Reactive carbonyls from tobacco smoke increase arterial endothelial layer injury

We hypothesized that reactive carbonyls generated from smoke exposure cause increased arterial low-density lipoprotein (LDL) accumulation and endothelial layer permeability. In addition, we hypothesized that estrogen supplementation was protective against chronic environmental tobacco smoke (ETS) exposure to the artery wall. Quantitative fluorescence microscopy was used to determine artery injury after exposure. For our chronic studies, ovariectomized rats treated with subcutaneous placebo or 17beta-estradiol pellets were exposed to ETS or filtered air for 6 wk. ETS exposure increased carotid artery LDL accumulation more than fourfold compared with filtered air exposure, an effect largely mediated by increased permeability. No protective effect of estradiol was observed. Acute ETS exposure of a buffer solution containing LDL resulted in a more than sixfold increase in the highly reactive carbonyl glyoxal. Perfusion of this solution through carotid arteries resulted in a 105% increase in permeability. Moreover, perfusion of glyoxal alone caused a 50% increase in carotid artery permeability. This endothelial damage and changes in lipid accumulation may serve as an initiating event in atheroma formation in individuals exposed to ETS.     

Effect of ovariectomy at two periods of the year on LH and FSH basal concentrations and pituitary response to LHRH in the brown hare (Lepus europaeus)

In the brown hare, fertile mating takes place from the beginning of December to September. Seasonal variations of basal concentrations of LH and FSH, and pituitary response to a monthly i.v. injection of LHRH were studied in intact control females and in females ovariectomized during the seasonal anoestrus (OVX1) or during the breeding season (OVX2). In intact females, both basal and LHRH-stimulated LH levels showed an annual variation, with minimal values during anoestrus. During the breeding season, the LH response to LHRH exhibited a biphasic pattern. In contrast, there was no clear seasonal variation in basal and LHRH-stimulated FSH concentrations. After ovariectomy during anoestrus, basal LH remained low for 2 months and began to increase in December. After ovariectomy during the breeding season, LH basal concentrations increased within a few days after the operation. Thereafter, LH values remained high in both groups of females until September, and decreased significantly as in intact females. The pattern of LH release after LHRH remained monophasic in the two groups of ovariectomized females. In OVX1 females, the LH response increased as early as October, was maximum from December to April and decreased progressively until October. IN OVX2 females, the LH response decreased regularly after ovariectomy to a minimum in October. In the 2 groups of ovariectomized females, basal FSH concentrations and pituitary response to LHRH rose rapidly after ovariectomy and did not vary significantly thereafter. These results showed a direct central effect of season on the regulation of basal concentrations of LH, modulated by a negative feed-back of ovarian secretions during the breeding season. In intact hares, the enhanced LH response after LHRH during the breeding season was related to an acute positive effect of ovarian secretions. The regulation of FSH was less dependent on season and remained under a negative control of the ovary throughout the year.     

Chronic estradiol treatment attenuates stiffening, glycoxidation, and permeability in rat carotid arteries

Aging-related changes in vascular stiffening and permeability are associated with cardiovascular disease. We examined the interaction of estradiol on the aging process in vascular tissue from rats by assessing the changes in endothelial layer permeability, arterial compliance, and glycoxidative damage levels. We isolated carotid arteries from ovariectomized (OVX) rats that underwent 1 yr of estrogen treatment with subcutaneous pellets and a subsequent 1 mo of cessation of treatment. Endothelial layer permeability and arterial compliance were determined using quantitative fluorescence microscopy. Endothelial layer permeability was reduced with estradiol treatment (estrogen groups, 2.58 +/- 0.21 ng dextran x min(-1) x cm(-2) vs. nonestrogen groups, 4.01 +/- 0.30 ng dextran x min(-1) x cm(-2); P < 0.05). Additionally, arteries from animals treated with estradiol had an increased compliance index (estrogen groups, 82.9 +/- 3.8 mm2. Torr vs. nonestrogen groups, 69.3 +/- 3.2 mm2. Torr; P < 0.05). Estradiol treatment also reduced levels of pentosidine, which is a specific marker of glycoxidative damage (estrogen groups, 0.11 +/- 0.03 pmol pentosidine/nmol collagen vs. nonestrogen groups, 0.20 +/- 0.03 pmol pentosidine/nmol collagen; P < 0.05). These results indicate that estradiol has multiple chronic vasculoprotective effects on the artery wall to maintain normal vascular wall function.     

