CBMN Assay Research Articles

Heterocyclic phytometabolites formononetin and arbutin prevent in vitro oxidative and alkylation-induced mutagenicity

Phenolic phytometabolites are promising bioactive compounds for management of genomic instability related diseases. Formononetin (FMN) and arbutin (ARB) are found in several plant sources. Our goal was to investigate the safety and efficacy of FMN and ARB using in vitro both standardized and alternative toxicogenetic methods. FMN and ARB were evaluated through the OECD’S guidelines No. 471 (Bacterial Reverse Mutation Test –Salmonella/microsome) and No. 487 (In vitro Mammalian Micronucleus Test – CBMN assay), accordingly to the mentioned recommendations. Also, antimutagenicity of FMN and ARB was assessed in S. Typhimurium strains TA98, TA100 and TA1535, following pre-, co- and post- treatment protocols. Liver human lineages HepG2 and F C3H were assayed for cytotoxicity after exposure to FMN and ARB (24, 48 and 72 h) using in vitro WST-1 test. ARB showed no mutagenicity in the Salmonella/microsome test under both metabolic conditions (in presence or absence of 4 % S9 mix), but FMN was cytotoxic to the TA97 and TA100 strains after metabolic activation. Under this same condition, FMN induced an increase in the mutagenic index of strain TA1535 at two of the highest tested concentrations. Even so, ARB and FMN exhibited protection against the induced alkylation of DNA in multiple action modes. In the antimutagenicity assay, FMN reached the maximum of 80 % of oxidative-provoked mutagenicity reduction in TA98 strain in co-treatment with known mutagen, besides 69 % of reduction in TA100 in the same exposure condition. ARB showed up to reduce induced mutagenicity in strains TA100 and TA1535, reaching percentages from 55 % to 100 % of antimutagenicity in all of the tested exposure models against alkylating agent. In the CBMN assay, no increase in micronuclei formation was observed. The results suggest that FMN and ARB prevent DNA from mutation using multi-targeted antimutagenic roles. Finally, our data suggests that FMN and ARB are not genotoxic and presented encouraging antimutagenicity action in vitro, being promising compounds for use in genomic instability-related diseases therapeutics.

Assessment of Biological Damage Induced during Multidetector Computed Tomography (MDCT) Examination.

Computed tomography (CT) is known for its non-invasiveness, fast procedure, and also for providing detailed diagnostic information to physicians. It also utilises low-dose-rate ionising radiation (X-rays) as a source for imaging. Multidetector computed tomography (MDCT) is an advanced system that uses iodinated contrast media for more accurate diagnostic results. Studies suggest using these contrasts will lead to greater radiation adsorption with significant DNA damage. No studies have been taken comparing the physical dose with the biological effect. The present study sheds light on the same by assessing the biological effect of CT with and without contrast intervention. The present study is timebound; thus, 21 participants attending for CT thorax and abdomen with no history of any cancer were included. The same participants underwent both pre-contrast and post-contrast studies. The blood sample was taken before the procedure and used as a control. Physical parameters like DLP and CTDI obtained from the instrument were compared with the MN frequency obtained (CBMN Assay). The study showed a significant increase (p-value < 0.05) in the Physical and MN frequency in the Post-Contrast group compared to the pre-contrast group. Although a positive correlation was observed between pre and post-contrast groups, the results were not found to be statistically significant (p-value < 0.05). The study confirms increased physical dose and MN frequency upon contrast intervention. This study recommends the judicial use of MDCT in disease diagnostics.

Aqueous sage leave extract attenuates inflammation and oxidant-induced genotoxicity in human peripheral blood mononuclear cells.

