Impact of maternal chromium restriction on glucose tolerance, plasma insulin and oxidative stress in WNIN rat offspring

in Journal of Molecular Endocrinology
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  • 1 Division of Endocrinology and Metabolism, National Centre for Laboratory Animal Sciences, National Institute of Nutrition, Jamai Osmania PO, Hyderabad 500 007, India

Robust evidence suggests that nutritional insult during fetal development could program the offspring to glucose intolerance, impaired insulin response and insulin resistance (IR). Considering the importance of chromium (Cr) in maintaining carbohydrate metabolism, this study determined the effect of maternal Cr restriction (CrR) on glucose metabolism and plasma insulin in Wistar/NIN (WNIN) rat offspring and the associated biochemical and/or molecular mechanisms. Female, weanling WNIN rats received ad libitum for 12 weeks, a control diet or the same with 65% restriction of Cr and mated with control males. Some of the Cr-restricted dams were rehabilitated from conception or parturition and their pups weaned on to control diet. At the time of weaning, half of the Cr restricted offspring were rehabilitated to control diet while others continued on Cr-restricted diet. Maternal CrR increased fasting plasma glucose, fasting insulin, homeostasis model assessment of IR, and area under the curve of glucose and insulin during oral glucose tolerance test in the offspring. Expression and activity of rate-limiting enzymes of glucose metabolism were comparable among different groups and expression of genes involved in insulin secretion was increased albeit in male offspring whereas antioxidant enzyme activities were decreased in offspring of both genders. Rehabilitation, in general, corrected the changes albeit partially. Maternal dietary CrR induced IR, impaired glucose tolerance in WNIN rat offspring and was associated with increased oxidative stress, which may predispose them to type 2 diabetes in their later life.

Abstract

Robust evidence suggests that nutritional insult during fetal development could program the offspring to glucose intolerance, impaired insulin response and insulin resistance (IR). Considering the importance of chromium (Cr) in maintaining carbohydrate metabolism, this study determined the effect of maternal Cr restriction (CrR) on glucose metabolism and plasma insulin in Wistar/NIN (WNIN) rat offspring and the associated biochemical and/or molecular mechanisms. Female, weanling WNIN rats received ad libitum for 12 weeks, a control diet or the same with 65% restriction of Cr and mated with control males. Some of the Cr-restricted dams were rehabilitated from conception or parturition and their pups weaned on to control diet. At the time of weaning, half of the Cr restricted offspring were rehabilitated to control diet while others continued on Cr-restricted diet. Maternal CrR increased fasting plasma glucose, fasting insulin, homeostasis model assessment of IR, and area under the curve of glucose and insulin during oral glucose tolerance test in the offspring. Expression and activity of rate-limiting enzymes of glucose metabolism were comparable among different groups and expression of genes involved in insulin secretion was increased albeit in male offspring whereas antioxidant enzyme activities were decreased in offspring of both genders. Rehabilitation, in general, corrected the changes albeit partially. Maternal dietary CrR induced IR, impaired glucose tolerance in WNIN rat offspring and was associated with increased oxidative stress, which may predispose them to type 2 diabetes in their later life.

Introduction

Fetal growth retardation caused by maternal undernutrition results in obesity, insulin resistance (IR), and associated adult onset diseases such as cardiovascular disease, type 2 diabetes mellitus, and hypertension (Barker et al. 1993). A possible reason for this could be a primary impairment of β-cell development (Dahri et al. 1991) and an overall underdevelopment of endocrine pancreas (Holness 1996). Several animal models have demonstrated that malnutrition during fetal development programs the endocrine pancreas and insulin-sensitive tissues, resulting in IR and diabetes in later life (Snoeck et al. 1990). In utero protein restriction reduces pancreatic β-cell number in rat offspring and blunts insulin secretion in response to oral glucose challenge (Berney et al. 1997, Reusens & Remacle 2001).

We showed earlier that maternal micronutrient restriction increased body fat% especially the central adiposity, impaired plasma lipids, altered glucose tolerance, impaired insulin secretion, increased oxidative stress, and appeared to predispose the Wistar/NIN (WNIN) rat offspring to IR and associated diseases in later life (Venu et al. 2004a,b, 2005, 2008, Padmavathi et al. 2009).

Trivalent chromium (Cr), also called glucose tolerance factor, regulates glucose and lipid metabolism (Mertz 1969). Acting as a cofactor for insulin, it enhances glucose utilization by insulin target tissues (Anderson 1992). It facilitates insulin binding to its receptors, activates insulin receptor kinases and inhibits insulin receptor phosphatases (Vincent 2000). Chromodulin, the Cr-binding oligopeptide activities tyrosine kinase activity of insulin receptor in response to insulin (Davis & Vincent 1997). Although epidemiological evidence for the incidence of Cr deficiency per se is limited, several studies in humans and experimental animals report the beneficial effects of Cr supplementation on glucose tolerance and insulin sensitivity (Anderson 1989, Amoikon et al. 1995, Kitchalong et al. 1995, Ravina et al. 1995). Recently, we reported that maternal Cr restriction (CrR) increased body fat% (central adiposity) and was associated with decreased muscle development in WNIN rat offspring (Padmavathi et al. 2010a,b).

Despite the known impact of Cr supplementation on glucose tolerance and insulin sensitivity, effects if any of maternal CrR in programming the offspring to altered glucose metabolism and plasma insulin levels have not yet been reported. Therefore, this study determined the effect of maternal CrR on glucose metabolism and plasma insulin levels in WNIN rat offspring.

Materials and methods

Study design

Weanling (21-day old), female rats (n=30) WNIN strain were obtained from National Centre for Laboratory Animal Sciences (NCLAS) with the approval of the Ethics Committee on animal experiments at National Institute of Nutrition, Hyderabad, India. They were housed individually in polypropylene cages with wire mesh bottom and maintained at 22±2 °C, under standard lighting conditions (12 h light:12 h darkness cycle). Diets were prepared according to AIN-93G formulation and their Cr content analyzed by atomic absorption spectrometer (Varian Atomic Absorption Spectrometer, Spectra AA 220; Varian India, Agilent Technologies, Forest Hills, VIC, Australia) using reduced flame. The animal feeding and experimental protocol for this study has been described previously (Padmavathi et al. 2010a).

Oral glucose tolerance test (OGTT) was performed (as given below) in WNIN female rats (mothers to be) after 3 months on their respective diets (i.e. before mating) and in the offspring of different groups at quarterly intervals from 3 months of their age. The offspring were sacrificed at 15 months of age after performing the OGTT, the liver and pancreas were dissected out, weighed, snap frozen in liquid nitrogen, and stored at −80 °C till analyzed.

Glucose tolerance and plasma insulin levels

Blood samples were collected from supra-orbital sinus of overnight fasted rats. Glucose was then administered orogastrically at a dose of 2.5 g/kg body weight and blood samples were collected from the tail vein at 30, 60, and 120 min under light ether anesthesia. Plasma glucose was estimated using the commercial kit (Biosystem, Barcelona, Spain) and insulin by the RIA kit (BRIT, Mumbai, India). The area under the curve (AUC) of glucose and insulin during OGTT were computed by the trapezoidal method (Mathews et al. 1990). The indices of IR such as homeostasis model assessment of IR (HOMA-IR) index and the ratio of glucose AUC to insulin AUC during OGTT were calculated as mentioned earlier (Padmavathi et al. 2009).