Influence of experimental diabetes on regulatory mechanisms of vascular response of rabbit carotid artery to acetylcholine

The purpose of this study was to analyse the influence of experimental diabetes on vascular response of rabbit carotid artery to acetylcholine (Ach). We compared the Ach-induced relaxant response of isolated arterial segments obtained from both control and diabetic animals. To assess the influence of the endothelium, this cell layer was mechanically removed in some of the arterial segments ("rubbed arteries") from each experimental group. Ach induced a concentration-related endothelium-mediated relaxation of carotid artery from control rabbits that was significantly higher with respect to that obtained in diabetic animals. Pre-treatment with N(G)-nitro-L-arginine (L-NA) induced a concentration-dependent inhibition of relaxant response to Ach, which was significantly higher in carotid arteries isolated from diabetic rabbits. Incubation of rubbed arteries with L-NA almost abolished the relaxant response to Ach in arterial segments from both control and diabetic animals. Indomethacin potentiated Ach-induced response of carotid arteries from control rabbits, without modifying that obtained in those from diabetic animals. Aminoguanidine did not significantly inhibit the relaxant action of Ach in arterial segments from either control or diabetic rabbits. These results suggest that diabetes impairs endothelial modulatory mechanisms of vascular response of rabbit carotid artery to Ach. This endothelial dysfunction is neither related with a lower release of nitric oxide (NO) or prostacyclin. Diabetes impairs the production of some arachidonic acid vasoconstrictor derivative. There has been observed an increased modulatory activity of NO, but this is not related with the expression of an inducible isoform of NO synthase.     

Does estradiol treatment normalize the hypothalamic-pituitary-adrenal axis in streptozotocin-induced ovariectomized diabetic female rats?

We recently found circulating corticosterone (CS) levels to be significantly lower in diabetic female rats as compared with proestrous control animals. This reduction in CS was correlated with the hypoestrogenic state of the diabetic female. It was the purpose of this study to evaluate basal and corticotropin releasing hormone (CRH)-stimulated CS secretion in ovariectomized (OVX) control (C) and streptozotocin-induced diabetic (D) rats given blank, 5 mcg and 20 mcg estradiol (E2) implants to determine if adrenal CS secretion in the diabetic is normalized by E2 treatment. After 3 weeks of diabetes, pituitary-adrenal function was assessed in rats from each group with a CRH stimulation test. The remaining rats were sacrificed for determination of CS, E2, testosterone and fructosamine in serum. Suppressed CS secretion in OVX female diabetic rats was partially restored with E2 therapy. Basal CS levels were significantly higher in 20 mcg E2 treated C and D rats compared with OVX rats. However, C rats had significantly higher basal CS compared with D rats in similarly E2 treated groups. The CS response to CRH stimulation was not different between OVX female diabetic and control rats. Estrogen enhanced the CS response to CRH stimulation in control animals but not in diabetic animals suggesting altered estrogen action at the pituitary level in diabetic animals.     

