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Biomedical subjects

J L Sagripanti

Publications and source records attributed to J L Sagripanti.

At least 19 recordsLinked to original sources
PubMed2000

Cytotoxicity of liquid disinfectants.

This study was prompted by toxic responses to disinfecting agents reported in patients after surgical procedures and in sensitized health care personnel. We evaluated the cytotoxicity of seven substances used in the formulation of common liquid chemical disinfectants and sterilants. We found that a standard method based on direct microscopic examination of cell cultures was insensitive and may result in an underestimation of the risk that disinfectants pose to health care personnel or patients who are exposed to these substances. Using independent quantitative tests measuring the integrity of the cellular membrane, metabolic activity, or cell growth, we found that there is a several-hundredfold difference in the relative toxicity of various disinfecting substances. The concentration toxic in 50% of the cell population (TC(50)) that was found for each disinfectant was similar in a variety of cell lines from human, monkey, or mouse origin. Statistical analysis of TC(50)s suggests that liquid disinfecting agents could be classified in three main groups according to their relative toxicity, with: (1) mild (TC(50) > 1 mM, including phenol, hydrogen peroxide, and formaldehyde); (2) moderate (1mM > TC(50) > 0.1 mM, sodium hypochlorite); and (3) severe (TC(50) < 0.1 mM, glutaraldehyde, cupric ascorbate, and peracetic acid) toxicity. These data suggest a vast difference in the potential risk of various disinfectants and sterilants. The data presented in this study should help to define the relative toxic risk of different disinfecting substances to patients and health care personnel and assist in the selection of safer microbicidal formulations.

Animals↗
PubMed1999

Iron chelators modulate the production of DNA strand breaks and 8-hydroxy-2'-deoxyguanosine.

The interaction of chelators and reducing agents is of particular importance in understanding iron-associated pathology since catalytic iron undergoes cyclic reduction and oxidation in vivo. Therefore, we treated plasmid DNA with free or chelated Fe(III) in the presence of biological reductants, and simultaneously measured the number of single strand breaks (SSBs) and oxidative base modification (8-hydroxy-2'-deoxyguanosine; 8-OHdG) by quantitative gel electrophoresis and HPLC with electrochemical detection, respectively. Production of SSBs and 8-OHdG was linearly correlated suggesting that these two different lesions share a common chemical mechanism. The levels of both lesions were enhanced when Fe(III) was chelated to citrate or nitrilotriacetic acid. Reducing agents showed different potency in inducing DNA damage catalyzed by chelated iron (L-ascorbate > L-cysteine > H2O2). Chelation increased SSB formation by approximately 8-fold and 8-OHdG production by approximately 4-fold. The ratio of SSB/8-OHdG catalyzed by chelated iron, which is twice as high as by unchelated iron, indicates that chelation affects iron-catalyzed oxidative DNA damage in a specific way favoring strand breakage over base modification. Since iron is mostly chelated in biological systems, the production of genomic and mitochondrial DNA damage, particularly strand breaks, in diseases involving iron overload is likely to be higher than previously predicted from studies using unchelated iron.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1999

Bacterial spores survive treatment with commercial sterilants and disinfectants.

This study compared the activity of commercial liquid sterilants and disinfectants on Bacillus subtilis spores deposited on three types of devices made of noncorrodible, corrodible, or polymeric material. Products like Renalin, Exspor, Wavicide-01, Cidexplus, and cupric ascorbate were tested under conditions specified for liquid sterilization. These products, at the shorter times indicated for disinfection, and popular disinfectants, like Clorox, Cavicide, and Lysol were also studied. Data obtained with a sensitive and quantitative test suggest that commercial liquid sterilants and disinfectants are less effective on contaminated surfaces than generally acknowledged.

Bacillus subtilis↗
PubMed1997

Molecular characteristics of Junin virus.

