Changes in the estrogen/testosterone balance at menopause may negatively influence the development of diabetic kidney disease. Furthermore, recent studies suggest that changes in hormone levels during perimenopause may influence disease development. Injection of 4-vinylcyclohexene diepoxide (VCD) in B 6C3F1 mice induces gradual ovarian failure, preserving both the perimenopausal (peri-ovarian failure) and menopausal (post-ovarian failure) periods. To address the impact of the transition into menopause on the development of diabetes and diabetic kidney damage, we used streptozotocin (STZ)-induced diabetes in the VCD model of menopause. After 6 wk of STZ-induced diabetes, blood glucose was significantly increased in post-ovarian failure (post-OF) diabetic mice compared with cycling diabetic mice. In peri-ovarian failure (peri-OF) diabetic mice, blood glucose levels trended higher but were not significantly different from cycling diabetic mice, suggesting a continuum of worsening blood glucose across the menopausal transition. Cell proliferation, an early marker of damage in the kidney, was increased in post-OF diabetic mice compared with cycling diabetic mice, as measured by PCNA immunohistochemistry. In post-OF diabetic mice, mRNA abundance of early growth response-1 (Egr-1), collagen-4␣1, and matrix metalloproteinase-9 were increased and 3␤-hydroxysteroid dehydrogenase 4 (3␤-HSD4) and transforming growth factor-␤2 (TGF-␤2) were decreased compared with cycling diabetic mice. In peri-OF diabetic mice, mRNA abundance of Egr-1 and 3␤-HSD4 were increased, and TGF-␤2 was decreased compared with cycling diabetic mice. This study highlights the importance and utility of the VCD model of menopause, as it provides a physiologically relevant system for determining the impact of the menopausal transition on diabetes and diabetic kidney damage. diabetes; 3␤-HSD4; perimenopause; real-time PCR; estrogen DIABETES IS ONE OF THE MOST prevalent and costly diseases afflicting developed countries, with estimates placing the current global cost of diabetes at $150 billion a year (1). Approximately one-third of all diabetics die of end-stage renal disease (33) due to progressive renal damage and hypertension.17␤-Estradiol is considered protective against the development and progression of many diseases, including cardiovascular (9) and renal disease (34). Premenopausal women have slower rates of progression of nondiabetic renal disease than age-matched men (23), a difference that seems to disappear after menopause (3). The impact of estrogen on diabetic renal disease is less clear, however. Several studies demonstrate a decreased incidence of diabetic renal disease in women, but others have found no difference between men and women (34).The 5-10 years preceding menopause is termed perimenopause, and during this time estrogen levels fluctuate, with periods of low estrogen interspersed with periods of very high estrogen (31). The periods of low estrogen become more frequent as a woman approaches menopause until circulating levels of 17␤...

Oxygen increases ductus arteriosus smooth muscle cytosolic calcium via release of calcium from inositol triphosphate-sensitive stores. Am J Physiol Lung Cell Mol Physiol 288: L917-L923, 2005. First published February 4, 2005 doi:10.1152 doi:10. /ajplung.00403.2004 shunts away from the lungs via the ductus arteriosus (DA) and the foramen ovale. After birth, the DA closes concomitant with increased oxygen tension. The present experimental series tests the hypothesis that oxygen directly increases DA smooth muscle cell (SMC) cytosolic calcium ([Ca 2ϩ ]i) through inactivation of a K ϩ channel, membrane depolarization, and entry of extracellular calcium. To test the hypothesis, DA SMC were isolated from late-gestation fetal lambs and grown to subconfluence in primary culture in low oxygen tension (25 Torr). DA SMC were loaded with the calcium-sensitive fluorophore fura-2 under low oxygen tension conditions and studied using microfluorimetry while oxygen tension was acutely increased (120 Torr). An acute increase in oxygen tension progressively increased DA SMC [Ca 2ϩ ]i by 11.7 Ϯ 1.4% over 40 min. ]i via release of calcium from intracellular calcium stores, and subsequent entry of extracellular calcium, and 2) potentiates the effect of contractile agonists. Prolonged patency of the DA may result from disordered intracellular calcium homeostasis. oxygen sensing; pulmonary hypertension potassium channels; vascular biology IN THE FETAL PULMONARY CIRCULATION oxygen tension and nitric oxide production are low (13). Fetal pulmonary blood flow is limited, and blood pressure is high. Blood is shunted away from the lungs through the foramen ovale and the ductus arteriosus (DA), a structure that connects the pulmonary artery to the aorta in utero. At birth, pulmonary blood flow immediately increases by 8-to 10-fold, and pulmonary blood pressure decreases by 50% within the initial 24 h of life (24). An acute increase in oxygen tension causes perinatal pulmonary vasodilation (3, 26), even while it results in constriction of the DA (1, 5). Initial closure of the DA in response to an increase in PO 2 is caused by vessel constriction (14), whereas long-term closure is accomplished through cell migration, apoptosis (25), and cell proliferation (4, 15, 18).In the fetal state, elevation of pulmonary vascular tone and patency of the DA is of critical importance. How pulmonary vascular tone remains elevated while DA tone remains low despite being adjacent vascular structures is unknown. In the fetal pulmonary circulation, endothelin, a powerful vasoconstrictor (29) produced by the endothelial cell (20), plays a key role in maintaining elevated tone, as endothelin inhibition results in sustained fetal pulmonary vasodilation without affecting either pulmonary or aortic pressures (17). This observation suggests that endothelin possesses site-specific properties in the fetal circulation. Whether the low oxygen tension environment of the normal fetus enables endothelin to modulate perinatal pulmonary vascular tone without affecting DA ...

