Nisha G. Kemse

A Combined Supplementation of Omega-3 Fatty Acids and Micronutrients (Folic Acid, Vitamin B12) Reduces Oxidative Stress Markers in a Rat Model of Pregnancy Induced Hypertension

Objectives Our earlier studies have highlighted that an altered one carbon metabolism (vitamin B12, folic acid, and docosahexaenoic acid) is associated with preeclampsia. Preeclampsia is also known to be associated with oxidative stress and inflammation. The current study examines whether maternal folic acid, vitamin B12 and omega-3 fatty acid supplementation given either individually or in combination can ameliorate the oxidative stress markers in a rat model of pregnancy induced hypertension (PIH). Materials and Methods Pregnant Wistar rats were assigned to control and five treatment groups: PIH; PIH + vitamin B12; PIH + folic acid; PIH + Omega-3 fatty acids and PIH + combined micronutrient supplementation (vitamin B12 + folic acid + omega-3 fatty acids). L-Nitroarginine methylester (L-NAME; 50 mg/kg body weight/day) was used to induce hypertension during pregnancy. Blood Pressure (BP) was recorded during pregnancy and dams were dissected at d20 of gestation. Results Animals from the PIH group demonstrated higher (p<0.01 for both) systolic and diastolic BP; lower (p<0.01) pup weight; higher dam plasma homocysteine (p<0.05) and dam and offspring malondialdehyde (MDA) (p<0.01), lower (p<0.05) placental and offspring liver DHA and higher (p<0.01) tumor necrosis factor–alpha (TNF–ά) levels as compared to control. Individual micronutrient supplementation did not offer much benefit. In contrast, combined supplementation lowered systolic BP, homocysteine, MDA and placental TNF-ά levels in dams and liver MDA and protein carbonyl in the offspring as compared to PIH group. Conclusion Key constituents of one carbon cycle (folic acid, vitamin B12 and DHA) may play a role in reducing oxidative stress and inflammation in preeclampsia.

Maternal omega-3 fatty acid supplementation on vitamin B12 rich diet improves brain omega-3 fatty acids, neurotrophins and cognition in the Wistar rat offspring

INTRODUCTION The consequences of wide spread vegetarianism due to low vitamin B12 on brain development and functioning is gaining importance. However, there are no studies which have evaluated exclusively vitamin B12 supplementation during pregnancy on brain growth. A series of our animal studies have documented adverse effects of maternal micronutrient imbalance on brain neurotrophins and its amelioration by omega-3 fatty acids. Therefore, the present study investigated the effect of maternal supplementation with vitamin B12 alone and B12 plus omega-3 fatty acid on pup brain fatty acids and neurotrophins at birth and 3 mo of age. METHODS AND RESULTS Pregnant Wistar rats and their male offspring were assigned to 3 dietary groups: Control (normal vitamin B12 (25 μg/kg), vitamin B12 supplemented (BS) (50 μg/kg), vitamin B12 supplemented with omega-3 fatty acid (BSO) till 3 month of age. Maternal vitamin B12 supplementation (BS) increased brain BDNF (protein and mRNA) and DHA levels in pups at birth and in the hippocampus at 3 month of age (BDNF only). These effects were further enhanced by omega-3 fatty acid supplementation to vitamin B12 supplemented group. The spatial memory performance was found to be enhanced in BSO group which was characterised by less number of errors in radial eight arm maze. CONCLUSION Our results indicate that a combination of omega-3 fatty acid and vitamin B12 enriched diet may exert beneficial effects on synaptic plasticity and cognition, which may prove beneficial for mental health, particularly in preventing neurocognitive disorders.

Supplementation of maternal omega-3 fatty acids to pregnancy induced hypertension Wistar rats improves IL10 and VEGF levels.

OBJECTIVES Our recent study demonstrates the beneficial effect of a combined supplementation of vitamin B12, folic acid, and docosahexaenoic acid in reducing the severity of pregnancy induced hypertension (PIH). It is also known to be associated with angiogenic imbalance and inflammation. The current study examines whether the individual/combined supplementation of folic acid, vitamin B12 and omega-3 fatty acid during pregnancy can ameliorate the inflammatory markers and restore the angiogenic balance in a rat model of PIH. MATERIALS AND METHODS There were total of six groups, control and five treatment groups: PIH Induced; PIH+vitamin B12; PIH+folic acid; PIH+Omega-3 fatty acids and PIH+combined micronutrient supplementation (vitamin B12+folic acid+omega-3 fatty acids). Hypertension during pregnancy was induced using L- Nitroarginine methylester (L-NAME; 50mg/kg body weight/day). Dams were dissected at d20 of gestation and placental tissues were collected for further analysis. RESULTS Animals from the PIH induced group demonstrated lower (p<0.01 for both) IL-10 and VEGF levels as compared to control. However, PIH induction did not alter the protein levels of eNOS, IL-6, Flt and mRNA levels of VEGF and VEGFR-1/ Flt-1. Individual micronutrient supplementation of vitamin B12 and folate did not offer benefit. In contrast individual omega-3 fatty acid as well as combined micronutrient supplementation showed IL-10 and VEGF levels comparable to that of control. CONCLUSION Omega 3 fatty acid supplementation plays a key role in reducing inflammation in pregnancy induced hypertension.

