PubMed Health⌕ Search

Biomedical subjects

C J Conti

Publications and source records attributed to C J Conti.

At least 127 records · Page 7Linked to original sources
PubMed1989

Induction of terminal differentiation-resistant epidermal cells in mouse skin and in papillomas by different initiators during two-stage carcinogenesis.

Carcinogen treatment of normal mouse epidermal cells causes some cells, if cultured under the appropriate conditions, to continue to proliferate instead of terminally differentiate, forming foci at 37 degrees C in medium with a calcium level above 0.1 mM. We have examined these Calcium (Ca)-resistant cells formed in the skin of SENCAR mice after treatment with the carcinogen initiator 7,12-dimethylbenz[a]anthracene (DMBA) followed by tumor promotion with 12-O-tetradecanoylphorbol-13-acetate (TPA). Although in our previous studies TPA promotion initially increased the size but reduced the number of foci caused by the carcinogen initiator N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), TPA promotion of DMBA-treated mice increased the size but had no effect on the number of foci. Papillomas resulting from DMBA plus TPA treatment contained many rapidly growing Ca-resistant cells, corroborating our earlier results with MNNG. Permanent cell lines prepared from papilloma-derived foci formed squamous cell carcinomas in nude mice after relatively short periods in culture. These data provide further evidence that Ca-resistant cells may be papilloma (and perhaps carcinoma) precursors in vivo. In addition, since TPA tends to reduce the number of early Ca-resistant cells caused by MNNG but not by DMBA, this may at least partially explain why treatment with DMBA plus TPA is much more effective in producing papillomas in SENCAR mice than is treatment with MNNG plus TPA.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1989

Sequential trisomization of chromosomes 6 and 7 in mouse skin premalignant lesions.

Using a direct cytogenetic technique, we identified a nonrandom trisomy of chromosome 6 in 12 of 12 aneuploid mouse skin papillomas and in 10 of 11 squamous cell carcinomas induced by chemical carcinogenesis. The second most common abnormality observed was trisomy of chromosome 7 found in most dysplastic papillomas and in 10 of 11 carcinomas. The two trisomies were the only abnormalities found in all aneuploid papillomas and in several carcinomas. Mutation at codon 61 of the Ha-ras gene, which resides on chromosome 7, was also a common feature of the tumors sampled. Extensive homology exists between mouse chromosome 6 and human chromosome 7, the trisomy of which was recently suggested as a primary cytogenetic event in several human epithelial cancers. We propose a multistep model of tumor progression in which a sequence of specific nonrandom chromosomal abnormalities appear to be required for malignant transformation.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1989

Multistage chemical carcinogenesis protocols produce spindle cell carcinomas of the mouse skin.

Spindle cell carcinomas were identified using polyacrylamide gel electrophoresis and immunoblotting of proteins extracted from paraffin-embedded tissue sections. Immunohistochemistry using rabbit monospecific antisera against the mouse 55 kd keratin polypeptide also identified these tumors. A group of 53 SENCAR mice initiated with 7,12-dimethylbenz[a]anthracene (DMBA) and promoted with 12-O-tetradecanoylphorbol-13-acetate (TPA) yielded, after one year, four spindle cell carcinomas (0.07/mouse), whereas another group of 31 mice treated with a three-stage carcinogenesis protocol (initiation with DMBA and promotion for 10 weeks with TPA followed by 10 weeks of benzoyl peroxide) gave rise to six spindle cell carcinomas (0.19/mouse). The number of keratin-positive tumor cells and the intensity of the immunostain varied markedly, but all tumors expressed the 55 kd polypeptide. Although other carcinogens, mainly UV radiation, have been able to induce spindle cell tumors, the present data indicate that chemical carcinogenesis protocols are able to induce the formation of this highly malignant variant of skin carcinoma.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1988

Tumor progression in Sencar mouse skin as a function of initiator dose and promoter dose, duration, and type.

