RESUMEN
The potential of clinical drug diclofenac sodium which is routinely used in clinics as non-steroid anti-inflammatory drugs opens a new insight in development of radioprotector. The drug has shown its potential radioprotective efficacy in clonogenic cell survival in Chinese hamster V79 cells with a DMF of 1.4. The pBR322 plasmid DNA gets damaged by radiation in which the supercoiled form gradually disappears with increasing radiation dose. Diclofenac sodium has shown its radioprotective potential by scavenging radiation induced free radicals which are depicted by its ability in restoring the fraction of supercoiled form of plasmid DNA back to normal. 250µM concentration of the drug provides GSSB yield of 3.65±0.5×10(-9)Da(-1)Gy(-1). This drug has shown to have a free radical scavenging activity because the radical cation with blue color is converted to the colorless neutral form by addition of diclofenac sodium in ABTS assay. Whole body survival study has shown it to protect 45.5% of C57BL/6 mice at a lethal irradiation dose of 9Gy. Therefore, this molecule offers a potential radioprotective ability besides being used routinely as analgesic and anti inflammatory compound.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacología , Diclofenaco/farmacología , Fibroblastos/efectos de los fármacos , Depuradores de Radicales Libres/farmacología , Protectores contra Radiación/farmacología , Animales , Línea Celular/efectos de los fármacos , Línea Celular/efectos de la radiación , Radioisótopos de Cobalto , Ensayo de Unidades Formadoras de Colonias , Cricetinae , Cricetulus , Relación Dosis-Respuesta en la Radiación , Evaluación Preclínica de Medicamentos , Fibroblastos/efectos de la radiación , Rayos gamma , Masculino , Ratones , Ratones Endogámicos C57BL , Estructura Molecular , Plásmidos/efectos de los fármacos , Plásmidos/efectos de la radiación , Traumatismos Experimentales por Radiación/etiología , Traumatismos Experimentales por Radiación/prevención & control , Tolerancia a Radiación/efectos de los fármacos , Especies Reactivas de Oxígeno/análisis , Irradiación Corporal Total/efectos adversosRESUMEN
The cobalt(III) complexes Et(4)N[Co(L(1))(2)] and [Co(L(2))(3)] [H(2)L(1) is 2,6-bis(N-(2-pyridyl)carbamoyl)pyridine and HL(2) is 2-(N-(2-pyridyl)carbamoyl)pyridine] were used as the building blocks for preparing a series of {M(2+)-Co(3+)-M(2+)} (where M is Zn, Cd, or Hg) and {Co(3+)-M(2+)} (where M is Zn or Cd) heterometallic complexes. All heterometallic complexes were characterized using a host of spectroscopic methods (IR, NMR, and UV/vis spectroscopy and mass spectrometry), elemental analysis, and conductivity measurements. One of the representative compounds, {Hg(2+)-Co(3+)-Hg(2+)}, was characterized crystallographically, and it was revealed that two Hg(II) ions are coordinated within the clefts created by the building block Et(4)N[Co(L(1))(2)]. The results of screening for anticancer activity against the human brain tumor U87 cell line and antibacterial activity against a range of resistant (Pseudomonas aeruginosa and Proteus vulgaris) as well as standard (Staphylococcus aureus SA 96, P. aeruginosa MTCC 1688, Klebsiella planticola MTCC 2272, and Escherichia coli T7) bacterial strains indicate promising activities. Notably, the observed activity was found to vary with the type of building block and the secondary metal ion present in the heterometallic complex. Treatment-induced cell death [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, MTT and macrocolony assay), growth inhibition, cytogenetic damage, cell cycle delay, and apoptosis were studied as the parameters for cellular response.
Asunto(s)
Antibacterianos/síntesis química , Antibacterianos/farmacología , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Antibacterianos/química , Antineoplásicos/química , Bacterias/efectos de los fármacos , Cadmio/química , Línea Celular , Supervivencia Celular/efectos de los fármacos , Complejos de Coordinación/síntesis química , Complejos de Coordinación/química , Complejos de Coordinación/farmacología , Cristalografía por Rayos X , Citometría de Flujo , Humanos , Mercurio/química , Modelos Moleculares , Zinc/químicaRESUMEN
Radiation induced skin wound/dermatitis is one of the common side effects of radiotherapy or interventional radiobiology. In order to combat impaired healing of radiation wounds, alternative therapy to use sildenafil citrate (SC) topical hydrogel as a therapeutic option was proposed that has known to enhance nitric oxide in wounds. Our aim was to develop a radiation induced skin wound model and to investigate the wound healing efficacy of 5% SC hydrogel formulation in Sprague-Dawley rats. In the present study, the radiation wound inducing dose was optimized using a multi-dose localized γ-radiation trail with 10-55Gy range (15Gy interval). Optimal irradiation dose for wound induction was selected based on radiation skin damage assessment criteria followed the relative change from <35Gy or>55Gy showed significant variation and median 45Gy γ-dose was selected for studying acute effects of radiation on wound healing. Significant (p<0.05) higher wound contraction (88±1.02%), skin damage reduction (81±0.82%), tensile strength (45±1.61%), nitric oxide and protein recovery (53±0.72%) at dermal level prove the wound healing efficacy of 5% SC hydrogel formulation as compared to Rad 45Gy control. In addition, the dose modifying factor (DMF) for SC hydrogel treatment was found to be 1.83 and 1.57 with respect to total wound area contraction and skin damage reduction. Skin histopathology in treated tissues showed improved granulation tissue formation, less inflammatory infiltrates and mature collagen fibres in the dermis. Thus, the modality could help to improve delayed wound healing in irradiated skin tissues.
