadectomized male and female Wistar rats

A low-salt diet is known to decrease and salt excess to increase blood pressure in humans and rodents. Sex steroids seem to play a role in salt dependent hypertension. However, little is known about sex differences in mineralocorticoid receptor blockade between male and female rats. The objective of the work was at first to investigate the effects of a low-salt vs. a high-salt diet on blood pressure without the influence of gonadal steroids in male and female rats. Second, to determine the sex-specific effects of mineralocorticoid receptor blockade by spironolactone in high-salt and low-salt fed gonadectomized male and female animals. Normotensive male and female Wistar rats were gonadectomized and put on a low (NaCl<0.03%) or high (NaCl=4%) salt diet. On each diet animals received spironolactone or placebo. Blood pressure was measured by tail-cuff-method; 24-h urine samples were collected in metabolic cages and blood was collected for hormonal measurements. High-salt diet significantly increased systolic blood pressure in both sexes. This effect could be blocked effectively by spironolactone only in male rats. Spironolactone treatment significantly increased aldosterone levels in males and females independent of the sodium content of the diet. High sodium diet significantly increased relative kidney weight, which was not altered by spironolactone treatment. Independently of gonadal steroids a high-salt diet increased blood pressure in gonadectomized male and female rats. Spironolactone lowered blood pressure only in male not in female rats on a high-salt diet clearly indicating sex-specific effects of the mineralo-corticoid antagonist spironolactone.     

Sexual Dimorphism in Urinary Angiotensinogen Excretion During Chronic Angiotensin II−Salt Hypertension

BackgroundThe intrarenal renin−angiotensin system contributes to hypertension by regulating sodium and water reabsorption throughout the nephron. Sex differences in the intrarenal components of the renin−angiotensin system have been involved in the greater incidence of high blood pressure and progression to kidney damage in males than females.ObjectiveThis study investigated whether there is a sex difference in the intrarenal gene expression and urinary excretion of angiotensinogen (AGT) during angiotensin II (Ang II)−dependent hypertension and high-salt (HS) diet.MethodsMale and female Sprague-Dawley rats were divided into 5 groups for each sex: Normal-salt control, HS diet (8% NaCl), Ang II−infused (80 ng/min), Ang II−infused plus HS diet, and Ang II−infused plus HS diet and treatment with the Ang II receptor blocker, candesartan (25 mg/L in the drinking water). Rats were evaluated for systolic blood pressure (SBP), kidney AGT mRNA expression, urinary AGT excretion, and proteinuria at different time points during a 14-day protocol.ResultsBoth male and female rats exhibited similar increases in urinary AGT, with increases in SBP during chronic Ang II infusion. HS diet greatly exacerbated the urinary AGT excretion in Ang II−infused rats; males had a 9-fold increase over Ang II alone and females had a 2.5-fold increase. Male rats displayed salt-sensitive SBP increases during Ang II infusion and HS diet, and female rats did not. In the kidney cortex, males displayed greater AGT gene expression than females during all treatments. During Ang II infusion, both sexes exhibited increases in AGT gene message compared with same-sex controls. In addition, HS diet combined with Ang II infusion exacerbated the proteinuria in both sexes. Concomitant Ang II receptor blocker treatment during Ang II infusion and HS diet decreased SBP and urinary AGT similarly in both sexes; however, the decrease in proteinuria was greater in the females.ConclusionDuring Ang II−dependent hypertension and HS diet, higher intrarenal renin-angiotensin system activation in males, as reflected by higher AGT gene expression and urinary excretion, indicates a mechanism for greater progression of high blood pressure and might explain the sex disparity in development of salt-sensitive hypertension.     

