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T J Slaga

Publications and source records attributed to T J Slaga.

At least 109 records · Page 6Linked to original sources
PubMed1987

Terminal differentiation-resistant epidermal cells in mice undergoing two-stage carcinogenesis.

We have used an in vivo-in vitro approach to investigate the cellular aspects of two-stage skin carcinogenesis. Female SENCAR mice initiated with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) were promoted twice weekly with 12-O-tetradecanoylphorbol-13-acetate (TPA). Epidermal cultures from untreated or TPA-treated mice had few focus-forming cells resistant to calcium-induced terminal differentiation. Cultures from mice treated with MNNG alone formed numerous foci. Brief promotion (four TPA treatments) of MNNG-treated mice produced fewer but statistically larger foci, suggesting that TPA was selecting against more slowly growing cells. MNNG plus TPA-treated mice with very early papillomas produced more and larger foci than those due to MNNG treatment alone, suggesting that the papillomas may have comprised calcium-resistant cells. These cells may indeed be initiated cells since a permanent cell line arising after MNNG plus brief TPA treatment eventually formed histological papillomas in vivo. If calcium-resistant cells are initiated, then there were many more initiated cells in the skin (with or without TPA treatment) than papillomas expected, implying that either some initiated cells never formed papillomas, or that a significant accumulation of initiated cells had already occurred in the skin within 2 weeks of MNNG treatment. Subsequent TPA promotion of these cells apparently produced a toxic response that passively selected for more rapidly growing initiated cells, which eventually accumulated into papillomas.

Animals↗
PubMed1987

Metastasis from squamous cell carcinomas of SENCAR mouse skin produced by complete carcinogenesis.

The incidence of metastasis was evaluated in female SENCAR mice after induction of squamous cell carcinomas by repetitive applications of either benzo [a] pyrene (B [a] P) or N-methyl-N'-nitro-N-nitrosogaunidine (MNNG). Between 41 and 50 weeks 50% of the animals with carcinomas in the B [a] P group had metastases, whereas 20% had metastases in the MNNG group. Very few metastases were observed before 40 weeks of treatment. The major site of metastasis was the lungs; however, metastatic tumors were also found in lymph nodes, adrenal glands and kidneys.

Animals↗
PubMed1987

Initiation, promotion and complete carcinogenesis by N-methyl-N'-nitro-N-nitrosoguanidine or ethylnitrosourea in the Sencar mouse skin tumorigenesis model.

Five doses of either N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or ethylnitrosourea (ENU) were tested as complete carcinogens, tumor initiators and tumor promoters in the SENCAR skin tumorigenesis model. As tumor initiators, MNNG-induced papillomas at all doses tested, while ENU was active from 10-40 mumol. As complete carcinogens, MNNG from doses of 0.5-5.0 mumol and ENU from doses of 10 mumol-40 mumol were potent inducers of both papillomas and carcinomas indicating that these agents are active as both tumor initiators and tumor promoters.

Animals↗
PubMed1987

A new medium for primary cultures of adult murine epidermal cells: application to experimental carcinogenesis.

A culture medium has been formulated to support the long-term proliferation and differentiation of normal murine epidermal cells in primary cultures. This formulation is based upon MCDB-151 modified by the addition of 1.2 mM Ca2+, 1% fetal bovine serum (FBS) or 0.1% purified bovine serum albumin (BSA), trace elements, and growth supplements. For 4-12 weeks, the cultures demonstrate desmosomes, keratin fibers, basal proliferation, and stratification characteristic of keratinocytes. The addition of 12-O-tetradecanoylphorbol-13-acetate (TPA) resulted in approximately 70% cellular detachment, but with continued treatment, the remaining cells proliferated to confluence and stratified. This culture medium has application to a number of studies related to skin carcinogenesis in the murine model system that have not previously been possible.

Animals↗
PubMed1987

Progressive dysplasia and aneuploidy are hallmarks of mouse skin papillomas: relevance to malignancy.

