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. 2021 Mar 18;24(2):145–152. doi: 10.1298/ptr.E10064

Monophasic Pulsed Current Stimulation of Duty Cycle 10% Promotes Differentiation of Human Dermal Fibroblasts into Myofibroblasts

Mikiko UEMURA 1, Masaharu SUGIMOTO 2, Yoshiyuki YOSHIKAWA 3, Terutaka HIRAMATSU 4, Taketo INOUE 5
PMCID: PMC8419484  PMID: 34532210

Abstract

Objective: Many clinical trials have shown the therapeutic effects of electrical stimulation (ES) in various conditions. Our previous studies showed that ES (200 μA and 2 Hz) promotes migration and proliferation of human dermal fibroblasts (HDFs). However, the effective duty cycle and the effect of ES on myofibroblast differentiation are unclear. This study aimed to investigate the relationship between duty cycle and myofibroblast differentiation. Methods: HDFs were subjected to ES (200 μA and 2 Hz) for 24 h with the duty cycle adapted at 0% (control), 10%, 50%, or 90%. α-smooth muscle actin (SMA) and transforming growth factor (TGF)-β1 mRNA and α-SMA protein expressions were assessed. Collagen gel contraction was observed for 48 h after ES initiation and the gel area was measured. Cell viability and pH of culture medium were analyzed for cytotoxicity of the ES. Results: Cell viabilities were decreased in the 50% and the 90% groups but ES did not influence on pH of culture media. ES with a duty cycle of 10% significantly promoted the mRNA expression of α-SMA and TGF-β1. α-SMA protein expression in the 10% group was also significantly higher than that of the control group. Collagen gel subjected to ES with a duty cycle of 10% was contracted. Conclusion: Duty cycle can influence on myofibroblast differentiation and ES with a duty cycle 10% is the effective for wound healing.

Keywords: Electrotherpy, Pressure ulcer, Microcurrent, Wound healing

Pressure injury is a chronic wound that decreases the quality of life of patients because of the associated pain and treatment costs1). Thus, there is a need for an effective and low-cost treatment for pressure injury. Electrical stimulation (ES) treatment is recommended for pressure injury healing2), and clinical studies have shown its therapeutic effects against pressure injuries3-6). Healing of chronic wounds is delayed because their inflammation period is prolonged and granulation tissue formation is inhibited7). Granulation tissue formation is crucial for wound closure, and human dermal fibroblasts (HDFs) are a key factor in this process because their migration toward wound sites, proliferation, and differentiation into myofibroblasts are necessary for granulation tissue formation8). Our previous studies showed that a monophasic-pulsed current (intensity; 200 μA, frequency; 2 Hz) promotes the migration of HDFs toward the cathode and their proliferation9,10), and that ES with the same conditions has a good therapeutic effect on pressure injuries11). Thus, we revealed the optimum intensity and frequency of ES for pressure injury healing.

Myofibroblasts have a strong contractile ability and are involved in the wound healing process; the differentiation from fibroblasts to myofibroblasts results in granulation tissue contraction leading to wound closure8). Myofibroblasts are characterized by the expression of α-smooth muscle actin (α-SMA), and transforming growth factor-β (TGF-β) influences myofibroblasts to promote collagen synthesis in granulation tissue12). The effects of ES on the differentiation of HDFs have been demonstrated in in vivo and in vitro studies13,14). Direct current upregulates TGF-β1 and collagen I/III expression in mouse fibroblasts and promotes α-SMA expression in HDFs13). However, a pulse width of 300 ms within 600 ms promotes higher α-SMA expression in HDFs than a pulse width of 10 ms within 1200 ms14). Thus, the effects of ES on HDFs are influenced by pulse width. The pulse width is a result of the duty cycle, which is the on-and-off ratio of one stimulation (Fig. 1); a duty cycle of 100% indicates direct current, and the effect of duty cycle on HDF differentiation with an ES of 200 μA and 2 Hz, which promotes migration and proliferation in HDFs, is unclear.

Fig. 1.

Fig. 1

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Duty cycle.

Duty cycle is the on-off ratio of stimulation. Both figures present a 2-Hz monophasic pulsed current. Pulse width indicates stimulation duration during one stimulation cycle, and this is determined by the duty cycle and frequency. (A) The waveform with a duty cycle of 50% for which the on-off cycle is 1:1 and pulse width is 250 ms. (B) Duty cycle of 90%, with an on-off cycle of 9:1 and pulse width of 450 ms. The waveform approaches direct current as the duty cycle reaches 100%.