Activity of enzymes of glucose metabolism

Liver was homogenized and processed as described earlier (Venu et al. 2005) to get the microsomal and cytosolic fractions. Protein content was estimated by the bicinchoninic acid method (Smith et al. 1985) and the activities of the rate-limiting enzymes of glucose metabolism, antioxidant enzymes, and oxidative stress markers were determined as follows.

Hepatic glucokinase (GCK) activity was assayed in the cytosolic fraction by the reduction of NADP at 340 nm (Pilkis 1975). One unit of GCK represents the phosphorylation of 1 μmol of glucose per minute. Pyruvate kinase (PK) activity was determined in liver cytosol by the oxidation of NADH at 340 nm (Bucher & Pfleiderer 1955) and one unit of PK is the nanomoles of phosphoenolpyruvate hydrolyzed per minute.

Glucose-6-phosphatase (G6PC) activity was monitored at 510 nm in the microsomal fraction by the formation of quinoneimine. One unit of G6PC is defined as 1 μmol of glucose-6-phosphate formed per minute (Gierow & Jergil 1982). Fructose-1,6-bisphosphatase (FBP) was determined in the cytosolic fraction by following the rate of NADPH formation at 340 nm (Marcus et al. 1982) and one unit of this enzyme represents the formation of 1 μmol of fructose-6-phosphate per minute.

Gene expression in liver and pancreas by semi-quantitative PCR

Total RNA was isolated from 100 mg each of liver and pancreatic tissues using Qiazol reagent. This was followed by the synthesis of cDNA from 2 μg of total RNA using Invitrogen Kit (Invitrogen Life Technologies). Primers were designed with the aid of primer quest software (Integrated DNA Technologies, Coralville, IA, USA). Semi-quantitative PCR was conducted to analyze the expression of Gck (5′-CCTTAGACCTGGGAGGAACC-3′; 5′-ACGATGTTGTTCCCTTCTGC-3′), Pklr (5′-CCAAGGGACCTGAGATACGA-3′; 5′-AGGTCCACCTCAGTGTTTGG-3′), phosphoenolpyruvate carboxykinase (Pck; 5′-CCCAGGAGTCACCATCACTT-3′; 5′-TTCGTAGACAAGGGGGACAC-3′), G6pc (5′-AGCTCCGTGCCTCTGATAAA-3′; 5′-AAAGTGAGCCGCAAGGTAGA-3′), preproinsulin 1 (Ins1; 5′-CACCTTTGTGGTCCTCACCT-3′; 5′-CCAGTTGGTAGAGGGAGCAG-3′), and Ins2 (5′-CAGCACCTTTGTGGTTCTCA-3′; 5′-CAGTGCCAAGGTCTGAAGGT-3′) with the internal standard 18S rRNA (5′-CCAGAGCGAAAGCATTTGCCAAGA-3′; 5′-AATCAACGCAAGCTTATGACCCGC-3′). The amplified products were resolved on 1.2% agarose gel electrophoresis and the image was quantified with the Bio-Rad gel documentation system using Quantity One Software (Bio-Rad Laboratories). Results are expressed as the ratio of the intensities of the band of the gene of interest to that of the 18S rRNA.

Oxidative stress and antioxidant defense markers

Oxidative stress and antioxidant status were determined in liver homogenate. Thiobarbituric acid reactive substances was determined as a measure of lipid peroxidation (Balasubramanian et al. 1988) and protein carbonyls were quantified spectrophotometrically at 370 nm using 2,4-dinitrophenylhydrazine (Uchida et al. 1998). Reduced glutathione (GSH) and oxidized glutathione (GSSG) were estimated spectrofluorimetrically at excitation and emission wavelengths of 350 and 420 nm, respectively, using ortho-phthalaldehyde (Hissin & Hilf 1976).

Superoxide dismutase (SOD) activity was estimated in the cytosolic fraction (100 000 g supernatant; Marklund & Marklund 1974) and one unit of SOD activity is the amount of the enzyme that inhibits the rate of auto-oxidation of pyrogallol by 50% per minute. The activity of catalase was measured by the reduction of hydrogen peroxide (Aebi 1984) whereas glutathione peroxidase (GPx) was determined in the cytosolic fraction by the oxidation of reduced glutathione by cumene hydroperoxide (Paglia & Valentine 1967). One unit of GPx is the micromoles of NADPH oxidized per minute.

Statistical analysis

All values are presented as mean±s.e.m. Data were analyzed using unpaired Student's t-test to identify differences between control and restricted mothers. One-way ANOVA followed by the multiple range test or least significant difference method was used appropriately to analyze data in the offspring. Wherever heterogeneity of variance was observed, differences between the groups were tested by non-parametric Mann–Whitney U test. The differences were considered significant at P<0.05.

Results

Glucose tolerance and plasma insulin levels in female WNIN rats on CrR

Plasma Cr was lower (P<0.05) in CrR than CrC (chromium control) rats (Padmavathi et al. 2010a). However, there was no difference between CrC and CrR rats in the levels of fasting plasma glucose and insulin or glucose AUC and insulin AUC during OGTT (Table 1). In line with these observations, IR as assessed by HOMA index or the ratio of glucose AUC to insulin AUC during OGTT was comparable between the female CrC and CrR rats (mother to be) before mating (Table 1).

Table 1

Glucose tolerance and insulin resistance indices in female Wistar/NIN rats. Values are mean±s.e.m. (n=6)

CrCCrR
Parameter
Fasting glucose (mmol/l)4.73±0.3874.84±0.167
Fasting insulin (pmol/l)398±83.8411±54.5
HOMA-IR11.6±1.3612.7±1.50
Glucose AUC (mmol/l per h)11.9±0.71411.7±0.508
Insulin AUC (pmol/l per h)815±146801±88.0
Glucose AUC/insulin AUC0.015±0.0020.015±0.001

Growth characteristics of control, Cr-restricted and rehabilitated offspring

Food intake was comparable among the offspring of different groups at all the time points studied (Table 2). Plasma Cr levels were lower (P<0.05) in CrR than CrC offspring and three rehabilitation regimes restored the levels to control from 3 months of their age (Padmavathi et al. 2010a). However, body weight was higher (P<0.05) in CrR than CrC offspring of both the sexes from 12 months of age and rehabilitation appeared to correct the body weight change partially albeit at 12 months of age but not later (Padmavathi et al. 2010a).

Table 2

Daily food intake in male and female offspring of different groups at 9 and 15 months of their age. Values are mean±s.e.m. (n=6)

MonthsCrCCrRCrRCCrRPCrRW
Males
 Food intake (g)915.0±0.34115.7±0.20915.5±0.36215.1±0.31615.4±0.306
1515.1±0.34115.9±0.20915.4±0.32614.9±0.31615.5±0.306
Females
 Food intake (g)912.4±0.47613.4±0.15812.7±0.51912.5±0.42112.2±0.374
1512.2±0.47813.0±0.16012.0±0.51812.8±0.42212.4±0.379

Glucose tolerance and plasma insulin levels to glucose challenge

Males

CrR offspring had higher (P<0.05) fasting plasma glucose and insulin at 9 and 15 months of age (Figs 1A and B and 2) than controls. While rehabilitation restored glucose levels, it had varied effects on plasma insulin levels. At 9 months of age, Cr rehabilitated from parturition (CrRP) and Cr rehabilitated from weaning (CrRW) appeared to mitigate the increased plasma insulin levels whereas Cr rehabilitated from conception (CrRC) had only a partial effect. On the other hand, at 15 months of age CrRC but not CrRP and CrRW restored these changes (Figs 1A and B and 2). HOMA-IR was higher (P<0.05) in CrR at 9 and 15 months of age and the changes in general appeared to be corrected by rehabilitation (Fig. 1C).

