Biomedical subjects

C S Baxter

Publications and source records attributed to C S Baxter.

At least 19 recordsLinked to original sources

PubMed1992

Stimulation of TGF-beta 1 mRNA concentration in mouse skin treated with benzo[a]pyrene.

Topical application of benzo[a]pyrene (B[a]P) at dose rates of 32 or 64 micrograms/week to the dorsal skin of female Swiss (ICR) mice resulted in a marked and rapid increase in concentration of RNA for transforming growth factor beta 1 (TGF-beta 1) in epidermis. Two RNA species 1.9 and 2.5 kb, detected by a mouse TGF-beta 1 cDNA probe, were coordinately expressed. The concentration of these species appeared to be maximal 6-12 h after application, and returned to control levels after 48 h. A second, less intense maximum was observed 72-96 h after treatment. Similar effects were observed in CD-1 and HRS (both hr/hr and hr/+) mice, which are also sensitive to B[a[] tumorigenesis. In comparison with 32 and 64 micrograms/week a dose rate of 16 micrograms/week was essentially without activity in increasing TGF-beta 1 RNA concentration. All three dose rates induced an increase in epidermal RNA for ornithine decarboxylase, however, and with kinetics similar to those observed with the potent tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate. The results obtained support other findings made in this laboratory, that at high dose rates above 16 micrograms tumorigenesis by B[a]P involves a strong tumor-promoting component. The latter further appears to be mediated by increased TGF-beta 1 expression.

Animals↗

PubMed1992

Carba-prostacyclin inhibits 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced transformation in sensitive murine epidermal JB6 cells.

The ability of carba-prostacyclin (cPGI2), a stable analog of prostacyclin (PGI2), to inhibit 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced transformation of JB6 cells was investigated. JB6 cells sensitive (P+) and resistant (P-) to TPA-induced transformation to anchorage-independent growth were plated in soft agar in the presence or absence of cPGI2 for 14-21 days. Transformation frequencies were determined by recording colony numbers. cPGI2 was found to inhibit TPA-induced transformation of P+ cells in a dose-dependent fashion with 1 microM cPGI2 producing approximately 50% inhibition of colonies in soft agar. Our findings are consistent with the hypothesis that TPA-induced transformation in JB6 cell variants is mediated by PGI2 via regulation of adenylate cyclase activity and cAMP accumulation, with resultant inhibition of expression of the transformed phenotype, reflected in anchorage-independent growth.

Animals↗

PubMed1992

JB6 murine epidermal cell lines sensitive and resistant to 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced transformation exhibit differential arachidonic acid metabolism in response to TPA and the calcium ionophore A23187.

In a study of arachidonic acid metabolism in murine epidermal JB6 cells, promoter-sensitive (P+) and promoter-resistant (P-) variants, labeled with [3H]arachidonic acid, were treated successively with 12-O-tetradecanoylphorbol-13-acetate (TPA) and the calcium ionophore A23187. Released radiolabel was separated by HPLC and identified by coelution of standards. Prostacyclin release was then quantified by radioimmunoassay for 6-keto prostaglandin (PG)F1 alpha. A23187 alone resulted in a small but significant enhanced release of radiolabel from both cell variants (0.7 +/- 0.2% for P- and 0.6 +/- 0.3% for P+ cells; mean +/- SD). Treatment with TPA and subsequent treatment with A23187 resulted in a synergistically enhanced release of radiolabel from both cell variants (4.1 +/- 0.8% for P- and 3.4 +/- 0.9% for P+ cells) relative to that with either agent alone. Although the predominant product for each treatment regimen was prostaglandin E2 (PGE2), the TPA-resistant cells (P-) released significantly more 6-keto PGF1 alpha, a stable breakdown product of PGI2, than did the TPA-sensitive (P+) cells. These results indicate differential arachidonic acid metabolism between JB6 cell variants resistant and sensitive to TPA-induced transformation.

