Biomedical subjects

R K Johnson

Publications and source records attributed to R K Johnson.

At least 163 records · Page 9Linked to original sources

PubMed1988

A murine model to evaluate the ability of in vitro clonogenic assays to predict the response to tumors in vivo.

The use of the human tumor cloning assay as a predictor of clinical response of human tumors to drugs is predicated on the hypothesis that the in vivo response of a tumor to a drug can be correlated with the in vitro response of cells derived from the tumor. To test this hypothesis, we utilized a murine tumor model in which the in vivo and in vitro responses of a tumor can be accurately and reproducibly compared. Drug activity was assessed in P388 leukemia with the standard in vivo antitumor assay (i.p. tumor/i.p. drug administration) and an in vitro assay wherein the ascites tumor cells are removed from mice, treated with a drug, and directly cloned in soft agar to measure clonogenic capacity. The response of P388 cells to analogues within four separate classes of antitumor agents, anthracyclines, anthraquinones, platinum(II) coordination complexes, and phosphinogold(I) complexes was evaluated. The clonogenic assay failed to discriminate between highly active in vivo antitumor agents and analogues with only marginal in vivo efficacy (i.e., doxorubicin and daunorubicin versus rhodomycins A and B, ametantrone versus NSC 276740, cisplatin versus transplatin, [Au(dppe)2]Cl versus [Au(depe)2]PF6. Furthermore, the in vitro clonogenic assay failed to detect carboplatin which was a highly active agent in vivo. The basis for these discrepancies was explored by a more detailed comparison of doxorubicin and rhodomycin B. In vivo or in vitro drug exposure with subsequent measurement of cell kill by the in vitro clonogenic and in vivo tumorigenic assay demonstrated that the in vitro assay overestimated the cytotoxic potency of the drugs relative to the tumorigenic assay. Treatment of tumors in vivo with doxorubicin at doses below the maximally tolerated dose in mice resulted in multiple log cell kill as measured in vitro or in vivo, whereas rhodomycin B was cytotoxic only at dose levels exceeding its maximally tolerated dose. The results indicate that a subset of tumor stem cells capable of forming colonies in soft agar are significantly more sensitive to the cytotoxic effects of anthracyclines than are in vivo tumorigenic stem cells. Cytotoxic potency as measured by an in vitro soft agar clonogenic assay is not an accurate predictor of in vivo antitumor efficacy even in a model in which ascites tumor cells are directly exposed to i.p. drug. The in vitro cytotoxicity assay is useful only as a nonselective prescreen and must be used in combination with other indicators of tumor cell selectivity and dose-limiting organ toxicity.

Animals↗

PubMed1988

Tumorigenicity of the cyc- variant of the S49 murine lymphoma deficient in the Gs-alpha subunit of adenylate cyclase.

S49 cyc- lymphoma cells contain a mutation resulting in loss of a functional guanine nucleotide regulatory protein rendering their adenylate cyclase refractory to most stimuli. S49 wild-type and cyc- clones were used in the present study to investigate the possible association of altered cAMP metabolism with tumorigenicity and metastatic potential. The S49 clones were implanted i.v., i.p., and intracerebrally in both athymic nude mice and syngeneic, immunocompetent BALB/c mice. Both S49 clones gave rise to tumors when inoculated into athymic mice, and no differences were observed in the tumorigenicity or metastatic potential of S49 wild-type and cyc- cells. Implantation of S49 clones in syngeneic BALB/c mice gave rise to few tumors except when administered intracerebrally, where wild-type cells were more tumorigenic than cyc- cells. This raises the possibility of differences in immunogenicity between the S49 clones. Analysis of cell lines derived from tumors grown in athymic mice showed that they retained the phenotype of the S49 clones used for inoculations. The results indicate that, despite differences in adenylate cyclase responsiveness, S49 wild-type and cyc- cells are both highly tumorigenic and metastatic.

Adenylyl Cyclases↗

PubMed1988

A comparison of EMG procedures in the carpal tunnel syndrome with clinical-EMG correlations.

