Tumor Promoting Agent 12-O-tetradecanoylphorbol-13-acetate Research Articles

No Involvement of Lysophosphatidic Acid Receptor-3 in Cell Migration of Mouse Lung Tumor Cells Stimulated by 12-O-Tetradecanoylphorbol-13-acetate

The tumor promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA) stimulates cell migration of several tumor cells. Recently, we reported that loss of lysophosphatidic acid (LPA) receptor-3 (LPA3) enhanced cell migration of murine lung tumor LL/2 cells. In the present study, we investigated whether LPA3 is involved in cell migration of mouse lung tumor cells stimulated by TPA. Exogenous LPA3 gene (Lpar3)-expressing (LL/2-a3) cells and LL/2-AB cells as a vector control generated from LL/2 cells were used. In a cell migration assay, TPA treatment significantly stimulated cell migration of LL/2-AB and LL/2-a3 cells, while the cell migration abilities of LL/2-a3 were markedly lower than those of LL/2-AB cells. Using quantitative real-time reverse transcription (RT)-polymerase chain reaction (PCR) analysis, no effect of TPA treatment on the expression levels of LPA1, LPA2 and LPA3 genes was detected in either type of cells. These results suggest that the LPA3 may not be involved in the enhanced migration ability by TPA in mouse lung tumor cells.

Nordihydroguaiaretic Acid from Creosote Bush (Larrea tridentata) Mitigates 12‐O‐Tetradecanoylphorbol‐13‐Acetate‐Induced Inflammatory and Oxidative Stress Responses of Tumor Promotion Cascade in Mouse Skin

Nordihydroguaiaretic acid (NDGA) is a phenolic antioxidant found in the leaves and twigs of the evergreen desert shrub, Larrea tridentata (Sesse and Moc. ex DC) Coville (creosote bush). It has a long history of traditional medicinal use by the Native Americans and Mexicans. The modulatory effects of topically applied NDGA was studied on acute inflammatory and oxidative stress responses in mouse skin induced by stage I tumor promoting agent, 12-O-tetradecanoylphorbol-13-acetate (TPA). Double TPA treatment adversely altered many of the marker responses of stage I skin tumor promotion cascade. Pretreatment of NDGA in TPA-treated mice mitigated cutaneous lipid peroxidation and inhibited production of hydrogen peroxide. NDGA treatment also restored reduced glutathione level and activities of antioxidant enzymes. Elevated activities of myeloperoxidase, xanthine oxidase and skin edema formation in TPA-treated mice were also lowered by NDGA indicating a restrained inflammatory response. Furthermore, results of histological study demonstrated inhibitory effect of NDGA on cellular inflammatory responses. This study provides a direct evidence of antioxidative and anti-inflammatory properties of NDGA against TPA-induced cutaneous inflammation and oxidative stress corroborating its chemopreventive potential against skin cancer.

Evaluation of Diuron (3-[3,4-dichlorophenyl]-1,1-dimethyl urea) in a Two-stage Mouse Skin Carcinogenesis Assay

Diuron (3-[3,4-dichlorophenyl]-1,1-dimethyl urea) is an herbicide with carcinogenic activity in rats and mice, which have developed respectively urothelial and mammary gland tumors in long-term studies. Accordingly, diuron has been categorized as a "likely human carcinogen" by the U.S. Environmental Protection Agency. Although the carcinogenesis-initiating activity of diuron has been reported in an early initiation-promotion mouse skin study, its genotoxic potential has been disputed. It is necessary to clarify the mode of action through which it has caused rodent neoplasia and verify its relevance to humans. Herein, two experiments were developed to verify the initiating and promoting potentials of diuron in a twenty-three- and a twenty-one-week-long mouse skin carcinogenesis protocol. In one, dimethylsulfoxide (DMSO) was the solvent for the herbicide; in the other, acetone was the alternative solvent in order to verify whether DMSO had inhibitory influence on a potential cutaneous carcinogenic activity. The adopted schedule for the tumor-promoting agent 12-O-tetradecanoylphorbol 13-acetate (TPA) resulted in skin ulcers, which demonstrates the need for careful selection of TPA dose levels and frequency of application in this model. In both studies, diuron did not exert any influence on the skin carcinogenesis process, in contrast with results already reported in the literature.

