Anti-Inflammatory Agents ; pharmacology ; Brain ; drug effects ; metabolism ; Humans ; NF-kappa B ; metabolism ; Neurons ; drug effects ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism ; alpha-MSH ; pharmacology

OBJECTIVETo observe the effect of basic fibroblast growth factor (bFGF) and alpha-melanocyte stimulating hormone (alpha-MSH) on adhesion and migration of melanocytes in vitro.

METHODSHuman melanocytes were obtained from normal human foreskins. Culture dishes covered with fibronectin were used to perform melanocytes adhesion assay, and cell motility was assessed using the Transwell micropore filter method.

RESULTbFGF and alpha-MSH increased melanocytes adhesion on culture dishes covered with fibronectin. bFGF stimulated melanocytes migration through micropore filter while alpha-MSH had no significant effects.

CONCLUSIONbFGF and alpha-MSH could promote the adhesion and migration of melanocytes, which suggests that two agents may play a role in the repigmentation of vitiligo.

Cell Adhesion ; drug effects ; Cell Movement ; drug effects ; Cells, Cultured ; Fibroblast Growth Factor 2 ; pharmacology ; Humans ; Melanocytes ; cytology ; alpha-MSH ; pharmacology

The pathogenesis of chronic cyclosporine A (CsA) nephrotoxicity has not been elucidated, but apoptosis is thought to play an important role in CsA induced tubular atrophy. Recently Fas-Fas ligand system mediated apoptosis has been frequently reported in many epithelial cells as well as in T lymphocytes. We investigated the ability of CsA to induce apoptosis in cultured human proximal tubular epithelial cells and also the effect of -MSH on them. Fas, Fas ligand, and an intracellular adaptor protein, Fas-associating protein with death domain (FADD) expression, and poly-ADP ribose polymerase (PARP) cleavage were also studied. CsA induced apoptosis in cultured tubular epithelial cells demonstrated by increased number of TUNEL positive cells and it was accompanied by a significant increase in Fas mRNA and Fas ligand protein expressions. FADD and the cleavage product of PARP also increased, indicating the activation of caspase. In -MSH co-treated cells, apoptosis markedly decreased with downregulation of Fas, Fas ligand and FADD expressions and also the cleavage product of PARP. In conclusion, these data suggest that tubular cell apoptosis mediated by Fas system may play a role in tubular atrophy in chronic CsA nephrotoxicity and pretreatment of -MSH may have a some inhibitory effect on CsA induced tubular cell apoptosis.
Antigens, CD95/genetics ; Apoptosis/*drug effects ; Carrier Proteins/biosynthesis ; Caspases/physiology ; Cells, Cultured ; Cyclosporine/*toxicity ; Human ; Immunosuppressive Agents/*toxicity ; Kidney Tubules, Proximal/cytology/*drug effects/metabolism ; Membrane Glycoproteins/biosynthesis ; NAD+ ADP-Ribosyltransferase/metabolism ; RNA, Messenger/analysis ; alpha-MSH/*pharmacology