One hundred and twenty-two patients were selected on the basis of symptoms of finger numbness and tingling thought to suggest carpal tunnel syndrome. The severity of the symptoms was graded and the patients were studied with seven established EMG procedures, motor inching to the abductor pollicis brevis and second lumbrical muscles, and sensory inching. Symptom severity was compared with the degree of nerve conduction abnormality and results from symptomatic hands with those abnormal results from patient's asymptomatic hands. Motor inching to the abductor pollicis brevis muscle was the most sensitive study and, when combined with sensory inching, gave a sensitivity of 97-100% (depending on symptom severity). Motor inching to the second lumbrical muscle alone detected the lesion site in several severely involved nerves. Guidelines are suggested with which one can estimate the likelihood that a patient's finger sensory symptoms are on the basis of carpal tunnel syndrome.

Adolescent↗

PubMed1988

Antimicrobial and anticancer activity of tetrahedral, chelated, diphosphine silver(I) complexes: comparison with copper and gold.

Tetrahedral, bischelated Ag(I) diphosphine complexes [Ag(P-P)2]NO3, where P-P is Ph2P(CH2)2PPh2 (dppe), Et2P(CH2)2PPh2 (depe), and cis-Ph2P(CH = CH)PPh2 (dppey), are potently cytotoxic to B16 melanoma cells in vitro (IC50 4 microM) and exhibit good activity against ip P388 leukemia in mice. The complex [Ag(dppe)2]NO3 is active against M5076 reticulum cell sarcoma. The antibacterial and antifungal activities of Ag(I) diphosphine and related Cu(I) and Au(I) complexes were assessed. The complexes [Au(dppey)2]Cl, [Au(dppp)2]Cl and (CuCl)2(dppe)3 show modest activity against three of the 12 bacterial strains tested, but all complexes exhibit antifungal activity against three strains of C. albicans in a "defined" medium, [Ag(depe)2]NO3 and [Au(dppp)2]Cl having comparable activity to fungizone. Antifungal activity of the complexes is reduced in Sabouraud's broth medium, and lost altogether for the Ag(I) complexes. Reactions of some of the Ag(I) complexes with glutathione and blood plasma were studied by 31P NMR.

Animals↗

PubMed1988

cis-4-Carboxy-6-(mercaptomethyl)-3,4,5,6-tetrahydropyrimidin-2(1 H)-one , a potent inhibitor of mammalian dihydroorotase.

A series of cis- and trans-4-carboxy-3,4,5,6-tetrahydropyrimidin-2(1H)-ones possessing either a carboxy, hydroxymethyl, or mercaptomethyl substituent at C-6 were prepared and tested for their ability to inhibit mammalian dihydroorotase. Of these compounds, only the cis-6-mercaptomethyl compound, cis-1, was found to be a potent competitive inhibitor of the enzyme (Ki = 140 nM at pH 7.4 and 8.5) when assayed in the direction of dihydro-L-orotate hydrolysis. These results suggest that the inhibition arises from the ligation of the thiolate to the zinc atom which is thought to be located in the enzyme's active site. Although analysis of cis-1 with 2,2'-dithiobis(5-nitrobenzoic acid) revealed significant loss of the free thiol group under enzymatic assay conditions, the addition of the reducing agent, dithiothreitol, to the enzymatic reaction mixtures afforded cis-1 complete protection against this chemical decomposition, as evidenced by lowering of the inhibition constant in the presence of dithiothreitol. Compound cis-1 had no significant antiproliferative activity against B16 melanoma cells in tissue culture, possibly due to the rapid decomposition of the compound or poor permeability into cells.

Amidohydrolases↗

PubMed1988

Effect of sex ratio of the birth litter on subsequent reproductive performance of gilts.

Records on age at puberty from 1,555 gilts and total number of pigs born in litters of 1,187 gilts from the Nebraska gene pool population were used to evaluate the effects of uterine environment on subsequent reproductive performance. Independent variables were line, year, line x year, proportion of males in the birth litter (sex ratio), number born in the birth litter (fraternity size) and sex ratio x fraternity size. Sex ratio, fraternity size and their interaction influenced age at puberty (P less than .01) but not number born (P greater than .2). Partial regression coefficients indicated that age at puberty tended to decrease as sex ratio increased, particularly in small litters. Although the regression coefficients were relatively large, sex ratio, fraternity size and their interaction accounted for only 1.3% of the variation in age at puberty within line x year subclass. These results offer little encouragement for the use of sex ratio as a phenotypic selection criterion for improvement of reproductive performance in gilts. Results suggest that female swine are similar to rodents in response to uterine environmental effects.

