Imrana Naseem

Protective effect of quercetin on hyperglycemia, oxidative stress and DNA damage in alloxan induced type 2 diabetic mice

AIMS Quercetin is a natural polyphenolic flavonoid and acts as a quencher for reactive oxygen species generated by any physical or chemical action. In type 2 diabetes mellitus (T2DM) the basic characteristic feature is hyperglycemia which leads to complications involving oxidative stress. In view of this, the present study was conducted to examine the effect of quercetin in T2DM. MAIN METHODS A total of 18 mice were divided into three groups, vis control, diabetic and diabetic treated with quercetin. Fasting blood glucose (FBG) levels and anti-oxidant enzyme activity were assayed. Creatinine, urea, lipid peroxidation, GLUT4 expression and DNA damage were also measured. KEY FINDINGS A significant decrease in FBG level and liver and kidney marker enzymes was observed in the quercetin treated group as compared to the diabetic one. Glutathione, SOD, catalase, and glutathione-S-transferase levels were also found to be increased on quercetin supplementation. Thiobarbituric acid-reactive substance level was decreased while GLUT4 expression levels were increased in the treated group. DNA damage was also affected positively by quercetin when subjected with single cell alkaline gel electrophoresis. Thus, we may suggest an anti-oxidant potential and protective effect of quercetin in T2DM mice. SIGNIFICANCE From this study, we conclude that quercetin ameliorates hyperglycemia and oxidative stress, by blunting free radical induced toxicity in T2DM.

Ameliorative effect of riboflavin on the cisplatin induced nephrotoxicity and hepatotoxicity under photoillumination.

Cisplatin is a widely used anticancer drug. It is documented that it elicits major side effects like nephrotoxicity and hepatotoxicity due to oxidative stress forcing the patients to limit its clinical use in long term treatment. Riboflavin (vitamin B(2)) is a strong photosensitizer because it generates reactive oxygen species (ROS) upon photoillumination. We have tried to trap its photosensitizing property to ameliorate the cisplatin induced nephrotoxicity and hepatotoxicity in mice. They were treated with riboflavin and cisplatin separately as well as with their combination under photoilluminated condition. The status of major antioxidant enzymes, antioxidant proteins, functional markers, lipid peroxidation and protein oxidation was studied in liver, kidneys and serum samples of all the groups. Cisplatin treated group showed significantly compromised level of antioxidant enzymes and the proteins with higher extent of lipid and protein oxidation. Similar but less pronounced pattern was observed in the riboflavin treated group. The groups treated with the combination of cisplatin and riboflavin showed all the parameters tended towards normal levels in a dose dependent manner. Hence, it can be hypothesized that riboflavin shows ameliorative effect on the cisplatin induced nephrotoxicity and hepatotoxicity under the mentioned treatment conditions.

Synthesis and characterization of transition metal 2,6-pyridinedicarboxylic acid derivatives, interactions of Cu(II) and Ni(II) complexes with DNA in vitro

Mononuclear complexes M(L)Cl(2) where M=Mn(II), Fe(II), Co(II), Ni(II) and Cu(II) and (L=N,N-diethylpiperazinyl,2,6-pyridinedicarboxylate), have been synthesized and characterized by elemental analysis, FT-IR, (1)H NMR spectroscopy, UV-vis, magnetic moment, TGA/DSC, cyclic voltammetry and conductivity measurement data. The spectral data suggests that the dipicolinic acid acts as a bidentate ligand and is coordinated to the metal ion through the carboxylate oxygen. The cyclic voltammogram for Cu(L)Cl(2) complex was found to display two reversible Cu(II)/Cu(I) and Cu(II)/Cu(III) redox couple. The ligand exhibits a two-step thermolytic pattern while the complexes decompose in three stages respectively. An octahedral geometry has been proposed for both the complexes. The investigation of the interaction of the complexes with calf thymus DNA has been performed with absorption spectroscopy and fluorescence quenching experiments, which showed that the complexes are avid binders of calf thymus DNA. Also the interaction of the Cu(II) and Ni(II) complexes with plasmid DNA (pUC 19) was studied using agarose gel electrophoresis. The results revealed that these complexes can act as effective DNA cleaving agents resulting in the nicked form of DNA (pUC 19) under physiological conditions. The gel was run both in the absence and presence of an oxidizing agent (H(2)O(2)). The ligand and its complexes have also been screened against microbes in order to study their antibacterial action. The results revealed that the Cu(II) complex has activity comparable with the reference drugs gentamycin and flucanzole.