Asunto(s)
Hidrogeles/farmacología , Inhibidores de Fosfodiesterasa 5/farmacología , Radiodermatitis , Repitelización/efectos de los fármacos , Citrato de Sildenafil/farmacología , Piel/efectos de los fármacos , Animales , Modelos Animales de Enfermedad , Rayos gamma/efectos adversos , Óxido Nítrico/metabolismo , Ratas , Piel/metabolismo , Piel/patología , Piel/efectos de la radiaciónRESUMEN
BACKGROUND & OBJECTIVE: Hippophae rhamnoides L. has been widely exploited for medicinal purposes and an extract of its whole berries coded as RH-3 has been found to render radioprotection. Effect of pre-irradiation treatment of up to 10 microg/ml RH-3 was studied in U 87 cells using MTT assay. This study aims at unraveling the mechanism of action of RH-3 in amelioration of radiation-induced cytotoxicity in vitro. METHODS: Most effective doses selected were studied further for the elucidation of radiomodifying properties of RH-3, especially with respect to early and late events of apoptosis. RESULTS: RH-3 at concentrations of 7.5 and 10 microg/ml (-15 min) were found most effective in protecting against 2 Gy induced cytotoxicity in terms of MTT reducing ability in U 87 cells. RH-3 was observed to mitigate radiation-induced cellular and mitochondrial free radicals. Mitochondrial membrane potential depletion (studied up to 12 h) was prevented by RH-3 pre-irradiation administration. It could also restore the level of antiapoptotic protein Bcl-2 at 24 and 48 h comparable to the control value. RH-3 also prevented radiation-induced increase in mitochondrial mass at 48 and 72 h post-treatment and the values were comparable to that of control cells. Annexin-V-FITC assay at 12 and 24 h time intervals indicated significant protection against radiation-induced apoptosis by RH-3 pre-irradiation treatment. INTERPRETATION & CONCLUSION: Our findings showed that probably RH-3 acts as an antioxidant preventing cellular and mitochondrial free radical generation that could contribute to its ability to inhibit radiation-induced apoptosis and cytotoxicity.
Asunto(s)
Hippophae , Plantas Medicinales , Protectores contra Radiación/farmacología , Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Línea Celular , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , Radicales Libres/metabolismo , Rayos gamma/efectos adversos , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Potencial de la Membrana Mitocondrial/efectos de la radiación , Extractos Vegetales/farmacologíaRESUMEN
Sildenafil Citrate (SC) is a US FDA approved drug, have been used to treat wounds due to their nitric oxide (NO) stimulating activity in the tissue. But, there are only a few studies about the topical effect of this drug on the healing of traumatic wounds. The purpose of the study is to develop topical SC hydrogel (SCH) and to investigate its dermal toxicity and wound healing efficacy in Sprague dawley rats. In the present study, hydrogel containing SC showed no change and stable with respect to pH, homogeneity, spreadability and effiecient encapsulation. SEM analysis represents the uniform texture of the SCH. Acute dermal toxicity of the SCH exhibited that the formulations are devoid of any toxic effects and safe to be used. Percentage of wound contraction, re-epithelization, tensile strength and biochemical parameters such as hydroxyproline, collagen, total protein and NO content at dermal level prove the wound healing efficacy of prepared SCH. In addition, histopathology confirmed that the SCH promoted re-epithelization, collagen synthesis, deposition and regeneration of skin appendages. Results demonstrated that SCH has no dermal toxicity and promoted wound healing. Thus, prepared SCH shows promising skin wound healing property against traumatic wounds.