Figure 1
Figure 1

Effect of maternal Cr restriction and rehabilitation on glucose tolerance and plasma insulin levels in WNIN male offspring at different ages. Panel A, fasting glucose; panel B, fasting insulin; panel C, HOMA-IR; panel D, glucose AUC during OGTT; panel E, insulin AUC during OGTT; panel F, glucose AUC/insulin AUC during OGTT. Values are mean±s.e.m. (n=6). Bars without a common superscript (‘a and b’) are different at P<0.05 by one-way ANOVA followed by post hoc least significant difference (LSD) test.

Citation: Journal of Molecular Endocrinology 47, 3; 10.1530/JME-11-0010

Figure 2
Figure 2

Effect of maternal Cr restriction and rehabilitation on plasma glucose and insulin levels at various time points (0, 30, 60, and 120 min) during oral glucose tolerance test in WNIN male offspring. Panel A, plasma glucose kinetics at 9 months of age; panel B, plasma glucose kinetics at 15 months of age; panel C, plasma insulin kinetics at 9 months of age; panel D, plasma insulin kinetics at 15 months of age. Values are mean (n=6).

Citation: Journal of Molecular Endocrinology 47, 3; 10.1530/JME-11-0010

Glucose AUC and insulin AUC during OGTT were higher (P<0.05) in CrR than CrC at 9 but not 15 months of age (Figs 1D and E and 2) and rehabilitation in general mitigated the changes. Nevertheless, the ratio of glucose AUC to insulin AUC was comparable among different groups at both the time points (Fig. 1F).

Females

Fasting plasma glucose was higher (P<0.05) in CrR than CrC both at 9 and 15 months of age. Only CrRC appeared to correct the change at 9 months of age while all rehabilitation regimes did so at 15 months of age (Figs 3A and 4). On the other hand, fasting insulin was higher (P<0.05) in CrR than CrC offspring albeit at 15 months of age only and rehabilitation did not appear to correct the change (Figs 3B and 4). As a consequence, HOMA-IR was higher (P<0.05) in CrR than CrC both at 9 and 15 months of age. Curiously, CrRP and CrRW but not CrRC appeared to correct the change partially at 9 months, while no rehabilitation regime appeared to mitigate the change at 15 months of age (Fig. 3C).

Figure 3
Figure 3

Effect of maternal Cr restriction and rehabilitation on glucose tolerance and plasma insulin levels in WNIN female offspring at different ages. Panel A, fasting glucose; panel B, fasting insulin; panel C, HOMA-IR; panel D, glucose AUC during OGTT; panel E, insulin AUC during OGTT; panel F, glucose AUC/insulin AUC during OGTT. Values are mean±s.e.m. (n=6). Bars without a common superscript (‘a and b’) are different at P<0.05 by one-way ANOVA followed by post hoc least significant difference (LSD) test.

Citation: Journal of Molecular Endocrinology 47, 3; 10.1530/JME-11-0010

Figure 4
Figure 4

Effect of maternal Cr restriction and rehabilitation on plasma glucose and insulin levels at various time points (0, 30, 60, and 120 min) during OGTT in WNIN female offspring. Panel A, plasma glucose kinetics at 9 months of age; panel B, plasma glucose kinetics at 15 months of age; panel C, plasma insulin kinetics at 9 months of age; panel D, plasma insulin kinetics at 15 months of age. Values are mean (n=6).

Citation: Journal of Molecular Endocrinology 47, 3; 10.1530/JME-11-0010

Glucose AUC and insulin AUC during OGTT were higher (P<0.05) in CrR than CrC offspring at 15 but not 9 months of age. While all three rehabilitation regimes appeared to restore the glucose AUC to control levels, only CrRP but not CrRC and CrRW appeared to correct insulin AUC (Figs 3D and E and 4). The ratio of glucose AUC to insulin AUC was decreased (P<0.05) in CrR offspring compared with CrC but only at 15 months of age and rehabilitation in general mitigated this change (Fig. 3F).

Gene expression and enzyme activity

Gene expression of hepatic GCK, PK, Pck, and G6PC, the enzymes important in glycolysis and gluconeogenesis was comparable among offspring of different groups (in both genders; Fig. 5A and B).

Figure 5
Figure 5

Effect of maternal Cr restriction and rehabilitation on expression of genes of glucose metabolism by semi-quantitative PCR in liver in WNIN male (A) offspring (at 18 months of age) and female (B) offspring (at 15 months of age); gel picture for each gene is the representation of different groups. Values are mean±s.e.m. (n=6).

Citation: Journal of Molecular Endocrinology 47, 3; 10.1530/JME-11-0010

The activity of hepatic GCK was lower (P<0.05) in female CrR than CrC offspring and all rehabilitation regimes corrected this change. Unlike females, GCK activity was comparable among male offspring of different groups. However, activities of PK, G6PC, and FBP were comparable among offspring of both sexes in different groups (Table 3).

Table 3

Enzymatic activity in liver of Wistar/NIN rat offspring fed different diets. Values are mean±s.e.m. (n=6)

CrCCrRCrRCCrRPCrRW
Males
 G6PC (units/mg)0.014±0.0040.020±0.0020.022±0.0020.014±0.0050.024±0.002
 FBP (units/mg)0.017±0.0010.020±0.0010.018±0.0000.021±0.0020.019±0.001
 GCK (units/mg)0.006±0.0000.005±0.0010.007±0.0000.005±0.0000.007±0.000
 PK (units/mg)0.141±0.0300.109±0.0110.181±0.0030.127±0.0070.102±0.003
Females
 G6PC (units/mg)0.034±0.0020.028±0.0020.036±0.0060.043±0.0050.038±0.000
 FBP (units/mg)0.020±0.0010.019±0.0020.022±0.0010.021±0.0010.021±0.000
 GCK (units/mg)0.012±0.001a0.005±0.001b0.011±0.000a0.009±0.002a0.012±0.003a
 PK (units/mg)0.105±0.0050.100±0.0080.096±0.0070.109±0.0100.108±0.009

Means without a common superscript (‘a and b’) are significantly different at P<0.05 by one-way ANOVA followed by post hoc least significant difference (LSD) test.

Expression of Ins1 and Ins2 genes was higher (P<0.05) in the pancreas of male CrR than CrC offspring. While no rehabilitation regime could restore the change in Ins1 expression, CrRP and CrRW but not CrRC corrected the change in Ins2 expression (Fig. 6A). In female offspring, expression of Ins1 was comparable among groups while curiously, expression of Ins2 was lower (P<0.05) in CrR than CrC offspring. However, rehabilitation in general corrected changes in Ins2 gene expression (Fig. 6B).

Figure 6
Figure 6

Effect of maternal Cr restriction and rehabilitation on expression of genes involved in insulin secretion by semi-quantitative PCR in pancreas in WNIN male (A) offspring (at 18 months of age) and female (B) offspring (at 15 months of age); gel picture for each gene is the representation of different groups. Values are mean±s.e.m. (n=6).

Citation: Journal of Molecular Endocrinology 47, 3; 10.1530/JME-11-0010

Oxidative stress and antioxidant status

Males

Malondialdehyde (MDA) levels were higher (P<0.05) in CrR than CrC offspring and CrRC and CrRP restored this change (Table 4). However, protein carbonyls, glutathione (reduced and oxidized) levels, and catalase activity were comparable among different groups. Interestingly, SOD and GPx activities were lower (P<0.05) in CrR than CrC offspring (Table 4). While CrRP and CrRW corrected the change only in SOD activity, all rehabilitation regimes corrected the reduced GPx activity.

