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. 2022 Jan 26;14(2):287.
doi: 10.3390/pharmaceutics14020287.

In Vivo Quantification of the Effectiveness of Topical Low-Dose Photodynamic Therapy in Wound Healing Using Two-Photon Microscopy

Hala Zuhayri  1 , Viktor V Nikolaev  1 , Anastasia I Knyazkova  1 , Tatiana B Lepekhina  1 , Natalya A Krivova  1 , Valery V Tuchin  1   2 , Yury V Kistenev  1
Affiliations

In Vivo Quantification of the Effectiveness of Topical Low-Dose Photodynamic Therapy in Wound Healing Using Two-Photon Microscopy

Hala Zuhayri et al. Pharmaceutics. .

Abstract

The effect of low-dose photodynamic therapy on in vivo wound healing with topical application of 5-aminolevulinic acid and methylene blue was investigated using an animal model for two laser radiation doses (1 and 4 J/cm2). A second-harmonic-generation-to-auto-fluorescence aging index of the dermis (SAAID) was analyzed by two-photon microscopy. SAAID measured at 60-80 μm depths was shown to be a suitable quantitative parameter to monitor wound healing. A comparison of SAAID in healthy and wound tissues during phototherapy showed that both light doses were effective for wound healing; however, healing was better at a dose of 4 J/cm2.

Keywords: low-dose photodynamic therapy; photosensitizer; second-harmonic-generation-to-auto-fluorescence aging index of the dermis; two-photon microscopy; wound healing.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
A full-thickness cutaneous wound.
Figure 2
Figure 2
Digital photograph assessment of healing progression from day 0 to day 14: (a) control group; (b) LDPDT–5-ALA/1 J/cm2; (c) LDPDT–5-ALA/4 J/cm2; (d) LDPDT–MB/1 J/cm2; (e) LDPDT–MB/4 J/cm2.
Figure 3
Figure 3
Wound healing rate.
Figure 4
Figure 4
SHG (red) and AF (green) images of wound healing. (a) Control group; (b) LDPDT–5-ALA/1 J/cm2; (c) LDPDT–5-ALA/4 J/cm2; (d) LDPDT–MB/1 J/cm2; (e) LDPDT–MB/4 J/cm2.
Figure 5
Figure 5
(a) SHG (red) and AF (green) of healthy skin at depths from 4 to 80µm; (b) SHG (red) and AF (green) of a wound at depths from 4 to 80 µm.
Figure 6
Figure 6
SAAID for healthy skin.
Figure 7
Figure 7
(a) SAAID for control group; (b) SAAID for LDPDT–5-ALA/4 J/cm2 group.
Figure 8
Figure 8
(a) SAAID for LDPDT–5-ALA (4 J/cm2 and 1 J/cm2) groups; (b) SAAID for LDPDT–MB (4 J/cm2 and 1 J/cm2) groups.
Figure 9
Figure 9
SAAID for LDPDT–5-ALA and LDPDT–MB/4 J/cm2 groups.
Figure 10
Figure 10
Index SAAID on day 14 at 40 μm, 60 μm and 80 μm for healthy skin, control, LDPDT–5-ALA/1 J/cm2, LDPDT–5-ALA/4 J/cm2, LDPDT–MB/1 J/cm2, and LDPDT–MB/4 J/cm2.
Figure 11
Figure 11
p-values of SAAID for all groups at depths from 40 to 60 μm and 80 μm on day 14.
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