Estrogen responsiveness of renal calbindin-D28k gene expression in rat kidney

In women, calcium excretion in the urine rises after menopause and falls with estrogen replacement therapy. The amount of calcium lost in the urine following estrogen therapy is less than should occur based on changes in serum calcium and the amount of calcium filtered by the kidney. This suggests there may be a direct effect of estrogen therapy to increase renal calcium reabsorption. Calbindin D_28k is a putative calcium ferry protein located in the distal renal tubules which has been shown to increase transcellular calcium transport. We proposed that estrogen loss after menopause may diminish gene expression of renal calbindin D_28k and subsequently diminish renal calcium reabsorption. We used the ovariectomized rat model of estrogen deficiency to investigate changes at the messenger RNA level of calbindin D_28k in ovariectomized rats (OVX), sham ovariectomized rats (S-OVX), and estrogen treated ovariectomized rats (E-OVX). We have demonstrated that ovariectomy in rats diminishes the gene expression of renal calbindin D_28k. The mRNA levels were approximately three times lower in OVX rats than S-OVX rats. Administration of 17β estradiol to OVX rats produced a significant increase in mRNA level to greater than the S-OVX rats by 4 h. Measurement of serum 1,25 dihydroxyvitamin D₃ showed lower level in OVX rats than S-OVX rats but no significant change in E-OVX animals. In conclusion, our results indicate that estrogen increases renal calbindin D_28k mRNA levels, by a mechanism independent of changes in 1,25 dihydroxyvitamin D₃. This may result in increased expression of calbindin D_28k protein which may have a role in reducing renal calcium excretion. J. Cell. Biochem. 65:340–348. © 1997 Wiley-Liss, Inc.     

Altered Kidney CYP2C and Cyclooxygenase‐2 Levels Are Associated with Obesity‐Related Albuminuria

Objective: To determine cytochrome P450 (CYP450) and cyclooxygenase (COX) expression and metabolite regulation and renal damage in the early stages of obesity‐related hypertension and diabetes. Research Methods and Procedures: Obese and lean Zucker rats at 10 to 12 weeks of age were studied. Blood pressure was measured in the conscious state using radiotelemetry. Blood glucose levels and body weight were measured periodically. Protein expression of CYP450 and COX enzymes in the kidney cortex, renal microvessels, and glomeruli was studied. The levels of CYP450 and COX metabolites in urine were measured, and urinary albumin excretion, an indicator of kidney damage, was measured. Results: Body weight and blood glucose averaged 432 ± 20 grams and 105 ± 5 mg/dl, respectively, in obese Zucker rats as compared with 320 ± 8 grams and 91 ± 5 mg/dl, respectively, in age‐matched 10‐ to 12‐week‐old lean Zucker rats. Renal microvascular CYP4A and COX‐2 protein levels were increased 2.3‐ and 17.0‐fold, respectively, in obese Zucker rats. The protein expression of CYP2C11 and CYP2C23 was decreased 2.0‐fold in renal microvessels isolated from obese Zucker rats when compared with lean Zucker rats. The urinary excretion rate of thromboxane B₂ was increased significantly in obese Zucker as compared with lean Zucker rats (22.0 ± 1.8 vs. 13.4 ± 1.0 ng/d). Urinary albumin excretion, an index of kidney damage, was increased in the obese Zucker rat at this early age. Discussion: These results suggest that increased CYP4A and COX‐2 protein levels and decreased CYP2C11 and CYP2C23 protein levels occur in association with microalbuminuria during the onset of obesity‐related hypertension and type 2 diabetes.     

PPAR-alpha activator fenofibrate increases renal CYP-derived eicosanoid synthesis and improves endothelial dilator function in obese Zucker rats

