5 International and 24 th National Conference on ...

5th International and 24th National

Conference on Environmental Health

December 14-16, 2021

Kashan, Iran

Seasonally varied Proinflammatory effects of urban particulate matter-

induced(PM2.5) on human lung epithelial cells (A549) in vitro at the most

industrial cities of Iran

Abstract

In ambient air, particulate matter, especially fine particles (PM2.5), can induce irreversible impacts on human

health. In this research, cells were individually exposed to three concentrations of PM2.5 (25, 50, and 100

μg/mL) and three times (12, 24, and 48h). We assessed in winter (wPM2.5) and summer (sPM2.5) PM2.5

sample and exposed cells of A549 to concentrations of PM2.5 samples to measure its cell viability and release

of pro-inflammatory cytokines. We assessed the liberation of pro-inflammatory cytokines (interleukin-6 and

interleukin-8) by the ELISA method and cell viability by MTT assay [3- (4, 5-dimethylthiazol-2-yl) -2, 5-

diphenyltetrazolium bromide] . The toxicological outcomes of this research determined that raising the

concentration of PM2.5 particulates and contact time with it decreases cell viability and raises inflammatory

answers. Seasonal cytotoxicity of PM2.5 particles in the summer season compared to the winter season was

lower. The lowest percent viability was observed at two days of exposure and 100 μg/mL exposure in the

winter sample. Moreover, PM2.5 particles were important in the value of IL-8 and IL-6 . The medium release

level of IL-6 and IL-8 in the winter and the large exposure time and concentrations (48h–100 μg/mL) was

much higher than in the summer . These values were double as high for winter PM2.5 samples as for summer

samples. The compounds in PM2.5 at varied seasons can cause some biological impressions. The samples’

chemical components in two seasons presented that the PMs were various in chemical properties. In general,

heavy metals and polycyclic aromatic hydrocarbons were more in the winter samples . However, the samples

of wPM2.5 had a lower mass quota of metals such as aluminum, iron, copper, zinc, and magnesium.

Concentrations of chromium, cadmium, arsenic, mercury, nickel, and lead were more significant in the sample

of wPM2.5.

Keywords: Pro-inflammatory cytokine. Lung epithelial cells (A549). In vitro.

1- Introduction

Inflammation of the airways is one of the short-term effects on humans' lungs due to airborne PM [1].

Inflammation when it happens with severity or for a long time at the airway's bottom, cause breathing system

illnesses like chronic obstructive pulmonary disease (COPD) and asthmatic [2, 3]. This process kills pro-

inflammatory intermediate using macrophages of alveolar (AMs) and epithelial lung cells (ECs). Cytokines are

signaling proteins involved in regulating physiological factors that will function as a pro-inflammatory or anti-

inflammatory mediator. Chemokines are secreted only by cell damage or by multiple stimuli to absorb and

activate immune cells [4]. Two known proinflammatory cytokines are interleukin 6 (IL-6) and 8 (IL-8). These

proteins have special functions. Since the first defense response is IL-8, it is released rapidly after stimulation;

it immediately triggers the body's immune cells' part, especially some white blood cells at the site of infection.

IL-8 continues inactive form after being released, therefore its effect is fast and long-enduring [5]. IL-6 is a




Kashan, Iran

cytokine which is associated with inflammatory and infection responses and the regulation of metabolic,

regenerative, and neurological behaviors [6].

Particulate matter is one of the leading air pollutants, which is produced from natural and human-made sources

[7-9]. Exclusively particles with an aerodynamic diameter of less than 2.5 microns (PM2.5) in mall-scale and

powerful penetrative ability is damaging to human safety [10, 11], which they can be an increased risk of lung

cancer, respiratory disease, heart disease and brain disease [12-14]. Toxicity research by some cellular

mechanisms has shown the destructive effects of PM2.5 particles (such as cytotoxicity and inflammatory

products of cytokines) [15, 16]. The compounds bonded with PM2.5 particles has intricate compositions from

various provenance; therefore, theycan have different effects on human health [17-19]. Aerosol levels may vary

in different seasons[20-22]. Since the concentration and compositions bonded with PM2.5 determine health

hazards by various aerosol sources, particulate matter's toxic effects were critical to air pollution control and

environmental management [23, 24]. Karaj is the center for connecting major cities in the north and center of

Iran. Problems of ecological, including air pollution, are caused by multiple daily trips (about 70,000 vehicles

per day) [25]. In this study, we investigated the toxicity and combination of PM2.5 in two seasons (summer and

winter) at traffic sites in Karaj for accurate assessment the health dangers of PM2.5 in various concentrations.

We measured the toxicological influences of PM2.5 on epithelial lung cells (A549) and analyzed their

association with PM2.5 constituents (metals and PAHs).

2- Material and Methods

Sampling and extraction PM2.5

Particulates matter with a diameter of less than 2.5 microns was collected in the high-traffic stations of Karaj

in two seasons of the year 2018-19 (winter and summer) Fig.1. high-volume peripheral pump (Leland Legacy

(SKC)) with 3 L/min flow rate for 24 h which contains polytetrafluoroethylene (PTFE) filter with the pore size

of 1 micron and diameter of 37 mm on the health centers building's roof was located; and the height from the

ground for sampling was about 3 meters. Sampling was done from each station for 24 hours (8.00 am until 8.00

am days later). Sampling time was in summer from 1 to 27 July 2019) and in winter from 1 march to 9 february

2019. The filters were placed in a desiccator for 48 hours (before and after sampling) and weighed to determine

the particle matter weight with a balance (0.0001 mg accuracy, Mettler Toledo AB204-N). We saved the filters

gathered from the two seasons in the freezer at -20 °C away from light (for chemical and biological analysis).