Effects of dietary protein restriction on nephron number in the mouse

In rats, maternal protein restriction reduces nephron endowment and often leads to adult hypertension. Sex differences in these responses have been identified. The molecular and genetic bases of these phenomena can best be identified in a mouse model, but effects of maternal protein restriction on kidney development have not been examined in mice. Therefore, we determined how combined prenatal and postnatal protein restriction in mice affects organ weight, glomerular number and dimensions, and renal expression of angiotensin receptor mRNA, in both male and female offspring. C57/BL6/129sv mice received either a normal (20% wt/wt; NP) or low (9% wt/wt; LP) protein diet during gestation and postnatal life. Offspring were examined at postnatal day 30. Protein restriction retarded growth of the kidney, liver, spleen, heart, and brain. All organs except the brain weighed less in female than male offspring. Protein restriction increased normalized (to body weight) brain weight, with females having relatively heavier brains than males. The effects of protein restriction were not sex dependent, except that normalized liver weight was reduced in males but increased in females. Glomerular volume, but not number, was greater in female than in male mice. Maternal protein restriction reduced nephron endowment similarly in male and female mice. Renal expression of AT(1A) receptor mRNA was approximately sixfold greater in female than male NP mice, but similar in male LP and female LP mice. We conclude that maternal protein restriction reduces nephron endowment in mice. This effect provides a basis for future studies of developmental programming in the mouse.     

Major molecular differences between mammalian sexes are involved in drug metabolism and renal function

Many anatomical differences exist between males and females; these are manifested on a molecular level by different hormonal environments. Although several molecular differences in adult tissues have been identified, a comprehensive investigation of the gene expression differences between males and females has not been performed. We surveyed the expression patterns of 13,977 mouse genes in male and female hypothalamus, kidney, liver, and reproductive tissues. Extensive differential gene expression was observed not only in the reproductive tissues, but also in the kidney and liver. The differentially expressed genes are involved in drug and steroid metabolism, osmotic regulation, or as yet unresolved cellular roles. In contrast, very few molecular differences were observed between the male and female hypothalamus in both mice and humans. We conclude that there are persistent differences in gene expression between adult males and females. These molecular differences have important implications for the physiological differences between males and females.     

Sex‐Dependent Dietary Obesity,Induction of UCPs,and Leptin Expression in Rat Adipose Tissues

Objective: The aim of this study was to determine the sex‐dependent differences in the response of key parameters involved in thermogenesis and control of body weight in brown adipose tissue (BAT) and white adipose tissue (WAT) in postcafeteria‐fed rats, a model of dietary obesity. Research Methods and Procedures: BAT and WAT were obtained from male and female control and postcafeteria‐fed Wistar rats. Postcafeteria‐fed rats were initially fed with cafeteria diet from day 10 of life until day 110 (cafeteria period) and with standard chow diet from then until day 180 of life (postcafeteria period). Body mass and energy intake were evaluated. Biometric parameters were analyzed in interscapular BAT (IBAT). Levels of uncoupling protein 1 (UCP1), α₂‐adrenergic receptor (AR), and β₃‐AR proteins and UCP1, UCP2, UCP3, β₃‐AR, and leptin mRNAs, in IBAT or WAT, were studied by Western blot and Northern blot analyses, respectively. Results: Rats attained 59% (females) and 39% (males) increase in body weight at the end of the cafeteria period. During the postcafeteria period, the rats showed a loss of body weight, which was higher in females. Postcafeteria‐fed female rats also presented higher activation of thermogenic parameters in IBAT, including UCP1, UCP2, and UCP3 mRNAs. Female control rats showed lower levels of both α2 and β₃‐ARs in BAT compared with male rats, but these levels in postcafeteria‐fed female and male rats were the same, because males tended to down‐regulate them. Levels of leptin mRNA in response to the postcafeteria state depended on gender and the specific WAT depot studied. Discussion: It is suggested that in postcafeteria‐fed female rats, BAT thermogenic capacity becomes more efficiently activated than in males. Female rats also showed a bigger weight loss. The parallel regulation of the levels of UCP2 and UCP3 mRNAs, with respect to UCP1 mRNA, with higher activation in female postcafeteria‐fed rats, suggests a possible role of both UCP2 and UCP3 in the regulation of energy expenditure and in the control of body weight. The distinct responses to overweight of α2 and β₃‐ARs—which were sex dependent—and leptin mRNA—which depended on both sex and WAT depot—also support the different response of thermogenesis‐related parameters between overweight males and females.     

