Repurposing of genistein as anti-sickling agent: elucidation by multi spectroscopic, thermophoresis, and molecular modeling techniques.

Sickle cell disease (SCD) is a major medical problem in which mono-therapeutic interventions have so far shown only limited effectiveness. We studied the repurpose of genistein, which could prevent sickle hemoglobin from polymerizing under hypoxic conditions in this disease. Genistein an important nutraceutical molecule found in soybean. The present study examines the repurposing genistein as an anti- sickling agent. Genistein shows inhibition of Hb S polymerization as well as a sickle reversal. Also, we have explored the interaction of the genistein with sickle hemoglobin (Hb S), using fluorescence, far-UV-CD spectroscopy, MicroScale Thermophoresis (MST), FTIR, combined with molecular modeling computations. The quenching constant decreases with increasing temperature, a characteristic that coincides with the static type of quenching mechanism. Temperature-dependent fluorescence measurements and molecular modeling studies reveal that apart from the hydrogen bonding, electrostatic interactions also play a crucial role in genistein and Hb S complex formation. In silico, distribution prediction of adsorption, digestion, metabolism, excretion, and toxicity (ADME/Tox) based on physical and chemical properties show that genistein is nontoxic and has ideal drug properties. The helicity and thermophoretic mobility of Hb S was a change in the presence of genistein, which leads to the destabilizing the Hb S polymer was examined using CD and MST, respectively. Our results open up the possibility for a promising therapeutic approach for the SCD by repurposed genistein as an anti-sickling agent.Communicated by Ramaswamy H. Sarma.

Potential of isoquercitrin as antisickling agent: a multi-spectroscopic, thermophoresis and molecular modeling approach.

Sickle cell disease is an inherited disease caused by point mutation in hemoglobin (ß-globin gene). Under oxygen saturation, sickle hemoglobin form polymers, leading to rigid erythrocytes. The transition of the blood vessels is altered and initiated by the adhesion of erythrocytes, neutrophils and endothelial cells. Sickle Hemoglobin (HbS) polymerization is a major cause in red blood cells (RBC), promoting sickling and destruction of RBCs. Isoquercitrin, a medicinal bioactive compound found in various medicinal plants, has multiple health benefits. The present study examines the potential of isoquercitrin as an anti-sickle agent, showing a significant decrease in the rate of polymerization as well as sickling of RBCs. Isoquercitrin-induced graded alteration in absorbance and fluorescence of HbS, confirmed their interaction. A negative value of ΔG° strongly suggests that it is a spontaneous exothermic reaction induced by entropy. Negative ΔH° and positive ΔS° predicted that hydrogen and hydrophobic binding forces interfered with a hydrophobic microenvironment of ß6Val leading to polymerization inhibition of HbS. HbS-Isoquercitrin complex exhibits helical structural changes leading to destabilization of the HbS polymer as confirmed by CD spectroscopy. MST and DSC results indicate greater changes in thermophoretic mobility and thermal stability of sickle hemoglobin in the presence of isoquercitrin, respectively. These findings were also supported by molecular simulation studies using DOCK6 and GROMACS. Hence, we can conclude that isoquercitrin interacts with HbS through hydrogen bonding, which leads to polymerization inhibition. Consequently, isoquercitrin could potentially be used as a medication for the treatment of sickle cell disease.Communicated by Ramaswamy H. Sarma.

Alizarin interaction with sickle hemoglobin: elucidation of their anti-sickling properties by multi-spectroscopic and molecular modeling techniques.

Polymerization of hemoglobin S is a major cause of morbidity and mortality in sickle cell disease, which leads to sickling and destruction of red blood cell. Alizarin, a bioactive compound from Rubia cordifolia, is reported to be blood purifier. This study investigates the potential of alizarin as an anti-sickling agent, showing a significant decrease in the rate of polymerization, therefore inhibiting the rate of sickling with increasing concentration. Interaction studies indicated that the fluorescence intensity of sickle hemoglobin (Hb S) decreases gradually with increasing alizarin concentration. This suggests the static quenching, where binding constant and the number of binding sites were deduced at different temperatures. The negative values of Gibbs energy change (ΔG0) strongly suggest that it is entropy-driven spontaneous and exothermic reaction. Negative enthalpy (ΔH0) and positive entropy (ΔS0) stipulated that hydrogen and hydrophobic bonding forces were interfering in a hydrophobic micro-environment of ß6Val leading to Hb S polymerization inhibition. In circular dichroism (CD) spectra, Hb S in the presence of alizarin shows helical structural changes leading to destabilization of Hb S polymer. These findings were also supported by molecular docking simulation studies using DOCK6 and GROMACS. So, from these findings, we may conclude that alizarin interacts with Hb S through hydrogen bonding and leading to inhibition of Hb S polymerization. Consequently, alizarin may have potential use as an anti-sickle cell medication for sickle cell disorder. Communicated by Ramaswamy H. Sarma.

