Raised concentrations of lipid peroxidation products (LPO) in pregnant women with impaired glucose tolerance.

INTRODUCTION: Lipid peroxidation (LPO) results from oxidative damage to membrane lipids. Whereas LPO rises in normal pregnancy, the effect of gestational diabetes mellitus (GDM) on this process has not been clearly defined. MATERIALS AND METHOD: Fasting blood concentrations of malondialdehyde+4-hydroxyalkenals (MDA+4-HDA), as LPO index, TNFa soluble receptors (sTNF-R1 and sTNF-R2), and soluble adhesion molecules (sICAM-1, sVCAM-1), were measured in 51 women at 28 weeks of gestation. The women were divided according to the results of 50.0 g glucose challenge test (GCT) and 75.0 g oral glucose tolerance test (OGTT): Controls (n=20), normal responses to both GCT and OGTT; Intermediate Group (IG) (n=15), abnormal GCT but normal OGTT; GDM group (n=16), abnormal both GCT and OGTT. RESULTS: Glucose concentrations in women diagnosed with GDM were within the range of impaired glucose tolerance. There were no significant differences in concentrations of either TNF a soluble receptors R1 and R2, or sICAM-1 or sVCAM-1. LPO concentrations [MDA+4-HDA (nmol/mg protein)] were significantly higher in women with GDM than in the other two groups [64.1±24.3 (mean±SD), 39.3±23.1, 47.0±18.1, for GDM, IG and Controls, respectively; p<0.05]. In multivariate analysis, the only significant independent correlation was between LPO level and glucose at 120 minutes of OGTT (rs=0.42; p=0.009). CONCLUSIONS: Oxidative damage to membrane lipids is increased in GDM and might result directly from hyperglycaemia. Physiological significance of this phenomenon remains to be elucidated.

Limited penetration of perfluorocarbon in porcine pancreas preserved by two-layer method with (19)fluorine magnetic resonance spectroscopy and headspace gas chromatography.

The mechanism of the two-layer method (TLM) of pancreas preservation is unclear. Facilitating oxygen diffusion into preserved pancreas has been suggested, but direct measurements of tissue pO(2) have yielded conflicting results. The degree of penetration of perfluorocarbon (PFC) into the pancreas during TLM storage is unknown. Segments of porcine pancreas (7.5 cm in length) were preserved either in University of Wisconsin solution (UW) alone (n = 6) or in TLM for 24 h (n = 6). Pancreatic samples were analyzed using Varian INOVA 9.4 T MR scanner. External PFC standard was introduced for quantification. Four consecutive transverse images of 4 mm thickness were obtained using a spin-echo sequence. (19)Fluorine magnetic resonance spectroscopy ((19)F MRS) was performed with the same parameters except with more averages. MR data were confirmed by headspace chromatography. PFC standard was readily detected in (19)F MR images. There was no signal from pancreas in (19)F MR images following either UW or TLM storage. (19)F MR spectra typical of PFC were not obtained from either UW- or TLM-preserved pancreas with nonlocalized (19)F MRS. Mean concentration of PFC in TLM pancreas measured by head space chromatography was 0.011 nl/g (SD ± 0.006), not significantly different from background concentration (0.012 nl/g, SD ± 0.006) in UW pancreas (p = 0.42). There was no evidence of penetration of PFC into pancreas tissues investigated either by MR or chromatography in organs preserved at hypothermia by TLM, and mechanisms of TLM remain speculative.

Serum levels of matrix metalloproteinases MMP-2 and MMP-9 and their inhibitors in women with glucose intolerance in pregnancy and normal controls.