Traditional medicine has used sage (Salvia officinalis L.) preparations for centuries to prevent and treat various inflammatory and oxidative stress-induced conditions. The aim of this in vitro study was to determine the bioactive properties of a sage leave extract obtained with environmentally friendly aqueous extraction and lyophilisation in primary human peripheral blood cells. To that end we measured the total phenolic and flavonoid content (TPC and TFC, respectively) with gas chromatography-mass spectrometry (GC-MS). Non-cytotoxic concentrations determined with the trypan blue assay were used to assess the antioxidant (DPPH, ABTS, and PAB assay), antigenotoxic (CBMN assay), immunomodulatory (IL-1β and TNF-α), and neuroprotective effects (AChE inhibition). The extract contained high TPC (162 mg GAE/g of dry extract) and TFC (39.47 mg QE/g of dry extract) concentrations, while β-thujone content was unexpectedly low (below 0.9 %). Strong radical-scavenging activity combined with glutathione reductase activation led to a decrease in basal and H2O2-induced oxidative stress and DNA damage. A decrease in TNF-α and increase in IL-1β levels suggest complex immunomodulatory response that could contribute to antioxidant and, together with mild AChE inhibition, neuroprotective effects. Overall, this study has demonstrated that aqueous sage leave extract reduces the levels of thujone, 1,8-cineole, pinene, and terpene ketones that could be toxic in high concentrations, while maintaining high concentrations of biologically active protective compounds which have a potential to prevent and/or treat inflammatory and oxidative stress-related conditions.

RABiT-III: an Automated Micronucleus Assay at a Non-Specialized Biodosimetry Facility.

Micronuclei, detected through the cytokinesis-block micronucleus assay, are valuable indicators of ionizing radiation exposure, especially in short-term lymphocyte cultures. The peripheral human blood lymphocyte assay is recognized as a prime candidate for automated biodosimetry. In a prior project at the Columbia University Center for Radiological Research, we automated this assay using the 96-well ANSI/SLAS microplate standard format and relied on established biotech robotic systems named Rapid Automated Biodosimetry Tool (RABiT). In this study, we present the application of a similar automated biotech setup at an external high-throughput facility (RABiT-III) to implement the same automated cytokinesis-block micronucleus assay. Specifically, we employed the Agilent BRAVO liquid-handling system and GE IN Cell Analyzer 6000 imaging system in conjunction with the PerkinElmer Columbus image data storage and analysis system. Notably, this analysis system features an embedded PhenoLOGIC machine learning module, simplifying the creation of cell classification algorithms for CBMN assay image analysis and enabling the generation of radiation dose-response curves. This investigation underscores the adaptability of the RABiT-II CBMN protocol to diverse RABiT-III biotech robotic platforms in non-specialized biodosimetry centers. Furthermore, it highlights the advantages of machine learning in rapidly developing algorithms crucial for the high-throughput automated analysis of RABiT-III images.

Effect of iron and calcium on radiation sensitivity in prostate cancer patients relative to controls.

High intake of red meat and/or dairy products may increase the concentration of iron and calcium in plasma-a risk factor for prostate cancer (PC). Despite our understandings of nutrients and their effects on the genome, studies on the effects of iron and calcium on radiation sensitivity of PC patients are lacking. Therefore, we tested the hypothesis that high plasma levels of iron and calcium could increase baseline or radiation-induced DNA damage in PC patients relative to healthy controls. The present study was performed on 106 PC patients and 132 age-matched healthy individuals. CBMN assay was performed to measure mi-cronuclei (MN), nucleoplasmic bridges (NPBs), and nuclear buds (NBuds) in lymphocytes. Plasma concentrations of iron and calcium were measured using inductively coupled plasma atomic emission spectroscopy. MN, NPBs, and NBuds induced by radiation ex vivo were significantly higher in PC patients with high plasma iron (P = .004, P = .047, and P = .0003, respectively) compared to healthy controls. Radiation-induced MN and NBuds frequency were also significantly higher in PC patients (P = .001 and P = .0001, respectively) with high plasma calcium levels relative to controls. Furthermore, radiation-induced frequency of NBuds was significantly higher in PC patients (P < .0001) with high plasma levels of both iron and calcium relative to controls. Our results support the hypothesis that high iron and calcium levels in plasma increases the sensitivity to radiation-induced DNA damage and point to the need of developing nutrition-based strategies to minimize DNA damage in normal tissue of PC patients undergoing radiotherapy.

A standardized and automated CBMN assay for biological dosimetry: the CytoRADx™ system.

