Particulate matter (PM) is known to increase the risk of diseases in major organs and PM may also slow the complex process of wound healing. We investigated the effect of PM on wound healing
Twelve 10-week-old Sprague-Dawley male rats were punctured with three 8 mm-diameter holes each in the skin on their backs. The holes were divided into three groups according to treatment: PM 50 µg/mL (group A), PM 100 µg/mL (group B) and saline (control). For histologic analysis, three rats were sacrificed on each of the 1st, 3rd, 7th, and 28th days, and a total of 36 samples were collected. Inflammatory cell infiltration and neovascularization were evaluated for comparison among groups.
Inflammation increased in groups A and B on day 1, but no significance was observed. On the 28th day, increased inflammation was observed in groups A and B compared to the control group, with significant difference. Angiogenesis increased in groups A and B compared to the control group on day 1, but no significance was observed. On the 3rd day, there was a decrease in group B, with statistical significance. On day 28, it was observed that angiogenesis significantly increased in group B compared to the control group.
Inflammation increased in the PM groups compared with the control group and angiogenesis decreased, then slowly increased in the PM groups compared with the control group. Our findings suggest that PM increases inflammation and delays angiogenesis in the wound healing process.
Particulate matter (PM) in the air is a complex mixture of particles and chemicals, and the particle size, concentration, and chemical content of PM vary widely. It is composed of a wide range of chemical elements including nitrates, sulfates, hydrocarbons, organic compounds, biological chemicals, and metals [
There has been continuous epidemiological research showing that airborne PM produces a variety of negative health outcomes including respiratory and cardiovascular disorders [
Wound healing is a normal biological process of skin tissues in the body [
While pollutants such as tobacco smoke and air pollution have been intensively studied for their harmful effects on human skin, the effects of PM on wound healing have not been studied. Therefore, we hypothesized that PM would inhibit wound healing and investigated the effect of PM in
Fine PM NIST 2786 (PM4.0) was purchased from Sigma-Aldrich, Inc. (St. Louis, MO, USA). A PM4 stock solution diluted with 0.5 mL normal saline was prepared, and was sonicated to avoid agglomeration of the suspended PM4 particles.
Twelve 10-week-old male Sprague-Dawley rats weighing 330 g were used in this study. After a 1-week acclimatization period, the rats were anesthetized with sevoflurane for inhalation of anesthesia. The fur was shaved using an electric shaver and disinfected with 10% povidone-iodine and 70% alcohol solutions. The animal experiments were approved by the Institutional Animal Care and Use Committee of Wonkwang University (IACUC No. WKU21-48).
A biopsy punch (Kai Medical, Chiyoda, Japan) was used to create three 8 mm physical holes in the skin on the back of the rats. Normal saline (control), PM 50 µg/mL (group A), and PM 100 µg/mL (group B) solutions were applied to each of the three holes of the rats. The wounds were then covered with a transparent film dressing (Tegaderm; 3M, Saint Paul, MN, USA) (
To observe histological changes, three rats were euthanized on each of days 1, 3, 7, and 28. Tissue samples were collected for each hole, and a total of 36 samples were subjected to analysis. The tissues were collected to include the normal tissue surrounding the defects. They were fixed with 10% formalin solution, embedded in paraffin, cut into slices, and stained with hematoxylin and eosin (H&E) and Masson’s trichrome stains. The stained slides were scanned on a digital slide scanning device, and the images were visualized using CaseViewer software version 2.4 (3DHISTECH, Budapest, Hungary). A descriptive analytical study of the inflammatory infiltrate and neovascularization was performed and a comparative analysis among groups was carried out. The penetration of polymorphonuclear and mononuclear cells was presented as a morphological score of inflammation, and the new vascular formation of the tissues was measured for scoring angiogenesis. The data were obtained using a semi-quantitative scoring system described previously [
The histological sections were examined by two examiners, who were blinded to the group information while performing all the histological analyses in this study, and their average score value was used for the comparative tissue analysis.
Statistical data and graphs were generated via performing analyses of variance using StatView software (SAS Institute Inc., Cary, NC, USA). Values were expressed as the mean±standard deviation. The significance of the differences between groups was determined by using Fisher’s least significant difference post-hoc analyses.
At 4 weeks after wound generation, all three groups were in a completely healed state, and showed no complications such as infection.