Animals↗

PubMed1988

Correlated responses of carcass and reproductive traits to selection for rate of lean growth in swine.

Mass selection for an index of increased postweaning average daily gain and decreased backfat thickness was practiced for five generations. Litter size and weight for 221 gilt litters, birth weight and nipple number for 2,242 piglets and weaning weight at 42 d of age for 2,111 pigs were recorded. Carcass measurements were taken on 331 pigs. Differences between means of the lines (select control) were regressed on cumulative selection differential of the index. These regression coefficients were negative (P greater than .10) for total number born, number born alive, number weaned per litter, nipple number and carcass backfat thickness. Coefficients were positive (P greater than .10) for individual pig and litter weights at birth and weaning and for the carcass traits of length, longissimus muscle area and percentage of ham and loin. Absolute values of realized genetic correlations of index with traits evaluated were all .35 or less except the correlation with carcass backfat, which was -.84. None of these was significant; therefore, index selection for lean growth should have little effect on litter size and weight but may have a beneficial effect on carcass backfat.

Adipose Tissue↗

PubMed1988

Evidence that DNA topoisomerase I is necessary for the cytotoxic effects of camptothecin.

The budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe are both sensitive to camptothecin, an inhibitor of DNA topoisomerase I. An S. cerevisiae DNA repair mutant, rad52, is hypersensitive to the drug. In both species, topoisomerase I mutants totally lacking the enzyme are completely resistant to the drug. A strain with a mutation leading to a temperature-sensitive topoisomerase I exhibits temperature dependence in its in vivo response to camptothecin. A strain carrying a plasmid that overproduces topoisomerase I is hypersensitive to the drug. The rad52 mutant is killed by overproduction of the enzyme, even in the absence of the drug. The response of several of these strains to camptothecin analogs, to DNA topoisomerase II inhibitors, and to other drugs is reported. The cytotoxic effects of camptothecin are discussed in terms of the drug extending the lifetime of a topoisomerase I-DNA covalent intermediate, which is recognized as DNA damage by a DNA repair system.

Camptothecin↗

PubMed1987

Purification of topoisomerase II from amsacrine-resistant P388 leukemia cells. Evidence for two forms of the enzyme.

Topoisomerase II was purified from an amsacrine-resistant mutant of P388 leukemia. A procedure has been developed which allows the rapid purification of nearly homogeneous enzyme in quantities sufficient for enzyme studies or production of specific antisera. The purified topoisomerase II migrated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis as two bands with apparent molecular masses of 180 (p180) and 170 kDa (p170); both proteins unknotted P4 DNA in an ATP-dependent manner and displayed amsacrine-stimulated covalent attachment to DNA. Staphylococcus V8 protease cleavage patterns of p170 and p180 showed distinct differences. Specific polyclonal antibodies to either p170 or p180 recognized very selectively the form of the enzyme used to generate the antibodies. Immunoblotting with these specific antibodies showed that both p180 and p170 were present in cells lysed immediately in boiling sodium dodecyl sulfate. Comparison of the purified topoisomerase II from amsacrine-resistant P388 with that from amsacrine-sensitive P388 demonstrated that each cell type contained both p180 and p170; however, the relative amounts of the two proteins were consistently different in the two cell types. The data strongly suggest that p170 is not a proteolytic fragment of p180. Thus, P388 cells appear to contain two distinct forms of topoisomerase II.

Amsacrine↗

PubMed1987

Relationship between the intracellular effects of camptothecin and the inhibition of DNA topoisomerase I in cultured L1210 cells.

