Determination of clobetasol propionate, tazarotene and tazarotenic acid in Bama mini-pig skin by UPLC-MS/MS: Application to pharmacokinetic and drug-drug interaction studies.

Tazarotene and clobetasol propionate are efficacious for the treatment of psoriasis. The plasma pharmacokinetic assessments of tazarotene or clobetasol propionate have been reported. However, the pharmacokinetic characteristics of tazarotene and clobetasol propionate in skin when used together have not been studied. In the present study, sensitive and rapid methods were developed for the determination of clobetasol propionate, tazarotene and its active metabolite tazarotenic acid in Bama mini-pig skin by UPLC-MS/MS. After homogenization and pretreatment of skin samples, the separation was performed on a WondaSiL C18 column (4.6 × 150 mm, 5 µm) for tazarotene and clobetasol propionate. The separation of tazarotenic acid was achieved on a BDS HYPERSIL C18 column (4.6 × 100 mm, 2.4 µm). All the analytes were quantified with positive electrospray ionization and multiple reactions monitoring mode. The assay was validated in the range of 22-1111 ng/g for tazarotene and clobetasol propionate, 2-111 ng/g for tazarotenic acid in skin samples. The methods were fully validated to meet the requirements for bioassay in accuracy, precision, recovery, reproducibility, stabilities and matrix effects, and successfully applied to evaluate the novel combination ointment of tazarotene and clobetasol propionate. The obtained intradermal content-time curves characterized the dermal absorption and metabolism features of the combination ointment. It was also found that there was no significant drug-drug interaction trend between tazarotene and clobetasol propionate. The obtained results would be essential for the development and clinical applications of this novel combination ointment.

Evaluation of a 13-hexyl-berberine hydrochloride topical gel formulation.

13-hexyl-berberine hydrochloride (HB-13) is a derivative from berberine which finds widespread applications in the treatment of infectious pathogens including fungi, bacteria, parasites and viruses. As our continuing efforts for treatment of herpes simplex virus (HSV), we studied the topical delivery and safety of HB-13 in a gel formulation (0.5%) in a pig model. Our studies demonstrated the maximal HB-13 concentration was 2.51 µg/mL, which was more than the half maximal inhibitory concentration (IC50) as we previously reported. In addition, there was no sign of irritation or histological aberrance for stripped skin continuously applied with 0.5% HB-13 gel for 21 days. In conclusion, 0.5% HB-13 gel can achieve effective anti-HSV concentration in the dermis and it is safe to use.

[Effects of triptolide on the production of interferon-gamma in human peripheral blood mononuclear cell and phosphorylation of signal transducer and activator of transcription-1 and production of interleukin-8].

OBJECTIVE: To investigate the effects of triptolide on the production of interferon-gamma (IFN-gamma) in human peripheral blood mononuclear cell (PBMC) and interleukin-8 (IL-8) in HaCaT keratinocytes and phosphorylation of signal transducer and activator of transcription-1 (STAT1) of IFN-gamma signal transduction pathways in HaCaT cells. METHODS: Human PBMC was induced by phytohaemagglutinin (PHA-L) and HaCaT cells were stimulated by recombinant human IFN-gamma (rhIFN-gamma). The productions of IFN-gamma and IL-8 in cells were detected by ELISA. The expression of STAT1 and its phosphorylation were analyzed by Western blot. RESULTS: Triptolide inhibited the production of IFN-gamma in human PBMC induced by PHA-L in a dose-dependent manner (P < 0.05, P < 0.01, P < 0.001) and the 50% inhibitory concentration (IC50) value was 5.96 x 10(-11) mol/L. IL-8 production in HaCaT cells induced by rhIFN-gamma in vitro was also inhibited by triptolide (P < 0.001) and the IC50 value was about 1.15 x 10(-13) mol/L. The expressions of phosphorylated STAT1 in HaCaT cells stimulated by rhIFN-gamma was inhibited by triptolide (P < 0.01) and the IC50 value was about 9.45 x 10(-11) mol/L. CONCLUSION: Triptolide can inhibit the production of IFN-gamma in human PBMC and downregulate IL-8 level in HaCaT keratinocytes induced by rhIFN-gamma. Triptolide can inhibit the phosphorylations of STAT1 of IFN-gamma signal pathway in HaCaT keratinocytes stimulated by IFN-gamma.

Inhibition of the nuclear factor-kappaB signaling pathway by leflunomide or triptolide also inhibits the anthralin-induced inflammatory response but does not affect keratinocyte growth inhibition.

We performed this study to determine the relationship between activation of nuclear factor (NF)-kappaB and inhibition of keratinocyte growth by anthralin, which not only might be useful for a better understanding of the role of NF-kappaB in the pathogenesis of psoriasis, but also indicate whether the inflammatory reaction induced by anthralin is inseparable from its antipsoriatic activity. The involvement of NF-kappaB was assessed using the antipsoriatic drugs leflunomide and triptolide (T0) as effectors, since they can inhibit NF-kappaB activation induced by anthralin. The results showed that the inhibition of keratinocyte growth by anthralin was not related to the activation of NF-kappaB. Using sodium salicylate, a known NF-kappaB inhibitor, further confirmed this conclusion. Thus it might be possible to inhibit the inflammatory response induced by anthralin via repression of NF-kappaB activation. We found that leflunomide or T0 could significantly inhibit the mRNA overexpression of interleukin-8 and intercellular adhesion molecule-1 in keratinocytes induced by anthralin. Taken together, our data indicate that the growth inhibition of anthralin is related to the NF-kappaB-independent signaling pathway, and that leflunomide or T0 could control proinflammatory cytokine expression induced by anthralin via inhibiting the activation of NF-kappaB.