We report a systematic histopathologic study of papillomas at different times during promotion, correlating the results with those from cytogenetic analysis of the same tumors. Papillomas were induced in SENCAR mice by two-stage carcinogenesis (7,12-dimethylbenz[a]anthracene and phorbol 12-myristate 13-acetate). Individual tumors were randomly sampled at different times during promotion, and histopathologic and cytogenetic studies were carried out on every tumor. Early during promotion (10 weeks), most papillomas were well-differentiated hyperplastic lesions with mild or no cellular atypia. No tumors showed severe dysplastic changes. By 20 weeks of promotion, a dramatic drop had occurred in the number of lesions with no dysplasia. Most of the tumors presented moderate dysplasia, and some already showed severe dysplastic changes. At later stages (30-40 weeks), most of the papillomas were classified as moderately or severely dysplastic papillomas, and several were considered to be intrapapillomatous carcinomas. This histopathologic evaluation was supported by nuclear measurements performed on papillomas at different time points. Chromosomal abnormalities followed a similar trend. Papillomas seem to start as diploid lesions, but between 10 and 20 weeks of promotion, hyperdiploid cells can be observed in almost every tumor. In some cases the stem line was taken over by aneuploid clones. At 40 weeks of promotion, all papillomas were aneuploid, most of them with hyperdiploid stem lines. A positive correlation was found between the histological and cytogenetic studies, with the most aggressive and atypical tumors being the more aneuploid. These results support the idea that most, if not all, papillomas are truly premalignant lesions in different stages of the potential progression toward malignancy. Chromosomal abnormalities might play an important role in the sequence of events leading to malignancy.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1987

Characterization of an inbred strain of the SENCAR mouse that is highly sensitive to phorbol esters.

An inbred strain of SENCAR mice was developed that is more sensitive to two-stage skin carcinogenesis protocols than the outbred parental stock. These mice, registered as SSIN/UTSP, were compared to the SENCAR in initiation-promotion protocols using the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) twice weekly for 22 weeks at dose levels of 0.5, 1.0 and 2.0 micrograms. With 0.5 microgram of TPA, the number of papillomas in the SSIN was 3-fold higher than in the SEN-CAR; the tumor incidence was 100 versus 60%, respectively. Similar, although less dramatic, results were found with 1.0 and 2.0 micrograms of TPA. In two-step promotion protocols using TPA twice weekly for 2 weeks and 2 micrograms mezerein twice weekly for 15 weeks the SSIN produced approximately 50% more tumors than the SENCAR at 0.5 microgram of TPA. At 2 micrograms of TPA the tumor response between the two mice was not significantly different. Epidermal hyperplasia was considerably greater in the SSIN at 0.5 microgram of TPA as was ornithine decarboxylase activity. These TPA-sensitive inbred mice should be useful in investigations on the biochemical and genetic factors involved in skin tumor promotion.

Animals↗
PubMed1986

Phosphorylation of histones is stimulated by phorbol esters in quiescent Reuber H35 hepatoma cells.

Histones isolated from Reuber H35 rat hepatoma cells treated with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) were examined for possible alterations in phosphorylation. Incorporation of 32P orthophosphate into individual acid-extracted histones was monitored by autoradiography and scintillation counting of polyacrylamide gels or by reverse-phase high performance liquid chromatography. Treatment of quiescent H35 cells (arrested by serum starvation) with submicromolar doses of TPA resulted in a rapid and specific increase in phosphorylation of histones H2B and H1(0). Smaller increases in phosphorylation were observed for H4. No significant change in phosphorylation of the major H1 histones or H2A were observed after 1 h of treatment. The phosphorylation was TPA dose-dependent, with a maximum increase of approximately 14-fold for H2B, 11-fold for H1(0), and 2-fold for H4 achieved at 0.8 M TPA. The nonpromoting parent compound phorbol did not induce any of these changes. Furthermore, the mitogenic hormone insulin did not cause a similar pattern of histone phosphorylation, suggesting that the effect observed was not due to a general mitogenic response in the H35 hepatoma cells. Addition of 8-Br-cAMP also failed to reproduce the effect of TPA on histone phosphorylation, suggesting that cAMP-dependent protein kinases are not likely to be involved in mediating this response to TPA.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1986

A direct cytogenetic technique for mouse skin carcinomas and papillomas.