Moreover, safety is mandatory for medical instruments including the ES device. In our previous studies, we did not assess the effects of long-term stimulation in HDFs. Low-intensity or small-electric fields within 10 h showed good effects on HDFs, promoting migration and proliferation9,10,15,16). Zhang et al. and Wang et al. showed that longer ES has good effects on the migration and secretion of pro-healing cytokines14,15); however, the therapeutic effects and adverse effects of long-term micro current stimulation in HDFs are not clear. Therefore, in this study, we adopted >10-h stimulation and confirmed the effects of ES on cell viability. In clinical trials, it is necessary to decide the pulse width, as well as waveform, intensity, and frequency, to conduct ES. Thus, we hypothesized that the duty cycle might have an effect on promoting α-SMA expression in HDFs and their differentiation into myofibroblasts.

Method

Cell culture and electrical stimulation

Primary HDFs (CC-2511; Clonetics, San Diego, CA, USA) derived from a 33-year-old woman were used. HDFs were cultured in low-glucose Dulbecco's modified eagle medium (DMEM; Wako, Osaka, Japan) supplemented with 10% fetal bovine serum (Nichirei, Tokyo, Japan) and 5% penicillin-streptomycin Solution (Wako) in a CO2 incubator at 37°C. HDFs that had undergone 3-7 passages were used in the experiments. HDFs were seeded in a 35-mm tissue culture dish (Wako) and cultured for 24 h. The culture medium was refreshed and platinum electrodes were set on both sides of a dish (Fig. 2). Thereafter, HDFs were subjected to monophasic-pulsed current stimulation (intensity, 200 μA; frequency, 2 Hz) for 24 h in a CO2 incubator. HDFs without ES were used as controls. Duty cycles of 10%, 50%, or 90% were adapted to confirm the influence of differences in duty cycles.

Fig. 2.

Fig. 2

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Electrical stimulation.

Human dermal fibroblasts (HDFs) were seeded in a 35-mm tissue culture dish and cultured for 24 h in a CO2 incubator at 37°C. Electrodes were set on both sides of a dish and connected to the electrical stimulation (ES) device. Then, HDFs were exposed to the ES for 24 h in a CO2 incubator at 37°C.

Cell viability and cell number

The effect of ES on cytotoxicity was assessed because the ES that induces cell death cannot be clinically applied even if it promotes pro-fibrotic factors and the contractile ability of HDFs. Cell viability and cell numbers were analyzed after ES by trypan blue staining17,18). ES that induced cell death was not used to analyze gene and protein expression.

TaqMan real-time RT-PCR

To detect whether ES promoted fibroblast differentiation into myofibroblasts, the mRNA expression of α-SMA and TGF-β1 were analyzed. After ES, the total RNA was extracted from HDFs using the High Pure RNA Isolation Kit (Roche, Basel, Switzerland). RNA was reverse-transcribed into cDNA. The mRNA expression of α-SMA, TGF-β1, and GAPDH was then analyzed by TaqMan real time RT-PCR. Relative gene expression was calculated using the ΔΔCt method after normalization to GAPDH expression.

Western blotting

The expression of α-SMA is an indicator of myofibroblasts, and the expression of α-SMA was assessed by western blotting. After ES, protein was extracted from HDFs using pro-prep TM (Cosmo Bio, Tokyo, Japan) according to the manufacturer's instruction. Pooled samples containing the same amount of protein were subjected to SDS-polyacrylamide gel electrophoresis (BIO-RAD, CA, USA) and transferred onto membranes (Thermo Fisher Scientific, MA, USA). After blocking with blocking reagents (GE Healthcare, Buckinghamshire, UK), the membranes were incubated overnight at 4°C with the following primary antibodies: anti-α-SMA antibody (ab5694; Abcam, Cambridge, UK) or anti-GAPDH antibody (G8795; Sigma-Aldrich, MO, USA). The membranes were then incubated with the appropriate secondary antibodies, anti-rabbit IgG antibody (NA934V; GE Healthcare) or anti-mouse IgG antibody (NA931V; GE Healthcare), for 1 h at room temperature. The membranes were incubated with ECL mix (GE Healthcare), and the blots were quantified by densitometry (Chemi Doc XRS; BIO-RAD). Western blotting was performed four times using pooled samples (n = 4 per group). Data were normalized using GAPDH and by relative expression and compared with levels in the control group with Image J (National Institutes of Health, Bethesda, MD, USA).

