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Papers by David Grdina

Abstract 507: Elevated manganese superoxide dismutase (SOD2) activity as a mechanism for the low dose dadiation- and thiol-induced adaptive responses in human RKO36 colon carcinoma cells and BFS2C mouse fibrosarcoma cells

Exposure of cells to low non-lethal doses of ionizing radiation (≤ 10 cGy) or WR1065, the active ... more Exposure of cells to low non-lethal doses of ionizing radiation (≤ 10 cGy) or WR1065, the active free thiol form of amifostine, can induce pro-survival pathways that result in protection against the damaging effects of a 2 Gy dose of ionizing radiation. One such signaling pathway involves the elevation of active manganese superoxide dismutase (SOD2). SOD2 is a mitochondrial matrix protein that serves as the primary mitochondrial defense against superoxide formation. Its primary function is to facilitate the dismutation of two molecules of superoxide anion (O2−) produced by normal respiratory processes or following exposure to ionizing radiation into water and hydrogen peroxide. To characterize the role of SOD2 in the radiation- and thiol-induced adaptive responses, RKO36 human colon carcinoma cells and BFS2C mouse fibrosarcoma cells were exposed to 10 cGy x-rays and 40 μM or 4 mM WR1065 and SOD2 activity measured 24 h later. Significant increases in SOD2 activity in RKO36 cells (3.7-fold, P < 0.001) and BFS2C cells (3.0-fold, P = 0.005) were observed 24 h after exposure to 10 cGy. SOD2 activity was also observed to be significantly elevated in RKO36 cells (3.5-fold, P = 0.014 and 3.4-fold, P = 0.017) and BFS2C cells (3.7-fold, P = 0.007 and 2.5 fold, P = 0.021) 24 h after treatment with 40 μM or 4 mM WR1065, respectively. The protective effect of the low dose radiation- and thiol-induced elevation in active SOD2 was examined using the endpoint of micronuclei formation as a measure of chromosomal damage. Exposure of RKO36 and BFS2C cells to 2 Gy resulted in a significant increase in the mean number of micronuclei observed relative to unirradiated control cells (8.8 versus 2.2, P < 0.001; 5.4 versus 2.1, P < 0.001, respectively). The frequency of micronuclei formation was significantly reduced in both RKO36 cells (P = 0.003) and BFS2C cells (P = 0.011) exposed to 10 cGy 24 h prior to the 2 Gy dose. A significant reduction in micronuclei formation was also observed in RKO36 and BFS2C cells treated with 40 μM or 4 mM WR1065 30 min or 24 h prior to irradiation with 2 Gy. Treatment of both cell lines with SOD2 siRNA reduced SOD2 activity relative to mock-transfected control cells which resulted in the abrogation of both the radiation- and thiol-induced protective effect. This work was supported by NIH/NCI grant R01 CA132998 and DOE grant DE- PS02-08ER08-21 (D.J.G.). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 507.

Molecular Aspects of Medicine, 1991

Experientia, May 1, 1981

Tumor cells isolated from a murine fibrosarcoma were grown in primary culture for two days and th... more Tumor cells isolated from a murine fibrosarcoma were grown in primary culture for two days and then separated on a basis of size by velocity sedimentation. Centrifugal elutriation and STAPUT methods were compared for their ability to isolate biophysically unique tumor subpopulations. The isolated cell fractions were assayed for cell number, incorporation of triatiated thymidine and Coulter volume. Both methods were comparable with regard to ability to separate tumor cells on a basis of size. Elutriation had the advantage of speed but required sophisticated equipment. The STAPUT method was less expensive but required somewhat longer times for separation.

Cytotoxic effect of adriamycin in vivo on synchronized murine fibrosarcoma cellst

International Journal of Radiation Oncology Biology Physics, Aug 1, 1979

Abstract The cytotoxic effect of Adriamycin (ADR) on synchronized fibrosarcoma (FSa) cells lodged... more Abstract The cytotoxic effect of Adriamycin (ADR) on synchronized fibrosarcoma (FSa) cells lodged in the lungs of specific pathogen free C 3 Hf/Bu mice was determined. FSa cells from primary asynchronous cultures were separated and synchronized on the basis of cell size by centrifugal elutriation. Cell cycle parameters were determined by flow micro-fluorometry (FMF). Viable tumor cells from each elutriator fraction were injected intravenously into recipient mice. Twenty minutes later, ADR (10 mg/kg) was administered intravenously into half of the animals in each group. After fourteen days the animals were killed, and the resulting lung colonies were counted. Under these conditions, the S-phase enriched populations were found to be the most sensitive to ADR. Cell survival ranged from a maximum of 8% to a minimum of 0.3% for the Gi and S-phase enriched populations, respectively. Dose response curves were also determined for selected populations treated in vivo . While no great difference in Do was observed between the survival curves representing each of these populations (range, 0.65–0.71 mg/kg), the extrapolation numbers varied from 0.41 (51% S-phase) to 2.1 (9% S-phase).

