Lysophosphatidic acid type 2 receptor agonists in targeted drug development offer broad therapeutic potential.

The growth factor-like lipid mediator, lysophosphatidic acid (LPA), is a potent signaling molecule that influences numerous physiologic and pathologic processes. Manipulation of LPA signaling is of growing pharmacotherapeutic interest, especially because LPA resembles compounds with drug-like features. The action of LPA is mediated through activation of multiple types of molecular targets, including six G protein-coupled receptors that are clear targets for drug development. However, the LPA signaling has been linked to pathological responses that include promotion of fibrosis, atherogenesis, tumorigenesis, and metastasis. Thus, a question arises: Can we harness, in an LPA-like drug, the many beneficial activities of this lipid without eliciting its dreadful actions? We developed octadecyl thiophosphate (OTP; subsequently licensed as Rx100), an LPA mimic with higher stability in vivo than LPA. This article highlights progress made toward developing analogs like OTP and exploring prosurvival and regenerative LPA signaling. We determined that LPA prevents cell death triggered by various cellular stresses, including genotoxic stressors, and rescues cells condemned to apoptosis. LPA2 agonists provide a new treatment option for secretory diarrhea and reduce gastric erosion caused by nonsteroidal anti-inflammatory drugs. The potential uses of LPA2 agonists like OTP and sulfamoyl benzoic acid-based radioprotectins must be further explored for therapeutic uses.

Cutaneous exposure to lewisite causes acute kidney injury by invoking DNA damage and autophagic response.

Lewisite (2-chlorovinyldichloroarsine) is an organic arsenical chemical warfare agent that was developed and weaponized during World Wars I/II. Stockpiles of lewisite still exist in many parts of the world and pose potential environmental and human health threat. Exposure to lewisite and similar chemicals causes intense cutaneous inflammatory response. However, morbidity and mortality in the exposed population is not only the result of cutaneous damage but is also a result of systemic injury. Here, we provide data delineating the pathogenesis of acute kidney injury (AKI) following cutaneous exposure to lewisite and its analog phenylarsine oxide (PAO) in a murine model. Both agents caused renal tubular injury, characterized by loss of brush border in proximal tubules and tubular cell apoptosis accompanied by increases in serum creatinine, neutrophil gelatinase-associated lipocalin, and kidney injury molecule-1. Interestingly, lewisite exposure enhanced production of reactive oxygen species (ROS) in the kidney and resulted in the activation of autophagic and DNA damage response (DDR) signaling pathways with increased expression of beclin-1, autophagy-related gene 7, and LC-3A/B-II and increased phosphorylation of γ-H2A.X and checkpoint kinase 1/2, respectively. Terminal deoxyribonucleotide-transferase-mediated dUTP nick-end labeling-positive cells were detected in renal tubules along with enhanced proapoptotic BAX/cleaved caspase-3 and reduced antiapoptotic BCL2. Scavenging ROS by cutaneous postexposure application of the antioxidant N-acetyl-l-cysteine reduced lewisite-induced autophagy and DNA damage. In summary, we provide evidence that topical exposure to lewisite causes AKI. The molecular mechanism underlying these changes involves ROS-dependent activation of autophagy and DDR pathway associated with the induction of apoptosis.

Epigenetic modulation of intestinal Na+/H+ exchanger-3 expression.