Particle extraction from the filters was more than 75% of PM2.5 main weight on filters. For biological research,

filters were immersed in 5 ml of Mili-Q water in a sterile falkon, and three rounds of 20 min were oultrasound

bath applied for each filter group (wPM2.5 and sPM2.5). Water containing suspended particles was dried by

lyophilization, then we stored the extracted PMs at -80 °C [26].

Chemical characterization

Half filters are used to analyze for PAHs, a quarter filter to analyze for heavy metals , and a quarter filter for

biological analysis. In total, we gathered 16 samples of PM2.5 in two seasons (winter and summer) for chemical

and biological analysis [27, 28].




Kashan, Iran

Fig.1. Geographical location and sampling points of the study area

Measurement of PAH compounds

PAHs were extracted by placing filters in acetone and dichloromethane (3: 1) on wet ultrasound for one-third

of an hour. The solution was filtered to eliminate insoluble fraction, and 16 PAHs analyzed and measured by

using gas chromatography-mass spectrometry (Agilent model 7890B, Agilent-MS 5975B, Model [H1] EI ) by

an Agilent Capillary HP-5MS Column (30 m, 0.25 mm, 0.25 um)[29]. PAHs compounds included the

following: Naphthalene (NaP), Acenaphthylene (Acy), Acenaphthene (Ace), Ffluorene (Flu), Phenanthrene

(Phen), Fluoranthene (Flrt), Anthracene (Anth), Pyrene (Pyr), Chrysene (Chr), Benzo[a] anthracene (BaA),

Benzo[b]fluoranthene (BbF), Benzo[k]fluoranthene (BkF), Benzo[a]pyrene (BaP), Dibenzo[a,h]anthracene

(DahA), Indeno[1,2,3-cd]pyrene (IcdP), Benzo[ghi]perylene (BghiP).

Measurements of heavy metals

Heavy metals bounded with PM2.5 were analyzed using the ICP-OES tool. In total, we examined 11 heavy

metals (Pb, Al, Zn, Ar, Ag, Cr, Mg, Fe, Ni, Cu, Cd). The specifications of the ICP-OES device were as follows:

inductively coupled plasma – optical emission spectrometry (ICP – OES) on a perkin elmer instrument model

optima 8000(Sheltin, USA) with axial torch view UV- sensitive, dual backside –illuminated charge- coupled

device( CCD) array detector was applied for determination of the target elements. The optimal instrumental

conditions and the emission lines, which were selected for determination of the analyte via ICP- OES, were as

follows: RF generator power:1.5Kw, frequency of RF generator: 40 MHz, plasma gas flow rate: 8 L/min, pump

rate:1Ml/min.




Kashan, Iran

Cell culture and treatment

In this research, human lung cells (A549) were used for cytotoxicity. They proliferated in DMEM F12 culture

medium with 1% penicillin-streptomycin antibiotic and 10% bovine fetal serum. When cell growth density in

the 5% CO2 atmosphere reached 80-90% at 37 ° C, 0.25% digestion was performed with trypsin. The cells

were cultured several times, and after passing through the primary cells and reaching the exponential growth

period, they were used to determine toxicity. A549 cells were placed in 104 cells per well in 96 microplates.

After 24 hours of incubation, three concentrations of PM2.5 (25, 50, and 100 μg/ml) were added to the

microplate separately from summer and winter [30, 31].

Cell viability

Metabolic activity of cells was done by MTT assay [3- (4, 5-dimethylthiazol-2-yl) -2, 5-diphenyltetrazolium

bromide][32]. MTT method used to estimate the viability of A549 cells after three time periods (12, 24, and 48

hours) and three concentrations in two seasons (25, 50, and 100μg/ml exposed to summer PM2.5 and winter

PM2.5). MTT by-product was dissolved in 1 ml DMSO via cellular metabolism (crystals of insoluble formazan).

The absorption of each sample was calculated in an ELISA reader with a wavelength of 570 nm. The resulting

data were compared and reported with the control group(which is 100% viable).

Cytokine release

After three-time exposed to PM2.5(12 h, 1 and 2 days), cell culture media were gathered and centrifuged for 10

min at 250 gr to eliminate cell residue and remaining PM. The final supernatants were saved at -70°C till use.

We used human ELISA (BD biosciences pharmingen) kits to measure interleukin levels 6 and 8 in cellular

procedures. The concentration of cytokines is calculated in pg/ml.

Statistical analyses

Results analysis was performed through the SPSS (V 23.0). Variations were estimated to be statistically

meaningful at P-value < 0.05. Moreover, prism 8.0 was employed for describing the graphs detailed as means

± standard error (SE). The data were shown as mean values and standard deviations. We analyzed data from

control wells and winter and summer PMs for exposure duration (12, 24, and 48 h). Additionally, wilcoxon

signed-rank test was utilized to distinguish the variations within PM of winter and summer in the value of

cytokines released.