Diabetes abolishes the vascular protective effects of estrogen in female rats

Estrogen is known to exert a protective effect against cardiovascular disease. However, women with diabetes have three times the risk as compared with age-matched non-diabetic women. Our previous study on aortic rings of ovariectomized (OVX) female rats treated with 17-beta-estradiol (E2) demonstrated that the beneficial effect of estrogen is related to the basal release of NO from endothelial cells. In the present study, in order to understand why estrogen protection is abolished in diabetes, we tested vascular responses in OVX, streptozotocin-diabetic female rats and their non-diabetic controls receiving or not E2 replacement. Concentration-response curves to norepinephrine (NE) showed attenuation of the contractile response in E2-treated diabetic, with respect to non-diabetic preparations. This response was further impaired in diabetic, E2-deprived rats. The basal release of NO, as evaluated by concentration-related responses to N(G)-methyl-L-arginine acetate in NE-precontracted aortic rings, was found to be impaired in E2-treated diabetic rats, no further effect being induced by E2 deprivation. The endothelium-dependent relaxation produced by carbachol did not change between groups, whereas the relaxation produced by histamine was enhanced by both diabetes and E2 deprivation. However, E2 treatment counteracted the response to histamine only in preparations from non-diabetic animals. Finally, the relaxation induced by sodium nitroprusside, an endothelium-independent relaxant agent, was comparable between groups. These findings suggest that the lack of protective effects of estrogen in diabetes may be mainly ascribed to the failure of estrogen to reverse the impaired basal release of NO and the abnormal relaxation to histamine, which are observed in the aorta of diabetic rats.     

Effects of ovarian input on GnRH and LH secretion immediately postovulation in pony mares

The potential involvement of ovarian factors in regulating GnRH and LH postovulation was studied in ovarian intact (Group 1; n=3) and ovariectomized (OVX; Group 2; n=3) mares (OVX within 12 hr of ovulation). Blood samples were collected every 10 min for 6 hr from jugular vein (JV) and intercavernous sinus (ICS) during estrus and on Day 8 postovulation for LH and GnRH analysis. Additionally, JV samples were collected twice daily (12-hr intervals) for 30 days for LH and progesterone (P4) analysis. A significant treatment x day effect (P<0.0001) describes declining plasma LH concentrations in intact mares, and regression analysis indicated that response curves were not parallel (P<0.001). Plasma LH concentrations remained elevated in OVX mares. LH increased further in OVX mares by Day 8 post-OVX (P<0.06), reflecting the increased (P<0.07) LH episode amplitude. GnRH decreased from estrus to Day 8 in both groups reflecting an effect of sampling period (P<0.03). GnRH episode amplitude declined (P<0.08) from estrus (62.8+/-3.1 pg/mL) to Day 8 (46.3+/-3.1 pg/mL) in OVX mares, but not in control mares (intact estrus, 36.5+/-6.4; intact Day 8, 37.5+/-7.3; OVX estrus, 62.8+/-3.1; OVX Day 8, 46.3+/-3.1 pg/mL). In conclusion, we propose that postovulatory LH decline requires ovarian feedback in mares, and that OVX alters GnRH secretory dynamics such that LH concentrations does not decline postovulation and, in fact, is further elevated with time after OVX.     

Morphologic and histochemical study of blood capillaries in boar testes: effects of abdominal cryptorchidism

BACKGROUND: Few data exist about the features of testicular microvasculature under normal and pathologic conditions. METHODS: The morphology and lectin affinity of testicular capillaries were examined in healthy boars and in unilateral and bilateral abdominal cryptorchid boars. RESULTS: The capillaries of scrotal testes contained a) the endothelial layer formed by two cells, b) the basal lamina constituted by collagen fibers and glycoconjugates with fucosyl, galactosyl, glucosyl, and neuraminic acid residues, and c) the pericyte layer formed by a single cell. These components participated in substrate exchange between blood and testicular tissue. The abdominal testes showed increased numbers of capillaries, which could exhibit a mature appearance, but also angiogenic or degenerative patterns. Angiogenesis was manifested in interstitial capillaries and was characterized by a) proliferation of endothelial cells, b) decreased thickness and decreased content of collagen fibers and glycoconjugates in the basal lamina, and c) lack of pericytes. Degenerative capillaries lay in association with seminiferous tubules and showed a) pyknotic endothelial cells; b) thickening, collagenization, and altered glycoconjugate content in the basal lamina; and c) increased development of pericytes. The angiogenesis of interstitial capillaries resulted in high vascular permeability, and the degeneration of intertubular capillaries led to defective substrate exchange between blood and seminiferous tubules. CONCLUSIONS: Unilateral abdominal cryptorchidism did not alter the morphology and function of capillaries in the scrotal testis. Unilateral and bilateral abdominal cryptorchidism resulted in increased numbers and abnormal morphology and function of capillaries in abdominal testes. The proliferation of interstitial capillaries correlated with the immaturity of Leydig cells, and the degeneration of intertubular capillaries correlated with the thickening of the lamina propria.     