A method was developed for obtaining direct chromosome preparations from SENCAR mouse skin tumors induced by chemical carcinogenesis protocols. Papillomas and squamous cell carcinomas were mechanically dispersed immediately after resection and were placed in a modified Hanks' solution with collagenase, trypsin, hyaluronidase, bovine albumin, and Colcemid. Total exposure to Colcemid did not exceed 1 hr. Metaphases were obtained in 100% of the analyzed specimens, allowing chromosome counting screening for double minutes and, in 50% of the cases, useful G-banded slides. The technique described has produced, for this type of tumor, a higher number of successful G-banded preparations than other previously reported methods for solid tumors. This procedure may be applicable for the study of human solid tumors that are histologically similar to our murine model, such as squamous cell carcinoma of cervix or lung.

Animals↗
PubMed1986

Malignant progression of mouse skin papillomas treated with ethylnitrosourea, N-methyl-N'-nitro-N-nitrosoguanidine, or 12-O-tetradecanoylphorbol-13-acetate.

Mouse skin tumors were induced by a single topical application of 7,12-dimethylbenzanthracene (DMBA), followed by biweekly promotion with 12-O-tetradecanoylphorbol-13-acetate (TPA). After 20 weeks of promotion, mice were treated twice weekly for 2 weeks with either ethylnitrosourea (ENU), N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or TPA. Thereafter all groups were treated biweekly with TPA. The ENU-treated group had a higher percentage of animals with carcinomas and developed 217% more cumulative carcinomas per group than TPA-treated controls. The percentage of mice with carcinomas and the cumulative number of carcinomas per group in MNNG-treated mice was higher than TPA-treated controls but was less than ENU-treated mice. The ratio of cumulative carcinomas to cumulative papillomas in ENU treated, MNNG-treated and TPA-treated mice was 16%, 9% and 6%, respectively. Histological examination of tumors remaining at the termination of the experiment revealed the presence of keratoacanthomas, some of which stained positive for gamma-glutamyltransferase (GGT), in the ENU-treated and MNNG-treated, but not the TPA-treated groups. The fact that no new papillomas developed during the progression stage indicated that enhanced carcinogenesis resulted from the progression of pre-existing tumors. Enhanced progression of benign skin tumors in mice by only a few treatments of an agent may serve as a potential model for studies into the mechanisms and the inhibition of malignant progression. The model also allows for a comparison of the potency of agents in enhancing malignant progression.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1986

Aneuploidy, an early event in mouse skin tumor development.

It has been suggested that chromosomal alterations can play a role in mouse skin carcinogenesis. These changes have been proposed as a possible mechanism of action of tumor promoters and they have also been related to the conversion of benign papillomas to carcinomas. However, direct evidence showing the chromosomal constitution of these tumors has not been previously described. Here we show the presence of aneuploid cells in very early papillomas and the eventual displacement of the diploid stem line by aneuploid clones at later stages. Squamous cell carcinomas (SCC) induced by the same protocol were highly aneuploid. These results suggest that genomic imbalance produced by aneuploidy may be related to the malignant conversion of the benign papillomas.

Aneuploidy↗
PubMed1986

The intracellular fate of [3H]benzo[a]pyrene diol epoxide diastereomers localized in vivo in SENCAR mouse epidermis: a quantitative electron microscopic study.

The intracellular localization of topically applied anti and syn diastereomers of racemic benzo[a]pyrene-7,8-diol-9,10-epoxide (BPDE) was investigated in SENCAR mouse epidermis using conventional electron microscopic (EM) autoradiography. The shaved backs of adult mice were treated in vivo with the active skin tumor initiator anti [3H]BPDE or with its stereoisomer syn [3H]BPDE, which is inactive as an initiator. After 3, 12 and 24 h of exposure, thin sections of the treated skin were prepared for high resolution autoradiography to determine the intracellular distributions of radioactivity bound within the epidermis. For each treatment and exposure time, the interfollicular area of epidermis examined by EM encompassed 350-700 keratinocytes, and extended from the basal lamina to the innermost keratinizing layer at a magnification that permitted basal and suprabasal cell populations to be distinguished in situ following autoradiography. Statistical analysis of the distributions of silver grains localized over keratinocytes at 3-, 12- and 24-h timepoints revealed that 27.2% of the carcinogenic anti [3H]BPDE grains occurred over epidermal nuclei, which was significantly more (P less than 0.001) than 20.0% of the non-carcinogenic syn [3H]BPDE grains associated with nuclei at all timepoints examined. In the basal cell population at the 3-h time point 33% of both racemic diastereomers were found within nuclei; at 12 and 24 h of exposure the nuclear association of anti [3H]BPDE grains increased (P less than 0.01) to 39% and 47% respectively, while the nuclear-associated syn [3H]BPDE grains remained at 29% and 33%. The localization of [3H]BPDE observed in our experiments suggests that the epidermal cell nucleus may be an important intracellular target for anti BPDE.