The influence of initiator dose and promoter dose, duration, and type on the progression of papillomas to carcinomas was examined in Sencar mice. A good dose-response relationship for promotion of papilloma formation by 12-O-tetradecanoylphorbol-13-acetate (TPA) [following initiation with 6.5 micrograms of 7,12-dimethylbenz(a)anthracene (DMBA)] was observed in the range of 0.125 to 2.0 micrograms/mouse. A maximal papilloma response was induced with 2 micrograms/mouse (24 papillomas/mouse). When adjusted for mortality, the carcinoma incidence after 60 wk of promotion was essentially the same (approximately 80%) for doses above 0.5 micrograms/mouse. In a related experiment, mice were given an initiation dose of either 2 or 20 micrograms of DMBA followed by applications of 2 micrograms of TPA for 3, 5, 7, or 60 wk. Papilloma formation was proportional to length of treatment, with a maximum of 29 papillomas/mouse (20-micrograms initiating dose of DMBA) and 10 papillomas/mouse (2-micrograms initiating dose of DMBA) occurring between 10 and 15 wk of promotion. In this experiment, the carcinoma incidence was clearly proportional to the duration of promoter treatment at the low initiation dose of DMBA. The carcinoma incidence, on the other hand, was similar (approximately 70%) in groups of mice given an initiation dose of 20 micrograms of DMBA and promotion treatment for greater than or equal to 5 wk. Thus, the initiator dose had a dramatic effect on the outcome of these experiments. Additional experiments were performed to compare tumor progression with the anthrone promoter, chrysarobin. At optimal promoting doses, chrysarobin treatment produced a maximum number of papillomas that was approximately 1/3 that produced by TPA (6.4 versus 17.0 papillomas per mouse, respectively). However, the carcinoma response was very similar in these two treatment groups, confirming previous work from this laboratory. In addition, chrysarobin treatment following 10 wk of TPA promotion did not enhance the progression of preexisting papillomas to carcinomas. The data presented in this paper are consistent with a model in which several types or stages of papillomas are initially produced during two-stage carcinogenesis in mouse skin with different probabilities of progressing to carcinomas. However, the data indicate that optimal doses of promoter and initiator exist and can influence interpretation of tumor progression studies in mouse skin.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1988

Sequential development of aneuploidy, keratin modifications, and gamma-glutamyltransferase expression in mouse skin papillomas.

To elucidate the role and timing of expression of different premalignant and malignant markers in tumor promotion, we correlated alterations in keratin patterns and gamma-glutamyltransferase (GGT) expression with the chromosomal status of individual mouse skin papillomas. Papillomas were induced by 7,12-dimethylbenz[a]anthracene initiation and 12-O-tetradecanoylphorbol-13-acetate promotion. Individual tumors were randomly sampled at 20 and 35 weeks of promotion. Each tumor was cytogenetically analyzed and serial paraffin sections were used for GGT detection, immunoblotting, and immunohistochemistry studies. Monospecific antibodies elicited against keratins K1 (Mr 67,000) and K14 (Mr 55,000) were used to analyze keratin modifications. Most tumors at 20 weeks of promotion, although exhibiting aneuploidy, still presented high levels of the K1 differentiation-associated keratin. Later during promotion those tumors bearing the highest aneuploidy indexes were those that showed a marked decrease in or absence of the K1 protein. Furthermore, those same tumors with the highest levels of genomic alterations also exhibited foci of GGT activity. These results support the idea that the majority of papillomas under continuous promotion are progressing toward malignancy. Aneuploidy seems to precede detectable keratin modifications, and GGT activity appears to be the latest marker to be expressed.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1988

In vivo behavior of murine epidermal cell lines derived from initiated and noninitiated skin.

