Increase In Sister Chromatid Exchanges Research Articles

Haloperidol's Cytogenetic Effect on T Lymphocytes of Systemic Lupus Erythematosus and Rheumatoid Arthritis Patients: An In Vitro Study.

Investigating haloperidol's cytogenetic behavior in cultured human T lymphocytes of patients with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). Four haloperidol solutions were added in cultures of peripheral blood lymphocytes of healthy individuals, SLE, and RA patients. After 72 hours of incubation, the cultured lymphocytes were plated on glass slides, and stained with the fluorescence plus Giemsa method, and sister chromatid exchanges (SCEs), proliferation rate index (PRI), and mitotic index (MI) were measured with the optical microscope. Result analysis revealed: (a) a statistically significant (p=0.001) dose-dependent increase of SCEs in SLE patients compared to healthy individuals; (b) a statistically significant (p=0.001) dose-dependent decrease of SCEs in RA patients for haloperidol concentrations 5, 10μg/mL; (c) a statistically significant (p=0.001) dose-dependent increase of SCEs in RA patients for haloperidol concentrations 20, 100μg/mL; and (d) a statistically significant (p=0.001) dose-dependent reduction of PRI and MI in both patient groups compared to healthy individuals. Furthermore, a correlation was observed between (a) SCE and PRI index variations, (b) MI and SCE index variations, and (c) PRI and MI index variations. Haloperidol affects T lymphocytes from SLE and RA patients by modifying DNA replication procedures, DNA damage response, and ferroptosis. Considering the wide use of haloperidol in neuropsychiatric symptoms of SLE and RA patients, further studies with more immune cell subsets are needed to evaluate its effects on human genetic material.

Olanzapine’s Cytogenetic Effect on T Lymphocytes in Systemic Lupus Erythematosus and Rheumatoid Arthritis Patients: In Vitro Study

This study will investigate olanzapine's cytogenetic behavior in cultured human T lymphocytes in patients with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). Three olanzapine solutions were added in cultures of peripheral blood lymphocytes of healthy individuals, SLE, and RA patients. After 72 hours of incubation, the cultured lymphocytes were plated on glass slides and stained with the fluorescence plus Giemsa method. Sister chromatid exchanges (SCEs), proliferation rate index (PRI), and mitotic index (MI) were measured with the optical microscope. There was a statistically significant (p=0.001) dose-dependent increase of SCEs in SLE and RA patients compared to healthy individuals and a statistically significant (p=0.001) reduction of PRI and MI in the highest concentration in the SLE group. Moreover, Spearman's rank correlation coefficient was applied to calculate the correlation between SCEs, PRI, and MI. Negative significant correlations were noticed for both patient groups concerning SCEs-PRI alterations and SCEs-MI alterations. Conversely, positive correlations were noticed for both patient groups for PRI-MI alterations. Conclusions: Olanzapine affects T lymphocytes from SLE and RA patients by modifying DNA replication procedures and DNA damage response. Considering the use of olanzapine in neuropsychiatric symptoms of SLE, further in vivo studies are necessary to evaluate its effect on human DNA.

Bloom Syndrome Patients Display Accelerated Epigenetic Aging in Blood Lymphocytes