Sex-dependence of the metabolism of aldosterone in adrenalectomized and intact rats

The metabolism of aldosterone at physiological levels was shown to be significantly different in male and female rats: more aldosterone was metabolized in the male rats leading to significantly higher quantities of non-extractable polar derivatives of aldosterone in all subcellular fractions of kidney, particularly in the cytosol fractions. These results may be correlated with our findings in which it was shown that the physiological responses to aldosterone in both adrenalectomized and intact rats were significantly greater in the males than the females. These sex differences support the concept that the metabolism of aidosterone may be essential for the components of the physiological response to aldosterone to occur. Furthermore, the sex-dependence of the metabolism of aldosterone appears to be independent of the presence of the previously identified protein receptor-hormone complexes. At all dosages within the physiological range, no significant differences were observed between the extent of the ³H-aldosterone labeling of these binding proteins in the kidney cytosol fractions of male and female adrenalectomized rats, even at dosages where no physiological response was demonstrable in the females.     

Gonadectomy reveals sex differences in circadian rhythms and suprachiasmatic nucleus androgen receptors in mice

In mammals, it is well established that circadian rhythms in physiology and behavior, including the rhythmic secretion of hormones, are regulated by a brain clock located in the suprachiasmatic nucleus (SCN) of the hypothalamus. While SCN regulation of gonadal hormone secretion has been amply studied, the mechanisms whereby steroid hormones affect circadian functions are less well known. This is surprising considering substantial evidence that sex hormones affect many aspects of circadian responses, and that there are significant sex differences in rhythmicity. Our previous finding that "core" and "shell" regions of the SCN differ in their expression of clock genes prompted us to examine the possibility that steroid receptors are localized to a specific compartment of the brain clock, with the discovery that the androgen receptor (AR) is concentrated in the SCN core in male mice. In the present study, we compare AR expression in female and male mice using Western blots and immunochemistry. Both of these methods indicate that ARs are more highly expressed in males than in females; gonadectomy eliminates and androgen treatment restores these sex differences. At the behavioral level, gonadectomy produces a dramatic loss of the evening activity onset bout in males, but has no such effect in females. Treatment with testosterone, or with the non-aromatizable androgen dihydrotestosterone, restores male locomotor activity and eliminates sex differences in the behavioral response. The results indicate that androgenic hormones regulate circadian responses, and suggest an SCN site of action.     

Estrogen upregulates renal angiotensin II AT1 and AT2 receptors in the rat

We studied renal AT1 and AT2 receptors in male, female, ovariectomized and ovariectomized-estrogen-treated Wistar-Hanover and Wistar-Kyoto rats. AT1 receptors and AT1A receptor mRNA predominated, with no significant differences between males and females. AT2 receptor expression was restricted in female rats to the capsule, the transition zone between outer and inner medulla, the endothelium lining the papilla, and arcuate arteries and veins. There were no AT2 receptors in male rats, while male mice express substantial numbers of estrogen-dependent AT2 receptors. Arcuate arteries and veins expressed AT1B mRNA in males and females, and AT2 mRNA in females only. AT1 receptor and AT2 receptor expression were estrogen-dependent, with increases in AT1 and AT2 receptor expression after estrogen treatment in ovariectomized rats. Estrogen treatment increased prostaglandin E2 (PGE2) and cGMP concentrations in the renal medulla, and eNOS expression in cortical arteries. In rodents, expression of renal Angiotensin II receptor types is estrogen-dependent, with significant species, strain and area differences. Our results support an important role for AT2 receptors in the regulation of renal function and in the protective effects of estrogen in the kidney.     

Meta-analysis of sex differences in gene expression in schizophrenia

Schizophrenia is a severe psychiatric disorder which influences around 1 % of the worldwide population. Differences between male and female patients with schizophrenia have been noted. There is an earlier age of onset in males compared with females with this diagnosis, and in addition, there are differences in symptom profiles between the sexes. The underlying molecular mechanism of sex difference remains unclear. Here we present a comprehensive analysis to reveal the sex differences in gene expression in schizophrenia with stringent statistics criteria. We compiled a data set consisting of 89 male controls, 90 male schizophrenia patients, 35 female controls and 32 female schizophrenia patients from six independent studies of the prefrontal cortex (PFC) in postmortem brain. When we tested for a sex by diagnosis interaction on gene expression, 23 genes were up-regulated and 23 genes were down-regulated in the male group (q-value?<?0.05), several genes are related to energy metabolism, while 4 genes are located on sex chromosome. No genes were statistically significant in the female group when multiple testing correction were conducted (q-value <0.05), most likely due to the small sample size. Our protocol and results from the male group provide a starting point for identifying the underlying different mechanism between male and female schizophrenia patients.     

