Cytotoxic effect and role of exogenous antioxidants in carpet dust mediated toxicity in rat hepatocytes in vitro

Toxicology in Vitro 18 (2004) 419–425

www.elsevier.com/locate/toxinvit

Cytotoxic effect and role of exogenous antioxidants in carpetdust mediated toxicity in rat hepatocytes in vitro

Mohamed Ameen, Iqbal Ahmad, M. Syed Musthapa, Qamar Rahman *

Fibre Toxicology Division, Industrial Toxicology Research Centre, Post Box No. 80, M.G. Marg, Lucknow 226 001, India

Received 9 August 2003; accepted 3 December 2003

Abstract

Carpet industries bear a great deal of economic and commercial significance in India. In order to safe guard the workers against

the health hazards caused by dust in their occupational environment; it necessitates studying the biological importance of these

dusts. The present study was designed to investigate the toxicity of carpet dust (knotted and tuffted) on isolated rat hepatocytes. The

hepatocytes were isolated by collagenase perfusion method and cells were incubated with different concentration of carpet dust (100–

5000 lg/106 cells) with various time (30–180 min) intervals. An exogenous antioxidant vitamin-E also used to find out the role ofantioxidants and free radical production in carpet dust mediated toxicity. Cell viability by trypan blue exclusion and leakage of

enzyme lactate dehydrogenase (LDH) were determined. Reduced glutathione (GSH), formation of thiobarbituric acid reactive

substance (TBARS) were also measured. A significant decrease in the cell viability was observed after 60, 180 min upon incubation

with tuffted carpet dust, while knotted carpet dust caused a significant decrease in the viability after 180 min. LDH leakage was

parallel to the cell viability. Thiobarbituric acid reactive substance was significantly increased at 30 and 60 min with carpet dust

treated hepatocytes. Dust at 1000 and 5000 lg dose level showed significantly increased formation of TBARS at 30 min incubation.However, when hepatocytes were co-incubated with carpet dust and Vit-E (10, 15 lM), a significant decrease in LDH release andTBARS production was observed while 15 lM Vit-E showed an enhanced protection than 10 lM Vit-E treated hepatocytes. The

effect of carpet dust on cell viability, LDH leakage, TBARS production, GSH depletion was time and dose-dependent. Moreover,

we observed that tuffted carpet dust causes greater effect than knotted one on the above mentioned parameters. Our studies also

revealed that Vit-E in culture media diminishes the carpet dust mediated toxicity.

� 2003 Elsevier Ltd. All rights reserved.

Keywords: Antioxidant; Carpet dust; Cytotoxicity; Hepatocytes; Reactive oxygen species; Vitamin-E

1. Introduction

Prolonged inhalation of carpet dusts by the worker

from the polluted occupational environment during

carpet weaving leads to a variety of lung diseases

(Ozesmi et al., 1987). A unique form of interstitial lungdisease has been reported in carpet workers, and respi-

rable carpet shred (including fibres) were identified in

Abbreviations: Vitamin-E, Vit-E; Vitamin-C, Vit-C; Lactate dehy-

drogenase, LDH; Reduced glutathione, GSH; Thiobarbituric acid

reactive substance, TBARS; Alveolar macrophages, AMs; Reactive

oxygen species, ROS; Reactive oxygen intermediates, ROIs; 5,5-

Dithiobis-2-nitrobenzoic acid, DTNB; 2-Thiobarbituric acid, TBA;

Kreb–Hansleit buffer, KH; Trichloroacetic acid, TCA; Hydrochloric

acid, HCl; Malonaldehyde, MDA; Lipopolysaccharides, LPS*Corresponding author. Tel.: +91-522-227-2833/221-3786x314; fax:

+91-522-222-8227/221-1547.

E-mail address: [email protected] (Q. Rahman).

0887-2333/$ - see front matter � 2003 Elsevier Ltd. All rights reserved.

doi:10.1016/j.tiv.2003.12.003

workplace air-samples. Moreover, in carpet industries

weaving is the dustiest procedure where maximum

number of workers is engaged while weaving, the dust

becomes air-borne and causes direct exposure to the

population engaged in it. In order to assure welfare,

these workers are to be safe guarded against the healthhazards caused by dusts in their occupational environ-

ment, thereby necessitating the studies on biological

importance of these dusts. Light microscopic observa-

tion of carpet dust revealed that it contains fibres of

cotton, wool and their contaminants with endotoxin

(Ameen et al., 2002, 2003a,b). Due to scarcity of liter-

ature regarding carpet dust and its toxic potential, it is

necessary to review the components responsible for itstoxicity. Earlier study from our laboratory has shown

that dust collected from carpet weaving units i.e. tibbati,

knotted and tuffted have the ability to cause cytotoxic,

inflammatory and oxidant nature in rat lung in vivo

mail to: [email protected]