Chemotaxis of granulocytes across bovine pulmonary artery intimal explants without endothelial cell injury

Emigration of granulocytes from vessel lumen to a site of injury is a hallmark of acute inflammation but whether this migration is necessarily associated with vascular damage is not clear. To follow the structural changes associated with granulocyte migration across an intact endothelial cell layer and to assess changes in vascular permeability, an in vitro technique was developed in which intimal explants were stripped from bovine pulmonary artery and mounted in chemotaxis chambers. All explants studied had granulocytes and trace amounts of 3H-water, 14C-sucrose and 125I-albumin in the upper well of the chambers. Experimental explants had zymosan-activated plasma in the lower well and control explants had either serum in the lower well or zymosan-activated plasma in the upper well. Explants were incubated at 37 degrees C for periods from 15 min to 3 hr. When the chemoattractant was added to the lower well, granulocytes migrated into the explants. Transmission and scanning electron microscopy showed an orderly sequence of granulocyte--endothelial interactions throughout which the two cell types maintained close opposition--granulocyte adherence to and exploration of the endothelial surface; penetration and migration through the interendothelial cell junction; reapposition and reformation of the luminal 'tight' junctions and finally passage of granulocytes through the endothelial basal lamina. After 60 min incubation, the majority of granulocytes seen in each section was through the endothelial cell layer and after 2 hr, they were through the basal lamina. Structural evidence of granulocyte or endothelial cell damage was not found at any of the times examined, neither was there any demonstrable increase in intimal permeability. In control explants, granulocyte migration was strikingly less frequent at 2 hr (approximately 10% of that seen towards the chemoattractant). Thus, granulocyte migration across an endothelial cell layer towards a chemoattractant is not necessarily associated with structural evidence of endothelial cell injury or increased vascular permeability.     

The effects of different exercise modes for preventing endothelial dysfunction of arteries and bone loss in ovariectomized rats

[Purpose]

Several epidemiological studies have demonstrated that there are positive correlations between vascular disorders and bone loss in postmenopausal women. The aim of the present study was to examine the effect of different types of exercise (e.g., climbing and swimming) for preventing endothelial dysfunction of arteries and bone loss in ovariectomized rats.

[Methods]

Twenty Sprague-Dawley female rats were randomly divided into three groups: ovariectomy (OVX) plus treatment with vitamin D₃ and nicotine (VDN) (control rats [Con], n = 7), which is an animal model for endothelial dysfunction and bone loss; voluntary climbing resistance exercise with OVX plus VDN (climbing rats [Clim], n = 6), and swimming exercise with OVX plus VDN (swimming rats [Swim], n = 7). The period of exercise training was 8 weeks.

[Results]

The endothelin-1 (ET-1) protein levels were significantly lower in the Clim and Swim groups than in the Con. The endothelial nitric oxide synthase protein levels were significantly higher in the Swim group than in the Con, but they did not differ between the Clim and Con groups. The cortical bone mineral density in the tibia and breaking energy of the femur were significantly higher in the Clim group than in the Con, but this positive effect was not seen in the Swim group.

