Myricitrin, a Glycosyloxyflavone in Myrica esculenta Bark Ameliorates Diabetic Nephropathy via Improving Glycemic Status, Reducing Oxidative Stress, and Suppressing Inflammation.

The present study evaluated the therapeutic potential of myricitrin (Myr), a glycosyloxyflavone extracted from Myrica esculenta bark, against diabetic nephropathy. Myr exhibited a significant hypoglycemic effect in high fat-fed and a single low-dose streptozotocin-induced type 2 diabetic (T2D) rats. Myr was found to improve glucose uptake by the skeletal muscle via activating IRS-1/PI3K/Akt/GLUT4 signaling in vitro and in vivo. Myr significantly attenuated high glucose (HG)-induced toxicity in NRK cells and in the kidneys of T2D rats. In this study, hyperglycemia caused nephrotoxicity via endorsing oxidative stress and inflammation resulting in the induction of apoptosis, fibrosis, and inflammatory damages. Myr was found to attenuate oxidative stress via scavenging/neutralizing oxidative radicals and improving endogenous redox defense through Nrf-2 activation in both in vitro and in vivo systems. Myr was also found to attenuate diabetes-triggered renal inflammation via suppressing NF-κB activation. Myr inhibited hyperglycemia-induced apoptosis and fibrosis in renal cells evidenced by the changes in the expressions of the apoptotic and fibrotic factors. The molecular docking predicted the interactions between Myr and different signal proteins. An in silico absorption, distribution, metabolism, excretion, and toxicity (ADMET) study predicted the drug-likeness character of Myr. Results suggested the possibility of Myr to be a potential therapeutic agent for diabetic nephropathy in the future.

Rosmarinic Acid Attenuates Cadmium-Induced Nephrotoxicity via Inhibition of Oxidative Stress, Apoptosis, Inflammation and Fibrosis.

The present investigation was executed to reveal the protective mechanism of rosmarinic acid (RA) against cadmium (Cd)-induced nephrotoxicity. RA exhibited a concentration-dependent anti-apoptotic effect against CdCl2 in isolated mouse proximal tubular epithelial cells. Cd treatment significantly (p < 0.01) imparted oxidative stress to the renal cells via excessive ROS production, triggering NO level, NADPH oxidase activation, and impairment of cellular redox defense system. Cd-mediated oxidative stress significantly (p < 0.01) endorsed apoptosis to the murine kidney cells by triggering NF-κB/PKC-δ/TNFR2 activation. In addition, CdCl2 induced renal fibrosis by triggering TGF-ß1/SMAD3/α-SMA/collagen signaling within renal cells. On the other hand, RA significantly (p < 0.05-0.01) attenuated Cd-provoked oxidative stress and associated pathological signal transduction in murine renal cells. RA treatment also could significantly (p < 0.05-0.01) reciprocate Cd-mediated pathological changes in blood and urine parameters in mice. In addition, histological data supported the pharmacological findings. In silico chemometric analyses predicted the possible interactions between RA and different signal proteins and anticipated drug-likeness characteristics of RA. Hence, RA can potentially be applied as a therapeutic agent to treat Cd-mediated nephrotoxicity in future.

Carnosic Acid Attenuates Cadmium Induced Nephrotoxicity by Inhibiting Oxidative Stress, Promoting Nrf2/HO-1 Signalling and Impairing TGF-ß1/Smad/Collagen IV Signalling.

Cadmium (Cd) imparts nephrotoxicity via triggering oxidative stress and pathological signal transductions in renal cells. The present study was performed to explore the protective mechanism of carnosic acid (CA), a naturally occurring antioxidant compound, against cadmium chloride (CdCl2)-provoked nephrotoxicity employing suitable in vitro and in vivo assays. CA (5 µM) exhibited an anti-apoptotic effect against CdCl2 (40 µM) in normal kidney epithelial (NKE) cells evidenced from cell viability, image, and flow cytometry assays. In this study, CdCl2 treatment enhanced oxidative stress by triggering free radical production, suppressing the endogenous redox defence system, and inhibiting nuclear factor erythroid 2-related factor 2 (Nrf2) activation in NKE cells and mouse kidneys. Moreover, CdCl2 treatment significantly endorsed apoptosis and fibrosis via activation of apoptotic and transforming growth factor (TGF)-ß1/mothers against decapentaplegic homolog (Smad)/collagen IV signalling pathways, respectively. In contrast, CA treatment significantly attenuated Cd-provoked nephrotoxicity via inhibiting free radicals, endorsing redox defence, suppressing apoptosis, and inhibiting fibrosis in renal cells in both in vitro and in vivo systems. In addition, CA treatment significantly (p < 0.05-0.01) restored blood and urine parameters to near-normal levels in mice. Histological findings further confirmed the protective role of CA against Cd-mediated nephrotoxicity. Molecular docking predicted possible interactions between CA and Nrf2/TGF-ß1/Smad/collagen IV. Hence, CA was found to be a potential therapeutic agent to treat Cd-mediated nephrotoxicity.

