Synaptophysin Chaperones the Assembly of 12 SNAREpins under each Ready-Release Vesicle.

The synaptic vesicle protein Synaptophysin has long been known to form a complex with the v-SNARE VAMP, but a more specific molecular function or mechanism of action in exocytosis has been lacking because gene knockouts have minimal effects. Utilizing fully-defined reconstitution and single-molecule measurements, we now report that Synaptophysin functions as a chaperone that determines the number of SNAREpins assembling between a ready-release vesicle and its target membrane bilayer. Specifically, Synaptophysin directs the assembly of 12 ± 1 SNAREpins under each docked vesicle, even in the face of an excess of SNARE proteins. The SNAREpins assemble in successive waves of 6 ± 1 and 5 ± 2 SNAREpins, respectively, tightly linked to oligomerization of and binding to the vesicle Ca++ sensor Synaptotagmin. Templating of 12 SNAREpins by Synaptophysin is likely the direct result of its hexamer structure and its binding of VAMP2 dimers, both of which we demonstrate in detergent extracts and lipid bilayers.

Rapid Quantification of First and Second Phase Insulin Secretion Dynamics using an In vitro Platform for Improving Insulin Therapy.

High-throughput quantification of the first- and second-phase insulin secretion dynamics is intractable with current methods. The fact that independent secretion phases play distinct roles in metabolism necessitates partitioning them separately and performing high-throughput compound screening to target them individually. We developed an insulin-nanoluc luciferase reporter system to dissect the molecular and cellular pathways involved in the separate phases of insulin secretion. We validated this method through genetic studies, including knockdown and overexpression, as well as small-molecule screening and their effects on insulin secretion. Furthermore, we demonstrated that the results of this method are well correlated with those of single-vesicle exocytosis experiments conducted on live cells, providing a quantitative reference for the approach. Thus, we have developed a robust methodology for screening small molecules and cellular pathways that target specific phases of insulin secretion, resulting in a better understanding of insulin secretion, which in turn will result in a more effective insulin therapy through the stimulation of endogenous glucose-stimulated insulin secretion.

Native Planar Asymmetric Suspended Membrane for Single-Molecule Investigations: Plasma Membrane on a Chip.

Cellular plasma membranes, in their role as gatekeepers to the external environment, host numerous protein assemblies and lipid domains that manage the movement of molecules into and out of cells, regulate electric potential, and direct cell signaling. The ability to investigate these roles on the bilayer at a single-molecule level in a controlled, in vitro environment while preserving lipid and protein architectures will provide deeper insights into how the plasma membrane works. A tunable silicon microarray platform that supports stable, planar, and asymmetric suspended lipid membranes (SLIM) using synthetic and native plasma membrane vesicles for single-molecule fluorescence investigations is developed. Essentially, a "plasma membrane-on-a-chip" system that preserves lipid asymmetry and protein orientation is created. By harnessing the combined potential of this platform with total internal reflection fluorescence (TIRF) microscopy, the authors are able to visualize protein complexes with single-molecule precision. This technology has widespread applications in biological processes that happen at the cellular membranes and will further the knowledge of lipid and protein assemblies.

Chebulagic acid attenuates HFD/streptozotocin induced impaired glucose metabolism and insulin resistance via up regulations of PPAR γ and GLUT 4 in type 2 diabetic rats.

The aim of the present study was to evaluate the effects of chebulagic acid on enzymes involved in carbohydrate metabolism in high-fat diet/streptozotocin-induced diabetic rats. Diabetes was induced in albino male Wistar rats by feeding them with a high-fat diet comprising of 84.3% standard laboratory rat chow, 5% lard, 10% yolk powder, 0.2% cholesterol, and 5% bile salt for 2 weeks. After 2 weeks, the animals were kept in an overnight fast and injected with a low dose of streptozotocin (35 mg/kg, dissolved in 0.1 M sodium citrate buffer, pH 4.5). At the end of the experimental periods, the blood glucose, plasma insulin, hemoglobin, glycated hemoglobin levels, homeostatic model of insulin resistance, glycogen content, glycogen synthase, glycogen phosphorylase were measured in high-fat diet and streptozotocin-induced diabetic rats whereas the glucose metabolic enzymes such as hexokinase, glucose 6 phosphate dehydrogenase, glucose 6 phosphatase, fructose 1,6 bisphosphatase and protein expression of peroxisome proliferator-activated receptor-γ (PPAR-γ) and Glucose transporter 4 (GLUT4) were analyzed. Administration of chebulagic acid to HFD/STZ -induced diabetic rats for 30 days significantly decreased the levels of plasma glucose, homeostatic model assessment of insulin resistance, glycogen phosphorylase and glycosylated hemoglobin and increased the levels of insulin, glycogen content and glycogen synthase. On the other hand, the altered activity of carbohydrate metabolic enzymes, PPAR-γ and GLUT4 were brought back to near-normal levels on treatment with chebulagic acid. The effect produced by chebulagic acid on various parameters was comparable to that of metformin.