AIM: Matrix metalloprotenases (MMPs) are proteolytic enzymes active in inflammatory states. We have examined MMP-9, MMP-2, and their respective tissue inhibitors: TIMP-1 and TIMP-2 in sera of women with gestational diabetes mellitus (GDM) and various degrees of insulin resistance (IR) in the third trimester of pregnancy. METHODS: Fasting serum levels of MMP-9, MMP-2, TIMP-1 and TIMP-2 were measured in 26th-28th week of gestation in 51 women divided according to their response to a 50-g glucose challenge test (GCT) and a 75-g OGTT: controls (n = 20): both tests normal; the GDM group (n = 16) both tests abnormal; the intermediate group (IG; n = 15) abnormal GCT and normal OGTT. MMPs and TIMPs were correlated with the parameters of IR: homeostasis model assessment (HOMA) and insulin resistance index (IRI). RESULTS: MMP-9, MMP-2, TIMP-1 and MMP-9/TIMP-1 ratio were not different among the groups. TIMP-2 levels were significantly higher in the GDM and IG groups than in controls (p < 0.01). MMP-2/TIMP-2 ratio was lower in the GDM group than in the other groups (p < 0.01) and was correlated to HOMA and IRI (r = -0.465 and r = -0.43 respectively, p < 0.01). CONCLUSIONS: Serum MMP levels do not reflect inflammation in GDM. Elevated TIMP-2 and consequently lower MMP-2/TIMP-2 levels in GDM need to be clarified, but are unlikely to be a consequence of inflammation.

Mutation in the TBCE gene is associated with hypoparathyroidism-retardation-dysmorphism syndrome featuring pituitary hormone deficiencies and hypoplasia of the anterior pituitary and the corpus callosum.

CONTEXT: Hypoparathyroidism-retardation-dysmorphism (HRD) syndrome, an autosomal recessive disorder characterized by distinct clinical, biochemical, and genetic abnormalities, is characterized by severe short stature, the etiology of which is unclear. Homozygous mutation of the tubulin cofactor E (TBCE) gene leading to loss of four amino acids (c.155-166del12; p.del 52-55) in the TBCE protein has been associated with the syndrome. AIM: The aim of the study was to describe the clinical, biochemical, and neuroradiological features of children with genetically proven HRD syndrome. METHODS: Six children from four independent Middle Eastern pedigrees with clinical features of HRD syndrome were confirmed to have the previously reported homozygous mutation in TBCE (c.155-166del12) and were investigated with magnetic resonance imaging (MRI) of the brain and standard pituitary function testing. RESULTS: Cranial MRI in all children showed severe hypoplasia of the anterior pituitary and corpus callosum, with decreased white matter bulk. Four of five children tested had subnormal GH and cortisol responses to glucagon, and plasma IGF-I concentration was low in all six children. Cortisol response to synacthen was suboptimal in one of three patients tested. Male children (n = 3) had clinical features suggestive of hypogonadotropic hypogonadism. CONCLUSION: GH insufficiency, hypocortisolemia, and abnormal cranial MRI appear to be associated with HRD syndrome and may contribute in part to the short stature. Our data support the need for longer term monitoring for evolving pituitary hormone deficiencies and raise the possibility that TBCE may play a role in development of the anterior pituitary, corpus callosum, and white matter in addition to the parathyroid glands.

Highly selective intraportal transplantation of pancreatic islets.

BACKGROUND: Histological assessment of intraportally transplanted islets in experimental rodent models is limited by the wide dissemination of islets throughout the liver. We describe a technique of highly selective intraportal islet transplantation, which increases the density of transplanted islets and hence facilitates their histological analysis and validate this model as a means of quantitative histological analysis of islet graft survival. We also compared the number of islets visualized histologically with that of nonabsorbable dextran beads, representing the number of transplanted islets there would have been if there had been no graft loss. MATERIALS AND METHODS: Diabetic Lewis rats or nondiabetic Sprague-Dawley rats were transplanted with 500 or 1000 syngeneic islets or an equivalent number of beads either into the main branch (MB), or selectively into the right branch (RB) of the portal vein. RESULTS: Islet transplantation led to an identical fall in blood glucose levels whichever technique was used. The graft area and number of islets recovered for histological analysis was 3- to 4-fold higher when islets were transplanted using the RB technique. Quantitative histological graft analysis demonstrated that 46% to 61% of intraportally transplanted islets were lost compared with corresponding bead graft sizes. Fewer islets were lost when a greater islet mass was transplanted. CONCLUSIONS: Selective islet transplantation increases the concentration of islets and hence facilitates islet recovery after intraportal transplantation without detrimental effects on transplantation outcome. This technique, when combined with bead transplantation and digital image analysis, provides an accurate method for estimating the number of islets surviving intra-portal transplantation.