Collagen gel contraction assay

The collagen gel contraction assay was performed to analyze the contractile ability of HDFs using a collagen-based cell contraction assay kit (Cell Biolabs, CA, USA) according to the manufacturer's protocol. First, 240 × 104 cells were mixed with collagen solution and incubated for 1 h at 37°C. The solution was then added to 1 mL of DMEM and incubated for 48 h at 37°C in a CO2 incubator. Collagen gels were separated from the dishes with spatula and subjected to ES for 24 h. Gel contractions were observed at 24 and 48 h after ES initiation. The gel areas were measured using image J and the area change rates were calculated for each dish.

Measurement of culture medium pH

pH was measured in each culture medium after ES. A pH probe (Mettler Toledo, Zurich, Switzerland) coupled with a pH meter (Mettler Toledo) was used to measure pH. The pH probe was calibrated with a standard pH buffer solution (pH 4.01, 6.86, and 9.18; AS ONE, Osaka, Japan) before immersing the probe in culture medium to measure pH.

Statistical analysis

All data were tested using the Shapiro-Wilk test and F-test. Student's t-tests were used to analyze the data that followed a normal distribution with equal variances, and the results with p < 0.05 were considered statistically significant. Bonferroni correction was used and p < 0.0125 was considered statistically significant when the data were compared among more than three groups. When the data did not follow a normal distribution, the Mann-Whitney U test was used to evaluate data. Results with p < 0.05 were considered statistically significant. All statistical analyses were performed using EZR (Saitama Medical Center, Jichi Medical University, Saitama, Japan), which is a graphical user interface for R (The R Foundation for Statistical Computing, Vienna, Austria).

Ethical approval

In this study, we used a primary cell culture sold by Clonetics, and therefore, the study did not require ethical approval.

Results

ES with high duty cycle has adverse effects on HDFs

To better understand the adverse effect of ES on fibroblasts, we examined the cell viability and cell numbers after ES. Cell viability was decreased in the 50% and 90% groups (p < 0.0125 vs. control) and the viability of the 90% group was less than 90% (Fig. 3). However, ES with a 10% duty cycle did not decrease cell viability. Cell number after ES in the 90% group was decreased to less than 70%, but there were no significant differences between control and stimulus groups (v.s. 10%; p = 0.237, v.s. 50%; p = 0.339, and v.s. 90%; p = 0.021, respectively). These results indicate that cell toxicity might occur as the duty cycle approaches 100%, and direct current and high duty cycle could suppress cell proliferation. Next, to confirm ionization by ES, we measured the pH of each culture medium. The pH values of culture media were 7.77 ± 0.14 (control), 7.93 ± 0.09 (duty cycle 10%), 7.91 ± 0.05 (duty cycle 50%), and 7.96 ± 0.08 (duty cycle 90%), with no significant differences between the control and stimulus groups.

Fig. 3.

Fig. 3

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Adverse effects of electrical stimulation on human dermal fibroblasts.

The cell viability of human dermal fibroblasts after electrical stimulation is shown. Data are presented as the mean ± SD; n = 4 per group. **, p < 0.01, significant difference between the electrical stimulation group and the control group. The statistical differences between the control and the treated groups were tested by Student's t-tests with a Bonferroni correction.

ES promotes the expression of α-SMA and TGF-β1

The mRNA and protein expression levels were compared between the control and the 10% groups because ES with a duty cycle of 50% and 90% introduced cell death. ES with a 10% duty cycle significantly promoted the mRNA expression of α-SMA and TGF-β1 (Fig. 4, p = 0.029 and p = 0.029). The mRNA expression of α-SMA in the 10% group was more than twice that in the control group. The protein level of α-SMA in the 10% group was similarly significantly higher than that of the control group (Fig. 5, p = 0.046). Thus, ES with a duty cycle of 10% might promote the differentiation of fibroblasts into myofibroblasts.

Fig. 4.

Fig. 4

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The expression of α-SMA and TGF-β1 in human dermal fibroblasts after electrical stimulation.

Relative expression of (A) α-SMA and (B) TGF-β1 mRNA. Data are presented as box-whisker plots. The statistical differences between the control and the 10% group were tested by Mann-Whitney U test; n = 4 per group. *, p < 0.05, significant difference between the 10% group and the control group.

Fig. 5.

Fig. 5

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α-SMA protein expression in human dermal fibroblasts after electrical stimulation.