Antioxidant and antihyperlipidemic activities of catechol derivatives and biflavonoid isolated from Semecarpus anacardium seeds.

Semecarpus anacardium Linn. (Family: Anacardiaceae), commonly known marking nuts has been used in various traditional system of medicines for various ailments (such as antiatherogenic, antiinflammatory, antimicrobial, hypoglycemic, anticarcinogenic etc) since ancient times.Based on the wide pharmacological activities of this plant, the present study was aimed to explore the antioxidant and antihyperlipidemic potential in high fat diet fed rats using catechol derivatives I-IV and biflavonoid isolated from seeds of Semecarpus anacardium. Oral administration of catechol derivatives I-IV and biflavonoid at a concentration of 50 mg/kg b.wt to high fat diet fed rats for a period of 30 days significantly decreased the lipid profiles, body weight gain and organ weight when compared to untreated hypercholesterolemic rats. However, biflavonoid treated hypercholesterolemic rats showed more pronounced effects in all the parameters tested when compared to all catechol derivatives (I-IV) treated hypercholesterolemic rats. The effect produced by biflavonoid on various parameters was comparable to that of simvastastin- a standard drug. In vitro antioxidant activities were also conducted using these five compounds in which biflavonoid showed more significant antioxidant potential at a concentration of 1000 µg/ml when compared to catechol derivatives (I-IV). The pronounced antioxidant potential of biflavonoid might have contributed to the hypolipidemic action in hypercholesterolemic rats and improved oil red O staining of thoracic aorta has also supported the parameters investigated. Further, the molecular mechanism of cholesterol lowering potential of this drug is needed.

Vesicle capture by membrane-bound Munc13-1 requires self-assembly into discrete clusters.

Munc13-1 is a large banana-shaped soluble protein that is involved in the regulation of synaptic vesicle docking and fusion. Recent studies suggest that multiple copies of Munc13-1 form nano-assemblies in active zones of neurons. However, it is not known whether such clustering of Munc13-1 is correlated with multivalent binding to synaptic vesicles or specific plasma membrane domains at docking sites in the active zone. The functional significance of putative Munc13-1 clustering is also unknown. Here, we report that nano-clustering is an inherent property of Munc13-1 and is indeed required for vesicle binding to bilayers containing Munc13-1. Purified Munc13-1 protein reconstituted onto supported lipid bilayers assembled into clusters containing from 2 to ˜ 20 copies as revealed by a combination of quantitative TIRF microscopy and step-wise photobleaching. Surprisingly, only clusters containing a minimum of 6 copies of Munc13-1 were capable of efficiently capturing and retaining small unilamellar vesicles. The C-terminal C2 C domain of Munc13-1 is not required for Munc13-1 clustering, but is required for efficient vesicle capture. This capture is largely due to a combination of electrostatic and hydrophobic interactions between the C2 C domain and the vesicle membrane.

Munc13 binds and recruits SNAP25 to chaperone SNARE complex assembly.

Synaptic vesicle fusion is mediated by SNARE proteins-VAMP2 on the vesicle and Syntaxin-1/SNAP25 on the presynaptic membrane. Chaperones Munc18-1 and Munc13-1 cooperatively catalyze SNARE assembly via an intermediate 'template' complex containing Syntaxin-1 and VAMP2. How SNAP25 enters this reaction remains a mystery. Here, we report that Munc13-1 recruits SNAP25 to initiate the ternary SNARE complex assembly by direct binding, as judged by bulk FRET spectroscopy and single-molecule optical tweezer studies. Detailed structure-function analyses show that the binding is mediated by the Munc13-1 MUN domain and is specific for the SNAP25 'linker' region that connects the two SNARE motifs. Consequently, freely diffusing SNAP25 molecules on phospholipid bilayers are concentrated and bound in ~ 1 : 1 stoichiometry by the self-assembled Munc13-1 nanoclusters.