Table 4

Oxidative stress and antioxidant status in liver of Wistar/NIN rat offspring of different groups. Values are mean±s.e.m. (n=6)

CrCCrRCrRCCrRPCrRW
Males
 MDA (nmol/mg)0.540±0.025a0.848±0.044b0.565±0.077a0.606±0.048a0.821±0.025a
 Protein carbonyls (nmol/mg protein)2.32±0.1532.09±0.1172.17±0.0712.11±0.0942.40±0.102
 GSH (μmol/mg)3.58±0.1574.11±0.2123.39±0.4583.24±0.1864.03±0.096
 GSSG (μmol/mg)6.98±0.4528.18±0.6827.56±0.8217.97±0.8037.13±0.745
 Catalase (units/mg)0.110±0.0060.102±0.0080.107±0.0080.114±0.0060.099±0.006
 SOD (units/mg)7.79±0.428a6.12±0.553b6.21±0.249b7.30±0.553a6.94±0.299a
 GPx (units/mg)0.228±0.014a0.165±0.024b0.184±0.016a0.259±0.029a0.215±0.017a
Females
 MDA (nmol/mg)0.358±0.0530.347±0.0200.405±0.0330.340±0.0230.327±0.024
 Protein carbonyls (nmol/mg protein)2.44±0.1432.53±0.0542.38±0.1352.65±0.0612.52±0.103
 GSH (μmol/mg)3.74±0.1403.40±0.2683.60±0.2013.60±0.3613.17±0.367
 GSSG (μmol/mg)6.93±0.7588.00±0.6606.64±0.6567.01±0.4138.63±0.575
 Catalase (units/mg)0.074±0.004a0.049±0.002b0.070±0.003a0.066±0.006a0.051±0.002b
 SOD (units/mg)7.79±0.423a5.97±0.393b6.18±0.296b6.49±0.366b6.30±0.184b
 GPx (units/mg)0.293±0.018a0.207±0.026b0.177±0.012b0.179±0.019b0.208±0.067b

Means without a common superscript (‘a and b’) are significantly different at P<0.05 by one-way ANOVA followed by post hoc least significant difference (LSD) test.

Females

Malondialdehyde, protein carbonyls and glutathione (reduced and oxidized) levels were comparable among female offspring of different groups. However, catalase, SOD, and GPx activities were lower (P<0.05) in CrR than CrC. While no rehabilitation regime could correct the decrease in SOD and GPx activities, CrRC and CrRP but not CrRW could correct the reduced catalase activity (Table 4).

Discussion

We showed earlier that micronutrient restriction in utero predisposed rat offspring to glucose intolerance and altered insulin secretion (Venu et al. 2004a,b, 2005, 2008, Padmavathi et al. 2009). Cr supplementation is known to modulate body composition, improve glucose tolerance and insulin sensitivity (Anderson 1989, Mertz 1993). Recently, we reported that chronic maternal dietary CrR (65% restriction) irreversibly increased body fat% (especially visceral adiposity) and decreased muscle mass (Padmavathi et al. 2010a,b) in WNIN rat offspring, despite comparable (to controls) food intake suggesting the effects to be due to maternal CrR-induced programming of the fetal body composition (Padmavathi et al. 2010c, Vincent & Rasco 2010). Although there was some difference in plasma Cr levels between mothers and offspring, at the levels of dietary CrR employed, they were significantly lower in CrR than controls in both the mothers (35%) and the offspring (57%). In this study, effects of CrR per se and of maternal CrR on glucose metabolism and plasma insulin levels were elucidated in the WNIN rat model.

In line with our reports in maternal magnesium or zinc restricted rat offspring (Venu et al. 2005, Padmavathi et al. 2009), this study showed that chronic dietary CrR per se did not affect glucose tolerance and IR in WNIN female rats. However, they are at variance with the reported effects of Cr deficiency in human subjects (Offenbacher & Pi-Sunyer 1980). This lack of effect of CrR per se could be due to moderate Cr deficiency observed in CrR rats and/or insufficient duration of Cr deficiency and/or the use of a different species (rat).

Chronic maternal dietary CrR induced fasting hyperglycemia, hyperinsulinemia increased HOMA-IR, altered the kinetics of blood glucose and insulin during the OGTT, impaired glucose tolerance and resulted in postprandial hyperinsulinemia to glucose challenge and mitigation of these changes by rehabilitation was albeit partial. These observations are in agreement with studies reporting impaired glucose tolerance in the later life of diet-restricted rat offspring (Dahri et al. 1991, Langley et al. 1994). However, they disagree with the decreased capacity of insulin secretion to a glucose challenge in the offspring born to WNIN rat dams on magnesium and zinc deficiencies (Venu et al. 2005, 2008, Padmavathi et al. 2009). Considering that IR and/or altered insulin secretion exist before the onset of fasting and postprandial hyperglycemia which lead to type 2 diabetes (Weyer et al. 1999, Dostou & Gerich 2001), our findings suggest the predisposal of the CrR offspring to type 2 diabetes. The finding that the changes observed at 9 months of age were not seen later in male offspring is in agreement with our similar findings in Mg restricted rat offspring (Venu et al. 2008). Although the reasons for the transient nature of the effect remains to be understood, the findings do indicate the importance of maternal Cr status in regulating glucose tolerance and plasma insulin levels in later life of the offspring.

In general, blood glucose levels are regulated by a balance between glucose uptake by peripheral tissues and glucose secretion by the liver. An imbalance between systemic glucose delivery (endogenous glucose production) and glucose utilization leads to hyperglycemia (Mevorach et al. 1998). Despite the fasting hyperglycemia and impaired glucose tolerance observed in the CrR offspring, that there were no changes in the gene expression of the rate-limiting enzymes of glycolytic and gluconeogenic pathways, probably indicates that maternal CrR may not alter the expression of these enzymes at transcription level and the observed phenotypic changes could be due to post-transcriptional and/or translational and/or post-translational modifications.

Although enzyme protein expression was not assessed, the observation that activities of the key enzymes of glycolysis and gluconeogenesis were in general comparable among different groups (but for lower GCK in CrR than CrC female offspring and rehabilitation corrected the defect) appear to suggest that maternal CrR may not affect glucose metabolism in the offspring. On the other hand, decreased GCK activity in CrR female offspring could lower glucose catabolism raising intracellular glucose levels that in turn could impair the clearance of circulating glucose and result in hyperglycemia. Such increase in plasma glucose levels may not be conducive to hepatic gluconeogenesis and hence the comparable activities of gluconeogenic enzymes in these offspring. These findings not only agree with mRNA expression data but may also suggest impaired glucose metabolism (decreased glycolysis?) as a possible reason for the fasting hyperglycemia. Although the actual mechanism(s) need to be studied, the present results are the first to the best of our knowledge to demonstrate the effect of maternal CrR on glucose tolerance in the offspring.

Insulin genes in rodents form a two-gene system (Soares et al. 1985, Wentworth et al. 1986) composed of preproinsulin 2 (Ins2), an ortholog to the insulin genes in the other mammals and Ins1, a rodent-specific retrogene. Ins1 and Ins2 are expressed in pancreas and both encode proinsulin peptides that include signal peptide, B chain, C-peptide, and A chain. In line with the observed hyperinsulinemia, expression of Ins1 and Ins2 genes were increased in CrR male offspring indicating that maternal CrR may affect the transcription of the insulin genes, resulting in hyperinsulinemia (fasting and post-glucose). However, it was surprising that in female CrR offspring Ins2 expression was indeed downregulated compared with CrC while expression of Ins1 was unaffected. The reasons for these discrepancies and gender differences need to be deciphered.