Bloom Syndrome (BSyn) is an autosomal recessive disorder caused by variants in the BLM genes at locus 15q26.1. The BLM gene encodes a protein that is part of the DNA RECQ helicase family, which has a role in homologous recombination, DNA structural integrity, DNA repair responses, and telomere maintenance. Patients with BSyn present with poor growth, sun sensitivity, mild immunodeficiency, diabetes, and increased risk of cancer, most commonly leukemias. Additionally, patients with BSyn develop myelodysplasia and life-threatening side effects from standard chemotherapy and radiation, often due to bone marrow toxicity. Cells from patients with BSyn display increased sister chromatid exchange, telomere defects, and telomere-dependent chromosome fusions. Despite these aging-associated findings, patients with BSyn do not appear to have typical premature aging characteristics such as graying hair, osteoporosis, and early-onset cardiovascular disease. We hypothesized that patients with BSyn may have increased epigenetic aging in blood, which may be related to their increased risk of hematologic dysfunction and malignancies. In this study, we aimed to use DNA methylation in order to identify an aging signature that could be a potential biomarker of disease risk. In partnership with the Bloom Syndrome Registry (BSR) as well as JScreen and our own clinical practice, we extracted 49 DNA samples from blood lymphocytes of patients with BSyn as well as carriers. The samples were then processed per standard Illumina protocol for the Infinium MethylationEPIC BeadChip kit. We utilized the online DNA Methylation Age Calculator (https://dnamage.genetics.ucla.edu/) to calculate multiple measures of methylation aging for each patient, including the Horvath pan-tissue epigenetic clock DNAmAge, the Hannum clock, the GrimAge lifespan predictor, and the PhenoAge phenotypic age estimate. Simple linear regression was used to determine statistical significance. Of the 49 total samples, 18 samples were from patients with BSyn (ages 1-38 years) and 31 samples were from carriers with one variant in BLM (ages 23-69 years). We discovered that BSyn patients had increased DNAm age compared to carriers measured by the pan-tissue clock (p=0.0054). Similarly, epigenetic clocks that were designed to predict human mortality risk (GrimAge and PhenoAge) indicate significant age acceleration as well, p<0.0088 and p<0.0001 respectively. Finally, DNA methylation-based estimators of telomere length show significantly shorter telomere lengths in BSyn patients compared to carriers, p<0.0001. Epigenetic age can be a better predictor of health and disease over biologic or chronologic age. In this study, we show for the first time that patients with BSyn have evidence of epigenetic aging across several measures in blood lymphocytes. Future studies are ongoing to investigate underlying mechanisms for this and whether epigenetic aging in these patients can predict hematologic dysfunction and cancer risk.

Alternative end-joining originates stable chromosome aberrations induced by etoposide during targeted inhibition of DNA-PKcs in ATM-deficient tumor cells.

ATM and DNA-PKcs coordinate the DNA damage response at multiple levels following the exposure to chemotherapy. The Topoisomerase II poison etoposide (ETO) is an effective chemotherapeutic agent that induces DNA double-strand breaks (DSB), but it is responsible from the chromosomal rearrangements frequently found in therapy-related secondary tumors. Targeted inhibition of DNA-PKcs in ATM-defective tumors combined with radio- or chemotherapy has been proposed as relevant therapies. Here, we explored the DNA repair mechanisms and the genetic consequences of targeting the non-oncogenic addiction to DNA-PKcs of ATM-defective tumor cells after exposure to ETO. We demonstrated that chemical inhibition of DNA-PKcs followed by treatment with ETO resulted in the accumulation of chromatid breaks and decreased mitotic index in both A-T cells and ATM-knocked-down (ATMkd) tumor cells. The HR repair process in DNA-PKcs-inhibited ATMkd cells amplified the RAD51 foci number, with no correlated increase in sister chromatid exchanges. The analysis of post-mitotic DNA lesions presented an augmented number of persistent unresolved DSB, without alterations in the cell cycle progression. Long-term examination of chromosome aberrations revealed a strikingly high number of chromatid and chromosome exchanges. By using genetic and pharmacological abrogation of PARP-1, we demonstrated that alternative end-joining (alt-EJ) repair pathway is responsible for those chromosome abnormalities generated by limiting c-NHEJ activities during directed inhibition of DNA-PKcs in ATM-deficient cells. Targeting the non-oncogenic addiction to DNA-PKcs of ATM-defective tumors stimulates the DSB repair by alt-EJ, which is liable for the origin of cells carrying stable chromosome aberrations that may eventually restrict the therapeutic strategy.

Assessment of the genotoxic potential of tetrachlorvinphos insecticide by cytokinesis-block micronucleus and sister chromatid exchange assays.