3- Results and Discussion

Concentration of PM2.5 on different seasons

Seasonal variaition concentrations of PM2.5 particulates samples are showed in Fig.2. Seasonal variations

behold with the highest concentrations of PM2.5 during winter. Therefore, the wPM2.5 sample concentration is

more elevated than the sPM2.5 samples (W > S). The extracted PM mass from the filters ranged 811 μg for the

sPM2.5 filters and 1704 μg for wPM2.5 samples based on these data. The average daily concentration of PM2.5

is 2.25 times higher in the winter and 1.08 times higher in the summer than the WHO guidline. (WHO guide

value: 25µg/m3) .The average concentration of PM2.5 particles was 56.4 μg/m3 and 26.9 μg/m3 in winter and

summer, respectively. Development of Karaj city, due to connect large cities in the north and center, has

significant daily traffic, which leads to the appearance of many environmental issues including air pollution




Kashan, Iran

and increasing of particle concentrations [33]. The maximum daily concentration of PM2.5 particles in winter

and summer was 95.8 μg/m3 and 48.6 μg/m3, and the maximum and minimum was 18.5 μg/m3 and 11.2 μg/m3

is respectively. many studies in the countries of the world showed that concentration of PM2.5 particles in cold

season higher than in hot season. The scientists found that the source and composition of PM2.5 in a region vary

in different seasons, and the topographic effect is a long-term effect with minor changes in short-term periods

[34-37]. Therefore, citizens of Karaj face a high concentration of PM2.5 in winter. We found that the daily

concentration of PM2.5 and its fluctuations increase in cold seasons, and the duration of air pollution increases.

Fig.2. Concentrations of PM2.5 particulate in winter and summer seasons

Concentration of heavy metals in different seasons

Changes in heavy metals concentration in the studied seasons are reported in table 1. The average of heavy

metals in two seasons has been shown in fig.3. According to the data, the amount of heavy metals differed

considerably with the seasons changed. table 1 shows the mean and standard deviation of heavy metals in

PM2.5. The measured values indicate that high concentrations of heavy metals (Pb, Hg, As, Cd, Cr, and Ni) in

winter can be related to high levels of PM2.5 in this season, which leads to more aggregation of these metals in

the particulate matter. According to the data obtained for both studied seasons, aluminum and iron had the

highest abundance among other metals adsorbed on PM2.5. The average of aluminum in season of summer

were 479 ng/m3 and 537.1 ng/m3 and winter, and average of iron in season of summer and winter is 266.8

ng/m3, 303.9 ng/m3, respectively.




Kashan, Iran

Table 1. Heavy metals concentration changes in winter and summer seasons

Season

Heavy

Metals Summer Winter

SD Min Max Mean SD Min Max Mean

70.1 52.6 232.9 122.6 71.3 56.6 247.8 149.5 Pb

172.2 117.1 549 275.7 128.2 91.4 448.8 215.5 Zn

16.5 8.8 50.4 28 37.5 9.1 118.6 52.0 Hg

18.3 24.1 66.3 40.8 38.0 15.3 107.8 54.5 As

37.5 62.2 177.1 130 42.9 49.3 182.9 107.0 Mn

54 20.8 169.8 98 70.5 80.1 284.0 141.4 Cd

13.3 12.7 47.4 35.2 38.7 12.4 130.2 66.6 Cr

44.2 31 134.3 77.8 57.9 5.7 157.9 83.2 Ni

240 62.4 692 365.4 113.5 148.9 455.7 270.4 Cu

162.8 366.6 831.1 537.1 122.0 169.3 539.2 303.9 Fe

133.9 328.5 666.4 479 92.0 125.6 426.4 266.8 Al

According to the table 1, the highest concentrations of heavy metals in summer were related to Al and Fe, and

in winter season were related to Pb, Hg, As, Cd, Cr and Ni, respectively. The high levels of Al and Fe compared

to other elements are probably due to these two elements presence in the earth crust. In addition to natural

resources, other sources of Fe and Al pollutants are due to vehicles movement and motorcycles on the roads

[38, 39]. In general, according to the information obtained from both studied seasons, the average concentration

of heavy metals was less in summer than in winter. On the other, a meaningful variation (p <0.05) was observed

for heavy metal concentrations in winter compared to summer. Most heavy metals weigh in this study in winter

is higher than summer season (fig.3). The variation in the amount of adsorbed heavy metals bounded with PM2.5

particles in our study and schilirò's research may be due to differences in the composition of PM2.5 in different

regions and current conditions [40]. The discrepancy in the amount of concentration and kind of metals sorbed

on PM2.5 in our study and schilirò's study may be owing to PM type and condition differences. The discrepancy

in the amount of concentration and kind of heavy metals bounded on PM2.5 in our study and schilirò's study

may be owing to PM type and condition differences. heavy Metals such as Mg, Fe, Cu, Pb, and Zn were mostly

related to traffic emissions, road wear (Mg), and tire wear (Zn) [41] . Other studies [42, 43] showed that Pb, Zn,

Ni, and Cd could be part of vehicle exhaust. Coal burning is also an essential source of Cd, Ar, and Pb [44, 45].

Industrial processes are effective in emitting heavy metals such as Pb,Cu, Cd, Co, Se and Ag [46, 47]. The

value of (Cr, Cd, As, Hg, Ni, and Pb) in the winter was more than the summer concentrations. The highest of

heavy metals amount in summer belonged to Al, Fe, Cu, and the lowest levels also belonged to Hg. The value

of Pb, In winter was over than in the summer. Nevertheless, this concentration was lesser than the guidance

amount (500ng/m3). Increasing the these heavy metals in the cold season due to regular temperature inversion

and atmospheric stability limits the dispersion and accumulates of pollutants in the city [48, 49].