In line with our earlier reports on vitamin restricted rat offspring (Venu et al. 2004a), maternal CrR increased oxidative stress (MDA levels) and decreased activities of antioxidant enzymes in the offspring. These findings are in agreement with literature that maternal undernutrition is associated with stress in the offspring (Hostetler & Kincaid 2004) and agrees with increased corticosteroid stress we reported earlier in these offspring (Padmavathi et al. 2010a). The present results are in agreement with substantial evidence that increased oxidative stress and/or decreased antioxidant defense is associated with IR that could lead to type 2 diabetes at a later date (Giugliano et al. 1995, Vijayalingam et al. 1996, Urakawa et al. 2003). However, the actual mechanism(s) and/or the causal relationships among the increased oxidative stress, IR, and impaired glucose tolerance in the CrR offspring remain to be deciphered yet.

Overall, these findings stress the importance of maternal Cr status in modulating carbohydrate metabolism and plasma insulin levels in the offspring. Although much work needs to be done to decipher the underlying/associated mechanism(s), they have unequivocally demonstrated the effects of maternal CrR per se in programming the offspring to glucose intolerance and IR and the possible involvement of increased oxidative stress in the process. Taken together with our earlier demonstration of increased body fat%, especially the visceral adiposity (Padmavathi et al. 2010a) and decreased myogenesis and altered muscle function in CrR offspring (Padmavathi et al. 2010b), the present findings appear to suggest the probable predisposal of the CrR offspring to type 2 diabetes in their later life.

Declaration of interest

The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.

Funding

This work was supported by a research grant to M R from the Department of Biotechnology, Government of India, New Delhi, India (project no. BT/PR2832/Med/14/390/2001).

Acknowledgements

The authors acknowledge the Council for Scientific and Industrial Research (CSIR) and the Indian Council of Medical Research (ICMR) for awarding research fellowships.

References

  • Aebi H 1984 Catalase in vitro. Methods in Enzymology 105 121126.

  • Amoikon EK, Fernandez JM, Southern LL, Thompson DL Jr, Ward TL & Olcott BM 1995 Effect of chromium tripicolinate on growth, glucose tolerance, insulin sensitivity, plasma metabolites, and growth hormone in pigs. Journal of Animal Science 73 11231130.

    • Search Google Scholar
    • Export Citation
  • Anderson RA 1989 Essentiality of chromium in humans. Science of the Total Environment 86 7581. doi:10.1016/0048-9697(89)90196-4.

  • Anderson RA 1992 Chromium, glucose tolerance, and diabetes. Biological Trace Element Research 32 1924. doi:10.1007/BF02784583.

  • Balasubramanian KA, Manohar M & Mathan VI 1988 An unidentified inhibitor of lipid peroxidation in intestinal mucosa. Biochimica et Biophysica Acta 962 5158. doi:10.1016/0005-2760(88)90094-X.

    • Search Google Scholar
    • Export Citation
  • Barker DJ, Hales CN, Fall CH, Osmond C, Phipps K & Clark PM 1993 Type 2 (non-insulin-dependent) diabetes mellitus, hypertension and hyperlipidaemia (syndrome X): relation to reduced fetal growth. Diabetologia 36 6267. doi:10.1007/BF00399095.

    • Search Google Scholar
    • Export Citation
  • Berney DM, Desai M, Palmer DJ, Greenwald S, Brown A, Hales CN & Berry CL 1997 The effects of maternal protein deprivation on the fetal rat pancreas: major structural changes and their recuperation. Journal of Pathology 183 109115. doi:10.1002/(SICI)1096-9896(199709)183:1<109::AID-PATH1091>3.0.CO;2-B.

    • Search Google Scholar
    • Export Citation
  • Bucher T & Pfleiderer G 1955 Pyruvate kinase from muscle. Methods in Enzymology 1 435440.

  • Dahri S, Snoeck A, Reusens-Billen B, Remacle C & Hoet JJ 1991 Islet function in offspring of mothers on low-protein diet during gestation. Diabetes 40 (Supplement 2) 115120.

    • Search Google Scholar
    • Export Citation
  • Davis CM & Vincent JB 1997 Chromium oligopeptide activates insulin receptor tyrosine kinase activity. Biochemistry 36 43824385. doi:10.1021/bi963154t.

    • Search Google Scholar
    • Export Citation
  • Dostou J & Gerich J 2001 Pathogenesis of type 2 diabetes mellitus. Experimental and Clinical Endocrinology and Diabetes 109 (Suppl 2) S149S156. doi:10.1055/s-2001-18577.

    • Search Google Scholar
    • Export Citation
  • Gierow P & Jergil B 1982 Spectrophotometric method for glucose-6-phosphate phosphatase. Methods in Enzymology 89 4447.

  • Giugliano D, Ceriello A & Paolisso G 1995 Diabetes mellitus, hypertension, and cardiovascular disease: which role for oxidative stress? Methods 44 363368.

    • Search Google Scholar
    • Export Citation
  • Hissin PJ & Hilf R 1976 A fluorometric method for determination of oxidized and reduced glutathione in tissues. Analytical Biochemistry 74 214226. doi:10.1016/0003-2697(76)90326-2.

    • Search Google Scholar
    • Export Citation
  • Holness MJ 1996 The influence of sub-optimal protein nutrition on insulin hypersecretion evoked by high-energy/high-fat feeding in rats. FEBS Letters 396 5356. doi:10.1016/0014-5793(96)01067-8.

    • Search Google Scholar
    • Export Citation
  • Hostetler CE & Kincaid RL 2004 Maternal selenium deficiency increases hydrogen peroxide and total lipid peroxides in porcine fetal liver. Biological Trace Element Research 97 4356. doi:10.1385/BTER:97:1:43.

    • Search Google Scholar
    • Export Citation
  • Kitchalong L, Fernandez JM, Bunting LD, Southern LL & Bidner TD 1995 Influence of chromium tripicolinate on glucose metabolism and nutrient partitioning in growing lambs. Journal of Animal Science 73 26942705.

    • Search Google Scholar
    • Export Citation
  • Langley SC, Browne RF & Jackson AA 1994 Altered glucose tolerance in rats exposed to maternal low protein diets in utero. Comparative Biochemistry and Physiology. Physiology 109 223229. doi:10.1016/0300-9629(94)90124-4.

    • Search Google Scholar
    • Export Citation
  • Marcus F, Rittenhouse J, Chatterjee T & Hosey MM 1982 Fructose-1,6-bisphosphatase from rat liver. Methods in Enzymology 90 352357.

  • Marklund S & Marklund G 1974 Involvement of the superoxide anion radical in the autoxidation of pyrogallol and a convenient assay for superoxide dismutase. European Journal of Biochemistry 47 469474. doi:10.1111/j.1432-1033.1974.tb03714.x.

    • Search Google Scholar
    • Export Citation
  • Mathews JN, Altman DG, Campbell MJ & Royston P 1990 Analysis of serial measurements in medical research. British Medical Journal 300 230235. doi:10.1136/bmj.300.6719.230.

    • Search Google Scholar
    • Export Citation
  • Mertz W 1969 Chromium occurrence and function in biological systems. Physiological Reviews 49 163239.

