Clinical Study of Vitamin D for the Treatment of Vitiligo
the Biological Effect of Vitamin D on Melanocytes
Objective: To study the effects of 1α ,25-dihydroxyvitamin D3 on cell proliferation and melanin synthesis of normal human melanocytes.
Methods: Melanocytes of foreskins of healthy men were cultured and treated with various concentration of 1α , 25-dihydroxyvitamin D3. After incubation 48 hours,Cell proliferation was measured with MTI assay. The synthesis of melanin was determined by NaOH method . The tyrosinase activity was detected by DOPA-oxidation.
Results: 1α , 25-dihydroxyvitamin D3 could promote the proliferation of cultured melanocyte at 10-9- 10-5mol / L level. Especially,cells increased most at 10-7mol / L 1α ,25-dihydroxyvitamin D3 ,but the proliferation of cells is not significant compared with the untreated control （P>0.05） .1α ,25-dihydroxyvitamin D3 could significantly increase the tyrosinase activity and melanin synthesis at 10-9- 10-8mol / L Preincubation with 1α 25-dihydroxyvitamin D3 10-9- 10-8mol / L increased tyrosinase activity 137%, 123% （P<0.05 ） and melamin synthesis 40.63%, 18.75%,respectively （P<0.05 ） .The effect of 1α , 25-dihydroxyvitamin D3 on melanocyte was not dose-depended.
Conclusion: 1α, 25-dihydroxyvitamin D3 could increase the tyrosinase activity and melamin synthesis of melanocytes. Morover 1α, 25-dihydroxyvitamin D3 can promote proliferation of melanocyte.
Objective: Tyrosinase and microphthalmia associated transcription factor correlate with the differention of melanocytes. To explore the mechanism underliing promoting defferention action by 1α , 25-dihydroxyvitamin D3, the study was performed to investigate how 1α , 25-dihydroxyvitamin D3 effect the tyrosinase and microphthalmia associated transcription factor.
Method: The melanonacytes were treated with 10-9 10-7 10-5mol/l 1α , 25-dihydroxyvitamin D3 for 24 hours. Western blot and semiquanttitative reverse transcriptase polymerase chain reaction were performed to assess the protein and mRNA expression levels of microphthalmia-associated transcription factor tyrosinase.
Result: Microphthalmia associated transcription factor protein level increased 157.08%, 138.2%, 97.9%, respctively in melanocyte treated with 10-9 10-7 10-5mol/l 1α,25-dihydroxyvitamin D3 for 24h compared with untreated control （P<0.05） .Tyrosinase protein level increased 270.4%, 92.86% respectively, in melanocytes treated with 10-9, 10-7mol/l 1α , 25-dihydroxyvitamin D3 with untreated control （P<0.05） .But tyrosinase mRNA level did not increase following 1α, 25-dihydroxyvitamin D3 treatment.
Conclusion: These results show that 1α , 25-dihydroxyvitamin D3 fuctions by initiallystimulating protein levels of microphthalmia associated transcription factor andtyrosinase expression, which thereby leads to increases in melanin production bymelanocytes.
Objective:VitaminD3,one of the major regulators of calcium homeostasis was reportedthat it can induced an increase in intrtracellular free calcium（Cai）of keratinocyte, butwheather it effects the calcium homeostasis in melanocyte is still unknown. We usefluorescein stain to investigate the effects of 1α, 25-dihydroxyvitamin D3 on theconcentration of Cytosolic free Ca2+ in human melanocyte in vitro.
Methods: An in vitro incubation system was used; the concentration of Cytosolic freeCa2+ was quantitated using the fluorescent Ca2+ indicator Fura-4. Under OlympusConfocal microscope, the time course of the fluorescence response of Fluo4-AM wasobtained from time lapse image every 5s.
Results: 1α , 25-dihydroxyvitamin D3 can promote the increase of Cytosolic Ca2+concentration in human melanocyte at 10-5 10-7 10-9mol / L. 15second after 1α,25-dihydroxyvitamin D3 was added, the intrtracellular free calcium（Cai） inmelanocyte rised to （1.07±0.07） （1.11±0.04） （1.02±0.03）, and increase 6.6%,11.1 %和4.1 %respectivelly, compared with the control （P<0.05） .
Conclusion: 1α, 25-dihydroxyvitamin D3 can promote the increase of Cytosolic Ca2+concentration in human melanocyte at 10-9-10-5mol / L , especially at 10-7mol /L .
The result indicate that the increase of Cytosolic Ca2+ level involved in the signalparthway by which the melanocyte differentiation and melamin sythesis action of 1α ,25-dihydroxyvitamin D3.Objective: To assess the the safety and efficacy of topical tacacitol.
Methods: We performed a within patient controlled study. Thirty-four patients withvitiligo enrolled in the study. Symmetrical or near lesions were selected as referencelesions. There was no evidence of spontaneous repigmentation in the lesions. In thisrandomized comparison study, tacalcitol or placebo was applied twicely a day to thereference lesions. Patients were examined monthly intervals and record their responseto the treatment. All data were analysed by SPSS.
Results: Thirty patients （16 men, 14 women） were evaluated. Four patients failed tocomplete the study. Eight of the patients’s taget lesions were on faces and necks;sixteen of the taget lesins on trunk, six of the taget lesins on extremities.Treatment waswell tolerated. 73.3% of those treated with tacalcitol showed good repigmentation,36.7% of those treated with placebo （P<0.05） . Among the second and thirdmonth,53.3% and 76.7%of those treated with tacalcitol showed initial repigmentation ,26.7% and50% of those treated with placebo （P<0.05） respectively. There is nocorrelation between clinical feature and the efficacy of tacalcitol.
Conclusion: The initial repigmentation was seen earlier in lesions treated withtacalcitol than with placebo.Topical tacalcitol appears to be an effective andwell-tolerated treatment for vitiligo.
Objective: To study and compare the efficacy of combined tacalcitol and monochromatic excimer light （MEL） 308-nm therapy vs MEL 308-nm monotherapy in treating vitiligo.
Methods: Thirty-eight patients with vitiligo were enrolled in a within patient controlled clinical trial. Symmetrical or nearby lesions were randomly selected to be treated by either tacalcitol or placebo. Tacalcitol or placebo was applied twicely a day to the reference lesions. Each lesion was treated weekly by the MEL 308-nm with the same dosage on the two sides,for a total of 12 sessions.The ointment were applied 2h before the MEL 308-nm. Patients were examined at monthly intervals. The mean number of sessions and the cumulative dosage for initial and excellent repigmentation were calculated.
Result: Thirty-five patients （19 men, 16 women） were evaluated. Three patients failed to complete the study. Sixteen of the patients's taget lesions were on faces and necks, fifteen of taget lesions on trunk, four of target lesions on extremities.Treatment were well tolerated.The mean±SEM cumulative dose and number of excimer light exposures for initial repigmentation were 4.27±3.59J/ cm2 and 4.89±3.16 on the tacalcitol side, and 5.36±4.12J/cm2 and 5.69±3.29on the placebo side, respectively （P<0.05）. For excellent repigmentation, respective values were 7.72±5.64 J/ cm2 and 7.79±4.70 on the tacalcitol side, 8.18±4.868J/ cm2 and 8.4±3.92 on the placebo side （P>0.05）. Treatment with tacalcitol result in significantly higher percentage of repigmentation （71.4%）, compared with placebo （54.3%）.Conclusion: Our results have shown that concurrent topical tacalcitol potentiates the efficacy of the MEL 308-nm in the treatment of vitiligo, and that this combination achieves earlier dosage.