Results of filter elution assays of lesions produced in the DNA of cultured L1210 cells by the antineoplastic alkaloid camptothecin support the notion that topoisomerase I is an intracellular target of this drug. One to 10 microM camptothecin induced DNA single-strand, but not double-strand, breaks when incubated with intact cells or with their isolated nuclei. Approximately one half of the strand breakage was protein concealed, as judged by filter elution. Camptothecin-induced, protein-concealed DNA strand breaks disappeared rapidly after drug removal. DNA-protein cross-links were generated by camptothecin with frequencies approximately equal to those of protein-concealed DNA strand breaks. It is likely that camptothecin can inhibit topoisomerase I in intact cells in a manner similar to that in which other antineoplastic agents such as amsacrine or teniposide inhibit topoisomerase II. DNA-breaking lesions other than those resulting from trapped topoisomerase I-DNA complexes may also be generated by camptothecin. The yields of DNA strand breaks induced by camptothecin, amsacrine, or teniposide were approximately doubled when cells were incubated for 16 h with 3-aminobenzamide, an inhibitor of poly(ADP ribosylation) of proteins, prior to 1-h exposure to the antineoplastic compounds. 3-Aminobenzamide also enhanced the cytotoxic action of camptothecin, amsacrine, and teniposide. These results suggest that protein-concealed strand breaks can be lethal lesions and that intracellular topoisomerase I and II activity may be regulated coordinately through poly(ADP ribosylation).

Animals↗

PubMed1987

Antitumor activity of bis(diphenylphosphino)alkanes, their gold(I) coordination complexes, and related compounds.

Bisphosphines related to bis(diphenylphosphino)ethane (dppe) and their gold complexes are described that are active in a spectrum of transplantable tumor models. When administered ip on days 1-5 at its maximally tolerated dose (MTD) of 40 mumol/kg, dppe reproducibly gives 100% increase in life span (ILS) in mice bearing ip P388 leukemia. Coordination of chlorogold(I) to each phosphine in dppe gave a complex that had similar activity but at a much lower dose level than dppe; the MTD for the gold(I) complex was 7 mumol/kg. Among other metal complexes of dppe, the Au(III) complex was active (greater than 50% ILS) whereas Ag(I), Ni(II), Pt(II), Pd(II), and Rh(I) complexes were inactive. Among dppe analogues, replacement of phenyl groups with ethyl or benzyl groups resulted in inactivity for both ligands and the corresponding gold complexes whereas substitution with cyclohexyl or heterocyclic ring systems yielded ligands and/or gold complexes with antitumor activity. Among substituted-phenyl dppe and dppe(AuCl)2 analogues, 3-fluoro, 4-fluoro, perdeuterio, 4-methylthio, and 2-methylthio analogues were active; 4-methyl, 3-methyl, 4-methoxy, 4-dimethylamino, and 4-trifluoromethyl analogues were marginal or inactive. Analogues in which the ethane bridge of dppe or dppe(AuCl)2 was varied between one and six carbons, unsaturated or substituted, revealed that activity was maximal with ethane or cis-ethylene. Compounds with good P388 activity were also active in other animal tumor models.

Animals↗

PubMed1987

Growth performance for four breeds of swine: crossbred females and purebred and crossbred boars.

Purebred and two-breed cross (F1) boars were mated to F1 females to produce all possible three- and four-breed cross pigs involving the Duroc, Yorkshire, Landrace and Spotted breeds. Individual postweaning average daily gain (ADG), age at 100 kg (AGE) and probed backfat thickness at 100 kg (BF) data were collected on 3,456 pigs. A total of 213 pens with an average of 15.58 pigs per pen was evaluated for postweaning feed-to-gain ratio (F/G) and average daily feed consumption (ADF). Genotype X environment interactions, specifically breed X year-season farrowed and breed X parity (for ADG), were found to be highly significant. Certain results, however, were reasonably consistent across environments. Duroc-sired pigs grew more efficiently than other sire breed groups (3.11 vs 3.21 F/G), although there were no significant differences in ADF between sire groups. Duroc-sired pigs had less BF than other three-breed cross pigs, based upon within breed of dam comparisons, suggesting differences in composition between the more efficient Duroc-sired pigs and other breed groups. Landrace-sired pigs were fatter than other sire groups. No real differences between crossbred-sired pigs and the average of contemporary pure-bred-sired pigs were apparent for F/G, ADF, ADG, AGE, or BF. Assuming paternal heterosis to be zero, these results suggested recombination effects to be negligible for postweaning performance traits. Apart from via direct genetic effects, mating crossbred rather than purebred boars to females of different breeding should have little or no impact on feedlot performance of offspring produced.