Previous studies have shown that the synthesis of renal cytochrome P-450 (CYP)-derived eicosanoids is downregulated in genetic or high-fat diet-induced obese rats. Experiments were designed to determine whether fenofibrate, a peroxisome proliferator-activated receptor (PPAR)-alpha agonist, would induce renal eicosanoid synthesis and improve endothelial function in obese Zucker rats. Administration of fenofibrate (150 mg.kg(-1).day(-1) for 4 wk) significantly reduced plasma insulin, triglyceride, and total cholesterol levels in obese Zucker rats. CYP2C11 and CYP2C23 proteins were downregulated in renal vessels of obese Zucker rats. Consequently, renal vascular epoxygenase activity decreased by 15% in obese Zucker rats compared with lean controls. Chronic fenofibrate treatment significantly increased renal cortical and vascular CYP2C11 and CYP2C23 protein levels in obese Zucker rats, whereas it had no effect on epoxygenase protein and activity in lean Zucker rats. Renal cortical and vascular epoxygenase activities were consequently increased by 54% and 18%, respectively, in fenofibrate-treated obese rats. In addition, acetylcholine (1 microM)-induced vasodilation was significantly reduced in obese Zucker kidneys (37% +/- 11%) compared with lean controls (67% +/- 9%). Chronic fenofibrate administration increased afferent arteriolar responses to 1 microM of acetylcholine in obese Zucker rats (69% +/- 4%). Inhibition of the epoxygenase pathway with 6-(2-propargyloxyphenyl)hexanoic acid attenuated afferent arteriolar diameter responses to acetylcholine to a greater extent in lean compared with obese Zucker rats. These results demonstrate that the PPAR-alpha agonist fenofibrate increased renal CYP-derived eicosanoids and restored endothelial dilator function in obese Zucker rats.     

Chronic administration of adrenomedullin attenuates the hypertension and increases renal nitric oxide synthase in Dahl salt-sensitive rats

Adrenomedullin reduces systemic blood pressure and increases urinary sodium excretion partly through the release of nitric oxide. We hypothesized that chronic adrenomedullin infusion ameliorates salt-sensitive hypertension and increases the expression of renal nitric oxide synthase (NOS) in Dahl salt-sensitive (DS) rats, because the reduced renal NOS expression promotes salt sensitivity. DS rats and Dahl salt-resistant (DR) rats were fed a high sodium diet (8.0% NaCl) for 3 weeks. The high sodium diet resulted in an increase in blood pressure and a reduction of urinary sodium excretion in association with increased renal adrenomedullin concentrations and decreased expression of renal neuronal NOS (nNOS) and renal medullary endothelial NOS (eNOS) in DS rats compared with DR rats. Chronic adrenomedullin infusion partly inhibited the increase of blood pressure and proteinuria in association with a restoration of renal nNOS and medullary eNOS expression in DS rats under the high sodium diet. The immunohistochemical analysis revealed that the restored renal nNOS expression induced by chronic adrenomedullin infusion may reflect the restoration of nNOS expression in the macula densa and inner medullary collecting duct. These results suggest that adrenomedullin infusion has beneficial effects on this hypertension probably in part through restored renal NOS expression in DS rats.     

Expression of cytochrome P-450 4 enzymes in the kidney and liver: Regulation by PPAR and species-difference between rat and human

Members of the cytochrome P-450 4 (CYP4) family catalyze the ω-hydroxylation of fatty acids, and some of them have the PPAR response element in the promoter area of the genes. The localization of CYP4A and PPAR isoforms and the effect of PPAR agonists on CYP4A protein level and activity were determined in rat kidney and liver. Immunoblot analysis showed that CYP4A was expressed in the liver and proximal tubule, with lower expression in the preglomerular microvessel, glomerulus and thick ascending limb (TAL), but the expression was not detected in the collecting duct. PPARα was expressed in the liver, proximal tubule and TAL. PPARγ was expressed in the collecting duct, with lower expression in the TAL, but no expression in the proximal tubule and liver. The PPARα agonist clofibrate induced CYP4A protein levels and activity in the renal cortex and liver. The PPARγ agonist pioglitazone did not modulate them in these tissues. The localization of CYP4A and CYP4F were further determined in human kidney and liver by immunohistochemical technique. Immunostainings for CYP4A and CYP4F were observed in the hepatocytes of the liver lobule and the proximal tubules, with lower stainings in the TALs and collecting ducts, but no staining in the glomeruli or renal vasculatures. These results indicate that the inducibility of CYP4A by PPAR agonists in the rat tissues correlates with the expression of the respective PPAR isoforms, and that the localization of CYP4 in the kidney has a species-difference between rat and human.     