Results suggest that protein, glycerophospolipid, galactoside, and sialyl glycoside residues are present in Junin virus (JV), are accessible to enzymatic digestion, and play an important role in infection. Four major protein bands with molecular masses (Mr) 64 +/- 2, 56 +/- 2, 52 +/- 3 (mean +/- standard deviation, n = 4) and approximately 12-18 kDa were consistently detected after denaturing gel electrophoresis analysis of purified attenuated JV. The 52 kDa protein showed a diffuse tail in the 52-56 kDa range and was considered to be the JV glycoprotein. By Western blotting, the 64 kDa protein bound a JV neutralizing antibody and was considered to be the viral nucleoprotein. Additional bands corresponding to larger proteins (approximately 200, 96, 86, and 78 80 kDa in size), as well as fainter and broader bands in the 23-44 kDa region were also present in purified JV preparations. The relative resistance of virus infectivity to RNase digestion demonstrates that the genome of JV is protected from enzymatic attack. Analysis of purified JV virions by electrophoresis resolved the viral small (S) RNA and large (L) RNA species, 3636 +/- 54 bases and 7667 +/- 154 bases long, respectively (average length +/- range, in four determinations). The (S) RNA of attenuated JV appeared slightly larger than that reported for a pathogenic strain, ruling out a large sequence deletion as a reason for attenuation.

Animals↗
PubMed1997

Comparative sensitivity of 13 species of pathogenic bacteria to seven chemical germicides.

BACKGROUND: The relative resistance of diverse human bacterial pathogens to commonly used germicidal agents has not been established. METHODS: We measured by titration the survival of thirteen different bacteria after exposure to glutaraldehyde, formaldehyde, hydrogen peroxide, peracetic acid, cupric ascorbate, sodium hypochlorite, or phenol. RESULTS: Our comparative experiments allowed classification of the organisms' survival into four groups: (a) Pseudomonas aeruginosa and Staphylococcus aureus showed the most resistance, (b) Clostridium perfringens, Salmonella typhimurium, Staphylococcus epidermidis, and Escherichia coli O157:H7 showed intermediate resistance, (c) Listeria monocytogenes, Shigella sonnei, and Vibrio parahaemolyticus survived some treatments with chemical agents only in the presence of protecting protein (serum albumin), and (d) Vibrio cholerae, Vibrio vulnificus, Bacillus cereus, and Yersinia enterocolitica did not survive any of the treatments applied. CONCLUSION: We found species that more frequently survived exposure to germicidal agents were also those most commonly reported in association with hospital infections. Our findings suggest that resistance to disinfectants may be more important than pathogenicity in determining the relative prominence of an organism as an agent responsible for nosocomial infections.

Bacteria↗
PubMed1997

Mechanism of copper-mediated inactivation of herpes simplex virus.

The inactivation of herpes simplex virus (HSV) by copper was enhanced by the following reducing agents at the indicated relative level: ascorbic acid >> hydrogen peroxide > cysteine. Treatment of HSV-infected cells with combinations of Cu(II) and ascorbate completely inhibited virus plaque formation to below 0.006% of the infectious virus input, while it maintained 30% viability for the host mammalian cells. The logarithm of the surviving fraction of HSV mediated by 1 mg of Cu(II) per liter and 100 mg of reducing agent per liter followed a linear relationship with the reaction time, in which the kinetic rate constant for each reducing agent was -0.87 min(-1) (r = 0.93) for ascorbate, -0.10 min(-1) (r = 0.97) for hydrogen peroxide, and -0.04 min(-1) (r = 0.97) for cysteine. The protective effects of metal chelators and catalase, the lack of effect of superoxide dismutase, and the partial protection conferred by free-radical scavengers suggest that the mechanism of copper-mediated HSV inactivation is similar to that previously reported for copper-mediated DNA damage. The sensitivity exhibited by HSV to Cu(II) and reducing agents, particularly ascorbate, might be useful in the development of therapeutic antiviral agents.

Animals↗
PubMed1996

Cupric and ferric ions inactivate HIV.

Human immunodeficiency virus (HIV-1) was inactivated by either cupric or ferric ions when the virus was free in solution and also 3 hr after cell infection. Fifty percent inactivation of cell-free HIV was achieved with Cu(II) at a concentration between 0.16 and 1.6 mM, or by 1.8 to 18 mM Fe(III). Thus, the dose to inactivate 50% of infectious HIV (D50) by Cu(II) or Fe(III) is higher than that reported for glutaraldehyde (0.1 mM); between the D50 reported for sodium hypochlorite (1.3 mM) and sodium hydroxide (11.5 mM), and significantly lower than that required for HIV inactivation by ethanol (360 mM). Treatment of infected cells for 30 min at 20 degrees C with 6 mM Cu(II) or Fe(III) completely inhibited the formation of syncytia and the synthesis of virus-specific p24 antigen in HIV-infected cells, while still preserving cell viability. The virucidal properties of cupric and ferric ions could be exploited for the development of novel virucidal formulations efficient against HIV.