(A) A representative blot is shown. (B) Western blot quantitation. Data are presented as box-whisker plots; n = 4 per group. The statistical difference between the control and the 10% group was tested by the Student's-t test.

ES promotes collagen gel contraction

To confirm the effect on wound closure, we investigated the contractile capacity of fibroblasts using the three-dimensional (3D) collagen gel contraction assay. Collagen gel contraction rates were compared between control and 10% groups. The areas of collagen gels were reduced after stimulation (Fig. 6A). Fig. 6B, 6C show the rate of change in the gel areas since the beginning of ES in gels with HDFs contracted for 24 and 48 h. The gel subjected to ES with a 10% duty cycle contracted more than the control at 24 and 48 h after ES initiation (p = 0.0096, p = 0.0115, respectively), and the gel of the 10% group was contracted by approximately 50% from the beginning. These results indicate that ES of a 10% duty cycle promoted the contractile ability of HDFs with myofibroblast differentiation, and this was maintained both during and after ES.

Fig. 6.

Fig. 6

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Collagen gel contraction.

(A) The collagen gel contraction in human dermal fibroblasts 24 and 48 h after electrical stimulation (ES) initiation. (B) Quantification of collagen gel contraction 24 h after ES initiation. (C) Quantification of collagen gel contraction 48 h after ES initiation. Data are presented as the mean ± SD; n = 4 per group. **, p < 0.01, significant difference between the ES with a duty cycle of 10% group and the control group. The statistical difference between the control and the 10% group was tested by Student's t-test.

Discussion

The present study showed a novel way to establish the optimum duty cycle of ES therapy for healing pressure injuries by analyzing the effects of duty cycle on HDFs. The monophasic-pulsed microcurrent of a 10% duty cycle promoted α-SMA and TGF-β1 expression and collagen gel contraction without cytotoxicity. These results suggest that a monophasic-pulsed current promotes fibroblast differentiation into myofibroblasts and that this effect is related to the duty cycle.

Viability of HDFs exposed to ES with a duty cycle of 50% and 90% was decreased to less than 90%, and cell number in the 90% group was decreased. This result indicated that an ES with a >50% duty cycle has toxic effects for HDFs. We hypothesized that the cell toxicity of ES was related to electrolysis and thus measured the pH of each culture medium. The pH of culture media in ES groups were alkalized (around 7.9), but there were no significant differences between control and ES groups. These results indicated that the cell toxicity of ES was not due to the change in pH. Some studies have shown that ES induces cell apoptosis and necrosis depending on voltage or pulse length with electroporation in the cell membrane and influx of Ca2+ 19-21). Pulse length is decided by the duty cycle and a high duty cycle means a long pulse length. Thus, ES with a high duty cycle might alter the permeability of cell membranes, inducing cell death. Moreover, the pH of the wound surface is higher with severe pressure injury and the pH of the wound surface is increased when pressure injury deteriorates22,23). Therefore, it is necessary to use electrodes in which ionization tendency is large in clinical trials because ES might increase the pH of tissues to avoid alkalization of wound surface.

Regarding the contractile ability of HDFs with ES, that with a duty cycle of 10% promoted the contraction of collagen gel. Wang et al14), who adopted 24-h ES, reported that duty cycle of approximately 8% (pulse width of 10 ms within 1200 ms) with 50 mV/mm promotes collagen gel contraction; however, duty cycle of 50% (pulse width of 300 ms within 600 ms) with 50 mV/mm did not promote this. This is similar to the result of the present study. However, ES with 100 mV/mm promoted collagen gel contraction in both conditions14). This could be due to the difference in intensity. Thus, duty cycle might be involved in the contractile ability of HDFs. ES with a duty cycle of 10% also promoted α-SMA and TGF-β1 expression. As shown previously, α-SMA expression is an indicator of myofibroblast, and collagen gel contractile ability reflects wound contraction, which is necessary for wound closure. ES with a duty cycle of 10% promoted both α-SMA expression and collagen gel contraction. Thus, fibroblasts might differentiate into myofibroblasts with ES. Moreover, TGF-β1 is a key factor in wound contraction that functions by promoting α-SMA expression, differentiation, and the secretion of collagen12). The present study results suggest that ES might induce differentiation of myofibroblasts and wound contraction by autocrine secretion of TGF-β1and α-SMA in HDFs. However, this study did not assess temporal changes in the expressions of mRNA and protein. α-SMA expression could be higher within 24 h after ES initiation, but the relationship between the expression of α-SMA and the duration of ES is unclear in this study. Yoshikawa et al10) showed that 1-h ES with a duty cycle of 50% promotes cell proliferation. Thus, the pro-fibrotic effects of ES on HDFs might be influenced by stimulation duration. Moreover, it is not clear which cell sensor received ES to promote TGF-β1 and α-SMA expression. TGF-β1-mediated signals are enhanced via integrin β1, which is a cell surface receptor24) and contributes to some outside-in signals that regulate certain cellular functions including differentiation25).