Administration, Topical↗
PubMed1986

Elevated expression of Ha-ras is an early event in two-stage skin carcinogenesis in SENCAR mice.

Alterations in the expression of the Ha-ras oncogene were investigated in SENCAR mice epidermis at various stages of initiation and promotion during two-stage skin carcinogenesis in SENCAR mice. Adult SENCAR mice were treated with 200 nmol of the (+) enantiomer of benzo[a]pyrene 7,8-diol 9,10-epoxide-anti (BPDE-anti), a potent initiating agent, followed by repetitive treatments with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). Other mice were 'sham'-initiated with the (-) enantiomer of BPDE-anti, which is inactive as a tumor initiator. Polyadenylated RNA was isolated from pre-tumor epidermis and tumors at eight different stages of tumorigenesis and analyzed for changes in Ha-ras expression using Northern blot hybridization. Significantly enhanced levels of Ha-ras RNA were observed in TPA-promoted papillomas as early as 7 weeks after initiation. Only trace amounts of Ha-ras RNA were present in untreated epidermis or epidermis treated with the (+) or (-) enantiomer followed by 2-12 treatments with TPA (pre-papilloma stage). Southern blot hybridization of tumor DNA indicated that the increased expression of the Ha-ras oncogene was not due to gene amplification. We conclude that elevated levels of Ha-ras expression can occur at an early stage of tumor development in mouse epidermis in vivo and may play a role in tumorigenesis.

Animals↗
PubMed1986

Allogeneic transplantation of normal epidermal cells and squamous cell carcinomas in SENCAR mice.

A method used in our laboratory for the transplantation of single cell suspensions of epidermis and skin carcinomas is described. Silicone chambers were placed into a subcutaneous granulation tissue formed by the implantation of a glass disk. The skin was closed on top of the chamber by using Michael autoclips. Cells were injected 24 to 48 hr later through the skin using a syringe and needle. The neoformation of epithelia with adnexa was observed when newborn epidermal cells were injected. When a fresh cell suspension of squamous cell carcinoma was used, a typical differentiated carcinoma was formed within 2 to 4 weeks. However, after 4 weeks, some of the grafts showed mild signs of rejection. The technique described is a useful system to transplant squamous cell carcinomas and can also be used as a rapid assay for malignancy.

Animals↗
PubMed1986

Comparison of the tumorigenic response of SENCAR and C57BL/6 mice to benzo(a)pyrene and the inter-experimental variability over a three-year period.

SENCAR and C57BL/6 mice were compared for their ability to produce tumors after benzo(a)pyrene [B(a)P] initiation and 12-O-tetradecanoylphorbol-13-acetate (TPA) promotion. SENCAR mice initiated with 101 micrograms/mouse B(a)P and promoted with TPA (2 micrograms/mouse, twice weekly) produced large numbers of papillomas, whereas C57BL/6 mice produced none after 26 weeks of promotion. Continued treatment of the B(a)P-initiated C57BL/6 mice with TPA up to 52 weeks did not induce any papillomas nor did higher doses of B(a)P. Application of increased doses of TPA (10 micrograms/mouse, twice weekly) to B(a)P-initiated C57BL/6 mice (404 micrograms/mouse) for 50 weeks produced few papillomas. Substantial papilloma formation in C57BL/6 mice was observed after weekly treatment with B(a)P (101 micrograms/mouse), with maximal production occurring at weeks 39 to 41 of treatment. In contrast, SENCAR mice treated according to the same protocol produced an equivalent response with maximal papilloma formation occurring 12 to 13 weeks earlier. Therefore, C57BL/6 mice exposed to B(a)P are capable of producing papillomas under certain experimental conditions. The inter-experimental variability of B(a)P-induced (50.5 micrograms/mouse) papilloma formation after 30 weeks of TPA promotion (2 micrograms/mouse, twice weekly) was examined in SENCAR mice over a 37-month period. Low statistical variation was observed in papilloma multiplicity, papilloma incidence, or papilloma latency. Male and female SENCAR mice produced equal values in the three parameters: 4.4 +/- 1.6 papillomas/mouse, 87% +/- 10% of the mice bearing papillomas, and 9.6 +/- 1.3 weeks (time at which 10% of the mice bore papillomas). The numbers of papillomas per mouse did not follow a Poisson distribution.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1986