Supplementation with omega-3 fatty acids during gestation and lactation to a vitamin B12-deficient or -supplemented diet improves pregnancy outcome and metabolic variables in Wistar rats

Maternal vitamin B12 deficiency leads to an adverse pregnancy outcome and increases the risk for developing diabetes and metabolic syndrome in mothers in later life. Our earlier studies have demonstrated that vitamin B12 and n-3 polyunsaturated fatty acids (PUFA) are interlinked in the one carbon cycle. The present study for the first time examines the effect of maternal n-3 PUFA supplementation to vitamin B12 deficient or supplemented diets on pregnancy outcome, fatty-acid status and metabolic variables in Wistar rats. Pregnant dams were assigned to one of the following groups: control, vitamin B12 deficient, vitamin B12 supplemented, vitamin B12 deficient + n-3 PUFA or vitamin B12 supplemented + n-3 PUFA. The amount of vitamin B12 in the supplemented group was 0.50 μg kg(-1) diet and n-3 PUFA was alpha linolenic acid (ALA) 1.68, eicosapentaenoic acid 5.64, docosahexaenoic acid (DHA) 3.15 (g per 100g fatty acids per kg diet). Our findings indicate that maternal vitamin B12 supplementation did not affect the weight gain of dams during pregnancy but reduced litter size and weight and was ameliorated by n-3 PUFA supplementation. Vitamin B12 deficiency or supplementation resulted in a low percentage distribution of plasma arachidonic acid and DHA. n-3 PUFA supplementation to these diets improved the fatty-acid status. Vitamin B12 deficiency resulted in higher homocysteine and insulin levels, which were normalised by supplementation with either vitamin B12 or n-3 PUFA. Our study suggests that maternal vitamin B12 status is critical in determining pregnancy outcome and metabolic variables in dams and that supplementation with n-3 PUFA is beneficial.

Nutraceutical properties of dietary plants extracts: Prevention of diabetic nephropathy through inhibition of glycation and toxicity to erythrocytes and HEK293 cells

Abstract Context: Glycated albumin is reported to elicit pathobiologic effects in diabetic nephropathy and abrogating its biologic effects has novel therapeutic potential. Objective: This study examines the effects of dietary plants extracts (Laurus nobilis, Carum carvi, Coccinia grandis, Mentha arvensis, Phaseolus vulgaris) against albumin glycation and its toxicity to erythrocytes and HEK293 cells. Materials and methods: Albumin (10 mg/ml) was incubated with fructose (250 mM) in PBS along with aqueous plant extracts (1% w/v) for 4 d. After incubation, the antiglycation potential of extracts was estimated by measuring AGEs, fructosamine, amyloids, carbonyls, free amino groups, and antioxidant potential of albumin. The glycation extent of the treated samples was determined by boronate affinity chromatography. Effect of extracts against glycation induced cytotoxicity in erythrocytes and HEK 293 cells was assessed by estimating viability, glutathione, and antioxidant capacity. Plant extracts were tested for their phenolic content and antioxidant potential (reducing potential, DPPH, ABTS, NO, and H2O2 radical scavenging activities). Results: Plant extracts significantly decreased the AGEs formation and amyloid aggregation in glycated BSA (p < 0.001). Further, fructosamine and carbonyls were reduced to 55–72% and 83–89%, respectively. Free amino group and antioxidant activity of albumin were also preserved by 1.25–1.40-fold and 1.75–1.8-fold, respectively. Further, co-incubation of extracts with glycated albumin, protected erythrocytes, and HEK293 cells as they inhibited cellular hemolysis/toxicity (p < 0.001) by upregulating cellular antioxidants. Discussion and conclusion: Plant co-incubation reversed many modifications in albumin glycation, cellular dysfunction indicating that dietary sources with antiglycating and antioxidant potential could be considered for the effective management of diabetic nephropathy.