This study revealed that myofibroblast differentiation mediated by ES is influenced by duty cycle and that ES with a duty cycle of 10% promotes cell differentiation, whereas a duty cycle >50% induces cell death. Some studies revealed that difference in intensity or frequency affects cell behavior such as migration, proliferation, and the secretion of some cytokines9,13,14,26). Thus, the effects of ES on cell behavior differed with these parameters. In this study, we examined the effects of differences in the duty cycle on HDF differentiation into myofibroblasts and viability. The results indicated that a duty cycle of 10% promotes wound contraction with myofibroblast differentiation. However, a higher duty cycle and long-term stimulation might have adverse effects. Therefore, it is necessary to set stimulation parameters, such as duty cycle, duration, intensity, frequency, and polarity, according to the purpose of ES for pressure injuries in clinical trials.

Limitation

The reason for the decrease in cell viability in the 50% and 90% groups was unclear. A 24-h ES with a duty cycle of 10% promoted myofibroblast differentiation, but this study used only 24-h stimulation and did not reveal the relationship between ES duration and fibrotic effects. Therefore, 10% might not be the optimum duty cycle for HDFs with short duration. TGF-β1 mRNA expression was increased by ES with a duty cycle of 10%, but other cytokines and the signaling pathway that accelerates the secretion of fibrotic factors induced by ES are also unclear. Therefore, further study is needed to investigate the effect of duration on differentiation and which pathway is influenced by ES.

Conclusion

This study suggests that the duty cycle influences myofibroblast differentiation and HDF viability and shows that 10% is the effective duty cycle of monophasic-pulsed micro-currents for granulation tissue formation, which induces pressure injury healing.

Conflict of Interest

The authors declare no conflicts of interest.

Acknowledgments

We would like to thank Dr. Matsuo for lending us measuring equipment.