Animals↗

PubMed1987

Effect of level of protein and supplemental choline on reproductive performance of gilts fed sorghum diets.

A total of 214 gilts was used (two trials) to determine the effect of protein level and choline supplementation during gestation on weight gain, conception rate and subsequent reproductive performance. The gilts were fed either a 12 or 16% crude protein sorghum-soybean meal diet containing either a high supplemental choline level or no supplemental choline in a 2 X 2 factorial arrangement of treatments. Conception rate was not influenced by either protein or choline level. Choline supplementation increased pig weight at 42 d of age (P less than .14) and litter weight at 21 (P less than .12) and 42 d (P less than .1). Gilts fed the 16% protein diet produced larger pigs at 42 d (P less than .13) and heavier litters at birth, (P less than .1) 21 d (P less than .14) and 42 d (P less than .05) than gilts fed the 12% protein diet. A larger choline effect on litter size and pig and litter weight was observed for gilts fed the 12% protein diet than for those fed the 16% gestation diet, although the protein-choline interaction was not significant for any traits measured. The incidence of spraddle leg condition was low and was not affected by level of dietary protein or supplemental choline.

Animal Feed↗

PubMed1987

Effect of unilateral hysterectomy and ovariectomy on puberty, uterine size and embryo development in swine.

Eighty crossbred gilts were assigned randomly to treatments: 1) removal of an ovary and ipsilateral uterine horn (UHO) at 130 d of age and removal of the remaining ovary and uterine horn 12 d post-puberty; 2) UHO at 130 d of age, mated and reproductive tracts recovered when slaughtered at 30 d of gestation; 3) UHO 12 d post-puberty, mated and slaughtered at 30 d of gestation and 4) unoperated controls that were mated and slaughtered at 30 d of gestation. Age of puberty was not affected by treatments. Gilts in treatment 1 had a mean ovulation rate at the pubertal estrus comparable to gilts in treatment 3. But, gilts in treatments 2 and 3 had 16% fewer (P less than .01) corpora lutea at 30 d of gestation than control gilts. Length and weight of the remaining uterine horn at 12 d post-puberty for gilts treated at 130 d of age were similar to the averages of gilts left intact. Gilts with one uterine horn had 2.2 fewer live embryos at 30 d of gestation than control gilts (P less than .01). But, the proportion of corpora lutea represented by live embryos did not differ significantly among treatments. Gilts with one uterine horn had 1.1 fewer live embryos (P less than .15) after adjustment for number of corpora lutea, less uterine space occupied by each embryo (P less than .01) and less total placental membrane per embryo (P less than .05) than control gilts.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1987

Characterization of a subline of P388 leukemia resistant to amsacrine: evidence of altered topoisomerase II function.

Sensitive (P388/S) and amsacrine-resistant (P388/amsacrine) sublines of P388 leukemia were cloned in vitro and tested for differential chemosensitivity against a panel of drugs. P388/amsacrine, resistant both in vivo and in vitro to amsacrine, was cross-resistant to other putative topoisomerase II inhibitors including teniposide, etoposide, bisantrene, and doxorubicin. P388/amsacrine, was however, as sensitive as cloned P388/S to camptothecin, an inhibitor of topoisomerase I. The pattern of cross-resistance suggested that an alteration in topoisomerase II may be involved in the resistance of P388/amsacrine to these drugs. No differences in the uptake of amsacrine were detected between the two sublines. Cross-resistance to vinblastine was evident in P388/amsacrine; however resistance to vinblastine was associated with alterations in uptake or efflux of the drug. The number of protein-concealed single-strand breaks induced in whole cells by amsacrine, teniposide, bisantrene, and camptothecin was measured. Diminished numbers of strand breaks in the resistant subline were consistent with decreases in DNA-protein crosslinks. In the absence of drug treatment, resistant cells sustained approximately one-half as many single-strand breaks and DNA-protein crosslinks as the sensitive cells during preparation of nuclei. As measured by the P4 phage DNA unknotting assay, 0.35 M NaCl nuclear extracts from P388/S contained approximately 2.3-fold more topoisomerase II catalytic activity than did extracts from P388/amsacrine. The amount of protein that immunoreacted with a specific antibody to calf thymus topoisomerase II was also decreased in the resistant cells. These data suggest that alterations in topoisomerase II which lead to differential drug sensitivities are partially responsible for the resistance of P388/amsacrine to a specific group of drugs.