12-O-Tetradecanoylphorbol-13-acetate (TPA)-induced c-Jun N-terminal Kinase (JNK) Phosphatase Renders Immortalized or Transformed Epithelial Cells Refractory to TPA-inducible JNK Activity

c-Jun N-terminal kinase (JNK) regulates gene expression in response to various extracellular stimuli. JNK can be activated by the tumor promoting agent, 12-O-tetradecanoylphorbol-13-acetate (TPA) in normal human oral keratinocytes but not in human keratinocytes that have been immortalized (HOK-16B and HaCaT) or transformed (HOK-16B-Bap-T) nor in a cervical carcinoma cell line (HeLa). The refractory JNK activation response to TPA is not due a defect in the JNK pathway, because JNK can be activated by other stimuli, e.g. UV irradiation and an alkylating agent N-methyl-N'-nitrosoguanidine in these immortalized or transformed cells. More importantly, the refractory JNK and JNKK activation response to TPA can be restored by treatment of the cells with a combination of TPA and a protein-tyrosine phosphatase inhibitor, sodium orthovanadate. Furthermore, pretreatment of cells with TPA partially inhibited UV- or N-methyl-N'-nitrosoguanidine-induced JNK activity. These results suggest that a TPA-inducible, orthovanadate-sensitive protein-tyrosine phosphatase may specifically down-regulate JNK signaling pathway in these immortalized/transformed epithelial cells. In contrast, ERK and p38/Mpk2 are not regulated by this TPA-induced phosphatase. This putative protein-tyrosine phosphatase appears to be JNK pathway-specific.

Replacement of Ser657 of protein kinase C-alpha by alanine leads to premature down regulation after phorbol-ester-induced translocation to the membrane.

Protein kinase C (PKC) is activated at the cell membrane by interacting with both the acidic lipid phosphatidylserine and the second messenger diacylglycerol. A direct activation of the kinase is also possible by substituting diacylglycerol with phorbol esters such as the tumor promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA). Transphosphorylation of the activation loop followed by autophosphorylation at sites located on various domains of the protein have been suggested to be required as permissive activation of the alpha and beta isoforms of PKC [Cazaubon, S., Bornancin, F. & Parker, P. (1994) Biochem. J. 301, 443-448; Keranen, L. M., Dutil, E. M. & Newton, A. C. (1995) Curr. Biol. 5, 1393-1403]. Ser657, located near the C-terminus of PKC-alpha, represents a site which is very conserved among the members of the PKC protein family. Circumstantial evidence suggested that this residue represents a possible site of phosphorylation. The conversion of Ser657 to alanine caused a 70% loss of the catalytic activity as well as a drastically increased down regulation upon translocation of this isozyme to the membrane when induced by phorbol ester. The faster electrophoretic mobility of the mutant protein compared to that of the wild-type enzyme suggested that Ser657 represents a phosphorylation site.

Characterization of the gap junction protein, connexin45

Connexin45 is a gap junction protein which forms channels with unique characteristics. RNA blots demonstrated that connexin45 is expressed in a number of cell lines including WB, SK Hep1, BHK, A7r5, CLEM, and BWEM cells. Connexin45 was further studied in BWEM cells using specific affinity-purified antibodies directed against a synthetic peptide representing amino acids 285-298 of its sequence. Immunofluorescence experiments demonstrated that the BWEM cells expressed both connexin43 and connexin45 and that these connexins colocalized. Connexin45 polypeptide, immunoprecipitated from BWEM cells metabolically labeled with [35S]-methionine, consisted of a predominant 48 kD polypeptide. Connexin45 and connexin43 contained radioactive phosphate when immunoprecipitated from BWEM cells metabolically labeled with [32P]-orthophosphoric acid. This phosphate label was removed from connexin45 by alkaline phosphatase digestion. Treatment of BWEM cells with the tumor promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA) inhibited intercellular passage of microinjected Lucifer yellow. While TPA treatment induced phosphorylation of connexin43 in these cells, it reduced the expression of connexin45. Furthermore, the connexin45 expressed after TPA treatment was not phosphorylated. These results suggest that treatments which alter protein phosphorylation may regulate connexin43 and connexin45 in BWEM cells by different mechanisms.

Effect of the ornithine decarboxylase inhibitor α,α-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.

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)

Effects of various chemical agents on the transformation of rat fibroblasts by an activated c-Ha-ras oncogene.

