tretinoin and 3-5-di-tert-butylchalcone-4--carboxylic-acid

It was recently revealed that epidermal growth following topical treatment with all-trans retinoic acid (atRA) was at least partly induced by heparin-binding epidermal growth factor-like growth factor (HB-EGF) released from suprabasal keratinocytes. Since proliferation of keratinocytes appears to be one of the critical roles of atRA in depigmentation treatment and promotion of wound healing, HB-EGF is considered suitable for assessing the therapeutic value of topical retinoids. In this study, HB-EGF mRNA expression in normal human keratinocytes after atRA treatment was examined, and the effects of a variety of natural and synthetic retinoids were compared. The results of reverse transcription polymerase chain reaction (RT-PCR) suggested that induction of differentiation increased HB-EGF mRNA expression in cultured keratinocytes. Real-time PCR analyses revealed that HB-EGF mRNA expression was elevated dose-dependently with atRA, peaking at 12 h. This elevation was more prominent in confluent keratinocytes than in subconfluent cells, suggesting that differentiated keratinocytes are more subject to stimulation of HB-EGF expression by atRA than proliferating keratinocytes. HB-EGF mRNA was upregulated in differentiation-induced keratinocytes by all retinoids used in this study at 1 micromol/l, and marked upregulation was seen when treated with three isotypes of retinoic acid (atRA, and 9-cis and 13-cis retinoic acid). RARalpha-selective agonists (Am80, Am580, ER-38925, and TAC-101) and a panagonist of RARs (Re80) caused relatively low elevation of HB-EGF transcripts, as did all-trans retinol (Rol) and all-trans retinal (Ral). Although another panagonist (Ch55) showed the highest elevation of HB-EGF mRNA, it was relatively cytotoxic at the concentration employed. Ral and Rol were found to upregulate HB-EGF when used at 100 micromol/l to 1 mmol/l, to a similar extent of atRA at 1-10 micromol/l. The capacity of retinoids to upregulate HB-EGF may be an important index for investigation and development of an ideal synthetic retinoid, which has maximum benefits and minimum side-effects

Topics: Cells, Cultured; Chalcone; Chalcones; Epidermal Growth Factor; Heparin-binding EGF-like Growth Factor; Humans; Intercellular Signaling Peptides and Proteins; Keratinocytes; Keratolytic Agents; Reference Values; Retinaldehyde; Retinoids; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tretinoin; Up-Regulation; Vitamin A

Retinoids, especially all-trans retinoic acid (RA), have been shown to inhibit the differentiation of preadipose cells. It is important to human health, especially to obesity, that the regulatory system for the differentiation of adipocytes is well defined. Previously, we have shown that retinoic acid receptor (RAR) gamma 2 gene expression is up-regulated by RA in 3T3-L1 preadipose cells. In this study, the RAR system was dissected and the RA-regulated function in 3T3-L1 cells was assigned to one given receptor. We used three synthetic retinoids; (1) Ro 41-5253, a selective RAR alpha antagonist, (2) Ch 55, an RAR alpha, beta and gamma agonist, and (3) Am 80, an RAR alpha and beta agonist, which has less affinity to RAR gamma. Ro 41-5253 reverted RA-induced inhibition of the differentiation of 3T3-L1 cells. However, there was no significant reversion in RA-induced RAR gamma mRNA level by treatment with Ro 41-5253. In the case of RAR agonists, both Am 80 and Ch 55 strongly inhibited the differentiation of 3T3-L1 cells. However, Am 80 weakly increased RAR gamma mRNA content less than did Ch 55. These findings suggest, that RAR alpha is involved in the prevention of adipose differentiation by RA in 3T3-L1 cells. Moreover, there seems no causal relationship between the prevention of adipose differentiation by RA and the up-regulation of RAR gamma 2 gene expression by RA in 3T3-L1 cells. We have shown the functional heterogeneity of RA action through different RARs in 3T3-L1 cells.