. Chronic intrauterine pulmonary hypertension selectively modifies pulmonary artery smooth muscle cell gene expression. Am J Physiol Lung Cell Mol Physiol 290: L426 -L432, 2006; doi:10.1152/ajplung.00281.2005.-Pulmonary artery smooth muscle cell (PASMC) relaxation at birth results from an increase in cytosolic cGMP, cGMP-dependent and kinase-mediated activation of the Ca 2ϩ -sensitive K ϩ channel (KCa), and closure of voltage-operated Ca 2ϩ channels (VOCC). How chronic intrauterine pulmonary hypertension compromises perinatal pulmonary vasodilation remains unknown. We tested the hypothesis that chronic intrauterine pulmonary hypertension selectively modifies gene expression to mitigate perinatal pulmonary vasodilation mediated by the cGMP kinase-KCa-VOCC pathway. PASMC were isolated from late-gestation fetal lambs that had undergone either ligation of the ductus arteriosus (hypertensive) or sham operation (control) at 127 days of gestation and were maintained under either hypoxic (ϳ25 Torr) or normoxic (ϳ120 Torr) conditions in primary culture. We studied mRNA levels for cGMP kinase I␣ (PKG-1␣), the ␣-chain of VOCC (Cav1.2), and the ␣-subunit of the KCa channel. Compared with control PASMC, hypertensive PASMC had decreased VOCC, KCa, and PKG-1␣ expression. In response to sustained normoxia, expression of VOCC and KCa channel decreased and expression of PKG-1␣ increased. In contrast, sustained normoxia had no effect on PKG-1␣ levels and an attenuated effect on VOCC and KCa channel expression in hypertensive PASMC. Protein expression of PKG-1␣ was consistent with the mRNA data. We conclude that chronic intrauterine pulmonary hypertension decreases PKG expression and mitigates the genetic effects of sustained normoxia on pulmonary vasodilation, because gene expression remains compromised even after sustained exposure to normoxia. fetal; oxygen sensing; nitric oxide IN UTERO, oxygen tension is low and pulmonary vascular resistance is greater than systemic vascular resistance (36). At birth, the pulmonary circulation undergoes an unprecedented and unparalleled transition, given that pulmonary blood flow increases 8-to 10-fold and arterial pressure decreases by 50% within 24 h, concomitant with an increase in oxygen tension, establishment of an air-liquid interface, and rhythmic distention of the lung (9,15,44).Recent data suggest that activation of the large-conductance Ca 2ϩ -sensitive K ϩ channel (K Ca , also known as BK Ca or MaxiK) plays a critically important role in mediating the response to perinatal pulmonary vasodilator stimuli such as oxygen (11) ] i ) and promoting vasodilation (20). PKG also may directly activate the K Ca channel by phosphorylation of its ␣-chain (42). Activation of the channel results in membrane hyperpolarization, closure of voltage-gated Ca 2ϩ channels, a decrease in cytosolic Ca 2ϩ , and vasodilation. Recent data indicate the existence of several different isoforms of cGMP-dependent protein kinase. In vascular smooth muscle cells, relaxation is contingent on activation of the type I ...