Tetrachlorvinphos is an organophosphate that is classified as a carcinogen in humans by several authorities. Due to very limited data regarding the genotoxic potential, we aimed to comprehensively investigate in vitro genotoxic potential of tetrachlorvinphos. We performed our study by applying the cytokinesis-block micronucleus cytome and sister chromatid exchange (SCE) assays to human peripheral blood lymphocytes. We evaluated micronucleus (MN) and SCE frequencies and cytokinesis-block proliferation index in both exposed and non-exposed lymphocytes. We also calculated the chromosomal instability level in response to exposure by combining the results of MN and SCE. We found that MN frequency did not increase with exposure to tetrachlorvinphos (0-50µg/ml). In contrast, we observed that SCE frequencies significantly increased with exposure to ≥5µg/ml tetrachlorvinphos. Furthermore, exposure to tetrachlorvinphos at concentrations of 50µg/ml induced a significant increase in chromosomal instability level (p < 0.05). Cytokinesis-block proliferation index level did not significantly decrease in response to tetrachlorvinphos exposure. Our findings reveal that tetrachlorvinphos resulted in different DNA damages that were measured by two assays. Furthermore, our findings suggested that exposure to tetrachlorvinphos increased chromosomal instability that is a hallmark of many malignancies. We conclude that although tetrachlorvinphos does not significantly increase the MN level, the significant increase of both SCE and CIN frequencies indicates the genotoxic potential of tetrachlorvinphos in human peripheral lymphocytes. Additionally, tetrachlorvinphos is not cytotoxic in the range of tested concentrations.

Predominant cellular mitochondrial dysfunction in the TOP3A gene-caused Bloom syndrome-like disorder

TOP3A promotes processing of double Holliday junction dissolution and also plays an important role in decatenation and segregation of human mtDNA. Recently, TOP3A mutations have been reported to cause Bloom syndrome-like disorder. However, whether the two function play equal roles in the disease pathogenesis is unclear. We retrospectively studied the disease progression of two siblings with Bloom-like syndrome caused by two novel mutations of TOP3A, p.Q788* and p.D479G. Beside the common clinical manifestations, our patients exhibited liver lipid storage with hepatomegaly. In cellular and molecular biological studies, TOP3A deficiency moderately increased sister chromatid exchanges and decreased cell proliferation compared with BLM or RMI2 deficiency. These changes were rescued by ectopic expression of either of the wildtype TOP3A or TOP3A-D479G. In contrast, reduced mitochondrial ATP generation and oxygen consumption rates observed in TOP3A defective cells were rescued by over-expression of the wildtype TOP3A, but not TOP3A-D479G. Considering the different impact of the TOP3A-D479G mutation on the genome stability and mitochondrial metabolism, we propose that the impaired mitochondrial metabolism plays an important role in the pathogenesis of TOP3A-deficient Bloom-like disease.

Germline BLM mutations and metastatic prostate cancer.

Biallelic loss-of-function BLM mutations result in Bloom syndrome: a genetic disorder characterized by growth deficiencies, photosensitivity, and multiple cancer susceptibilities. There are conflicting reports about whether or not heterozygous BLM carriers are at a higher risk of various cancers. Without BLM protein functionality, there is evidence of increased sister chromatid exchange and chromosomal instability. Metastatic prostate cancer patients (N = 796) underwent germline genetic testing as part of routine care at three academic centers. Patients with heterozygous BLM mutations were identified. Tumor tissue was analyzed for somatic alterations in those patients who had a germline pathogenic mutation. Control data using a population sample were extracted from the Genome Aggregation Database. Heterozygous BLM germline mutations in 5 of 796 patients (prevalence, 0.63%). All mutations were loss-of-function truncating alterations. None of the mutations were BLMAsh . The control population (gnomAD) frequency of pathogenic or likely pathogenic BLM mutations was 0.18% (212 of 116 653). The relative risk (RR) of BLM mutations in metastatic prostate cancer patients was 3.4 (95% CI, 1.42-8.33; P < .0062) compared to gnomAD controls. Tumor DNA sequencing in the BLM carriers showed no evidence of somatic BLM mutations. Interestingly, 3 of 5 BLM germline carriers had bi-allelic BRCA2 inactivation evident on tumor sequencing. One patient had both germline and somatic mutations in BRCA2. Excluding the patient with the germline BRCA2 mutation (BLM prevalence, 4 of 796: 0.50%) still yielded a statistically significant finding vs the gnomAD controls (RR, 2.8; 95% CI, 1.02-7.39; P < .04). Truncating BLM germline mutations occur at a higher frequency in patients with advanced prostate cancer as compared to control populations. Though no biallelic loss of BLM was no noted in cancers, a surprising number of the BLM germline heterozygotes had pathogenic BRCA2 mutations in their tumor.