International Journal of Radiation Oncology Biology Physics, Nov 1, 1990

V79 cells grown as spheroids are more radioresistant than those grown as monolayers. Viable cells... more V79 cells grown as spheroids are more radioresistant than those grown as monolayers. Viable cells from spheroid culture contain restraints to ethidium bromide driven rewinding of DNA supercoils that are absent in monlayer cells. Spheroid cells also contain a DNA-protein matrix that is more resistant to detergent-induced degradation. The increase in structural integrity may be related to a 55-60 kD protein in the nucleoids of spheroid, but not monolayer cells. Spheroid cell radioresistance may therefore be related to a more stable chromatin platform for high fidelity repair of DNA damage.

Cell Proliferation, Nov 1, 1981

Murine fibrosarcoma (FSa) cells form at least five unique subpopulations after centrifugation in ... more Murine fibrosarcoma (FSa) cells form at least five unique subpopulations after centrifugation in linear Renografin density gradients. Each of these subpopulations has been characterized with respect to selected kinetic parameters using pulse-labelling techniques and flow microfluorometry (FMF) analysis. Tumourbearing mice were first injected intraperitoneally with a pulse label of tritiated thymidine (13H]TdR, 50 pCi). Following 15, 30, 60 min or 24 hr these animals were injected with cold thymidine. Animals were killed, their tumours removed and made into suspension, and separated by density gradient centrifugation. Each gradient was fractionated and the density, cell number, tritium activity, and labelling index (LI) per fraction were determined. These data were then compared to F M F data for selected cell density bands. The results indicated a relatively higher uptake of [ 3H1TdR in the cells recovered at the lighter (1.06-1.12 g/cm3) as compared to the heavier (> 1.12 g/cm3) densities. Following a 30-min pulse, the LI's of light cells ((1.12 g/cm3) ranged from 25 to 30%, while the heavier cells (> 1.12 g/cm3) had LI's between 10 and 15%. The unseparated control cells had an LI of 19%. Comparable results were found at the other times tested. In contrast, the F M F profiles describing the DNA contents of the cells banding in the gradient showed no difference in proportion of S-phase cells among the separated subpopulations. This lack of correlation between the F M F determination of S-phase cells and labelling index for the denser cell populations implies that D N A content alone is not an effective measurement of the functional activity of cells in solid tumours. Finally, the relatively reduced uptake of [3HITdR by these denser cells suggests that they may have resided at relatively large distances from the functional vasculature in the tumour.

Reversion of radiosensitivity in azacytidine-treated xrs5 cells does not result in full radioprotection by WR-1065

International Journal of Radiation Oncology Biology Physics, 1992

A series of cell lines were previously generated from the radiation sensitive Chinese hamster ova... more A series of cell lines were previously generated from the radiation sensitive Chinese hamster ovary line xrs5 after treatment with azacytidine. Six of these lines have been examined for their resistance to killing by 0 to 20 Gray of 60Co gamma rays and the amount of radioprotection afforded by treatment with the drug 2-[(aminopropyl)amino]ethanethiol (WR-1065). As xrs5 cells have lost the ability to be protected by WR-1065, studies were performed to determine whether reversion to radio-resistance correlated with recovery of aminothiol radioprotection. Treatment of azacytidine-treated, radiation sensitive and resistant cells with four millimolar WR-1065 30 minutes prior to irradiation enhanced survival after exposure to gamma radiation, although the enhancement in survival was less than for wild type Chinese hamster ovary K1 cells. The data suggest that there is not an absolute linkage between recovery of gamma ray radiation resistance and protection by WR-1065 and other factors, such as chromatin organization, must play a role.