Asunto(s)
Hidrogeles/administración & dosificación , Citrato de Sildenafil/administración & dosificación , Piel/efectos de los fármacos , Cicatrización de Heridas/efectos de los fármacos , Administración Tópica , Animales , Composición de Medicamentos , Femenino , Hidrogeles/química , Hidrogeles/toxicidad , Ratas , Ratas Sprague-Dawley , Citrato de Sildenafil/química , Citrato de Sildenafil/toxicidad , Piel/metabolismo , Piel/patología , Resultado del Tratamiento , Vasodilatadores/administración & dosificación , Vasodilatadores/química , Vasodilatadores/toxicidad , Cicatrización de Heridas/fisiologíaRESUMEN
Earlier studies have shown that 2-deoxy-D-glucose (2-DG), a glucose analogue and inhibitor of glycolytic ATP production selectively enhances radiation-induced damage in cancer cells by inhibiting the energy (ATP) dependent postirradiation DNA and cellular repair processes. A reduction in radiation induced cytogenetic damage has been reported in normal cells viz., peripheral blood lymphocytes and bone marrow cells. Since induction of apoptosis plays a major role in determining the radiosensitivity of some most sensitive normal cells including splenocytes and thymocytes, we investigated the effects of 2-DG on radiation induced apo tosis in these cells in vitro. Thymocytes and splenocytes isolated from normal Swiss albino mouse were irradiated with Co60 gamma-rays and analyzed for apoptosis at various post-irradiation times. 2-DG added at the time of irradiation was present till the termination of cultures. A time dependent, spontaneous apoptosis was evident in both the cell systems, with nearly 40% of the cells undergoing apoptosis at 12 hr of incubation. The dose response of radiation-induced apoptosis was essentially similar in both the cell systems and was dependent on the incubation time. More than 70% of the splenocytes and 60% of the thymocytes were apoptotic by 12 hr following an absorbed dose of 2 Gy. Presence of 2-DG marginally reduced the fraction of splenocytes undergoing apoptosis at all absorbed doses, while no change was observed in thymocytes. Presence of 2-DG did not significantly alter either the level or the rate of induction of spontaneous apoptosis in both these cell systems. These results are consistent with the earlier findings on radiation-induced cytogenetic damage in human PBL in vitro and mouse bone marrow cells and lend further support to the proposition that 2-DG does not enhance radiation damage in normal cells, while radiosensitizing the tumors and hence is an ideal adjuvant in the radiotherapy of tumors.
Asunto(s)
Antimetabolitos/farmacología , Apoptosis , Desoxiglucosa/farmacología , Rayos gamma , Bazo/citología , Timo/citología , Animales , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Células Cultivadas , ADN/metabolismo , Relación Dosis-Respuesta en la Radiación , Femenino , Ratones , Bazo/efectos de los fármacos , Bazo/efectos de la radiación , Timo/efectos de los fármacos , Timo/efectos de la radiaciónRESUMEN
Protection of γ-ray-induced injury in hematopoietic and gastrointestinal (GI) systems is the rationale behind developing radioprotectors. The objective of this study, therefore, was to investigate the radioprotective efficacy and mechanisms underlying sesamol in amelioration of γ-ray-induced hematopoietic and GI injury in mice. C57BL/6 male mice were pre-treated with a single dose (100 or 50 mg/kg, 30 min prior) of sesamol through the intraperitoneal route and exposed to LD50/30 (7.5 Gy) and sublethal (5 Gy) dose of γ-radiation. Thirty-day survival against 7.5 Gy was monitored. Sesamol (100 mg/kg) pre-treatment reduced radiation-induced mortality and resulted survival of about 100% against 7.5 Gy of γ-irradiation. Whole-body irradiation drastically depleted hematopoietic progenitor stem cells in bone marrow, B cells, T cell subpopulations, and splenocyte proliferation in the spleen on day 4, which were significantly protected in sesamol pre-treated mice. This was associated with a decrease of radiation-induced micronuclei (MN) and apoptosis in bone marrow and spleen, respectively. Sesamol pre-treatment inhibited lipid peroxidation, translocation of gut bacteria to spleen, liver, and kidney, and enhanced regeneration of crypt cells in the GI system. In addition, sesamol pre-treatment reduced the radiation-induced pattern of expression of p53 and Bax apoptotic proteins in the bone marrow, spleen, and GI. This reduction in apoptotic proteins was associated with the increased anti-apoptotic-Bcl-x and PCNA proteins. Further, assessment of antioxidant capacity using ABTS and DPPH assays revealed that sesamol treatment alleviated total antioxidant capacity in spleen and GI tissue. In conclusion, the results of the present study suggested that sesamol as a single prophylactic dose protects hematopoietic and GI systems against γ-radiation-induced injury in mice.