Kashan, Iran

Fig.3. Concentrations of Heavy Metals in winter and summer seasons

Concentration of PAH compounds on different seasons

Table 2 statistically examines the composition of PAHs in two seasons. In the winter, BhgiP, DBahA, and Flrt

have a highest concentrations and IND have a lowest concentrations. In the summer, DBahA, Bap, and Bkf

gave the highest concentrations and IND the lowest. Depending on the carcinogenicity, PAHs compounds can

be divided into two categories (carcinogenic and non-carcinogenic).

Table 2. Concentration of PAHs in winter and summer seasons

Summer Winter PAH Compound

SD Min Max Mean SD Min Max Mean

0.29 0.19 1.08 0.26 0.27 0.12 0.89 0.28 IND

0.86 0.32 3.48 0.78 1.03 0.49 4.61 1.14 BghiP

0.95 0.15 1.45 0.90 0.69 0.51 3.25 1.1 DBahA

0.75 0.16 2.92 0.96 0.61 0.26 2.75 0.98 Bap

0.82 0.20 2.87 1.1 0.87 0.45 3.41 1 Bkf

0.8 0.55 2.63 0.81 0.90 0.23 3.51 1 Bbf

0.70 0.20 1.34 0.82 0.42 0.23 2.97 0.63 BaA

0.28 0.48 1.08 0.59 0.26 0.37 1.26 0.62 Chr

0.54 0.69 2.54 0.99 0.47 0.69 2.34 1 Pyr

0.43 0.64 1.38 0.76 0.80 0.46 3.25 1.2 Flrt

0.31 0.23 1.00 0.30 0.26 0.35 1.24 0.54 Anth

0.49 0.14 1.52 0.42 0.36 0.30 1.43 0.66 Phen

0.37 0.16 1.56 0.56 0.29 0.23 1.08 0.49 Flu

0.23 0.10 0.89 0.31 0.20 0.25 1.53 0.42 Ace

0.32 0.22 1.05 0.41 0.31 0.32 1.24 0.48 Acy

0.49 0.23 1.71 0.47 0.41 0.39 1.34 0.54 NaP

6.24 6.86 PAHs-human carcinogen

4.28 5.41 PAHs-non human carcinogen




Kashan, Iran

Fig.4 shows that the amount of carcinogenic compounds in winter was more than in summer season (about

52%). According to studies, in the summer, PAHs are mostly generating by traffic, but in the winter, owing to

residential heating are produced according to previous studies, traffic areas and industrial cities, especially in

winter and autumn, have higher PAH levels [50]. Changing seasons and sampling traffic conditions have

different effects on PAHs content and concentration and their derivatives. Particulate matter samples from cold

and hot seasons had a significant difference (p <0.05) [51]. However, the air stability and inversion in cold

seasons can be another reason for the increase in PAHs. However, increasing fossil fuel consumption by cars

and traffic can be one of PAHs leading causes. Other possible PAHs sources include industrial waste

incinerators, combustion of wood, power plants, and the iron and steel industries [52-54]. Combustion of PAHs

play a significant role in air pollution (e.g., home heating). Numerous studies have shown that particles with

smaller aerodynamic diameters contain a higher percentage of carcinogenic PAHs [54, 55].

Fig.4. PAHs compounds carcinogenic in winter and summer seasons

Cell toxicity

Significant reductions in cell viability were demonstrated after three exposure to particulate matter (12h, 1 and

2 days) with samples from two seasons Fig 5(a-c). The percentage of viable exposed to the A549 cell line with

wPM2.5 and sPM2.5 with concentrations (25, 50 and 100 μg/ml) is given in table3. These results showed that

A549 cell viability was reduced by particle types (sPM2.5 and wPM2.5) compared to the control group. Also,

cell viability decreased more with wPM2.5 samples than with sPM2.5. In other words, particulate matter can limit

cellular metabolic activity [30]. Particulate matter content causes reactive oxygen species (ROS) generation

and reduces mitochondrial function, ultimately causing cell damage, cell death, and cell toxicity [56]. The study

of Gualtieri et al., is consistent with our research. The decrease in cell viability in particulate matter collected

in winter was more significant than in summer in all concentrations [27]. There was a meaningful differentiation

between the exposure groups (50 and 100 mg/ml) and the control group (ANOVA; p <0.05). No significant

differences were observed between the exposure group of 25 μg/ml and the control group in both study seasons.

Cell viability at 25 μg/ml particulate matter was 88.3% and 5.81% in summer and winter, respectively.




Kashan, Iran

Fig.5 (a-c). cell viability in A549 cell exposed for three-time (12 h, 1 and 2 day) to increasing doses of PM2.5

Table 3. Percentage of viable cells exposure to concentration of PM2.5 (25, 50 and 100 µg/ml) in winter and summer

seasons

Pro-inflammatory response of A549 to PM2.5

Analysis of the inflammatory reply of interleukin 6 and 8 was estimated by the secretion of cytokines in the

culture medium. Interleukin 6 and 8 amount of secretion data are shown in fig 6 (a-d). We found that at three

exposure times (12h, 1 and 2 days) after treatment, interleukin 6 and 8 amount of secretion increased

significantly in the treated groups in comparison with the corresponding group without exposure(p <0.05).

Dose and time-dependency responses: show the dose-related and time-related effects of the PM2.5 of winter and

summer on interleukin 6 and 8 production in A549 cells. IL-8 secretion from contact with particulate matter

(wPM2.5 and sPM2.5) exhibited a dose-dependent increase compared to the control group. However, the response

of 100 μg/ml winter PM2.5 showed the most significant increase in the three exposure times. Also, the reaction

to PM2.5 was lower in summer than in winter (Fig.6(c-d)) .