Melanocortin is the downstream mediator of leptin signaling and absence of leptin signaling in ob/ob and db/db mice revealed the enhancement of bone formation through the central regulation. While alpha-melanocyte-stimulating hormone (alpha MSH) inhibits the secretion of interleukin-1 alpha and tumor necrosis factor-alpha from the inflammatory cells, alpha MSH can also enhance clonal expansion of pro B cells linked to stimulation of osteoclastogenesis. Therefore, we tested the effect of melanocortin on bones. alpha MSH analogues [6His] alpha MSH-ND and [6Asn] alpha MSH-ND were synthesized and the radio-ligand receptor binding- and cyclic AMP generating activity were analyzed in China Hamster Ovary cell line over- expressing melanocortin receptors. The EC50 of [6His] alpha MSH-ND measured from melanocortin-1, 3, 4 and 5 receptors were 0.008 0.0045, 1.523 0.707, 0.780 0.405, and 250.320 42.234 nM, respectively, and the EC50 of [6Asn] alpha MSH-ND were 16.8 6.94, 271.8 21.95, 8.0 1.21, and 1132.5 635.46 nM, respectively. Four weeks after the subcutaneous injection of the analogues, the body weights in the [6His] alpha MSH-ND and the [6Asn] alpha MSH-ND treated groups (346.0 20.63 g vs. 350.0 13.57 g) were lower than that of the vehicle treated group (375.8 17.31 g, p 0.05). There was no difference in the total femoral BMD measured by dual x-ray absorptiometry among the three groups. Among the three groups, there were no differences in the total numbers of crystal violet positive- or alkaline phosphatase positive colonies, in the expression of Receptor Activator of Nuclear Factor Kappa-B ligand on the tibia and the total number of multinucleated osteoclast-like cells differentiated from primary cultured bone marrow cells. From the above results, no evidence of bone gain or loss was found after treatment of the alpha MSH analogues peripherally.
Animal ; Body Weight/drug effects ; Bone and Bones/*drug effects ; CHO Cells ; Cyclic AMP/biosynthesis ; Eating/drug effects ; Hamsters ; Male ; Osteoblasts/drug effects/physiology ; Osteoclasts/drug effects/physiology ; Rats ; Rats, Sprague-Dawley ; Receptors, Corticotropin/physiology ; alpha-MSH/analogs & derivatives/*pharmacology

OBJECTIVE: To explore the effects of taurolithocholic acid (tLCA) and chenodeoxycholic acid (CDCA) on the expression of aorexigenic neuropeptide in mouse hypothalamus GT1-7 cells. METHODS: Mouse hypothalamic GT1-7 cells were treated with culture medium containing 10% FBS (control group, =3) or with 10 nmol/L, 100 nmol/L, 1 μmol/L and 10 μmol/L tLCA (tLCA group, =3) or CDCA (CDCA group, =3) for 12, 24 or 48 h. Real-time PCR was performed to determine the expression levels of proopiomelanocortin (POMC) mRNA in the cells, and the production levels of α-melanocyte-stimulating hormone (α-MSH) were assessed using an ELISA kit. Signal transduction and activator of transcription 3 phosphorylation (p-STAT3), threonine kinase phosphorylation (p-AKT), suppressor of cytokine signaling 3 (SOCS3), G protein-coupled bile acid receptor-1 (TGR5) and farnesoid X receptor (FXR) protein were detected by Western blotting. RESULTS: Western blotting results showed that mouse hypothalamic GT1-7 cells expressed two bile acid receptors, TGR5 and FXR, whose expressions were regulated by bile acids. Real-time PCR showed that the expression of POMC mRNA was significantly increased in the cells after treatment with 10 μmol/L tLCA or CDCA for 24 h. POMC-derived anorexigenic peptide α-MSH increased significantly in GT1-7 cells after treatment with 10 μmol/L tLCA or CDCA for 24 h. Treatment of the cells with tLCA or CDCA significantly increased the expressions of intracellular signaling proteins including p-STAT3, p-AKT and SOCS3. CONCLUSIONS Mouse hypothalamic GT1-7 cells express bile acid receptors TGR5 and FXR. Bile acids tLCA or CDCA can promote the expression of POMC mRNA and increase the production of the anorexigenic peptide α-MSH. The intracellular signaling proteins p-AKT, p-STAT3 and SOCS3 are likely involved in bile acid-induced anorexigenic peptide production.
Animals ; Cell Line ; Chenodeoxycholic Acid ; pharmacology ; Gene Expression Regulation ; drug effects ; Hypothalamus ; cytology ; Mice ; Neuropeptides ; genetics ; metabolism ; Pro-Opiomelanocortin ; genetics ; RNA, Messenger ; genetics ; STAT3 Transcription Factor ; metabolism ; Signal Transduction ; drug effects ; Suppressor of Cytokine Signaling 3 Protein ; metabolism ; Taurolithocholic Acid ; pharmacology ; alpha-MSH ; genetics