References

  • 1.Ohura T, Sanada H, et al.: Clinical activity-based cost effectiveness of traditional versus modern wound management in patients with pressure ulcers. Wounds. 2004; 16: 157-163. [Google Scholar]
  • 2.Liu L, Moody J, et al.: A quantitative, pooled analysis and systematic review of controlled trials on the impact of electrical stimulation settings and placement on pressure ulcer healing rates in persons with spinal cord injuries. Ostomy Wound Manage. 2016; 62: 16-34. [PubMed] [Google Scholar]
  • 3.European Pressure Ulcer Advisory Panel, National Pressure Injury Advisory Panel and Pan Pacific Pressure Injury Alliance: Prevention and Treatment of Pressure Ulcers/Injuries: Quick Reference Guide 2019. 2019. [cited 2020 Mar. 17]; Available from: http://www.internationalguideline.com/
  • 4.Kloth LC: Electrical stimulation technologies for wound healing. Adv Wound Care (New Rochelle). 2014; 3: 81-90. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Lala D, Spaulding SJ, et al.: Electrical stimulation therapy for the treatment of pressure ulcers in individuals with spinal cord injury: a systematic review and meta-analysis. Int Wound J. 2016; 13: 1214-1226. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6.Khouri C, Kotzki S, et al.: Hierarchical evaluation of electrical stimulation protocols for chronic wound healing: An effect size meta-analysis. Wound Repair Regen. 2017; 25: 883-891. [DOI] [PubMed] [Google Scholar]
  • 7.Grinnell F: Fibroblasts, Myofibroblasts, and Wound Contraction. J Cell Biol. 1994; 124: 401-404. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 8.Zhao R, Liang H, et al.: Inflammation in Chronic Wounds. Int J Mol Sci. 2016 Dec 11; 17: 2085. 10.3390/ijms17122085. PubMed PMID 27973441; PubMed Central PMCID PMC5187885. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 9.Uemura M, Maeshige N, et al.: Monophasic pulsed 200-μA current promotes galvanotaxis with polarization of actin filament and integrin α2β1 in human dermal fibroblasts. Eplasty. 2016 Jan 19; 16: e6. eCollection 2016. PubMed PMID: 26819649; PubMed Central PMCID: PMC4724796. [PMC free article] [PubMed] [Google Scholar]
  • 10.Yoshikawa Y, Sugimoto M, et al.: Monophasic pulsed microcurrent of 1-8 Hz increases the number of human dermal fibroblasts. progress in rehabilitation medicine. 2016; 1: 2016005. https//.org/10.2490/prm.20160005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Deguchi T, Iwakawa N, et al.: The promotional effect of electrotherapy on pressure ulcer healing. Japanese Journal of Electrophysical Agents. 2015; 22: 44-47. [Google Scholar]
  • 12.Desmoulière A, Geinoz A, et al.: Transforming growth factor-beta 1 induces alpha-smooth muscle actin expression in granulation tissue myofibroblasts and in quiescent and growing cultured fibroblasts. J Cell Biol. 1993; 122: 103-111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Rouabhia M, Park H, et al.: Electrical stimulation promotes wound healing by enhancing dermal fibroblast activity and promoting myofibroblast transdifferentiation. PLoS One. 2013; 19: e71660. 13.1371/journal.pone.0071660. PubMed PMID 23990967; PubMed Central PMCID PMC3747189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Wang Y, Rouabhia M, et al.: Pulsed electrical stimulation modulates fibroblasts' behaviour through the Smad signalling pathway. J Tissue Eng Regen Med. 2017; 11: 1110-1121. [DOI] [PubMed] [Google Scholar]
  • 15.Zhang J, Ren R, et al.: A Small Physiological Electric Field Mediated Responses of Extravillous Trophoblasts Derived from HTR8/SVneo Cells: Involvement of Activation of Focal Adhesion Kinase Signaling. PLoS One. 2014; 18: e92252. 10.1371/journal.pone.0092252. PubMed PMID 24643246; PubMed Central PMCID PMC3958492. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16.Guo A, Song B, et al.: Effects of Physiological Electric Fields on Migration of Human Dermal Fibroblasts. J Invest Dermatol. 2010; 130: 2320-2327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 17.Antov Y, Barbul A, et al.: Electroendocytosis: stimulation of adsorptive and fluid-phase uptake by pulsed low electric fields. Exp Cell Res. 2004; 297: 348-362. [DOI] [PubMed] [Google Scholar]
  • 18.Mamalis A, Garcha M, et al.: Light emitting diode-generated blue light modulates fibrosis characteristics: fibroblast proliferation, migration speed, and reactive oxygen species generation. Lasers Surg Med. 2015; 47: 210-215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 19.Matsuki N, Takeda M, et al.: Activation of caspases and apoptosis in response to low-voltage electric pulses. Oncol Rep. 2010; 23: 1425-1433. [DOI] [PubMed] [Google Scholar]
  • 20.Love MR, Palee S, et al.: Effects of electrical stimulation on cell proliferation and apoptosis. J Cell Physiol. 2018; 233: 1860-1876. [DOI] [PubMed] [Google Scholar]
  • 21.Tang LL, Sun CX, et al.: Steep pulsed electric fields modulate cell apoptosis through the change of intracellular calcium concentration. Colloids Surf B Biointerfaces. 2007; 57: 209-214. [DOI] [PubMed] [Google Scholar]
  • 22.Schneider LA, Korber A, et al.: Influence of pH on wound-healing: a new perspective for wound-therapy? Arch Dermatol Res. 2007; 298: 413-420. [DOI] [PubMed] [Google Scholar]
  • 23.Sharpe JR, Booth S, et al.: Progression of Wound pH During the Course of Healing in Burns. J Burn Care Res. 2013; 34: e201-e208. 10.1097/BCR.0b013e31825d5569. PMID 23128128. [DOI] [PubMed] [Google Scholar]
  • 24.Li Y, Li BS, et al.: Effects of integrin β1 in the treatment of stress urinary incontinence by electrical stimulation. Mol Med Rep. 2019; 19: 4727-4734. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25.Huang C, Fu X, et al.: The involvement of integrin β1 signaling in the migration and myofibroblastic differentiation of skin fibroblasts on anisotropic collagen-containing nanofibers. Biomaterials. 2012; 33: 1791-1800. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26.Urabe H, Akimoto R, et al.: Effects of pulsed electrical stimulation on growth factor gene expression and proliferation in human dermal fibroblasts. Mol Cell Biochem. 2020; Sep 23. 10.1007/s11010-020-03912-6. PMID 32968926. Online ahead of print. [DOI] [PubMed] [Google Scholar]

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