Rosiglitazone reduces blood pressure in female Dahl salt-sensitive rats

Postmenopausal women (PMW) are at greater risk for salt-sensitive hypertension and insulin resistance than premenopausal women. Peroxisome-proliferator-activated receptor-gamma (PPARγ) agonists reduce blood pressure (BP) and insulin resistance in humans. As in PMW, ovariectomy (OVX) increases salt sensitivity of BP and body weight in Dahl salt-sensitive (DS) rats. This study addressed whether rosiglitazone (ROSI), a PPARγ agonist, attenuates salt-sensitive hypertension in intact (INT) and OVX DS rats, and if so, whether insulin resistance, nitric oxide (NO), oxidative stress, and/or renal inflammation were contributing mediators. Telemetric BP was similar in OVX and INT on low salt diet (0.3% NaCl), but was higher in OVX than INT on high salt (8% NaCl). ROSI reduced BP in OVX and INT on both low and high salt diet, but only attenuated salt sensitivity of BP in OVX. Nitrate/nitrite excretion (NO_x; index of NO) was similar in INT and OVX on low salt diet, and ROSI increased NO_x in both groups. High salt diet increased NO_x in all groups but ROSI only increased NO_x in OVX rats. OVX females exhibited insulin resistance, increases in body weight, plasma leptin, cholesterol, numbers of renal cortical macrophages, and renal MCP-1 and osteopontin mRNA expression compared to INT. ROSI reduced cholesterol and macrophage infiltration in OVX, but not INT. In summary, PPARγ activation reduces BP in INT and OVX females, but attenuates the salt sensitivity of BP in OVX only, likely due to increases in NO and in part to reductions in renal resident macrophages and inflammation.     

Epoxyeicosatrienoic Acid inhibition alters renal hemodynamics during pregnancy

In this study we examined the expression of cytochrome P450 (CYP) 2C and CYP2J isoforms in renal proximal tubules and microvessels isolated from rats at different stages of pregnancy. We also selectively inhibited epoxyeicosatrienoic acid (EET) production by the administration of N-methanesulfonyl-6-(2-proparyloxyphenyl)hexanamide (MSPPOH 20 mg/kg/day iv) to rats during Days 14-17 of gestation and to age-matched virgin rats and determined the consequent effects on renal function. Western blot analysis showed that CYP2C11, CYP2C23, and CYP2J2 expression was significantly increased in the renal microvessels of pregnant rats on Day 12 of gestation. In the proximal tubules, CYP2C23 expression was significantly increased throughout pregnancy, while the expression of CYP2C11 was increased in early and late pregnancy and the expression of CYP2J2 was increased in middle and late pregnancy. MSPPOH treatment significantly increased pregnant rats' mean arterial pressure, renal vascular resistance, and sodium balance but significantly decreased renal blood flow, glomerular filtration rate, and urinary sodium excretion, as well as fetal pups' body weight and length. In contrast, MSPPOH treatment had no effect on renal hemodynamics or urinary sodium excretion in age-matched virgin rats. In pregnant rats, MSPPOH treatment also caused selective inhibition of renal cortical EET production and significantly decreased the expression of CYP2C11, CYP2C23, and CYP2J2 in the renal cortex, renal microvessels, and proximal tubules. These results suggest that upregulation of renal vascular and tubular EETs contributes to the control of blood pressure and renal function during pregnancy.     

Renal sodium retention in cirrhotic rats depends on glucocorticoid-mediated activation of mineralocorticoid receptor due to decreased renal 11beta-HSD-2 activity

Downregulation of the renal glucocorticoid-metabolizing enzyme 11beta-hydroxysteroid dehydrogenase type 2 (11beta-HSD-2) during liver cirrhosis may allow activation of the mineralocorticoid receptor (MR) by glucocorticoids and contribute to sodium retention. We tested this hypothesis in male Wistar rats with decompensated liver cirrhosis and ascites 7 wk after bile duct ligation (BDL). Renal 11beta-HSD-2 mRNA, protein, and activity were significantly decreased in decompensated rats. The urinary Na(+)/K(+) ratio was reduced by 40%. Renal epithelial sodium channel (ENaC) mRNA and immunostaining were only slightly affected. Complete metabolic studies, including fecal excretion, showed that the BDL rats had avid renal sodium retention. Treatment of the BDL rats with dexamethasone suppressed endogenous glucocorticoid production, normalized total sodium balance and renal sodium excretion, and reduced ascites formation to the same degree as direct inhibition of MR with K-canrenoate. Total potassium balance was negative in the BDL rats, whereas renal potassium excretion was unchanged. In the distal colon, expression of ENaC was increased in BDL rats. Fecal potassium excretion was increased in cirrhotic rats, and this was corrected by treatment with K-canrenoate but not dexamethasone. We conclude that development of sodium retention and decompensation in cirrhotic rats is associated with downregulation of renal 11beta-HSD-2 activity and inappropriate activation of renal sodium reabsorption by endogenous glucocorticoids. In addition, the overall potassium loss in the BDL model is due to increased fecal potassium excretion, which is associated with upregulation of ENaC in distal colon.     