Elevated concentrations of retinol-binding protein-4 (RBP-4) in gestational diabetes mellitus: negative correlation with soluble vascular cell adhesion molecule-1 (sVCAM-1).

BACKGROUND: Retinol-binding protein-4 (RBP-4) may increase insulin resistance (IR) in animals, with elevated levels reported in humans with obesity and type 2 diabetes. There are, however, few data on concentrations of RBP-4 in gestational diabetes mellitus (GDM). METHODS: We measured fasting serum levels of RBP-4, soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular cell adhesion molecule-1 (sVCAM-1) in 50 women at 28 weeks of gestation, divided according to the results of a 50 g glucose challenge test (GCT) and a 75 g oral glucose tolerance test (OGTT): (1) controls (n = 20), normal responses to both GCT and OGTT; (2) intermediate group (IG) (n = 15): false positive GCT, but normal OGTT; and (3) GDM group (n = 15), both GCT and OGTT abnormal. IR was assessed by homeostasis model assessment (HOMA-IR) and by insulin resistance index (IRI) based on glycemia and insulinemia during OGTT. RESULTS: All groups were matched for age and body mass index (BMI). RBP-4 levels (microg/ml, mean+/-standard deviation) were higher in women with GDM vs. controls (53.9 +/- 17.9 vs. 29.7 +/- 13.9, p < or = 0.001), with a trend towards higher RBP-4 in GDM compared with IG (38.0 +/- 19.3, p = 0.07). There was no significant correlation between RBP-4 and age, BMI, insulin, IRI or HOMA-IR, but there was a moderate, significant negative correlation between RBP-4 and sVCAM-1 (r(2) = 0.20, p = 0.001). CONCLUSIONS: RBP-4 levels are elevated in women with GDM, but do not correlate with IR indices and correlate negatively with sVCAM-1. The physiological significance of RBP-4 rise in women with GDM remains to be elucidated.

Time course of islet loss after intraportal transplantation.

A high percentage of islets fail to survive intraportal transplantation, but the absolute amount and time course of this loss are uncertain. We have devised a technique to directly quantitate the number of surviving islets using simultaneous selective transplantation of islets and inert beads into the posterior lobes of the liver. Islet:bead ratio did not change significantly within the first 2 hours, but fell progressively thereafter, giving calculated islet survival rates of 89, 43, and 66% at days 1, 7 and 28, respectively. This technique provides a baseline from which to develop strategies to improve the survival of intraportally transplanted islets.

The importance of treating multiple cardiometabolic risk factors in patients with Type 2 diabetes.

Type 2 diabetes is usually associated with a number of metabolic and cardiovascular (or cardiometabolic) risk factors that contribute to a high rate of vascular events in these patients. Adipose tissue is now known to secrete a number of pro-inflammatory adipokines that are thought to mediate the link between obesity, insulin resistance and atherosclerosis. Therefore, not only is abdominal obesity a major cardiometabolic risk factor per se, it has the potential to give rise to other emerging risk factors. Plasma concentrations of inflammatory markers, such as C-reactive protein, may provide additional information to guide management and may even represent therapeutic targets. Reducing the risk of cardiovascular events in patients with Type 2 diabetes will involve targeting traditional risk factors and probably novel cardiometabolic factors.

Time course and quantification of pancreatic islet revascularization following intraportal transplantation.

A large proportion of islets are lost after transplantation partly due to a lack of functional vasculature. Islets revascularize from host tissue but the process takes up to 2 weeks and has been suggested to result in reduced vascular density in engrafted islets. We describe a method for observing and quantifying the revascularization of intraportally transplanted islets that includes number, density, and branching of islet capillaries. Syngeneic islets were transplanted selectively into the two right posterior lobes of the liver of adult Lewis rats. Sections of the livers were dual stained for insulin and Bandeiraea simplicifolia and analyzed for islet morphology, area, and vascular density from day 0 to day 14 posttransplant and compared to native islets. Vascular density was 1431 +/- 75.7 vessels/mm2 in native islets and fell to 325.3 +/- 30.8 vessels/mm2 (p < 0.001) by day 1 posttransplant and subsequently increased until day 14 when it was significantly higher than in native islets (2612.5 +/- 107.8 vessels/mm2, p < 0.001). The percentage of islet area occupied by vascular space was 9.1 +/- 0.9% in native islets. After falling to 2.3 +/- 0.3% (p < 0.001) 1 day posttransplant this rose to supranormal levels (21.5 +/- 0.8%, p < 0.001) by day 14. The index of capillary branching was 0.771 +/- 0.017 in native islets and fell to 0.465 +/- 0.02 (p = 0.001) by day 3 but returned to native values by day 7 posttransplantation (0.726 +/- 0.03). This technique provides a robust method for tracking and quantifying the revascularization of intraportally transplanted islets, which should enable the comparison of different strategies aimed at accelerating islet revascularization.