Copper↗
PubMed1996

Association between 8-hydroxy-2'-deoxyguanosine formation and DNA strand breaks mediated by copper and iron.

Oxidative DNA damage is involved in diverse biological phenomena and consists of several kinds of lesions, mainly, strand breaks, base modifications, and DNA-protein crosslinking. However, little is known about the existence of a chemical relationship among them or the ratio by which these different types of lesions are produced. In the present study we investigated whether a relationship exists between DNA strand breakage and base modification. We selected cupric [Cu(II)] and ferric [Fe(III)] ions for this study because these transition metals are active catalysts of DNA damage in vivo. Supercoiled plasmid DNA pZ189 was treated with Cu(II) or Fe(III) in the presence of different reducing agents. We measured in each sample both the number of DNA single-strand breaks (SSB) by quantitative electrophoresis and the amount of a modified DNA base, 8-hydroxy-2'-deoxyguanosine (8-OHdG) by HPLC with simultaneous electrochemical (EC) and spectrophotometric detection. The number of DNA SSBs produced was linearly related to the number of 8-OHdG present. The regression of the number of SSBs as a function of the number of 8-OHdG is expressed by the equation [SSBs] = b x [8-OHdG], where b = 1.7, 2.0, 2.7, 1.7, and 9.4, for Cu(II) in the presence of H2O2, L-cysteine and L-ascorbate, and for Fe(III) in the presence of H2O2 and L-ascorbate, respectively. The linear correlation we observed between the production of SSB and 8-OHdG mediated by Fe(III) and by Cu(II) suggests that these products may arise via a common chemical mechanism and could allow an easier and more precise estimation of DNA breakage.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1996

Comparative sporicidal effect of liquid chemical germicides on three medical devices contaminated with spores of Bacillus subtilis.

BACKGROUND: The relatively limited variety of surfaces and geometries challenged in current sporicidal testing reduces the predictive value of these analyses when extrapolated to the wide variety of medical devices. The unknown spore load being challenged and the qualitative nature (growth/no growth) of those tests further prevent precise comparison among liquid chemical disinfectants. Hence, the relative activity of different chemical substances has not been clearly established, hindering selection of the best agent for each clinical situation. METHODS: A micromethod was developed to assess sporicidal activity against Bacillus subtilis spores deposited on three different medical devices; carbon steel dental burs, silicone-rubber medical catheters, and titanium-alloy dental abutment screws. The spore load on each device and the recovery after three analytical steps were quantitatively assessed with spores radiolabeled with carbon 14 methionine. RESULTS: The killing of 2 to 7 x 10(6) spores loaded on three different devices and exposed to glutaraldehyde, formaldehyde, copper ascorbate, hydrogen peroxide, peracetic acid, sodium hypochlorite, or phenol for 30 minutes at 20 degrees C ranged from a 10(3)-fold decrease for 10% hydrogen peroxide to zero decrease for 5% phenol. Our results suggest that the nature of the surface being challenged may affect the sporicidal activity of some chemical agents. CONCLUSION: The quantitative data presented allow comparison of the sporicidal effect of different liquid chemical agents. These findings may help prevent an overestimation of sporicidal activity and possible transmission of pathogens from the surface of improperly decontaminated medical devices.

Bacillus subtilis↗
PubMed1996

Comparative sporicidal effects of liquid chemical agents.

We compared the effectiveness of glutaraldehyde, formaldehyde, hydrogen peroxide, peracetic acid, cupric ascorbate (plus a sublethal amount of hydrogen peroxide), sodium hypochlorite, and phenol to inactivate Bacillus subtilis spores under various conditions. Each chemical agent was distinctly affected by pH, storage time after activation, dilution, and temperature. Only three of the preparations (hypochlorite, peracetic acid, and cupric ascorbate) studied here inactivated more than 99.9% of the spore load after a 30-min incubation at 20 degrees C at concentrations generally used to decontaminate medical devices. Under similar conditions, glutaraldehyde inactivated approximately 90%, and hydrogen peroxide, formaldehyde, and phenol produced little killing of spores in suspension. By kinetic analysis at different temperatures, we calculated the rate of spore inactivation (k) and the activation energy of spore killing (delta E) for each chemical agent. Rates of spore inactivation had a similar delta E value of approximately 20 kcal/mol (ca.83.68 kJ/mol) for every substance tested. The variation among k values allowed a quantitative comparison of liquid germicidal agents.