6-Ketoprostaglandin F1 alpha↗

PubMed1991

Effect of tumor-promoting agents on density and morphometric parameters of mouse epidermal Langerhans and Thy-1+ cells.

Topical application of tumor-promoting agents to the dorsal skin of female SENCAR mice on a twice-weekly basis resulted in a reduction in density per unit area of bone marrow-derived Thy-1+ dendritic cells. Activity was observed for well-established tumor-promoting doses of promoting agents of several different chemical types, including 12-O-tetradecanoylphorbol-13-acetate (TPA, diterpene diester), anthralin (dihydroxyanthrone), and n-dodecane (n-alkane). A reduction in density of the same cells was also observed on the basis of the asialoGM1 lipid as a surface marker after TPA treatment. No parallel effect was observed for epidermal Langerhans (Ia+) cells, the second major epidermal immunofunctional cell type, except in the case of anthralin, a finding which is consistent with the reported toxicity of this agent. The stage 2 promoting agent mezerein was unique in inducing a consistent increase in Langerhans cell densities, but did not affect the density of Thy-1+ cells when applied for a prolonged period unless applied following four doses of TPA. In contrast to the SENCAR strain, the promotion-resistant Balb/c and C56BL/6 strains showed no response with respect to TPA-induced reduction of Thy-1+ cell density. In addition to effects on density, the above tumor-promoting agents induced morphological changes in both Thy-1+ and Langerhans cells. When these changes were placed on a quantitative basis by the calculation of shape and area fraction parameters, marked and significant effects were observed for the above agents, but not for the partial promoting agent mezerein nor the non-promoting phorbol diester 4-O-methyl-TPA. The effects of TPA were largely blocked by the potent anti-promoting agent fluocinolone acetonide, moreover. These findings further support an important role for quantitative and qualitative alterations in dendritic epidermal cells in tumor promotion.

Alkanes↗

PubMed1991

Benzo[a]pyrene-induced skin damage and tumor promotion in the mouse.

Epidermal cell kinetics and DNA adduct levels, and skin morphological changes were measured following weekly topical applications for 29 weeks of high (16, 32 and 64 micrograms) benzo[a]pyrene (B[a]P) doses to female ICR/Harlan mice, in order to investigate the relationship of these parameters to the timing, incidence and morphology of the elicited tumors. During the tumor latency period, [3H]thymidine labeling index, mitotic index, epidermal cell stacking, incidence of pyknotic and dark basal keratinocytes and labeled mitoses were periodically measured, as were nuclear area and DNA content. DNA adducts in skin epidermis were measured by an ELISA method over a period of 9 weeks of single weekly applications of 64, 32, 16 or 8 micrograms B[a]P. There was an initial linear increase in DNA adducts with dose in the epidermis but the increase was much less steep above 32 micrograms/week. This did not correlate with the sharp rise in tumor response above the 32 micrograms/week dose rate. Cell kinetic changes in response to the 64 micrograms/week dose reached a plateau in the first few weeks of the tumor latent period. There was little epidermal hyperplasia but an associated dose-dependent increase in [3H]thymidine labeling index, mitotic index and incidence of pyknotic and dark cells. This evidence indicated that B[a]P produced extensive cytotoxicity and cell death with regenerative proliferation under these conditions. Giant keratinocytes occurred in all dose groups. Analysis of a labeled mitosis curve indicated that B[a]P produced a G2/M block. There was a marked inflammatory response in the dermis at all B[a]P doses. Mice were observed weekly for tumor formation. Virtually all of the tumors were papillomas on initial appearance and required an average of 8 weeks to convert to carcinomas. The substantial cell killing and regenerative proliferation, and the correspondence between the dose-response patterns for epidermal damage and tumors, together with the initial appearance of tumors in the benign form, a characteristic of the action of promoting agents, provided evidence that the tissue damage associated with the high dose levels of B[a]P used in this study reflected tumor-promoting activity in this mouse epidermal tumorigenesis model. The implication of the results for mathematical models of tumor formation are discussed.

