Vivian C. Yang

EFFECTS OF ZINC SUPPLEMENTATION ON THE PLASMA GLUCOSE LEVEL AND INSULIN ACTIVITY IN GENETICALLY OBESE (OB/OB) MICE

The effects of zinc supplementation (20 mM ZnCl2 from the drinking water for eight weeks) on plasma glucose and insulin levels, as well as its in vitro effect on lipogenesis and lipolysis in adipocytes were studied in genetically obese (ob/ob) mice and their lean controls (+/?). Zinc supplementation reduced the fasting plasma glucose levels in both obese and lean mice by 21 and 25%, respectively (p < 0.05). Fasting plasma insulin levels were significantly decreased by 42% in obese mice after zinc treatment. In obese mice, zinc supplementation also attenuated the glycemic response by 34% after the glucose load. The insulin-like effect of zinc on lipogenesis in adipocytes was significantly increased by 80% in lean mice. However, the increment of 74% on lipogenesis in obese mice was observed only when the zinc plus insulin treatment was given. This study reveals that zinc supplementation alleviated the hyperglycemia of ob/ob mice, which may be related to its effect on the enhancement of insulin activity.

Identification and expression of scavenger receptor SR-BI in endothelial cells and smooth muscle cells of rat aorta in vitro and in vivo

In this study, we used immunoelectron microscopy to investigate the subcellular localization of scavenger receptor class B type I (SR-BI) in the arterial walls of rats. The expression of SR-BI in cultured endothelial and smooth muscle cells of rat aorta after exposure to high-density lipoprotein (HDL) was also investigated by immunofluorescence microscopy and immunoblotting analysis. A peptide containing residues 495-509 from mouse SR-BI (mSR-BI) plus an NH2-terminal cysteine was coupled to hemocyanin to generate mSR-BI antiserum in rabbits. Reactivity of antiserum against the synthetic peptides was confirmed with an enzyme-linked immunosorbent assay (ELISA). The results showed that SR-BI was specifically localized on the surface of the endothelial cells and smooth muscle cells. SR-BI was also observed in the cytoplasm of smooth muscle cells. Immunoblotting analysis indicated that SR-BI was expressed in the cell membrane. The levels of SR-BI increased gradually from 1 to 3 h and decreased at 24 and 48 h after cholesterol-loaded cells were incubated in the culture medium containing HDL. We conclude that SR-BI, a functional receptor for HDL, is expressed in the aortic endothelial cells as well as in smooth muscle cells. This receptor also responds to the presence of HDL in the culture medium.

Uroflowmetry: its current clinical utility for women

Uroflowmetry, the simple, non-invasive measurement of urine flow over time during micturition, has a long and interesting history, clear definitions, a clear purpose in screening for voiding difficulty and, most importantly, technical accuracy. Data interpretation is currently limiting its clinical utility, despite appropriate analysis being available in long-standing existing research. The main clinically important numerical parameters are the maximum and average urine flow rates and the voided volume. Urine flow rates are strongly dependent on voided volume. Reference to established (Liverpool) nomograms will most accurately correct for this dependency. Nomograms will also optimise the validation of uroflowmetry data and the accurate assessment of its normality, compared with fixed urine flow rates and “cutoffs” for voided volume. Abnormally slow urine flow (under the 10th centile Liverpool Nomograms) is the most clinically significant abnormality. Repeat uroflowmetry, concomitant post-void residual measurement and voiding cystometry studies are appropriate options for evaluating any abnormal uroflowmetry.

Caveolin-1 Expression Is Associated with Plaque Formation in Hypercholesterolemic Rabbits

Caveolin-1, the major structural protein of caveolae, is present in several cell types known to play a role in the development of atherosclerosis. In this study, the distribution and expression of caveolin-1 in the arterial walls were studied in hypercholesterolemic rabbits. Immunohistochemical results indicated that the staining intensity of caveolin-1 reached a high level in the arterial intima at 5 weeks after high-cholesterol-diet treatment and decreased to a very low level at 8 weeks when atheromatous plaques appeared. Western blot analysis showed that in rabbits fed a high-cholesterol diet for 5 weeks, the expression of caveolin-1 reached its highest level and then decreased from 8 to 12 weeks. The proliferative activity of smooth muscle cells (SMCs) decreased to the lowest level at 5 weeks and then increased at 8 and 12 weeks. Nitric oxide synthase activity gradually decreased in animals fed a high-cholesterol diet throughout the experiment. These studies demonstrate that the change in abundance of caveolin-1 is associated with SMC proliferation in the formation of atheromatous plaque after hypercholesterolemia insult.