Antigenotoxic and Antimutagenic Effects of Andrographis paniculata, a Traditional Medicinal Herb against Genotoxicity of Cyclophosphamide: An In Vitro Study on Human Peripheral Lymphocytes.

Andrographis paniculata (family: Acanthaceae) is a medicinal herb-used in Indian system of medicine (Ayurveda, Siddha, and Unani), traditional and folk systems to treat various illnesses. This study examined the phytochemical constituents of ethanol extract from A. paniculata and its protective effect against genotoxicity caused by cyclophosphamide (CPA). Phytochemical screening and estimation of total phenolic content were analyzed using standard methods. The bioactive components from the ethanol extract of A. paniculata (EAP) were analyzed using gas chromatography-mass spectrometry. To investigate the protective effect of EAP against CPA-induced genotoxicity, human peripheral lymphocyte cultures were used. To test the antigenotoxic and antimutagenic effects of EAP, lymphocytes were treated with different concentrations of extract (50∼250 mg/mL) alone and co-treated along with CPA+EAP for 48 h. The cells were analyzed for structural chromosomal aberrations (CAs) and sister chromatid exchanges (SCEs) in control, CPA treated, and CPA+ EAP co-treated lymphocytes. Results of the study revealed that the lymphocyte cultures which had 48 h continuous exposure to EAP (50∼250 mg/mL) did not show any significant changes in CAs and SCE frequencies. These results substanti-ate the antimutagenic nature of the extract. Furthermore, the lymphocytes co-treated with CPA along with extract showed a significant reduction in CAs (reduced from 26.50±2.50% to 11.00±1.00%) and SCEs (reduced from 9.92±0.63 per cell to 4.56±0.18 per cell). These results suggest that A. paniculata is protective against CPA induced genotoxicity and put forward its possible use as a supplement with chemotherapeutic drugs.

Acyclic Isoprenoid Attenuates Lipid Anomalies and Inflammatory Changes in Hypercholesterolemic Rats.

The present study was aimed to explore the antihyperlipidemic and anti-inflammatory effect of acyclic isoprenoid on high fat diet fed rats. Hypercholesterolemia was induced by the diet comprising of the normal rat chow 84.3%, lard 5%, yolk powder 10%, cholesterol 0.2% and 0.5% bile salt were fed to the rats for the period of 8 weeks. The results showed that abnormally elevated levels of plasma lipid profiles. Three different doses of acyclic isoprenoid (20, 40 and 80 mg/kg b.w/day) were administered orally to hypercholesterolemia suffering rats for the period of 30 days. Among these three doses of acyclic isoprenoid, the dose 80 mg/kg b.w. was significantly decreased the plasma lipid profiles when compared to other two doses. The effect produced by acyclic isoprenoid (80 mg/kg b.w) was comparable to that of simvastatin. Therefore, 80 mg/kg b.w was fixed as a effective dose and used for further analyses. Acyclic isoprenoid administration reinstated the elevated levels of cardiac and inflammatory markers in both blood and serum of hypercholesterolemic rats. In addition, acyclic isoprenoid administration decreased activity of 3-hydroxy 3-methyl-glutaryl-CoA reductase and increased the activity of lecithin cholesterol acyl transferase. These findings suggest that the administration of acyclic isoprenoid was potentially ameliorated the cardiac marker enzymes and inflammatory markers in addition to its antihypercholesterolemic effect.

Chromosome Territorial Organization Drives Efficient Protein Complex Formation: A Hypothesis.

In eukaryotes, chromosomes often form a transcriptional kissing loop during interphase. We propose that these kissing loops facilitate the formation of protein complexes. mRNA transcripts from these loops could cluster together into phase-separated nuclear granules. Their export into the ER could be ensured by guided diffusion through the inter-chromatin space followed by association with nuclear baskets and export factors. Inside the ER, these mRNAs would form a translation hub. Juxtaposed translation of these mRNAs would increase the cis/trans protein complex assembly among the nascent protein chains. Eukaryotes might employ this pathway to increase complex formation efficiency.