Topics: 3T3 Cells; Adipocytes; Animals; Benzoates; Cell Differentiation; Chalcone; Chalcones; Chromans; Mice; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Retinoids; Tetrahydronaphthalenes; Tretinoin

all-trans-Retinoic acid (RA) is a potent inducer in vitro of the differentiation of the human acute myeloid leukemia cell line HL60. A mechanism for RA-induced differentiation of HL60 cells may involve retinoylation (RA acylation) which is a post-translational modification of proteins occurring in many eukaryotic cell lines. Here, we found that differentiation by the synthetic retinoid (E)4-[3-(3,5-di-tert-butylphenyl)-3-oxo-1-propenyl]-benzoic acid (Ch55) was dose-dependent in serum-free medium. The synthetic retinoid 4(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenylcarbamoyl) benzoic acid (Am80) did not induce differentiation. Ch55 bound covalently to proteins of HL60 cells. In contrast, covalent binding of Am80 to HL60 proteins was much lower. Two-dimensional gel electrophoresis patterns of proteins labeled covalently by RA and Ch55 were different with few proteins labeled by both retinoids. The level of retinoylation was increased by Am80 and combinations of RA with either Ch55 or Am80 synergistically induced differentiation of HL60 cells. These results suggest that covalent modification of proteins by a retinoid may play a role in inducing differentiation of HL60 cells. In addition, the synergy seen with combinations of RA and either Ch55 or Am80 suggests that some synthetic retinoids may be active because they displace RA from intracellular sites or because they inhibit RA catabolism.

Topics: Benzoates; Cell Differentiation; Chalcone; Chalcones; Drug Synergism; Electrophoresis, Gel, Two-Dimensional; Humans; Leukemia, Promyelocytic, Acute; Neoplasm Proteins; Protein Binding; Tetrahydronaphthalenes; Tretinoin; Tumor Cells, Cultured

RCJ C 5.18 (C 5.18) is a chondrogenic clonal cell line which, under standard culture conditions, develops chondroblastic features including the production of a cartilagenous matrix. Retinoic acid (RA) is known to inhibit the chondrogenic differentiation of C 5.18 cells and this may parallel the teratogenic effects of retinoids in vivo; however, the question as to which of the 3 retinoic acid receptors (RAR alpha, beta, gamma) or the 3 retinoid X receptors (RXR alpha, beta, gamma) mediate this RA-induced inhibition remains unanswered. We tested several retinoids with different receptor binding characteristics. Cartilage formation in C 5.18 cultures was evaluated by counting the number of cartilage nodules formed, and by quantitating the glycosaminoglycan content of the cultures using alcian blue staining. All of the retinoids prevented cartilage formation in a dose-dependent manner. Treatment with the retinoids did not affect cell number, thereby ruling out any toxic effects. RA, which binds to all 3 RARs with similar affinity, produced a 50% inhibition (IC50) of cartilage formation at 4 x 10(-10) M. We also tested Ch55, which also binds to all 3 RARs, but with higher affinity than RA. This compound was approximately 10 times more potent than RA (IC50 2 x 10(-11) M). 9-cis RA, which binds to the 3 RARs with affinities similar to RA and also binds to the 3 RXRs, was less active (IC50 8 x 10(-9) M), suggesting that RXR binding interferes with the inhibitory effect of ligand-activated RARs. 9-cis retinal, for which the binding characteristics are unknown, had the same effect as 9-cis RA.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Benzoates; Cartilage; Cell Differentiation; Cell Line; Chalcone; Chalcones; Dose-Response Relationship, Drug; Glycosaminoglycans; Rats; Receptors, Cytoplasmic and Nuclear; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Retinoic Acid Receptor gamma; Retinoid X Receptors; Retinoids; Teratogens; Tetrahydronaphthalenes; Transcription Factors; Tretinoin