Percent of Cell Viability

PM Concentration(µg/ml)

Winter Summer SD Mean SD Mean

1.52 87.65 6.38 91.60 25 12h 5.83 82.48 9.13 85.45 50

5.87 73.28 11.6 78.29 100 6.09 85.70 1.77 90.26 25

24h 3.62 81.39 8.38 82.38 50

6.13 71.17 7.23 77.96 100

4.13 81.32 6.36 88.44 25

48h

3.85 75.16 6.84 80.34 50

6.91 64.52 3.48 77.24 100




Kashan, Iran

There were considerable differences between the control group and IL-6 production in the sample of wPM2.5

and sPM2.5 in three treat times. A significant increment in IL-6 release induced by PM2.5 in both seasons was

connected with increased dose and time of PM2.5 exposure. According to the previous studies and our study,

particulate matter collected in winter has a higher potential for the secretion of pro-inflammatory cytokine IL-

6 (Fig.6(a-b)) [30, 57]. There is a meaningful variation within the PM exposure groups (winter and summer)

and the control group. According to the data, IL-8 secretion is higher than IL-6. Nevertheless, in general, the

secretion of both cytokines was significant. The effect of increasing exposure time to PM2.5 of lung cells (A549)

in winter and summer on IL-6 release is displayed . There was a time-dependent increase in cytokines (IL-8

and IL-6) release for both seasons PM2.5, but the response time of exposure 12 h was less than other times

exposure. In our study, at exposure times of 12 h, 1 and 2 days, a significant difference in IL-8 and IL-6

secretion was observed between the treatment of 25 and 100μg/ml of suspended particles for two seasons.

Fig.6(a-b) indicates that the increase in IL-6 release in PM and winter and summer was significant at 12 h of

exposure time compared to 2 days of exposure time at all. This increment in cytokine release was statistically

meaningful in winter and summer. The inflammatory response was evaluated in A549 cells by quantifying two

selected inflammatory cytokines (IL-6, IL-8) after 12 h, 1 and 2 days of exposure. A dose-dependent secretion

of significant interleukin 6 and 8 exposed for both concentrations to sample 50 and 100 µg/ml of PM2.5 was

observed in humane lung cells (A549). The wPM2.5 is a more vigorous inflammatory agent than sPM2.5. It has

beforehand been reported that the secretion of inflammatory intermediaries is significantly correlated with

winter PM2.5 exposure. This difference can be due to particulate different chemical compositions due to change

of seasons [27, 58]. Statistically significant release of IL-8 and IL-6 by human lung cells exposed to two

samples of PM2.5 (sPM2.5 and wPM2.5) against the control group at three exposure times (12 h, 1 and 2 days)

was significant.

In contrast, exposure of human lung cells to two samples of PM2.5 (sPM2.5 and wPM2.5) leading to a statistically

meaningful increase in the release of IL-8 and especially IL-6 against control groups. It is noteworthy that

interleukin 6 and 8 secretion from A549 cells treated to PM2.5 samples were dose-dependent and increased

rapidly at two days exposure time. Differences in cytokine release are observed between PM2.5 samples

collected in the cold season and samples collected in the warm season. Therefore, given these data, it can be

moderately hard to distinguish which chemical agent in the two PM2.5 samples is essentially involved in their

pro-inflammatory influences.




Kashan, Iran

Fig.6 (a-d). The amount of secretion an Interleukin 6 and 8

4- Conclusion

This research illuminated that the concentration of PM2.5, PAHs, and heavy metals was more in winter season.

Toxicity and inflammation of PM2.5 extract were more in winter than in summer season. Winter samples had

a more significant effect on IL-8 and IL-6 secretion than summer samples. Summer particulate matter samples

did less damage to the inflammatory properties of the cells. Inflammatory responses (IL-6 and IL-8) can be

caused by exposure to metals, endotoxins, and some carbonaceous varieties. PAHs adsorbed on the particulate

carbon perform an essential role in inflammation. In new research has shown that PAHs related to particles

carbon part can cause cell damage. This damage leads to cytotoxicity and pro-inflammatory response. The

proper functions of inflammatory reactions are necessary for sustaining tissue and limb homeostasis. Here we

studied the cell survival and inflammatory manners induced in Karaj city of wPM2.5 and sPM2.5 in lung epithelial

A549 cells by metabolic activity of cells and investigating two main interleukins 6 and 8. Both samples increase

interleukin secretion at all PM2.5 concentrations. However, wPM2.5 causes less viability and more inflammatory

responses in the studied cells than sPM2.5. It is necessary to note that various PM inflammatory potency is

highly variable and certainly much depending on the particular combination of chemicals adsorbed. Thus, the

various chemical compounds of PM2.5 urban air are more likely to explain different results. Particle composition

depends not only on origin and season but also on sampling and extraction methods. An increment in PM

amount in the air is usually seen in the cold season. It was because of the wind direction; PM entered the city

(northwest) in winter. The extracts toxicity can also be ascribed to the diversity in collection terms and seasons




Kashan, Iran

(winter more numerous than summer). The heavy metals and PAHs load in the aqueous extract can explain

most of the toxicity found in PMs. PM2.5 has been shown to produce inflammatory cytokines response in lung

epithelial cells. Besides, heavy metals in PM2.5 and residual oil fly ash can cause cytokine secretion.