Mechanical characterization of HIV-1 with a solid-state nanopore sensor.

Enveloped viruses fuse with cells to transfer their genetic materials and infect the host cell. Fusion requires deformation of both viral and cellular membranes. Since the rigidity of viral membrane is a key factor in their infectivity, studying the rigidity of viral particles is of great significance in understating viral infection. In this paper, a nanopore is used as a single molecule sensor to characterize the deformation of pseudo-type human immunodeficiency virus type 1 at sub-micron scale. Non-infective immature viruses were found to be more rigid than infective mature viruses. In addition, the effects of cholesterol and membrane proteins on the mechanical properties of mature viruses were investigated by chemically modifying the membranes. Furthermore, the deformability of single virus particles was analyzed through a recapturing technique, where the same virus was analyzed twice. The findings demonstrate the ability of nanopore resistive pulse sensing to characterize the deformation of a single virus as opposed to average ensemble measurements.

Acute and sub acute studies of catechol derivatives from Semecarpus anacardium.

The present study was aimed at evaluating the acute and subacute toxicity of catechol derivatives (I-IV, isolated from Semecarpus anacardium nuts) in Wistar Albino rats. In acute study (14 days), catechol derivatives I-IV 800 mg/kg caused no behavioral adverse effects and mortality. Fifty percent (LD50) of mortality was observed in catechol derivatives I-III (1600 mg/kg b.wt) and catechol derivative IV (1250 mg/kg b.wt). In subacute study, daily oral administration of catechol derivatives I-IV (300 mg/kg b.wt) for 30 days did not result in death or significant changes in the body weight and organ weight, In hematological and some biochemical analysis showed few beneficial effects particularly in catechol derivatives I and IV treated rats that is transient rise in WBC count and HDL cholesterol and decrease in LDL, plasma and tissue lipid profile. These results indicate the impact of catechol derivatives in boosting the immune system and reducing cardiovascular risk factors and thereby they possess cardio protective and immunopotentiating effect. Further, histopathological examination of liver and kidney showed normal architecture that suggests no morphological disturbances. Based on the results obtained, it may be concluded that the catechol derivatives are potentially toxic but therapeutically effective.

Targeting cell surface HIV-1 Env protein to suppress infectious virus formation.

HIV-1 Env protein is essential for host cell entry, and targeting Env remains an important antiretroviral strategy. We previously found that a peptide triazole thiol KR13 and its gold nanoparticle conjugate AuNP-KR13 directly and irreversibly inactivate the virus by targeting the Env protein, leading to virus gp120 shedding, membrane disruption and p24 capsid protein release. Here, we examined the consequences of targeting cell-surface Env with the virus inactivators. We found that both agents led to formation of non-infectious virus from transiently transfected HEK293T cells. The budded non-infectious viruses lacked Env gp120 but contained gp41. Importantly, budded virions also retained the capsid protein p24, in stark contrast to p24 leakage from viruses directly treated by these agents and arguing that the agents led to deformed viruses by transforming the cells at a stage before virus budding. We found that the Env inactivators caused gp120 shedding from the transiently transfected HEK293T cells as well as non-producer CHO-K1-gp160 cells. Additionally, AuNP-KR13 was cytotoxic against the virus-producing HEK293T and CHO-K1-gp160 cells, but not untransfected HEK293T or unmodified CHO-K1 cells. The results obtained reinforce the argument that cell-surface HIV-1 Env is metastable, as on virus particles, and provides a conformationally vulnerable target for virus suppression and infectious cell inactivation.

Isolation and Characterization of an Acyclic Isoprenoid from Semecarpus anacardium Linn. and its Antibacterial Potential in vitro: - Antimicrobial Activity of Semecarpus anacardium Linn. Seeds.

