RESUMEN
Numerous metals are well-known human bladder carcinogens. Despite the significant occupational and public health concern of metals and bladder cancer, the carcinogenic mechanisms remain largely unknown. Chromium, in particular, is a metal of concern as incidences of bladder cancer have been found elevated in chromate workers, and there is an increasing concern for patients with metal hip implants. However, the impact of hexavalent chromium (Cr(VI)) on bladder cells has not been studied. We compared chromate toxicity in two bladder cell lines; primary human urothelial cells and hTERT-immortalized human urothelial cells. Cr(VI) induced a concentration- and time-dependent increase in chromosome damage in both cell lines, with the hTERT-immortalized cells exhibiting more chromosome damage than the primary cells. Chronic exposure to Cr(VI) also induced a concentration-dependent increase in aneuploid metaphases in both cell lines which was not observed after a 24h exposure. Aneuploidy induction was higher in the hTERT-immortalized cells. When we correct for uptake, Cr(VI) induces a similar amount of chromosome damage and aneuploidy suggesting that the differences in Cr(VI) sensitivity between the two cells lines were due to differences in uptake. The increase in chromosome instability after chronic chromate treatment suggests this may be a mechanism for chromate-induced bladder cancer, specifically, and may be a mechanism for metal-induced bladder cancer, in general.
Asunto(s)
Cromo/toxicidad , Inestabilidad Cromosómica/efectos de los fármacos , Inestabilidad Cromosómica/fisiología , Urotelio/efectos de los fármacos , Urotelio/fisiología , Células Cultivadas , Daño del ADN/efectos de los fármacos , Daño del ADN/fisiología , Relación Dosis-Respuesta a Droga , Células Epiteliales/efectos de los fármacos , Células Epiteliales/patología , Células Epiteliales/fisiología , Humanos , Urotelio/patologíaRESUMEN
Cobalt exposure is increasing as cobalt demand rises worldwide due to its use in enhancing rechargeable battery efficiency, super-alloys, and magnetic products. Cobalt is considered a possible human carcinogen with the lung being a primary target. However, few studies have considered cobalt-induced toxicity in human lung cells. Therefore, in this study, we sought to determine the cytotoxicity and genotoxicity of particulate and soluble cobalt in human lung cells. Cobalt oxide and cobalt chloride were used as representative particulate and soluble cobalt compounds, respectively. Exposure to both particulate and soluble cobalt induced a concentration-dependent increase in cytotoxicity, genotoxicity, and intracellular cobalt ion levels. Based on intracellular cobalt ion levels, we found that soluble cobalt was more cytotoxic than particulate cobalt while particulate and soluble cobalt induced similar levels of genotoxicity. However, soluble cobalt induced cell cycle arrest indicated by the lack of metaphases at much lower intracellular cobalt concentrations compared to cobalt oxide. Accordingly, we investigated the role of particle internalization in cobalt oxide-induced toxicity and found that particle-cell contact was necessary to induce cytotoxicity and genotoxicity after cobalt exposure. These data indicate that cobalt compounds are cytotoxic and genotoxic to human lung fibroblasts, and solubility plays a key role in cobalt-induced lung toxicity.
