[Oral administration of medications to infants: Implementation of a preliminary risk analysis in a hospital service of pediatrics]. / Administration orale de médicaments au nourrisson : réalisation d'une analyse préliminaire des risques dans un service d'hospitalisation pédiatrique.

PURPOSE: The objective of this study was to reduce the risk of errors when administering oral medications to infants aged 28 days to 2 years. MATERIAL AND METHODS: The method of the preliminary risk analysis (PRA) was implemented by a multidisciplinary group in a hospital service of pediatrics. The study focused on the phase of preparation of drugs by nurses before administration. RESULTS: This analysis revealed 41 scenari, 16 were criticality unacceptable. In particular, their analysis highlighted the impact of the drug dosage form, the lack of scientific information and the human factor on this preparation. Eleven action sheets have been written. DISCUSSION: The risk management requires significant human investment, material resources and organizational solutions: formations, information, i.e. computerized prescribing, dispensing and administering system, centralized drug preparations, automated drugs cabinets or unit drug daily dispensing system. CONCLUSION: Control these risks means to get specific actions at pediatric wards, enhance dispensing system by the hospital pharmacist and the support of the pharmaceutical industry to get commercially available pediatric drugs.

[The study assessed the prevalence of TPS and its associations with psychopathy in a population of forensic violent patients in a Belgian security hospital]. / Evaluation du trouble de la personnalité sadique en relation avec la psychopathie auprès de patients internés en hôpital psychiatrique sécuritaire.

The population was composed of 76 male patients (mean age=36.14). All of them having committed a violent offence indexed in their institutional file: (1) sexual offences on children; (2) rapes of adult women; (3) homicide offence; and (4) assaults and batteries. TPS was defined by the following 8 diagnostic criteria as described in DSM III-R: 1) has used physical cruelty or violence for the purpose of establishing dominance in a relationship; 2) humiliates or demeans people in the presence of others; 3) has treated or disciplined someone under his or her control unusually harshly; 4) is amused by, or takes pleasure in, the psychological or physical suffering of others; 5) has lied for the purpose of harming or inflicting pain on others 6) gets other people to do what he or she wants by frightening them 7) restricts the autonomy of people with whom he or she has a close relationship; 8) is fascinated by violence, weapons, martial arts, injury, or torture. These criteria were assessed from (a) clinical and institutional files and (b) clinical collateral informations. TPS assessment was conducted by two -trainees in clinical psychology (kappa=0.87; n=20). The assessment of psychopathy was conducted according to the guidelines of the Hare psychopathy checklist manual (PCL-R, 1991, 2003): coding of clinical and institutional files and semi-structural clinical interviews. The PCL-R is mainly composed by 2 factors: factor 1 "Emotional detachment" describing the core psychological component of psychopathy, and factor 2 "Chronically antisocial factor" reflecting behavioral instability and antisocial life style. The total cut-off score for the inclusion of the diagnosis was 25. The prevalence of TPS in the population was 25% (n=19) and is congruent with the large range described in the literature (0.5 to 33%). The most frequent criteria were 6 (gets other people to do what he or she wants by frightening them), 1 (has used using physical cruelty or violence for the purpose of establishing dominance in a relationship) and 3 (has treated or disciplined someone under his or her control unusually harshly). The most sensible criteria were: 7 (restricts the autonomy of people with whom he or she has a close relationship), 8 (major interest for violence) and 4 (pleasure in the psychological or physical suffering of others). The most specific criteria were: 3 (has treated or disciplined someone under his or her control unusually harshly), 6 (gets other people to do what her or she wants by frightening them), 4 (takes pleasure in the psychological or physical suffering of others) and 1 (has used physical -cruelty or violence for the purpose of establishing dominance in a relationship). As concerns psychopathy, the mean of factor 1, factor 2 and the PCL-R total scores were 7.40, 9.08 and 18.67. Thus, 38% of patients were considered as "low psychopaths", 36% were considered as "moderate psychopaths" and 26% were considered as "high psychopaths". In spite of few significant positive correlations between some TPS and PCL-R criteria, TPS diagnosis was not significantly correlated with factors 1, factor 2, nor with total score of the PCL-R. The mean psychopathy total score did not differ between sadistic and non-sadistic patients. Moreover, a two ways ANOVA comparing PCL-R factors 1 and 2 did not reveal any differences between sadism and non-sadism. Again, these comparisons did not support hypothesis of a strong association between TPS and psychopathy.