Effect of inspired oxygen on portal and hepatic oxygenation: effective arterialization of portal blood by hyperoxia.

Because hypoxia may compromise the survival of intraportally transplanted pancreatic islets, we have measured portal blood flow and both portal and hepatic oxygenation in normal and diabetic rats breathing graded inspired oxygen concentrations. Portal blood flow and hepatic tissue oxygenation were measured using a transonic flowmeter and near infrared spectroscopy while gas analysis was carried out on portal venous blood samples. The effects of breathing 13%, 21%, 50%, or 100% oxygen were compared in animals with steptozotocin-induced diabetes and in controls. In diabetic rats breathing 21% oxygen, portal blood flow was significantly lower than in controls (7.2+/-0.7 vs. 9.1+/-0.8 ml/min, p < 0.05). In both groups, breathing 100% oxygen significantly increased portal flow (to 8.4+/-1.0 and 12.2+/-0.7 ml/min, respectively). This effect was not secondary to hepatic arterial vasoconstriction because it was not prevented by hepatic artery ligation. In controls, breathing 100% oxygen increased portal pO2 from 5.0+/-0.9 to 14.4+/-1.4 kPa (p < 0.05) and portal venous oxygen saturation (PSaO2) from 53.9+/-12.1% to 92.9+/-1.4% (p < 0.05), a value not significantly different from peripheral (arterial) saturation. Similarly, in diabetic animals pO2 rose from 5.6+/-0.3 to 11.7+/-0.4 kPa (p < 0.01) and SO2 from 55.5+/-5.2% to 88.5+/-0.6% (p < 0.05). Hepatic oxyhemoglobin rose and deoxyhemoglobin fell reciprocally as a function of the inspired oxygen concentration. Improved hepatic oxygenation observed in animals breathing oxygen-enriched gas mixtures results from an increase in splanchnic blood flow coupled with a marked increase in portal oxygen saturation. This effective arterialization of portal blood may have important consequences for the success of intraportal transplantation of pancreatic islets.

Hyperoxia improves the survival of intraportally transplanted syngeneic pancreatic islets.

BACKGROUND: Hypoxia in the portal vein may compromise the survival of intraportally transplanted pancreatic islets. We therefore examined the effect of inspired oxygen on the outcome of islet transplantation. METHODS: Blood glucose concentrations, glucose tolerance, and the size and number of surviving islets were measured in diabetic rats housed for 48 hr under hyperoxic (100% O(2)), hypoxic (11% O(2)), or normoxic (21%O(2)) conditions after intraportal transplantation of 350, 500, 700, or 1,000 syngeneic islets. RESULTS: In normoxic diabetic rats, the smallest graft size to consistently restore normoglycemia was 1,000 islets. A graft size of 700 islets was effective in only three of nine animals, whereas 500 islets were ineffective in all eight animals undergoing transplantation. In contrast, in hyperoxically housed rats, graft sizes of 700 or 500 islets restored normoglycemia in eight of nine or five of eight animals, respectively. In those animals that became normoglycemic, the glucose tolerance of the hyperoxically treated rats receiving 700 islets was almost identical to that of normoxically housed animals receiving 1,000 islets. The average size of the islets 6 weeks after transplantation was the same in livers of hyperoxic and control rats. However, the total islet area and number of islets engrafted in hyperoxic rats was significantly increased when compared with livers from normoxic animals receiving the same graft size, so the area in hyperoxic rats receiving 700 islets was not significantly different from normoxic recipients of 1,000 islets. CONCLUSIONS: Hyperoxia posttransplantation increases the number of islets that survive the engraftment process and allows normalization of plasma glucose levels with a smaller number of transplanted islets.