Asunto(s)
Carcinógenos Ambientales/toxicidad , Cobalto/toxicidad , Pulmón/efectos de los fármacos , Mutágenos/toxicidad , Transporte Biológico , Carcinógenos Ambientales/análisis , Carcinógenos Ambientales/química , Carcinógenos Ambientales/metabolismo , Ciclo Celular , Línea Celular , Supervivencia Celular/efectos de los fármacos , Células Clonales , Cobalto/análisis , Cobalto/química , Cobalto/metabolismo , Fragmentación del ADN/efectos de los fármacos , Humanos , Pulmón/química , Pulmón/metabolismo , Mutágenos/análisis , Mutágenos/química , Mutágenos/metabolismo , Concentración Osmolar , Óxidos/análisis , Óxidos/química , Óxidos/metabolismo , Óxidos/toxicidad , Tamaño de la Partícula , Fagocitosis/efectos de los fármacos , SolubilidadRESUMEN
Depleted uranium (DU) is extensively used in both industry and military applications. The potential for civilian and military personnel exposure to DU is rising, but there are limited data on the potential health hazards of DU exposure. Previous laboratory research indicates DU is a potential carcinogen, but epidemiological studies remain inconclusive. DU is genotoxic, inducing DNA double strand breaks, chromosome damage and mutations, but the mechanisms of genotoxicity or repair pathways involved in protecting cells against DU-induced damage remain unknown. The purpose of this study was to investigate the effects of homologous recombination repair deficiency on DU-induced genotoxicity using RAD51D and XRCC3-deficient Chinese hamster ovary (CHO) cell lines. Cells deficient in XRCC3 (irs1SF) exhibited similar cytotoxicity after DU exposure compared to wild-type (AA8) and XRCC3-complemented (1SFwt8) cells, but DU induced more break-type and fusion-type lesions in XRCC3-deficient cells compared to wild-type and XRCC3-complemented cells. Surprisingly, loss of RAD51D did not affect DU-induced cytotoxicity or genotoxicity. DU induced selective X-chromosome fragmentation irrespective of RAD51D status, but loss of XRCC3 nearly eliminated fragmentation observed after DU exposure in wild-type and XRCC3-complemented cells. Thus, XRCC3, but not RAD51D, protects cells from DU-induced breaks and fusions and also plays a role in DU-induced chromosome fragmentation.
Asunto(s)
Aberraciones Cromosómicas/efectos de la radiación , Cromosomas de los Mamíferos/efectos de la radiación , Mutágenos/farmacología , Reparación del ADN por Recombinación , Uranio/farmacología , Animales , Células CHO , Cromosomas de los Mamíferos/química , Cricetulus , Roturas del ADN de Doble Cadena/efectos de los fármacos , Proteínas de Unión al ADN/deficiencia , Proteínas de Unión al ADN/genética , Prueba de Complementación GenéticaRESUMEN
Inappropriate survival signaling after DNA damage may facilitate clonal expansion of genetically compromised cells, and it is known that protein tyrosine phosphatase (PTP) inhibitors activate key survival pathways. In this study we employed the genotoxicant, hexavalent chromium [Cr(VI)], which is a well-documented carcinogen of occupational and environmental concern. Cr(VI) induces a complex array of DNA damage, including DNA double strand breaks (DSBs). We recently reported that PTP inhibition bypassed cell cycle arrest and abrogated Cr(VI)-induced clonogenic lethality. Notably, PTP inhibition resulted in an increase in forward mutations at the HPRT locus, supporting the hypothesis that PTP inhibition in the presence of DNA damage may lead to genomic instability (GIN), via cell cycle checkpoint bypass. The aim of the present study was to determine the effect of PTP inhibition on DNA DSB formation and chromosomal integrity after Cr(VI) exposure. Diploid human lung fibroblasts were treated with Cr(VI) in the presence or absence of the PTP inhibitor, sodium orthovanadate, for up to 24h, and cells were analyzed for DNA DSBs and chromosomal damage. Cr(VI) treatment induced a rapid increase in DNA DSBs, and a significant increase in total chromosomal damage (chromatid breaks and gaps) after 24h. In sharp contrast, PTP inhibition abrogated both DNA DSBs and chromosomal damage after Cr(VI) treatment. In summary, PTP inhibition in the face of Cr(VI) genotoxic stress decreases chromosomal instability (CIN) but increases mutagenesis, which we postulate to be a result of error-prone DNA repair.