We have previously reported that the potent tumor-promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA) and a factor from fetal calf serum (FCS) markedly enhance the transformation of mouse C3H 10T1/2 and Rat 6 fibroblasts, when added to cultures following transfection with plasmid pT24 DNA that contains an activated c-Ha-ras oncogene. In the present study, we examined possible enhancing or inhibiting effects of various chemicals on the transformation of Rat 6 fibroblasts by T24 DNA when tested in the presence of calf serum, calf serum plus TPA or FCS. We found that, like TPA, the chemicals mezerein, 1-oleoyl-2-acetylglycerol, and phospholipase C increased the yield of T24-induced foci, thus further implicating protein kinase C as a critical constituent in this process. Low concentrations (10(-6)-10(-7)M) of retinoic acid (both trans and 13-cis) also stimulated cell transformation. Several compounds inhibited T24-induced transformation. These included nontoxic concentrations of the calcium ionophore A23187, indomethacin, and epsilon-amino-n-caproic acid. Compounds that failed to exert a significant reproducible effect included vasopressin, vitamin D3, selenium, antipain, Bowman-Birk inhibitor, vitamin B12, epidermal growth factor, platelet-derived growth factor, insulin, and transferrin. These findings suggest that this simple in vitro system might be useful for detecting enhancers and inhibitors of ras oncogene-induced cell transformation and also elucidating their mechanisms of action.

The effect of calcium and 3′,5′cAMP on the induction of ornithine decarboxylase activity in bone and bone cells

The interrelationship between mediators of hormone action (cAMP and calcium) and induction of ornithine decarboxylase (ODC) activity was investigated in bone and bone cells. Stimuli that enhanced the cAMP level in both osteoblastlike (BL) and osteoclastlike (CL) cells and in intact calvaria only induced ODC activity in the isolated cell populations. Moreover, addition of fresh medium induced ODC activity in BL cells and not in calvaria, without an effect on the cAMP level. The tumor-promoting agent 12-O-tetradecanoyl-phorbol-13-acetate (TPA), however, induced in both systems ODC activity, without an effect on the cAMP concentration. From these data it was concluded that induction of ODC activity may not be mediated by cAMP. The role of Ca in ODC induction was also investigated. Experiments with BL cells, incubated in media with various Ca concentrations, as well as experiments with the Ca blocker D600 showed that basal and hormone-induced ODC activity was dependent on the intracellular Ca concentration. A regulatory role of cAMP, however, in concert with Ca, cannot be excluded.

Stimulation of ornithine decarboxylase activity and DNA synthesis by phorbol esters or bile acids in rat colon.

The changes of colonic epithelial ornithine decarboxylase (ODC) activity and DNA synthesis following intrarectal administration of a tumor-promoting agent, 12-O-tetradecanoylphorbol-13-acetate (TPA), or various bile acids to male noninbred rats were studied. A single instillation of TPA, at a dose as low as 16 nmol, led to a significant (about 10-fold) increase in colonic ODC activity. Peak ODC activity was observed at 4 hr, and the enzyme activity returned to the control level about 24 hr after intrarectal TPA. This pattern was almost the same as that observed after sodium deoxycholate treatment. TPA showed more potent induction of ODC activity than deoxycholate, although the maximal induction was greater in the case of deoxycholate treatment. Both TPA and deoxycholate stimulated DNA synthesis at 2 days after intrarectal instillation, after an initial depression at 4-12 hr. A structure-activity study of 26 bile acids revealed that 5 beta-cholanoic acid with alpha-hydroxy groups in two of the 3 alpha, 7 alpha, 12 alpha positions and 5 beta-cholanoic acid with a 3 alpha-hydroxy group induced colonic ODC activity significantly, while the 3 alpha, 6 alpha-dihydroxy acid did not. Replacement of hydroxy groups by keto groups or a change from alpha to beta configuration decreased the ODC-inducing activities. Tri-substituted 5 beta-cholanoic acid derivatives, whether hydroxy or keto, did not stimulate ODC. These data indicate that a specific bile acid structure with a definite spatial relationship of the hydroxy groups is required for induction of colonic ODC activity.

Fatal lymphoproliferative disease in a common marmoset ( Callithrix jacchus) following inoculation of Ag876 strain of Epstein-Barr virus and a tumor-promoting agent: Preliminary report

We report here the development of a fatal lymphoproliferative disease in a common marmoset ( Callithrix jacchus) following inoculation of the Ag876 strain of Epstein-Barr virus (EBV) and of the tumor promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA). The marmoset showed antibodies against EBV capsid and early antigens (VCA, EA) but not against nuclear antigen (EBNA). EBNA was detected in impression smears from lymph nodes and liver, and the presence of the EBV genome was detected from the same organs by hydridization techniques. To our knowledge this is the first report of the oncogenicity of this strain of EBV. The need for further studies to assess the in vivo interaction between EBV and tumor promoting agents in the development of malignancies is stressed.