Major nuclear accidents can result in many casualties, and it is important to assess the absorbed radiation dose to support treatment decisions. Biological dosimetry (BD) allows retrospective determination of dose using biological markers. To achieve consistent cytogenetic assay results across labs, the current practice requires each lab to generate periodic, unique calibration curves using in vitro dose-effect experiments. Here, we present CytoRADx™, a standardized biodosimetry system that integrates automated dose calculation in a high-throughput platform without the need for lab-specific calibration curves. CytoRADx consists of an improved, standardized Cytokinesis Block Micronucleus assay combined with automated analysis utilizing an established slide scanning device. We tested CytoRADx for accuracy and reproducibility across different instruments, sites, days and operators. Our results demonstrate that CytoRADx eliminates the time-consuming, lab-specific calibration curves, allowing multiple laboratories to obtain consistent results and to distribute the testing burden in the event of a large-scale accident.

Evaluating the role of cytokinesis-block micronucleus assay as a biomarker for oxidative stress-inducing DNA damage in type 2 diabetes mellitus patients

BackgroundType 2 Diabetes Miletus (T2DM) is a common metabolic and lifestyle disorder leading to increased oxidative stress and DNA damage. The present study aims to evaluate the feasibility of utilizing the cytokinesis-block micronucleus assay (CBMN) as a biomarker for assessing the DNA damage induced due to variations in oxidative stress.MethodologyThe study group includes diabetic (n = 50) and non-diabetic (n = 50) subjects. The assays for the diabetes-like fasting blood sugar, postprandial glucose and hemoglobin A1c (HbA1c), lipid profiling, and serum ferritin level along with c-reactive protein (CRP) were applied. Further, the CBMN assay was performed to evaluate the micronuclei present in the lymphocytes of control and T2DM groups.ResultsSignificant imbalance in the glycaemic index, dyslipidemia, increased ferritin levels, and CRP levels, with a significant increase of micronucleus frequency, was found in T2DM patients compared with the control group. Results suggest a trend of positive correlation between HbA1c and the micronuclei, indicating the assay’s potential importance as a biomarker for T2DM-induced risk assessment.ConclusionFrom the observed results, it can be suggested that the CBMN assay could be used to assess the risk of oxidative stress-induced DNA damage in high glycaemic index diabetic patients.

Measuring blood cell DNA damage using the PIG-A mutation and CBMN assay in pancreatic cancer patients: a pilot study.

Pancreatic cancer still has one of the worst prognoses of all solid malignancies, despite developments in cancer knowledge and care. Research into pancreatic cancer has not fully translated into clinical improvements and as a result, fewer than 1% of patients survive 10 years post-diagnosis. This bleak outlook for patients could be improved by earlier diagnosis. The human erythrocyte phosphatidylinositol glycan class A (PIG-A) assay monitors the mutation status of the X-linked PIG-A gene by measuring glycosyl phosphatidylinositol (GPI)-anchored proteins on the extracellular surface. We have previously identified an elevated PIG-A mutant frequency in oesophageal adenocarcinoma patients and here investigate whether this could be seen in a pancreatic cancer cohort, given the urgent need for novel pancreatic cancer biomarkers. In our pilot study, an elevated PIG-A mutant frequency (5.775 × 10-6 (95% CI 4.777-10) mutants per million) was seen in pancreatic cancer patients (n = 30) when compared to the non-cancer control group (n = 14) who had an erythrocyte mutant frequency of 4.211 × 10-6 (95% CI 1.39-5.16) mutants per million (p = 0.0052). A cut-off value of 4.7 mutants per million provided an AUROC of 0.7595 with a sensitivity of 70% and specificity of 78.57%. A secondary measure of DNA damage in an alternative blood cell population also showed an increase in peripheral lymphocytes using the cytokinesis-block micronucleus assay (p = 0.0164) (AUROC = 0.77, sensitivity = 72.22%, specificity = 72.73%). The micronucleus frequency and PIG-A status show some potential as blood-based biomarkers of pancreatic cancer, but further investigations of these DNA damage tests are required to assess their utility in pancreatic cancer diagnosis.