Inflammation increased in groups A (3.00±0.00) and B (2.83±0.41) compared to that in the control group (2.50±0.93), and angiogenesis increased in group A (2.00±1.16) compared to that in the control group (1.00±0.00) and group B (1.00±0.00), but the difference was not significant on day 1. Three days after injecting PM, angiogenesis decreased in group B (2.14±1.07) compared to that in the control group (2.88±0.35) and group A (2.88±0.35), and the difference was significant (P=0.029). Seven days after injecting PM, angiogenesis decreased in group A (2.50±0.76) compared to that in the control group (3.00±0.00), but the difference was not significant (P=0.105). Lastly, after 28 days of injecting PM, inflammation increased in groups A (2.00 ±0.00) and B (1.50±0.55) compared to that in the control group (1.00± 0.00), and the difference was significant (P<0.001, P=0.027). Angiogenesis also tended to increase in groups A (0.50±0.58) and B (1.83±0.98) compared to that in the control group (0.50±0.55), and the difference was significant only in group B (P=0.004) (
H&E staining (
Air pollution, which is widespread globally, has an increasingly harmful effect on human health [
There are four main mechanisms through which PM adversely affects human health: oxidative stress, genotoxicity, cell death, and inflammation [
Wound healing is a normal but complex process, involving the phases of hemostasis, inflammation, proliferation, and remodeling. Each phase must proceed reliably and with consistency, and if abnormal progression occurs at any one phase, wound healing may be prolonged or the wound may become chronic. A number of different types of cells are involved in these phases [
In consideration of the aforementioned, we conducted an experiment assuming that PM could affect the wound healing process, especially in the inflammation phase. We performed this study using an
This increased inflammation is believed to be caused by prolonged elevation of pro-inflammatory cytokines such as interleukin-1 and tumor necrosis factor-α.
This study had several limitations as a preliminary study. First, because the physiological structures of rats are different from those of humans, it is questionable whether the study results could be generalized to humans. Two- and three-dimensional
In conclusion, while previous studies have reported various harmful effects of PM on human health, this study is meaningful in that it is the first study on the effect of PM on wound healing. The present study revealed that PM affects wound healing by increasing inflammation and delaying angiogenesis. The results can support further investigation on the effect of PM on wound healing.
This work was supported by Wonkwang University in 2021. Young Cheon Na is an editorial board member of the journal but was not involved in the peer reviewer selection, evaluation, or decision process of this article. No other potential conflicts of interest relevant to this article were reported.
Wound creation on back of rat. (A) Wounds were created using a biopsy punch to make three 8 mm-diameter holes. (B) The wounds were covered with a transparent film dressing after introducing saline (control), PM 50 µg/mL (group A), and PM 100 µg/mL (group B) in each punch hole.
Histologic findings. The particulate matter-contaminated groups A and B both displayed increased inflammatory scores and delayed angiogenesis compared to the control group (H&E, ×20).
Comparison of inflammation progression using the semi-quantitative scoring system
Inflammation | Control group (saline) | Group A (PM 50 µg/mL) | Group B (PM 100 µg/mL) | P-value (control vs. group A) | P-value (control vs. group B) | P-value (group A vs. group B) |
---|---|---|---|---|---|---|
Day 1 | 2.50±0.93 | 3.00±0.00 | 2.83±0.41 | 0.129 | 0.305 | 0.629 |
Day 3 | 3.00±0.00 | 3.00±0.00 | 2.75±0.46 | - | 0.161 | 0.161 |
Day 7 | 3.00±0.00 | 3.00±0.00 | 3.00±0.00 | - | - | - |
Day 28 | 1.00±0.00 | 2.00±0.00 | 1.50±0.55 | <0.001 |
0.027 |
0.045 |
Values are presented as mean±SD. Scoring: 0, absence of inflammation; 1, presence of few inflammatory cells; 2, many inflammatory cells; 3, exaggerated inflammatory cellularity.
PM, particulate matter.
P<0.05 is considered statistically significant.
Comparison of angiogenesis progression using the semi-quantitative scoring system
Angiogenesis | Control group (saline) | Group A (PM 50 µg/mL) | Group B (PM 100 µg/mL) | P-value (control vs. group A) | P-value (control vs. group B) | P-value (group A vs. group B) |
---|---|---|---|---|---|---|
Day 1 | 1.00±0.00 | 2.00±1.16 | 1.00±0.00 | 0.071 | - | 0.109 |
Day 3 | 2.88±0.35 | 2.88±0.35 | 2.14±1.07 | - | 0.029 |
0.029 |
Day 7 | 3.00±0.00 | 2.50±0.76 | 3.00±0.00 | 0.105 | - | 0.105 |
Day 28 | 0.50±0.55 | 0.50±0.58 | 1.83±0.98 | - | 0.004 |
0.008 |
Values are presented as mean±SD. Scoring: 0, normal vascularization; 1, discrete vascular formation; 2, moderate vascular formation; 3, high vascular formation.
PM, particulate matter.
P<0.05 is considered statistically significant.