Integrating tacrolimus into eutectic oil-based microemulsion for atopic dermatitis: simultaneously enhancing percutaneous delivery and treatment efficacy with relieving side effects

Figures & data

Figure 1 The AFM image of FK506 MCE ME.

Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Table 1 Cumulative permeated amount, flux, and retention of FK506 from different formulations after 24-hr penetration in vitro (mean ± SD, n=6)

	Permeated amount (μg· cm^−2)	Flux (μg· cm^−2· hr^−1)	Retention (μg· cm^−2)
PC	0.32±0.04	0.10	1.83±0.44
Oil free	0.34±0.05	0.12	1.83±0.12
Ointment	4.76±0.60**	0.28**	2.41±0.29
MCE ME	4.39±0.54**	0.28**	3.69±0.47^++
MCE ME gel	4.46±0.73**	0.29**	3.79±0.26^++

Notes: **P<0.001 in comparison with PC or oil free; ⁺⁺P<0.001 in comparison with PC, oil free, or ointment. Data represented mean ± SD, n=6/group.

Abbreviations: FK506, tacrolimus; PC, the solution of 0.1% tacrolimus (FK506,w/v) dissolved in propylene carbonate; oil free, 0.1% (w/v) FK506 solution containing the same surfactant and cosurfactant as microemulsion but without menthol/camphor eutectic; ointment, 0.1% (w/w) FK506 commercial ointment; MCE ME, 0.1% (w/v) FK506 microemulsion based on menthol/camphor eutectic; MCE ME gel, 0.1% (w/w) FK506 microemulsion gel based on menthol/camphor eutectic.

Figure 2 In vitro skin penetration of FK506 from different formulations (A) and skin retention of FK506 after 24 h penetration (B).

Note: **P<0.001.

Abbreviations: FK506, tacrolimus; PC, The solution of 0.1% tacrolimus (FK506,w/v) dissolved in propylene carbonate; Oil free, 0.1% (w/v) FK506 solution containing the same surfactant and cosurfactant as microemulsion but without menthol/camphor eutectic; Ointment, 0.1%(w/w) FK506 commercial ointment; MCE ME, 0.1% (w/v) FK506 microemulsion based on menthol/camphor eutectic; MCE ME gel, 0.1% (w/w) FK506 microemulsion gel based on menthol/camphor eutectic; h, hours; SD, standard deviation.

Figure 3 Cell viability of HaCaT cells after incubation with MCE ME vehicle of different concentrations for 12 h and 24 h, respectively.

Abbreviations: HaCaT, immortal human keratinocytes; MCE, ME vehicle; Microemulsion based on menthol/camphor eutectic without FK506; SD, standard deviation.

Figure 4 Schematic diagram of the experimental procedure (A); effects of different formulations containing 0.1% FK506 on dermatitis scores throughout the experiment (B); and clinical features of AD-like skin lesions in the end of experiment (C).

Notes: *P<0.05 in comparison with DNCB; **P<0.001 in comparison with DNCB; ^#P<0.05 in comparison with DNCB + Ointment; ^##P<0.001 in comparison with DNCB + Ointment; ⁺⁺P<0.001.

Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment; AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free, AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle, AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506; SD, standard deviation.

Figure 5 The ear thickness (A), spleen index (B) on day 28, and TEWL (C) of the mice treated with different formulations throughout the experiment.

Notes: *P<0.05 in comparison with DNCB; **P<0.001 in comparison with DNCB; ^#P<0.05 in comparison with DNCB + Ointment; ^##P<0.001 in comparison with DNCB +Ointment; ⁺P<0.05; ⁺⁺P<0.001.

Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; TEWL, transdermal water loss; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment: AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free, AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle; AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506; SD, standard deviation.

Figure 6 Histological features of the skin treated with different formulations in AD mouse model. The sections were stained with hematoxylin and eosin (A) and toluidine blue (B), respectively.

Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment; AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free; AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle, AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506.

Figure 7 Effects of different formulations on serum total IgE.

Notes: *P<0.05 in comparison with DNCB; **P<0.001 in comparison with DNCB; ^#P<0.05 in comparison with DNCB + Ointment; ^##P<0.001 in comparison with DNCB +Ointment; ⁺P<0.05; ⁺⁺P<0.001.

Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment: AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free, AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle, AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506; SD, standard deviation.

Figure 8 Inhibition effects of different formulations on scratch frequency (A) and substance P expression (B).

Notes: *P<0.05 in comparison with DNCB; **P<0.001 in comparison with DNCB; ^#P<0.05 in comparison with DNCB + Ointment; ^##P<0.001 in comparison with DNCB +Ointment; ⁺P<0.05; ⁺⁺P<0.001.

Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment: AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free, AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle, AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506; SD, standard deviation.