Ascorbic Acid↗
PubMed1995

Cytotoxic and mutagenic effects of ferric nitrilotriacetate on L5178Y mouse lymphoma cells.

An iron chelate, ferric nitrilotriacetate (Fe-NTA), induces renal proximal tubular necrosis that leads to a high incidence of renal adenocarcinoma in rodents. Others have shown that Fe-NTA induces modified DNA base products both in vitro and in vivo. However, Fe-NTA is negative in the Ames Salmonella test with or without S9 activation. The goal of this project was to determine if Fe-NTA is cytotoxic and mutagenic using the L5178Y (TK +/-) mouse lymphoma assay. Our experiments showed a relationship between the concentration of Fe-NTA (0 to 1 mM) and the decrease in relative survival. An exposure-dependent increase in the number of mutations was observed with increasing concentrations of Fe-NTA. At 14% relative survival, there was about a 4-fold increase in mutations (trifluorothymidine resistance) over unexposed, control cells. Ferric nitrate or nitrilotriacetic acid alone induced a relatively low 1.5- or 1.1-fold increase in mutation, respectively. Our results establish that Fe-NTA is mutagenic in the L5178Y mouse lymphoma assay system.

Animals↗
PubMed1994

Increased 8-hydroxydeoxyguanosine in kidney and liver of rats continuously exposed to copper.

Copper is a ubiquitous metal in the environment, it is a component of dental casting gold alloys and dental amalgams, and it is a main component in some intrauterine contraceptive devices (IUDs). Since copper materials implanted in the human body corrode and release ions into the surrounding tissue, the potential toxicity caused by contact of this metal with bodily fluids needs to be evaluated. We implanted male Wistar rats with osmotic mini pumps that continuously administered saline, CuCl2, or a copper chelate, cupric nitrilotriacetate (Cu-NTA), at a rate of 4 mg copper/kg body wt/day. This experimental design maintained serum copper concentrations at a level 30-70% (CuCl2) or 100-120% (Cu-NTA) higher than in untreated controls. At different times postimplantation, we measured the levels of 8-hydroxydeoxyguanosine (8-OHdG) in DNA of kidney, liver, and tissue surrounding the pump implant, since production of 8-OHdG has been associated with mutagenesis and carcinogenesis. Hepatic and renal levels of 8-OHdG in CuCl2- or Cu-NTA-treated animals were significantly higher than in control animals. In contrast, histopathologic changes in kidneys and livers of rats exposed to CuCl2 and Cu-NTA were limited to mild changes involving hepatic focal necrosis and slightly increased mitotic activity in the renal proximal tubules. These observations suggest that levels of 8-OHdG could be an early marker of copper toxicity. It is unlikely that the high levels of copper at which we observed DNA modification will be encountered after occupational or environmental exposure. A different situation could be found around medical devices that include copper, particularly IUDs, where the amount of copper administered in our experiments could be released in the uterus of women after a few months of continued IUD use.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1993

Replication and physical parameters important for preparing purified Junin virus.

Junin virus (JV) is an Arenavirus and the causative agent of Argentine hemorrhagic fever (AHF), an often fatal human disease. The attenuated strain XJ-clone 3 (XJC13) of JV, after being tested in humans, has been considered a promising vaccine. We found that synthesis of JV XJC13 reaches a peak 2 days after infection and the kinetics of synthesis are little affected by the multiplicity of infection (MOI) in a range from 0.125 to 1.00. Virus synthesis is sensitive to actinomycin D, indicating that cellular biosynthesis is required for viral replication. Combined precipitation and ultracentrifugation of supernatant from virus-infected cells yielded large amounts of concentrated and purified virion that banded in sucrose as a single peak with average density 1.177 +/- 0.015 g/ml. Purified virions have an average diameter of 203 +/- 23 nm by electron microscopy and an average sedimentation coefficient of 454 +/- 27 S. The results from the present study should assist in the preparation of large amounts of attenuated Junin virus which are required for vaccination against and diagnosis of Argentine hemorrhagic fever.