Lithospermic acid B isolated from Salvia miltiorrhiza ameliorates ischemia/reperfusion-induced renal injury in rats

The present study was designed to examine whether lithospermic acid B (LSB) isolated from Salvia miltiorrhiza has an ameliorative effect on renal functional parameters in association with the expression of aquaporin 2 (AQP 2) and Na,K-ATPase in the ischemia-reperfusion induced acute renal failure (ARF) rats. LSB showed strong antioxidant activity against production of reactive oxygen species (ROS), ROS-induced hemolysis, and production of lipid peroxide in a dose-dependent manner. Polyuria caused by down-regulation of renal AQP 2 in the ischemia-reperfusion induced ARF rats was partially restored by administration of LSB (40 mg/kg, i.p.), restoring expression of AQP 2, in renal inner and outer medulla. The expression of Na,K-ATPase alpha1 subunit in outer medulla of the ARF rats was also restored in the ARF rats by administration of LSB, while beta1 subunit level was not altered. The renal functional parameters including creatinine clearance, urinary sodium excretion, urinary osmolality, and solute-free reabsorption were also partially restored in ischemia-ARF rats by administration of LSB. Histological study also showed that renal damages in the ARF rats were abrogated by administration of LSB. Taken together, these data indicate that LSB ameliorates renal defects in rats with ischemia-reperfusion induced ARF, most likely via scavenging of ROS.     

Overexpression of HIF Prolyl‐Hydoxylase‐2 transgene in the renal medulla induced a salt sensitive hypertension

Renal medullary hypoxia‐inducible factor (HIF)‐1α and its target genes, such as haem oxygenase and nitric oxide synthase, have been indicated to play an important role in the regulation of sodium excretion and blood pressure. HIF prolyl hydroxylase domain‐containing proteins (PHDs) are major enzymes to promote the degradation of HIF‐1α. We recently reported that high salt intake suppressed the renal medullary PHD2 expression and thereby activated HIF‐1α‐mediated gene regulation in the renal medulla in response to high salt. To further define the functional role of renal medullary PHD2 in the regulation of renal adaptation to high salt intake and the longer term control of blood pressure, we transfected PHD2 expression plasmids into the renal medulla in uninephrectomized rats and determined its effects on pressure natriuresis, sodium excretion after salt overloading and the long‐term control of arterial pressure after high salt challenge. It was shown that overexpression of PHD2 transgene increased PHD2 levels and decreased HIF‐1α levels in the renal medulla, which blunted pressure natriuresis, attenuated sodium excretion, promoted sodium retention and produced salt sensitive hypertension after high salt challenge compared with rats treated with control plasmids. There was no blood pressure change in PHD2‐treated rats that were maintained in low salt diet. These results suggested that renal medullary PHD2 is an important regulator in renal adaptation to high salt intake and a deficiency in PHD2‐mediated molecular adaptation in response to high salt intake in the renal medulla may represent a pathogenic mechanism producing salt sensitive hypertension.     

Retroperitoneal white adipose tissue mitochondrial function and adiponectin expression in response to ovariectomy and 17β-estradiol replacement

Sexual dimorphism has been previously found both in mitochondrial biogenesis and function and in adiponectin expression of retroperitoneal WAT. However, little is known about the E2 effects on WAT mitochondrial function. Accordingly, the aim of this study was to examine in greater depth the role of estrogens in sexual dimorphism. This was accomplished by studying the effects of ovariectomy and E2 replacement on retroperitoneal WAT mitochondrial function. Fourteen-week-old female and ovariectomized (OVX) female Wistar rats were used in this study. The ovariectomy was performed at 5 weeks of age and at 10 weeks of age OVX rats were divided into two experimental groups: OVX, and OVX treated with 17β-estradiol (E2) (OVX+E2). Subcutaneous injections of E2 (10 μg/kg/48 h) were administered to the OVX+E2 rats for 4 weeks previous to the sacrifice whereas OVX rats were treated only with the vehicle. Levels of the main markers for mitochondrial biogenesis and function and those representatives of the antioxidant defense system and insulin sensitivity were determined. Additionally, the mRNA levels of the α and β estrogen receptors and of some adipocyte differentiation markers were studied. Our results indicate that retroperitoneal WAT was able to adapt itself to ovariectomy without any changes in mitochondrial function markers or for the adiponectin levels. However, E2 supplementation led to an unexpected decrease in: TFAM protein levels, in LPL, PPARγ and adiponectin gene expression and in the systemic HMW adiponectin levels. This decrease is probably due to the down-regulation of the ERα mRNA expression to avoid an over-stimulation by E2.     