Genotoxicity of cisplatin and carboplatin in cultured human lymphocytes: a comparative study.

Cisplatin and carboplatin are integral parts of many antineoplastic management regimens. Both platinum analogues are potent DNA alkylating agents that robustly induce genomic instability and promote apoptosis in tumor cells. Although the mechanism of action of both drugs is similar, cisplatin appears to be more cytotoxic. In this study, the genotoxic potential of cisplatin and carboplatin was compared using chromosomal aberrations (CAs) and sister-chromatid exchange (SCE) assays in cultured human lymphocytes. Results showed that cisplatin and carboplatin induced a significant increase in CAs and SCEs compared to the control group (p<0.01). Levels of induced CAs were similar in both drugs; however, the magnitude of SCEs induced by cisplatin was significantly higher than that induced by carboplatin (p<0.01). With respect to the mitotic and proliferative indices, both cisplatin and carboplatin significantly decreased mitotic index (p<0.01) without affecting the proliferative index (p>0.05). In conclusion, cisplatin was found to be more genotoxic than carboplatin in the SCE assay in cultured human lymphocytes, and that might explain the higher cytotoxicity of cisplatin.

Etoposide. Documentation of proposed values of occupational exposure limits (OELs)

Etoposide. Documentation of proposed values of occupational exposure limits (OELs)

Assessment of Genotoxicity of Levosimendan in Human Cultured Lymphocytes.

Levosimendan is a positive inotropic and a vasodilator agent with pleotropic characteristics that include antioxidation, anti-inflammation and smooth muscle vasodilation. In this study, the effects of levosimendan (0, 0.1, 1, 10, and 20 µg/ml) on oxidative DNA damage and sister-chromatid exchanges (SCEs) were evaluated in human cultured lymphocytes. The results showed that levosimendan increased the frequency of SCEs in all examined concentrations (P<0.01) except for 0.1 µg/ml. On the other hand, levosimendan did not induce oxidative DNA damage as measured by the 8-OHdG biomarker (P > 0.05). In addition, neither mitotic arrest nor proliferation index was affected by levosimendan at all examined doses (P > 0.05). In conclusion, levosimendan might be associated with increases in sister-chromatid exchanges in cultured human lymphocytes. In vivo studies are required to confirm the present findings.

Genotoxic effects of Peganum harmala L. in relation to traditional use

Peganum harmala L. is a well-known medicinal plant, widely distributed and used in folk medicine. Recently, numerous reports have been published regarding the pharmacological and therapeutic effects of P. harmala and its major alkaloids components. However, mammalian toxicity due to P. harmala alkaloids extract has been rarely investigated. The present research aimed to study the genotoxic effects of harmala plant extract in relation to traditional use. Harmala crude alkaloid was extracted and the toxicity of the extract was examined on albino mice by intraperitoneal injection. The genotoxic effects of the alkaloid extract was tested on bone marrow cells isolated from sacrificed mice. The total seeds alkaloids showed moderate toxic effects on male albino mice with LD50 350 mg/ Kg body weight. P. harmala alkaloids extract showed significant reduction in the mitotic index (MI) and increased depression of mammalian cells division possibly through chromosomal aberrations. P. harmala alkaloids induced different types of chromosome aberrations including rings, breaks, polyploidy, sticky, laggards and bridges with the sticky form as the most abundant type. Furthermore, P. harmala alkaloids induced a significant increase in sister chromatid exchange (SCE) compared to untreated controls. The frequency of micronuclei was increased with increasing the concentration but was not affected by increasing the exposure time. The medicinal use of harmala should be under control since higher doses and/ or longer exposure is genotoxic. An amount of plant that contains ≥ 12 mg alkaloids cannot be safe for traditional use. Key words: Peganum harmala, genotoxicity, chromosome aberration, sister chromatid exchange.