HPLC, NMR based chemical profiling and biological characterisation of Indian propolis.

The present study aimed to investigate chemical profile, antioxidant and antimicrobial activities of Indian Melifera propolis (IMP) samples collected from 13 different states. Chemical characterisation of ethanolic extracts of IMP (EEMP) samples was carried out by using HPLC and 1HNMR spectroscopy. The antioxidant activity of EEMP samples was measured by DPPH, ABTS, and FRAP assay. Moreover, the antimicrobial activity of each EEMP sample tested against bacteria and yeast using a 96 well plate microdilution method. All EEMP samples had remarkable antioxidant and antimicrobial activities. The antioxidant potential of EEMP samples found to have a moderate positive correlation with their total phenolics and flavonoids content. Majority of EEMP samples had a minimum inhibitory concentration (MIC) ≤1mg/mL against Staphylococcus aureus. Chemometric analysis of 1HNMR data indicated that brown, green, green-brown, red and red-brown coloured IMP samples were chemically distinct from each other, and showed two separate clusters for northern and southern states propolis samples. HPLC analysis confirmed phenethyl caffeate was most common and abundant compound in IMP samples among studied compounds. In conclusion, this study may be helpful for defining the quality of IMP as a raw material, and also in finished food and health care products.

Association of metabolic syndrome with severity of coronary artery disease.

BACKGROUND: South Asians are more prone to develop metabolic syndrome (MetS). The additive predictive value of components of MetS for cardiovascular diseases is still debated. We undertook this study to evaluate the association of MetS and its components with severity of coronary artery disease (CAD). MATERIALS AND METHODS: Three hundred patients with known coronary disease above the age of 25 years were included in this study. Blood samples were collected for biochemical markers. Patients were stratified into subjects with and without MetS (International Diabetes Federation, IDF, criteria) and severity of CAD (number of vessel involved). RESULTS: Mean age of the patient in the study was 60.9 ± 12.4 years (male, M: 72%; female, F: 28%). MetS was present in 64% patients. Patients with MetS had more severe CAD compared to those without MetS. Triple vessel disease (TVD) was present in 62.5% of patients with MetS compared to 34.3% among without MetS (P < 0.0001). The percent number of patients with TVD showed increasing trend with increasing number of components of MetS (0-0%; 1-20%; 2-27.5%; 3-47.8%; 4-72.6%; 5-78.3%; Chi square for trend < 0.0001). Inflammatory markers [interleukin (IL) 6: 77.67 ± 79.48 vs. 41.21 ± 60.72 pg/ml, P < 0.0001; tumor nuclear factor (TNF)-α: 28.0 ± 47.49 vs 20.43 ± 24.5 pg/ml, P < 0.0001; high sensitive C-reactive protein (hsCRP): 14.30 ± 9.91 vs. 7.02 ± 7.18 mg/L, P < 0.0001], insulin resistance [homeostatic model analysis insulin resistance (HOMA-IR): 22.33 ± 23.37 vs. 10.86 ± 13.90, P < 0.0001] were higher and insulin sensitivity [quantitative insulin check index (QUICKI): 0.26 ± 0.03 vs. 0.30 ± 0.04, P < 0.0001] was significantly lower in subjects with MetS compared to subjects without MetS. Among lipids, total cholesterol were comparable but triglyceride (175 ± 42 vs. 179 ± 48 vs. 180 ± 47 mg/dl, P < 0.0001) was high and high-density lipoprotein (HDL; 44.72 ± 7.63 vs. 39.96 ± 8.70 vs. 36.05 ± 8.84, P < 0.0001) was low in subjects with TVD compared to others. Similarly, percentage of patients with diabetes (7.5% vs. 26.3% vs. 63.7%, P < 0.0001) and hypertension (34.3% vs. 56.6% vs. 77.7%, P < 0.0001) were higher in subjects with TVD compared to others. CONCLUSIONS: There is a strong correlation of MetS and its components with severity of CAD.

Study of pattern of dyslipidemia and its correlation with cardiovascular risk factors in patients with proven coronary artery disease.