Natural Products as Alternative Choices for P-Glycoprotein (P-gp) Inhibition.

Multidrug resistance (MDR) is regarded as one of the bottlenecks of successful clinical treatment for numerous chemotherapeutic agents. Multiple key regulators are alleged to be responsible for MDR and making the treatment regimens ineffective. In this review, we discuss MDR in relation to P-glycoprotein (P-gp) and its down-regulation by natural bioactive molecules. P-gp, a unique ATP-dependent membrane transport protein, is one of those key regulators which are present in the lining of the colon, endothelial cells of the blood brain barrier (BBB), bile duct, adrenal gland, kidney tubules, small intestine, pancreatic ducts and in many other tissues like heart, lungs, spleen, skeletal muscles, etc. Due to its diverse tissue distribution, P-gp is a novel protective barrier to stop the intake of xenobiotics into the human body. Over-expression of P-gp leads to decreased intracellular accretion of many chemotherapeutic agents thus assisting in the development of MDR. Eventually, the effectiveness of these drugs is decreased. P-gp inhibitors act by altering intracellular ATP levels which are the source of energy and/or by affecting membrane contours to increase permeability. However, the use of synthetic inhibitors is known to cause serious toxicities. For this reason, the search for more potent and less toxic P-gp inhibitors of natural origin is underway. The present review aims to recapitulate the research findings on bioactive constituents of natural origin with P-gp inhibition characteristics. Natural bioactive constituents with P-gp modulating effects offer great potential for semi-synthetic modification to produce new scaffolds which could serve as valuable investigative tools to recognize the function of complex ABC transporters apart from evading the systemic toxicities shown by synthetic counterparts. Despite the many published scientific findings encompassing P-gp inhibitors, however, this article stand alones because it provides a vivid picture to the readers pertaining to Pgp inhibitors obtained from natural sources coupled with their mode of action and structures. It provides first-hand information to the scientists working in the field of drug discovery to further synthesise and discover new P-gp inhibitors with less toxicity and more efficacies.

Carnosic acid attenuates doxorubicin-induced cardiotoxicity by decreasing oxidative stress and its concomitant pathological consequences.

This work aimed to reveal the protective mechanism of CA against Dox (doxorubicin)-induced cardiotoxicity. In isolated murine cardiomyocytes, CA showed a concentration-dependent cytoprotective effect against Dox. Dox treatment significantly (p < 0.01) increased the formation of reactive oxygen species (ROS), increased NO levels, activated NADPH oxidase, and inactivated the cellular redox defense mechanism in cardiac cells, resulting in augmented oxidative stress in cardiomyocytes and rat hearts. Dox-induced oxidative stress significantly (p < 0.01) upregulated several pathogenic signal transductions, which induced apoptosis, inflammation, and fibrosis in cardiomyocytes and murine hearts. In contrast, CA significantly (p < 0.05-0.01) reciprocated Dox-induced cardiac apoptosis, inflammation, and fibrosis by suppressing oxidative stress and interfering with pathological signaling events in both isolated murine cardiomyocytes and rat hearts. CA treatment significantly (p < 0.05-0.01) countered Dox-mediated pathological changes in blood parameters in rats. Histological examinations backed up the pharmacological findings. In silico chemometric investigations predicted potential interactions between CA and studied signal proteins, as well as the drug-like features of CA. Thus, it would be concluded that CA has the potential to be regarded as an effective agent to alleviate Dox-mediated cardiotoxicity in the future.

Carnosic Acid, a Natural Diterpene, Attenuates Arsenic-Induced Hepatotoxicity via Reducing Oxidative Stress, MAPK Activation, and Apoptotic Cell Death Pathway.

The present studies have been executed to explore the protective mechanism of carnosic acid (CA) against NaAsO2-induced hepatic injury. CA exhibited a concentration dependent (1-4 µM) increase in cell viability against NaAsO2 (12 µM) in murine hepatocytes. NaAsO2 treatment significantly enhanced the ROS-mediated oxidative stress in the hepatic cells both in in vitro and in vivo systems. Significant activation of MAPK, NF-κB, p53, and intrinsic and extrinsic apoptotic signaling was observed in NaAsO2-exposed hepatic cells. CA could significantly counteract with redox stress and ROS-mediated signaling and thereby attenuated NaAsO2-mediated hepatotoxicity. NaAsO2 (10 mg/kg) treatment caused significant increment in the As bioaccumulation, cytosolic ATP level, DNA fragmentation, and oxidation in the liver of experimental mice (n = 6). The serum biochemical and haematological parameters were significantly altered in the NaAsO2-exposed mice (n = 6). Simultaneous treatment with CA (10 and 20 mg/kg) could significantly reinstate the NaAsO2-mediated toxicological effects in the liver. Molecular docking and dynamics predicted the possible interaction patterns and the stability of interactions between CA and signal proteins. ADME prediction anticipated the drug-likeness characteristics of CA. Hence, there would be an option to employ CA as a new therapeutic agent against As-mediated toxic manifestations in future.