Mutations in TOP3A Cause a Bloom Syndrome-like Disorder

Bloom syndrome, caused by biallelic mutations in BLM, is characterized by prenatal-onset growth deficiency, short stature, an erythematous photosensitive malar rash, and increased cancer predisposition. Diagnostically, a hallmark feature is the presence of increased sister chromatid exchanges (SCEs) on cytogenetic testing. Here, we describe biallelic mutations in TOP3A in ten individuals with prenatal-onset growth restriction and microcephaly. TOP3A encodes topoisomerase III alpha (TopIIIα), which binds to BLM as part of the BTRR complex, and promotes dissolution of double Holliday junctions arising during homologous recombination. We also identify a homozygous truncating variant in RMI1, which encodes another component of the BTRR complex, in two individuals with microcephalic dwarfism. The TOP3A mutations substantially reduce cellular levels of TopIIIα, and consequently subjects’ cells demonstrate elevated rates of SCE. Unresolved DNA recombination and/or replication intermediates persist into mitosis, leading to chromosome segregation defects and genome instability that most likely explain the growth restriction seen in these subjects and in Bloom syndrome. Clinical features of mitochondrial dysfunction are evident in several individuals with biallelic TOP3A mutations, consistent with the recently reported additional function of TopIIIα in mitochondrial DNA decatenation. In summary, our findings establish TOP3A mutations as an additional cause of prenatal-onset short stature with increased cytogenetic SCEs and implicate the decatenation activity of the BTRR complex in their pathogenesis.

Laser Irradiation of Organic Tattoo Pigments Releases Carcinogens with 3,3′-Dichlorobenzidine Inducing DNA Strand Breaks in Human Skin Cells

Laser Irradiation of Organic Tattoo Pigments Releases Carcinogens with 3,3′-Dichlorobenzidine Inducing DNA Strand Breaks in Human Skin Cells

In vitro genotoxic and cytotoxic effects of doxepin and escitalopram on human peripheral lymphocytes

Antidepressants are drugs used for the treatment of many psychiatric conditions including depression. There are findings suggesting that these drugs might have genotoxic, carcinogenic, and/or mutagenic effects. Therefore, the present in vitro study is intended to investigate potential genotoxic and cytotoxic effects of the antidepressants escitalopram (selective serotonin reuptake inhibitor) and doxepin (Tricyclic antidepressant) on human peripheral lymphocytes cytokinesis-block micronucleus (CBMN), sister chromatid exchange (SCE), and single cell gel electrophoresis (alkaline comet assay) were used for the purpose of the study. In the study, four different concentrations of both drugs (1, 2.5, 5, and 10 µg/mL) were administered to human peripheral lymphocytes for 24 h. The tested concentrations of both drugs were found to exhibit no cytotoxic and mitotic inhibitory effects. SCE increase caused by 5 and 10 µg/mL of escitalopram was found statistically significant, while no statistically significant increase was observed in DNA damage and micronucleus (MN) formation. Moreover, the increase caused by doxepin in MN formation was not found statistically significant. Besides, 10 µg/mL of doxepin was demonstrated to significantly increase arbitrary unit and SCE formation. These findings suggest that the investigated concentrations of escitalopram and doxepin were non-cytotoxic but potentially genotoxic at higher concentrations.

Genotoxic Evaluation of Duloxetine II. The Effect on the Number of Sister Chromatid Exchanges, the Mitotic Index, and the Proliferation Kinetics in Mouse Bone Marrow.

