| Objective：To study the expression of granulocyte colony-stimulating factor receptor(G-CSFR) in human melanocytes and biologic impact of human recombinantgranulocyte colony-stimulating factor on human melanocytes.Methods：Human melanocvtes were primarily cultured and divided into two groupsto be cultured with or without the presence of G-CSF, hence to observe their growthand morphological changes. The expression of G-CSFR in human melanocytes、neutrophils and erythroleukemia(HEL92.1.7) were detected by flow cytometry.G-CSFR in the cellular localization was examined by immunofluorescence staining.Western Blotting and reverse transcription polymerase chain reaction(RT-PCR) wereemployed to measure G-CSFR protein and mRNA expression levels in humanmelanocytes、neutrophils and HEL92.1.7．Then melanocytes were treated withvarious concentrations(200、400、600、800μg/L)of rhG-CSF for72hoursrespectively，and MTT and Dopa-oxidase assay were used to detect the proliferationand tyrosinase activity of the treated melanocytes.Results：1. G-CSFR expression rates of human melanocytes(76.81±10.70%) weresignificantly higher than that of HEL92.1.7(2.53±1.54%)(P <0.01)，and lower thanneutrophils(85.76±15.71%)(P <0.05).2.The immunofluorescence staining showedG-CSFR positive expression in neutrophils and melanocytes, negative expression inHEL92.1.7cells. Its positive staining was mainly located in cell membrane and cytoplasm.3. The levels of G-CSFR protein and mRNA in melanocytes were lessthan neutrophils(P <0.01or P <0.05)，but significantly higher than HEL92.1.7(P <0.01). There was no significant relation between the levels of G-CSFR protein andG-CSFR mRNA with rhG-CSF(P﹥0.05).4. The proliferations of humanmelanocvtes were notably enhanced by the treatment with rhG-CSF at theconcentrations from200to800μg/L and600μg/L rhG-CSF(164.04±13.0%) had thestrongest capacity. The rhG-CSF at the concentrations of200to800μg/L had noeffect on tyrosinase activity of human melanocytes(P>0.05).Conclusion：G-CSFR is expressed in human melanocytes. Its positive staining ismainly located in cell membrane and cytoplasm. rhG-CSF can promote theproliferation of cultured human melanocytes，but it doesn′t contribute to tyrosinaseactivity of melanocytes. Objective：To study the expression of granulocyte colony-stimulating factor (G-CSF)and G-CSFR in human melanoma cells.Methods：The expression of G-CSFR in human melanoma cell lines (M14、A375)、human leukaemia cell line(U937) and erythroleukemia(HEL92.1.7) were detected byflow cytometry. G-CSFR in the cellular localization were examined byimmunofluorescence staining. Western Blotting and reverse transcription polymerasechain reaction(RT-PCR) were employed to measure G-CSFR protein and G-CSFRmRNA expression levels in M14、A375、U937and HEL92.1.7cells. G-CSF levels ofM14and A375cells were tested by enzyme-linked immune sorbent assay (ELISA).Results：1. G-CSFR expression rates of M14cells were(80.13±10.12%)，significantlyhigher than that of HEL92.1.7(2.69±1.74%)(P<0.01)，and slightly lower than U937(84.05±7.36%)(P <0.05). G-CSFR expression rates of A375cells were(56.29±7.38%)，higher than A375(P <0.05)，but lower than U937and M14cells(P <0.05).2.The immunofluorescence staining showed G-CSFR positive expression inU937、M14and A375cells, negative expression in HEL92.1.7cells. Its positivestaining was mainly located in cell membrane and cytoplasm.3. G-CSFR protein andmRNA were found in M14cells. Their expression levels were significantly higherthan HEL92.1.7(P <0.01), but slightly less than U937(P <0.05). The levels ofG-CSFR protein and mRNA in A375cells were less than U937and M14cells (P <0.05), but higher than HEL92.1.7(P <0.01).4. M14cells produced G-CSF in atime-dependent manner, G-CSF was not deteced in A375cells.Conclusion：G-CSFR is expressed in human melanoma cell lines M14and A375. Its positive staining is mainly located in cell membrane and cytoplasm.M14cells cansecrete G-CSF, but A375cells can′t produce G-CSF. G-CSF/G-CSFR autocrinepathway may exist in M14cells. |
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