Chemoprotective Effect of Phyllanthus emblica Molybdenum Disulphide (MoS2) Nanoparticles on Bleomycin-Induced Cytotoxicity in Human Lymphocytes

Following the world's largest nuclear accident at Fukushima in 2011, a lot of research has been conducted to try to minimize the potential health effects of radiation exposure. One way to do this is to use natural antioxidants to protect against oxidative DNA damage. We studied the effects of an Phyllanthus emblica fruit crude extract MoS2 NPs against bleomycin-induced cytotoxicity in human lymphocytes to see if it increases the overall chemoprotective mechanism. We analyzed the frequency of micronuclei (MN) and DNA damage after treatment with different concentrations of bleomycin. We observed a significant increase in cell viability in lymphocytes treated with Phyllanthus emblica fruit crude extract MoS2 NPs and then exposed to bleomycin induced cytotoxic cells. When lymphocytes were pretreated with Phyllanthus emblica fruit crude extract MoS2 NPs (80 and 100 μg/mL), the frequency of micronuclei was decreased at all doses of bleomycin and DNA damage was decreased at 20 μg/mL of bleomycin induced cytotoxicity human lymphocytes. These results suggest that Phyllanthus emblica fruit crude extract MoS2 NPs may have chemoprotective effects by performing CBMN assay.

The influence of vitamin D supplementation and strength training on health biomarkers and chromosomal damage in community-dwelling older adults

Older adults lack of proper physical activity which is often accompanied by vitamin D deficiency. Those factors are known to contribute to health issues in the later years of life. The main goal of this intervention study was to investigate the effect of different vitamin D supplementation strategies for 4 weeks solely or combined with a 10-week strength training program on chromosomal stability in peripheral blood mononuclear cells in community-dwelling older people. One hundred women and men (65–85 years) received either vitamin D3 daily (800 IU), a monthly dose (50.000 IU) or placebo for 17 weeks. All groups received 400 mg calcium daily. The fitness status of the study participants was measured using the 30- second chair stand test, the handgrip strength test and the 6-min walk test. The cytokinesis block micronucleus cytome (CBMN) assay was applied to analyze chromosomal anomalies, including cytotoxic and genotoxic parameters. Changes in antioxidant markers were measured in plasma.Walking distance and chair stand performance improved significantly. Increased levels of the parameters of the CBMN assay were detected for all intervention groups at study end. At baseline micronuclei (MNi) frequency correlated significantly with BMI in both sexes (females: r = 0.369, p = 0.034; males: r = 0.265, p = 0.035), but not with vitamin D serum levels. In females, body fat (r = 0.372, p < 0.001) and functional parameter using the 30-s chair stand test (r = 0.311, p = 0.002) correlated significantly with MNi frequency. Interestingly, not vitamin D supplementation but 10 weeks of resistance training increased MNi frequency indicating elevated chromosomal instability and also adverse effects on antioxidant markers including glutathione and FRAP were detected in the group of community-dwelling older adults.

Biocompatibility analysis of halloysite clay nanotubes

The biocompatibility of four different halloysite clay nanotubes (HNT-N,-U,-P,-S), from various deposits in the world, was evaluated. More precisely, we tested their cytogenetic effects through MTT assay (for cytotoxic effects), and CBMN assay (for genotoxic effects) on mammalian cell cultures (CHO, HeLa, and HepG2 cell lines). Our results indicated a generally high level of biocompatibility of these new-generation products, however at high concentrations, and long times of exposition, we observed a moderate level of toxicity. Thus, we suggest caution in the future use of great amounts of HNTs due to their effects on living organisms.

Toxicogenetic assessment of a pre-workout supplement: In vitro mutagenicity, cytotoxicity, genotoxicity and glutathione determination in liver cell lines and in silico ADMET approaches.