Ovariectomy worsens secondary hyperparathyroidism in mature rats during low-Ca diet

Estrogen deficiency impairs intestinal Ca absorption and induces bone loss, but its effects on the vitamin D-endocrine system are unclear. In the present study, calciotropic hormones levels, renal vitamin D metabolism, 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]-dependent intestinal calcium absorption, and bone properties in 3-mo-old sham-operated (sham) or ovariectomized (OVX) rats fed either a normal-Ca (NCD; 0.6% Ca, 0.65% P) or a low-Ca (LCD; 0.1% Ca, 0.65% P) diet for 2 wk were determined. LCD increased serum 1,25(OH)2D3 levels in both sham and OVX rats. Serum parathyroid hormone [PTH(1-84)] levels were highest in OVX rats fed LCD. Renal 25-hydroxyvitamin D1alpha-hydroxylase (1-OHase) protein expression was induced in both sham and OVX rats during LCD, while renal 1-OHase mRNA expression was highest in OVX rats fed LCD. Renal vitamin D receptor (VDR) and mRNA expressions in rats were induced by ovariectomy in rats fed NCD but suppressed by ovariectomy in rats fed LCD. The induction of intestinal calcium transporter-1 and calbindin-D9k mRNA expressions by LCD were not altered by ovariectomy. As expected, bone Ca content, cancellous bone mineral density, and bone strength index in proximal metaphysis of rat tibia were reduced by both ovariectomy and LCD (P<0.05) as analyzed by two-way ANOVA. Taken together, the data demonstrate that ovariectomy alters the responses of circulating PTH levels, renal 1-OHase mRNA expression, and renal VDR expression to LCD. These results suggest that estrogen is necessary for the full adaptive response to LCD mediated by both PTH and 1,25(OH)2D3.     

17beta-estradiol deficiency reduces potassium excretion in an angiotensin type 1 receptor-dependent manner

This study examined the effects of ovariectomy (OVX) and 17beta-estradiol (E(2)) replacement (OVX + E(2)) on renal function in Sprague-Dawley rats. OVX caused a 40% decrease in the fractional excretion of potassium (FE(K(+))) that was prevented by E(2) replacement [Sham, 24.2 +/- 2.9%; OVX, 14.5 +/- 2.1% (P < 0.05 vs. OVX + E(2)); and OVX + E(2), 26.2 +/- 2.7%; n = 7-11] and that corresponded to significant increases in plasma potassium [(in mmol/l): Sham, 3.15 +/- 0.087; OVX, 3.42 +/- 0.048 (P < 0.05 vs. OVX + E(2)); and OVX + E(2), 3.19 +/- 0.11; n = 7-11]. No effects of OVX were detected on plasma levels of sodium and aldosterone. Angiotensin II type 1 receptor (AT(1)R) densities in ovariectomized rats were 1.4-fold and 1.3-fold higher in glomerular [maximum binding capacity (B(max); in fmol/mg protein): Sham, 482 +/- 21; OVX, 666 +/- 20 (P < 0.05 vs. OVX + E(2)); and OVX + E(2), 504 +/- 26; n = 7-11] and proximal tubular [B(max) (in fmol/mg protein): Sham, 721 +/- 16; OVX, 741 +/- 24 (P < 0.05 vs. OVX + E(2)); and OVX + E(2), 569 +/- 23; n = 7-11] membranes compared with E(2) replete animals, respectively. Both the angiotensin-converting enzyme inhibitor captopril and the AT(1)R antagonist losartan prevented the OVX-induced decrease in the FE(K(+)) and the increase in renal AT(1)R densities, suggesting that E(2) deficiency reduces potassium excretion in an ANG II/AT(1)R-dependent manner. These findings may have implications for renal function in postmenopausal women as well as contribute to the reasons underlying the age-induced increase in susceptibility to hypertension-associated disease in women.     