Oxidized LDL inhibits vascular endothelial cell morphogenesis in culture.

SummaryHuman umbilical cord vein endothelial cells can be induced to undergo morphogenesis (tube formation) by phorbol ester (TPA) when cultured on or in three-dimensional collagen gels. Induction of morphogenesis by TPA is accompanied by increased activity of the collagenase gene transcription factors, ETS1 and AP1, and the elaboration of collagenase by the endothelial cells. In the present study, we used endothelial cell elongation as a measure of morphogenesis and showed that oxidized low density lipoprotein (oxLDL) inhibited endothelial cell migration in monolayer cultures and TPA-induced morphogenesis in collagen gels in a dose-dependent manner. Moreover, the inhibition was positively correlated with the extent of LDL oxidation. In contrast, native LDL stimulated cell migration and TPA-induced morphogenesis under the same culture conditions. However, in the absence of TPA, LDL showed no effect on EC morphogenesis. Further studies showed that inhibition of TPA-induced endothelial cell morphogenesis by oxLDL is correlated with suppression of the protein kinase C (PKC) and ETS1/AP1 activities. The results indicated that the inhibition of endothelial cell morphogenesis by oxLDL is probably mediated through inhibition of the TPA-activated PKC pathway and its subsequent suppression of the ETS1/AP1 activity. The results also indicated that EC migration can be mediated through PKC-dependent and independent pathways and only the former pathway can induce EC morphogenesis as well.

Ultrastructure and permeability of endothelial cells in branched regions of rat arteries

The ultrastructure and the permeability to macromolecules of the endothelia in the branched and unbranched regions of the arteries were compared using two different age groups (3 and 12 months) of rats. In the aortic arch, the endothelial cells were longer and thinner and contained fewer intracytoplasmic vesicles than those observed in the unbranched regions of aorta. Quantitative study revealed that the volume density of intracytoplasmic vesicles in the branched regions of aortic arch in 3-month-old rats was significantly (P < 0.01) lower than the density value in the unbranched regions of aorta. The volume densities of vesicles in both regions of the aorta were lower than those in the carotid artery. There was an apparent increase in the frequency of the simple type of interendothelial contacts and a decrease in the complex type in the branched regions as compared with those in the unbranched regions of aorta and carotid artery. In addition to the normal interendothelial contacts, several open junctions with increasing width (25-300 nm) were identified in the branched regions of aortic arch and the bifurcations of carotid artery. For rats at the age of 12 months, local areas of the subendothelial space were expanded. Basal lamina-like and electron-dense materials were accumulated in the subendothelium. The volume densities of vesicles in the aortic endothelia were significantly (P < 0.01) increased as compared with those in the 3-month-old group. The volume density of vesicles in the aortic arch was again significantly (P < 0.01) lower than that in the unbranched regions of aorta. Furthermore, the frequency of the simple type of intercellular contacts was increased, whereas that of the complex type was decreased in both regions of aorta. With regard to the junctional complexes, the frequencies of gap junctions and tight junctions were increased and the junctionless intercellular contacts were decreased compared with those of the 3-month-old group.

ABCA1 modulates the oligomerization and Golgi exit of caveolin-1 during HDL-mediated cholesterol efflux in aortic endothelial cells

Previously, the authors have shown that the molecular interaction between caveolin-1 and ATP-binding cassette transporter A1 (ABCA1) is associated with the high-density lipoprotein (HDL)-mediated cholesterol efflux pathway in aortic endothelial cells (ECs). This study analyzed the role ABCA1 plays in caveolin-1-mediated cholesterol efflux in aortic ECs. Knockdown of ABCA1 by siRNA in primary rat aortic ECs after cholesterol treatment did not affect caveolin-1 expression but led to the retention of caveolin-1 in the Golgi apparatus, impaired caveolin-1 oligomerization, and reduced cholesterol efflux. Immunoblotting assay and immunofluorescence microscopy demonstrated that HDL transiently up-regulated ABCA1 expression, induced caveolin-1 oligomerization, and promoted its Golgi exit, thereby enhancing cholesterol efflux. These HDL-induced events, however, were inhibited by down-regulation of ABCA1. It is concluded that HDL up-regulates ABCA1 expression, which in turn modulates the oligomerization and Golgi exit of caveolin-1 to enhance cholesterol efflux in aortic ECs.

