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Persistent Elevation and Metabolic Dependence of Circulating E-Selectin after Delivery in Women with Gestational Diabetes Mellitus

Department of Medicine III, Division of Endocrinology and Metabolism (A.K-W., P.F., W.W.), Department of Clinical Pharmacology (B.J.) and Department of Medical and Chemical Laboratory Diagnostics (O.F.W.), University of Vienna, Austria

Address all correspondence and requests for reprints to: Oswald F Wagner, Department of Clinical and Chemical Laboratory Diagnostics, University of Vienna, Währinger Gürtel 18–20, 1090 Vienna, Austria.

	Abstract

Top Abstract Introduction Subjects and Methods OGTT Statistical analyses Results Discussion References

 
The increased risk of premature atherosclerosis in noninsulin-dependent diabetes mellitus (NIDDM) might be related in part to augmented expression of endothelial adhesion molecules (AMs). So far it is, however, unknown whether increased circulating (c) AMs in NIDDM are only a consequence of this disease or also involved in its sequelae.

To determine the presence of cAMs in a population at increased risk for subsequent development of NIDDM, we analyzed fasting and postprandial [oral glucose tolerance test (OGTT): 100 g] serum concentrations of circulating E-selectin, vascular cell adhesion molecule-1 (cVCAM-1), and intercellular adhesion molecule-1 (cICAM-1) in pregnant women with either gestational diabetes (GDM) or normal glucose tolerance (NT) before and after delivery vs. nonpregnant healthy women (C). During pregnancy cE-selectin and cVCAM-1 were elevated in both GDM and NT vs. nonpregnant females (P < 0.01 vs. C). Following delivery, all GDM females regained normal glucose tolerance according to OGTT criteria, but showed slightly higher postprandial [area under the curve (AUC)_{180 min}] glycemia and HbA1c values than nonpregnant healthy women (P < 0.05), indicating persisting subtle abnormalities in carbohydrate metabolism. cE-selectin and cVCAM-1 remained increased in GDM (P < 0.01 vs.C) after delivery, but fell to normal in NT (P < 0.05 before vs. after delivery). Furthermore, a correlation was seen in GDM females between cE-selectin and HbA1c (P < 0.005), fasting glucose (P < 0.01), and insulin (P < 0.05) as well as postprandial (AUC_{180 min}) glucose and insulin concentrations (P < 0.05) during OGTTs, both before and after delivery. ICAM-1, however, did not differ significantly between groups.

In summary, GDM is characterized by persistently raised levels of cE-selectin and cVCAM-1 12 weeks after delivery. Whether these persistent elevations of cE-selectin and cVCAM-1 reflect early vascular injury or represent a risk factor for atherosclerosis in women at increased risk for NIDDM remains to be determined.

	Introduction

Top Abstract Introduction Subjects and Methods OGTT Statistical analyses Results Discussion References

 
THE adhesion molecules (AMs) E-selectin, vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) are supposed to play a major role in the pathogenesis of vascular disease (1, 2). It is assumed that their endothelial expression triggers adhesion of monocytes to the endothelium, an early step in the development of atherosclerosis (1, 3). Serum concentrations of circulating (c) AMs may reflect endothelial activity and/or damage (4), because endothelial cells release soluble forms of AMs in correlation to their surface expression (3, 5). The origin of individual cAMs differs. Surface expression of E-selectin is completely and that of VCAM-1 rather specific for endothelium. In contrast, the origin of ICAM-1 is widespread and thus does not allow conclusions on the endothelial state.

Noninsulin-dependent diabetes mellitus (NIDDM) is associated with an increased risk of premature atherosclerosis (6, 7). Recently, increased serum concentrations of AMs have been observed in NIDDM (8, 9, 10, 11). In some (8, 9, 10) but not all (11) of these studies a potential effect of glycemic control on cE-selectin was postulated. These controversial observations might be because of cross-sectional study design and heterogeneity of patients for the presence of diabetic complications and additional atherogenic risk factors.

Pregnancy is a hyperinsulinemic state, which may transgress into impaired glucose tolerance if insulin secretion is unable to compensate for pregnancy-associated insulin resistance (12, 13, 14). Although most women with gestational diabetes mellitus (GDM) regain normal glucose tolerance after delivery, many of them feature subtle disturbances in postpartum glucose homeostasis and are at high risk of later developing NIDDM (13, 14, 15, 16). Thus, GDM can be regarded a model for early NIDDM (16).

To investigate the hypothesis of a dependence of cE-selectin on glucose metabolism in the absence of coexistent atherosclerosis and additional atherogenic risk factors, cAMs were studied in GDM and pregnant women with normal glucose tolerance (NT) before and after delivery as well as in healthy nonpregnant women (C).