Animals↗
PubMed1993

Induction of oxidative single- and double-strand breaks in DNA by ferric citrate.

The relative risk of primary hepatocellular carcinoma in genetic hemochromatosis (GH) is estimated at over 200 times as that of control populations. Recently, ferric ion chelated to citrate (Fe-citrate) was identified as the major non-transferrin-bound iron in the serum of GH patients. We investigated whether low concentration of Fe-citrate plus reductant could damage supercoiled plasmid DNA under physiological pH and ionic strength. Incubation of Fe-citrate with either H2O2, L-ascorbate, or L-cysteine induced single- and double-strand breaks in supercoiled plasmid pZ189 in a concentration- and time-dependent fashion. DNA strand breaks produced by Fe-citrate plus H2O2 increased at reduced pH (< or = 6.9). Catalase and free radical scavengers inhibited the DNA breakage produced by Fe-citrate in combination with each reductant, suggesting that H2O2 and finally .OH are responsible DNA damaging species. The catalytic ability of Fe-citrate to induce DNA strand breaks, particularly double-strand breaks (DSBs), may contribute to the carcinogenic processes observed in GH.

Ascorbic Acid↗
PubMed1993

DNA single- and double-strand breaks produced by ferric nitrilotriacetate in relation to renal tubular carcinogenesis.

Fe(III) bound to a chelator, nitrilotriacetate (NTA), has been reported to induce a high frequency of adenocarcinoma localized to the proximal tubules of the kidney in rodents. In order to examine possible mechanisms for the carcinogenic activity, we investigated the in vitro production of single- and double-strand breaks in DNA mediated by iron alone or Fe-NTA chelate using supercoiled plasmid pZ189. Neither Fe(III) nor NTA alone broke DNA. Fe(III) plus NTA together mediated the efficient oxidative production of DNA single- and double-strand breaks in the presence of reducing agents (ascorbate >> H2O2 > cysteine). The Fe(III):NTA ratio (1:4) that was found to be optimal for DNA strand breakage was similar to the ratio that produced adenocarcinomas in rodents. Maximal Fe-NTA-mediated DNA damage in vitro was induced under conditions of neutral pH, low ionic strength, presence of reducing agent and absence of albumin. These conditions are present exclusively in the cortical proximal tubules of the kidney, the only location where toxicity and carcinogenicity of Fe-NTA has been observed. Thus, localized DNA damage may explain the anatomic site preferred by Fe-NTA-induced carcinogenesis.

Adenocarcinoma↗
PubMed1993

Virus inactivation by copper or iron ions alone and in the presence of peroxide.

Cupric and ferric ions were able to inactivate five enveloped or nonenveloped, single- or double-stranded DNA or RNA viruses. The virucidal effect of these metals was enhanced by the addition of peroxide, particularly for copper(II). Under the conditions of our test, mixtures of copper(II) ions and peroxide were more efficient than glutaraldehyde in inactivating phi X174, T7, phi 6, Junin, and herpes simplex viruses. The substances described here should be able to inactivate most, if not all, viruses that have been found contaminating medical devices.

Copper↗
PubMed1992

Detection of Junin virus by the polymerase chain reaction.

Argentine hemorrhagic fever is an often fatal human disease caused by Junin virus, an RNA-containing virus and member of the Arenavirus family. This virus was detected in vitro by the polymerase chain reaction (PCR) procedure. A pair of Junin virus-specific PCR DNA oligonucleotide primers and an oligonucleotide probe were designed from a known portion of the viral RNA sequence. RNA was isolated from Junin virus-infected monkey kidney cells and used to produce complementary DNA (cDNA) by reverse transcription. A DNA segment, 151 +/- 24 bp long, was amplified from this cDNA and characterized by agarose gel electrophoresis and Southern blot hybridization with the Junin virus-specific DNA probe. Sensitivity experiments showed that Junin virus could be detected with nanogram quantities of RNA isolated from virus-infected cells. The rapid and sensitive assay described here may contribute towards the development of a procedure for the early diagnosis of Argentine hemorrhagic fever.

Arenaviruses, New World↗