Asunto(s)
Inestabilidad Cromosómica/efectos de los fármacos , Inestabilidad Genómica/efectos de los fármacos , Proteínas Tirosina Fosfatasas/antagonistas & inhibidores , Cromo/toxicidad , Roturas del ADN de Doble Cadena , Daño del ADN , Fibroblastos , Humanos , Pulmón/citologíaRESUMEN
Amyotrophic lateral sclerosis (ALS) is characterized by motor neuron degeneration accompanied by aberrant accumulation and loss of function of the RNA-binding protein TDP43. Thus far, it remains unresolved to what extent TDP43 loss of function directly contributes to motor system dysfunction. Here, we employed gene editing to find whether the mouse ortholog of the TDP43-regulated gene STMN2 has an important function in maintaining the motor system. Both mosaic founders and homozygous loss-of-function Stmn2 mice exhibited neuromuscular junction denervation and fragmentation, resulting in muscle atrophy and impaired motor behavior, accompanied by an imbalance in neuronal microtubule dynamics in the spinal cord. The introduction of human STMN2 through BAC transgenesis was sufficient to rescue the motor phenotypes observed in Stmn2 mutant mice. Collectively, our results demonstrate that disrupting the ortholog of a single TDP43-regulated RNA is sufficient to cause substantial motor dysfunction, indicating that disruption of TDP43 function is likely a contributor to ALS.
Asunto(s)
Esclerosis Amiotrófica Lateral , Estatmina , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Modelos Animales de Enfermedad , Homocigoto , Ratones , Ratones Transgénicos , Neuronas Motoras/metabolismo , Unión Neuromuscular/metabolismo , Estatmina/genética , Estatmina/metabolismoRESUMEN
The cohesin complex plays an essential role in chromosome maintenance and transcriptional regulation. Recurrent somatic mutations in the cohesin complex are frequent genetic drivers in cancer, including myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). Here, using genetic dependency screens of stromal antigen 2-mutant (STAG2-mutant) AML, we identified DNA damage repair and replication as genetic dependencies in cohesin-mutant cells. We demonstrated increased levels of DNA damage and sensitivity of cohesin-mutant cells to poly(ADP-ribose) polymerase (PARP) inhibition. We developed a mouse model of MDS in which Stag2 mutations arose as clonal secondary lesions in the background of clonal hematopoiesis driven by tet methylcytosine dioxygenase 2 (Tet2) mutations and demonstrated selective depletion of cohesin-mutant cells with PARP inhibition in vivo. Finally, we demonstrated a shift from STAG2- to STAG1-containing cohesin complexes in cohesin-mutant cells, which was associated with longer DNA loop extrusion, more intermixing of chromatin compartments, and increased interaction with PARP and replication protein A complex. Our findings inform the biology and therapeutic opportunities for cohesin-mutant malignancies.
Asunto(s)
Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/metabolismo , Reparación del ADN/genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Mutación , Síndromes Mielodisplásicos/genética , Síndromes Mielodisplásicos/metabolismo , Animales , Línea Celular Tumoral , Cromatina/genética , Cromatina/metabolismo , Daño del ADN , Modelos Animales de Enfermedad , Femenino , Humanos , Células K562 , Leucemia Mieloide Aguda/tratamiento farmacológico , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones Mutantes , Ratones SCID , Ratones Transgénicos , Síndromes Mielodisplásicos/tratamiento farmacológico , Proteínas Nucleares/genética , Ftalazinas/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Células U937 , Ensayos Antitumor por Modelo de Xenoinjerto , CohesinasRESUMEN
Exposure to toxic and carcinogenic metals is widespread; however, their mechanisms of action remain largely unknown. One potential mechanism for metal-induced carcinogenicity and toxicity is centrosome amplification. Here we review the mechanisms for metal-induced centrosome amplification, including arsenic, chromium, mercury and nano-titanium dioxide.