	Subjects and Methods

Top Abstract Introduction Subjects and Methods OGTT Statistical analyses Results Discussion References

 
None of the subjects [GDM, NT, or control (C)] studied had a family history of diabetes, was taking any medication, or had any known disease. All women had normal renal and liver function, were negative for islet cell antibodies, and free of manifest infections, hypertension, hyperlipidemia, and obesity (Table 1). Cesarean section or any other surgical intervention were exclusion criteria. No pregnant woman had signs of placental insufficiency as documented by Doppler velocimetry and pH of umbilical cord. All females delivered healthy children appropriate for gestational age. The nonpregnant control subjects (C) were members of the paramedical staff. GDM was diagnosed by a 100-g oral glucose tolerance test (OGTT) following the criteria of O’Sullivan and of the 3rd International Workshop on Gestational Diabetes Mellitus (17, 18). A positive 100-g OGTT was defined as two or more values above 5.8 mmol/L in the fasting state, 10.6 mmol/L at 1 h, 9.2 mmol/L at 2 h, and/or 8.1 mmol/L at 3 h after glucose loading. The groups were matched for age and (preconceptual) body mass index (BMI). There were no significant differences in weight gain during pregnancy (12.0 ± 2.0 kg in GDM and 14.8+1.9 kg in NT) or BMI at term (30.3 + 1.8 vs. 29.0 + 0.9 kg/m²) between GDM and NT. Three months after delivery, BMI decreased to the preconceptual one in both groups (Table 1). Women with GDM, who were reinvestigated for reclassification of glucose tolerance after delivery using a 75-g standard OGTT (WHO criteria; Ref.19), featured normal glucose tolerance. In addition, 100-g OGTTs were repeated 12 weeks after delivery for comparison of glucose and insulin release vs. the pregnant state. cAMs were determined both in the fasting state and 180 min after glucose loading (100 g). The protocol has been approved by the ethics committee of the University of Vienna. Informed consent was obtained from all subjects studied.

	OGTT

Top Abstract Introduction Subjects and Methods OGTT Statistical analyses Results Discussion References

Serum samples for determination of cAMs were stored at -80 C and analyzed in duplicate by commercially available enzyme-linked immunosorbent assays (British Bio-Technology Product Ltd., Abdington, UK) as described previously (20) with both inter- and intraassay coefficients of variation being <6%.

	Statistical analyses

Top Abstract Introduction Subjects and Methods OGTT Statistical analyses Results Discussion References

 
Results are expressed as means and SEM unless otherwise designated. Group differences were determined by ANOVA; correlation coefficients were calculated by linear regression analysis. In addition, multiple regression analysis was computed (STAT Software), to determine independent regulators of cE-selectin levels. Differences within GDM and NT before vs. after delivery were calculated by the paired Student’s t test. A P value <0.05 was considered significant.

	Results

Top Abstract Introduction Subjects and Methods OGTT Statistical analyses Results Discussion References

 
Fasting (Fig. 1) and postprandial serum concentrations of cE-selectin and cVCAM-1 were elevated (P < 0.05) in GDM (fasting: cE-selectin +128%, cVCAM-1 +63%; postprandially: +110%, +55%, respectively) and NT (fasting: cE-selectin +145%, cVCAM-1 +69%; postprandially: +107%, +59%, respectively) compared with nonpregnant females (C), whereas fasting and stimulated cICAM-1 did not differ between groups. There were no differences between fasting and stimulated (180 min, 100-g OGTT) cAMs in each group.

View larger version (17K): [in this window] [in a new window]   Figure 1. cE-selectin, cVCAM-1, and cICAM-1 in pregnant women with GDM and those with normal glucose tolerance (NT) before and after delivery as well as in nonpregnant control females (C); shown for each single individual. #, P < 0.01 vs. C (ANOVA). *, P < 0.05, NT before vs. after delivery (paired Student’s t test).

View larger version (26K): [in this window] [in a new window]   Figure 2. Relationship between cE-selectin and HbA1c values (A), fasting plasma glucose concentrations (B), and AUCs during OGTT(AUC180 min; 100 g) for glucose (C) and insulin (D) in women with GDM during pregnancy (• and solid line) and 3 months after delivery ( and dotted line).

In GDM, cE-selectin correlated with fasting glucose, HbA1c, glucose_AUC, insulin_AUC (Fig. 2), and fasting insulin (r = 0.5, P = 0.047). In the entire group of females (GDM, NT, and C), cE-selectin also correlated with fasting glucose (r = 0.5, P < 0.05), insulin (r = 0.63, P < 0.02), glucose_AUC (r = 0.77, P < 0.0025), and insulin_AUC (r = 0.63, P < 0.02), as well as with HbA1c (r = 0.64, P < 0.03). Multiple regression analysis selected only glucose but not insulin levels as an independent predictor of cE-selectin levels (P < 0.05).