Duloxetine is an antidepressant which has showed valuable results, particularly in patients with major depression. This type of drugs is known to require genotoxic studies as part of their preclinical safety evaluation. In the case of duloxetine, however, there have been controversial results. Therefore, we considered it worthwhile to extend studies on the matter in an attempt to reach a conclusion. The present assay was made in mouse bone marrow to evaluate the capacity of the drug to induce sister chromatid exchanges (SCE), as well as to modify the proliferation kinetics and the mitotic index. Three doses of the antidepressant were tested (2, 20, and 200 mg/kg), besides the control mice were administered with purified water, and the positive treated animals administered with 1 mg/kg of doxorubicin. The results indicated a moderate but significant increase of SCE with the three tested doses, no effect regarding the mitotic index and a small reduction in the proliferation kinetics. Although in our assay the drug showed a lower effect, the present study agreed with a previous report that analyzed the amount of micronuclei in mouse peripheral blood, and it confirmed the relevance of evaluating the genotoxic effect of antidepressants, specifically duloxetine by applying diverse tests.

Pml Nuclear Body Disruption Cooperates in APL Pathogenesis, Impacting DNA Damage Repair Pathways

Acute promyelocytic leukemia (APL) is driven by the oncogene PML-RARA which is generated by fusion of the promyelocytic leukemia (PML) and retinoic acid receptor alpha (RARA) genes, and which strongly interferes with downstream signalling and the architecture of multiprotein structures known as PML nuclear bodies (NBs). NB disruption is a diagnostic hallmark of APL, yet the significance of this phenomenon to disease pathogenesis and treatment response remains poorly understood. The majority of APL patients can now be cured with combination therapy with arsenic trioxide (ATO) and ATRA (All Trans-Retinoic Acid), which synergize promoting re-formation of disrupted Pml NBs. To date, the importance of NB disruption has only been studied in vitro. To address this, we generated a knock-in mouse model with targeted NB disruption achieved through mutation of key zinc-binding cysteine residues (C62A/C65A) in the RING domain of Pml. Homozygous PmlC62A/C65A mice are viable, and developmentally normal. At a cellular level, Pml NB disruption was confirmed and treatment with ATO was associated with defective Pml SUMOylation and degradation. A key feature of APL fusion proteins is the capacity to homodimerise (mediated by the fusion partner e.g. PML), which is not a feature of wild-type RARα. This forced homodimerisation of RARα has been shown to be critical for APL pathogenesis. We investigated whether Pml NB disruption could cooperate in vivo with forced RARα homodimerisation (mediated artificially by linking RARα to the dimerisation domain of the NFκB p50 subunit). While no leukemias arose in PmlC62A/C65A mice, p50-RARα mice expressing PmlC62A/C65A presented a doubling in the rate of leukemia development (p<0.0001) compared to PmlWT-p50-RARα, leading to a penetrance comparable to that observed in previously published PML-RARα transgenic models. Moreover, the latency period to onset of leukemia was significantly reduced in the context of NB disruption (p=0.008). ATRA treatment significantly improved the survival of mice transplanted with PmlWT-p50-RARα or Pml-RARα leukemic blasts, but not with PmlC62A/C65A-p50-RARα. These data reveal not only the key role of PML-RARα expression-induced NB disruption in APL development, but also the importance of re-formation of NBs for an effective response to differentiating drug.While formation of the PML-RARA fusion is considered an initiating event in APL pathogenesis, it is insufficient for the full leukemic phenotype. Exome sequencing studies have consistently identified presence of cooperating mutations. Since Pml and Pml NB have established roles in DNA repair and in the maintenance of genomic stability, we speculated that loss of NB integrity could affect these functions. Whole exome sequencing revealed a pattern of higher genomic instability in PmlC62A/C65A-p50-RARα leukemia as compared to PmlWT-p50-RARα, with detection of mutations found in human APL, including Kras, Ptpn11 and Usp9y. Using DNA repair reporter assays, we demonstrated that DNA repair via both non-homologous end joining (NHEJ; p=0.01) and homologous recombination (HR; p=0.006) pathways was less efficient in PmlC62A/C65A primary cells than in PmlWT cells. Importantly, using a PML-RARα-inducible cell line, comparable defects in the NHEJ and HR pathways, which were PML-RARα dependent, were identified. These data were also supported by an increase in sister-chromatid exchange (p<0.0001) and chromosome abnormality (p=0.0002) in the context of PmlC62A/C65A versus PmlWT. Interestingly, the kinetic of repair of ionising radiation (IR)-induced DNA double-strand breaks, assessed by analysis of γH2AX foci formation and clearance, was not affected. None of the DNA repair players analysed (e.g. Blm, Rad51 and 53BP1) failed to form foci in response to IR. However, their basal levels of foci were significantly greater in the presence of PmlC62A/C65A (p<0.04; quantified using Amnis ImageStreamX Mk II imaging flow cytometer). Additionally, we found that Rad51 foci showed a defect in localisation post-IR when PmlC62A/C65A was expressed, with impairment of Rad51 co-localisation and interaction with γH2AX. Altogether, our data therefore highlight the significant contribution of Pml NB to the effectiveness of DNA damage repair processes, and the manner in which their disruption mediated by the PML-RARα oncoprotein can assist APL pathogenesis. DisclosuresHills:TEVA: Honoraria. Grimwade:TEVA: Research Funding.