[Conclusion]

Voluntary climbing exercise decreased arterial ET-1 protein levels and prevented bone loss in a postmenopause-model rat combining OVX and VDN. Conversely, swimming suppressed endothelial dysfunction of the arteries but did not prevent bone loss. Thus, the type of exercise should be cautiously chosen for enhancing vascular function and bone status, especially in females after menopause.     

Estrogen status and skeletal muscle recovery from disuse atrophy.

Although estrogen loss can alter skeletal muscle recovery from disuse, the specific components of muscle regrowth that are estrogen sensitive have not been described. The primary purpose of this study was to determine the components of skeletal muscle mass recovery that are biological targets of estrogen. Intact, ovariectomized (OVX), and ovariectomized with 17beta-estradiol replacement (OVX+E2) female rats were subjected to hindlimb suspension for 10 days and then returned to normal cage ambulation for the duration of recovery. Soleus muscle mass returned to control levels by day 7 of recovery in the intact animals, whereas OVX soleus mass did not recover until day 14. Intact rats recovered soleus mean myofiber cross-sectional area (CSA) by day 14 of recovery, whereas the OVX soleus remained decreased (42%) at day 14. OVX mean fiber CSA did return to control levels by day 28 of recovery. The OVX+E2 treatment group recovered mean CSA at day 14, as in the intact animals. Myofibers demonstrating central nuclei were increased at day 14 in the OVX group, but not in intact or OVX+E2 animals. The percent noncontractile tissue was also increased 29% in OVX muscle at day 14, but not in either intact or OVX+E2 groups. In addition, collagen 1a mRNA was increased 45% in OVX muscle at day 14 of recovery. These results suggest that myofiber growth, myofiber regeneration, and extracellular matrix remodeling are estrogen-sensitive components of soleus muscle mass recovery from disuse atrophy.     

Effects of D2 receptor agonist and antagonist on behavior of intact and ovariectomized rats

The involvement of D2 dopaminergic receptors in behavioral responses during ovary cycle was assessed in adult intact female rats and ovariectomized (OVX) female rats. Quinperole (0.1 mg/kg), D2 receptor agonist and sulpiride (10.0 mg/kg), D2 receptor antagonist were injected chronically to adult intact and ovariectomized (OVX) female rats either separately or in combination with 17beta-estradiol (0.5 microg) within 14 days. Behavior of these animals was assessed in the "open field" test, whereas passive avoidance performance served as a model of learning. In intact rats, the passive avoidance performance was observed only in metestrous and diestrous. Chronic quinperole administration to intact females resulted in the appearance of the passive avoidance performance in proestrous and estrous, as distinct from the control animals. The passive avoidance performance was not reproduced in OVX rats. Quinperole per se or in combination with 17beta-estradiol completely restored the passive avoidance performance in OVX rats. Moreover, quinperole or sulpiride administration to OVX rats increased horizontal locomotor activity, exploratory behavior, and grooming behavior.     

Basal lamina of avian ovarian follicle: influence on morphology of granulosa cells in-vitro