Asunto(s)
Antioxidantes/uso terapéutico , Benzodioxoles/uso terapéutico , Fenoles/uso terapéutico , Protectores contra Radiación/uso terapéutico , Animales , Antioxidantes/farmacología , Apoptosis , Benzodioxoles/farmacología , Rayos gamma , Masculino , Ratones , Ratones Endogámicos C57BL , Fenoles/farmacología , Protectores contra Radiación/farmacologíaRESUMEN
PURPOSE: Two deoxy-D-glucose (2-DG), an inhibitor of glucose transport and glycolysis, has been shown to differentially inhibit the repair of radiation damage in cancer cells by reducing the flow of metabolic energy. Since hematoporphyrin derivatives (Hpd) inhibit certain enzymes of the respiratory metabolism, resulting in an increase in the glucose usage and glycolysis, Hpd could possibly enhance the energy-linked radiosensitizing effects of 2-DG in cancer cells. The purpose of the present work was to verify this suggestion. METHODS AND MATERIALS: Two human tumor cell lines (cerebral glioma, BMG-1 and squamous cell carcinoma, 4197) and a murine tumor cell line (Ehrlich ascites tumor [EAT], F-15) in vitro were investigated. A commercially available preparation of Hpd, Photosan-3 (PS-3) was used in the present studies. Cells incubated with 0-10 microg/ml PS-3 for 0-24 h before irradiation were exposed to 2.5 Gy of Co-60 gamma rays and maintained under liquid holding conditions for 1-4 h to facilitate repair. 2-DG (0-5 mM) added at the time of irradiation was present during the liquid holding. Radiation-induced cytogenetic damage (micronuclei formation) and cell death (macrocolony assay) were analyzed as parameters of radiation response. Effects of these radiosensitizers on glucose usage and glycolysis were also studied by measuring the glucose consumption and lactate production using enzymatic assays. RESULTS: The glucose consumption and lactate production of BMG-1 cells (0.83 and 1.43 pmole/cell/h) were twofold higher than in the 4197 cells (0.38 and 0.63 pmole/cell/h). Presence of PS-3 (10 microg/ml) enhanced the rate of glycolysis (glucose consumption and lactate production) in these cells by 35% to 65%, which was reduced by 20% to 40% in the presence of 5 mM 2-DG. In exponentially growing BMG-1 and EAT cells, presence of 2-DG (5 mM; equimolar with glucose) for 4 hours after irradiation increased the radiation-induced micronuclei formation and cell death by nearly 40%, whereas no significant effects could be observed in 4197 cells. In EAT cells, radiation was also observed to induce apoptotic death, which was significantly increased in the presence of the combination (PS-3 + 2-DG). The combination (PS-3 + 2-DG) enhanced the radiation damage in all three cell systems by 60-100%. Furthermore, the radiosensitizing effects of the combination (PS-3 + 2-DG) were higher at pH 6.7 as compared to pH 7. 4. In the plateau phase, presence of 2-DG alone did not significantly influence the radiation response of either BMG-1 or of 4197 cells, whereas in combination with PS-3, 2-DG enhanced the radiation damage in both these cell lines by 40% to 50%. Furthermore, in BMG-1 cells, the effects of 2-DG were observed to be reversible to a very great extent, while that of the combination were mostly irreversible. CONCLUSION: The hematoporphyrin derivative PS-3 enhances the radiosensitizing effects of 2-DG in cancer cells, possibly by further reducing the energy supply leading to an irreversible inhibition of DNA repair, and increased cytogenetic damage and cell death. Since both these compounds have been used in clinical practice, further studies to investigate their use in improving radiotherapy of tumors are warranted.
Asunto(s)
Antimetabolitos/farmacología , Desoxiglucosa/farmacología , Neoplasias/radioterapia , Fármacos Fotosensibilizantes/farmacología , Fármacos Sensibilizantes a Radiaciones/farmacología , Animales , Carcinoma de Ehrlich/metabolismo , Carcinoma de Ehrlich/radioterapia , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/radioterapia , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Glucosa/metabolismo , Glucólisis , Hematoporfirinas , Humanos , Concentración de Iones de Hidrógeno , Ratones , Pruebas de Micronúcleos , Neoplasias/metabolismo , Radiobiología , Esferoides Celulares/efectos de los fármacos , Esferoides Celulares/metabolismo , Esferoides Celulares/efectos de la radiación , Células Tumorales Cultivadas/efectos de la radiación , Ensayo de Tumor de Célula MadreRESUMEN
PURPOSE: The glucose analog and glycolytic inhibitor, 2-deoxy-D-glucose (2-DG), has been shown to differentially enhance the radiation damage in tumor cells by inhibiting the postirradiation repair processes. The present study was undertaken to examine the relationship between 2-DG-induced modification of energy metabolism and cellular radioresponses and to identify the most relevant parameter(s) for predicting the tumor response to the combined treatment of radiation + 2-DG. METHODS AND MATERIALS: Six human tumor cell lines (glioma: BMG-1 and U-87, squamous cell carcinoma: 4451 and 4197, and melanoma: MeWo and Be-11) were investigated. Cells were exposed to 2 Gy of Co-60 gamma-rays or 250 kVP X-rays and maintained under liquid-holding conditions 2-4 h to facilitate repair. 2-DG (5 mM, equimolar with glucose) that was added at the time of irradiation was present during the liquid holding. Glucose utilization, lactate production (enzymatic assays), and adenine nucleotides (high performance liquid chromatography and capillary isotachophoresis) were investigated as parameters of energy metabolism. Induction and repair of DNA damage (comet assay), cytogenetic damage (micronuclei formation), and cell death (macrocolony assay) were analyzed as parameters of radiation response. RESULTS: The glucose consumption and lactate production of glioma cell lines (BMG-1 and U-87) were nearly 2-fold higher than the squamous carcinoma cell lines (4197 and 4451). The ATP content varied from 3.0 to 6.5 femto moles/cell among these lines, whereas the energy charge (0.86-0.90) did not show much variation. Presence of 2-DG inhibited the rate of glucose usage and glycolysis by 30-40% in glioma cell lines and by 15-20% in squamous carcinoma lines, while ATP levels reduced by nearly 40% in all the four cell lines. ATP:ADP ratios decreased to a greater extent ( approximately 40%) in glioma cells than in squamous carcinoma 4451 and MeWo cells; in contrast, presence of 2-DG reduced ADP:AMP ratios by 3-fold in the squamous carcinoma 4451, whereas an increase was noted in the glioma cell line BMG-1. 2-DG significantly reduced the initial rates of DNA repair in all cells, resulting in an excess residual damage after 2 h of repair in BMG-1, U-87, and 4451 cell lines, whereas no significant differences could be observed in the other cell lines. Recovery from potentially lethal damage was also significantly inhibited in BMG-1 cells. 2-DG increased the radiation-induced micronuclei formation in the melanoma line (MeWo) by nearly 60%, while a moderate (25-40%) increase was observed in the glioma cell lines (BMG-1 and U-87). Presence of 2-DG during liquid holding (4 h) enhanced the radiation-induced cell death by nearly 40% in both the glioma cell lines, while significant effects were not observed in others. CONCLUSIONS: The modifications in energetics and radiation responses by 2-DG vary considerably among different human tumor cell lines, and the relationships between energy metabolism and various radiobiologic parameters are complex in nature. The 2-DG-induced modification of radiation response does not strictly correlate with changes in the levels of ATP. However, a significant enhancement of the radiation damage by 2-DG was observed in cells with high rates of glucose usage and glycolysis, which appear to be the two most important factors determining the tumor response to the combined treatment of 2-DG + radiation therapy.