Effect of Alkylated and Intercalated DNA on the Generation of Superoxide Anion by Riboflavin

Superoxide anion (O2.−) was photogenerated upon illumination of riboflavin in fluorescent light. The rate of O2.− formation was stimulated by double stranded DNA but not by denatured DNA or RNA. Depurinated DNA, which was predominantly depleted in guanine residues, did not exhibit the stimulatory effect, indicating an interaction of riboflavin, or active oxygen species derived from it, with guanine bases. Also, the stimulation of O2.− photogeneration was not observed with ethidium bromide but was seen with proflavin-intercalated DNA. Since ethidium bromide intercalates preferentially between purines and pyrimidines, and proflavin prefers dA-dT rich sites, these results were interpreted to suggest that the interaction of riboflavin with DNA is mainly with GC or CG base pairs.

Riboflavin Ameliorates Cisplatin Induced Toxicities under Photoillumination

Background Cisplatin is an effective anticancer drug that elicits many side effects mainly due to induction of oxidative and nitrosative stresses during prolonged chemotherapy. The severity of these side effects consequently restricts its clinical use under long term treatment. Riboflavin is an essential vitamin used in various metabolic redox reactions in the form of flavin adenine dinucleotide and flavin mononucleotide. Besides, it has excellent photosensitizing property that can be used to ameliorate these toxicities in mice under photodynamic therapy. Methods and Findings Riboflavin, cisplatin and their combinations were given to the separate groups of mice under photoilluminated condition under specific treatment regime. Their kidney and liver were excised for comet assay and histopathological studies. Furthermore, Fourier Transform Infrared Spectroscopy of riboflavin-cisplatin combination in vitro was also conducted to investigate any possible interaction between the two compounds. Their comet assay and histopathological examination revealed that riboflavin in combination with cisplatin was able to protect the tissues from cisplatin induced toxicities and damages. Moreover, Fourier Transform Infrared Spectroscopy analysis of the combination indicated a strong molecular interaction among their constituent groups that may be assigned for the protective effect of the combination in the treated animals. Conclusion Inclusion of riboflavin diminishes cisplatin induced toxicities which may possibly make the cisplatin-riboflavin combination, an effective treatment strategy under chemoradiotherapy in pronouncing its antineoplastic activity and sensitivity towards the cancer cells as compared to cisplatin alone.

Multi-spectroscopic and molecular modelling approach to investigate the interaction of riboflavin with human serum albumin

Riboflavin (RF) plays an important role in various metabolic redox reactions in the form of flavin adenine dinucleotide and flavin mononucleotide. Human serum albumin (HSA) is an important protein involved in the transportation of drugs, hormones, fatty acid and other molecules which determine the biodistribution and physiological fate of these molecules. In this study, we have investigated the interaction of riboflavin RF with HSA under simulative physiological conditions using various biophysical, calorimetric and molecular docking techniques. Results demonstrate the formation of riboflavin–HSA complex with binding constant in the order of 104 M−1. Fluorescence spectroscopy confirms intermediate strength having a static mode of quenching with stoichiometry of 1:1. Experimental results suggest that the binding site of riboflavin mainly resides in sub-domain IIA of HSA and that ligand interaction increases the α-helical content of HSA. These parameters were further verified by isothermal titration calorimetry ITC which confirms the thermodynamic parameters obtained by fluorescence spectroscopy. Molecular docking was employed to suggest a binding model. Based on thermodynamic, spectroscopic and computational observations it can be concluded that HSA-riboflavin complex is mainly stabilized by various non-covalent forces with binding energy of −7.2 kcal mol−1.

Understanding the mechanism of non-enzymatic glycation inhibition by cinnamic acid: an in vitro interaction and molecular modelling study

Under hyperglycaemic conditions non-enzymatic glycation of proteins gives rise to advanced glycation end products (AGEs). The AGEs thus formed generate free radicals, which foster the development of diabetes and its associated complications. Inhibition of glycation is expected to play a role in controlling diabetes. Plant derived antioxidants like cinnamic acid (CA) are known for limiting AGE formation, however, the mechanism involved is poorly understood. Therefore, we aimed to investigate the possible mechanism of inhibition of AGEs formation by CA through various experimental approaches. Glycation of HSA was achieved by incubating the reaction mixture with glucose for 30 days at 37 °C. The protein samples were tested for levels of free lysine & thiol groups, carbonyl content and reactive oxygen species (ROS). Interaction between CA and HSA was also studied through various biophysical techniques. Thermodynamic studies showed a strong exothermic interaction between CA and HSA. The positive value of TΔS° and negative value of ΔH° indicates that the HSA–CA complex is mainly stabilized by a hydrophobic interaction and hydrogen bond. Further, molecular docking reveals that CA binds to HSA subdomain IIA (Sudlows site I) with a binding energy of −7.0 kcal mol−1, nearly the same as obtained in isothermal titration calorimetry (ITC) and fluorescence spectroscopy. The results of various spectroscopic techniques along with molecular docking and examination of many biomarkers highlights the role of CA in preventing disease progression.

