Shailendra Pratap Singh

Comparative study of genotoxicity and tissue distribution of nano and micron sized iron oxide in rats after acute oral treatment.

Though nanomaterials (NMs) are being utilized worldwide, increasing use of NMs have raised concerns over their safety to human health and environment. Iron oxide (Fe(2)O(3)) NMs have important applications. The aim of this study was to assess the genotoxicity of Fe(2)O(3)-30nm and Fe(2)O(3)-bulk in female Wistar rats. Fe(2)O(3)-30nm was characterized by using transmission electron microscopy, dynamic light scattering, laser Doppler velocimetry and surface area analysis. The rats were treated orally with the single doses of 500, 1000, 2000mg/kg bw of Fe(2)O(3)-30nm and Fe(2)O(3) -bulk. The genotoxicity was evaluated at 6, 24, 48 and 72h by the comet assay in leucocytes, 48 and 72h by micronucleus test (MNT) in peripheral blood cells, 18 and 24h by chromosomal aberration (CA) assay and 24 and 48h by MNT in bone marrow cells. The biodistribution of iron (Fe) was carried out at 6, 24, 48 and 72h after treatment in liver, spleen, kidney, heart, brain, bone marrow, urine and feces by using atomic absorption spectrophotometry. The % tail DNA, frequencies of micronuclei and CAs were statistically insignificant (p>0.05) at all doses. These results suggest that Fe(2)O(3)-30nm and Fe(2)O(3)-bulk was not genotoxic at the doses tested. Bioavailability of Fe was size and dose dependent in all the tissues from the groups exposed to Fe(2)O(3)-30nm. Fe(2)O(3) NMs were able to enter in the organs and the rats are biocompatible with much higher concentration of Fe. However, the accumulated Fe did not cause significant genotoxicity. This study provides additional knowledge about the toxicology of Fe(2)O(3) NMs.

Toxicity Study of Cerium Oxide Nanoparticles in Human Neuroblastoma Cells

The present study consisted of cytotoxic, genotoxic, and oxidative stress responses of human neuroblastoma cell line (IMR32) following exposure to different doses of cerium oxide nanoparticles (CeO2 NPs; nanoceria) and its microparticles (MPs) for 24 hours. Cytotoxicity was evaluated by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide and lactate dehydrogenase assays whereas genotoxicity was assessed using the cytokinesis-block micronucleus and comet assays. A battery of assays including lipid peroxidation, reactive oxygen species (ROS), hydrogen peroxide, reduced glutathione, nitric oxide, glutathione reductase, glutathione peroxidase, superoxide dismutase, catalase, and glutathione S-transferase were performed to test the hypothesis that ROS was responsible for the toxicity of nanoceria. The results showed that nanosized CeO2 was more toxic than cerium oxide MPs. Hence, further study on safety evaluation of CeO2 NPs on other models is recommended.

Toxicity assessment of manganese oxide micro and nanoparticles in Wistar rats after 28 days of repeated oral exposure

In the near future, nanotechnology is envisaged for large‐scale use. Hence health and safety issues of nanoparticles (NPs) should be promptly addressed. Twenty‐eight‐day oral toxicity, genotoxicity, biochemical alterations, histopathological changes and tissue distribution of nano and microparticles (MPs) of manganese oxide (MnO2) in Wistar rats was studied. Genotoxicity was assessed using comet, micronucleus and chromosomal aberration assays. The results demonstrated a significant increase in DNA damage in leukocytes, micronuclei and chromosomal aberrations in bone marrow cells after exposure of MnO2‐NPs at 1000, 300u2009mgu2009kg–1 bw per day and MnO2‐MPs at the dose of 1000u2009mgu2009kg–1 bw per day. Our findings showed acetylcholinestrase inhibition at 1000 as well as at 300u2009mgu2009kg–1 bw per day in blood and with all the doses in the brain indicating the toxicity of MnO2‐NPs. Further, the doses significantly inhibited different ATPases in the brain P2 fraction. Significant changes were observed in aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) in the liver, kidney and serum in a dose‐dependent manner. MnO2‐MPs at 1000u2009mgu2009kg–1 bw per day were found to induce significant alterations in biochemical enzymes. A significant distribution was found in all the tissues in a dose‐dependent manner. MnO2‐NPs showed a much higher absorptivity and tissue distribution as compared with MnO2‐MPs. A large fraction of MnO2‐NPs and MnO2‐MPs was cleared by urine and feces. Histopathological analysis revealed that MnO2‐NPs caused alterations in liver, spleen, kidney and brain. The MnO2‐NPs induced toxicity at lower doses compared with MnO2‐MPs. Further, this study did not display gender differences after exposure to MnO2‐NPs and MnO2‐MPs. Therefore, the results suggested that prolonged exposure to MnO2 has the potential to cause genetic damage, biochemical alterations and histological changes. Copyright

Biochemical alterations induced by acute oral doses of iron oxide nanoparticles in Wistar rats