Sex differences and similarities in the neuroendocrine regulation of social behavior in an African cichlid fish

An individual's position in a social hierarchy profoundly affects behavior and physiology through interactions with community members, yet little is known about how the brain contributes to status differences between and within the social states or sexes. We aimed to determine sex-specific attributes of social status by comparing circulating sex steroid hormones and neural gene expression of sex steroid receptors in dominant and subordinate male and female Astatotilapia burtoni, a highly social African cichlid fish. We found that testosterone and 17β-estradiol levels are higher in males regardless of status and dominant individuals regardless of sex. Progesterone was found to be higher in dominant individuals regardless of sex. Based on pharmacological manipulations in males and females, progesterone appears to be a common mechanism for promoting courtship in dominant individuals. We also examined expression of androgen receptors, estrogen receptor α, and the progesterone receptor in five brain regions that are important for social behavior. Most of the differences in brain sex steroid receptor expression were due to sex rather than status. Our results suggest that the parvocellular preoptic area is a core region for mediating sex differences through androgen and estrogen receptor expression, whereas the progesterone receptor may mediate sex and status behaviors in the putative homologs of the nucleus accumbens and ventromedial hypothalamus. Overall our results suggest sex differences and similarities in the regulation of social dominance by gonadal hormones and their receptors in the brain.     

Comparison of gene expression in male and female mouse blastocysts revealed imprinting of the X-linked gene, Rhox5/Pem, at preimplantation stages

Mammalian male preimplantation embryos develop more quickly than females . Using enhanced green fluorescent protein (EGFP)-tagged X chromosomes to identify the sex of the embryos, we compared gene expression patterns between male and female mouse blastocysts by DNA microarray. We detected nearly 600 genes with statistically significant sex-linked expression; most differed by 2-fold or less. Of 11 genes showing greater than 2.5-fold differences, four were expressed exclusively or nearly exclusively sex dependently. Two genes (Dby and Eif2s3y) were mapped to the Y chromosome and were expressed in male blastocysts. The remaining two (Rhox5/Pem and Xist) were mapped to the X chromosome and were predominantly expressed in female blastocysts. Moreover, Rhox5/Pem was expressed predominantly from the paternally inherited X chromosome, indicating sex differences in early epigenetic gene regulation.     

Sexual Dimorphism in the Expression of Mitochondria-Related Genes in Rat Heart at Different Ages

Cardiovascular disease (CVD) is the leading cause of mortality worldwide. Moreover, sex and age are considered major risk factors in the development of CVDs. Mitochondria are vital for normal cardiac function, and regulation of mitochondrial structure and function may impact susceptibility to CVD. To identify potential role of mitochondria in sex-related differences in susceptibility to CVD, we analyzed the basal expression levels of mitochondria-related genes in the hearts of male and female rats. Whole genome expression profiling was performed in the hearts of young (8-week), adult (21-week), and old (78-week) male and female Fischer 344 rats and the expression of 670 unique genes related to various mitochondrial functions was analyzed. A significant (p<0.05) sexual dimorphism in expression levels of 46, 114, and 41 genes was observed in young, adult and old rats, respectively. Gene Ontology analysis revealed the influence of sex on various biological pathways related to cardiac energy metabolism at different ages. The expression of genes involved in fatty acid metabolism was significantly different between the sexes in young and adult rat hearts. Adult male rats also showed higher expression of genes associated with the pyruvate dehydrogenase complex compared to females. In young and adult hearts, sexual dimorphism was not noted in genes encoding oxidative phosphorylation. In old rats, however, a majority of genes involved in oxidative phosphorylation had higher expression in females compared to males. Such basal differences between the sexes in cardiac expression of genes associated with energy metabolism may indicate a likely involvement of mitochondria in susceptibility to CVDs. In addition, female rats showed lower expression levels of apoptotic genes in hearts compared to males at all ages, which may have implications for better preservation of cardiac mass in females than in males.     