Na+/H+ exchanger-3 (NHE3) is crucial for intestinal Na+ absorption, and its reduction has been implicated in infectious and inflammatory bowel diseases (IBD)-associated diarrhea. Epigenetic mechanisms such as DNA methylation are involved in the pathophysiology of IBD. Whether changes in DNA methylation are involved in modulating intestinal NHE3 gene expression is not known. Caco-2 and HuTu 80 cells were used as models of human intestinal epithelial cells. Normal C57/BL6, wild-type, or growth arrest and DNA damage-inducible 45b (GADD45b) knockout (KO) mice were used as in vivo models. NHE3 gene DNA methylation levels were assessed by MBDCap (MethyMiner) assays. Results demonstrated that in vitro methylation of NHE3 promoter construct (p-1509/+127) cloned into a cytosine guanine dinucleotide-free lucia vector decreased the promoter activity in Caco-2 cells. DNA methyltransferase inhibitor 5-azacytidine (10 µM, 24 h) caused a significant decrease in DNA methylation of the NHE3 gene and concomitantly increased NHE3 expression in Caco-2 cells. Similarly, 5-azacytidine treatment increased NHE3 mRNA levels in HuTu 80 cells. 5-Azacytidine treatment for 3 wk (10 mg/kg body wt ip, 3 times/wk) also resulted in an increase in NHE3 expression in the mouse ileum and colon. Small-interfering RNA knockdown of GADD45b (protein involved in DNA demethylation) in Caco-2 cells decreased NHE3 mRNA expression. Furthermore, there was a significant decrease in NHE3 mRNA and protein expression in the ileum and colon of GADD45b KO mice. Our findings demonstrate that NHE3 gene expression is regulated by changes in its DNA methylation. NEW & NOTEWORTHY Our studies for the first time demonstrate that Na+/H+ exchanger-3 gene expression is regulated by an epigenetic mechanism involving DNA methylation.

Role of intracellular calcium and NADPH oxidase NOX5-S in acid-induced DNA damage in Barrett's cells and Barrett's esophageal adenocarcinoma cells.

Mechanisms whereby acid reflux may accelerate the progression from Barrett's esophagus (BE) to esophageal adenocarcinoma (EA) are not fully understood. Acid and reactive oxygen species (ROS) have been reported to cause DNA damage in Barrett's cells. We have previously shown that NADPH oxidase NOX5-S is responsible for acid-induced H2O2 production in Barrett's cells and in EA cells. In this study we examined the role of intracellular calcium and NADPH oxidase NOX5-S in acid-induced DNA damage in a Barrett's EA cell line FLO and a Barrett's cell line CP-A. We found that pulsed acid treatment significantly increased tail moment in FLO and CP-A cells and histone H2AX phosphorylation in FLO cells. In addition, acid treatment significantly increased intracellular Ca(2+) in FLO cells, an increase that is blocked by Ca(2+)-free medium with EGTA and thapsigargin. Acid-induced increase in tail moment was significantly decreased by NADPH oxidase inhibitor diphenylene iodonium in FLO cells, and by blockade of intracellular Ca(2+) increase or knockdown of NOX5-S with NOX5 small-interfering RNA (siRNA) in FLO and CP-A cells. Acid-induced increase in histone H2AX phosphorylation was significantly decreased by NOX5 siRNA in FLO cells. Conversely, overexpression of NOX5-S significantly increased tail moment and histone H2AX phosphorylation in FLO cells. We conclude that pulsed acid treatment causes DNA damage via increase of intracellular calcium and activation of NOX5-S. It is possible that in BE acid reflux increases intracellular calcium, activates NOX5-S, and increases ROS production, which causes DNA damage, thereby contributing to the progression from BE to EA.

Assessment of the deoxyribonucleic acid damage caused by occupational exposure to chemical compounds in Isfahan Polyacryl Company.

BACKGROUND: Chemical pollutants found in industrial environments can cause chronic genotoxicity in vulnerable individuals during the long-term exposure. The primary purpose of the present study was to assess the deoxyribonucleic acid (DNA) damage caused by occupational exposure to industrial chemicals and secondary purpose is to investigate the effect of possible risk factors of genotoxicity. MATERIALS AND METHODS: The blood samples of the workers of Isfahan Polyacryl Company were evaluated in terms of genotoxicity using the comet assay method. The percentage of DNA in the tail and tail moment were measured and DNA damage was evaluated. Furthermore, the effect of age, smoking, duration of working in the company and working in two parts of the company on the degree of vulnerability to genotoxicity was assessed. RESULTS: The amount of DNA damage in the target group (the production line workers) was significantly higher than the control group (the staffs), 3.87 versus 1.52 as tail moment, (P < 0.0001). DNA damage was significantly higher in smoker groups compared with non-smoker target group and control group, 4.18 versus 3.07 and 1.52 respectively as tail moment, (P < 0.0001). Furthermore, it was higher in person working in two different parts of the company compared to those work in one part and control group, 4.63 versus 3.74 and 1.52 respectively as tail moment, (P < 0.0001). CONCLUSION: Occupational exposure to Polyacryl caused DNA damage. Smoking and working in two parts of the company may have a significant role in DNA damage.