Asunto(s)
Desoxiglucosa/farmacología , Metabolismo Energético/efectos de los fármacos , Neoplasias/metabolismo , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/radioterapia , Daño del ADN , Reparación del ADN/efectos de la radiación , Metabolismo Energético/efectos de la radiación , Glioma/metabolismo , Glioma/radioterapia , Glucosa/metabolismo , Humanos , Ácido Láctico/metabolismo , Melanoma/metabolismo , Melanoma/radioterapia , Pruebas de Micronúcleos , Neoplasias/radioterapia , Dosis de Radiación , Radiobiología , Células Tumorales Cultivadas/metabolismo , Células Tumorales Cultivadas/efectos de la radiaciónRESUMEN
PURPOSE: Lepidopteran insect cells are known to exhibit very high radioresistance. Although very effective DNA excision-repair has been proposed as a contributing factor, a detailed understanding of insect cell radiation responses has not yet been obtained. Therefore, the study was carried out to understand the in vitro radiation responses of Sf9 lepidopteran cells. MATERIALS AND METHODS: Exponentially growing asynchronous Sf9 cells (derived from ovaries of Spodoptera frugiperda) were exposed to gamma-radiation doses of 2-200 Gy. Cell survival, growth inhibition, cell cycle progression delay, alterations in cell morphology as well as induction of DNA damage, micronuclei and apoptosis were studied at various post-irradiation time intervals. RESULTS: Biphasic survival response curves were obtained with D0 rising from 20 Gy (at doses < or = 60 Gy) to 85 Gy (between 60 and 200 Gy), corroborating earlier reports on lepidopteran cells. An additional downward deviation at 2 Gy indicated a hypersensitive response. Dose-dependent growth inhibition with a transient G2 delay starting 12 h and extending up to 48-96 h was observed at doses of 10-200 Gy, while a brief G1/S transition delay was observed only at higher doses (> or = 100 Gy). Significant DNA damage was detected only at 20 Gy and higher doses, in contrast with human cells that showed similar damage at 2 Gy. Interestingly, micronuclei were not induced at any of the doses tested, although spontaneous micronucleation was evident in <1% of cells. Lack of micronucleus induction even at doses that induced significant DNA damage and a transient G2 block (20-50 Gy) strongly indicated a role of holocentric lepidopteran chromosomes. Apoptosis was detected only in a small proportion of cells (3%) exposed to 200 Gy, and cell/nucleus size and granularity increased by 72-96 h post-irradiation in a dose-dependent manner. Sf9 nucleoids extracted at 2 M NaCl showed higher compactness than the nucleoids prepared from human cells. CONCLUSIONS: It is clearly shown that lepidopteran cells are highly resistant to the induction of DNA damage and micronuclei, and display very low induction of apoptosis at doses up to 200 Gy. While the lack of micronucleus induction seems to be primarily due to the holocentric nature of their chromosomes, certain unique signalling pathways might be responsible for the low induction of apoptosis. Factors causing protection of Sf9 cellular DNA from radiation-induced damage are presently being investigated.