Animals↗

PubMed1989

Effect of the ornithine decarboxylase inhibitor alpha, alpha-difluoromethylornithine on phorbol diester-induced inhibition of murine B lymphocyte differentiation.

The tumor-promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA) inhibits the differentiation of murine B lymphocytes to antibody-producing plasma cells, in unfractionated spleen cell cultures or enriched B lymphocyte cultures. To determine the role of polyamines in TPA-induced inhibition, unfractionated splenic lymphocytes, in culture with antigen, were incubated with alpha, alpha-difluoromethylornithine (DFMO, 0.10 mM), an irreversible inhibitor of ornithine decarboxylase (ODC). DFMO prevented the TPA-induced inhibition of antibody forming cell number in a 5-day in vitro immunization procedure as measured by a hemolytic plaque assay. In enriched B lymphocyte cultures, however, DFMO had no comparable effect. DFMO did not prevent TPA-induced inhibition of antibody production in unfractionated spleen cell cultures but itself inhibited the amount of antibody produced. Putrescine (0.1 mM), added on day 4 of immunization, reversed DFMO inhibition of antibody production but did not enable DFMO to prevent the TPA-induced inhibition. These findings suggest that TPA-induced inhibition of plasma cell number can be mediated indirectly through effects on T lymphocytes and/or macrophages or directly through effect on B lymphocytes.

Animals↗

PubMed1989

Comparative histomorphometric changes in SENCAR mouse epidermis in response to multiple treatments with complete and stage-specific tumor promoting agents.

Responses of various cells of the epidermis and dermis to topically applied agents have been implicated in the mechanism of multistage mouse tumorigenesis. These responses have been discussed almost entirely in the context of a single promoter treatment, although tumor expression is dependent on multiple applications. Responses of keratinocytes, epidermal dendritic non-keratinocytes and dermal leukocytes were therefore recorded following multiple topical applications of the potent complete tumor-promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA). In order to assess the importance of the response of individual cell types to the mechanisms and stages of promotion, responses to TPA were compared with those to agents with low complete promoting activity, but significant activity in individual stages of multistage promotion models. These included 4-O-methyl-TPA, a stage 1 promoting agent, mezerein and n-dodecane, stage 2 promoting agents of apparently different mechanism of action, and ethyl phenylpropiolate (EPP), a highly inflammatory stage 3 promoting agent. In agreement with previous findings, TPA induced a persistent epidermal hyperplasia and an increase in dark keratinocytes, although a similar finding was made for EPP and n-dodecane. The response to n-dodecane was significantly delayed, however, and that to EPP was accompanied by focal epidermal destruction and inflammation. The response to n-dodecane contrasted with that found for mezerein, supporting the suggestion that their mechanisms of action are distinct. Multiple treatments of 4-O-methylTPA caused no increase in dark cells, and mezerein induced no increase in numbers of pyknotic cells, whereas increases were expected in both cases on the basis of single dose experiments. Of the agents examined, only TPA induced a decrease in pale dendritic epidermal cells in the absence of marked toxicity, supporting the previous proposal that prolonged effects on this cell type are important in the promotion process. Some degree of persistent dermal leukocyte infiltration was observed with all agents excepting 4-O-methylTPA, although the extent of the response and its cellular characteristics appeared strongly dependent on the agent applied. In the case of TPA small mononuclear cells, neutrophils and macrophages all provided significant contributions to the total infiltrate. A similar phenomenon was observed with n-dodecane and EPP, with an additional increase in eosinophils which was not observed with TPA. Mezerein differed from both TPA and n-dodecane in inducing a significant increase only in eosinophils. As reported previously for single applications, prolonged TPA application caused a change in morphology and a considerable decrease in numbers of Thy-1+ epidermal dendritic cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetone↗

PubMed1988

Alterations in histone phosphorylation in HL-60 cells specific to monocytic differentiation.