	Discussion

Top Abstract Introduction Subjects and Methods OGTT Statistical analyses Results Discussion References

 
The elevations of cE-selectin and cVCAM-1 in GDM and NT vs. matched nonpregnant control women might suggest that such increase is characteristic for pregnancy or the insulin resistance in pregnancy. The responsible mechanisms for that phenomenon however remain to be elucidated. Because no evidence of VCAM-1 and E-selectin expression has been found in placentae from normal pregnancies as well as from those complicated by preeclampsia or intrauterine growth retardation (21), the increase in cE-selectin and cVCAM-1 could be influenced by the endocrine changes related to pregnancy.

In accordance with others reporting normalization of glucose tolerance in 90% of the women with former GDM postpartum (22), all GDMs had returned to normal glucose tolerance according to established OGTT criteria. Interestingly, contrary to NT, where cE-selectin and cVCAM-1 fell to normal after delivery, in former GDMs these cAMs remained elevated. Although it cannot be excluded that cE-selectin and cVCAM-1 might decrease to normal after a longer follow-up period in these subjects also, it is remarkable that their normalization within 3 months after delivery appeared in all women with normal glucose tolerance, whereas they remained unchanged in women with GDM. Because cE-selectin and cVCAM-1 even tended to increase (ns) in GDM by approximately 8–15% 12 weeks after delivery, ongoing endothelial dysfunction might be self-perpetuating after its initiation during pregnancy. In contrast to the endothelial-specific cAMs E-selectin and VCAM-1, cICAM-1, which is of nonspecific origin, did not differ between or within any group.

It has to be emphasized that a homogeneous group of GDM females without additional risk factors for NIDDM and atherosclerosis has been selected for evaluation of a relation of cAMs to glucose metabolism. Although the group might represent a subset of GDM subjects at lower risk for later development of NIDDM, these women, nevertheless, featured postpartum persisting metabolic abnormalities commonly described in former GDM such as mild hyperglycemia, slightly elevated basal insulin concentrations, and a defect in acute glucose-stimulated insulin release (13, 15, 23).

Increased cAM levels in general may reflect both changes in synthesis/release and clearance. Data on clearance sites and rates of cAMs are not available so far. However, in young women with normal kidney and liver function, increased levels are expected to primarily reflect synthesis/release (4).

In contrast to NIDDM, advanced atherosclerosis is not likely in young females lacking additional risk factors. However, microvascular dysfunctions can be present in otherwise healthy prediabetic subjects with mild fasting hyperglycemia and hyperinsulinemia related to insulin resistance (24) and therefore might account for the observed increase in E-selectin and VCAM-1.

A potential association between a high risk to develop NIDDM and cE-selectin is supported by the strong relation between cE-selectin and metabolic control in GDM subjects (Fig. 2). In a multiple regression analysis, E-selectin was independently related to glycemia and only secondarily to associated basal and total (basal and delayed postprandial) insulin release. The correlation between E-selectin and plasma glucose concentrations in the entire group of females might further support the hypothesis of a possible effect of chronic glycemia on cE-selectin expression. Although glucose has been shown to increase monocyte binding to human endothelial cells in vitro (25), it failed to directly stimulate E-selectin expression in cultured endothelial cells (26). As recently proposed by Cominacini and co-workers (10), glycemia might in vivo affect E-selectin plasma concentrations through its effect on oxidative stress.

Although cVCAM-1 was not directly related to glucose metabolism in GDM or NIDDM, additional metabolic aberrations as increased formation of advanced glycosylation end products may add to its increase (27).

The finding of elevated cE-selectin and cVCAM-1 in women at risk for NIDDM gains further relevance by the recently described direct angiogenic effect of both soluble E-selectin and VCAM-1 (28), which may also play an important role in the development of diabetic retinopathy and nephropathy. Indeed, higher cVCAM levels have been found in insulin- dependent diabetic patients with retinopathy and nephropathy than in those without these complications (29).

Although the relevance of elevated soluble AMs in vascular disease is not established until now, three models for a role of elevated cE-selectin and cVCAM-1 may be considered. First, elevated serum levels could reflect increased expression of the respective cAMs on the endothelial surface, thereby potentially stimulating monocyte recruitment into the vascular wall. Secondly, increased cAMs may themselves induce angiogenesis. Third, increased cAMs may simply be the result of early vascular abnormalities in GDM and therefore represent a consequence rather than a potential cause of early vascular lesions. Further longitudinal studies will be necessary to thoroughly investigate the behavior of cAMs and the potential relation of cAMs to vascular and metabolic changes in women with a history of GDM.

Received December 5, 1996.

Revised March 14, 1997.

Revised June 16, 1997.

Accepted August 19, 1997.

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