Deoxyribonucleic acid damage study in primary amenorrhea by comet assay and karyotyping.

AIM: This study aims at evaluating the chromosomal abnormalities and deoxyribonucleic acid (DNA) damage in cases with primary amenorrhea by karyotyping and comet assay. STUDY DESIGN: A total of 30 cases of primary amenorrhea were recruited. Secondary sexual characters were assessed by Tanner staging. Chromosomal analysis was performed by conventional phytohemagglutinin stimulated lymphocyte cell culture technique. Alkaline version of comet assay was used to evaluate DNA damage. RESULTS: The chromosomal pattern of 20 subjects (66.7%) was found to be normal (46,XX). Two subjects had 46,XY pattern and eight subjects had Turner syndrome (45,X or 45,X/46,XX). The comet parameters were found to be increased among subjects with 45,X monosomy, when compared to the rest of the study group and also in subjects with Tanner stage 1 when compared to stage 2. CONCLUSION: Comet assay revealed increased DNA damage in cases with 45,X monosomy, compared with subjects with 46,XX and 46,XY karyotype, which correlated with clinical features.

Deoxyribonucleic acid damage in Iranian veterans 25 years after wartime exposure to sulfur mustard.

BACKGROUND: More than 100,000 Iranian veterans and civilians still suffer from various long-term complications due to their exposure to sulfur mustard (SM) during the Iran-Iraq war in 1983-88. The aim of the study was to investigate DNA damage of SM in veterans who were exposed to SM, 23-27 years prior to this study. MATERIALS AND METHODS: Blood samples were obtained from the veterans and healthy volunteers as negative controls. Lymphocytes were isolated from blood samples and DNA breaks were measured using single-cell microgel electrophoresis technique under alkaline conditions (comet assay). Single cells were analyzed with "Tri Tek Comet Score version 1.5" software and DNA break was measured based on the percentage of tail DNA alone, or in the presence of H2O2 (25 µM) as a positive control. RESULTS: A total of 25 SM exposed male veterans and 25 male healthy volunteers with similar ages (44.66 ± 6.2 and 42.12 ± 5.75 years, respectively) were studied. Percentage of the lymphocyte DNA damage was significantly (P < 0.01) higher in the SM-exposed individuals than in the controls (6.47 ± 0.52 and 1.31 ± 0.35, respectively). Percentages of DNA damage in the different age groups of 35-39, 40-44, 45-49, and 50-54 years in SM-exposed veterans (5.48 ± 0.17, 6.7 3 ± 1.58, 6.42 ± 0.22, and 7.27 ± 0.38, respectively) were all significantly (P < 0.05) higher than the controls (1.18 ± 0.25, 1.53 ± 0.22, 1.27 ± 0.20, and 1.42 ± 0.10, respectively). The lymphocytes incubated with H2O2 had much higher DNA damage as expected. The average of tail DNA is 42.12 ± 2.75% for control cells + H2O2 and 18.48 ± 2.14% for patients cells + H2O2; P < 0.001. CONCLUSION: SM exposure of the veterans revealed DNA damage as judged by the comet assay.

Detection of Low Level Microwave Radiation Induced Deoxyribonucleic Acid Damage Vis-à-vis Genotoxicity in Brain of Fischer Rats.

BACKGROUND: Non-ionizing radiofrequency radiation has been increasingly used in industry, commerce, medicine and especially in mobile phone technology and has become a matter of serious concern in present time. OBJECTIVE: The present study was designed to investigate the possible deoxyribonucleic acid (DNA) damaging effects of low-level microwave radiation in brain of Fischer rats. MATERIALS AND METHODS: Experiments were performed on male Fischer rats exposed to microwave radiation for 30 days at three different frequencies: 900, 1800 and 2450 MHz. Animals were divided into 4 groups: Group I (Sham exposed): Animals not exposed to microwave radiation but kept under same conditions as that of other groups, Group II: Animals exposed to microwave radiation at frequency 900 MHz at specific absorption rate (SAR) 5.953 × 10(-4) W/kg, Group III: Animals exposed to 1800 MHz at SAR 5.835 × 10(-4) W/kg and Group IV: Animals exposed to 2450 MHz at SAR 6.672 × 10(-4) W/kg. At the end of the exposure period animals were sacrificed immediately and DNA damage in brain tissue was assessed using alkaline comet assay. RESULTS: In the present study, we demonstrated DNA damaging effects of low level microwave radiation in brain. CONCLUSION: We concluded that low SAR microwave radiation exposure at these frequencies may induce DNA strand breaks in brain tissue.