In order to examine the role of histone phosphorylation in regulation of the pathway of HL-60 cell differentiation, cells were labelled with [32P]phosphoric acid and histones fractionated by two-dimensional polyacrylamide gel electrophoresis. The monocytic inducer 12-O-tetradecanoylphorbol-13-acetate (TPA) was found to specifically stimulate phosphorylation of histone H2B in a concentration-dependent manner. At a concentration of 100 mM, H2B phosphorylation was stimulated 2.3-fold after 4 h. A second monocytic inducer 1,25-dihydroxy-cholecalciferol (100 nM) also induced phosphorylation specifically in histone H2B. In contrast, the granulocytic inducers DMSO (1.5%) or retinoic acid (1 microM) did not increase phosphorylation in any histone species.

Cell Differentiation↗

PubMed1988

Critical comparison of histological and morphometric changes in SENCAR mouse epidermis in response to n-dodecane, 12-O-tetradecanoylphorbol-13-acetate and mezerein.

n-Dodecane, a previously little-studied tumor-promoting agent and mezerein, a diterpenoid natural product, have both been reported to have activity primarily in Stage II of two stage tumor promotion in SENCAR mouse skin. Histological changes in this tissue were therefore investigated in response to these agents in order to determine whether changes could be identified which were common to Stage II promotion by both compounds, and specific in this respect compared to those induced by the complete promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA). All three agents were applied at doses which have previously been found active in multistage tumorigenesis studies in this strain. A single dose of 50 mg dodecane induced no increase in the number of interfollicular cell layers or epidermal thickness, nor any observable inflammation, 6-144 h after application. In contrast, marked increases were predictably observed with TPA and mezerein, maximal responses occurring after 48-72 h. n-Dodecane induced no increase in the number of keratinocytes with dense cytoplasm and increased affinity for basophilic dyes (dark cells), only TPA demonstrating this activity. The alkane likewise did not increase the number of pyknotic basal keratinocytes indicating that toxicity would not account for the low Stage I activity of this agent in the way proposed for mezerein which was the most active in this respect, inducing a significant increase 48 h after treatment. Like TPA and mezerein, n-dodecane induced a significant increase in large intra-mitochondrial densities. Forty-eight to seventy-two hours after application, dodecane induced a significant decrease in the numbers of dendritic epidermal cells, a response which was also observed for TPA and mezerein, although occurring somewhat more rapidly. All three agents appeared to induce these cells to retract their characteristic processes. After four applications of n-dodecane the number of epidermal cell layers and mitotic index were equal to or greater than those observed with TPA. These findings show that in SENCAR epidermis the previously uncharacterized tumor-promoting agent n-dodecane induced essentially no histologic changes in mouse skin in common with mezerein, a second agent with activity primarily in Stage II of two stage tumor promotion, which were not also shown by the complete promoter TPA. The only characteristic specific to Stage II promoting agents therefore remains an inability to induce increased numbers of dark cells. In most other respects dodecane induced responses similar to those observed for TPA, although a distinctly different temporal dependence was noted in several cases.(ABSTRACT TRUNCATED AT 400 WORDS)

Alkanes↗

PubMed1988

Qualitative and quantitative effects on epidermal Langerhans (Ia+) and Thy-1+ dendritic cells following topical application of phorbol diesters and mezerein.

Repeated twice-weekly applications of promoting doses of 12-O-tetradecanoylphorbol-13-acetate (TPA) to the dorsal skin of female adult SENCAR mice led to a reduction in numbers per unit area of epidermal Thy-1+ dendritic cells. Although no parallel effect was observed on Ia+ Langerhans cells, a concurrent reduction of dendritic morphology of both cell types was observed. Topical administration of TPA (2 micrograms) twice weekly for 4 or 8 weeks led to reductions in Thy-1+ cell numbers of 52 and 61%, respectively, whereas a single treatment was without effect. Similar effects were observed in animals initiated with 10 nmol 7,12-dimethylbenz[a]anthracene suggesting that the response was specific to the promotion, rather than the initiation, phase of two-stage tumorigenesis. All initiated animals bore tumors after 16 promoter treatments. In comparison with the potent promoter TPA, the weak overall, but stage-specific promoters 4-O-methylTPA and mezerein showed no effects on number or morphology of either dendritic cell type. These findings are therefore consistent with an important role for quantitative and qualitative alterations in epidermal non-keratinocytes of immune function in tumor promotion.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed1988

DNA strand breaks in embryonic mouse limbs exposed to methylnitrosourea.