Asunto(s)
Arsénico/toxicidad , Centrosoma/efectos de los fármacos , Centrosoma/metabolismo , Cromo/toxicidad , Mercurio/toxicidad , Titanio/toxicidad , Animales , Carcinógenos/toxicidad , Línea Celular , Exposición a Riesgos Ambientales , Humanos , Nanoestructuras/toxicidad , Titanio/químicaRESUMEN
Hexavalent chromium (Cr(VI)) compounds are well-established human lung carcinogens. Solubility plays an important role in their carcinogenicity with the particulate Cr(VI) compounds being the most carcinogenic. Epidemiology and animal studies suggest that zinc chromate is the most potent particulate Cr(VI) compound; however, there are few comparative data to support these observations. The purpose of this study was to compare the genotoxicity of zinc chromate with two other particulate Cr(VI) compounds, barium chromate and lead chromate, and one soluble Cr(VI) compound, sodium chromate. The clastogenic effects of barium chromate and zinc chromate were similar, but lead chromate induced significantly less damage. The levels of DNA damage measured by gamma-H2A.X foci formation were similar for the three particulate chromium compounds. Corrected for chromium uptake differences, we found that zinc chromate and barium chromate were the most cytotoxic, and lead chromate and sodium chromate were less cytotoxic. Zinc chromate was more clastogenic than all other chromium compounds, and lead chromate was the least clastogenic. There was no significant difference between any of the compounds for the induction of DNA double strand breaks. All together, these data suggest that the difference in the carcinogenic potency of zinc chromate over the other chromium compounds is not due solely to a difference in chromium ion uptake and that the zinc cation may in fact have an important role in its carcinogenicity.
Asunto(s)
Bronquios/citología , Cromo/toxicidad , Pruebas de Mutagenicidad , Carcinógenos Ambientales/toxicidad , Línea Celular , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , HumanosRESUMEN
Hexavalent chromium (Cr(VI)) compounds are known human lung carcinogens. Solubility plays an important role in its carcinogenicity with the particulate or insoluble form being the most potent. Of the particulate Cr(VI) compounds, zinc chromate appears to be the most potent carcinogen; however, very few studies have investigated its carcinogenic mechanism. In this study, we investigated the ability of chronic exposure to zinc chromate to induce numerical chromosome instability. We found no increase in aneuploidy after a 24 h exposure to zinc chromate, but with more chronic exposures, zinc chromate induced concentration- and time-dependent increases in aneuploidy in the form of hypodiploidy, hyperdiploidy, and tetraploidy. Zinc chromate also induced centrosome amplification in a concentration- and time-dependent manner in both interphase and mitotic cells after chronic exposure, producing cells with centriolar defects. Furthermore, chronic exposure to zinc chromate induced concentration- and time-dependent increases in spindle assembly checkpoint bypass with increases in centromere spreading, premature centromere division, and premature anaphase. Last, we found that chronic exposure to zinc chromate induced a G2 arrest. All together, these data indicate that zinc chromate can induce chromosome instability after prolonged exposures.
Asunto(s)
Centrosoma/efectos de los fármacos , Cromatos/toxicidad , Inestabilidad Cromosómica/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Pulmón/efectos de los fármacos , Huso Acromático/efectos de los fármacos , Compuestos de Zinc/toxicidad , Aneuploidia , Línea Celular , Humanos , Pulmón/citología , Tamaño de la Partícula , SolubilidadRESUMEN
Hexavalent chromium Cr(VI) is a respiratory toxicant and carcinogen, with solubility playing an important role in its carcinogenic potential. Zinc chromate, a water insoluble or 'particulate' Cr(VI) compound, has been shown to be carcinogenic in epidemiology studies and to induce tumors in experimental animals, but its genotoxicity is poorly understood. Our study shows that zinc chromate induced concentration-dependent increases in cytotoxicity, chromosome damage and DNA double strand breaks in human lung cells. In response to zinc chromate-induced breaks, MRE11 expression was increased and ATM and ATR were phosphorylated, indicating that the DNA double strand break repair system was initiated in the cells. In addition, our data show that zinc chromate-induced double strand breaks were only observed in the G2/M phase population, with no significant amount of double strand breaks observed in G1 and S phase cells. These data will aid in understanding the mechanisms of zinc chromate toxicity and carcinogenesis.