OBJECTIVES: Semecarpus anacardium Linn. is a plant well-known for its antimicrobial, antidiabetic and anti-arthritic properties in the Ayurvedic and Siddha system of medicine. This has prompted the screening of this plant for antibacterial activity. The main aims of this study were to isolate compounds from the plant's seeds and to evaluate their antibacterial effects on clinical bacterial test strains. METHODS: The n-butanolic concentrate of the seed extract was subjected to thin layer chromatography (TLC) and repeated silica gel column chromatography followed by elution with various solvents. The compound was identified based on observed spectral (IR, 1H NMR, 13C NMR and high-resolution mass spectrometry) data. The well diffusion method was employed to evaluate the antibacterial activities of the isolated acyclic isoprenoid compound (final concentration: 5 - 15 µg/mL) on four test bacterial strains, namely, Staphylococcus aureus (MTCC 96), Bacillus cereus (MTCC 430), Escherichia coli (MTCC 1689) and Acinetobacter baumannii (MTCC 9829). RESULTS: Extensive spectroscopic studies showed the structure of the isolated compound to be an acyclic isoprenoid (C21H32O). Moreover, the isoprenoid showed a remarkable inhibition of bacterial growth at a concentration of 15 µg/mL compared to the two other doses tested (5 and 10 µg/mL) and to tetracycline, a commercially available antibiotic that was used as a reference drug. CONCLUSION: The isolation of an antimicrobial compound from Semecarpus anacardium seeds validates the use of this plant in the treatment of infections. The isolated compound found to be active in this study could be useful for the development of new antimicrobial drugs.

Nanoparticle mechanics: deformation detection via nanopore resistive pulse sensing.

Solid-state nanopores have been widely used in the past for single-particle analysis of nanoparticles, liposomes, exosomes and viruses. The shape of soft particles, particularly liposomes with a bilayer membrane, can greatly differ inside the nanopore compared to bulk solution as the electric field inside the nanopores can cause liposome electrodeformation. Such deformations can compromise size measurement and characterization of particles, but are often neglected in nanopore resistive pulse sensing. In this paper, we investigated the deformation of various liposomes inside nanopores. We observed a significant difference in resistive pulse characteristics between soft liposomes and rigid polystyrene nanoparticles especially at higher applied voltages. We used theoretical simulations to demonstrate that the difference can be explained by shape deformation of liposomes as they translocate through the nanopores. Comparing our results with the findings from electrodeformation experiments, we demonstrated that the rigidity of liposomes can be qualitatively compared using resistive pulse characteristics. This application of nanopores can provide new opportunities to study the mechanics at the nanoscale, to investigate properties of great value in fundamental biophysics and cellular mechanobiology, such as virus deformability and fusogenicity, and in applied sciences for designing novel drug/gene delivery systems.

Impact of HIV-1 Membrane Cholesterol on Cell-Independent Lytic Inactivation and Cellular Infectivity.

Peptide triazole thiols (PTTs) have been found previously to bind to HIV-1 Env spike gp120 and cause irreversible virus inactivation by shedding gp120 and lytically releasing luminal capsid protein p24. Since the virions remain visually intact, lysis appears to occur via limited membrane destabilization. To better understand the PTT-triggered membrane transformation involved, we investigated the role of envelope cholesterol on p24 release by measuring the effect of cholesterol depletion using methyl beta-cyclodextrin (MßCD). An unexpected bell-shaped response of PTT-induced lysis to [MßCD] was observed, involving lysis enhancement at low [MßCD] vs loss of function at high [MßCD]. The impact of cholesterol depletion on PTT-induced lysis was reversed by adding exogenous cholesterol and other sterols that support membrane rafts, while sterols that do not support rafts induced only limited reversal. Cholesterol depletion appears to cause a reduced energy barrier to lysis as judged by decreased temperature dependence with MßCD. Enhancement/replenishment responses to [MßCD] also were observed for HIV-1 infectivity, consistent with a similar energy barrier effect in the membrane transformation of virus cell fusion. Overall, the results argue that cholesterol in the HIV-1 envelope is important for balancing virus stability and membrane transformation, and that partial depletion, while increasing infectivity, also makes the virus more fragile. The results also reinforce the argument that the lytic inactivation and infectivity processes are mechanistically related and that membrane transformations occurring during lysis can provide an experimental window to investigate membrane and protein factors important for HIV-1 cell entry.

Disulfide Sensitivity in the Env Protein Underlies Lytic Inactivation of HIV-1 by Peptide Triazole Thiols.