The alkaline elution assay was used to monitor DNA single strand breaks in hindlimbs of embryonic mice exposed to methylnitrosourea (MNU) on day 13 of gestation, a stage of development at which the embryotoxicity of MNU was minimal. DNA breakage was maximal by 1 h after exposure, and rapidly declined to control levels by 8 h. There was a reduction of fetal weights in the MNU-treated litters, but no increase in embryomortality. No malformations of the limbs were observed. The data are discussed in relation to an earlier study, where the same dose of MNU administered on day 11 of gestation was highly teratogenic and produced a substantially different pattern of DNA breakage in hindlimbs.

Animals↗

PubMed1987

Direct inhibition of murine B lymphocyte differentiation by 12-O-tetradecanoylphorbol-13-acetate.

The tumor-promoting agent, 12-O-tetradecanoylphorbol-13-acetate (TPA) inhibits the humoral immune response of lymphocytes to antigen. To test the hypothesis that this inhibition is due to a direct effect upon B lymphocytes, splenic lymphocytes or murine B lymphocytes, enriched by 'panning' splenic lymphocytes onto anti-IgM-coated petri dishes, were immunized in vitro with the thymus/accessory cell-independent antigen trinitrophenyl lipopolysaccharide (TNP-LPS) with or without TPA. The number of anti-TNP antibody-forming cells present in both lymphocyte populations after 5 days was almost completely inhibited to the same degree by TPA. These data unambiguously show that TPA can directly inhibit the differentiation of B lymphocytes to antibody-forming cells.

Animals↗

PubMed1987

Specific inhibition of phorbol diester-induced granulocyte-macrophage progenitor cell (GM-CFU) differentiation by lipoxygenase inhibitors.

The induction of differentiation of SENCAR murine granulocyte-macrophage precursor cells (GM-CFU) by the tumor-promoting phorbol diester 12-O--tetradecanoyl-phorbol-13-acetate (TPA) was inhibited by agents reported to inhibit specific aspects of arachidonic acid metabolism. These agents included phospholipase A2 inhibitors, and eicosatetraynoic acid (ETYA), a competitive inhibitor of arachidonic acid oxygenases. Whereas inhibitors reported specific for lipoxygenases were also active, no comparable effect was observed for inhibitors of the corresponding cyclooxygenase. These findings are therefore consistent with the hypothesis that induced differentiation of GM-CFU cells is regulated by products of arachidonic acid metabolism formed principally via lipoxygenase activity.

5,8,11,14-Eicosatetraynoic Acid↗

PubMed1987

Mechanism of mouse skin tumor promotion by n-dodecane.

Application of the alkane n-dodecane to the dorsal skin of 6-8 week old female SENCAR mice initiated with 10 nmol dimethylbenz[a]anthracene led to papilloma formation in the majority of treated animals. Compared to the potent phorbol diester 12-O-tetradecanoylphorbol-13-acetate (TPA), n-dodecane was several orders of magnitude less potent on a dose basis, and maximal papilloma response required more extended application (22 weeks for 50 mg dodecane compared to 12 weeks for 2 micrograms TPA). In two-stage promotion experiments n-dodecane appeared to act as a stage II promoting agent at appropriate doses, being comparable in activity to mezerein--an agent with well-characterized activity of this type. Dodecane, unlike mezerein, did not induce the formation of a significant number of pyknotic cells, however, suggesting that the weak promoting activity of dodecane in stage 1 was not a result of toxicity. In comparison with TPA, both mezerein and n-dodecane at promoting doses induced less sustained hyperplasia in SENCAR mouse skin, a finding also consistent with the proposal that n-dodecane is principally active in stage II of two-stage promotion models. Both agents induced ornithine decarboxylase activity in SENCAR mouse skin, the maximal induction being observed at apparently the same time after a single application.