Asunto(s)
Cromatos/toxicidad , Inestabilidad Cromosómica , Roturas del ADN de Doble Cadena , ADN/efectos de los fármacos , Fibroblastos/efectos de los fármacos , Pulmón/efectos de los fármacos , Mutágenos/toxicidad , Compuestos de Zinc/toxicidad , Proteínas de la Ataxia Telangiectasia Mutada , Ciclo Celular/efectos de los fármacos , Proteínas de Ciclo Celular/metabolismo , Línea Celular , Supervivencia Celular/efectos de los fármacos , ADN/metabolismo , Proteínas de Unión al ADN/metabolismo , Relación Dosis-Respuesta a Droga , Fibroblastos/metabolismo , Fibroblastos/patología , Humanos , Pulmón/metabolismo , Pulmón/patología , Proteína Homóloga de MRE11 , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Regulación hacia ArribaRESUMEN
Hexavalent chromium is a commonly used industrial metal that has been shown to induce lung cancer in workers having long term exposure. In the particulate form, Cr(VI) dissolves slowly in vivo, leading to an extended exposure of lung cells. Hexavalent chromium is taken into the cell and rapidly reduced to Cr(V), Cr(IV), Cr(III), and reactive oxygen species. Cells treated with Cr(VI) are subject to several types of DNA damage resulting from this reduction, including base modification, single-strand breaks, double-strand breaks, Cr-DNA adducts, DNA-Cr-DNA adducts, and protein-Cr-DNA adducts. These types of damage, if left unrepaired or are misrepaired, can lead to growth arrest, cytotoxicity, and apoptosis, as well as mutations leading to neoplastic transformation and ultimately tumorigenesis. Here we review the current literature on Cr-induced DNA damage and its repair.
Asunto(s)
Cromo/toxicidad , Daño del ADN , Reparación del ADN , Neoplasias Pulmonares/inducido químicamente , Contaminantes Ocupacionales del Aire/toxicidad , Humanos , Industrias , Neoplasias Pulmonares/patología , Enfermedades Profesionales/etiología , Exposición Profesional/efectos adversos , Estrés Oxidativo , Material Particulado/toxicidadRESUMEN
The endangered Western population of the Steller sea lion declined for three decades for uncertain reasons. We present baseline data of metal concentrations in pups as a first step towards investigating the potential threat of developmental exposures to contaminants. Seven metals were investigated: arsenic, cadmium, silver, aluminum, mercury, lead and vanadium. Vanadium was detected in only a single blubber sample. Mercury appears to be the most toxicologically significant metal with concentrations in the liver well above the current action level for mercury in fish. The concentrations of aluminum, arsenic, silver, cadmium and lead were present in one-fourth to two-thirds of all samples and were at either comparable or below concentrations previously reported. Neither gender nor region had a significant effect on metal burdens. Future work should consider metal concentrations in juveniles and adults and toxicological studies need to be performed to begin to assess the toxicity of these metals.
Asunto(s)
Metales Pesados/metabolismo , Leones Marinos/metabolismo , Contaminantes Químicos del Agua/metabolismo , Animales , Femenino , Masculino , Distribución TisularRESUMEN
Hexavalent chromium [Cr(VI)] compounds are established human lung carcinogens. The carcinogenicity of Cr(VI) is related to its solubility, with the most potent carcinogens being the insoluble particulate Cr(VI) compounds. However, it remains unknown why particulate Cr(VI) is more carcinogenic than soluble Cr(VI). One possible explanation is that particulates may provide more chronic exposures to chromate over time. We found that aneuploid cells increased in a concentration- and time-dependent manner after chronic exposure to lead chromate. Specifically, a 24-hour lead chromate exposure induced no aneugenic effect, whereas a 120-hour exposure to 0.5 and 1 microg/cm2 lead chromate induced 55% and 60% aneuploid metaphases, respectively. We also found that many of these aneuploid cells were able to continue to grow and form colonies. Centrosome defects are known to induce aneuploidy; therefore, we investigated the effects of chronic lead chromate exposure on centrosomes. We found that centrosome amplification in interphase and mitotic cells increased in a concentration- and time-dependent manner with 0.5 and 1 microg/cm2 lead chromate for 120 hours, inducing aberrant centrosomes in 18% and 21% of interphase cells and 32% and 69% of mitotic cells, respectively; however, lead oxide did not induce centrosome amplification in interphase or mitotic cells. There was also an increase in aberrant mitosis after chronic exposure to lead chromate with the emergence of disorganized anaphase and mitotic catastrophe. These data suggest that one possible mechanism for lead chromate-induced carcinogenesis is through centrosome dysfunction, leading to the induction of aneuploidy.