Amsacrine↗

PubMed1986

Correlation of the in vitro cytotoxic and in vivo antitumor activities of gold(I) coordination complexes.

A series of gold(I) coordination complexes including analogues of the antiarthritic agent auranofin 1 were evaluated for in vitro cytotoxic potency against both B16 melanoma cells and P388 leukemia cells and in vivo antitumor activity against P388 leukemia in mice. A number of the complexes showed potent cytotoxic activity in vitro and antitumor activity in vivo, with the phosphine-coordinated gold(I) thiosugar complexes demonstrating the greatest in vitro and in vivo activity. The data compiled for 63 complexes of the general structural formula LAuX provide the basis for the following observations: potent in vitro cytotoxic activity is observed for substituted (phosphine) gold complexes, lack of potency in vitro correlates well with lack of antitumor activity, potent cytotoxicity in vitro is not necessarily predictive of activity in vivo, in vivo antitumor activity is generally optimized by ligation of Au(I) with a substituted phosphine and a thiosugar.

Animals↗

PubMed1986

Genetic parameter estimates for reproductive traits of male and female littermate swine.

Reproductive traits of purebred and crossbred pigs produced in a four-breed diallel mating system using the Duroc, Landrace, Spotted and Yorkshire breeds were collected for five consecutive farrowing seasons (two farrowing seasons/year) beginning in fall 1976. Paternal half-sib heritabilities and genetic correlations for testicular traits (120 boars from 36 sires), serum testosterone (TE) and luteinizing hormone (LH) concentrations before and after treatment with gonadotropin releasing hormone (GnRH; 131 boars from 37 sires) and breeding performance traits (151 boars from 38 sires) were estimated. Heritability estimates were generally small to moderate except for sperm/gram testis (SGT), LH concentrations before (LHO) and at 3 h (LH3) after treatment with GnRH (.73 +/- .48, .61 +/- .46 and 1.19 +/- .45, respectively). A large positive genetic correlation was found for LHO with LH3 (.94 +/- .39), while a negative relationship existed for LH3 with TE concentrations at 3 h after GnRH injection. The genetic correlation between a boar's average first service conception rate and average conception rate also was significant (.82 +/- .54). Genetic correlations among littermate traits would suggest that selection for decreased age at puberty in gilts could cause an increase in LH concentrations in boar offspring, before and after GnRH injection, and may also have adverse effects on their ability to settle females. Selection for increased weight at puberty of gilts could cause TE concentrations of boar offspring to increase while having little effect on their breeding performance.

Animals↗

PubMed1986

Cellular pharmacology of mu-[1,2-bis(diphenylphosphino)ethane]bis[(1-thio-beta-D-gluco pyranosato-S)gold(I)]: a novel antitumor agent.

SK&F 102912 (mu-[1,2-bis(diphenylphosphino)ethane]bis[(1-thio-beta-D- glucopyranosato-S)gold(I)], [(Autg)2(dppe)]) has shown reproducible and significant activity in transplantable murine tumor models and represents a structurally unique class of antineoplastic agents. A number of in vitro studies were performed to elucidate the cellular pharmacology of this gold-containing complex. [(Autg)2(dppe)] is a potent cytotoxic agent in vitro as demonstrated by its ability to inhibit the clonogenic capacity of a variety of tumor cell lines following a brief exposure to the drug. Cell-cycle analysis using HL-60 cells showed that low concentrations (2 microM) of [(Autg)2(dppe)] induced an S-phase block and higher concentrations induced a secondary block at the G1/S boundary. [(Autg)2(dppe)] had several effects on DNA metabolism and structure including preferential inhibition in cells of DNA synthesis (relative to RNA and protein synthesis) and the production of DNA single- and double-strand breaks as measured by alkaline elution. The cytoxic mechanism of this gold complex appears to be distinct from that of the monophosphine-gold complex auranofin.

Antineoplastic Agents↗