Asunto(s)
ADN/efectos de la radiación , Tolerancia a Radiación/fisiología , Spodoptera/fisiología , Spodoptera/efectos de la radiación , Animales , Apoptosis/efectos de la radiación , División Celular/fisiología , Línea Celular , Tamaño de la Célula/fisiología , Supervivencia Celular/efectos de la radiación , Daño del ADN , Relación Dosis-Respuesta en la Radiación , Humanos , Micronúcleos con Defecto Cromosómico/efectos de la radiación , Dosis de Radiación , Spodoptera/citologíaRESUMEN
The ability of various acetoxy derivatives of 4-methylcoumarins to inhibit the genotoxic changes due to aflatoxin B(1) (AFB(1)) is reported here. Several 4-methylcoumarins (test compounds), such as 7,8-diacetoxy-4-methylcoumarin (DAMC), monoacetoxy-4-methylcoumarin (MAC), 5-N-acetyl-6-acetoxy-4-methylcoumarin (NAMC) and 7,8-dihydroxy-4-methylcoumarin (DHMC) were separately administered intraperitoneally (i.p.) to male wistar rats followed by AFB(1) administration i.p. or intratracheally (i.t.) (2-8 mg/kg b.wt.) and another dose of the test compound. The animals were sacrificed 26h after AFB(1) administration. From animals receiving AFB(1) i.p., bone marrow (BM) cells were isolated and stained with Mayer's haematoxylin and eosin. Micronuclei (MN) in BM were scored by light microscopy. From animals receiving AFB(1) i.t., bronchoalveolar lavage (BAL) was obtained, lung cells (LG) were isolated and stained with fluorochrome 6-diamidino-2-phenylindole (DAPI) for the analysis of MN, apoptotic bodies (AP) and cell cycle variations. Rats were separately treated with the vehicle DMSO to serve as the proper control. AFB(1) caused significant dose-dependent induction of MN in BM as well as LG. AP were observed in LG of rats receiving AFB(1) and was found to correlate with MN induction. DAMC injection caused significant decrease in AP due to AFB(1) in LG and MN in both BM and LG. The effectiveness of MAC was approximately half that of DAMC, thereby indicating that number of acetoxy groups on the coumarin molecule determine the efficacy. The fact that NAMC had no effect either on MN or AP indicate that neither acetoxy group at C-6 nor the N-acetyl group at C-5 facilitate the transfer of acetyl group to P-450 required for inhibition of AFB(1)-epoxidation. DHMC, the deacetylated product of DAMC had no normalizing effect on the induction of MN and AP. These findings confirm our earlier hypothesis that DAMC-mediated acetylation of microsomal P-450 (catalysing epoxidation of AFB(1)) through the action of microsomal transacetylase is responsible for the protective action of DAMC. The relative number and position of acetoxy groups on the coumarin nucleus determine the specificity to the transacetylase necessary for the chemopreventive action.
Asunto(s)
Aflatoxina B1/toxicidad , Antimutagênicos/farmacología , Cumarinas/farmacología , Mutágenos/toxicidad , Acetilación , Animales , Apoptosis , Células de la Médula Ósea/efectos de los fármacos , Sistema Enzimático del Citocromo P-450/metabolismo , Pulmón/citología , Pulmón/efectos de los fármacos , Masculino , Pruebas de Micronúcleos , Ratas , Ratas Wistar , Relación Estructura-ActividadRESUMEN
In the present studies, the role of oxidative stress in radiosensitization by a combination of 2-DG and 6-aminonicotinamide (6-AN) was examined in a human glioma cell line (BMG-1: wild type p53). Presence of 2-DG or 6-AN for 4 hr after irradiation (gamma ray 2.5 Gy) significantly enhanced the radiation-induced cell death by 18% and the combination (2-DG + 6-AN) enhanced the cell death by 35%. Neither 2-DG nor 6-AN had any further significant effect on the glutathione levels in irradiated cells. However, the combination (2-DG + 6-AN) caused a significant decrease in GSH content, increase in GSSG levels, and enhanced the superoxide radical generation under these conditions. The enhanced cell death caused by the combination (2-DG + 6-AN) mainly resulted by the process of apoptosis as revealed by annexin V binding and was associated with elevated levels of Cyclin B1. However, no significant change was observed in the levels of Bcl-2. Thus, for the first time, our results have demonstrated that the radiosensitizing effects of these modifiers could also be mediated through alterations in the oxidative stress besides energy limited inhibition of repair and recovery processes.
Asunto(s)
6-Aminonicotinamida/farmacología , Desoxiglucosa/farmacología , Estrés Oxidativo , Fármacos Sensibilizantes a Radiaciones/farmacología , 6-Aminonicotinamida/administración & dosificación , Línea Celular Tumoral , Desoxiglucosa/administración & dosificación , Humanos , Fármacos Sensibilizantes a Radiaciones/administración & dosificaciónRESUMEN
In the present studies, effects of glucose analogue, 2-deoxy-D-glucose (2-DG) on radiation-induced cell cycle perturbations were investigated in human tumor cell lines. In unirradiated cells, the levels of cyclin B1 in G2 phase were significantly higher in both the glioma cell lines as compared to squamous carcinoma cells. Upon irradiation with Co60 gamma-rays (2 Gy), the cyclin B1 levels were reduced in U87 cells, while no significant changes could be observed in other cell lines, which correlated well with the transient G2 delay observed under these conditions by the BrdU pulse chase measurements. 2-DG (5 mM, 2 hr) induced accumulation of cells in the G2 phase and a time-dependent increase in the levels of cyclin B1 in both the glioma cell lines, while significant changes could not be observed in any of the squamous carcinoma cell lines. 2-DG enhanced the cyclin B1 level further in all the cell lines following irradiation, albeit to different extents. Interestingly, an increase in the unscheduled expression of B1 levels in G1 phase 48 hr after irradiation was observed in all the cell lines investigated. 2-DG also increased the levels of cyclin D1 at 24 hr in BMG-1 cell line. These observations imply that 2-DG-induced alterations in the cell cycle progression are partly responsible for its radiomodifying effects.