INTRODUCTION: Dyslipidemia is a primary, widely established as an independent major risk factor for coronary artery disease (CAD). Asians differs in prevalence of various lipid abnormalities than non-Asians. Hence, this study was conducted with objective to evaluate the lipid abnormalities and there correlation with traditional and non-traditional risk factors in known subjects with CAD. MATERIALS AND METHODS: We studied the pattern and association of dyslipidemia with cardiovascular risk factors in 300 (Male: 216; Female: 84, age: 60.9 ± 12.4 years, range: 25-92 years) angiographically proved CAD patients. All patients were evaluated for anthropometry and cardiovascular risk factors and blood samples were collected for biochemical and inflammatory markers. RESULTS: Hypercholesterolemia, hypertriglyceridemia and low high density lipoprotein (HDL) was present in 23.3%, 63.0% and 54.6% in the total study population respectively. A total of 41.3% had atherogenic dyslipidemia (raised triglycerides [TG] and low HDL). Percentage of patients with type-2 diabetes mellitus and hypertension were higher in subjects with atherogenic dyslipidemia. Insulin sensitivity was low; insulin and insulin resistance (IR) along with inflammatory markers were high in subjects with atherogenic dyslipidemia. Patients with atherogenic dyslipidemia had significantly lower serum vitamin B12 levels and higher homocysteine (Hcy) levels. Hypertriglyceridemia was positively correlated with insulin, homeostasis model assessment of IR, Hcy, interleukin-6, Tumor necrosis factor-alpha, highly sensitive C-reactive protein and negatively with vitamin B12 and quantitative insulin check index and an opposite correlation of all quoted parameters was observed with low HDL. The correlation of traditional and non-traditional risk factors was stronger with low HDL and high TG compared with hypercholesterolemia. CONCLUSIONS: Hypertriglyceridemia and low HDL cholesterol is common in patients with CAD compared with hypercholesterolemia. This suggests that different preventive strategy is required in Indian patients with CAD.

Differences in traditional and non-traditional risk factors with special reference to nutritional factors in patients with coronary artery disease with or without diabetes mellitus.

INTRODUCTION: There is an increase in awareness about the role of nutritional factors in chronic non-communicable diseases. We therefore conducted this study with an aim to assess the relationship between nutritional factor (vitamin B12 and homocysteine [Hcy]) and its association with insulin resistance and inflammatory markers, and differences in traditional and non-traditional risk factors among diabetics and non-diabetics in known cases of coronary artery disease (CAD). MATERIALS AND METHODS: Three hundred consecutive patients with known coronary disease on coronary angiography, who were >25 years old were included in this study. All cases were interviewed using a questionnaire. Blood samples were analyzed for insulin, vitamin B12, Hcy and inflammatory markers (highly sensitive C-reactive protein [hsCRP], interleukin-6 [IL-6], Tumor necrosis factor-alfa [TNF-α]). Insulin resistance was calculated with homeostasis model assessment of insulin resistance (HOMA-IR). RESULTS: Mean age of the patients was 60.95 ± 12.3 years. Body mass index and waist hip ratio were comparable in both groups. Triglyceride, very low-density lipoprotein and HbA1C were significantly higher and high-density lipoprotein (HDL) was significantly lower in patients with diabetes. Patients with diabetes had significantly high levels of IL-6, hsCRP and TNF-α compared with non-diabetic patients. Insulin resistance was twofold higher in diabetic patients. Serum vitamin B12 levels were significantly lower and Hcy was significantly higher in the diabetic group compared with the non-diabetic patients. HbA1C, HOMA-IR and Hcy levels were positively correlated with inflammatory markers in the total study population and in the non-diabetic patients; but, in diabetic patients, HbA1C and Hcy showed this relation. CONCLUSIONS: Vitamin B12 deficiency is common in the diabetic population. Hcy levels were higher in diabetics compared with non-diabetics, and were related to glycemic level and insulin resistance in diabetic patients. Patients with diabetes had higher traditional risk factors than patients without diabetes in known patients with CAD. Glycemic status was associated with insulin resistance and inflammatory markers.

Vitamin B12 deficiency and hyperhomocysteinemia as correlates of cardiovascular risk factors in Indian subjects with coronary artery disease.