International Journal of Radiation Oncology Biology Physics, Feb 1, 1983

The survival of eeIIs from 2 humor subpop&tions after y-ray doses rangiog from 1 to 19 Gy was det... more The survival of eeIIs from 2 humor subpop&tions after y-ray doses rangiog from 1 to 19 Gy was determhd using a lung coboy assay. Metbykbohntbrene-hdoced Ibrosarcomas grown in the hind legs of QH/Kam ptbogen-free mice were imdiited in situ nben the tamers were 8-10 mm in dia. Single cell =w=io= Preps& from excised twmors were separated on a linear deosity gradient, aml the clomogaicity of predomilrrntly oxic BunI 2 (density 1.08 g/cm') and @omhaotly bypoxic Ed 4 (dmslty 1.14 p/cm') cclk was messwed. The sarviriqg fraction of celk 8fter doses of 1, z rmi 3 cy was estimated from that masmredaftertotal&sesofSCy -5 x 1 cy, IO Cy -5 x 2 Cy. and 15 Gy -5 x 3 Cy, under the assumption of equal effect per fractiuo (checked by estiartiog survivrl8t 3 Cy after different numbers of fractioos). Very little curvature was evident in tbe wrvival curves of Bawl 2 and Balld 4 celk (B/a -.0X+-.034 Cy-'). The initial segment of tbc shvinl curve of the pfehmhntly oxic B8ml2 cells was steeper (,D, -3.6 Cy) thn that of the predomiwttly bypoxic Bud 4 celk (,D, -5.2 Gyk both read& li8ar over 8 huge range+ to bses in excess of 3 Cy. These results imply that tbesc tumor smbpophth? Will be hmemitive, in their responw to multifractionated regimeq to changes in size of daee per fractioo in the raoge 0 to 3 Gy, a trait shed by two ?? cuteiy respoading normal tissues (murine testis rod jejunum). Tllmor cell wninl, Low dose, Ir situ, Equel effect per fr&ion.

British Journal of Cancer, Nov 1, 1980

The cytotoxic effects in vivo of single doses of either adriamycin (ADM), 1-p -D-arabinofuranosyl... more The cytotoxic effects in vivo of single doses of either adriamycin (ADM), 1-p -D-arabinofuranosylcytosine (Ara-C), bleomycin (BLM), cis-diamminedichloro- platinum (II) (cis-DDP), or cyclophosphamide (CY) on murine fibrosarcoma (FSa) cell populations were determined. Tumour cells were separated and synchronized by centrifugal elutriation. Viable tumour cells from selected elutriator fractions were then injected i.v. into whole-body-irradiated mice. Twenty minutes later selected doses of ADM, Ara-C, BLM, cis-DDP or CY were administered to selected groups of these animals. Fourteen days later the mice were killed. Killing of injected tumour cells by each of the chemotherapeutic agents was evidenced by a reduction in the lung colonies per cell injected in treated animals. Under these conditions the response of FSa cells in vivo to the 5 drugs tested differed both qualitatively and quantitatively. Ara-C was S-phase-specific in toxicity. ADM, BLM, and cis-DDP were preferentially toxic to S, G2+M and G1 cells respectively. CY, a drug requiring bioactivation to form alkylating metabolites, was found to be equally toxic to G1 and G2 + M enriched populations, but less effective in killing cell populations enriched with early-S cells.

Method for protection against tumor metastasis formation

Coactivation of ATM/ERK/NF-KB in the low-dose radiation-induced radioadaptive response in human skin keratinocytes

Free Radical Biology Medicine, 2009

On the a/� ratio and oer for clonogenic tumor cells

Int J Radiat Oncol Biol Phys, 1985

Application of an in vivo mutagenesis system to assess aminothiol effects on neutron-induced genotoxic damage in mouse spleenocytes

Abstract 67: A novel survivin-mediated adaptive response induced by very low dose imaging-level exposures and radiation therapy

Cancer Research, 2013

Exposure of cells to a very low dose of ionizing radiation can induce an enhanced resistance or a... more Exposure of cells to a very low dose of ionizing radiation can induce an enhanced resistance or adaptive response to a subsequent larger radiation dose as evidenced by an increase in cell survival. This expression of an adaptive response has been attributed to the result of signaling processes induced by very low radiation doses in the range of 0.5 to 100 mGy. The radiation-induced adaptive response is garnering considerable attention now due in part to the burgeoning increase in the use of imaging technologies such as computerized axial tomography and portal imaging to monitor tumor response and positioning during multi-dose standard radiation therapy protocols. The focus of this study was to investigate whether this process could occur when the delivery of the very small radiation dose was administered before or in between the administration of two typically used radiation therapy level doses. Specifically, could this signaling process be perturbed by the addition of a second 2 Gy...

WR-2721 [S-2-(3-aminopropylamino)ethylphosphorothioic acid] is an effective chemopreventive agent... more WR-2721 [S-2-(3-aminopropylamino)ethylphosphorothioic acid] is an effective chemopreventive agent. C57BL [times] BALB/c F[sub 1] female mice, were exposed to a single whole-body dose of 206 cGy from a [sup 60]Co photon source. Those groups treated with VATR-2721 (400 mg/kg) were administered the agent i.p. 30 min prior to irradiation. Over 90% of deaths were determined to be due to tumor involvement. WR-2721 afforded significant protection against life shortening due to radiation-induced tumors of connective tissue and epithelial tissue origins. Subsequent survival time in WR-2721-treated and irradiated animals as compared to matched irradiated-only controls was extended up to 59 days. A single exposure of animals to VVR-2721 did not affect the cumulative survival curves for unirradiated mice. WR-2721 possesses chemopreventive properties which can be clinically exploited to reduce the risk to therapy-induced secondary cancers in patients who otherwise would have an excellent prognos...