The role of protein tyrosine kinases in CYP1A1 induction by omeprazole and thiabendazole in rat hepatocytes.

Benzimidazoles compounds like omeprazole (OME) and thiabendazole (TBZ) mediate CYP1A1 induction differently from classical aryl hydrocarbon receptor (AhR) ligands, 3-methylcholanthrene (3-MC) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). To clarify the involvement of an intracellular signal pathway in CYP1A1 induction by OME and TBZ, the TBZ, OME and 3-MC signal-transducing pathways were compared by using specific protein tyrosine kinase inhibitors in primary culture of rat hepatocytes. The effect of OME and TBZ (75-250 microM) on cytochrome P450 1A1 (CYP1A1) expression was therefore studied in primary cultures of rat hepatocytes after 24 h, 48 h and 72 h of exposure. Both compounds provoked a dose- and time-dependent increase in CYP1A1 (EROD activity, protein and mRNA levels), but OME was less effective at all the concentrations and times tested. The mechanism of benzimidazole-mediated induction of CYP1A1 was investigated by comparison with 3-MC, a prototypical AhR ligand. As expected, OME and TBZ were unable to displace [(3)H]-TCDD from its binding sites to the AhR in competitive binding studies. Moreover, classic tyrosine kinase inhibitor herbimycin A (HA) inhibited the two benzimidazoles-mediated CYP1A1 inductions, but only partially inhibited the 3-MC-mediated one. Another two tyrosine kinase inhibitors, Lavendustin A (LA) and genistein (GEN), had no effect on CYP1A1 induction by benzimidazoles and 3-MC. These results are consistent with the implication of a tyrosine kinase, most probably the Src tyrosine kinase, in the mechanism of CYP1A1 induction in rat hepatocytes.

Induction of cytochrome P450 1A1 gene expression, oxidative stress, and genotoxicity by carbaryl and thiabendazole in transfected human HepG2 and lymphoblastoid cells.

Carbaryl and thiabendazole, two widely used pesticides, have been shown to induce cytochrome P450 1A1 (CYP1A1) expression, but neither compound is capable of displacing [3H] 2,3,7,8-tetrachlorodibenzo-P-dioxin from its aryl hydrocarbon receptor binding site. In the present study, we investigated the transcriptional regulation of CYP1A1 as well as other genes in various human hepatoma HepG2 cell lines stably transfected with the chloramphenicol acetyl transferase (CAT) reporter gene and cloned under the control of each of 14 promoters or response elements from relevant stress genes. Carbaryl and thiabendazole were found to activate CYP1A1 at the level of transcription, as demonstrated by the dose-dependent increase in reporter CAT and CYP1A1 mRNAs. Moreover, this effect appeared to be mediated via the xenobiotic responsive element (XRE), because both pesticides specifically activated various fusion constructs containing XRE sequences (CYP1A, glutathione S-transferase, and XRE). Carbaryl and to a lesser extent thiabendazole also activated other stress genes such as c-fos and NF-kappaBRE, HSP70 and GRP78, and GADD153 at a transcriptional level. These data suggest that these molecules induce early alert genes, including those known to be sensitive to oxidative stress. This led us to examine the genotoxic effect of carbaryl and thiabendazole by an in vitro DNA repair solid-phase assay. Both compounds provoked a strong DNA-damaging activity in the human lymphoblastoid cell line that constitutively expresses human CYP1A1 cDNA, but not in the parental line, indicating that CYP1A1 is chiefly implicated in carbaryl and thiabendazole genotoxicity. This effect was confirmed on HepG2 cells. These observations support the notion that intracellular signals leading to CYP1A1 induction, oxidative stress, and genotoxicity are intimately related.