Asunto(s)
Ciclo Celular/efectos de los fármacos , Ciclo Celular/efectos de la radiación , Desoxiglucosa/farmacología , Carcinoma de Células Escamosas/patología , Línea Celular Tumoral , Glioma/patología , HumanosRESUMEN
Radioprotective effects of bisbenzimidole derived DNA ligands Hoechst-33342 (H-342) and Hoechst-33258 (H-258) have been investigated in whole body irradiated stain-A and Balb/c mice (Co-60 Gamma-ray, absorbed doses of 2.5 to 10 Gy delivered at dose rates of 0.01 to 0.50 Gy/min). Biodistribution of Hoechst dyes (2 or 5 mg/kg, body wt., i.v.) and their effects on cell cycle kinetics in bone marrow were studied by flow cytometry. Protection against radiation-induced chromosomal aberrations, micronuclei formation, alterations in DNA content dispersion, inhibition of erythropoiesis and animal lethality were investigated. Significant amount of DNA bound Hoechst could be observed in liver, intestine, kidney and brain for more than 14 days after its administration, while in the bone marrow cells, a reduction in the bound Hoechst was noticed after 7 days. H-342 significantly reduced the radiation-induced chromosome aberrations mainly due to a decrease in the frequency of acentrics (nearly 30%), while a marginal decrease (10%) in the dicentrics was observed at all the dose rates studied. Both H-342 and H-258 reduced the radiation-induced micronuclei formation in a dose dependent manner (2-10 mg/kg body wt.) and this protective effect was observed up to 6 days after the administration. Neither of the two compounds induced any cytogenetic damage in the bone marrow cells of unirradiated animals nor induced tumours at the doses used here (< 5 mg/kg, body wt. i.v.). Reduction in cytogenetic damage of bone marrow cells led to a faster recovery of erythropoesis as observed by increased PCE/NCE ratio in the peripheral blood erythrocytes of the animals which received Hoechst before irradiation. H-258 (5 mg/kg body wt.) given 18 hr before irradiation reduced radiation-induced animal death (5-9 Gy), while no significant effect was observed at higher doses (10 Gy). However, H-342, which has a higher cell permeability, even at a lower dose (2 mg/kg body wt.) showed significant protection at 10 Gy. The protective effects could be enhanced further, by combining these DNA binding agents with the glucose analogue, 2-deoxy-D-glucose (2-DG) which has been shown earlier to protect bone marrow cells against radiation damage.
Asunto(s)
Bencimidazoles/uso terapéutico , Bisbenzimidazol/uso terapéutico , ADN/efectos de los fármacos , Traumatismos Experimentales por Radiación/prevención & control , Protectores contra Radiación/uso terapéutico , Animales , Bencimidazoles/farmacología , Bisbenzimidazol/farmacología , Ratones , Ratones Endogámicos BALB C , Irradiación Corporal TotalRESUMEN
To facilitate efficient handling of large samples, an attempt towards networking of laboratories in India for biological dosimetry was carried out. Human peripheral blood samples were exposed to (60)Co γ-radiation for ten different doses (0-5Gy) at a dose rate of 0.7 and 2Gy/min. The chromosomal aberrations (CA) were scored in Giemsa-stained and fluorescence in-situ hybridization with centromere-specific probes. No significant difference (p>0.05) was observed in the CA yield for given doses except 4 and 5Gy, between the laboratories, among the scorers and also staining methods adapted suggest the reliability and validates the inter-lab comparisons exercise for triage applications.
Asunto(s)
Bioensayo/métodos , Centrómero/genética , Centrómero/efectos de la radiación , Aberraciones Cromosómicas/efectos de la radiación , Linfocitos/fisiología , Linfocitos/efectos de la radiación , Radiometría/métodos , Colorantes Azulados/química , Células Cultivadas , Relación Dosis-Respuesta en la Radiación , Humanos , Hibridación Fluorescente in Situ/métodos , India , Dosis de Radiación , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
Exposure to ionizing radiations, whether medical, occupational or accidental, leads to deleterious biological consequences like mortality or carcinogenesis. It is considered that no dose of ionizing radiation exposure is safe. However, once the accurate absorbed dose is estimated, one can be given appropriate medical care and the severe consequences can be minimized. Though several accurate physical dose estimation modalities exist, it is essential to estimate the absorbed dose in biological system taking into account the individual variation in radiation response, so as to plan suitable medical care. Over the last several decades, lots of efforts have been taken to design a rapid and easy biological dosimeter requiring minimum invasive procedures. The metaphase chromosomal aberration assay in human lymphocytes, though is labor intensive and requires skilled individuals, still remains the gold standard for radiation biodosimetry. The current review aims at discussing the human lymphocyte metaphase chromosomal aberration assay and recent developments involving the application of molecular cytogenetic approaches and other technological advancements to make the assay more authentic and simple to use even in the events of mass radiation casualties.
