“…We therefore examined arsenite-induced mutagenesis in TK6 cells. Mutation in TK or HPRT genes can be positively selected using trifluorothymidine or 6-thioguanine, respectively, as each is present in TK6 cells in the haploid state (44,46,74). The TK mutation frequency (MF) increased 2.4-fold (P ϭ 0.007) in ATF4-deficient cells exposed to up to 50 M arsenite (Fig.…”
Arsenite is a human carcinogen causing skin, bladder, and lung tumors, but the cellular mechanisms underlying these effects remain unclear. We investigated expression of the essential base excision DNA repair enzyme apurinic endonuclease 1 (Ape1) in response to sodium arsenite. In mouse 10T 1 ⁄2 fibroblasts, Ape1 induction in response to arsenite occurred about equally at the mRNA, protein, and enzyme activity levels. Analysis of the APE1 promoter region revealed an AP-1/CREB binding site essential for arsenite-induced transcriptional activation in both mouse and human cells. Electrophoretic mobility shift assays indicated that an ATF4/c-Jun heterodimer was the responsible transcription factor. RNA interference targeting c-Jun or ATF4 eliminated arsenite-induced APE1 transcription. Suppression of Ape1 or ATF4 sensitized both mouse fibroblasts (10T 1 ⁄2) and human lymphoblastoid cells (TK6) to arsenite cytotoxicity. Expression of Ape1 from a transgene did not efficiently restore arsenite resistance in ATF4-depleted cells but did offset initial accumulation of abasic DNA damage following arsenite treatment. Mutagenesis by arsenite (at the TK and HPRT loci in TK6 cells) was observed only for ATF4-depleted cells, which was strongly offset by Ape1 expression from a transgene. Therefore, the ATF4-mediated up-regulation of Ape1 and other genes plays a key role against arsenite-mediated toxicity and mutagenesis.Epidemiological studies from Bangladesh, India (West Bengal), Taiwan, and South America indicate that arsenic ingestion, typically via contaminated drinking water, is associated with increased incidence of diverse human diseases, such as atherosclerosis, diabetes, and cancers of the skin, bladder, and lung (55). The arsenic found in drinking water is overwhelmingly one of the inorganic arsenic forms, mostly pentavalent arsenate and trivalent arsenite (55).Arsenite treatment has recently been introduced as therapy for acute promyelocytic leukemia (61). The pathological and therapeutic mechanisms of arsenic are actively debated, and DNA may be an important target for arsenic-related damage. For example, comet assays demonstrate that exposure to as little as 1 nM of sodium arsenite could rapidly (within 30 min) induce DNA base damage in human HeLa, neutrophilic NB4, and HL-60 cells (59, 69). For comparison, the reported mean blood arsenic concentration in people consuming highly contaminated water was 560 nM (49), while the mean plasma arsenite concentration for cancer treatment was 1,000 to 6,000 nM (60). However, many cell-based assays have failed to detect a significant increase in point mutations due to arsenite (55). One conclusion from these observations is that a strong cellular antimutagenesis mechanism counteracts the mutational potential of arsenite-induced DNA damage.Oxidized and other small base lesions are excised by DNA glycosylases, which channel damage into the base excision DNA repair (BER) pathway. In mammalian cells, glycosylase products are processed by the abasic (apurinic/apyrimidinic [AP]) end...
“…We therefore examined arsenite-induced mutagenesis in TK6 cells. Mutation in TK or HPRT genes can be positively selected using trifluorothymidine or 6-thioguanine, respectively, as each is present in TK6 cells in the haploid state (44,46,74). The TK mutation frequency (MF) increased 2.4-fold (P ϭ 0.007) in ATF4-deficient cells exposed to up to 50 M arsenite (Fig.…”
Arsenite is a human carcinogen causing skin, bladder, and lung tumors, but the cellular mechanisms underlying these effects remain unclear. We investigated expression of the essential base excision DNA repair enzyme apurinic endonuclease 1 (Ape1) in response to sodium arsenite. In mouse 10T 1 ⁄2 fibroblasts, Ape1 induction in response to arsenite occurred about equally at the mRNA, protein, and enzyme activity levels. Analysis of the APE1 promoter region revealed an AP-1/CREB binding site essential for arsenite-induced transcriptional activation in both mouse and human cells. Electrophoretic mobility shift assays indicated that an ATF4/c-Jun heterodimer was the responsible transcription factor. RNA interference targeting c-Jun or ATF4 eliminated arsenite-induced APE1 transcription. Suppression of Ape1 or ATF4 sensitized both mouse fibroblasts (10T 1 ⁄2) and human lymphoblastoid cells (TK6) to arsenite cytotoxicity. Expression of Ape1 from a transgene did not efficiently restore arsenite resistance in ATF4-depleted cells but did offset initial accumulation of abasic DNA damage following arsenite treatment. Mutagenesis by arsenite (at the TK and HPRT loci in TK6 cells) was observed only for ATF4-depleted cells, which was strongly offset by Ape1 expression from a transgene. Therefore, the ATF4-mediated up-regulation of Ape1 and other genes plays a key role against arsenite-mediated toxicity and mutagenesis.Epidemiological