RESUMO
The precise causes of psoriasis, a chronic skin disorder characterized by hyperproliferation of keratinocytes and incomplete keratinization, are unclear. It is known that expression of helix-loop-helix transcription factor Id1, which functions as an inhibitor of differentiation, is upregulated in psoriatic skin. We investigated the effect of the antipsoriatic drug dithranol on mRNA and protein expression levels of Id1 in the HaCaT keratinocyte cell line. Cultured HaCaT cells were treated with 0-0.5 µg/mL dithranol for 30 min. After 2 and 4 h, total cellular RNA and total proteins were isolated from HaCaT cells, and quantitative real-time reverse transcriptase (RT-PCR) and Western blot were used to determine the mRNA and protein levels of Id1, respectively. Changes in normalized Id1 mRNA levels were observed only after 4 h of dithranol treatment. There was reduced expression of Id1 mRNA transcripts in the HaCaT cells treated with 0.1 µg/mL dithranol, but the reduction was not significant. The expression of Id1 mRNA was significantly downregulated (almost 50%) when 0.25 or 0.5 µg/mL dithranol was applied to the HaCaT cells. However, the normalized Id1 protein levels were not significantly affected. The molecular mechanisms underlying this finding should be investigated further to help determine the therapeutic action of this drug.
Assuntos
Antralina/farmacologia , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Proteína 1 Inibidora de Diferenciação/genética , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Western Blotting , Linhagem Celular , Fármacos Dermatológicos/farmacologia , Humanos , Proteína 1 Inibidora de Diferenciação/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Although the precise causes of psoriasis remain to be elucidated, psoriasis has been known as a disorder in which factors in the immune system, enzymes and other biochemical substances that regulate skin cell division are functionally imbalanced, thereby resulting in rapid proliferation of keratinocytes and incomplete keratinization. The expression of candidate genes such as E2A and caspase-9, which have been recognized to play a critical role in cellular proliferation/differentiation and apoptosis, is of great interest. They may be therapeutically targeted by the antipsoriatic drug, dithranol. We examined the molecular effects of dithranol on the mRNA and protein expression levels of E2A and caspase-9 in the HaCaT keratinocyte cell line. The HaCaT cells were treated with 0-0.5 µg/mL dithranol for 30 min. After dithranol was washed out, the HaCaT cells were cultured for 2 h, and their total cellular RNA and proteins were isolated. Quantitative real-time reverse transcriptase-polymerase chain reaction and Western blot were performed to determine the mRNA and protein levels of these two genes. We found that dithranol treatment in the range of 0.25-0.5 µg/mL slightly upregulated the mRNA expression of E2A and caspase-9 approximately 1.5- and 1.2-fold, respectively. However, undetectable change and minor downregulation of the protein expression levels were observed for E2A and caspase-9, respectively. Consequently, these genes appear not to be viable therapeutic targets for dithranol.
Assuntos
Antralina/farmacologia , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Caspase 9/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Caspase 9/genética , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Fármacos Dermatológicos/farmacologia , Expressão Gênica/efeitos dos fármacos , Humanos , Psoríase/tratamento farmacológico , Psoríase/genética , Psoríase/metabolismo , RNA Mensageiro/metabolismoRESUMO
Although the precise causes of psoriasis are unclear, it is widely accepted that psoriasis is a disorder in which factors in the immune system, enzymes, and other biochemical substances that regulate skin cell division are impaired, leading to rapid proliferation of keratinocytes and incomplete keratinization. Expression of the helix-loop-helix transcription factor Id1 (inhibitor of differentiation/DNA binding), functioning as an inhibitor of differentiation, is known to increase in psoriatic skin. However, the molecular involvement of this particular biomarker in the psoriatic immune system remains to be elucidated. We measured Id1 mRNA expression in peripheral blood mononuclear cells (PBMCs) of psoriatic patients and healthy controls using semi-quantitative reverse transcriptase-PCR. The normalized level of Id1 transcripts in psoriatic patients was about 2-fold higher than that in controls (P < 0.05). When we examined the proliferation rate of PBMCs, the stimulation index obtained from the phytohemagglutinin stimulation assay was not significantly different in psoriatic patients. In patients with psoriasis, there was no correlation between the stimulation index and the psoriasis area severity index. We suggest that Id1 has a role in causing psoriatic immune cell symptoms.
Assuntos
Proteína 1 Inibidora de Diferenciação/sangue , Psoríase/sangue , Regulação para Cima , Adulto , Idoso , Sequência de Bases , Proliferação de Células , Primers do DNA , Feminino , Humanos , Proteína 1 Inibidora de Diferenciação/genética , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/sangue , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Expression of serotonin 2A receptor (5-HTR2A) is known to increase in psoriasis, a chronic inflammatory skin disease. We investigated a possible association between the -1438A/G single nucleotide polymorphism (rs6311) in the promoter region of 5-HTR2A gene and psoriasis in a Thai population. One hundred and twelve psoriatic patients and 151 unrelated healthy controls were included in our study. Genotyping was performed using the polymerase chain reaction and restriction fragment length polymorphism techniques. We found no overall differences in genotype distributions and allele frequencies when comparing between the two groups. When we analyzed a subset of psoriatic patients classified by onset and severity, only the -1438A allele was significantly increased in patients with late-onset psoriasis when compared with the healthy control group (chi(2) = 4.77, d.f. = 1, P = 0.029, odds ratio = 2.298 [95% confidence interval = 1.126-4.691]). This single nucleotide polymorphism may be involved in late-onset psoriasis in this Thai population.