The in vivo behavior of cell cultures derived from normal and carcinogen-treated mouse epidermis was studied by implanting the cultures in a s.c. vascularized bed protected by a silicone chamber. Cells derived from normal adult mouse epidermis as well as cells derived from tumor-promoter-treated skin were unable to grow in these systems. Conversely, cell lines derived from skin initiated with single doses of N-methyl-N'-nitro-N-nitrosoguanidine or 9,10-dimethyl-1,2-benzanthracene proliferated in these chambers, reforming an epithelial structure. The type of structure in the chambers varied, ranging from formation of almost normal epithelia to atypical invasive behavior. The variable in vivo behavior among the different cell lines may be attributed to the initiation agent, the number of passages of the cultures, random genetic events, the strain of mouse, or a combination of these factors. Most of the cell types used in this study and all the cell lines that were able to grow in these chambers were selected for resistance to Ca-induced terminal differentiation. However, resistance to terminal differentiation according to the Ca2+ switch does not always correlate with the ability to grow in the chambers, since cell lines derived from spontaneous foci of resistance failed to grow in this system. These studies showed some of the possibilities of the SC silicone chambers to study the histogenic potential of cell lines derived from carcinogen-treated epidermis. This system also appears suitable to study the complex relationship between epidermal cells and specialized (dermal) stroma.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1988

Cytogenetic profile of mouse skin tumors induced by the viral Harvey-ras gene.

Several investigators have postulated that the activation of the Harvey-ras (rasH) gene is the initiating event in mouse skin carcinogenesis. In support of this speculation, Roop et al. (Nature, 323, 822-824, 1986) have shown that papillomas developed from skin grafts of cultured keratinocytes in which the v-rasH gene had been introduced by a defective retroviral vector. Using the same technique we have analyzed the cytogenetic profile of primary mouse epidermal cultures bearing the activated ras gene and of the tumors generated by skin grafts of the same cells. The epidermal cultures infected with the replication-defective virus which incorporated the v-rasH gene into a packaging-defective Moloney murine leukemia virus did not show any detectable karyotypic abnormalities. These cells presented a diploid/tetraploid configuration similar to uninfected controls. Furthermore, papillomas produced by skin grafts of these cells also showed euploid chromosomal profiles. On the other hand, two carcinomas that arose spontaneously from pre-existing papillomas showed 80% aneuploid cells and aneuploid stem lines. It appears, from these results, that the activated rasH gene is able to generate benign lesions without the involvement of further gross genomic alterations, but that such alterations may be associated with malignant conversion.

Animals↗
PubMed1988

Polyacrylamide gel electrophoresis and immunoblotting of proteins extracted from paraffin-embedded tissue sections.

We show in this communication that polyacrylamide gel electrophoresis (PAGE) and immunoblotting of proteins can be performed using one to two 5-7 micron paraffin sections of tissues fixed in non-cross-linking fixatives (acetone, alcohol, or modified Carnoy's solution). Proteins for study were extracted from paraffin sections of mouse foot pad and liver. The presence of unaltered keratin polypeptides in tissues fixed with either acetone or alcohol was demonstrated in gels stained with Coomassie brilliant blue. The preservation of their antigenic determinants was demonstrated with immunoblotting. Furthermore, the immunoreactivity of soluble proteins, such as albumin, remained unaltered in immunoblots obtained from paraffin-embedded mouse liver sections. These data indicate that tissues embedded and stored in paraffin are useful for the above-mentioned biochemical and immunological studies and may therefore be an important technique for diagnostic pathology or retrospective studies.

Animals↗
PubMed1987

Characterization of skin tumor promotion and progression by chrysarobin in SENCAR mice.