Experiments were conducted to determine the influence of basal lamina on the morphology of ovarian granulosa cells in vitro. Pure and intact basal lamina was isolated from the large preovulatory follicles of the chicken ovary and designated basal lamina of avian ovarian follicle (BLAOF). Examination of the isolated basal lamina with electron microscope revealed an ultrastructure that is similar to that of basal lamina in the intact ovarian follicle. Pieces of the intact basal lamina were attached to the bottom of 32 mm culture dishes (BLAOF-coated dishes) in which differentiated granulosa cells isolated from the largest preovulatory follicle or undifferentiated granulosa cells isolated from immature small yellow chicken ovarian follicles were cultured; uncoated dishes served as controls. Granulosa cells incubated on intact basal lamina assumed spherical shape, whereas granulosa cells incubated directly on plastic in control dishes became highly flattened. Interestingly, granulosa cells that attached to plastic close to BLAOF (in BLAOF-containing dishes) became rounded. The storage of BLAOF-coated culture dishes at 4°C for 2 years had no apparent effect on its ability of the matrix material to induce changes in granulosa cell shape. Some components of the basal lamina could be solubilized with guanidine–HCl alone (fraction 1; 90–95% of total protein in BLAOF) with the remaining components solubilized with β-mercaptoethanol containing guanidine–HCl (fraction 2; 5–10% of total protein in BLAOF). Differentiated and undifferentiated chicken granulosa cells became rounded when incubated in fraction 1-pre-coated wells; whereas those incubated directly on plastic in control wells were flattened. Similarly, when fraction 1 of solubilized basal lamina was added as liquid to incubation mixture, it caused both differentiated and undifferentiated granulosa cells to assume spherical shapes. The storage of fraction 1-coated culture dishes at 4°C for 12 or more months had no apparent effect on its ability to influence granulosa cell shape. Fraction 1-induced changes in granulosa cell shape were similar to those observed for complete and intact basal lamina (BLAOF). These findings demonstrate that intact homologous basal lamina (BLAOF) or its solubilized (fluidized) form can induce normal (in vivo) morphology in granulosa cells. It is suggested that BLAOF or its solubilized form can be used to culture cells in experiments designed to examine the influence of the natural basal lamina microenvironment on cellular behavior and function.     

Lipogenesis in arterial wall and vascular smooth muscular cells: regulation and abnormalities in insulin-resistance

Background

Vascular smooth muscular cells (VSMC) express lipogenic genes. Therefore in situ lipogenesis could provide fatty acids for triglycerides synthesis and cholesterol esterification and contribute to lipid accumulation in arterial wall with aging and during atheroma.

Methods

We investigated expression of lipogenic genes in human and rat arterial walls, its regulation in cultured VSMC and determined if it is modified during insulin-resistance and diabetes, situations with increased risk for atheroma.

Results

Zucker obese (ZO) and diabetic (ZDF) rats accumulated more triglycerides in their aortas than their respective control rats, and this triglycerides content increased with age in ZDF and control rats. However the expression in aortas of lipogenic genes, or of genes involved in fatty acids uptake, was not higher in ZDF and ZO rats and did not increase with age. Expression of lipogenesis-related genes was not increased in human arterial wall (carotid endarterectomy) of diabetic compared to non-diabetic patients. In vitro, glucose and adipogenic medium (ADM) stimulated moderately the expression and activity of lipogenesis in VSMC from control rats. LXR agonists, but not PXR agonist, stimulated also lipogenesis in VSMC but not in arterial wall in vivo. Lipogenic genes expression was lower in VSMC from ZO rats and not stimulated by glucose or ADM.

Conclusion

Lipogenic genes are expressed in arterial wall and VSMC; this expression is stimulated (VSMC) by glucose, ADM and LXR agonists. During insulin-resistance and diabetes, this expression is not increased and resists to the actions of glucose and ADM. It is unlikely that this metabolic pathway contribute to lipid accumulation of arterial wall during insulin-resistance and diabetes and thus to the increased risk of atheroma observed in these situations.     

Differential effects of 17beta-estradiol and raloxifene on VSMC phenotype and expression of osteoblast-associated proteins