Evaluation of Deoxyribonucleic Acid Damage Using Neutral Comet Assay for High Radiation Doses: A Feasibility Study.

Purpose: This study aims to investigate the use of the neutral comet assay to assess deoxyribonucleic acid (DNA) damage in lymphocytes exposed to high doses of radiation. Materials and Methods: The research was conducted by obtaining informed consent, after which blood samples were taken from seven healthy individuals and this study was approved by the institutional ethics committee. At first, for the determination of dose-effect curves, samples obtained from the first five individuals were irradiated for doses ranging from 0 to 35 Gy after which they were processed under neutral comet assay. In order to verify the determined dose-effect curves, a test dose of 15 Gy was delivered to the samples obtained from the sixth and seventh individuals. The amount of DNA damage from the obtained comet assay images was analyzed using four comet assay parameters namely % tail DNA, tail length, tail moment (TM), and Olive TM (OTM). The most suitable comet assay parameter was evaluated based on the obtained dose-effect curves. Furthermore, the distribution of individual cells for each dose point was evaluated for all the four comet assay parameters to find the optimal parameter. Results: From our results, it was found that from 0 to 25 Gy all the four comet assay parameters fit well into a linear quadratic curve and above 25 Gy saturation was observed. Based on the individual cell distribution data, it was found that % tail DNA could be an optimal choice to evaluate DNA damage while using neutral comet assay for high-dose ionizing radiation. Conclusion: The neutral comet assay could be a potential tool to assess DNA damage from high doses of ionizing radiation greater than 5 Gy.

Investigations of the novel checkpoint kinase 1 inhibitor SRA737 in non-small cell lung cancer and colorectal cancer cells of differing tumour protein 53 gene status.

Aim: In response to DNA damage the serine/threonine-specific protein kinase checkpoint kinase 1 (CHK1) is activated allowing cells to enter S phase (S) and G2 phase (G2) cell-cycle arrest. CHK1 inhibitors are expected to prevent cells from entering such arrest, thereby enhancing DNA damage-induced cytotoxicity. In contrast, normal cells with intact ataxia-telangiectasia mutated (ATM), CHK2 and tumour suppressor protein 53 (P53) signalling are still able to enter cell-cycle arrest using the functioning G1/S checkpoint, thereby being rescued from enhanced cytotoxicity. The main objective of this work is to investigate the in vitro effects of the novel CHK1 inhibitor SRA737 on pairs of non-small cell lung cancer (NSCLC) and colorectal cancer (CRC) cell lines, all with genetic aberrations rendering them susceptible to replication stress but of differing tumour protein 53 (TP53) gene status, focusing on DNA damage induction and the subsequent effects on cell proliferation and viability. Methods: NSCLC cell lines H23 [TP53 mutant (MUT)] and A549 [TP53 wild-type (WT)] and CRC cell lines HT29 (TP53 MUT) and HCT116 (TP53 WT) were incubated with differing micromolar concentrations of SRA737 for 24 h and then analysed using alkaline comet and phosphorylated H2A.X variant histone (γH2AX)-foci assays to assess mostly DNA single strand break and double strand break damage, respectively. Cell-counting/trypan blue staining was also performed to assess cell proliferation/viability. Results: Clear concentration-dependent increases in comet formation and γH2AX-foci/cell were noted for the TP53 MUT cells with no or lower increases being noted in the corresponding TP53 WT cells. Also, greater anti-proliferative and cell killing effects were noted in the TP53 MUT cells than in the TP53 WT cells. Conclusions: This study's data suggests that P53 status/functioning is a key factor in determining the sensitivity of NSCLC and CRC cancer cells towards CHK1 inhibition, even in circumstances conducive to high replicative stress.