The characteristics of the skin tumor promotion response with anthrone derivatives has been further examined in SENCAR mice. Chrysarobin (1,8-dihydroxy-3-methyl-9-anthrone) was an effective skin tumor promoter when applied twice weekly with dose-dependent increases in both papillomas and squamous cell carcinomas between 25 and 100 nmol/mouse. A similar dose-response relationship for papilloma and carcinoma formation was observed when chrysarobin was applied once weekly. Interestingly, chrysarobin was approximately twice as active as a skin tumor promoter when applied once weekly versus twice weekly. Doses of 25,100, and 220 nmol/mouse gave maximal papilloma responses of 2.90, 8.15, and 9.38 versus 0.73, 4.70, and 5.42 papillomas/mouse, respectively, in mice initiated with 25 nmol 7,12-dimethylbenz(a)anthracene. Thus, unlike 12-O-tetradecanoylphorbol-13-acetate (TPA), where a twice weekly application frequency is optimal, application of anthrone promoters such as chrysarobin once weekly is a more optimal frequency for papilloma development. Chrysarobin was also a much more effective skin tumor promoter when the start of promotion was delayed by an additional 10 weeks. Thus, groups of mice initiated with 10 nmol 7,12-dimethylbenz(a)anthracene and having promotion started in either the 3rd or the 13th week after initiation had maximal responses of 5.6 or 11.0 papillomas/mouse, respectively. In addition, the rate of papilloma development was faster in the delayed promotion group. The progression of papillomas to carcinomas was examined in all chrysarobin-treated groups and compared with three groups of mice treated with 3.4 nmol TPA. After 60 weeks of promotion, the anthrone promoter-treated groups had carcinoma:papilloma ratios 2.5 to 5.0 times higher than the TPA-treated groups. This was due primarily to the fact that similar carcinoma responses were observed in both anthrone- and TPA-treated mice at optimal promoting doses whereas the papilloma responses were significantly lower in the former groups. The data suggest that anthrone derivatives are very efficient tumor promoters. The results are further discussed in terms of mechanisms of skin tumor promotion.

9,10-Dimethyl-1,2-benzanthracene↗
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

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

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

Regulation of cultured rat vaginal epithelial cells by 17 beta-estradiol and progesterone.

We have previously described a technique to obtain short-term cultures of epithelial cells from Wistar rat vaginae. In order to improve the efficiency and life span of these cultures, in the present study we have cultured the vaginal cells with lethally irradiated 3T3 cell feeder layers. Under this condition, cells can grow for several weeks while retaining epithelial characteristics and can eventually be subcultured. The proliferative effect of the ovarian hormones in these cultures was studied using two different approaches, [Methyl-3H]Thymidine (3HTdr) incorporation and increase in cell number. Both assays indicated a proliferative effect of 17 beta-estradiol and progesterone at physiological concentrations. This proliferative effect was also shown in feeder layer-free cultures, ruling out an indirect effect through the mesodermal cells. The capacity of the hormones to modify terminal differentiation in the culture was also studied, using colony stratification as an indicator of differentiation. Progesterone and fetal calf serum had an inhibitory effect on terminal differentiation, whereas 17 beta-estradiol induced a stimulatory action. This culture model allowed us to show a direct effect of the ovarian hormones on vaginal cells in vitro and seems to be a useful model to study hormone-cell interactions in vitro.

Animals↗
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

Immunodetection of involucrin in lesions of the oral mucosa.

The immunoperoxidase method for involucrin detection was applied to the study of the maturation of epithelial lesions of the oral mucosa that included specimens of leukoplakia, lichen planus, verrucous carcinoma, carcinoma in situ and invasive carcinoma. Areas of orthokeratinized, parakeratinized, and non-keratinized normal mucosa were also studied. Normal orthokeratinized epithelia showed intracytoplasmic or pericellular staining in the suprabasal epithelial layers in a pattern similar to that of the normal epidermis. Parakeratinized and non-keratinized epithelia were less stained. Intense staining was observed in leukoplakia, whereas the staining of lichen planus was less intense but exhibited a more homogeneous pericellular staining pattern than leukoplakia. Verrucous carcinoma was markedly and very irregularly stained. Carcinomas in situ and invasive carcinoma exhibited a slightly positive and patchy reaction. The distribution patterns of involucrin in the lesions correlated very well with the degree of epithelial differentiation. In addition, irregular patchy distribution correlated with the degree of atypia, and was especially evident in carcinomas.

Carcinoma↗
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↗