Sexual dimorphism in the renin-angiotensin system in aging spontaneously hypertensive rats

In young adult spontaneously hypertensive rats (SHR), mean arterial pressure (MAP) is higher in males than in females and inhibition of the renin-angiotensin system (RAS) eliminates this sex difference. After cessation of estrous cycling in female SHR, MAP is similar to that in male SHR. The purpose of this study was to determine the role of the RAS in maintenance of hypertension in aging male and female SHR. At 16 mo of age, MAP was similar in male and female SHR (183+/-5 vs. 193+/-8 mmHg), and chronic losartan (40 mg.kg-1.day-1 po for 3 wk) reduced MAP by 52% (to 90+/-8 mmHg, P<0.05 vs. control) in males and 37% (to 123+/-11 mmHg, P<0.05 vs. control) in females (P<0.05, females vs. males). The effect of losartan on angiotensin type 1 (AT1) receptor blockade was similar: MAP responses to acute doses of ANG II (62.5-250 ng/kg) were blocked to a similar extent in losartan-treated males and females. F2-isoprostane excretion was reduced with losartan more in males than in females. There were no sex differences in plasma renin activity, plasma angiotensinogen or ANG II, or renal expression of AT1 receptors, angiotensin-converting enzyme, or renin. However, renal angiotensinogen mRNA and protein expression was higher in old males than females, whereas renal ANG II was higher in old females than males. The data show that, in aging SHR, when blood pressures are similar, there remains a sexual dimorphism in the response to AT1 receptor antagonism, and the differences may involve sex differences in mechanisms responsible for oxidative stress with aging.     

Mapping of sex hormone receptors and their modulators along the nephron of male and female mice

Renal functions are regulated by steroid sex hormones, but the exhaustive identification of their receptors along the nephron is still lacking. Here, we have localized all known nuclear or membrane-bound sex hormone receptors and some of their activators along the nephron of male and female mice. Almost all receptors are present in male and female kidney, some of them having very restricted localization. Only one gene tested among 11 (ARA54) exhibits a gender difference in the level of its expression. This first “renal map” of sex steroid receptor expression may serve as a pre-requisite for investigating the role of these hormones on kidney functions.     

Sex Differences in Peripheral Mu-Opioid Receptor Mediated Analgesia in Rat Orofacial Persistent Pain Model

Unilateral ligation of the tendon of anterior superficial part of rat masseter muscle (TASM) leads to long-lasting allodynia. Sex differences in peripheral mu-opioid receptor (MOR)-mediated analgesia under persistent myogenic pain are not well understood. In this study, we examined (1) whether locally applied MOR agonists attenuate persistent pain following TASM ligation in a sex dependent manner, (2) whether there are sex differences of MOR expression changes in rat trigeminal ganglia (TG). The effects of MOR agonist, D-Ala2, N–Me-Phe4, Gly5-ol]-Enkephalin acetate salt (DAMGO), were assessed 14 days after TASM ligation in male, female and orchidectomized (GDX) male rats. MOR mRNA and protein levels in TG 14 days following tendon ligation were also determined. The mechanical thresholds of the injured side were significantly decreased in both male and female rats, from 3 days to 28 days after TASM ligation. A10 μg DAMGO significantly attenuated allodynia in male rats. A 10-fold higher dose of DAMGO was required in female and GDX male rats to produce the level of anti- allodynia achieved in male rats. The level of MOR mRNA in TG from male rats was significantly greater 14 days after TASM ligation compared with the sham-operated male rats, but not from female and GDX male rats. After TASM ligation, males had significantly more MOR immunoreactivity in TG compared to sham-operated males. The MOR levels increased to 181.8% of the sham level in male rats receiving tendon injury. But there was no significant change in female rats receiving tendon injury compared to the sham female rats. Taken together, our data suggest that there were sex differences in the effects of peripheral MOR agonists between male and female rats under TASM ligation developing long-lasting pain condition, which is partly mediated by sex differences in the changes of MOR expressions and testosterone is an important factor in the regulation of MOR.