Is CYP1A1 induction always related to AHR signaling pathway?

Humans are daily subjected to ever increasing amounts of exogenous compounds. Some of them are capable of inducing cytochrome P450s, a process that allows the cell to adapt to changes in its chemical environment. One of the most widely CYP studied is CYP1A1 because it metabolises a large number of xenobiotics to cytotoxic and/or mutagenic derivatives. To date, results from the literature indicate that induction of CYP1A1 does not only involve the classical activation cascade of the Ah receptor, e.g. binding of the ligand to the AhR, heterodimerisation with Arnt protein, constitution of a complex with XRE responsive element and subsequent gene activation. Indeed, some xenobiotics do activate CYP1A1 gene expression in spite of their inability to compete with TCDD for binding to the AhR. Other signaling pathways must therefore also be considered. Firstly, the CYP1A1 inducer compounds could be very weak AhR ligands or may be metabolized into a form which is in turn capable of binding to the Ah receptor. A second hypothesis would be that these molecules could act through other signaling cascades. At this time, two of them seem to be implicated. One concerns the RARs signal transduction pathway, as already described for retinoic acid. The second may involve tyrosine kinase activation, but the precise relationship between this activation and CYPA1 induction remains yet to be established. For the moment there is still a black box which needs to be investigated.

Diflubenzuron, a benzoyl-urea insecticide, is a potent inhibitor of TCDD-induced CYP1A1 expression in HepG2 cells.

Diflubenzuron (DFB) belongs to a group of compounds called benzoyphenyl ureas acting as chitin synthesis inhibitors, which also inhibit growth of B16 murine melanomas. The present study was designed to investigate the effect of this insecticide, on CYP1A1 expression and induction in human hepatoma cells HepG2. Treatment of HepG2 cells over 72 h with noncytotoxic concentrations of DFB resulted in a strong dose-dependent decrease in constitutive ethoxyresorufin-O-deethylase activity. Moreover, DFB significantly decreased CYP1A1 induction by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) after 24 h exposure, as demonstrated by ethoxyresorufin-O-deethylase (EROD) activity and Northern blot analysis. Additional studies were performed both on parental HepG2 cells and HepG2-241c.1, which were stably transfected with the chloramphenicol acetyltransferase (CAT) reporter gene, cloned under the control of the human CYP1A1 promoter (-1140 to +59). Ribonuclease protection assays (RPA) analysis clearly demonstrated an inhibition of CYP1A1 transcription in both cell lines. Surprisingly, in corresponding experiments using 3-methylcholanthrene (3-MC) as a CYP1A1 inducer, DFB was less effective. Finally, in competitive binding studies using a 9S-enriched fraction of HepG2 cytosol, DFB was capable of displacing [(3)H]-2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) from its Ah receptor binding site. Taken together, these results support the involvement of a transcriptional mechanism in the inhibition of CYP1A1 expression in HepG2 cells by DFB, possibly via an Ah receptor antagonism.

Cytochrome 1A1 induction by primaquine in human hepatocytes and HepG2 cells: absence of binding to the aryl hydrocarbon receptor.

Malaria remains the most prevalent infectious disease of tropical and subtropical areas of the world. It represents a crucial problem in public health care, affecting 750 million people annually, of whom at least two million die. Various antimalarials currently used were studied for their capability to induce expression of the cytochrome P450 1A1 (CYP1A1) gene, an enzyme that plays an important role in the activation of xenobiotics to genotoxic derivatives. Studies on human hepatocytes and HepG2 cell lines showed that primaquine was capable of dose dependently increasing both the ethoxyresorufin-O-deethylase activity and CYP1A1 mRNAs, suggesting a transcriptional activation of this gene. Moreover, alpha-naphthoflavone, a partial aryl hydrocarbon receptor (AhR) antagonist, and 8-methoxypsoralen, which interferes with the binding of activated AhR to the xenobiotic responsive element, were shown to suppress CYP1A1 induction when added to the cultures. However, neither primaquine nor its metabolites were able to displace [3H]2,3,7,8-tetrachlorodibenzo-p-dioxin from AhR in competitive binding studies using 9S-enriched fractions of human cytosol. These data, together with the induction of CYP1A1 promoter-directed chloramphenicol acetyl transferase gene expression, suggest that CYP1A1 induction involves the participation of the AhR but not a direct primaquine-receptor interaction. This supports the notion that an alternative ligand-independent mechanism has to be considered. Given the pharmaco-toxicological significance of CYP1A1 induction, these findings may have important implications in the treatment of malaria with primaquine and new analogs.