RESUMEN
The development of an approach based on the energy-linked modification of DNA repair and cellular recovery processes using 2-deoxy-D-glucose (2-DG; inhibitor of glycolytic ATP production) has shown promising results in a number of model systems of cancer. Following encouraging results on the tolerance and toxicity (acute as well as late effects) of the combination (2-DG and hypofractionated radiotherapy) in Phase I and II clinical trials, its efficacy is currently under evaluation in Phase III clinical trials for glioma patients. Since heterogeneous physiologic and metabolic status in tumors as well as host-tumor interactions influence the local tumor control, which coupled with systemic disturbances could determine the cure (long-term tumor free survival), investigations on the in vivo responses of tumors to the combined treatment have received considerable attention. This communication provides a brief overview on the in vivo studies related to radio- and chemosensitization of tumors by 2-DG, besides the normal tissue toxicity induced by the combined treatment of 2-DG and radiation or chemotherapeutic drugs.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Desoxiglucosa/administración & dosificación , Neoplasias Experimentales/metabolismo , Neoplasias Experimentales/terapia , Radioterapia/métodos , Animales , Terapia Combinada , Glucosa/metabolismoRESUMEN
Normal tissue toxicity is one of the major limiting factors in cancer therapy. Damage to normal tissues and critical organs restricts the use of higher therapeutic doses thereby compromising the efficacy. The glucose analog 2-deoxy-D-glucose (2-DG), an inhibitor of glycolytic ATP production has been shown to enhance radiation- and chemotherapeutic drug-induced damage in a number of cancer cells under in vitro and in vivo conditions while sparing or protecting normal cells. This review summarizes current understanding on the protection of normal cells and tissues against radiation- and chemotherapeutic drug-induced damage by 2-DG that makes this glucose analog an ideal adjuvant in cancer therapy.
Asunto(s)
Desoxiglucosa/farmacología , Neoplasias/metabolismo , Neoplasias/terapia , Tolerancia a Radiación , Fármacos Sensibilizantes a Radiaciones/farmacología , Animales , HumanosRESUMEN
The AT specific minor grove DNA binding ligands bisbenzimidazole derivatives like hoechst-33342 and hoechst-33258 which scavenge free radicals and stabilize macromolecular structure have been shown to afford radioprotection by reducing the induction of DNA damage. However, their ability to inhibit topoisomerases I & II, which play important roles in damage response pathways including DNA repair can enhance radiation damage under certain conditions. Since pool sizes of the topoisomerases differ not only between normal and tumor cells, but also among different tumors, it is anticipated that radiosensitization by hoechst-33342 can vary among tumors. The present studies were, therefore, undertaken to verify this proposition in human glioma (BMG-1 &U-87) and squamous carcinoma (4197 &4451) cell lines which differ in their biological behavior (ploidy, p53, cyclins, bcl, bax etc). Isotoxic concentrations of hoechst-33342 (IC50 i.e producing 50% cell kill) administered immediately following irradiation resulted in the radiosensitization of all cell lines, with a 4&7 fold increase in the cell death (loss of clonogenic cell survival) in U-87&BMG-1 and a 3 fold increase in 4197 &4451 cells. Growth inhibition and increase in cytogenetic damage (micronuclei formation) as well as delayed apoptosis observed under these conditions corroborated well with the enhanced cell death. The ligand induced a significant cell cycle delay, particularly in the late S and G2 phases of BMG-1, U-87 and 4197 cells, while no significant changes could be observed in 4451 cells. Higher endogenous levels of cyclin B1 found in both the glioma cell lines, was enhanced further by the ligand as compared to the squamous carcinoma cells. These results clearly demonstrate that the radiosensitizing effects of the ligand are indeed heterogeneous among different human tumor cell lines. The radiaosensitization is p53 independent and accompanied by enhanced mitotic death (linked to cytogenetic damage) as well as induction of cyclin B1 mediated apoptosis.
Asunto(s)
Bencimidazoles/farmacología , Fármacos Sensibilizantes a Radiaciones/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Carcinoma de Células Escamosas/patología , Carcinoma de Células Escamosas/radioterapia , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/efectos de la radiación , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , Ciclina B/biosíntesis , Ciclina B1 , Daño del ADN , Glioma/patología , Glioma/radioterapia , Humanos , Micronúcleos con Defecto CromosómicoRESUMEN
Camphor has been reported to exhibit radiomodifying properties for bacteria and solid tumours of mice. Therefore its radiomodifying effect was evaluated in the testis which is a less vascularised and hypoxic tissue. Young adult strain 'A' male mice were taken up for these studies. Camphor was administered at the rate of 0.1 mg/g body weight intraperitoneally and whole body irradiation was done after 45 minutes under normal aerated conditions. Doses of 0.5, 1.0 and 2.0 Gy were delivered to different groups with the help of 60Co gamma cell. Testicular germ cell renewal system which seems to be a very good model system for the study of chemical radiomodifier was used for this assay. Resting primary spermatocytes (RPS) were counted at different time intervals and comparison was made among different treatment groups and controls. It was observed that RPS counts significantly declined in radiation + camphor treated groups in comparison to radiation alone or control groups. Radiomodifying effect of camphor was significantly evident during the recovery period, i.e., on day 8 after 0.5 Gy irradiation and day 6 onward after 1.0 Gy and 2.0 Gy.