The benefits of practicing physical activity, such as weight loss and control, are commonly associated with caloric restriction diets and may be improved by the ingestion of thermogenic and ergogenic supplements. However, there is a lack of safety data on commonly marketed nutritional supplements. Therefore, this investigation aims to evaluate a pre-workout supplement for mutagenicity using the Ames test, hepatocytoxicity in HepG2 and F C3H cells after 24h, 48h and 72h, genotoxicity using the CBMN assay, determination of gluthatione activity and computational prediction of the three major isolated compounds present in the supplement. The mutagenicity test showed a mutagenic response in TA98 His+ revertants of 5mg/plate in the presence of metabolic activation, cytotoxicity in TA98 of 5mg/plate in the absence of metabolic conditions, and in TA102 of 0.5mg/plate both in the presence and absence of metabolic activation. In our in vitro eukaryotic cell viability, WST-1, LDH and alkaline phosphatase assays, the supplement showed hepatocytotoxicity both dose-dependently and time-dependently. In the cytokinesis blocking micronuclei assay, the supplement induced micronuclei, nuclear buds, nucleoplasmatic, bridge formation, and a decreased in nuclear division. In addition, the supplement decreased intra and extracellular GSH. Computational analysis showed that the three isolated compounds most present in the supplement have the potential to cause hepatotoxicity. In the present investigation, the pre-workout supplement induced mutagenic, genotoxic, and cytotoxic responses and GSH decrease. Thus, considering food safety and public health sanitary vigilance, the consumption of this pre-workout supplement may harm the health of its consumers.

Excessive Folic Acid Mimics Folate Deficiency in Human Lymphocytes.

Food fortification with synthetic folic acid (FA), along with supplementation, results in a marked increase in the population total of serum folates and unmetabolized folic acid (UMFA). Despite the success in reducing neural tube defects at birth in the intended target population (women of childbearing age), the potential deleterious effects of chronically high levels of UMFA in susceptible segments of the population require further investigation. In this study, we examine the effects of FA concentrations, ranging from depletion to supraphysiological levels, on markers of proliferation, DNA methylation, and DNA damage and repair in a human lymphoblastoid cell line (LCL). We note that both low and high levels of FA similarly impact global DNA methylation, cytome biomarkers measured through the CBMN assay, DNA damage induced by oxidative stress, and DNA base excision repair gene expression.

Cytotoxicity and Genotoxicity of Sunset Yellow and Potassium Sorbate in Jurkat Cell Line

Aims: Industrial processed food, pharmaceutical and cosmetical products contain numerous substances the safety of which has been widely concerned. Potassium sorbate (PS) is a common preservative used in a wide-range of products. Sunset yellow (SY) is one of the most commonly used dye in food and pharmaceutical industry. Several studies suggested cytotoxic and genotoxic potential of SY and PS in different cell lines. These effects can lead to organ damage and cancer development. The aim of this study is to investigate cytotoxic and genotoxic potential of two widely used food additives, SY and PS, in Jurkat cell line.&#x0D; Methodology: PrestoBlue assay was used to assess the cytotoxic potential of SY and PS. For observation of DNA damage, cytostatic and cytotoxic effects, the cytokinesis-block micronucleus cytome (CBMN) assay was performed.&#x0D; Results: Decrease of cell viability in Jurkat cell line was observed after 24-hour exposure to both SY and PS. CBMN assay has revealed significant increase of necrotic cells (P&lt;0.05). Genotoxic biomarkers were in physiological range after 24-hour exposure to both analyzed additives.&#x0D; Conclusion: Our findings suggest that SY, as well as PS have cytotoxic potential in Jurkat cell line, as a result of increased number of necrotic cells. Higher cytotoxic effect was caused by SY compared to PS. However, genotoxic potential was not recorded for any of the food additives analyzed.

In vitro and in silico assessment of cytotoxicity and chromosome instability induced by saxitoxin in human derived neural cell line