studies from Bangladesh, India (West Bengal), Taiwan, and South America indicate that arsenic ingestion, typically via contaminated drinking water, is associated with increased incidence of diverse human diseases, such as atherosclerosis, diabetes, and cancers of the skin, bladder, and lung (55). The arsenic found in drinking water is overwhelmingly one of the inorganic arsenic forms, mostly pentavalent arsenate and trivalent arsenite (55).Arsenite treatment has recently been introduced as therapy for acute promyelocytic leukemia (61). The pathological and therapeutic mechanisms of arsenic are actively debated, and DNA may be an important target for arsenic-related damage. For example, comet assays demonstrate that exposure to as little as 1 nM of sodium arsenite could rapidly (within 30 min) induce DNA base damage in human HeLa, neutrophilic NB4, and HL-60 cells (59, 69). For comparison, the reported mean blood arsenic concentration in people consuming highly contaminated water was 560 nM (49), while the mean plasma arsenite concentration for cancer treatment was 1,000 to 6,000 nM (60). However, many cell-based assays have failed to detect a significant increase in point mutations due to arsenite (55). One conclusion from these observations is that a strong cellular antimutagenesis mechanism counteracts the mutational potential of arsenite-induced DNA damage.Oxidized and other small base lesions are excised by DNA glycosylases, which channel damage into the base excision DNA repair (BER) pathway. In mammalian cells, glycosylase products are processed by the abasic (apurinic/apyrimidinic [AP]) end...
“…The expression of stem cell markers on day 21 was compared to values derived on day 7 in order to detect a possible kinetic of expression changes. The measured mode of cell death was selected to represent the dominating mechanism in cells of haematopoietic and epithelial origin, respectively: apoptosis in AHH-1 cells 21 and senescence in VH10 cells 22 . Figure 1 ( A ) Schematic representation of treatment scheme.…”
Evidence on the impact of chemotherapy on radiotherapy-induced second malignant neoplasms is controversial. We estimated how cisplatin modulates the in vitro response of two normal cell types to fractionated radiation. AHH-1 lymphoblasts and VH10 fibroblasts were irradiated at 1 Gy/fraction 5 and 3 times per week during 12 and 19 days, respectively, and simultaneously treated with 0.1, 0.2, 0.4, 0.8, 1.7 and 3.3 µM of cisplatin twice a week. Cell growth during treatment was monitored. Cell growth/cell death and endpoints related to accumulation of DNA damage and, thus, carcinogenesis, were studied up to 21 days post treatment in cells exposed to radiation and the lowest cisplatin doses. Radiation alone significantly reduced cell growth. The impact of cisplatin alone below 3.3 µM was minimal. Except the lowest dose of cisplatin in VH10 cells, cisplatin reduced the inhibitory effect of radiation on cell growth. Delayed cell death was highest in the combination groups while the accumulation of DNA damage did not reveal a clear pattern. In conclusion, fractionated, concomitant exposure to radiation and cisplatin reduces the inhibitory effect of radiation on cell proliferation of normal cells and does not potentiate delayed effects resulting from accumulation of DNA damage.
“…In animal or human cell cultures, anthracyclins induce chromosomal alterations rather than mutations (El-Mahdy Sayed Othman, 2000;Baumgartner et al, 2004), often through homologous recombination mechanisms (Lehmann et al, 2003). The altered cells then either engage their apoptotic programme or accumulate genetic alterations (Morris et al, 1995). Alkylating agents transfer alkyl groups to cellular constituents, inducing DNA adducts as well as DNA crosslinking that may lead to point mutations (Sanderson and Shield, 1996;Hunter et al, 2006).…”
In advanced breast cancers, TP53 mutation is highly predictive of complete response to high-dose epirubicin/cyclophosphamide chemotherapy. In these tumours with an altered control of genomic stability, accumulation of chemotherapy-induced genetic alterations may contribute to cell death and account for complete response. To explore the effects of chemotherapy on stability of the tumour genome, allelic profiles were obtained from microdissected tumour samples before and after chemotherapy in 29 unresponsive breast cancers (9 with TP53 mutation). Ninety-four per cent allelic profiles remained unchanged after treatment. Interestingly, 11 profiles (6%) showed important changes after treatment; allelic imbalances significantly increased (four cases) or decreased (seven cases) after chemotherapy in three distinct experiments, two of which using laser microdissected tumour cells. These genetic changes were not linked to the TP53 status, but one tumour showed complete disappearance of TP53-mutated cells in the residual tumour after treatment. Altogether, these observations carry important implications for the clonal evolution of breast cancers treated with DNA-damaging agents, as they point both to the importance of tumour heterogeneity and chemotherapydriven selection of subclones.
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