The first embryonic lower mouse molar was used as a model system to investigate the effects of two retinoids, retinoic acid (RA) and a synthetic analogue, Ch55, on morphogenesis and cytodifferentiations in vitro. Exogenous retinoids were indispensable for morphogenesis of bud, cap and bell-stage molars in serum-free, chemically-defined, culture media. Transferrin and RA or transferrin and Ch55 acted synergistically in promoting morphogenesis from bud and cap-stage explants. Transferrin, per se, had no morphogenetic effect. Epithelial histogenesis, odontoblast functional differentiation and ameloblast polarization always occurred in RA-depleted explants. Comparison of the distributions of bromodeoxyuridine (BrdU) incorporation between explants cultured in the absence or presence of RA revealed that RA could modify the patterns of cell proliferation in the inner dental epithelium and dental mesenchyme. Inner dental epithelium cell proliferation is regulated by the dental mesenchyme through basement membrane-mediated interactions, and tooth morphogenesis is controlled by the dental mesenchyme. Laminin is a target molecule of retinoid action. Using a monospecific antibody, we immunolocalized laminin and/or structurally-related molecules sharing the laminin B chain in the embryonic dental mesenchyme and in the dental basement membrane and showed that RA could promote the synthesis or secretion of these molecules. Based on previous in situ hybridization data, it was speculated that CRABPs might regulate the effects of RA on embryonic dental cell proliferation. The fact that Ch55, a retinoid which does not bind to CRABPs, is 100 times more potent than RA in promoting tooth morphogenesis in vitro seems to rule out this hypothesis. On the other hand, the stage-specific inhibition of tooth morphogenesis by excess RA is consistent with the hypothesis that CRABPs might protect embryonic tissues against potentially teratogenic concentrations of free retinoids.

Topics: Animals; Cell Differentiation; Cell Division; Chalcone; Chalcones; Dose-Response Relationship, Drug; Drug Synergism; Mice; Molar; Morphogenesis; Organ Culture Techniques; Transferrin; Tretinoin

Four retinoids, i.e. retinol (vitamin A), retinoic acid, retinyl acetate and synthetic chalcone carboxylic acid (Ch 55), were examined for their effects on embryonic angiogenesis using 4.5-day chorioallantoic membranes of chick embryo. The effects of these retinoids were compared with that of antibiotic herbimycin A, which was the most powerful inhibitor of the angiogenesis reported previously. The four retinoids strongly inhibited embryonic angiogenesis; the order of inhibitory activity was Ch 55 greater than retinoic acid greater than herbimycin A greater than retinyl acetate based on the dose required for the half-maximal inhibitory effect. The present results suggest that retinoids are effective inhibitors of angiogenesis, and can be applied for the management of certain diseases accompanied by aberrant angiogenesis, particularly that which occurs during progressive growth of solid tumors.

Topics: Animals; Benzoquinones; Chalcone; Chalcones; Chick Embryo; Diterpenes; Dose-Response Relationship, Drug; In Vitro Techniques; Lactams, Macrocyclic; Neovascularization, Pathologic; Quinones; Retinoids; Retinyl Esters; Rifabutin; Tretinoin; Vitamin A

The treatment of 311 cells, a pluripotent mouse teratocarcinoma cell line, with a new type of inducer, 3,5-di-tert-butylchalcone 4'-carboxylic acid (Ch55), results in the suppression of the c-mos gene, accompanied by early marker changes associated with cell differentiation, i.e., the enhanced secretion of plasminogen activator and the decrease in peanut agglutinin receptors and glucose transport. This indicates that Ch55 is a potent inducer of teratocarcinoma cells and suppresses c-mos expression.

Topics: 3-O-Methylglucose; Animals; Cell Differentiation; Chalcone; Chalcones; Gene Expression Regulation; Lectins; Methylglucosides; Mice; Molecular Weight; Peanut Agglutinin; Plasminogen Activators; Propiophenones; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-mos; Proto-Oncogenes; RNA, Messenger; Teratoma; Tretinoin; Tumor Cells, Cultured