BACKGROUND AND PURPOSE: Folate and vitamin B12 are essential components in the metabolism of homocysteine (Hcy). Hyperhomocysteinemia has been implicated in endothelial dysfunction and cardiovascular disease. However, the association of Hcy, vitamin B12, and folic acid with cardiovascular risk factors in patients with coronary artery disease (CAD) has not been studied in Indian patients. This study was conducted with the aim to evaluate the relationship of vitamin B12, folic acid, and Hcy levels with cardiovascular risk factors in subjects with known CAD. METHODS AND SUBJECTS: Three hundred patients (216 men; 84 women; aged 25-92 years) who had CAD on angiography were included in this study consecutively. All patients were evaluated for anthropometry and cardiovascular risk factors, and blood samples were collected for biochemical, nutritional, and inflammatory markers. RESULTS: Percentage of vitamin B12 and folate deficiency was 86.7% and 2.7%, respectively. Hyperhomocysteinemia was present in 95.3% patients. Vitamin B12 levels were significantly lower and Hcy levels were significantly higher in subjects with dyslipidemia, DM, and/or hypertension. Serum vitamin B12 was inversely associated with triglyceride and very low-density lipoprotein (VLDL) and positively with high-density lipoprotein (HDL). Hcy was positively associated with triglyceride and VLDL and negatively with HDL. Vitamin B12 was inversely correlated with inflammatory markers (high-sensitivity C-reactive protein and interleukin-6) directly related to insulin resistance whereas Hcy showed the opposite pattern. CONCLUSIONS: Serum vitamin B12 deficiency and hyperhomocysteinemia are related with cardiovascular risk factors in Indian patients with CAD.

Is hypomagnesaemia a coronary risk factor among Indians with coronary artery disease?

INTRODUCTION: Magnesium is an essential element that has numerous biological functions in the cardiovascular system. Hence, three hundred patients with known cardiovascular disease above the age of 25 years were studied to evaluate association between dietary and serum magnesium with cardiovascular risk factors. MATERIALS AND METHODS: Patients were divided into three groups according to serum magnesium levels; ≤1.6 (Group 1), >1.6-2.6 (Group 2) and: >2.6 mg/dl (Group 3), and into two groups according to dietary magnesium intake; ≤350 mg/day (Group 1) and >350 mg/day (Group 2), respectively. RESULTS: Mean age of patients was 60.97 ± 12.5 years. Total cholesterol, triglycerides, VLDL, and LDL were significantly higher and HDL cholesterol significantly lower in group 1 when compared with group 2 and group 3. Diabetes, dyslipidemia, and hypertension were negatively correlated with serum magnesium levels; which were maintained even after adjustment with age, sex, and anthropometric parameters in multiple regression analysis. Similar observations were observed in dietary magnesium intake except LDL and total cholesterol. Dietary magnesium was positively correlated with serum magnesium. CONCLUSIONS: Hypomagnesaemia and low dietary intake of magnesium are strongly related to cardiovascular risk factors among known subjects with coronary artery disease. Hence, magnesium supplementation may help in reducing cardiovascular disease.

Association of constitutional type of Ayurveda with cardiovascular risk factors, inflammatory markers and insulin resistance.

CONTEXT: Ayurveda propounds that diseases manifest from imbalance of doshas. There, have been attempts to indicate biochemical basis of constitutional types described in Ayurveda. AIMS: The study was intended to assess the association of constitutional types (Prakriti) with cardiovascular risk factors, inflammatory markers and insulin resistance in subjects with coronary artery disease (CAD). SETTINGS AND DESIGN: Hospital based cross sectional study. MATERIALS AND METHODS: Three hundred patients with CAD >25 years were studied. Assessment of Prakriti was done by using Ayusoft software. Biochemical parameters, inflammatory markers (hsCRP, TNF-alpha and IL-6) and insulin resistance (HOMA-IR) were measured. STATISTICAL ANALYSIS: Was done using EPI INFO, version 3.5.3. RESULTS: Mean age of patients was 60.97±12.5 years. Triglyceride, VLDL and LDL was significantly higher (P<0.0001, P<0.0001 and 0.0355, respectively) and HDL cholesterol (P<0.0001) significantly lower in vatta kapha (VK) Prakriti when compared with other constitution type. VK Prakriti was correlated with diabetes mellitus (r=0.169, P=0.003), hypertension (r=0.211, P≤0.0001) and dyslipidemia (r=0.541, P≤0.0001). Inflammatory markers; IL6, TNF alpha, hsCRP and HOMA IR was highest in VK Prakriti. Inflammatory markers were correlated positively with both VK and Kapha group. CONCLUSIONS: There is strong relation of risk factors (diabetes, hypertension, dyslipidemia), insulin resistance, and inflammatory markers with Vata Kapha and Kapha Prakriti.