Magnetic iron oxide nanoparticles with appropriate surface chemistry have been widely used with potential new applications in biomedical industry. Therefore, the aim of this study was to assess the size-, dose-, and time-dependent effects, after acute oral exposure to iron oxide-30 NP (Fe2O3-30), on various biochemical enzyme activities of clinical significances in a female Wistar rat model. Rats were exposed to three different doses (500, 1,000, and 2,000u2009mg/kg) of Fe2O3-30 and Fe2O3-Bulk along with control. Fe2O3-30 had no effect on growth, behavior, and nutritional performance of animals. Fe2O3-30 caused significant inhibition of acetylcholinestrase in red blood cells as well as in brains of treated rats. Further, more than 50% inhibition of total, Na+-K+, Mg2+, and Ca2+-ATPases activities, as observed in brains of exposed female rats, may be the result of disturbances in cellular physiology and the iono-regulatory process. Activation of the hepatotoxicity marker enzymes, aspartate aminotransferase and alanine aminotransferase, was recorded in serum and liver, whereas inhibition was observed in kidney. Similarly, enhancement of lactate dehydrogenase activity was observed in serum and liver; however, a decrease in enzyme levels was observed in kidneys of Fe2O3-30-treated rats. On the other hand, Fe2O3-Bulk did not depict any significant changes in these biochemical parameters, and alterations were near to control. Therefore, this study suggests that exposure to nanosize particles at acute doses may cause adverse changes in animal biochemical profiles. The use of the rat model signifies the correlation with the human system.

Synthesis, biological evaluation and molecular modeling studies of some novel thiazolidinediones with triazole ring

A new series of thiazolidinedione derivatives were synthesized and evaluated for in vitro α-glucosidase inhibition and anticancer activities. Compounds 3d, 3e and 3j showed potential α-glucosidase inhibition with IC₅₀ values ranging between 0.1 and 0.3 μg/ml whereas compounds 3i, 3j and 3k have showed better anticancer activity towards human cancer cell lines IMR-32 (neuroblastoma), Hep-G2 (hepatoma) and MCF-7 (breast). Molecular docking studies revealed compounds 3d, 3e and 3j are potent inhibitors of α-glucosidase and also showed compliance with standard parameters of drug likeness.

Development of a cell-based nonradioactive glucose uptake assay system for SGLT1 and SGLT2

Sodium-dependent glucose cotransporters (SGLT1 and SGLT2), which have a key role in the absorption of glucose in the kidney and/or gastrointestinal tract, have been proposed as a novel therapeutic strategy for diabetes and cardiomyopathy. Here we developed a simple cell-based, nonradioactive method for functional screening of SGLT1 and SGLT2 inhibitors. Stable cell lines expressing human SGLT1 and SGLT2 were established by transfecting HEK293 cells with vectors (pCMV6-Neo) having full-length human SGLT1 and SGLT2 and selecting the positive clones following neomycin treatment. We confirmed the gene expression of SGLT1 and SGLT2 by reverse transcription polymerase chain reaction (RT-PCR) and immunoblotting. Furthermore, to analyze the function of SGLTs, we incubated stable cell lines with 2-deoxyglucose or fluorescent d-glucose analog (2-NBDG) and performed glucose uptake assay. A significant (P<0.001) increase in glucose uptake was observed in both cell lines. The increased glucose uptake in both cell lines was completely inhibited when treated with nonspecific SGLT1/SGLT2 inhibitors and phlorizin (100μM), but not when treated with nonspecific sodium-independent facilitative glucose transporter (GLUT) inhibitors (100μM). Taken together, our data suggest that cell-based methods developed for screening SGLT1/SGLT2 inhibitors are phlorizin sensitive and specific for respective glucose transporters. This assay provides a simple and rapid method for identifying novel and selective SGLT inhibitors.

Genotoxicity of nano- and micron-sized manganese oxide in rats after acute oral treatment.

The use of nanotechnology has led to rapid growth in various areas. Manganese oxide (MnO2) nanomaterials (NMs) are typically used for biomedical applications. However, characterizing the potential human health effects of MnO2 NMs is required before fully exploiting these materials. The aim of this study was to investigate the acute oral toxicity of MnO2 NMs and MnO2-bulk particles in female albino Wistar rats. The genotoxic effects were examined using comet, micronucleus and chromosomal aberration assays. Nanosized MnO2 (45nm) significantly (p<0.01) increased DNA damage in peripheral blood leukocytes and micronuclei and enhanced chromosomal aberrations in the bone marrow cells at 1000mg/kg bw. These findings showed that the neurotoxicity of MnO2-45nm in the brain and red blood cells, as determined through acetylcholinesterase activity, was significantly (p<0.01) inhibited at 1000 and 500mg/kg bw doses. MnO2-45nm disrupted the physicochemical state and neurological system of the animals through alterations in ATPases via the total Na(+)-K(+), Mg(2+) and Ca(2+) levels in the brain P2 fraction. In addition, 500 and 1000mg/kg bw doses of MnO2-45nm caused significant changes in AST, ALT and LDH levels in the liver, kidney and serum of treated rats. Significant tissue distribution was found in all tissues in a dose- and time-dependent manner. MnO2-45nm exhibited much higher absorptivity and tissue distribution compared with MnO2-bulk. A large fraction of MnO2-45nm was cleared in the urine and feces. The histopathological analysis revealed that MnO2-45nm caused alterations in the liver, spleen and brain. These findings will provide fundamental information regarding the potential toxicities and biodistribution of nano and bulk MnO2 generated through acute oral treatment.