In freshwater, saxitoxins (STX) are produced by different cyanobacteria genera, including Raphidiopsis. Data regarding cytogenotoxicity effects of STX on human cells are scarse, this merit further studies of its toxicology. This study assessed the cytotoxicity and the chromosome instability of STX on SHSY-5Y human cell line. The CBMN assay allows the detection of chromosome breaks and abnormal chromosomal segregation. Additionally, in silico systems biology approach, used to search for known and predicted interaction networks, was applied to study the interactions between STX and SHSY-5Y cellular components. The results of the CBMN assay demonstrated that STX concentrations of 2.5 - 10 µg/L induced cytostasis and chromosome instability in a dose-response relationship. Apoptosis was detected after exposure of SHSY-5Y cultured cells to STX concentration of 10 µg/L. The results of the systems biology analysis revealed the interaction of STX with proteins related with acetylcoline pathway, cell cycle regulation and apoptosis. Furthermore, combining the in vitro and in silico approachs, it was possible to suggest a mechanism of action of STX in SHSY-5Y cells. Overall, the data demonstrated the cytotoxicity and mutagenicity of environmentally relevant concentrations of STX. These results should be considered when setting up guidelines for monitoring STX in water supply.

Novel Prognostic Biomarkers in Metastatic and Locally Advanced Colorectal Cancer: Micronuclei Frequency and Telomerase Activity in Peripheral Blood Lymphocytes.

PurposeDue to the current practice on colorectal cancer (CRC) management, chemoresistance is most often recognized at the end of the treatment. Therefore, effective and easy-to-use prognostic biomarkers are needed.Experimental DesignWe evaluated the prognostic significance of two novel CRC biomarkers: a) micronuclei frequency (MNf) in 55 metastatic CRC (mCRC) and 21 locally advanced rectal cancer (laRC) patients using cytokinesis block micronucleus assay (CBMN assay) and b) telomerase activity (TA) in 23 mCRC and five laRC patients using TRAP-ELISA. Both biomarkers were evaluated in peripheral blood lymphocytes (PBLs) before, at the middle, and at the end of the therapy (approximately 0, 3, and 6 months) for mCRC patients before, at the end of the therapy, and after surgery for laRC patients.ResultsOverall, MNf demonstrated significant prognostic value since a decrease of MNf less than 29% between middle and initial MNf measurements can discriminate between progressive and stable/responsive disease with sensitivity of 36% and specificity of 87.0% while being able to identify responsive disease with sensitivity of 72.7% and specificity of 59.3%. On the other hand, TA presented a significant trend of increase (p = 0.07) in patients with progressive disease at the middle measurement.ConclusionsThe findings of this study suggest that the MN frequency may serve as a promising prognostic biomarker for the monitoring of the treatment response of patients with CRC, while TA should be evaluated in a larger group of patients to further validate its significance.

Gasoline-station workers in Brazil: Benzene exposure; Genotoxic and immunotoxic effects

Chronic exposure to benzene is a risk factor for hematological malignancies. Gasoline-station workers are exposed to benzene in gasoline, via both inhalation and dermal contact (attendants and managers) or inhalation (workers in the on-site convenience stores and offices). We have studied the exposure of these workers to benzene and the resulting genotoxic and immunotoxic effects. Levels of urinary trans, trans-muconic acid were higher among gasoline-station workers than among office workers with no known exposure to benzene (comparison group). Among the exposed workers, we observed statistically significant biological effects, including elevated DNA damage (comet assay); higher frequencies of micronuclei and nuclear buds (CBMN assay); lower levels of T-helper lymphocytes and naive Th lymphocytes; lower CD4 / CD8 ratio; and higher levels of NK cells and memory Th lymphocytes. Both groups of exposed workers (inhalation and inhalation + dermal routes) showed similar genotoxic and immunotoxic effects.

Comparative Evaluation of Genotoxicity in Tobacco Users versus Nontobacco Users.

Background:Many of the contents of cigarette smoke are genotoxic in nature, and consequently, cytogenetic injury seems to be a trustworthy biomarker for deciding the influence of exposure to chromosome damaging agents in smoke. The cytokinesis-block micronucleus assay (CBMN assay) has been proven to be an effectual tool for the study of micronuclei (MN) that will help in estimating the genotoxicity in tobacco users alone which will further help in early cancer detection.Objective:The objective is to find out whether there is pronounced contrast in genotoxicity between tobacco users and nonusers by determining MN number in peripheral blood lymphocytes using CBMN assay.Methodology:MN frequency in peripheral blood lymphocytes was estimated in 5 ml of fresh blood obtained from sixty individuals using tobacco either smoking, chewing, or combination of both and also from thirty individuals with no habit of tobacco use. All were in the age group of 20–40 years.Results:There was a significant increase in genotoxicity in tobacco users when compared to that of nontobacco users. A positive correlation was also obtained between smoking index and MN frequency in the study.Conclusion:Approximation of frequency of MN by CBMN assay can be used to evaluate the genotoxicity present in blood and helps in identifying tobacco users who are at a high risk for the presence of cancer even before the appearance of clinical changes.