Furthermore, research inhaling high doses of urban PM2.5 displayed lung action disorder. Other investigations

have connected ambient PM2.5 to meaningful cardiopulmonary changes, blood parameters, and raised blood

pressure. In conclusion, here, we display that wPM2.5 exposure can create inflammatory responses and decrease

cell viability.

5- Acknowledgment

The authors gratefully acknowledge the financial support of the Research Center for Environmental Health

Technology, Iran University of Medical Sciences, Tehran, Iran.

6- References:

[1] Schwartz C, Bølling AK, Carlsten C. Controlled human exposures to wood smoke: a synthesis of the evidence. Particle and

fibre toxicology. 2020;17(1):1-17.

[2] Eapen MS, Myers S, Walters EH, Sohal SS. Airway inflammation in chronic obstructive pulmonary disease (COPD): a true

paradox. Expert Rev Respir Med. 2017;11(10):827-39.

[3] Fu H, Liu X, Li W, Zu Y, Zhou F, Shou Q, et al. PM2.5 Exposure Induces Inflammatory Response in Macrophages via the

TLR4/COX-2/NF-kappaB Pathway. Inflammation. 2020;43(5):1948-58.

[4] Ramgolam K, Chevaillier S, Marano F, Baeza-Squiban A, Martinon L. Proinflammatory effect of fine and ultrafine

particulate matter using size-resolved urban aerosols from Paris. Chemosphere. 2008;72(9):1340-6.

[5] Kocbach A, Herseth JI, Låg M, Refsnes M, Schwarze PE. Particles from wood smoke and traffic induce differential pro-

inflammatory response patterns in co-cultures. Toxicology and applied pharmacology. 2008;232(2):317-26.

[6] Longhin E, Holme JA, Gualtieri M, Camatini M, Ovrevik J. Milan winter fine particulate matter (wPM2.5) induces IL-6 and

IL-8 synthesis in human bronchial BEAS-2B cells, but specifically impairs IL-8 release. Toxicol In Vitro. 2018;52:365-73.

[7] Madureira J, Paciência I, Rufo J, Severo M, Ramos E, Barros H, et al. Source apportionment of CO2, PM10 and VOCs levels

and health risk assessment in naturally ventilated primary schools in Porto, Portugal. Building and Environment. 2016;96:198-205.

[8] Hajizadeh Y, Jafari N, Fanaei F, Ghanbari R, Mohammadi A, Behnami A, et al. Spatial patterns and temporal variations of

traffic-related air pollutants and estimating its health effects in Isfahan city, Iran. Journal of Environmental Health Science and

Engineering. 2021:1-11.

[9] Farrokhzadeh H, Jafari N, Sadeghi M, Alipour MT, Amin MM, Abdolahnejad A. Estimation of spatial distribution of PM10,

lead, and radon concentrations in Sepahanshahr, Iran using Geographic Information System (GIS). Journal of Mazandaran University

of Medical Sciences. 2018;27(159):84-96.

[10] Jordanova D, Jordanova N, Lanos P, Petrov P, Tsacheva T. Magnetism of outdoor and indoor settled dust and its utilization

as a tool for revealing the effect of elevated particulate air pollution on cardiovascular mortality. Geochemistry, Geophysics,

Geosystems. 2012;13(8).

[11] Hajizadeh Y, Mokhtari M, Faraji M, Abdolahnejad A, Mohammadi A. Biomonitoring of airborne metals using tree leaves:

Protocol for biomonitor selection and spatial trend. MethodsX. 2019;6:1694-700.

[12] Hadei M, Hopke PK, Nazari SSH, Yarahmadi M, Shahsavani A, Alipour MR. Estimation of mortality and hospital

admissions attributed to criteria air pollutants in Tehran metropolis, Iran (2013-2016). Aerosol and air quality research.

2017;17(10):2474-81.

[13] Dabass A, Talbott EO, Rager JR, Marsh GM, Venkat A, Holguin F, et al. Systemic inflammatory markers associated with

cardiovascular disease and acute and chronic exposure to fine particulate matter air pollution (PM2. 5) among US NHANES adults

with metabolic syndrome. Environmental research. 2018;161:485-91.

[14] Hajizadeh Y, Jafari N, Mohammadi A, Momtaz SM, Fanaei F, Abdolahnejad A. Concentrations and mortality due to short-

and long-term exposure to PM 2.5 in a megacity of Iran (2014–2019). Environmental Science and Pollution Research.

2020;27(30):38004-14.

[15] Luo X, Zhao Z, Xie J, Luo J, Chen Y, Li H, et al. Pulmonary bioaccessibility of trace metals in PM2. 5 from different

megacities simulated by lung fluid extraction and DGT method. Chemosphere. 2019;218:915-21.




Kashan, Iran

[16] Chen Q, Luo X-S, Chen Y, Zhao Z, Hong Y, Pang Y, et al. Seasonally varied cytotoxicity of organic components in PM2. 5

from urban and industrial areas of a Chinese megacity. Chemosphere. 2019;230:424-31.

[17] Liu Q, Baumgartner J, Zhang Y, Schauer JJ. Source apportionment of Beijing air pollution during a severe winter haze event

and associated pro-inflammatory responses in lung epithelial cells. Atmospheric Environment. 2016;126:28-35.

[18] Kermani M, Jonidi Jafari A, Gholami M, Taghizadeh F, Masroor K, Abdolahnejad A, et al. Characterisation of PM2. 5–

bound PAHs in outdoor air of Karaj megacity: the effect of meteorological factors. International Journal of Environmental Analytical

Chemistry. 2021:1-19.