Cytotoxic effects and induction of cytochromes P450 1A1/2 by insecticides, in hepatic or epidermal cells: binding capability to the Ah receptor.

Insecticides deserve particular attention since the general population is potentially exposed to such chemicals through many routes. We therefore tested the comparative acute and chronic toxicity of chemicals belonging to the major insecticides families (DDT, malathion and tetrachlorvinphos, carbaryl, cypermethrin, diflubenzuron), in hepatocytes, HepG2 and HaCaT cell lines. Two kinds of end-points were used: cytotoxicity parameters and CYP1A1 induction. Except for cypermethrin and diflubenzuron, all these chemicals exerted a cytotoxic effect in hepatocytes and HaCaT, but not in HepG2 cells. However, the induction of the EROD activity appeared more sensitive since a response was detected at lower concentrations. Significant differences were observed between the cell types and the insecticides. Furthermore, these chemicals were unable to displace [3H]TCDD from its binding sites, suggesting that they would not be a ligand of the Ah receptor. The experimental approach used herein may be a good means for predicting the acute and chronic toxicity of pesticides.

Toxic effects of several types of antifouling paints in human and rat hepatic or epidermal cells.

Fouling is the successive development of marine organisms on immersed surfaces, a process which has heavy negative economic impacts. Several antifouling technologies, generally based on the leaching of biocides from painted surfaces, have been developed, but these biocides are toxic to the environment. Hence, we compared the toxicity of several currently used paint lixiviats in rat hepatocytes, human HepG2 and HaCaT cells. Acute toxicity was assessed by the Neutral Red and MTT assays. Chronic effect was tested using induction of the 7-ethoxyresorufin-O-deethylase (EROD) activity as a marker. Large variations were observed among the various cell types or the antifouling formulations, both in terms of IC50 values (from approximately 0.5 to approximately 10%, v/v) and EROD induction (from approximately 1 to 10-fold over control). These differences appear to be related to variable biocide (copper compounds, organotins, etc...) concentrations in the different paint formulations, or to the specific metabolic capabilities of the cell system used.

Carbaryl induces CYP1A1 gene expression in HepG2 and HaCaT cells but is not a ligand of the human hepatic Ah receptor.

In spite of increasing numbers of insecticides used in agriculture, there are serious concerns regarding the potential risks of exposure to these agents. Carbaryl is one of the most important carbamate insecticides and has been used for about 30 years to control a wide range of pests. The study was designed to investigate if, among various insecticides currently used in world agriculture, this compound could induce human CYP1A1, an enzyme known to play an important role in the chemical activation of xenobiotics to genotoxic derivatives. Studies on HepG2 and HaCaT cell lines showed that carbaryl is capable of increasing, in a dose-dependent manner, both the ethoxyresorufin rufin-O-dec, O-deethylase activity and the steady-state concentrations of CYP1A1 mRNA, suggesting a transcriptional activation of this gene. When alpha-naphthoflavone, a partial Ah receptor (AhR) antagonist, and 8-methoxypsoralen, which interferes with the binding of activated AhR to the xenobiotic responsive element (XRE), were added to the cultures, CYP1A1 induction was suppressed. However, competitive binding studies using the 9S enriched fraction of human cytosol indicated that carbaryl did not displace [3H]TCDD from AhR. These data, together with the activation of a XRE-directed CAT reporter gene by carbaryl, suggest that induction of CYP1A1 involves the participation of the AhR and the XRE, but is not mediated by a direct carbaryl-receptor interaction. An alternative ligand-independent mechanism should be considered. Therefore, although carbaryl concentration in food is very low, care should be taken because of its possible adverse effects in human health through liver and skin, given the well established toxicological importance of CYP1A1 induction.