Several studies demonstrate an association between osteoporosis and arterial calcific disease, both of which being common in elderly women. Estradiol and raloxifene, a selective estrogen receptor modulator, prevent bone loss in postmenopausal women. Little is known regarding how these agents affect arterial calcification. The aim of this study was to determine whether or not 17beta-estradiol and raloxifene reduced vascular smooth muscle cell (VSMC) differentiation and expression of bone-associated proteins during phosphate-induced calcification in vitro. Aortic VSMC were cultured from adult, gonadally intact, and ovariectomized (OVX) female pigs. Calcifying medium was added, and cells were treated with solvent (control), 17beta-estradiol (E(2)), or raloxifene. Extent of calcification and phenotypic expression of bone-associated proteins [matrix gla protein (MGP), osteoprotegerin (OPG), and bone sialoprotein (BSP)] were examined at 3-day intervals over 2 wk. Calcium content increased in all groups but was greater in VSMC derived from intact compared with OVX animals. E(2) reduced calcification and preserved a contractile phenotype. Expression of OPG significantly decreased with time; this decrease was significantly greater in VSMC derived from OVX compared with gonadally intact pigs. E(2) and raloxifene preserved expression of OPG only in VSMC from intact pigs. Expression of MGP increased significantly with time and was not affected by E(2) or raloxifene treatments. E(2) treatment significantly inhibited synthesis of BSP in cells from both groups. In conclusion, E(2) slows differentiation of VSMC induced by excess phosphate. Effectiveness of raloxifene to preserve expression of bone cell-associated proteins depends on the hormonal status of the tissue donor.     

Influence of secretin and L-NAME on vascular permeability in the coronary circulation of intact and diabetic rats

The permeability in the intact and diabetic rat coronary circulation after administration of secretin (3.0 micromol/kg i.v.), an inhibitor of NOS (nitric oxide synthase), and L-NAME (N(G)-nitro-L-arginine-methyl ester hydrochloride) (1 mg/kg i.v.), and both substances given together, were studied. To measure protein extravasation Evans blue dye was used as a marker of vascular permeability. The vascular permeability of the left ventricle did not differ in intact and diabetic rats. In the diabetes state increased permeability of atria was observed. Administration of secretin did not influence the coronary vascular permeability in either the intact or the diabetic rats. L-NAME increased the atria permeability and did not change left ventricle permeability. In diabetes, injection of L-NAME caused a decrease in the permeability in both the atria and left ventricle. In intact rats secretin diminished the L-NAME effect in the atria. In diabetic rats co-administration of secretin+L-NAME increased the permeability of the atria and left ventricle, but L-NAME administered alone decreased them. Secretin modified the effect of L-NAME on coronary permeability in intact and diabetic rats.     

Insulinotropic effect of ovarian steroid hormones in streptozotocin diabetic female mice

To investigate the protective effect of ovarian sex steroids against streptozotocin diabetes, groups of ovariectomized streptozotocin diabetic female mice were treated orally with estradiol-17 beta (5 and 500 ug/kg/day) or progesterone (1 mg/kg/day) for 10 weeks. Streptozotocin produced a more severe hyperglycaemia and a greater fall in plasma insulin concentrations in ovariectomized mice than intact female mice. The estradiol-17 beta and progesterone treatments reduced both the severity of the hyperglycaemia and the fall in plasma insulin. Loss of pancreatic insulin after streptozotocin administration was reduced in intact mice and in mice treated with estradiol-17 beta. These observations suggest that ovarian sex steroids reduce the severity of streptozotocin diabetes at least partly by countering the cytotoxic effect of the drug on the islet B-cells, thereby reducing the fall in pancreatic and plasma insulin.     

Estradiol increases somatomedin-C concentrations in adolescent rhesus macaques (Macaca mulatta)