Monitoring of oxidative stress in nurses occupationally exposed to antineoplastic drugs

Antineoplastic drugs (ANDs) have been in clinical usage for more than five decades. The nonselective mechanism of action of ANDs between cancerous and noncancerous cells had well documented side effects such as acute symptoms, reproductive health issues, and potential cancer development in healthcare workers as a result of occupational exposure. The anticancer mechanism of ANDs is the generation of reactive oxygen species (ROS) which are responsible for various side effects in patients undergoing chemotherapy and the healthcare personnel occupationally exposed to them. ROS have potential to damage lipids, DNA, proteins, and so on leading to oxidative stress condition. The aim of this study was to evaluate the possible oxidative stress effect of antineoplastic drugs in nurses who routinely handle ANDs in an oncology hospital in south India. Malondialdehyde levels, reduced glutathione content, and glutathione S-transferase activity were analyzed in serum collected from 60 female nurses handling ANDs and compared with equal number of healthy volunteers matched by age and sex except AND exposure. The results showed statistically significant (P < 0.05) increase in malondialdehyde levels in the serum of exposed nurses. However, glutathione content and glutathione S-transferase activity was significantly decreased in these nurses. Our study suggests that the nurses occupationally exposed to ANDs were susceptible to the oxidative stress and emphasizes the need for a harmonized safe handling approach that assures minimal risk to the working nurses.

In vivo assessment of genotoxic effects of Annona squamosa seed extract in rats.

Widespread use of pesticides represents a potential risk to human and environmental health. Hence, biopesticides from plants are some of the future strategies for plant protection. In this regard, a seed extract of Annona squamosa was prepared and found to be a promising pesticide. In order to establish the inherent toxicity and non-target safety required for registration and marketing of pesticides, toxicological studies are conducted. The genotoxicity potential was evaluated in rats with 75, 150 and 300 mg/kg Annona squamosa by the comet assay in leucocytes, micronucleus and chromosomal aberration tests in bone marrow. We also studied the effects of 300 mg/kg of extract on lipid peroxidation, reduced glutathione level and glutathione S transferase activity in liver, lungs, brain, kidneys, heart and spleen of treated rats. The comet assay showed a statistically significant dose related increase in DNA migration. The micronucleus and chromosomal aberration tests revealed a significant induction in frequency of micronuclei and chromosomal aberrations at 150 and 300 mg/kg. Annona squamosa treatment significantly enhanced lipid peroxidation, decreased glutathione and glutathione S transferase levels revealing the oxidative stress condition. Our results warrant careful use of Annona squamosa seed extract as a biopesticide till more tests are carried out.

Genotoxic effects of chromium oxide nanoparticles and microparticles in Wistar rats after 28 days of repeated oral exposure

The nanotechnology industry has advanced rapidly in the last 10xa0years giving rise to the growth of the nanoparticles (NPs) with great potential in various arenas. However, the same properties that make NPs interesting raise concerns because their toxicity has not been explored. The in vivo toxicology of chromium oxide (Cr2O3)–NPs is not known till date. Therefore, this study investigated the 28-day repeated toxicity after 30, 300 and 1000xa0mg/kg body weight (bw)/day oral treatment with Cr2O3–NPs and Cr2O3 microparticles (MPs) in Wistar rats. The mean size of Cr2O3–NPs and Cr2O3–MPs was 34.89u2009±u20092.65xa0nm and 3.76u2009±u20093.41xa0μm, respectively. Genotoxicity was assessed using comet, micronucleus and chromosomal aberration (CA) assays. The results revealed a significant increase in DNA damage in peripheral blood leucocytes and liver, micronuclei and CA in bone marrow after exposure of 300 and 1000xa0mg/kg doses of Cr2O3–NPs and Cr2O3–MPs only at 1000xa0mg/kg bw/day. Cr biodistribution was observed in all the tissues in a dose-dependent manner. The maximum amount of Cr was found in the kidneys and least in the brain of the treated rats. More of the Cr was excreted in the faeces than in the urine. Furthermore, nanotreated rats displayed much higher absorption and tissue accumulation. These findings provide initial data of the probable genotoxicity and biodistribution of NPs and MPs of Cr2O3 generated through repeated oral treatment.