The role of natural indigo dye in alleviation of genotoxicity of sodium dithionite as a reducing agent.

Indigo blue is a natural dye used for thousands of years by civilizations to dye fabric blue and it is naturally obtained from Isatis tinctoria. I. tinctoria is not only used for extraction of indigo blue color but also used medicinally in Traditional Chinese Medicine because of its active compounds. Sodium dithionite (Na2S2O4) is used in dye bath for indigo blue extraction, but this reducing agent and its derivatives are major pollutants of textile industry and subsequently have hazardous influences on public health. Herein, the present study was designed to obtain the high yield of natural indigo dye but with low possible toxic effect. In this context, genotoxic effects of particular combinations of natural dye solutions obtained from Isatis tinctoria subsp. tomentolla with Na2S2O4 as reducing agent were investigated. Dye solutions were obtained using two different pH levels (pH 9 and 11) and three different concentrations of Na2S2O4 (2.5, 5 and 10mg/ml). In addition to the dye solutions and reducing agent, aqueous extracts of I. tinctoria were assessed for their genotoxicity on human lymphocytes. For in vitro testing of genotoxicity, chromosomal aberrations (CAs), sister chromatid exchanges (SCEs) and mitotic indexes (MI) assays were used. Accordingly, Na2S2O4 caused significant increases in CA and SCE as well decrease in MI but the genotoxic effects of sodium dithionite were reduced with natural indigo dye. As a result, aqueous extracts of Isatis leaves removed the toxic effects of sodium dithionite and showed anti-genotoxic effect. For the optimal and desired quality but with less toxic effects of natural dye, 2.5mg/ml (for wool yarn) and 5mg/ml (for cotton yarn) of Na2S2O4 doses were found to be the best doses for reduction in the dye bath at Ph 9.

PARP Inhibitors in Clinical Use Induce Genomic Instability in Normal Human Cells.