Use of Gain-of-Function Study to Delineate the Roles of crumbs in Drosophila Eye Development

The cell polarity gene, crumbs(crb), has been shown to participate in the development and degeneration of the Drosophila retina. Mutations in CRB1, the human homologue of Drosophilacrb, also result in retinitis pigmentosa and Leber congential amaurosis. In this study, we used the gain-of-function approach to delineate the roles of crb in developing Drosophila eye. In the third-instar larval stage, eye development is initiated with photoreceptor differentiation and positioning of photoreceptor nuclei in the apical cellular compartment of retinal epithelium. In the pupal stage, differentiated photoreceptors begin to form the photosensitive structures, the rhabdomeres, at their apical surface. Using GMR-Gal4 to drive overexpression of the Crb protein at the third-instar eye disc, we found that differentiation of photoreceptors was disrupted and the nuclei of differentiated photoreceptors failed to occupy the apical compartment. Using hs-Gal4 to drive Crb overexpression in pupal eyes resulted in interference with extension of the adherens junctions and construction of the rhabdomeres, and these defects were stage-dependent. This gain-of-function study has enabled us to delineate the roles of Crb at selective stages of eye development in Drosophila.

A prototype tissue engineered blood vessel using amniotic membrane as scaffold

In this study, we used amniotic membrane (AM), a natural extracellular matrix, as a scaffold for the fabrication of tissue engineered blood vessels (TEBVs). The inner surface of the denuded glutaraldehyde cross-linked AM tube was endothelialized with porcine vascular endothelial cells (ECs) and subjected to a physiological (12 dynecm(-2)) shear stress (SS) for 2 and 4 days. The results showed that after applying SS, an intact EC monolayer was maintained in the lumen surface of the TEBV. The ECs were aligned with their long axis parallel to the blood flow. The immunofluorescent microscopy showed that the intercellular junctional proteins, PECAM-1 and VE-cadherin, were surrounding the EC periphery and were better developed and more abundant in SS-treated TEBVs than the static controls. The Western blot indicated that the expressions of PECAM-1 and VE-cadherin were increased by 72 ± 9% and 67 ± 7%, respectively, after shear stress treatment. The distribution pattern of integrin β1 was mainly at the interface of ECs and AM in static TEBVs but it was extended to the cell-cell junctions after SS treatment. The SS promoted the expression of integrin α(v)β(3) without altering its distribution in TEBV. The results suggest that glutaraldehyde cross-linked AM tube can potentially be used as a scaffold biomaterial for TEBV fabrication. Most importantly, the use of an AM tube shortened the TEBV fabrication.

High glucose impairs EDHF-mediated dilation of coronary arterioles via reduced cytochrome P450 activity.

Endothelium-derived hyperpolarizing factor (EDHF) is an important vasodilator that regulates the vasomotor function. However, it remains unclear whether diabetes/hyperglycemia-induced vascular impairments extend to the EDHF. The present study aims to determine the effect of high glucose (HG) on EDHF-mediated arteriolar dilation and the underlying mechanism. Porcine coronary arterioles were isolated and pressurized for vasomotor study. Cultured porcine coronary artery endothelial cells (ECs) were used for molecular and biochemical analysis. Our results demonstrate that bradykinin (BK)-simulated arteriolar dilation is mediated by nitric oxide (NO) and EDHF pathways. Direct incubation of HG impaired vasodilation to BK but not to sodium nitroprusside (endothelium-independent vasodilator). In the presence of inhibitors of endothelial NO synthase (eNOS) and cyclooxygenase, the EDHF-mediated dilation was reduced by HG incubation. The inhibitory effect of HG was prevented by treating the vessels with superoxide scavenger Tempol. In cultured coronary endothelial cells, HG reduced endothelial epoxyeicosatrienoic acid (EET) production as well as cytochrome P450 epoxygenase (CYP) activity. Furthermore, the superoxide production was elevated in ECs after HG incubation. Pretreatment with Tempol before HG incubation prevented the increase of cellular superoxide and abolished the decrease of CYP activity. Collectively, our results suggest that, in addition to NO-mediated pathway, HG impairs the EET/EDHF-mediated vasodilation in coronary arterioles via the elevated level of superoxide leading to inhibition of CYP activity in coronary ECs.