Edible leaf extract of Ipomoea aquatica Forssk. (Convolvulaceae) attenuates doxorubicin-induced liver injury via inhibiting oxidative impairment, MAPK activation and intrinsic pathway of apoptosis.

Ipomoea aquatica Forssk. (Convolvulaceae) is an aquatic vegetable traditionally employed against toxic effects of xenobiotics. The present study has been designed to investigate the molecular mechanism underlying the beneficial role of the edible (aqueous) leaf extract of I. aquatica (AEIA) against doxorubicin (Dox)-induced liver injury. AEIA exhibited a dose-dependent (∼400 µg/ml) increase in cell viability against Dox (1 µM) in isolated rodent hepatocytes. AEIA (400 µg/ml) prevented the Dox-induced increase in ROS, redox imbalance, and activation of mitogen activated protein kinases (MAPK) and intrinsic pathway of apoptosis in hepatocytes. In the in vivo assay, administration of AEIA (100 mg/kg, p.o.) against Dox (3 mg/kg, i.p.) also reduced the oxidative impairment, DNA fragmentation, ATP formation, and up-regulated the mitochondrial co-enzymes Qs in the liver tissues of Wistar rats. Histological assessments were in agreement with the biochemical findings. Substantial quantities of phyto-antioxidants in AEIA may mediate its beneficial function against Dox-induced liver injury.

Protocatechuic Acid, a Phenolic from Sansevieria roxburghiana Leaves, Suppresses Diabetic Cardiomyopathy via Stimulating Glucose Metabolism, Ameliorating Oxidative Stress, and Inhibiting Inflammation.

Persistent hyperglycemia, impairment of redox status and establishment of inflammatory pathophysiology integrally play important role in the pathogenesis of diabetic cardiomyopathy (DC). Present study examined the therapeutic potential of protocatechuic acid isolated from the Sansevieria roxburghiana rhizomes against DC employing rodent model of type 2 diabetes (T2D). T2D was induced by high fat diet + a low-single dose of streptozotocin (35 mg/kg, i.p.). T2D rats exhibited significantly (p < 0.01) high fasting blood glucose level. Alteration in serum lipid profile (p < 0.01) and increased levels of lactate dehydrogenase (p < 0.01) and creatine kinase (p < 0.01) in the sera of T2D rats revealed the occurrence of hyperlipidemia and diabetic pathophysiology. A significantly (p < 0.01) high levels of serum C-reactive protein and pro-inflammatory mediators revealed the establishment of inflammatory occurrence in T2D rats. Besides, significantly high levels of troponins in the sera revealed the establishment of cardiac dysfunctions in T2D rats. However, protocatechuic acid (50 and 100 mg/kg, p.o.) treatment could significantly reverse the changes in serum biochemical parameters related to cardiac dysfunctions. Molecular mechanism studies demonstrated impairment of signaling cascade, IRS1/PI3K/Akt/AMPK/p 38/GLUT4, in glucose metabolism in the skeletal muscle of T2D rats. Significant (p < 0.01) activation of polyol pathway, enhanced production of AGEs, oxidative stress and up-regulation of inflammatory signaling cascades (PKC/NF-κB/PARP) were observed in the myocardial tissue of T2D rats. However, protocatechuic acid (50 and 100 mg/kg, p.o.) treatment could significantly (p < 0.05-0.01) stimulate glucose metabolism in skeletal muscle, regulated glycemic and lipid status, reduced the secretion of pro-inflammatory cytokines, and restored the myocardial physiology in T2D rats near to normalcy. Histological assessments were also in agreement with the above findings. In silico molecular docking study again supported the interactions of protocatechuic acid with different signaling molecules, PI3K, IRS, Akt, AMPK PKC, NF-κB and PARP, involved in glucose utilization and inflammatory pathophysiology. In silico ADME study predicted that protocatechuic acid would support the drug-likeness character. Combining all, results would suggest a possibility of protocatechuic acid to be a new therapeutic agent for DC in future.

Cytoprotective and Antioxidant Effects of an Edible Herb, Enhydra fluctuans Lour. (Asteraceae), against Experimentally Induced Lead Acetate Intoxication.