Ovariectomy is protective against renal injury in the high-salt-fed older mRen2. Lewis rat

Studies in experimental animals and younger women suggest a protective role for estrogen; however, clinical trials may not substantiate this effect in older females. Therefore, the present study assessed the outcome of ovariectomy in older mRen2. Lewis rats subjected to a high-salt diet for 4 wk. Intact or ovariectomized (OVX, 15 wk of age) mRen2. Lewis rats were aged to 60 wk and then placed on a high-salt (HS, 8% sodium chloride) diet for 4 wk. Systolic blood pressures were similar between groups [OVX 169 +/- 6 vs. Intact 182 +/- 7 mmHg; P = 0.22] after the 4-wk diet; however, proteinuria [OVX 0.8 +/- 0.2 vs. Intact 11.5 +/- 2.6 mg/mg creatinine; P < 0.002, n = 6], renal interstitial fibrosis, glomerular sclerosis, and tubular casts were lower in OVX vs. Intact rats. Kidney injury molecule-1 mRNA, a marker of tubular damage, was 53% lower in the OVX HS group. Independent from blood pressure, OVX HS rats exhibited significantly lower cardiac (24%) and renal (32%) hypertrophy as well as lower C-reactive protein (28%). Circulating insulin-like growth factor-I (IGF-I) levels were not different between the Intact and OVX groups; however, renal cortical IGF-I mRNA and protein were attenuated in OVX rats [P < 0.05, n = 6]. We conclude that ovariectomy in the older female mRen2. Lewis rat conveys protection against salt-dependent increase in renal injury.     

Endothelin-1 contributes to the sexual differences in renal damage in DOCA-salt rats

We investigated whether gender differences in renal damage in DOCA-salt hypertension are associated with effects of ovarian hormones and/or endothelin-1 (ET-1). Renal injuries and renal pre-pro-ET-1 mRNA expression were enhanced in male and female ovariectomized (OVX) DOCA rats versus female DOCA rats. Treatment with estrogen plus progesterone or progesterone, but not estrogen alone, attenuated renal damage and pre-pro-ET-1 mRNA expression in OVX DOCA rats. The ETA antagonist BMS182874 greatly ameliorated renal damage in male and OVX DOCA rats. In conclusion, the ovarian hormones have a protective role on the renal structural alterations in female DOCA rats by modulating effects of ET-1, via ETA receptors.     

The effects of long-term exposure to extremely low-frequency magnetic fields on bone formation in ovariectomized rats

The effects of long‐term extremely low‐frequency magnetic field (ELF‐MF) exposure on bone formation and biochemical markers were investigated in ovariectomized rats. Sixty mature female Sprague–Dawley rats were randomly divided into four different groups (n = 15): (i) unexposed control (CTL); (ii) ovariectomized only (OVX); (iii) non‐ovariectomized, exposed (SHAM + ELF‐MF); and (iv) ovariectomized, exposed (OVX + ELF‐MF). The third and fourth groups were exposed to 1.5 mT ELF‐MF for 4 h a day for 6 months. Bone mineral density (BMD) was determined using dual energy X‐ray absorption (DEXA) measurements. The formation and resorption of bone were evaluated using bone‐specific alkaline phosphatase (BAP), osteocalcin, osteoprotogerin, and N‐telopeptide. After 6 months of ELF‐MF therapy, BMD values were significantly lower in the OVX group and higher in the OVX + ELF‐MF and SHAM + ELF‐MF groups than they were before therapy (P < 0.001). Although there was no significant difference in BMD values among the groups before therapy, the BMD values increased significantly after 6 months in the OVX + ELF‐MF and SHAM + ELF‐MF groups and were reduced in the OVX group compared to the CTL group (P < 0.001). The concentrations of BAP, osteocalcin, osteoprotogerin, and N‐telopeptide in the three experimental groups also changed in a significant way compared to the CTL group. The results of the present study suggest that osteoporosis can be inhibited by ELF‐MF stimulation treatments. It was also concluded that ELF‐MF may be useful in the prevention of osteoporosis in ovariectomized rats. Bioelectromagnetics 33:543–549, 2012. © 2012 Wiley Periodicals, Inc.     