Asunto(s)
Aneuploidia , Centrosoma/efectos de los fármacos , Cromatos/toxicidad , Plomo/toxicidad , Pulmón/efectos de los fármacos , Línea Celular , Centrosoma/fisiología , Humanos , Pulmón/fisiología , Pulmón/ultraestructura , Mitosis/efectos de los fármacosRESUMEN
Particulate hexavalent chromium (Cr(VI)) is a well-established human lung carcinogen with widespread exposure among people in occupational settings and the general public. However, no studies have examined the chromate-induced malignant transformation of human lung epithelial cells, its predominant target. Human papillomavirus-immortalized human bronchial epithelial (BEP2D) cells were used to better understand the mechanisms involved in human bronchial carcinogenesis induced by particulate chromate. We found that aneuploid cells increased in a concentration-dependent manner after chronic exposure to lead chromate. Moreover, chronic exposure to lead chromate induced BEP2D cell transformation. Transformed BEP2D cells developed through a series of sequential steps, including altered cell morphology, loss of cell contact inhibition and anchorage-independent growth. Specifically, a 5-day exposure to lead chromate induced foci formation with 0, 1, 5, and 10 microg/cm2 lead chromate inducing 0, 7, 3, and 15 foci in 10 dishes. Anchorage independence was observed in cell lines derived from these foci. These foci-derived cells also showed centrosome amplification and increases in aneuploid metaphases. Our study demonstrates that particulate Cr(VI) is able to transform human bronchial epithelial cells, and that chromosome instability may play an important role in particulate Cr(VI)-induced neoplastic transformation.
Asunto(s)
Bronquios/efectos de los fármacos , Carcinógenos/toxicidad , Transformación Celular Neoplásica/inducido químicamente , Cromatos/toxicidad , Plomo/toxicidad , Bronquios/patología , Línea Celular Transformada , Transformación Celular Neoplásica/genética , Inestabilidad Cromosómica/efectos de los fármacos , HumanosRESUMEN
Particulate hexavalent chromium (Cr(VI)) is a well-established human lung carcinogen. It is currently a major public health concern, there is widespread exposure to it in occupational settings and to the general public. However, despite the potential widespread exposure and the fact that the lung is its target organ, few studies have considered the toxic effects of particulate Cr(VI) in human lung cells. Accordingly, we used lead chromate as a model particulate Cr(VI) compound and determined its cytotoxicity and genotoxicity in cultured human bronchial epithelial cells, using BEP2D cells as a model cell line. We found that lead chromate induced concentration-dependent cytotoxicity in BEP2D cells after a 24h exposure. Specifically, the relative survival was 78, 59, 53, 46 and 0% after exposure to 0.5, 1, 5, 10 and 50 microg/cm(2) lead chromate, respectively. Similarly, the amount of chromosome damage increased with concentration after 24h exposure to lead chromate. Specifically, 0.5, 1, 5 and 10 microg/cm(2) damaged 10, 13, 20 and 28% of metaphase cells with the total amount of damage reaching 11, 15, 24 and 36 aberrations per 100 metaphases, respectively. Lead chromate (50 microg/cm(2) lead chromate) induced profound cell cycle delay and no metaphases were found. In addition we investigated the effects of soluble hexavalent chromium, sodium chromate, in this cell line. We found that 1, 2.5, 5 and 10 microM sodium chromate induced 66, 35, 0 and 0% relative survival, respectively. The amount of chromosome damage increased with concentration after 24h exposure to sodium chromate. Specifically, 1, 2.5 and 5 microM damaged 25, 34 and 41% of metaphase cells with the total amount of damage reaching 33, 59 and 70 aberrations per 100 metaphases, respectively. Ten micromolar sodium chromate induced profound cell cycle delay and no metaphases were found. Overall the data clearly indicate that hexavalent Cr(VI) is cytotoxic and genotoxic to human lung epithelial cells.