[19] Heydari G, Taghizdeh F, Fazlzadeh M, Jafari AJ, Asadgol Z, Mehrizi EA, et al. Levels and health risk assessments of

particulate matters (PM 2.5 and PM 10) in indoor/outdoor air of waterpipe cafés in Tehran, Iran. Environmental Science and Pollution

Research. 2019;26(7):7205-15.

[20] Huebert BJ, Ming‐Xing W, WEI‐XIU L. Atmospheric nitrate, sulfate, ammonium and calcium concentrations in China.

Tellus B. 1988;40(4):260-9.

[21] Wang F, Wang J, Han M, Jia C, Zhou Y. Heavy metal characteristics and health risk assessment of PM2. 5 in students’

dormitories in a university in Nanjing, China. Building and Environment. 2019;160:106206.

[22] Kermani M, Jafari AJ, Gholami M, Arfaeinia H, Shahsavani A, Fanaei F. Characterization, possible sources and health risk

assessment of PM2. 5-bound Heavy Metals in the most industrial city of Iran. Journal of Environmental Health Science and

Engineering. 2021:1-13.

[23] Guo H, Gu X, Ma G, Shi S, Wang W, Zuo X, et al. Spatial and temporal variations of air quality and six air pollutants in

China during 2015–2017. Scientific reports. 2019;9(1):1-11.

[24] Kermani M, Arfaeinia H, Masroor K, Abdolahnejad A, Fanaei F, Shahsavani A, et al. Health impacts and burden of disease

attributed to long-term exposure to atmospheric PM10/PM2. 5 in Karaj, Iran: effect of meteorological factors. International Journal of

Environmental Analytical Chemistry. 2020:1-17.

[25] Vahidi MH, Fanaei F, Kermani M. Long-term health impact assessment of PM2. 5 and PM10: Karaj, Iran. International

Journal of Environmental Health Engineering. 2020;9(1):8.

[26] Wang J, Zhang WJ, Xiong W, Lu WH, Zheng HY, Zhou X, et al. PM2.5 stimulated the release of cytokines from BEAS-2B

cells through activation of IKK/NF-kappaB pathway. Hum Exp Toxicol. 2019;38(3):311-20.

[27] Gualtieri M, Øvrevik J, Holme JA, Perrone MG, Bolzacchini E, Schwarze PE, et al. Differences in cytotoxicity versus pro-

inflammatory potency of different PM fractions in human epithelial lung cells. Toxicology in vitro. 2010;24(1):29-39.

[28] Karbasdehi VN, Dobaradaran S, Nabipour I, Arfaeinia H, Mirahmadi R, Keshtkar M. Data on metal contents (As, Ag, Sr,

Sn, Sb, and Mo) in sediments and shells of Trachycardium lacunosum in the northern part of the Persian Gulf. Data in brief. 2016;8:966-

71.

[29] Hoseini M, Yunesian M, Nabizadeh R, Yaghmaeian K, Ahmadkhaniha R, Rastkari N, et al. Characterization and risk

assessment of polycyclic aromatic hydrocarbons (PAHs) in urban atmospheric Particulate of Tehran, Iran. Environ Sci Pollut Res Int.

2016;23(2):1820-32.

[30] Chen Q, Luo XS, Chen Y, Zhao Z, Hong Y, Pang Y, et al. Seasonally varied cytotoxicity of organic components in PM2.5

from urban and industrial areas of a Chinese megacity. Chemosphere. 2019;230:424-31.

[31] Gualtieri M, Ovrevik J, Holme JA, Perrone MG, Bolzacchini E, Schwarze PE, et al. Differences in cytotoxicity versus pro-

inflammatory potency of different PM fractions in human epithelial lung cells. Toxicol In Vitro. 2010;24(1):29-39.

[32] Niu X, Ho SSH, Ho KF, Huang Y, Sun J, Wang Q, et al. Atmospheric levels and cytotoxicity of polycyclic aromatic

hydrocarbons and oxygenated-PAHs in PM2. 5 in the Beijing-Tianjin-Hebei region. Environmental pollution. 2017;231:1075-84.

[33] Qishlaqi A, Beiramali F. Potential sources and health risk assessment of polycyclic aromatic hydrocarbons in street dusts of

Karaj urban area, northern Iran. Journal of Environmental Health Science and Engineering. 2019:1-16.

[34] Yin X, Sun Z, Miao S, Yan Q, Wang Z, Shi G, et al. Analysis of abrupt changes in the PM2.5 concentration in Beijing during

the conversion period from the summer to winter half-year in 2006–2015. Atmospheric Environment. 2019;200:319-28.

[35] Wang F, Guo Z, Lin T, Rose NL. Seasonal variation of carbonaceous pollutants in PM2.5 at an urban 'supersite' in Shanghai,

China. Chemosphere. 2016;146:238-44.

[36] Zhang HH, Li Z, Liu Y, Xinag P, Cui XY, Ye H, et al. Physical and chemical characteristics of PM2.5 and its toxicity to

human bronchial cells BEAS-2B in the winter and summer. J Zhejiang Univ Sci B. 2018;19(4):317-26.

[37] Arfaeinia H. Evaluation of public health impacts related to urban air pollution in Shiraz and Bushehr, Iran. 2015.

[38] Sowlat MH, Naddafi K, Yunesian M, Jackson PL, Shahsavani A. Source apportionment of total suspended particulates in an

arid area in southwestern Iran using positive matrix factorization. Bull Environ Contam Toxicol. 2012;88(5):735-40.