Poly(ADP-ribose) polymerases (PARPs) are the first proteins involved in cellular DNA repair pathways to be targeted by specific inhibitors for clinical benefit. Tumors harboring genetic defects in homologous recombination (HR), a DNA double-strand break (DSB) repair pathway, are hypersensitive to PARP inhibitors (PARPi). Early phase clinical trials with PARPi have been promising in patients with advanced BRCA1 or BRCA2-associated breast, ovary and prostate cancer and have led to limited approval for treatment of BRCA-deficient ovary cancer. Unlike HR-defective cells, HR-proficient cells manifest very low cytotoxicity when exposed to PARPi, although they mount a DNA damage response. However, the genotoxic effects on normal human cells when agents including PARPi disturb proficient cellular repair processes have not been substantially investigated. We quantified cytogenetic alterations of human cells, including primary lymphoid cells and non-tumorigenic and tumorigenic epithelial cell lines, exposed to PARPi at clinically relevant doses by both sister chromatid exchange (SCE) assays and chromosome spreading. As expected, both olaparib and veliparib effectively inhibited poly-ADP-ribosylation (PAR), and caused marked hypersensitivity in HR-deficient cells. Significant dose-dependent increases in SCEs were observed in normal and non-tumorigenic cells with minimal residual PAR activity. Clinically relevant doses of the FDA-approved olaparib led to a marked increase of SCEs (5-10-fold) and chromatid aberrations (2-6-fold). Furthermore, olaparib potentiated SCE induction by cisplatin in normal human cells. Our data have important implications for therapies with regard to sustained genotoxicity to normal cells. Genomic instability arising from PARPi warrants consideration, especially if these agents will be used in people with early stage cancers, in prevention strategies or for non-oncologic indications.

Cytotoxicity and genotoxicity of iron oxide nanoparticles: An in vitro biosafety study

With the development of nanotechnology and the wide use of iron oxide nanoparticles, it has become necessary to assess the potential adverse biological effects of magnetite. This study investigated the cytotoxicity, genotoxicity and oxidative damage of different concentrations of magnetite (0 to 1000 mg/L) in human whole blood cultures. After supplementation of magnetite, the blood samples were incubated for 72 h. Cell viability was assessed by the 3-(4,5-dimethyl-thiazol-2-yl) 2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) release assays. The total antioxidant capacity (TAC) and total oxidant status (TOS) were determined to evaluate the dose-dependent effects of magnetite on the oxidant/antioxidant balance and to evaluate the potential oxidative injury due to increased oxidative stress. Genotoxicity was estimated by by the sister chromatid exchange (SCE), micronuclei (MN) and chromosome aberration (CA) assays and determination of 8-oxo-2-deoxyguanosine (8-OH-dG) levels. The results of MTT and LDH assays showed that the higher concentrations of magnetite (100, 150, 300, 500 and 1000 mg/L) decreased cell viability. Concentrations of magnetite higher than 10 mg/L increased TOS levels and decreased TAC levels in human blood cells. Increasing concentrations of magnetite caused significant increases in MN, SCE and CA rates and 8-OH-dG levels. The obtained results showed that magnetite exerted dose-dependent effects on oxidative damage, genotoxicity and cytotoxicity in human blood cells.

GENOTOXICITY OF FORMALDEHYDE IN PERIPHERAL BLOOD LYMPHOCYTES AMONG PATHOLOGY LABORATORY WORKERS

Introduction: Formaldehyde (FA) is a chemical, traditionally used in pathology and anatomy laboratories as a tissue preservative. Many studies clearly indicated that FA can induce genotoxic effects in peripheral blood lymphocytes. Aim of work: to evaluate the genotoxic effects of formaldehyde among medical and paramedical personnel in The Histopathology Laboratory. Materials and Methods: The study was conducted in histopathology laboratory including 30 workers occupationally exposed to formaldehyde and 29 persons from other lab in kasr Al Aini hospital matching the exposed group for age, sex and socioeconomic status with no history of occupational exposure to formaldehyde. A questionnaire was done including inquiries about age, sex, occupational history and special habits. All participants were subjected to the following laboratory investigations: measuring DNA protein crosslink (DPC), sister chromatid exchange (SCE) and micronucleus (Mn) frequency in peripheral blood lymphocytes. Environmental monitoring of the work place was done to measure the level of FA at different place in the pathology lab. Results: This study showed statistically significant increase of DPC with significant increase in sister chromatid exchange (SCE) and micronucleus MN in peripheral blood lymphocytes. Conclusion: The studied group was exposed to formaldehyde level above the exposure limits recommended by NIOSH, ACGIH and OSHA with adverse health effects in the form of increase in DPC, SCE and Mn as genotoxicity biomarkers. These genotoxicity biomarkers are considered to play an important role in the carcinogenesis of FA.