Asunto(s)
Cromo/farmacología , Células Epiteliales/efectos de los fármacos , Pulmón/efectos de los fármacos , Carcinógenos Ambientales/química , Carcinógenos Ambientales/farmacología , Ciclo Celular/efectos de los fármacos , Línea Celular , Supervivencia Celular/efectos de los fármacos , Cromatos/farmacología , Cromo/química , Aberraciones Cromosómicas/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Células Epiteliales/citología , Células Epiteliales/metabolismo , Humanos , Plomo/farmacología , Pulmón/citología , Pulmón/metabolismo , Metafase/efectos de los fármacos , Tamaño de la Partícula , Compuestos de Sodio/farmacología , SolubilidadRESUMEN
Hexavalent chromium (Cr(VI)) is a well-designated human lung carcinogen, with solubility playing an important role in its carcinogenic potential. Although it is known that particulate or water-insoluble Cr(VI) compounds are more potent than the soluble species of this metal, the mechanisms of action are not fully elucidated. In this study, we investigated the hypothesis that the difference in potency between particulate and soluble Cr(VI) is due to more chronic exposures with particulate chromate because it can deposit and persist in the lungs while soluble chromate is rapidly cleared. Chronic exposure to both insoluble lead chromate and soluble sodium chromate induced a concentration and time-dependent increase in intracellular Cr ion concentrations in cultured human lung fibroblasts. Intracellular Pb levels after chronic exposure to lead chromate increased in a concentration-dependent manner but did not increase with longer exposure times up to 72 h. We also investigated the effects of chronic exposure to Cr(VI) on clastogenicity and found that chronic exposure to lead chromate induces persistent or increasing chromosome damage. Specifically, exposure to 0.5 microg/cm(2) lead chromate for 24, 48 and 72 h induced 23, 23 and 27% damaged metaphases, respectively. Contrary to lead chromate, the amount of chromosome damage after chronic exposure to sodium chromate decreased with time. For example, cells exposed to 1 microM sodium chromate for 24, 48 and 72 h induced 23, 13 and 17% damaged metaphases, respectively. Our data suggest a possible mechanism for the observed potency difference between soluble and insoluble Cr(VI) compounds is that chronic exposure to particulate Cr(VI) induces persistent chromosome damage and chromosome instability while chromosome damage is repaired with chronic exposure to soluble Cr(VI).
Asunto(s)
Cromo/farmacología , Pulmón/efectos de los fármacos , Carcinógenos Ambientales/química , Carcinógenos Ambientales/farmacología , Línea Celular , Cromatos/farmacología , Cromo/química , Aberraciones Cromosómicas/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Humanos , Plomo/farmacología , Pulmón/citología , Pulmón/metabolismo , Metafase/efectos de los fármacos , Tamaño de la Partícula , Compuestos de Sodio/farmacología , Solubilidad , Factores de TiempoRESUMEN
Cobalt is a toxic metal used in various industrial applications leading to adverse lung effects by inhalation. Cobalt is considered a possible human carcinogen with the lung being a primary target. However, few studies have considered cobalt-induced toxicity in human lung cells, especially normal lung epithelial cells. Therefore, in this study, we sought to determine the cytotoxicity and genotoxicity of particulate and soluble cobalt in normal primary human lung epithelial cells. Cobalt oxide and cobalt chloride were used as representative particulate and soluble cobalt compounds, respectively. Exposure to both particulate and soluble cobalt induced a concentration-dependent increase in cytotoxicity, genotoxicity, and intracellular cobalt ion levels. Based on intracellular cobalt ion levels, we found that soluble and particulate cobalt induced similar cytotoxicity while soluble cobalt was more genotoxic than particulate cobalt. These data indicate that cobalt compounds are cytotoxic and genotoxic to human lung epithelial cells.