Comparative study of CYP1A1 induction by 3-methylcholanthrene in various human hepatic and epidermal cell types.

Hepatocytes and keratinocytes are among the most widely used cells in pharmaco-toxicology, but a limitation of these models is the provision of human tissues on a regular basis. The suitability of HepG2, HaCaT and HESV cell lines as an acceptable substitute for primary cultures was examined. In these cell types, the effects of 3-methylcholanthrene (3-MC) were analysed on CYP1A1 gene expression, a crucial CYP subfamily in the activation of chemical carcinogens. Ethoxyresorufin O-deethylase (EROD) activity was never detected in HESV cells, but in other cell types it was stimulated in a concentration-dependent manner (maximal induction, 1-2.5 mum). Above this peak induction the effect fell rapidly. Northern blot analysis of CYP1A1 mRNA agreed with the trends obtained for EROD values. However, the decrease of the EROD activity observed at the highest 3-MC concentrations was not correlated with CYP1A1 mRNA reduction. This study also demonstrated that 3-MC is capable of significantly inducing CYP1A1 in HaCaT cells (17-fold over control), as in human hepatocytes (six- to 18-fold) and HepG2 (fourfold). Therefore, in contrast to SV40-immortalized keratinocytes (HESV), spontaneously immortalized keratinocytes (HaCaT) may constitute a valuable tool for studying epidermal CYP1A1 gene regulation by xenobiotics.

Evaluation of the toxicological risk to humans of caulerpenyne using human hematopoietic progenitors, melanocytes, and keratinocytes in culture.

The extensive growth of Caulerpa taxifolia in the Mediterranean sea produces important quantities of bioactive secondary metabolites unable to enter the food chain. The cytotoxic effects of caulerpenyne, the major secondary metabolite from C. taxifolia, was studied in different in vitro models: skin cells, primary cultures of melanocytes and keratinocytes, immortalized keratinocytes (HaCaT and HESV), and bone marrow cells (hematopoietic progenitors CFU-GM). Typical dose-response curves from neutral red uptake and MTT assays were recorded in all models with IC50 ranging from 6 to 24 microM. Hematopoietic progenitors were more sensitive to caulerpenyne than melanocyte and keratinocyte cell lines, which could be due to their higher proliferative rate. The distribution of aggregates in colonies, macroclusters, and microclusters of hematopoietic progenitors was also altered in the presence of caulerpenyne. From our evaluation of the caulerpenyne concentrations required to result in cellular toxicity, the risks of cutaneous and/or food intoxication to humans may be considered minimal.

Alteration of Ca2+ homeostasis of sea urchin embryos by retinoid CD 367, dual effect on egg cleavage and embryonic development.

The effect of a light stable retinoid (CD 367) was studied on sea urchin embryos. CD 367 did not affect sperm-egg interaction. In a range of concentrations between 10 and 100 microM, CD 367 delayed the first and the second cleavages. When added after fertilization, micromolar amounts of CD 367 delayed hatching and produced embryonic abnormalities in a dose-dependent manner. Mesodermal cells, primary (PMC) and secondary (SMC) mesenchyme cells migration was particularly disturbed, leading to exogastrulations and calcified spicules malformations. Concentrations of CD 367 higher than 8 microM were embryolethal. Micromolar amount of CD 367 increased plasmalemma Ca2+ permeability of fertilized eggs but not of unfertilized eggs. CD 367 inhibited ATP-dependent intracellular sequestration of Ca2+ in a range of concentrations similar to those affecting egg cleavage and embryonic structures. Since we were unable to detect nuclear receptors for CD 367 in sea urchin eggs and ovocytes, these effects probably are not related to interaction of the retinoid with members of the RAR family, to which CD 367 has a high affinity, but rather to its toxicity by the means of some unknown mechanisms.