We investigated the mode of action underlying lytic inactivation of HIV-1 virions by peptide triazole thiol (PTT), in particular the relationship between gp120 disulfides and the C-terminal cysteine-SH required for virolysis. Obligate PTT dimer obtained by PTT SH cross-linking and PTTs with serially truncated linkers between pharmacophore isoleucine-ferrocenyltriazole-proline-tryptophan and cysteine-SH were synthesized. PTT variants showed loss of lytic activity but not binding and infection inhibition upon SH blockade. A disproportionate loss of lysis activity vs binding and infection inhibition was observed upon linker truncation. Molecular docking of PTT onto gp120 argued that, with sufficient linker length, the peptide SH could approach and disrupt several alternative gp120 disulfides. Inhibition of lysis by gp120 mAb 2G12, which binds at the base of the V3 loop, as well as disulfide mutational effects, argued that PTT-induced disruption of the gp120 disulfide cluster at the base of the V3 loop is an important step in lytic inactivation of HIV-1. Further, PTT-induced lysis was enhanced after treating virus with reducing agents dithiothreitol and tris (2-carboxyethyl)phosphine. Overall, the results are consistent with the view that the binding of PTT positions the peptide SH group to interfere with conserved disulfides clustered proximal to the CD4 binding site in gp120, leading to disulfide exchange in gp120 and possibly gp41, rearrangement of the Env spike, and ultimately disruption of the viral membrane. The dependence of lysis activity on thiol-disulfide interaction may be related to intrinsic disulfide exchange susceptibility in gp120 that has been reported previously to play a role in HIV-1 cell infection.

Macrocyclic Envelope Glycoprotein Antagonists that Irreversibly Inactivate HIV-1 before Host Cell Encounter.

We derived macrocyclic HIV-1 antagonists as a new class of peptidomimetic drug leads. Cyclic peptide triazoles (cPTs) retained the gp120 inhibitory and virus-inactivating signature of parent PTs, arguing that cyclization locked an active conformation. The six-residue cPT 9 (AAR029b) exhibited submicromolar antiviral potencies in inhibiting cell infection and triggering gp120 shedding that causes irreversible virion inactivation. Importantly, cPTs were stable to trypsin and chymotrypsin compared to substantial susceptibility of corresponding linear PTs.

Peptide Triazole Inactivators of HIV-1 Utilize a Conserved Two-Cavity Binding Site at the Junction of the Inner and Outer Domains of Env gp120.

We used coordinated mutagenesis, synthetic design, and flexible docking to investigate the structural mechanism of Env gp120 encounter by peptide triazole (PT) inactivators of HIV-1. Prior results demonstrated that the PT class of inhibitors suppresses binding at both CD4 and coreceptor sites on Env and triggers gp120 shedding, leading to cell-independent irreversible virus inactivation. Despite these enticing anti-HIV-1 phenotypes, structural understanding of the PT-gp120 binding mechanism has been incomplete. Here we found that PT engages two inhibitor ring moieties at the junction between the inner and outer domains of the gp120 protein. The results demonstrate how combined occupancy of two gp120 cavities can coordinately suppress both receptor and coreceptor binding and conformationally entrap the protein in a destabilized state. The two-cavity model has common features with small molecule gp120 inhibitor binding sites and provides a guide for further design of peptidomimetic HIV-1 inactivators based on the PT pharmacophore.

Effect of tangeretin, a polymethoxylated flavone on glucose metabolism in streptozotocin-induced diabetic rats.

The present study was designed to evaluate the antihyperglycemic potential of tangeretin on the activities of key enzymes of carbohydrate and glycogen metabolism in control and streptozotocin induced diabetic rats. The daily oral administration of tangeretin (100mg/kg body weight) to diabetic rats for 30 days resulted in a significant reduction in the levels of plasma glucose, glycosylated hemoglobin (HbA1c) and increase in the levels of insulin and hemoglobin. The altered activities of the key enzymes of carbohydrate metabolism such as hexokinase, pyruvate kinase, lactate dehydrogenase, glucose-6-phosphatase, fructose-1,6-bisphosphatase, glucose-6-phosphate dehydrogenase, glycogen synthase and glycogen phosphorylase in liver of diabetic rats were significantly reverted to near normal levels by the administration of tangeretin. Further, tangeretin administration to diabetic rats improved hepatic glycogen content suggesting the antihyperglycemic potential of tangeretin in diabetic rats. The effect produced by tangeretin on various parameters was comparable to that of glibenclamide - a standard oral hypoglycemic drug. Thus, these results show that tangeretin modulates the activities of hepatic enzymes via enhanced secretion of insulin and decreases the blood glucose in streptozotocin induced diabetic rats by its antioxidant potential.