Asunto(s)
Cobalto/toxicidad , Citotoxinas/toxicidad , Células Epiteliales/efectos de los fármacos , Pulmón/citología , Pulmón/efectos de los fármacos , Mutágenos/toxicidad , Óxidos/toxicidad , Línea Celular , Aberraciones Cromosómicas/inducido químicamente , Daño del ADN/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Humanos , Pulmón/metabolismo , Pulmón/patología , Tamaño de la Partícula , SolubilidadRESUMEN
BRCA1/2 proteins function in homologous recombination (HR)-mediated DNA repair and cooperate with Fanconi anemia (FA) proteins to maintain genomic integrity through replication fork stabilization. Loss of BRCA1/2 proteins results in DNA repair deficiency and replicative stress, leading to genomic instability and enhanced sensitivity to DNA-damaging agents. Recent studies have shown that BRCA1/2-deficient tumors upregulate Polθ-mediated alternative end-joining (alt-EJ) repair as a survival mechanism. Whether other mechanisms maintain genomic integrity upon loss of BRCA1/2 proteins is currently unknown. Here we show that BRCA1/2-deficient tumors also upregulate FANCD2 activity. FANCD2 is required for fork protection and fork restart in BRCA1/2-deficient tumors. Moreover, FANCD2 promotes Polθ recruitment at sites of damage and alt-EJ repair. Finally, loss of FANCD2 in BRCA1/2-deficient tumors enhances cell death. These results reveal a synthetic lethal relationship between FANCD2 and BRCA1/2, and they identify FANCD2 as a central player orchestrating DNA repair pathway choice at the replication fork.
Asunto(s)
Proteína BRCA1/deficiencia , Proteína BRCA2/deficiencia , Reparación del ADN por Unión de Extremidades , Replicación del ADN , Proteína del Grupo de Complementación D2 de la Anemia de Fanconi/metabolismo , Neoplasias/genética , Neoplasias/patología , Animales , Proteína BRCA1/metabolismo , Proteína BRCA2/metabolismo , Proteínas Portadoras/metabolismo , Línea Celular Tumoral , Supervivencia Celular , Reparación del ADN por Unión de Extremidades/genética , Replicación del ADN/genética , ADN Polimerasa Dirigida por ADN/metabolismo , Endodesoxirribonucleasas , Inestabilidad Genómica , Humanos , Ratones Desnudos , Mutación/genética , Proteínas Nucleares/metabolismo , Poli(ADP-Ribosa) Polimerasas/metabolismo , Ubiquitinación , Regulación hacia Arriba/genética , ADN Polimerasa thetaRESUMEN
Hexavalent chromium (Cr(VI)) is a widespread environmental contaminant and a known human carcinogen, generally causing bronchial cancer. Recent studies have shown that the particulate forms of Cr(VI) are the potent carcinogens. Particulate Cr(VI) is known to induce a spectrum of DNA damage such as DNA single strand breaks, Cr-DNA adducts, DNA-protein crosslinks and chromosomal aberrations. However, particulate Cr(VI)-induced DNA double strand breaks (DSBs) have not been reported. Thus, the aim of this study was to determine if particulate Cr(VI)-induces DSBs in human bronchial cells. Using the single cell gel electrophoresis assay (comet assay), showed that lead chromate-induced concentration dependent increases in DSBs with 0.1, 0.5, 1 and 5 microg/cm2 lead chromate inducing a 20, 50, 67 and 109% relative increase in the tail integrated intensity ratio, respectively. Sodium chromate at concentrations of 1, 2.5 and 5 microM induced 38, 78 and 107% relative increase in the tail integrated intensity ratio, respectively. We also show that genotoxic concentrations of lead chromate activate the ataxia telangiectasia mutated (ATM) protein, which is thought to play a central role in the early stages of DSB detection and controls cellular responses to this damage. The H2A.X protein becomes rapidly phosphorylated on residue serine 139 in cells when DSBs are introduced into the DNA by ionizing radiation. By using immunofluorescence, we found that lead chromate-induced concentration-dependent increases in phosphorylated H2A.X (r-H2A.X) foci formation with 0.1, 0.5, 1, 5 and 10 microg/cm2 lead chromate inducing a relative increase in the number of cells with r-H2A.X foci formation of 43, 51, 115 and 129%, respectively.