[39] Lim J-M, Lee J-H, Moon J-H, Chung Y-S, Kim K-H. Source apportionment of PM10 at a small industrial area using Positive

Matrix Factorization. Atmospheric Research. 2010;95(1):88-100.

[40] Schilirò T, Bonetta S, Alessandria L, Gianotti V, Carraro E, Gilli G. PM10 in a background urban site: chemical

characteristics and biological effects. Environmental toxicology and pharmacology. 2015;39(2):833-44.




Kashan, Iran

[41] Jiang N, Liu X, Wang S, Yu X, Yin S, Duan S, et al. Pollution Characterization, Source Identification, and Health Risks of

Atmospheric-Particle-Bound Heavy Metals in PM10 and PM2.5 at Multiple Sites in an Emerging Megacity in the Central Region of

China. Aerosol and Air Quality Research. 2019;19(2):247-71.

[42] Rai P, Chakraborty A, Mandariya AK, Gupta T. Composition and source apportionment of PM1 at urban site Kanpur in India

using PMF coupled with CBPF. Atmospheric research. 2016;178:506-20.

[43] Haghnazari L, Mirzaei N, Arfaeinia H, Karimyan K, Sharafi H, Fattahi N. Speciation of As (ΙΙΙ)/As (V) and total inorganic

arsenic in biological fluids using new mode of liquid-phase microextraction and electrothermal atomic absorption spectrometry.

Biological trace element research. 2018;183(1):173-81.

[44] Liu J, Chen Y, Chao S, Cao H, Zhang A, Yang Y. Emission control priority of PM2. 5-bound heavy metals in different

seasons: a comprehensive analysis from health risk perspective. Science of the total environment. 2018;644:20-30.

[45] Dai Q-L, Bi X-H, Wu J-H, Zhang Y-F, Wang J, Xu H, et al. Characterization and source identification of heavy metals in

ambient PM10 and PM2. 5 in an integrated iron and steel industry zone compared with a background site. Aerosol and Air Quality

Research. 2014;15(3):875-87.

[46] Bi C, Chen Y, Zhao Z, Li Q, Zhou Q, Ye Z, et al. Characteristics, sources and health risks of toxic species (PCDD/Fs, PAHs

and heavy metals) in PM2. 5 during fall and winter in an industrial area. Chemosphere. 2020;238:124620.

[47] Soleimani M, Amini N, Sadeghian B, Wang D, Fang L. Heavy metals and their source identification in particulate matter

(PM2. 5) in Isfahan City, Iran. Journal of environmental sciences. 2018;72:166-75.

[48] Kurosaki Y, Mikami M. Recent frequent dust events and their relation to surface wind in East Asia. Geophysical Research

Letters. 2003;30(14).

[49] Huremović J, Žero S, Bubalo E, Dacić M, Čeliković A, Musić I, et al. Analysis of PM10, Pb, Cd, and Ni atmospheric

concentrations during domestic heating season in Sarajevo, Bosnia and Herzegovina, from 2010 to 2019. Air Quality, Atmosphere &

Health. 2020;13(8):965-76.

[50] Liu X, Li C, Tu H, Wu Y, Ying C, Huang Q, et al. Analysis of the effect of meteorological factors on PM2. 5-associated

PAHs during autumn-winter in urban Nanchang. Aerosol Air Qual Res. 2016;16:3222-9.

[51] Lyu Y, Su S, Wang B, Zhu X, Wang X, Zeng EY, et al. Seasonal and spatial variations in the chemical components and the

cellular effects of particulate matter collected in Northern China. Sci Total Environ. 2018;627:1627-37.

[52] Yang H-H, Lai S-O, Hsieh L-T, Hsueh H-J, Chi T-W. Profiles of PAH emission from steel and iron industries. Chemosphere.

2002;48(10):1061-74.

[53] Yang H-H, Lee W-J, Chen S-J, Lai S-O. PAH emission from various industrial stacks. Journal of Hazardous materials.

1998;60(2):159-74.

[54] Pehnec G, Jakovljevic I. Carcinogenic Potency of Airborne Polycyclic Aromatic Hydrocarbons in Relation to the Particle

Fraction Size. Int J Environ Res Public Health. 2018;15(11).

[55] Hassanvand MS, Naddafi K, Faridi S, Nabizadeh R, Sowlat MH, Momeniha F, et al. Characterization of PAHs and metals

in indoor/outdoor PM10/PM2. 5/PM1 in a retirement home and a school dormitory. Science of the Total Environment. 2015;527:100-

10.

[56] Roig N, Sierra J, Rovira J, Schuhmacher M, Domingo JL, Nadal M. In vitro tests to assess toxic effects of airborne PM10

samples. Correlation with metals and chlorinated dioxins and furans. Science of the total environment. 2013;443:791-7.

[57] Chen Y, Luo XS, Zhao Z, Chen Q, Wu D, Sun X, et al. Summer-winter differences of PM2.5 toxicity to human alveolar

epithelial cells (A549) and the roles of transition metals. Ecotoxicol Environ Saf. 2018;165:505-9.

[58] Ndong Ba A, Cazier F, Verdin A, Garcon G, Cabral M, Courcot L, et al. Physico-chemical characterization and in vitro

inflammatory and oxidative potency of atmospheric particles collected in Dakar city's (Senegal). Environ Pollut. 2019;245:568-81.

5 International and 24 th National Conference on ...

Documents

Transcript of 5 International and 24 th National Conference on ...