LncRNA NORAD mediates KMT2D expression by targeting miR-204-5p and affects the growth of gastric cancer.

Background: Long non-coding ribonucleic acids (lncRNAs) are a class of non-coding RNAs implicated in the development of many malignancies, including gastric cancer (GC). In this study, we investigated the functions and molecular mechanisms of non-coding RNA activated by deoxyribonucleic acid damage (NORAD) in GC. Methods: NORAD expression at the messenger RNA levels was determined by quantitative reverse transcriptase (RT)-polymerase chain reaction assays. Cell proliferation, migration, and invasion were detected by Cell Counting Kit-8 assays, in-vivo tumor formation assays, and Transwell assays. Cell-cycle distribution was detected by a flow cytometry analysis. NORAD location was detected by nucleocytoplasmic fractionation assays. The interaction between NORAD and the microRNA-204-5p (miR-204-5p)/Lysine Methyltransferase 2D (KMT2D) axis was verified by dual-luciferase reporter gene assays and RNA binding protein immunoprecipitation (RIP) assays. Western blot was used to study the phosphatase and tensin homolog (PTEN)/phosphoinositide 3-kinases (PI3K)/protein kinase B (AKT) signaling pathway. Results: NORAD was upregulated in the GC tissues and cell lines. The silencing of NORAD repressed cell proliferation and the Growth 2 (G2)/Mitosis (M) cell-cycle transition in GC. NORAD also regulated KMT2D expression by targeting miR-204-5p and mediated PTEN/PI3K/AKT signaling in GC. Conclusions: We found that NORAD acts as an oncogene in GC. Our findings might provide a novel therapeutic target for GC.

Changes in the levels of comet parameters before and after fluoxetine therapy in major depression patients.

Major depression belongs to mood disorders and characterized by worthlessness, no interest or happiness in any activity; lasting for atleast two weeks. Etio-pathological changes of major depression include oxidative stress leading to free radical synthesis which causes damage to carbohydrates, proteins, lipids and nucleic acids. Nucleic acid damage can be identified by either single or double strand breaks and for quantitative estimation of the same, neutral or alkaline comet assay is performed. Fluoxetine is the drug of choice for treatment of major depression having antioxidant function. In the current study eighty drug naïve major depression patients were recruited and comet parameters namely total comet length, head diameter and tail length were measured before starting the treatment and after completion of eight week fluoxetine therapy. The levels of comet parameters were higher in females than males suggesting higher prevalence of major depression among females. On categorizing into three age groups, the numbers of major depression patients belonging to 18-30 year age group were higher than 31-40 and 41-50 year age groups. All the parameters of deoxyribonucleic acid damage were reduced after eight week of fluoxetine therapy indicating that fluoxetine has anti-oxidant action along with its antidepressant properties, which cause reversal of oxidative stress induced damage occurring during major depression.

Changes in Deoxyribonucleic Acid Methylation Contribute to the Pathophysiology of Multiple Sclerosis.

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system characterized by loss of coordination, weakness, dysfunctions in bladder capacity, bowel movement, and cognitive impairment. Thus, the disease leads to a significant socioeconomic burden. In the pathophysiology of the disease, both genetic and environmental risk factors are involved. Gene x environment interaction is modulated by epigenetic mechanisms. Epigenetics refers to a sophisticated system that regulates gene expression with no changes in the DNA sequence. The most studied epigenetic mechanism is the DNA methylation. In this review, we summarize the data available from the current literature by grouping sets of differentially methylated genes in distinct biological categories: the immune system including innate and adaptive response, the DNA damage, and the central nervous system.

Cardiac Nuclear High-Mobility Group Box 1 Ameliorates Pathological Cardiac Hypertrophy by Inhibiting DNA Damage Response.

High-mobility group box 1 (HMGB1) is a deoxyribonucleic acid (DNA)-binding protein associated with DNA repair. Decreased nuclear HMGB1 expression and increased DNA damage response (DDR) were observed in human failing hearts. DNA damage and DDR as well as cardiac remodeling were suppressed in cardiac-specific HMGB1 overexpression transgenic mice after angiotensin II stimulation as compared with wild-type mice. In vitro, inhibition of HMGB1 increased phosphorylation of extracellular signal-related kinase 1/2 and nuclear factor kappa B, which was rescued by DDR inhibitor treatment. DDR inhibitor treatment provided a cardioprotective effect on angiotensin II-induced cardiac remodeling in mice.