Estrogen rapidly modulates mustard oil-induced visceral hypersensitivity in conscious female rats: A role of CREB phosphorylation in spinal dorsal horn neurons

This study investigated the effect of sex hormones on mustard oil (MO)-induced visceral hypersensitivity in female rats and analyzed possible involved signaling pathways. Female rats, either intact or ovariectomized (OVX), were prepared for abdominal muscle electromyography in response to colorectal distension after intracolonic instillation of MO. The effect of MO intracolonic sensitization was evaluated in intact rats, OVX rats, and OVX rats pretreated with a single injection of 17beta-estradiol (E), progesterone (P), E+P, or vehicle. cAMP-responsive element-binding protein (CREB) and phosphorylated CREB (pCREB) were detected in the superficial dorsal horn of L6 and S1 in MO or mineral oil-treated OVX rats with/without colorectal distension and estrogen replacement. The distal colorectum was removed for histological evaluation of inflammatory severity in MO-treated intact or OVX rats. The MO-treated rats had significantly higher visceromotor reflex than controls (enhanced visceral hypersensitivity), whereas OVX eliminated this hypersensitivity. After a single injection of E or E+P, the rats rapidly restored MO-induced visceral hypersensitivity within 2 h. Estrogen also rapidly induced a dose-dependent increase in pCREB expression in the superficial dorsal horn neurons in MO-treated, but not mineral oil-treated, OVX rats. The present study suggests that estrogen can rapidly modulate visceral hypersensitivity induced by MO intracolonic instillation in conscious female rats, which may involve spinal activation of the cAMP response element-mediated gene induction pathway.     

Pulsed electromagnetic fields prevent osteoporosis in an ovariectomized female rat model: a prostaglandin E2-associated process

With the use of Helmholtz coils and pulsed electromagnetic field (PEMF) stimulators to generate uniform time varying electromagnetic fields, the effects of extremely low frequency electromagnetic fields on osteoporosis and serum prostaglandin E(2) (PGE(2)) concentration were investigated in bilaterally ovariectomized rats. Thirty-five 3 month old female Sprague-Dawley rats were randomly divided into five different groups: intact (INT), ovariectomy (OVX), aspirin treated (ASP), PEMF stimulation (PEMF + OVX), and PEMF stimulation with aspirin (PEMF + ASP) groups. All rats were subjected to bilateral ovariectomy except those in INT group. Histomorphometric analyses showed that PEMF stimulation augmented and restored proximal tibial metaphyseal trabecular bone mass (increased hard tissue percentage, bone volume percentage, and trabecular number) and architecture (increased trabecular perimeter, trabecular thickness, and decreased trabecular separation) in both PEMF + OVX and PEMF + ASP. Trabecular bone mass of PEMF + OVX rats after PEMF stimulation for 30 days was restored to levels of age matched INT rats. PEMF exposure also attenuated the higher serum PGE(2) concentrations of OVX rats and restored it to levels of INT rats. These experiments demonstrated that extremely low intensity, low frequency, single pulse electromagnetic fields significantly suppressed the trabecular bone loss and restored the trabecular bone structure in bilateral ovariectomized rats. We, therefore, conclude that PEMF may be useful in the prevention of osteoporosis resulting from ovariectomy and that PGE(2) might relate to these preventive effects.     

Influence of gender and estrous cycle on plasma and renal catecholamine levels in rats

Several studies have demonstrated that gonadal hormones show significant effects on the brain and signaling pathways of effector organs/cells that respond to neurotransmitters. Since little information is available concerning the impact of male and female gonadal hormones on the renal and peripheral sympathetic system, the objective of this study was to further assess whether and how the renal content and plasma concentration of catecholamines are influenced by gender and the estrous cycle in rats. To achieve this, males Wistar rats were divided into 4 groups: (i) sham (i.e., control), (ii) gonadectomized, (iii) gonadectomized and nandrolone decanoate replacement at physiological levels or (iv) gonadectomized and nandrolone decanoate replacement at high levels. Female Wistar rats were divided into 6 groups: (i) ovariectomized (OVX), (ii) estrogen replacement at physiological levels and (iii) estrogen replacement at at high levels, (iv) progesterone replacement at physiological levels and (v) progesterone replacement at at high levels, and (vi) sham. The sham group was subdivided into four subgroups: (i) proestrus, (ii) estrus, (iii) metaestrus, and (iv) diestrus. Ten days after surgery, the animals were sacrificed and their plasma and renal catecholamine levels measured for intergroup comparisons. Gonadectomy led to an increase in the plasma catecholamine concentration in females, as well as in the renal catecholamine content of both male and female rats. Gonadectomized males also showed a lower level of plasma catecholamine than the controls. The urinary flow, and the fractional excretion of sodium and chloride were significantly increased in gonadectomized males and in the OVX group when compared with their respective sham groups.