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Photobiomodulation induces microvesicle release in human keratinocytes: PI3 kinase-dependent pathway role

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Abstract

Microvesicles (MVs, 100–1000 nm diameter) are released into the extracellular environment by mammalian cells. MVs interact with near or remote cells through different mechanisms; in particular, MVs from human keratinocytes accelerate wound healing. Photobiomodulation by laser improves wound healing, but no information is available about its effects on MV release from human keratinocyte. Human-immortalized keratinocytes (human adult low-calcium high-temperature, HaCaT) were starved for 24 h and then irradiated using a 980-nm energy density of 0, 16.2, 32.5, and 48.7 J/cm2. After 24 h, MVs released in the conditioned medium were isolated, stained, and quantified using flow cytometry. MVs were distinguished from exosomes on the basis of their volume (forward scatter signals). In some experiments, phosphatidylinositol 3-kinase (PI-3K) activity, involved in MV release and stimulated by laser light, was inhibited by pre-treating cells with Wortmannin (WRT, 10 μg/mL). MVs were observed in HaCaT-conditioned medium both in basal- and laser-stimulated conditions. Photobiomodulation therapy, also known as PBMT, was able to increase MV release from human keratinocytes reaching a maximum effect at 32.5 J/cm2 with a stimulation of (148.6 ±15.1)% of basal (p<0.001). PI-3K activity inhibition strongly reduced both basal- and laser-induced MV release; but PBMT by laser still increased MV release, compared to basal values in the presence of WRT. In vitro near infrared photobiomodulation increased the releasing of MVs from human keratinocytes, while Wortmannin, a PI-3K inhibitor, negatively affects both basal- and laser-induced releasing. Laser-induced MV release could be a new effect of biostimulation on the wound healing process.

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Acknowledgements

The authors thank Dr. Laura Begani from Wound Healing Laboratory for her helpful technical assistance and Dr. Joseph A. Simonpietri for the linguistic review of the paper.

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  1. Innovative Research Laboratory for Wound Healing, Health Sciences Department, Università del Piemonte Orientale, via Solaroli 17, 28100, Novara, Italy

    Flavia Lovisolo, Flavia Carton, Sarah Gino & Filippo Renò

  2. Dental Clinic, Health Sciences Department, Università del Piemonte Orientale, via Solaroli 17, 28100, Novara, Italy

    Mario Migliario

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  1. Flavia Lovisolo
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  2. Flavia Carton
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Correspondence to Filippo Renò.

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Flavia Lovisolo and Flavia Carton shared first authorship

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Lovisolo, F., Carton, F., Gino, S. et al. Photobiomodulation induces microvesicle release in human keratinocytes: PI3 kinase-dependent pathway role. Lasers Med Sci 37, 479–487 (2022). https://doi.org/10.1007/s10103-021-03285-2

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