A study of radiation injury in rat C6 glioma cell line by 1H-nuclear magnetic resonance spectroscopy / 中华放射肿瘤学杂志

Objective To study the radiation injury of rat C6 glioma cell line by high resolution,1 H-nuclear magnetic resonance (1 H NMR) spectroscopy,and to preliminarily investigate its mechanism.Methods Metabolite concentrations in C6 cells were determined by 1 H NMR spectroscopy.Comet assay was used to evaluate DNA damage.Flow cytometry was used to determine the cell cycle and apoptosis rate.Colony-forming assay was used to measure the colony-forming rate and preliminarily investigate the mechanism of radiation injury.The resuhs were analyzed by one-way analysis of variance and Pearson correlation analysis.Results With the increase in radiation dose from 0 Gy to 1,5,10,and 15 Gy,DNA damage was enhanced in a dose-dependent manner (P=0.000-0.690);the percentage of cells in G1 phase increased (P =0.026-0.749);the apoptosis rate significantly increased (all P =0.000);the colony-forming rate significantly declined (P =0.000-0.004);the Lac/Cr ratio significantly decreased (P =0.000-0.015),which had a negative linear correlation with DNA damage parameters (tail length,r=-0.971;％DNA in the tail,r =-0.998;tail moment,r =-0.995) and apoptosis rate (r =0.978).Conclusions 1 H NMR spectroscopy reveals that the change in the Lac/Cr ratio is associated with injury and apoptosis of C6 cells after radiation.1 H NMR spectroscopy has the potential to predict radiation injury of glioma.

Cellular response to fludarabine treatment in combination with different ionizing radiation in renal carcinoma 786-O cells / 中华放射肿瘤学杂志

Objective To investigate DNA double-strand breaks and radiosensitization in renal carcinoma 786-O cells induced by fludarabine (FA) combined with different ionizing radiations.Methods The 786-O cells were exposed to FA combined with X-ray or heavy ion beam irradiation.Flow cytometry was used to evaluate the percentage of γH2AX-positive cells and cell cycle.The neutral comet assay was used to detect DNA double-strand breaks.The colony-forming assay was used to evaluate the effects of different treatments on cell survival.Comparison between groups was made by one-way analysis of variance or Dunnet' s t test.Results Compared with FA alone or irradiation alone,FA combined with different ionizing radiations increased DNA double-strand breaks as shown by significantly increased levels of γH2AX (P=0.007,0.001);FA combined with heavy ion beam irradiation lead to a cell cycle block at the radiosensitive G2/M phase and significantly increased the expression of γH2AX in the G2/M phase (P=0.000,0.000);the neutral comet assay revealed that FA combined with irradiation significantly increased DNA sublethal damage (P=0.020,0.060);FA significantly reduced the colony-forming rate after irradiation (P=0.000,0.030;0.001,0.040).Conclusions FA enhances the effects induced by X-ray and heavy ion beam irradiation with different properties.Particularly,FA substantially enhances the cell death induced by heavy ion beam irradiation.

Cytotoxicity and DNA damage in the neutrophils of patients with sickle cell anaemia treated with hydroxyurea

Hydroxyurea (HU) is the most important advance in the treatment of sickle cell anaemia (SCA) for preventing complications and improving quality of life for patients. However, some aspects of treatment with HU remain unclear, including their effect on and potential toxicity to other blood cells such as neutrophils. This study used the measurement of Lactate Dehydrogenase (LDH) and Methyl ThiazolTetrazolium (MTT) and the comet assay to investigate the cytotoxicity and damage index (DI) of the DNA in the neutrophils of patients with SCA using HU.In the LDH and MTT assays, a cytoprotective effect was observed in the group of patients treated, as well as an absence of toxicity. When compared to patients without the treatment, the SS group (n=20, 13 women and 07 men, aged 18-69 years), and the group of healthy individuals (AA) used as a control group (n=52, 28 women and 24 men, aged 19-60 years), The SSHU group (n=21, 11 women and 10 men, aged 19-63 years) showed a significant reduction (p<0.001) in LDH activity and an increase in the percentage of viable cells by the MTT (p<0.001). However, the SSHU group presented significantly higher DI values (49.57±6.0 U/A) when compared to the AA group (7.43 ± 0,94U/A) and the SS group (22.73 ±5.58 U/A) (p<0.0001), especially when treated for longer periods (>20 months), demonstrating that despite the cytoprotective effects in terms of cell viability, the use of HU can induce DNA damage in neutrophils.