  • Mertz W 1993 Chromium in human nutrition: a review. Journal of Nutrition 123 626633.

  • Mevorach M, Giacca A, Aharon Y, Hawkins M, Shamoon H & Rossetti L 1998 Regulation of endogenous glucose production by glucose per se is impaired in type 2 diabetes mellitus. Journal of Clinical Investigation 102 744753. doi:10.1172/JCI2720.

    • Search Google Scholar
    • Export Citation
  • Offenbacher EG & Pi-Sunyer FX 1980 Beneficial effect of chromium-rich yeast on glucose tolerance and blood lipids in elderly subjects. Diabetes 29 919925. doi:10.2337/diabetes.29.11.919.

    • Search Google Scholar
    • Export Citation
  • Padmavathi IJ, Kishore YD, Venu L, Ganeshan M, Harishankar N, Giridharan NV & Raghunath M 2009 Prenatal and perinatal zinc restriction: effects on body composition, glucose tolerance and insulin response in rat offspring. Experimental Physiology 94 761769. doi:10.1113/expphysiol.2008.045856.

    • Search Google Scholar
    • Export Citation
  • Padmavathi IJ, Rao KR, Venu L, Ganeshan M, Kumar KA, Rao ChN, Harishankar N, Ismail A & Raghunath M 2010a Chronic maternal dietary chromium restriction modulates visceral adiposity: probable underlying mechanisms. Diabetes 59 98104. doi:10.2337/db09-0779.

    • Search Google Scholar
    • Export Citation
  • Padmavathi IJ, Rao KR, Venu L, Ismail A & Raghunath M 2010b Maternal dietary chromium restriction programs muscle development and function in the rat offspring. Experimental Biology and Medicine 235 349355. doi:10.1258/ebm.2009.009199.

    • Search Google Scholar
    • Export Citation
  • Padmavathi IJ, Venu L & Raghunath M 2010c Response to comment on: Padmavathi et al. (2010) chronic maternal dietary chromium restriction modulates visceral adiposity: probable underlying mechanisms. Diabetes; 59: 98–104. Diabetes 59 e3 doi:10.2337/db10-0116.

    • Search Google Scholar
    • Export Citation
  • Paglia DE & Valentine WN 1967 Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. Journal of Laboratory and Clinical Medicine 70 158169.

    • Search Google Scholar
    • Export Citation
  • Pilkis SJ 1975 Glucokinase of rat liver. Methods in Enzymology 42 3139.

  • Ravina A, Siezak L, Rubal A & Mirsky N 1995 Clinical use of the trace element chromium (III) in the treatment of diabetes mellitus. Journal of Trace Elements in Experimental Medicine 8 183190.

    • Search Google Scholar
    • Export Citation
  • Reusens B & Remacle C 2001 Intergenerational effect of an adverse intrauterine environment on perturbation of glucose metabolism. Twin Research 4 406411. doi:10.1375/1369052012597.

    • Search Google Scholar
    • Export Citation
  • Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ & Klenk DC 1985 Measurement of protein using bicinchoninic acid. Analytical Biochemistry 150 7685. doi:10.1016/0003-2697(85)90442-7.

    • Search Google Scholar
    • Export Citation
  • Snoeck A, Remacle C, Reusens B & Hoet JJ 1990 Effect of a low protein diet during pregnancy on the fetal rat endocrine pancreas. Biology of the Neonate 57 107118. doi:10.1159/000243170.

    • Search Google Scholar
    • Export Citation
  • Soares MB, Schon E, Henderson A, Karathanasis SK, Cate R, Zeitlin S, Chirgwin J & Efstratiadis A 1985 RNA-mediated gene duplication: the rat preproinsulin I gene is a functional retroposon. Molecular and Cellular Biology 5 20902103.

    • Search Google Scholar
    • Export Citation
  • Uchida K, Kanematsu M, Sakai K, Matsuda T, Hattori N, Mizuno Y, Suzuki D, Miyata T, Noguchi N & Niki E 1998 Protein-bound acrolein: potential markers for oxidative stress. PNAS 95 48824887. doi:10.1073/pnas.95.9.4882.

    • Search Google Scholar
    • Export Citation
  • Urakawa H, Katsuki A, Sumida Y, Gabazza EC, Murashima S, Morioka K, Maruyama N, Kitagawa N, Tanaka T & Hori Y 2003 Oxidative stress is associated with adiposity and insulin resistance in men. Journal of Clinical Endocrinology and Metabolism 88 46734676. doi:10.1210/jc.2003-030202.

    • Search Google Scholar
    • Export Citation
  • Venu L, Harishankar N, Krishna TP & Raghunath M 2004a Does maternal dietary mineral restriction per se predispose the offspring to insulin resistance? European Journal of Endocrinology 151 287294. doi:10.1530/eje.0.1510287.

    • Search Google Scholar
    • Export Citation
  • Venu L, Harishankar N, Prasanna Krishna T & Raghunath M 2004b Maternal dietary vitamin restriction increases body fat content but not insulin resistance in WNIN rat offspring up to 6 months of age. Diabetologia 47 14931501. doi:10.1007/s00125-004-1506-4.

    • Search Google Scholar
    • Export Citation
  • Venu L, Kishore YD & Raghunath M 2005 Maternal and perinatal magnesium restriction predisposes rat pups to insulin resistance and glucose intolerance. Journal of Nutrition 135 13531358.

    • Search Google Scholar
    • Export Citation
  • Venu L, Padmavathi IJ, Kishore YD, Bhanu NV, Rao KR, Sainath PB, Ganeshan M & Raghunath M 2008 Long-term effects of maternal magnesium restriction on adiposity and insulin resistance in rat pups. Obesity 16 12701276. doi:10.1038/oby.2008.72.

    • Search Google Scholar
    • Export Citation
  • Vijayalingam S, Parthiban A, Shanmugasundaram KR & Mohan V 1996 Abnormal antioxidant status in impaired glucose tolerance and non-insulin-dependent diabetes mellitus. Diabetic Medicine 13 715719. doi:10.1002/(SICI)1096-9136(199608)13:8<715::AID-DIA172>3.0.CO;2-Z.

    • Search Google Scholar
    • Export Citation
  • Vincent JB 2000 The biochemistry of chromium. Journal of Nutrition 130 715718.

  • Vincent JB & Rasco JF 2010 Comment on: Padmavathi et al. (2010) chronic maternal dietary chromium restriction modulates visceral adiposity: probable underlying mechanisms. Diabetes; 59: 98–104. Diabetes 59 e2 doi:10.2337/db10-0043.

    • Search Google Scholar
    • Export Citation
  • Wentworth BM, Schaefer IM, Villa-Komaroff L & Chirgwin JM 1986 Characterization of the two nonallelic genes encoding mouse preproinsulin. Journal of Molecular Evolution 23 305312. doi:10.1007/BF02100639.

    • Search Google Scholar
    • Export Citation
  • Weyer C, Bogardus C, Mott DM & Pratley RE 1999 The natural history of insulin secretory dysfunction and insulin resistance in the pathogenesis of type 2 diabetes mellitus. Journal of Clinical Investigation 104 787794. doi:10.1172/JCI7231.