Acetone↗

PubMed1986

Phorbol diester synergistically stimulates agonist-induced lipoxygenase product formation in murine macrophages.

Murine peritoneal macrophages were preincubated with 12-O-tetradecanoylphorbol-13-acetate (100 ng/ml) and then exposed to various concentrations of calcium ionophore A23187. This combined treatment resulted in an elevated release of arachidonic acid metabolites that was synergistic in nature. Resolution of the [3H]products released into the medium revealed that the effect on the cyclooxygenase products 6-keto PGF1 and PGE2 were additive to less than additive (A23187 concentration dependent), the production of lipoxygenase products was synergistically enhanced, and the largest effect was on 12-HETE production.

Animals↗

PubMed1986

Decreased sulfation of cellular chondroitin sulfate in response to activators of protein kinase C.

The sulfation of cellular chondroitin sulfate in human promyelocytic leukemia HL60 cells was inhibited by a number of phorbol diesters, which concurrently induced differentiation into monocytic cells. Inhibition was dependent on concentration, and was 90% complete at 10 nM 12-0-tetradecanoylphorbol-13-acetate (TPA), the most active ester. Maximal effects were seen within 2-4 hours following initiation of treatment. The degree of inhibition observed correlated well with the ability of the esters to induce differentiation, and with their reported affinity for a "receptor", identified as protein kinase C associated with certain lipids. Chondroitin sulfation was also inhibited in cells treated with sn-1,2-dioctanoylglycerol, a lipid which is considered to be an endogenous activator of protein kinase C. Our findings therefore indicate that monocytic differentiation of HL60 cells occurs subsequent to reduced glycosaminoglycan sulfation via activation of the calcium-activated, phospholipid-dependent protein kinase.

Cell Differentiation↗

PubMed1986

Topical application of a tumor promoter induces proliferation of an adherent cell population in murine spleen.

The tumor-promoting phorbol esters have proven to be potent immunomodulatory agents in vitro. It has not been possible to assess the role of phorbol ester-induced alterations of immune function in tumor promotion however, due to a lack of in vivo studies. Studies were therefore designed to assess proliferative responses of murine leukocytes after in vivo exposure to these agents. Topical application of 12-O-tetradecanoylphorbol-13-acetate (TPA) was found to cause an increase in division of murine spleen cells. The effect was dose-related up to 14 micrograms per application and generally reached a peak two days after TPA application. Cell separation experiments suggested TPA was acting on an adherent cell population distinct from splenic lymphocytes. Inflammatory reactions to TPA followed a similar time-course to that observed for spleen cell proliferation. The increased levels of proliferation observed could, therefore, be due to increased division of neutrophil and/or macrophage precursors residing in the murine spleen and appeared also to be associated with the inflammatory reaction induced by TPA.

Administration, Topical↗

PubMed1985

Specific stimulation of histone H2B and H4 phosphorylation in mouse lymphocytes by 12-O-tetradecanoylphorbol 13-acetate.

This report demonstrates that the potent tumor promoter 12-O-tetradecanoylphorbol 13-acetate rapidly stimulates the phosphorylation of histones H2B and H4 in a cell cycle-independent manner. This effect was observed in primary cultures of BALB/c mouse splenocytes, a population of noncycling, G0 cells which are not stimulated to divide by 12-O-tetradecanoylphorbol 13-acetate treatment alone. The biological nature of this cell system allowed the analysis of histone phosphorylation in the absence of a background of cell cycle-dependent changes and in response to a nonmitogenic agent. The phosphorylation of H2B was determined with high resolution through the use of two-dimensional gel electrophoresis. In contrast to 12-O-tetradecanoylphorbol 13-acetate, the mitogen from pokeweed did not induce stimulation of H2B and H4 phosphorylation, but did, however, elicit increases in the phosphorylation of histones H1, H2A, and H3, in parallel with changes in rate of DNA synthesis.

Animals↗