Estradiol (E2) may enhance somatomedin-C (Sm-C) secretion during puberty in female rhesus monkeys. The present study evaluated the importance of age and acute changes in E2 on Sm-C secretion. Intact (INT) females at their first ovulation (age 3.5 yr; n = 6) had higher levels of Sm-C across the ovulatory cycle than did intact adults (ADT) (n = 5). Levels of Sm-C were similar for both groups during the follicular and luteal phases despite higher follicular phase levels of E2. Young, ovariectomized, E2-treated (E2OVX) females (age 3.5 yr; n = 5; E2 = 50 pg/ml) had higher basal levels of Sm-C than did either age-matched ovariectomized (OVX) females (n = 3), ovariectomized adults (OXA), or E2-treated ovariectomized adults (E2A) (E2 = 100 pg/ml). When ovariectomized groups were given E2 to induce ovulatory increases, no changes in serum Sm-C occurred. Comparisons among age-mates revealed that basal levels of Sm-C were similar between INT and E2OVX, yet these levels were higher than those for OVX. Sm-C levels were similar among all adult groups. Serum growth hormone (GH) was highest in E2OVX, next highest in INT and OVX, and lowest in all adults. Higher Sm-C levels in young animals are, thus, related to these age differences in GH concentrations and are further enhanced by basal levels of E2 and not by acute changes in this steroid. Low Sm-C secretion in adults is associated with low GH levels. Thus, the facilitory effect of basal E2 on Sm-C release is observed during conditions when basal GH levels are elevated, a situation normally limited to adolescence.     

Improvement of vascular function by acute and chronic treatment with the GPR30 agonist G1 in experimental diabetes mellitus

The G-protein coupled estrogen receptor 30 (GPR30) is a seven-transmembrane domain receptor that mediates rapid estrogen responses in a wide variety of cell types. This receptor is highly expressed in the cardiovascular system, and exerts vasodilatory effects. The objective of the present study was to investigate the effects of GPR30 on vascular responsiveness in diabetic ovariectomized (OVX) rats and elucidate the possible mechanism involved. The roles of GPR30 were evaluated in the thoracic aorta and cultured endothelial cells. The GPR30 agonist G1 induced a dose-dependent vasodilation in the thoracic aorta of the diabetic OVX rats, which was partially attenuated by the nitric oxide synthase (NOS) inhibitor, nitro-L-arginine methylester (L-NAME) and the GPR30-selective antagonist G15. Dose-dependent vasoconstrictive responses to phenylephrine were attenuated significantly in the rings of the thoracic aorta following the acute G1 administration in the diabetic OVX rats. This effect of G1 was abolished partially by L-NAME and G15. The acute administration of G1 increased significantly the eNOS activity and the concentration of NO in the endothelial cells exposed to high glucose. G1 treatment induced an enhanced endothelium-dependent relaxation to acetylcholine (Ach) in the diabetic OVX rats. Further examination revealed that G1 induced vasodilation in the diabetic OVX rats by increasing the phosphorylation of eNOS. These findings provide preliminary evidence that GPR30 activation leads to eNOS activation, as well as vasodilation, to a certain degree and has beneficial effects on vascular function in diabetic OVX rats.     

The blood-brain barrier in a reptile, Anolis carolinensis

An electron microscopic study was made of the ultrastructure and permeability of the capillaries in the cerebral hemispheres of the lizard, Anolis carolinensis. The brain of Anolis is vascularized by a loop-type pattern consisting exclusively of arteriovenous capillary loops. The ultrastructure of the endothelium and the arrangement of the various layers from the capillary lumen to the central nervous tissue is similar to that of mammals. The endothelial cells form a continuous layer around the lumen and are joined by tight interendothelial junctions. The basal lamina of the endothelium is also continuous and encloses pericyte processes. The cells of the nervous tissue rest directly on the basal lamina of the capillary and are separated from each other by a 200 Å space. Intravenously injected horseradish peroxidase (MW 40,000) and ferritin (MW 500,000) were used to study the permeability of the capillaries. The entry of horseradish peroxidase and ferritin into the intercellular spaces of the brain is restricted by the tightness of the interendothelial junctions. No vesicular transport of either tracer occurs; however, ferritin does enter the endothelial cells in vacuoles. No tracer molecules are present in the basal lamina, pericytes, or nervous tissue. The different responses of the endothelial cell to the tracers used in this study suggest that endocytotic activities of endothelial cells involve different processes. Vacuoles formed by marginal folds, vacuoles formed by endothelial surface projections or deep invaginations of the plasma membrane, 600–800 Å vesicles, and coated vesicles all seem to differ in the nature of the substances which they endocytose.