    • Search Google Scholar
    • Export Citation

*(I J N Padmavathi and K R Rao contributed equally to this work)

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    Effect of maternal Cr restriction and rehabilitation on glucose tolerance and plasma insulin levels in WNIN male offspring at different ages. Panel A, fasting glucose; panel B, fasting insulin; panel C, HOMA-IR; panel D, glucose AUC during OGTT; panel E, insulin AUC during OGTT; panel F, glucose AUC/insulin AUC during OGTT. Values are mean±s.e.m. (n=6). Bars without a common superscript (‘a and b’) are different at P<0.05 by one-way ANOVA followed by post hoc least significant difference (LSD) test.

  • View in gallery

    Effect of maternal Cr restriction and rehabilitation on plasma glucose and insulin levels at various time points (0, 30, 60, and 120 min) during oral glucose tolerance test in WNIN male offspring. Panel A, plasma glucose kinetics at 9 months of age; panel B, plasma glucose kinetics at 15 months of age; panel C, plasma insulin kinetics at 9 months of age; panel D, plasma insulin kinetics at 15 months of age. Values are mean (n=6).

  • View in gallery

    Effect of maternal Cr restriction and rehabilitation on glucose tolerance and plasma insulin levels in WNIN female offspring at different ages. Panel A, fasting glucose; panel B, fasting insulin; panel C, HOMA-IR; panel D, glucose AUC during OGTT; panel E, insulin AUC during OGTT; panel F, glucose AUC/insulin AUC during OGTT. Values are mean±s.e.m. (n=6). Bars without a common superscript (‘a and b’) are different at P<0.05 by one-way ANOVA followed by post hoc least significant difference (LSD) test.

  • View in gallery

    Effect of maternal Cr restriction and rehabilitation on plasma glucose and insulin levels at various time points (0, 30, 60, and 120 min) during OGTT in WNIN female offspring. Panel A, plasma glucose kinetics at 9 months of age; panel B, plasma glucose kinetics at 15 months of age; panel C, plasma insulin kinetics at 9 months of age; panel D, plasma insulin kinetics at 15 months of age. Values are mean (n=6).

  • View in gallery

    Effect of maternal Cr restriction and rehabilitation on expression of genes of glucose metabolism by semi-quantitative PCR in liver in WNIN male (A) offspring (at 18 months of age) and female (B) offspring (at 15 months of age); gel picture for each gene is the representation of different groups. Values are mean±s.e.m. (n=6).

  • View in gallery

    Effect of maternal Cr restriction and rehabilitation on expression of genes involved in insulin secretion by semi-quantitative PCR in pancreas in WNIN male (A) offspring (at 18 months of age) and female (B) offspring (at 15 months of age); gel picture for each gene is the representation of different groups. Values are mean±s.e.m. (n=6).

  • Aebi H 1984 Catalase in vitro. Methods in Enzymology 105 121126.

  • Amoikon EK, Fernandez JM, Southern LL, Thompson DL Jr, Ward TL & Olcott BM 1995 Effect of chromium tripicolinate on growth, glucose tolerance, insulin sensitivity, plasma metabolites, and growth hormone in pigs. Journal of Animal Science 73 11231130.

    • Search Google Scholar
    • Export Citation
  • Anderson RA 1989 Essentiality of chromium in humans. Science of the Total Environment 86 7581. doi:10.1016/0048-9697(89)90196-4.

  • Anderson RA 1992 Chromium, glucose tolerance, and diabetes. Biological Trace Element Research 32 1924. doi:10.1007/BF02784583.

  • Balasubramanian KA, Manohar M & Mathan VI 1988 An unidentified inhibitor of lipid peroxidation in intestinal mucosa. Biochimica et Biophysica Acta 962 5158. doi:10.1016/0005-2760(88)90094-X.

    • Search Google Scholar
    • Export Citation
  • Barker DJ, Hales CN, Fall CH, Osmond C, Phipps K & Clark PM 1993 Type 2 (non-insulin-dependent) diabetes mellitus, hypertension and hyperlipidaemia (syndrome X): relation to reduced fetal growth. Diabetologia 36 6267. doi:10.1007/BF00399095.

    • Search Google Scholar
    • Export Citation
  • Berney DM, Desai M, Palmer DJ, Greenwald S, Brown A, Hales CN & Berry CL 1997 The effects of maternal protein deprivation on the fetal rat pancreas: major structural changes and their recuperation. Journal of Pathology 183 109115. doi:10.1002/(SICI)1096-9896(199709)183:1<109::AID-PATH1091>3.0.CO;2-B.

    • Search Google Scholar
    • Export Citation
  • Bucher T & Pfleiderer G 1955 Pyruvate kinase from muscle. Methods in Enzymology 1 435440.

  • Dahri S, Snoeck A, Reusens-Billen B, Remacle C & Hoet JJ 1991 Islet function in offspring of mothers on low-protein diet during gestation. Diabetes 40 (Supplement 2) 115120.

    • Search Google Scholar
    • Export Citation
  • Davis CM & Vincent JB 1997 Chromium oligopeptide activates insulin receptor tyrosine kinase activity. Biochemistry 36 43824385. doi:10.1021/bi963154t.

    • Search Google Scholar
    • Export Citation
  • Dostou J & Gerich J 2001 Pathogenesis of type 2 diabetes mellitus. Experimental and Clinical Endocrinology and Diabetes 109 (Suppl 2) S149S156. doi:10.1055/s-2001-18577.

    • Search Google Scholar
    • Export Citation
  • Gierow P & Jergil B 1982 Spectrophotometric method for glucose-6-phosphate phosphatase. Methods in Enzymology 89 4447.

  • Giugliano D, Ceriello A & Paolisso G 1995 Diabetes mellitus, hypertension, and cardiovascular disease: which role for oxidative stress? Methods 44 363368.

    • Search Google Scholar
    • Export Citation
  • Hissin PJ & Hilf R 1976 A fluorometric method for determination of oxidized and reduced glutathione in tissues. Analytical Biochemistry 74 214226. doi:10.1016/0003-2697(76)90326-2.

    • Search Google Scholar
    • Export Citation
  • Holness MJ 1996 The influence of sub-optimal protein nutrition on insulin hypersecretion evoked by high-energy/high-fat feeding in rats. FEBS Letters 396 5356. doi:10.1016/0014-5793(96)01067-8.

    • Search Google Scholar
    • Export Citation
  • Hostetler CE & Kincaid RL 2004 Maternal selenium deficiency increases hydrogen peroxide and total lipid peroxides in porcine fetal liver. Biological Trace Element Research 97 4356. doi:10.1385/BTER:97:1:43.

    • Search Google Scholar
    • Export Citation
  • Kitchalong L, Fernandez JM, Bunting LD, Southern LL & Bidner TD 1995 Influence of chromium tripicolinate on glucose metabolism and nutrient partitioning in growing lambs. Journal of Animal Science 73 26942705.

    • Search Google Scholar
    • Export Citation
  • Langley SC, Browne RF & Jackson AA 1994 Altered glucose tolerance in rats exposed to maternal low protein diets in utero. Comparative Biochemistry and Physiology. Physiology 109 223229. doi:10.1016/0300-9629(94)90124-4.

    • Search Google Scholar
    • Export Citation
  • Marcus F, Rittenhouse J, Chatterjee T & Hosey MM 1982 Fructose-1,6-bisphosphatase from rat liver. Methods in Enzymology 90 352357.

  • Marklund S & Marklund G 1974 Involvement of the superoxide anion radical in the autoxidation of pyrogallol and a convenient assay for superoxide dismutase. European Journal of Biochemistry 47 469474. doi:10.1111/j.1432-1033.1974.tb03714.x.