Interaction between latent papovavirus genomes and the tumor promoter TPA

The tumor promoting agent 12-O-tetradecanoylphorbol-13acetate (TPA) enhances both the transformation of rat embryo cells by adenovirus and the cloning efficiency of such cells [ 11. Induction of Epstein-Barr virus (EBV) antigens and stimulation of particle synthesis by treatment of the EBV-carrying cell lines with TPA was reported in [ 2,3]. Here we report that TPA exclusively enhances the viral DNA content of cells which harbor papovavirus genomes in a latent episomal state. On the other hand, no effect is exerted by TPA on the copy number of the viral DNA in productively infected cells and on integrated papovavirus DNA in transformed cells.

TPA-resistance in friend erythroleukemia cells: Role of membrane lipid fluidity

Summary Fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene was used to examine membrane lipid dynamics in clones of Friend erythroleukemia cells displaying sensitivity or resistance to the tumor promoting agent 12-O-tetradecanoyl-phorbol-13-acetate (TPA). Two resistant clones yielded higher fluorescence anisotropy values, indicative of decreased lipid fluidity, as compared to two sensitive clones. This difference in fluorescence anisotropy was abolished by treatment of the sensitive clones with cholesteryl hemisuccinate. The treatment also removed another phenotypic difference between the sensitive and resistant clones. Cell adherence to plastic culture dishes in the presence of TPA, a property of sensitive but not of resistant clones, was markedly diminished when the former were enriched with the cholesteryl ester. No significant differences were observed in the binding of a phorbol ester to high affinity saturable receptors between sensitive versus resistant cells. These results implicate membrane lipids as determinants of the actions of phorbol ester tumor promoters.

Analysis of developmentally homogeneous neural crest cell populations in vitro: II. A tumor-promoter (TPA) delays differentiation and promotes cell proliferation

The tumor-promoting agent 12- O-tetradecanoylphorbol-13-acetate (TPA) delayed pigmentation in developmentally homogeneous quail neural crest cell populations in vitro. Doses below 5 × 10 −8 M resulted only in a delay in pigmentation for up to 2 days without changing the rate of pigmentation, whereas intermediate doses (5 × 10 −8–10 −7 M) delayed the initiation and decreased the rate of pigmentation but had no effect on the ultimate fate. Doses of TPA above 2 × 10 −7 M irreversibly prevented pigmentation in some of the cells. TPA did not influence growth of the undifferentiated cell population. Since TPA delayed the decline in the cell proliferation rate that normally occurred with pigmentation, however, the final number of pigmented cells could be increased up to 10-fold in TPA-treated cultures without changing the homogeneity of the cultures. A part from the effects on cell differentiation, TPA caused an immediate change in cellular association within the clusters.

Epidermal growth factor, like tumor promoters, enhances viral and radiation-induced cell transformation.

The polypeptide hormone epidermal growth factor (EGF) has been shown to enhance adenovirus type 5 transformation of a cloned culture of rat embryo cells (CREF) and X-ray or u.v.-light induced transformation of 10T1/2 mouse embryo cells. In both systems, the degree of enhancement was quantitatively similar to that observed in treated rats grown in the presence of the potent tumor promoting agent 12-O-tetradecanoyl-phorbol-13-acetate (TPA). An increase in viral transformation was also observed in cells continuously exposed to phorbol esters with known promoting activity on mouse skin, but not structurally related analogs, inactive or weakly active in the two-stage mouse skin carcinogenesis assay. In addition, the appearance of transformed foci was accelerated and colonies tended to be larger in cultures grown in the presence of EGF or TPA. The present studies suggest the possibility that EGF may function as an endogenous promoter of carcinogenesis and further indicates that in vitro cell transformation systems may prove useful in identifying such agents.

Increased infectivity of oncogenic herpes viruses of primates with tumor promoter 12-O-tetradecanoylphorbol-13-acetate.

AbstractCell cultures of simian and human origin infected with two strains each of herpesvirus saimiri (HVS) and herpesvirus ateles (HVA) were compared for production of infectious virus and early and late antigens (EA, LA) in the presence and absence of the tumor promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA). Second, Epstein-Barr virus (EBV) infected human and simian lymphoblastoid cell lines of high and low cell passages were also compared for enhanced production of early antigen and virus capsid antigen (VCA) using two concentrations of TPA, owl monkey kidney (OMK) and squirrel monkey lung. Cell cultures infected with HVS and HVA with 20 ng/ml of TPA exhibited higher percentages of early and late antigen producing cells and contained 1.0-1.6 logs more virus. Such cells also had earlier cytopathic effects, larger plaques, and a 3-fold increase in the number of plaques. The TPA also enhanced HVS-EA, and LA both in OMK and human skin fibroblast (HSF) cells. The enhancement of EA was approxima...