BACKGROUND: Enhydra fluctuans Lour. (Asteraceae), an edible aquatic herb, is traditionally employed against toxic effects of heavy metals in India. The present study was planned to discover the protective effect of edible extract of E. fluctuans (AEEF) against Pb toxicity. METHODS: The cytoprotective role of AEEF was determined on murine hepatocytes employing MTT assay and Hoechst staining. The effects on lipid peroxidation, protein carbonylation, endogenous redox systems and the transcription levels of apoptotic proteins were studied after incubating the hepatocytes with AEEF (400 µg/ml) + Pb-acetate (6.8 µM). The defensive role of AEEF (100 mg/kg) against Pb-acetate (5 mg/kg) intoxication was measured in mice by in vivo assays. Biochemical, haematological and histological parameters, intracellular Pb burden and redox status were measured. RESULTS: AEEF exhibited a concentration dependent cytoprotective effect against Pb-induced cytotoxicity in vitro. Pb-acetate incubation significantly (p < 0.01) altered the extents of ROS production ↑, protein carbonylation ↑, lipid peroxidation ↑, endogenous antioxidant enzymes ↓ and GSH ↓ in vitro. Besides, Pb-acetate significantly (p < 0.01) induced apoptosis in the hepatocytes apparent from the altered expressions of apoptotic proteins viz. Apaf-1 ↑, Bad ↑, Bcl-2 ↓, Cyt C ↑, cleaved caspases↑, Bid ↑ and Fas ↑. However, AEEF (400 µg/ml) could significantly (p < 0.05-0.01) attenuate the Pb-acetate mediated toxic manifestation in vitro. In in vivo assay, Pb-acetate (5 mg/kg) treated mice exhibited significantly (p < 0.01) high intracellular Pb content. A high Pb-burden within the tissues caused significant (p < 0.05-0.01) patho-physiological alterations viz. ROS production ↑, protein carbonylation↑, lipid peroxidation ↑, DNA fragmentation ↑, ATP formation ↑, mitochondrial co-enzymes Q ↓, endogenous antioxidant enzymes ↓ and GSH ↓ within the selected tissues. The haematological and serum biochemical parameters were significantly (p < 0.05-0.01) different in the Pb-acetate treated mice. Finally, histological assessment imposed significant toxic occurrence within the organs of Pb-intoxicated animals. However, concurrent administration of AEEF (100 mg/kg) could significantly (p < 0.05-0.01) reinstate the Pb-acetate mediated toxicity. CONCLUSION: Presence of metal chelators and phyto-antioxidants within AEEF would offer overall protection through promoting Pb clearance coupled with restoring redox balance.

Water Spinach, Ipomoea aquatic (Convolvulaceae), Ameliorates Lead Toxicity by Inhibiting Oxidative Stress and Apoptosis.

BACKGROUND: Ipomoea aquatica (Convolvulaceae), an aquatic edible plant, is traditionally used against heavy metal toxicity in India. The current study intended to explore the protective role of edible (aqueous) extract of I. aquatica (AEIA) against experimentally induced Pb-intoxication. METHODS: The cytoprotective role of AEIA was measured on mouse hepatocytes by cell viability assay followed by Hoechst staining and flow cytometric assay. The effect on ROS production, lipid peroxidation, protein carbonylation, intracellular redox status were measured after incubating the hepatocytes with Pb-acetate (6.8 µM) along with AEIA (400 µg/ml). The effects on the expressions of apoptotic signal proteins were estimated by western blotting. The protective role of AEIA was measured by in vivo assay in mice. Haematological, serum biochemical, tissue redox status, Pb bioaccumulation and histological parameters were evaluated to estimate the protective role of AEIA (100 mg/kg) against Pb-acetate (5 mg/kg) intoxication. RESULTS: Pb-acetate treated hepatocytes showed a gradual reduction of cell viability dose-dependently with an IC50 value of 6.8 µM. Pb-acetate treated hepatocytes exhibited significantly enhanced levels (p < 0.01) of ROS production, lipid peroxidation, protein carbonylation with concomitant depletion (p < 0.01) of antioxidant enzymes and GSH. However, AEIA treatment could significantly restore the aforementioned parameters in murine hepatocytes near to normalcy. Besides, AEIA significantly reversed (p < 0.05-0.01) the alterations of transcription levels of apoptotic proteins viz. Bcl 2, Bad, Cyt C, Apaf-1, cleaved caspases [caspase 3, caspase 8 and caspase 9], Fas and Bid. In in vivo bioassay, Pb-acetate treatment caused significantly high intracellular Pb burden and oxidative pressure in the kidney, liver, heart, brain and testes in mice. In addition, the haematological and serum biochemical factors were changed significantly in Pb-acetate-treated animals. AEIA treatment restored significantly the evaluated-parameters to the near-normal position. CONCLUSION: The extract may offer the protective effect via counteracting with Pb mediated oxidative stress and/or promoting the elimination of Pb by chelating. The presence of substantial quantities of flavonoids, phenolics and saponins would be responsible for the overall protective effect.