Binding activity and biologic effect of a novel alpha-melanocyte-stimulating hormone analogue were tested on cells transiently expressing the human melanocortin-1 (MC1), MC3, MC4, and MC5 receptors. The human MC1 and MC5 receptor genes were cloned into the expression vector pcDNA3. 1/ myc-his(-) B. The vectors were transferred to HEK-293 cells by the calcium phosphate method. Stable receptor populations were generated using G418 selection (900 microg x mL(-1)) for subsequent bioassay analysis. K(i) values of the novel alpha-MSH analogue for MC1, MC3, MC4, and MC5 receptors were obtained in competition with [125I]-NDP-MSH for binding studies. The cyclic AMP level was tested by using [3H]-cyclic AMP kit. It is showed that K(i) values of the novel alpha-MSH analogue for MC1, MC3, MC4, and MC5 receptors were (0.159 +/- 0.040), (35.430 +/- 6.743), (19.293 +/- 2.780) and (2.230 +/- 0.670) nmol L(-1), respectively. Its EC50 values for MC1, MC3, MC4, and MC5 receptors were (0.45 +/- 0.07), (7.80 +/- 0.65), (2.55 +/- 0.23) and (0.33 +/- 0.09) nmol L(-1), respectively. In these tests, the novel alpha-MSH analogue is a MC1R and MC5R selective agonist.
Amino Acid Sequence ; Binding, Competitive ; Cell Line ; Cell Line, Tumor ; Cyclic AMP ; metabolism ; Genetic Vectors ; Humans ; Iodine Radioisotopes ; Kinetics ; Molecular Sequence Data ; Plasmids ; genetics ; Radioligand Assay ; Receptor, Melanocortin, Type 1 ; agonists ; genetics ; metabolism ; Receptors, Corticotropin ; agonists ; genetics ; metabolism ; Receptors, Melanocortin ; agonists ; genetics ; metabolism ; Transfection ; Tritium ; alpha-MSH ; analogs & derivatives ; chemistry ; metabolism ; pharmacology

Xanthohumol (XH), the principal prenylflavonoid of the hop plant (Humulus lupulus L.), dose-dependently inhibited isobutylmethylxanthine (IBMX)-induced melanogenesis in B16 melanoma cells, with little cytotoxicity at the effective concentrations. Decreased melanin content was accompanied by reduced tyrosinase enzyme activity, protein and mRNA expression. The levels of tyrosinase-related protein 1 and 2 mRNAs were decreased by XH. XH also inhibited alpha-melanocyte stimulating hormone- or forskolin-induced increases in melanogenesis, suggesting an action on the cAMP-dependent melanogenic pathway. XH downregulated the protein and mRNA expression of microphthalmia-associated transcription factor (MITF), a master transcriptional regulator of key melanogenic enzymes. These results suggest that XH might act as a hypo-pigmenting agent through the downregulation of MITF in the cAMP-dependent melanogenic pathway.
1-Methyl-3-isobutylxanthine/pharmacology ; Animals ; Cell Line ; Cell Survival/drug effects ; Dose-Response Relationship, Drug ; Down-Regulation ; Drug Antagonism ; Forskolin/pharmacology ; *Humulus ; Intramolecular Oxidoreductases/antagonists & inhibitors/biosynthesis ; Melanins/antagonists & inhibitors/*biosynthesis ; Melanocytes/*drug effects/*metabolism ; Melanoma, Experimental ; Membrane Glycoproteins/antagonists & inhibitors/biosynthesis ; Mice ; Microphthalmia-Associated Transcription Factor/antagonists & inhibitors ; Monophenol Monooxygenase/antagonists & inhibitors/biosynthesis/genetics ; Oxidoreductases/antagonists & inhibitors/biosynthesis ; Propiophenones/*pharmacology ; Signal Transduction/drug effects ; alpha-MSH/metabolism