A hidroxiuréia (HU) constitui o avanço mais importante no tratamento da anemia falciforme (AF) por prevenir complicações e aumentar a qualidade de vida dos pacientes. Entretanto, alguns aspectos do tratamento com HU permanecem obscuros, incluindo a sua ação e potencial toxicidade em outras células sanguíneas, tais como neutrófilos. Este estudo utilizou a mensuração da lactato desidrogenase (LDH) e do metil tiazoltetrazólio (MTT) e o ensaio do cometa para investigar a citotoxicidade e índice de dano (ID) ao DNA em neutrófilos de pacientes com AF em uso do medicamento. Nos ensaios de LDH e MTT, observou-se além de ausência de toxicidade, uma ação citoprotetora no grupo de pacientes tratados, Grupo SSHU (n=21, 11 mulheres e 10 homens, com idades entre 19-63 anos), quando comparados aos pacientes sem tratamento, Grupo SS (n=20, 13 mulheres e 07 homens, 18-69 anos), e grupo de indivíduos saudáveis (AA) usado como controle (n=52, 28 mulheres e 24 homens, 19-60 anos), com redução significativa (p<0,001) na atividade de LDH e aumento no percentual de células viáveis pelo MTT (p<0,001). Entretanto, o grupo SSHU apresentou valores de ID significativamente elevados (49,57±6,0 U/A), quando comparados ao grupo AA (7,43 ± 0,94U/A) e grupo SS (22,73 ±5,58 U/A) (p<0,0001), especialmente quando tratados por períodos mais longos (>20 meses), demonstrando que apesar dos efeitos citoprotetores quanto à viabilidade celular, o uso da HU pode induzir lesão ao DNA de neutrófilos.

The effect of β-elemene combined with irradiation on DNA damage and repair in A549 cells / 中华放射肿瘤学杂志

Objective To study if β-elemene can increase radiation-induced deoxyribonucleic acid (DNA) damage and decrease the damage repair.Methods Exponentially growing human lung adenocarcinoma cells (A549) were exposed to 10 or 20 μg/ml β-elemene for 24 h before irradiation.The effect of β-elemene on the in vitro radiosensitivity of A549 cells was evaluated using clonogenic assay.DNA damage and repair were evaluated using comet assay.Results Exposure to β-elemene before irradiation increased the radiosensitivity of A549 cells.The SERD0 for 10 μg/ml and 20 μg/ml β-elemene was 1.55 and 1.64, respectively.The SERDq for 10 μg/ml and 20 μg/ml β-elemene was 1.43 and 1.75, respectively.Combined treatment, comparing to irradiation or β-elemene treatment alone, induced higher levels of DNA damage and slower rate of damage repair.A549 cells exposed to 20 μg/ml β-elemene followed by irradiation showed a higher levels of tail moment (TM) than those exposed to irradiation or β-elemene alone at 0 h,2 h,6 h and 24 h after irradiation.The TM of the three groups at 0 h,2 h,6 h and 24 h after irradiation was 7.16±2.61,0.95±0.65 and 1.81±1.23(F=231.24,P<0.01), 3.65±2.06,0.11±0.07 and 1.58±1.40(F=90.22,P<0.01), 2.09±0.83,0.1±0.05 and 0.45±0.25(F=238.44,P<0.01), 1.45±1.37,0.11±0.08 and 0.60±0.40(F=38.94,P<0.01), respectively. Conclusions β-elemene can enhance the radiosensitivity of A549 cells through the enhancement of DNA damage and the inhibition of DNA damage repair.