Near-infrared low-level laser stimulation of telocytes from human myometrium

Abstract

Telocytes (TCs) are a brand-new cell type frequently observed in the interstitial space of many organs (see www.telocytes.com ). TCs are defined by very long (tens of micrometers) and slender prolongations named telopodes. At their level, dilations-called podoms (~300 nm), alternate with podomers (80-100 nm). TCs were identified in a myometrial interstitial cell culture based on morphological criteria and by CD34 and PDGF receptor alpha (PDGFRα) immunopositivity. However, the mechanism(s) of telopodes formation and/or elongation and ramification is not known. We report here the low-level laser stimulation (LLLS) using a 1,064-nm neodymium-doped yttrium aluminum garnet (Nd:YAG) laser (with an output power of 60 mW) of the telopodal lateral extension (TLE) growth in cell culture. LLLS of TCs determines a higher growth rate of TLE in pregnant myometrium primary cultures (10.3 ± 1.0 μm/min) compared to nonpregnant ones (6.6 ± 0.9 μm/min). Acute exposure (30 min) of TCs from pregnant myometrium to 1 μM mibefradil, a selective inhibitor of T-type calcium channels, determines a significant reduction in the LLLS TLE growth rate (5.7 ± 0.8 μm/min) compared to LLLS per se in same type of samples. Meanwhile, chronic exposure (24 h) completely abolishes the LLLS TLE growth in both nonpregnant and pregnant myometria. The initial direction of TLE growth was modified by LLLS, the angle of deviation being more accentuated in TCs from human pregnant myometrium than in TCs from nonpregnant myometrium. In conclusion, TCs from pregnant myometrium are more susceptible of reacting to LLLS than those from nonpregnant myometrium. Therefore, some implications are emerging for low-level laser therapy (LLLT) in uterine regenerative medicine.

Fig. 1

TCs in myometrial cell culture (fourth passage, day 3). a Phase contrast microscopy of typical a TC with very long telopodes. b Distribution of CD 34 immunopositivity in the same TC. c Cells that display the TC morphology express PDGFRα. Scale bar = 50 μm

Fig. 2

Comparative LLLS effect on TCs from nonpregnant and pregnant myometrium cell cultures (fourth passage, day 3). a TLE growth of TCs from nonpregnant myometrium. The angle of the TLE deviation is ≤30°. The time course of this effect is 20 s. b TLE growth of TCs from pregnant myometrium. The angle of the TLE deviation is between 30° and 72°, while the time course of this effect is longer—1 min and 6 s. Scale bar = 10 μm. c Telopodal local thickening upon optical tweezer stimulation was obtained in 25 % of TCs from pregnant myometrial cell culture. Scale bar = 40 μm. Yellow arrows indicate the TLE subjected to LLLS. The black arrows indicate the direction of the TLE. Each red square evidences the region of interest for the LLLS effect

Fig. 3

The comparative average of telopodal growth rate upon LLLS between TCs from nonpregnant and pregnant myometrium. *p < 0.01, unpaired Student’s t test

Fig. 4

Mibefradil effect on TLE upon LLLS in pregnant myometrium (myometrial interstitial cell culture at fourth passage, day 3). a Untreated TCs exposed to LLLS were considered as control. The time course of LLLS effect in these images (a–c) is 36 s. We can observe how a TLE grows (yellow arrow). b Mibefradil (1 μM) was perfused for 30 min, and afterwards, TCs were re-exposed to LLLS. Comparison of the TLE growth rate reveals that in 1 min and 4 s, the length of TLE is approximately the same as that in control and that the angle of the deviation is slightly above 30°. Yellow arrows indicate the TLE subjected to LLLS. The black arrows indicate the direction of the TLE. Each red square evidences the region of interest for the LLLS effect. Scale bar = 10 μm

Fig. 5

The comparative average growth rate of TLE upon LLLS between TCs from pregnant myometrium before and after acute mibefradil (30 min) treatment. *p < 0.05, paired Student’s t test