Radiation Response of Cell Populations Irradiated in Situ and Separated from a Fibrosarcoma

Radiation Research, 1976

Radiation Response of Cell Populations Irradiated in Situ and Separated from a Fibrosarcoma'... more Radiation Response of Cell Populations Irradiated in Situ and Separated from a Fibrosarcoma' ... DAVID J. GRDINA, IVAN BASIC,2 SHARON GUZZINO, AND KATHRYN A. MASON ... The University of Texas System Cancer Center, MD Anderson Hospital and ...

Cyclic-radiation response of murine fibrosarcoma cells grown as pulmonary nodules

International Journal of Radiation Oncology*Biology*Physics, 1982

ABSTRACT

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Protective effects of 2-[(aminopropyl)amino] ethanethiol against bleomycin and nitrogen mustard-induced mutagenicity in V79 cells

International Journal of Radiation Oncology*Biology*Physics, 1986

This study examines the effects of the radioprotector 2-[(aminopropyl)amino] ethanethiol (WR-1065... more This study examines the effects of the radioprotector 2-[(aminopropyl)amino] ethanethiol (WR-1065) on bleomycin (BLM) and nitrogen mustard- (HN2) induced cytotoxicity, DNA damage, and mutagenesis at the hypoxanthine-guanine phosphoribosyl transferase (HGPRT) locus in V79 Chinese hamster cells. The anti-mutagenic effect of WR-1065 on cis-diamminedichloroplatinum (cis-DDP) and radiation- (XRT) induced HGPRT mutations was also evaluated for comparative purposes. WR-1065 (4 mM) was added prior to exposure of cells to therapy agents. All exposure times were 30 min. and both cell survival and mutagenesis were assayed. WR-1065 was effective in protecting against both effects. The induction of mutants corrected for background by BLM, HN2, cis-DDP, or XRT was linear in all cases. Mutation frequencies without WR-1065 were 78 X 10(-6) per unit BLM, 66 X 10(-7) per microgram HN2, 25 X 10(-7) per microgram cis-DDP; and 87 X 10(-7) per Gy of XRT. With WR-1065, these were reduced to 37 X 10(-6) per unit BLM, 40 X 10(-7) per microgram HN2, 1 X 10(-7) per microgram cis-DDP, and 44 X 10(-7) per Gy of XRT. Mutation protection factors (MPF), a ratio of the corresponding slopes of the mutation induction curves, with and without WR-1065 were: BLM, MPF = 2.8; HN2, MPF = 3.4; cis-DDP, MPF = 7.1; and XRT, MPF = 5.1. Single-strand-break (SSB) formation in DNA by BLM or HN2, assayed by alkaline elution, was protected against by WR-1065. WR-1065 did not induce SSB in control cells. The reduction of the mutagenic effects of agents used in radiation and chemotherapy by radioprotectors may be an important additional benefit for consideration in their use in cancer treatment.

Protection against AZT-induced mutagenesis at the HGPRT locus in a human cell line by WR-151326

International Journal of Radiation Oncology*Biology*Physics, 1992

The anti-AIDS agent, AZT (3'-azido-3'-deoxythymidine), is mutagenic in a cultured... more The anti-AIDS agent, AZT (3'-azido-3'-deoxythymidine), is mutagenic in a cultured human hepatoma cell line designated HepG2 at the HGPRT (hypoxanthine-guanine phosphoribosyl transferase) locus. Using an exposure time of 3 hr, the number of mutants per 10(6) surviving cells increased as a function of AZT dose from 125 to 520. Chinese hamster ovary cells, in contrast, are not affected with respect to this endpoint when similar concentrations of AZT are used (i.e., 0.1 to 10 mg/ml). The aminothiol WR-151326 [3-(3-methylaminopropylamino) propanethiol dihydrochloride] was evaluated as a possible antimutagen for use with AZT. At a concentration of 4 mM, WR-151326 was added either concomitantly or following exposure of HepG2 cells to a 5 mg/ml concentration of AZT. Regardless of the treatment condition, WR-151326 was effective in reducing the mutagenic effects of AZT by about a factor of 2. Correcting for background mutations, the mutation frequencies determined were: AZT only for 3 hr, 110 x 10(-6) (S.E.M. +/- 6.0 x 10(-6)); AZT together with WR-151326 for 3 hr, 57 x 10(-6) (S.E.M. +/- 3.0 x 10(-6)); and AZT for 3 hr followed by WR-151326 for 3 hr, 68 x 10(-6) (S.E.M. +/- 5.0 x 10(-6)). This study demonstrates that AZT is mutagenic to a cell line of human origin and that WR-151326 can protect against this mutagenic process.

International Journal of Radiation Oncology*Biology*Physics, 1983