Effects of dose and duration of treatment with the tumor-promoting agent, 12-O-tetradecanoylphorbol-13-acetate on mouse skin carcinogenesis

The effects of dose and duration of treatment with the potent tumor-promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA) on the formation of skin tumors in Charles River CD-1 mice was studied. Mice were initiated with a single application of 0.2 micromol of 7,12-dimethylbenz[a]anthracene (DMBA) in 0.2 ml acetone. Beginning two weeks after initiation, mice were treated twice weekly with various doses (0.01 - 20 nmol) of TPA in 0.2 ml acetone. Application of either 0.01 or 0.1 nmol of TPA did not elicit tumors during the 50 weeks duration of treatment. A dose-dependent increase in the number of papillomas was observed through the range of 1 to 10 nmol of TPA. Twice weekly applications of 20 nmol of TPA did not further enhance the papilloma incidence. A good correlation was observed between the induction of ornithine decarboxylase (ODC) activity and the formation of skin tumors by various doses of TPA. To determine the effect of promotion duration on the incidence of papillomas and carcinomas, mice were treated with 10 nmol of TPA for various durations (6, 12, 18, 24, 30, or 36 weeks) beginning 2 weeks after initiation with 0.2 micromol of DMBA. Mice promoted for only 6 weeks developed papillomas and carcinomas after promotion had been discontinued. There was an intermediate incidence of tumors in the group treated for 12 weeks. Promotion for 18, 24, 30, or 36 weeks elicited virtually identical yields of papillomas. The incidence of carcinomas was proportional to promotion duration times of 6, 12, and 18 weeks, but carcinoma incidence was less than maximal in mice promoted for 24 weeks or longer. The results indicate that a) the incidence of papillomas serves as a rapid (18 weeks) index for subsequent appearance of carcinomas, b) twice weekly applications of 10 nmol of TPA for 18 weeks following initiation of female CD-1 mice with 0.2 micromol of DMBA is an appropriate protocol for maximum tumor yield in initiation-promotion experiments, and c) ODC induction may be an important component of the mechanism of skin tumor promotion by TPA.

Lymphocyte Activation by the Tumor-Promoting Agent 12-O-Tetradecanoylphorbol-13-Acetate (TPA)

Abstract In “Lymphocyte Activation by the Tumor-Promoting Agent 12-O-Tetradecanoylphorbol-13-Acetate (TPA)” by Jochen Abb, Gary J. Bayliss, and Friedrich Deinhardt in the May 1979 issue of the Journal of Immunology, Volume 122, No. 5, page 1639, the authors learned that similar studies had been reported previously by other investigators listed below: Estensen, R. D., J. W. Hadden, E. M. Hadden, F. Touraine, J. L. Touraine, M. K. Haddox, and N. D. Goldberg. 1974. In Control of Proliferation in Animal Cells. Edited by B. Clarkson and R. Baserga. Cold Spring Harbor Press, Cold Spring Harbor, N. Y. P. 627.Touraine, J. L., J. W. Hadden, F. Touraine, E. M. Hadden, R. Estensen, and R. A. Good. 1977. J. Exp. Med. 145:460.

Lymphocyte Activation by the Tumor-Promoting Agent 12-O-Tetradecanoylphorbol-13-Acetate (TPA)

TPA, a highly active tumor-promoting agent, is an effective mitogen for primate peripheral blood lymphocytes. Optimal stimulation of human lymphocytes was obtained 4 days after the addition of TPA at a concentration of 7.5 ng/ml. Lymphocyte fractionation experiments demonstrated that both T and B cells incorporated 3H-thymidine significantly in response to TPA. Lymphocyte blastogenesis was not due to the reactivation of latent herpesviruses by the tumor promoter, since similar responses to TPA were obtained with virus-genome positive or negative cells. Increased levels of DNA synthesis were observed when TPA was added to marmoset, baboon, rhesus monkey, or chimpanzee peripheral blood lymphocytes. Canine peripheral blood lymphocytes and spleen cells from guinea pigs, rats, and mice were not stimulated by TPA. These observations suggest that TPA-induced lymphocyte blastogenesis may be useful for studies of lymphocyte activation and of the molecular mechanisms of action of tumor-promoting phorbol esters.