Identification of synthetic retinoids with selectivity for human nuclear retinoic acid receptor gamma.

The action of retinoids on gene regulation is mediated by three distinct nuclear retinoic acid receptor (RAR) subtypes called RAR alpha, beta and gamma. Since RAR gamma is predominantly expressed in adult skin, specific ligands for this subtype could (i) represent valuable tools to evaluate the biological role of RAR gamma in skin and (ii) provide therapeutic entities with a higher therapeutic index at lower teratogenic risk. Using in vitro binding studies and a functional transactivation assay, we have identified three compounds with high RAR gamma selectivity.

Selective high affinity retinoic acid receptor alpha or beta-gamma ligands.

Biological effects of retinoic acid (RA) are mediated through its binding to three closely related nuclear receptors (RAR alpha, RAR beta, and RAR gamma) belonging to the steroid-thyroid nuclear receptor family. RARs are able to modulate the transcription of specific genes by binding to responsive elements located in the promoter-enhancer region of these genes. As demonstrated by in situ hybridization, the distribution of each RAR type in the developing embryo, as well as in the adult, is not uniform. In this context, synthetic retinoids that would behave as selective ligands would be invaluable for studying the respective roles of each RAR type in cultured cells, whole animals, and embryos. Moreover, from a pharmacological point of view, such selective compounds may possess a higher therapeutic index and a lower teratogenic risk, because they might affect specific tissues and spare some others. As an approach to this problem, we have set up two complementary assays, (i) an in vitro binding assay to determine the Kd values of retinoids for RAR alpha, RAR beta, and RAR gamma and (ii) a functional assay in cultured cells to evaluate the potential of retinoids to transactivate, through their binding to one type of RAR, a reporter gene. The binding assay uses nuclear extracts of COS-7 cells transfected with vectors expressing RAR alpha, RAR beta, or RAR gamma. The functional assay is a measure of chloramphenicol acetyltransferase (CAT) activity in HeLa cells co-transfected with the expression vectors used in the binding assay and the reporter gene TRE-tk-CAT. Selective agonists for RAR alpha (Am80 and Am580) and RAR beta-RAR gamma (CD495 and CD564) were identified. However, compounds with pure RAR beta or RAR gamma selectivity have not yet been identified.

Differentiation of F9 embryonal carcinoma cells by synthetic retinoids: amplitude of plasminogen activator production does not depend on retinoid potency or affinity for F9 nuclear retinoic acid receptors.

Retinoic acid and analogues (retinoids) are able to induce the differentiation of F9 murine embryonal carcinoma stem cells into endoderm-like cells. The secretion of plasminogen activator (PA) which accompanies this differentiation is a good index of the biological response of F9 cells to retinoids. We have previously reported that the potency of a series of natural and synthetic retinoids, evaluated by the concentration which provokes half-maximal induction of PA, correlates well with the affinity of these compounds for the endogenous F9 nuclear retinoic acid receptors, but not for the cytosolic retinoic acid binding protein, CRABP. In this paper we show that various retinoids differ, not only in terms of potency, i.e. the dilution at which they are active, but also in terms of the amount of PA that they induce. This parameter, called amplitude, is used to quantify the extent of PA induction by a given retinoid relative to retinoic acid. The amplitude parameters of synthetic retinoids are found to vary over a wide range and are independent of both potency and binding affinity for F9 retinoic acid receptors. It is proposed that the amplitude of the biological response to a given retinoid is the resultant of three factors: (i) the total or partial agonist character of the retinoid; (ii) the binding spectrum of the retinoid for the various types of retinoic acid receptors; (iii) the chemical and metabolic stability of the retinoid in the test system.

Effect of diterpene esters on actin cytoskeleton of SV40-transformed keratinocytes is not reproduced by diacylglycerols.

The tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) induces a disorganization of the actin cytoskeleton and its redistribution at the periphery of the cells in SV40-transformed human keratinocytes. This phenomenon is induced by other diterpene esters, such as 4-O-methyl TPA, 12-O-ethacrynylphorbol-13-acetate (EPA), 12-O-retinoylphorbol-13-acetate (RPA) and mezerein, which exert either convertogenic or promoting effects in skin tumor development in vivo. Two diacylglycerols: oleyl-acetyl glycerol (OAG) and dioctanoyl-glycerol (DOG) do not induce the disorganization of actin. Thus, the effects of these compounds, as they are found in SV40-transformed human keratinocytes, do not exhibit clear-cut correlations to either their protein kinase C activating abilities, or their effects on different stages of multistage carcinogenesis in mouse skin in vivo.

Biological activity of retinoids correlates with affinity for nuclear receptors but not for cytosolic binding protein.

In order to better understand the respective roles of the nuclear retinoic acid receptors (RARs) and the cytosolic retinoic acid binding protein (CRABP) in the mode of action of retinoic acid (RA), several types of RA analogs have been synthesized. Representative compounds have been radiolabeled to a high specific activity and their binding (direct and competition) to RARs and CRABP was determined. Their biological activity on F9 embryonal carcinoma cell differentiation has been determined by a quantitative assay of plasminogen activator (PA). All biologically active analogs studied in this work bound to RARs. A good correlation was found between PA induction and affinity for the RARs, with the exception of RA itself which was a good ligand but a moderate inducer of F9 differentiation. Two biologically active analogs (compounds II and III) did not bind to the CRABP. One biologically inactive analog (compound VIII) bound to CRABP. These results strongly suggest that retinoids must bind to RARs but not necessarily to CRABP in order to induce cell differentiation in F9 cells.

Effects of antiinflammatory agents on edema and DNA synthesis induced by 12-O-tetradecanoylphorbol-13-acetate in the guinea pig.

Inflammation and hyperplasia are frequently associated in skin diseases. In order to verify this relationship, we studied the antagonistic effect of different classes of antiinflammatory agents on the inflammatory and hyperplasiogenic responses elicited by one topical application of 12-O-tetradecanoylphorbol-13-acetate (TPA) to the ear of the guinea-pig. Edema and DNA synthesis were chosen as relevant parameters. All antiinflammatory agents tested significantly inhibited DNA synthesis induced by TPA. Moreover, all compounds except quinacrine and phenylbutazone also inhibited edema formation. In conclusion, our results demonstrate that while edema and hyperplasia are frequently associated, this is not always the case.

Transformation of human keratinocytes by SV40 virus alters their response to the tumor promoter TPA.

Recently, we have shown that transformation of human keratinocytes by SV40 virus induces the re-expression of characters found in fetal epidermis and monostratified epithelia. In the present study, TPA was found to alter some features of the human keratinocyte phenotype in a similar manner to SV40 transformation. Indeed, 12-O-tetradecanoyl-13-phorbol acetate (TPA) treatment induced the expression of cytokeratin no. 8, recognized by the monoclonal antibody TROMA-1, which is present in fetal epidermis and/or monostratified epithelia only, and fibronectin expression. However, TPA and SV40 transformation had specific non-overlapping effects. For example, TPA did not prevent terminal differentiation and stratification, as SV40-transformation did, but stimulated these processes. Moreover, TPA was found to induce additional changes in SV40-transformed keratinocytes. In particular it provoked the individualization of cells within the colonies. This effect was seen as little as 1 h after treatment and was reversible. This cellular alteration was accompanied by the reorganization of actin and by a decrease in the number of desmosomes; these changes were not observed after treatment of normal keratinocytes with TPA. These observations lead to the conclusion that TPA is able to trigger cellular responses which cannot be induced by SV40 products alone, but that TPA and some SV40 products can cooperate to elicit new responses, which could reflect a higher state of malignancy.