A STUDY ON SOMATIC DNA DAMAGES IN HYPERTENSIVE YOUNG ADULTS AND ITS CORRELATION WITH BIOCHEMICALAND DEMOGRAPHIC PARAME

Introduction: Incidence of hypertension is increasing in young population. Aggressive control is mandatory to preserve and protect public health in India. Several socio-demographic, anthropometric, environmental, lifestyle, biochemical and genetic factors contribute for the development of hypertension in young adults. Hypertension and its chronic complications are also linked to oxidative stress and DNA damage. Methods: The cross sectional study involves 180 young hypertensives between the age group of 18 – 39 years and 140 apparently healthy age and sex matched controls. After obtaining relevant history, anthropometric measurements were taken. Socio-demographic characters were recorded using proforma. After taking written informed consent, five ( 5 ml) of venous blood was collected after 8-12 hours of fasting. 2 ml of blood was transferred aseptically to a sodium heparinised vacutainer for evaluating somatic DNA damages by Cytokinesis-block Micronuclei assay (CBMN assay). Remaining 3 ml of blood is allowed to clot; serum separated and was used for other biochemical investigations such as fasting blood sugar, lipid profile, urea, creatinine and malondialdehyde (MDA). Statistical analysis done using SPSS software. Results: Fasting blood sugar, Total cholesterol, LDL cholesterol, triglycerides, urea and creatinine were significantly elevated in young hypertensives. Oxidative stress and DNA damage is significantly elevated in young hypertensives compared to control group. BMI and abdominal circumference are also significantly elevated in young hypertensive adults compared to control group. Conclusion: Increased oxidative stress and DNA damage in young hypertensives contribute to the complications associated with hypertension.

Cytogenetically-based biodosimetry after high doses of radiation.

Dosimetry is an important tool for triage and treatment planning following any radiation exposure accident, and biological dosimetry, which estimates exposure dose using a biological parameter, is a practical means of determining the specific dose an individual receives. The cytokinesis-blocked micronucleus assay (CBMN) is an established biodosimetric tool to measure chromosomal damage in mitogen-stimulated human lymphocytes. The CBMN method is especially valuable for biodosimetry in triage situations thanks to simplicity in scoring and adaptability to high-throughput automated sample processing systems. While this technique produces dose-response data which fit very well to a linear-quadratic model for exposures to low linear energy transfer (LET) radiation and for doses up for 5 Gy, limitations to the accuracy of this method arise at larger doses. Accuracy at higher doses is limited by the number of cells reaching mitosis. Whereas it would be expected that the yield of micronuclei increases with the dose, in many experiments it has been shown to actually decrease when normalized over the total number of cells. This variation from a monotonically increasing dose response poses a limitation for retrospective dose reconstruction. In this study we modified the standard CBMN assay to increase its accuracy following exposures to higher doses of photons or a mixed neutron–photon beam. The assay is modified either through inhibitions of the G2/M and spindle checkpoints with the addition of caffeine and/or ZM447439 (an Aurora kinase inhibitor), respectively to the blood cultures at select times during the assay. Our results showed that caffeine addition improved assay performance for photon up to 10 Gy. This was achieved by extending the assay time from the typical 70 h to just 74 h. Compared to micronuclei yields without inhibitors, addition of caffeine and ZM447439 resulted in improved accuracy in the detection of micronuclei yields up to 10 Gy from photons and 4 Gy of mixed neutrons-photons. When the dose-effect curves were fitted to take into account the turnover phenomenon observed at higher doses, best fitting was achieved when the combination of both inhibitors was used. These techniques permit reliable dose reconstruction after high doses of radiation with a method that can be adapted to high-throughput automated sample processing systems.