Aqueous extract of some indigenous medicinal plants inhibits glycation at multiple stages and protects erythrocytes from oxidative damage–an in vitro study

Azadirachta indica, Emblica officinalis, Syzygium cumini and Terminalia bellirica are common in Indian system of traditional medicine for the prevention of diabetes and its complications. The aim of the present study was to comprehensively and comparatively investigate the antiglycation potential of these plant extracts at multiple stages and their possible protective effect against glycated albumin mediated toxicity to erythrocytes. Antiglycation activities of these plant extracts was measured by co-incubation of plant extract with bovine serum albumin-fructose glycation model. The multistage glycation markers- fructosamines (early stage), protein carbonyls (intermediate stage) and AGEs (late stage) are investigated along with measurement of thiols and β aggregation of albumin using amyloid-specific dyes–Congo red and Th T. Protection of erythrocytes from glycated albumin induced toxicity by these plant extracts was assessed by measuring erythrocytes hemolysis, lipid peroxidation, reduced glutathione and intracellular antioxidant capacity. Total phenolics, reducing power and antioxidant activities of the plant extracts were also measured. In vitro glycation assays showed that plant extracts exerted site specific inhibitory effects at multiple stages, with T. bellirica showing maximum attenuation. In erythrocytes, along with the retardation of glycated albumin induced hemolysis and lipid-peroxidation, T. bellirica considerably maintained cellular antioxidant potential. Significant positive correlations were observed between erythrocyte protection parameters with total phenolics. These plant extracts especially T. bellirica prevents glycation induced albumin modifications and subsequent toxicity to erythrocytes which might offer additional protection against diabetic vascular complications.

Attenuation of glycation-induced multiple protein modifications by Indian antidiabetic plant extracts

Abstract Context: Protein glycation is the major contributing factor in the development of diabetic complications. The antiglycation potential of medicinal plants provides a promising opportunity as complementary interventions for complications. Objective: To investigate the antiglycation potential of 19 medicinal plants extracts using albumin by estimating different indicators: (1) glycation (early and late), (2) albumin oxidation, and (3) amyloid aggregation. Materials and methods: The effect of aqueous plant extracts (1% w/v) on protein glycation was assessed by incubating albumin (10 mg/mL) with fructose (250 mM) for 4 days. Degree of protein glycation in the absence and presence of plant extracts was assessed by estimating fructosamine, advanced glycation end products (AGEs), carbonyls, free thiol group and β-amyloid aggregation. Results: Petroselinum crispum, Boerhavia diffusa, Terminalia chebula, Swertia chirayita and Glycyrrhiza glabra showed significant antiglycating activity. P. crispum and A. barbadensis inhibited the carbonyl stress and protected the thiol group from oxidative damage. There was significant correlation between protein thiols and amyloid inhibition (R = −.69, p < .001). Conclusion: P. crispum, B. diffusa and T. chebula had the most potent antiglycation activity. These plant exerted noticeable antiglycation activity at different glycation modifications of albumin. These findings are important for identifying plants with potential to combat diabetic complications.

Altered breast milk components in preeclampsia; An in-vitro proton NMR spectroscopy study.

OBJECTIVE To investigate the metabolic profile of milk on day 3 and at the 6th month of lactation in mothers with preeclampsia (PE) and normotensive mothers. STUDY DESIGN Women with PE (n=29) and control women (n=31) were recruited for this study. Milk was collected on day 3 and at the 6th month of lactation. Proton NMR spectroscopy was used to identify 25 milk metabolites (alpha-lactose, beta-lactose, oligosaccharides, myo-inositol, alanine, glutamate, glutamine, glycine, histidine, isoleucine, leucine, lysine, phenylalanine, tyrosine, valine, acetone, citrate, creatine, phosphocreatine, acetate, choline, lactate, lipid, phosphocholine and glycerophosphocholine). Principle component analysis (PCA) and Partial Least Square Discriminant Analysis (PLS-DA) were carried out to identify differences in milk metabolite composition between both the groups. RESULTS The levels of milk metabolites varied between the control and PE groups. Alpha and beta-lactose, glycine, glycerophosphocholine (p<0.01 for all); glutamate, glutamine and phosphocholine levels (p<0.05 for all) were increased at the 6th month as compared to day 3 of lactation in the control group. However, in the PE group, only glycerophosphocholine level showed an increase (p<0.01) at the 6th month. The levels of acetate, acetone (p<0.05 for both) and creatine (p<0.01) decreased at the 6th month as compared to day 3 of lactation in both groups. However, the levels of oligosaccharides were similar between groups and also similar at day 3 and at the 6th month of lactation. CONCLUSION Our data indicates differential levels of metabolites in the milk of women with PE. Future studies are required to investigate the associations between milk components and infant growth and development.