SENCAR mouse skin tumorigenesis model versus other strains and stocks of mice.

The SENCAR mouse stock was selectively bred for eight generations for sensitivity to skin tumor induction by the two-stage tumorigenesis protocol using 7,12-dimethylbenz(a)anthracene (DMBA) as the initiator and 12-O-tetradecanoylphorbol-13-acetate (TPA) as the promoter. The SENCAR mouse was derived by crossing Charles River CD-1 mice with skin-tumor-sensitive mice (STS). The SENCAR mice are much more sensitive to both DMBA tumor initiation and TPA tumor promotion than CD-1, BALB/c, and DBA/2 mice. An even greater difference in the sensitivity to two-stage skin tumorigenesis is apparent between SENCAR and C57BL/6 mice when using DMBA-TPA treatment. However, the SENCAR and C57BL/6 mice have a similar tumor response to DMBA-benzoyl peroxide treatment, suggesting that TPA is not an effective promoter in C57BL/6 mice. The DBA/2 mice respond in a similar manner to the SENCAR mice when using N-methyl-N-nitro-N-nitrosoguanidine (MNNG)-TPA treatment. The SENCAR mouse model provides a good dose-response relationship for many carcinogens used as tumor initiators and for many compounds used as tumor promoter. When compared to other stocks and strains of mice, the SENCAR mouse has one of the largest data bases for carcinogens and promoters.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1986

Effects of chronic topical application of 12-O-tetradecanoylphorbol-13-acetate on the skin and internal organs of SENCAR mice.

Repetitive topical applications of 2 micrograms 12-O-tetradecanoylphorbol-13-acetate (TPA) twice weekly for 37 to 52 weeks induced a sustained epidermal hyperplasia, hyperplasia of hair follicles, and increased dermal cellularity in SENCAR mice. In addition, after 52 weeks of protracted promoter treatment most animals developed generalized amyloidosis involving liver and spleen, as well as interstitial nephritis. Severe pyelonephritis and papillary necrosis were also frequently seen. Reactive lymphoid hyperplasia was also a frequent finding. Chronic administration of TPA is not an innocuous treatment affecting only the interfollicular epidermis. The general effect of the promoter on the animals was a marked decrease in their longevity, probably through impairment of the immune system.

Administration, Topical↗
PubMed1986

Enhanced malignant progression of mouse skin tumors by the free-radical generator benzoyl peroxide.

Chemical carcinogenesis in mouse skin can be divided into the processes of initiation, promotion, and progression. The free-radical generator benzoyl peroxide is moderately active during the promotion stage. Repetitive treatment of mouse benign skin tumors (papillomas) with benzoyl peroxide (20 mg, twice weekly) increased the number of cumulative carcinomas per group by 325% and the number of keratoacanthomas by 44% compared to tumor-bearing Sencar mice treated with the promoter 12-O-tetradecanoylphorbol-13-acetate. The lack of increase in the number of cumulative papillomas per group due to benzoyl peroxide treatment suggests that benzoyl peroxide enhanced the progression of preexisting papillomas. The ability of benzoyl peroxide to enhance the progression of benign tumors to cancer should be considered when determining the human risk from exposure to this widely used chemical agent; in addition, biological assays specifically testing malignant progression may be essential and beneficial for determining an agent's carcinogenic risk.

9,10-Dimethyl-1,2-benzanthracene↗