Effect of 1, 25(OH)2 vitamin D3 on glucose homeostasis and DNA damage in type 2 diabetic mice

AIMS The purpose of the study was to examine the effect of 1, 25(OH)₂ Vitamin D₃ supplementation on type 2 diabetic (T2DM) mice. MATERIALS AND METHODS A total of 24 mice were taken and divided into three groups of control; diabetic and diabetic + vitamin D supplemented ones. Serum calcium level, fasting blood glucose level (FBG), hexokinase activity, glucose-6-phosphatse and fructose 1,6 bisphosphatase activity were measured to establish a relevant correlation between vitamin D supplementation and hyperglycemia in T2DM. RESULTS There occurred an increase in FBG levels (250 ± 0.41 mg/dl) and a significant decrease in serum calcium levels in the diabetic group (8.63 ± 0.40 mg/ml) both of which reached near control levels on vitamin D₃ supplementation. The activity of the glucose metabolic enzymes was also assayed in diabetic group and was found to be deviated from control group; hexokinase (0.0241 ± 0.014 μg/mg/ml) FBPase (0.433 ± 0.002 μg/mg/ml) and G6Pase (0.918 ± 0.02 μg/mg/ml). However, the activity of these enzymes returned to near control values with hexokinase activity reaching 0.717 ± 0.003 μg/mg/ml on vitamin D₃ supplementation. The FBPase and G6Pase activities were decreased to 0.2733 ± 0.008 μg/mg/ml and G6Pase 0.71 ± 0.01 μg/mg/ml respectively. In addition to enzymatic analysis, the organs of all three groups of mice were subjected to comet assay. The diabetic group receiving vitamin D supplementation showed a marked recovery exhibiting shorter tail length both in liver (21.80 ± 2.40 μm) and pancreatic cells (19.25 ± 1.90 μm) as compared to the diabetic group exhibiting a tail length of 30.41 ± 2.50 μm and 32.45 ± 2.87 μm in liver and pancreatic cells respectively. CONCLUSION The present study shows that vitamin D₃ supplementation is positively correlated with decrease in blood glucose level and serum calcium level in fasting condition. This suggests a positive influence of vitamin D on glucose homeostasis. Besides, the activity of various glucose metabolic enzymes (hexokinase, FBPase and G6Pase) as shown by our results and the remarkable shortening of DNA tail length in vitamin D supplemented diabetic group as compared to diabetic group without supplementation further support the idea that vitamin D supplementation might be an add-on therapy for patients with T2DM.

White light-mediated Cu (II)–5FU interaction augments the chemotherapeutic potential of 5-FU: an in vitro study

Abstract5-Fluorouracil (5-FU) is a potent photosensitizer used in colon and rectal cancers. 5-FU on galvanostatic electrolysis or radiation-induced oxidation of aqueous solution yields N1–C5-linked dimmer hydrate of 5-FU. Copper is presently associated with chromatin; in cancer cells the concentration of copper is very high. It has been shown to be capable of mediating the action of several anticancer drugs through the production of reactive oxygen species (ROS). The objective of the present study is to determine the Cu (II)-mediated anticancer mechanism of 5-FU under photo-illumination as well as 5-FU alone. We have shown that a pro-oxidant action was enhanced when Cu (II) was used with 5-FU as compared to 5-FU alone. This may be due to the inhibition of dimerization of 5-FU when present in combination with Cu (II) under photo-illumination. It was also shown that 5-FU alone as well as in combination with Cu (II) was able to generate oxidative stress in lymphocyte which is inhibited by scavengers of ROS. Moreover, the results of Fourier-transformed infrared spectra lead to the conclusion that the dimerization of 5-FU was inhibited when used in combination with Cu (II). It was due to the interaction of 5-FU with Cu (II). Hence, we propose that during chemoradiotherapy with 5-FU, the endogenous copper is mobilized by 5-FU, leading to the generation of ROS which cause oxidative stress and possibly cancer cell death by apoptosis.

Riboflavin-mediated cellular photoinhibition of cisplatin-induced oxidative DNA breakage in mice epidermal keratinocytes.

Background/purpose: Cisplatin is one of the most effective chemotherapeutic agents used in the treatment of various kinds of malignant tumors. A major drawback associated with cisplatin chemotherapy is its cytotoxicity/genotoxicity towards normal tissues. The cytotoxicity has been attributed, in part, to the oxidative stress generated by the drug. Riboflavin is a micronutrient known for its photosensitization characteristics. Owing to the emergence of photodynamic therapy as a modality for the treatment of solid tumors and other accessible lesions in terms of opthalmic, dermatological, cardiovascular and urological diseases, the therapeutic window of riboflavin as a sensitizer has been used to avoid the toxic side effects of cisplatin.