    • Search Google Scholar
    • Export Citation
  • Mathews JN, Altman DG, Campbell MJ & Royston P 1990 Analysis of serial measurements in medical research. British Medical Journal 300 230235. doi:10.1136/bmj.300.6719.230.

    • Search Google Scholar
    • Export Citation
  • Mertz W 1969 Chromium occurrence and function in biological systems. Physiological Reviews 49 163239.

  • Mertz W 1993 Chromium in human nutrition: a review. Journal of Nutrition 123 626633.

  • Mevorach M, Giacca A, Aharon Y, Hawkins M, Shamoon H & Rossetti L 1998 Regulation of endogenous glucose production by glucose per se is impaired in type 2 diabetes mellitus. Journal of Clinical Investigation 102 744753. doi:10.1172/JCI2720.

    • Search Google Scholar
    • Export Citation
  • Offenbacher EG & Pi-Sunyer FX 1980 Beneficial effect of chromium-rich yeast on glucose tolerance and blood lipids in elderly subjects. Diabetes 29 919925. doi:10.2337/diabetes.29.11.919.

    • Search Google Scholar
    • Export Citation
  • Padmavathi IJ, Kishore YD, Venu L, Ganeshan M, Harishankar N, Giridharan NV & Raghunath M 2009 Prenatal and perinatal zinc restriction: effects on body composition, glucose tolerance and insulin response in rat offspring. Experimental Physiology 94 761769. doi:10.1113/expphysiol.2008.045856.

    • Search Google Scholar
    • Export Citation
  • Padmavathi IJ, Rao KR, Venu L, Ganeshan M, Kumar KA, Rao ChN, Harishankar N, Ismail A & Raghunath M 2010a Chronic maternal dietary chromium restriction modulates visceral adiposity: probable underlying mechanisms. Diabetes 59 98104. doi:10.2337/db09-0779.

    • Search Google Scholar
    • Export Citation
  • Padmavathi IJ, Rao KR, Venu L, Ismail A & Raghunath M 2010b Maternal dietary chromium restriction programs muscle development and function in the rat offspring. Experimental Biology and Medicine 235 349355. doi:10.1258/ebm.2009.009199.

    • Search Google Scholar
    • Export Citation
  • Padmavathi IJ, Venu L & Raghunath M 2010c Response to comment on: Padmavathi et al. (2010) chronic maternal dietary chromium restriction modulates visceral adiposity: probable underlying mechanisms. Diabetes; 59: 98–104. Diabetes 59 e3 doi:10.2337/db10-0116.

    • Search Google Scholar
    • Export Citation
  • Paglia DE & Valentine WN 1967 Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. Journal of Laboratory and Clinical Medicine 70 158169.

    • Search Google Scholar
    • Export Citation
  • Pilkis SJ 1975 Glucokinase of rat liver. Methods in Enzymology 42 3139.

  • Ravina A, Siezak L, Rubal A & Mirsky N 1995 Clinical use of the trace element chromium (III) in the treatment of diabetes mellitus. Journal of Trace Elements in Experimental Medicine 8 183190.

    • Search Google Scholar
    • Export Citation
  • Reusens B & Remacle C 2001 Intergenerational effect of an adverse intrauterine environment on perturbation of glucose metabolism. Twin Research 4 406411. doi:10.1375/1369052012597.

    • Search Google Scholar
    • Export Citation
  • Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ & Klenk DC 1985 Measurement of protein using bicinchoninic acid. Analytical Biochemistry 150 7685. doi:10.1016/0003-2697(85)90442-7.

    • Search Google Scholar
    • Export Citation
  • Snoeck A, Remacle C, Reusens B & Hoet JJ 1990 Effect of a low protein diet during pregnancy on the fetal rat endocrine pancreas. Biology of the Neonate 57 107118. doi:10.1159/000243170.

    • Search Google Scholar
    • Export Citation
  • Soares MB, Schon E, Henderson A, Karathanasis SK, Cate R, Zeitlin S, Chirgwin J & Efstratiadis A 1985 RNA-mediated gene duplication: the rat preproinsulin I gene is a functional retroposon. Molecular and Cellular Biology 5 20902103.

    • Search Google Scholar
    • Export Citation
  • Uchida K, Kanematsu M, Sakai K, Matsuda T, Hattori N, Mizuno Y, Suzuki D, Miyata T, Noguchi N & Niki E 1998 Protein-bound acrolein: potential markers for oxidative stress. PNAS 95 48824887. doi:10.1073/pnas.95.9.4882.

    • Search Google Scholar
    • Export Citation
  • Urakawa H, Katsuki A, Sumida Y, Gabazza EC, Murashima S, Morioka K, Maruyama N, Kitagawa N, Tanaka T & Hori Y 2003 Oxidative stress is associated with adiposity and insulin resistance in men. Journal of Clinical Endocrinology and Metabolism 88 46734676. doi:10.1210/jc.2003-030202.

    • Search Google Scholar
    • Export Citation
  • Venu L, Harishankar N, Krishna TP & Raghunath M 2004a Does maternal dietary mineral restriction per se predispose the offspring to insulin resistance? European Journal of Endocrinology 151 287294. doi:10.1530/eje.0.1510287.

    • Search Google Scholar
    • Export Citation
  • Venu L, Harishankar N, Prasanna Krishna T & Raghunath M 2004b Maternal dietary vitamin restriction increases body fat content but not insulin resistance in WNIN rat offspring up to 6 months of age. Diabetologia 47 14931501. doi:10.1007/s00125-004-1506-4.

    • Search Google Scholar
    • Export Citation
  • Venu L, Kishore YD & Raghunath M 2005 Maternal and perinatal magnesium restriction predisposes rat pups to insulin resistance and glucose intolerance. Journal of Nutrition 135 13531358.

    • Search Google Scholar
    • Export Citation
  • Venu L, Padmavathi IJ, Kishore YD, Bhanu NV, Rao KR, Sainath PB, Ganeshan M & Raghunath M 2008 Long-term effects of maternal magnesium restriction on adiposity and insulin resistance in rat pups. Obesity 16 12701276. doi:10.1038/oby.2008.72.

    • Search Google Scholar
    • Export Citation
  • Vijayalingam S, Parthiban A, Shanmugasundaram KR & Mohan V 1996 Abnormal antioxidant status in impaired glucose tolerance and non-insulin-dependent diabetes mellitus. Diabetic Medicine 13 715719. doi:10.1002/(SICI)1096-9136(199608)13:8<715::AID-DIA172>3.0.CO;2-Z.

    • Search Google Scholar
    • Export Citation
  • Vincent JB 2000 The biochemistry of chromium. Journal of Nutrition 130 715718.

  • Vincent JB & Rasco JF 2010 Comment on: Padmavathi et al. (2010) chronic maternal dietary chromium restriction modulates visceral adiposity: probable underlying mechanisms. Diabetes; 59: 98–104. Diabetes 59 e2 doi:10.2337/db10-0043.

    • Search Google Scholar
    • Export Citation
  • Wentworth BM, Schaefer IM, Villa-Komaroff L & Chirgwin JM 1986 Characterization of the two nonallelic genes encoding mouse preproinsulin. Journal of Molecular Evolution 23 305312. doi:10.1007/BF02100639.

    • Search Google Scholar
    • Export Citation
  • Weyer C, Bogardus C, Mott DM & Pratley RE 1999 The natural history of insulin secretory dysfunction and insulin resistance in the pathogenesis of type 2 diabetes mellitus. Journal of Clinical Investigation 104 787794. doi:10.1172/JCI7231.

    • Search Google Scholar
    • Export Citation