Resnik ER, Keck M, Sukovich DJ, Herron JM, Cornfield DN. Chronic intrauterine pulmonary hypertension increases capacitative calcium entry in fetal pulmonary artery smooth muscle cells. Am J Physiol Lung Cell Mol Physiol 292: L953-L959, 2007. First published December 8, 2006 doi:10.1152/ajplung.00327.2006.-Oxygen causes perinatal pulmonary dilatation. Although fetal pulmonary artery smooth muscle cells (PA SMC) normally respond to an acute increase in oxygen (O2) tension with a decrease in cytosolic calcium ([Ca 2ϩ ]i), an acute increase in O2 tension has no net effect on [Ca 2ϩ ]i in PA SMC derived from lambs with chronic intrauterine pulmonary hypertension (PHTN). The present experimental series tests the hypothesis that an acute increase in O2 tension decreases capacitative calcium entry (CCE) in normal, but not hypertensive, fetal PA SMC. PA SMC were isolated from late-gestation fetal lambs after either ligation of the ductus arteriosus (PHTN) or sham (control) operation at 127 days gestation. PA SMC were isolated from the distal PA (Ն4th generation) and maintained under hypoxic conditions (ϳ25 Torr) in primary culture. After fura 2 loading, apparent [Ca 2ϩ ]i in PA SMC was determined as the ratio of 340-to 380-nm fluorescence intensity. Under both hypoxic and normoxic conditions, cyclopiazonic acid (CPA) increased [Ca 2ϩ ]i more in PHTN than in control PA SMC. CCE was determined in PA SMC under hypoxic and normoxic conditions, after superfusion with zero extracellular Ca 2ϩ and intracellular store depletion with CPA, followed by superfusion with Ca 2ϩ -containing solution, in the presence of the voltage-operated calcium channel blockade. CCE was increased in PHTN compared with control PA SMC under conditions of both acute and sustained normoxia. Transient receptor potential channel gene expression was greater in control compared with PHTN PA SMC. PHTN may compromise perinatal pulmonary vasodilation, in part, by modulating PA SMC CCE. cytosolic calcium; oxygen sensing; persistent pulmonary hypertension of the newborn IN UTERO, OXYGEN TENSION IS low and pulmonary vascular resistance is greater than systemic vascular resistance (11). At birth, the pulmonary circulation normally undergoes a dramatic transition as pulmonary blood flow increases 8-to 10-fold, and arterial pressure decreases concomitant with an increase in oxygen tension, establishment of an air-liquid interface, and rhythmic distention of the lung (5, 24).Fetal stress can compromise neonatal pulmonary vasodilation. Intrauterine hypoxemia, infection, hyperglycemia, and drug exposure can result in an incomplete response to perinatal vasodilator stimuli and persistently elevated pulmonary arterial blood pressure, limited pulmonary blood flow, and severe central hypoxemia. Persistent pulmonary hypertension of the newborn (PPHN) is characterized by extrapulmonary shunting of blood across the patent foreman ovale and through the patent ductus arteriosus (DA), severe hypoxemia, high levels of circulating endothelin-1, decreased nitric oxide prod...

Aquaporin (AQP) 1 null mice have a defect in the renal concentrating gradient because of their inability to generate a hyperosmotic medullary interstitium. To determine the effect of vasopressin on renal medullary gene expression, in the absence of high local osmolarity, we infused 1-deamino-8-d-arginine vasopressin (dDAVP), a V(2) receptor (V(2)R)-specific agonist, in AQP1 null mice for 7 days. cDNA microarray analysis was performed on the renal medullary tissue, and 5,140 genes of the possible 12,000 genes on the array were included in the analysis. In the renal medulla of AQP1 null mice, 245 transcripts were identified as increased by dDAVP infusion and 200 transcripts as decreased (1.5-fold or more). Quantitative real-time PCR measurements confirmed the increases seen for cyclin D1, early growth response gene 1, and activating transcription factor 3, genes associated with changes in cell cycle/growth. Changes in mRNA expression were correlated with changes in protein expression by semiquantitative immunoblotting; cyclin D1 and ATF3 were increased significantly in abundance following dDAVP infusion in the renal medulla of AQP1 null mice (161 and 461%, respectively). A significant increase in proliferation of medullary collecting ducts cells, following V(2)R activation, was identified by proliferating cell nuclear antigen immunohistochemistry; colocalization studies with AQP2 indicated that the increase in proliferation was primarily observed in principal cells of the inner medullary collecting duct (IMCD). V(2)R activation, via dDAVP, increased AQP2 and AQP3 protein abundance in the cortical collecting ducts of AQP1 null mice. However, V(2)R activation did not increase AQP2 protein abundance in the IMCD of AQP1 null mice.