420 M. Ameen et al. / Toxicology in Vitro 18 (2004) 419–425

(Ameen et al., 2002). Similar investigation has also

shown an alteration in the cellular marker enzyme

profile tempting to speculate that the progression of

pulmonary disease occurs in carpet dust exposed ratlung (Ameen et al., 2003a). Several researchers have

documented that working with cotton, wool, textile is

classically associated with pulmonary health hazards to

the workers such as obstructive airway diseases (OADs),

byssinosis, lung tightness and asthma (Simpson et al.,

1998; Fishwick et al., 1994; Fishwick and Pickering,

1992; Ozesmi et al., 1987). Moreover, occupational lung

diseases associated with hemp, flex, jute and organicdust were well recognized, which contains large number

of biologically active agents (Buick and Magee, 1999;

Heeder et al., 2000; Melbostad and Edward, 2001).

Brown and Donaldson (1991) and Brown et al. (1989)

documented that dust collected from ledges in wool mill

has the ability to cause injury to lung epithelial cells,

alveolar macrophages in vitro. Toxic potential study

with rat alveolar macrophages (AMs) exposed to carpetdusts in vitro, also showed that carpet dust has the

ability to cause cytotoxic and reactive oxygen species

(ROS) production with a dose dependent manner

(Ameen et al., 2003a,b).

Primary culture of rat hepatocytes are used in many

toxicological studies, and early toxicity prediction by

such primary culture is well documented (Paillard et al.,

1999). These cells were chosen mainly for their epithelialcharacter and the capacity to maintain a sufficient level

of xenobiotic metabolism in vivo (Rogiers and Ver-

cruysse, 1993). Hepatocytes have been proved to be a

useful model for studies of toxicological importance.

These cells have been used for the evaluation of cyto-

toxicity (Fleury et al., 1983; Aslam et al., 1992), car-

cinogenicity (Rahman and Casciano, 1985) and

genotoxicity (Denizean et al., 1985) of fibres/particles.In this context, we aimed to investigate possible

mechanisms by which carpet dust causes dust mediated

pulmonary health hazard. We have studied the state of

cytotoxic markers by cytoplasmic enzyme release (LDH)

and oxidative tone in cells incubated with carpet dust,

the formation of thiobarbituric acid reactive substances

(TBARS) and antioxidant estimation (reduced gluta-

thione; GSH) in primary culture of rat hepatocytesincubated with and without Vit-E to evaluate the role of

exogenous antioxidant and its protective efforts in car-

pet dust mediated toxicity.

2. Materials and methods

2.1. Chemicals

Collagenase type IV, vitamin-E (Vit-E) and 5,5-

dithiobis-2-nitrobenzoic acid (DTNB) were purchased

from Sigma Chemical Co., (St. Louis, MO, USA). 2-

Thiobarbituric acid (TBA) from BDH Chemical, Eng-

land. All the other chemicals/reagents were of analytical

grade and purchased from Sisco Research Laboratory

or Spectrochem Pvt., Ltd., Mumbai, India.

2.2. Dust collection

Dust sampling was performed on various carpet

weaving units (i.e., knotted and tuffted) following theprocedure of Milton et al. (1995) with slight modifica-

tion as mentioned elsewhere (Ameen et al., 2003a,b).

Dust generated during weaving of carpet that has fallen

on and around the looms were collected and prepared to

make it as fibre size of <10 lm for the use in toxico-

logical evaluation according to Zaidi (1969). The carpet

dusts were autoclaved overnight to eliminate endotoxin

and suspensions were made with phosphate buffer saline(PBS). They were then sonicated and vigorously vor-

texed to ensure an uniform suspension prior to use.

2.3. Animals

Male Wister rats were (average body weight 250 g)

obtained from animal breeding facility, Industrial Tox-

icology Research Centre, Lucknow, India. The animals

were maintained under standard animal husbandry

conditions. They were maintained on standard pellet

diet (Amrut feeds, Pune, India) and given tap water

ad libitum.

2.4. Isolation of hepatocytes

Hepatocytes were isolated by collagenase perfusion

technique by the method of Moldeus et al. (1978). The

isolated hepatocytes were placed in Kreb–Hansleit (KH)buffer (pH 7.4) containing 0.5% bovine serum albumin

(BSA). The cells were dispersed by gentle stirring. All

buffers were bubbled with O2 (95%) and CO2 (5%) prior

to use. The dispersed cells were filtered through four

layer of cotton gauge to remove the connective tissues

and clamps of cells. Then the filtrate was centrifuged

(50g) for 2 min at 4 �C. The cells were counted in ahaemocytometer. The viability was tested using trypanblue exclusion test and viability of the cells was 85–90%

throughout the study.

2.5. Treatment and incubation of hepatocytes

Kreb–Hansleit (KH) buffer with 0.5% BSA was used

as an incubation medium. Isolated hepatocytes (106

cells/ml) were incubated with 100, 500, 1000 or 5000 lg/ml dust samples at 37 �C in a metallic shaker with

moderate shaking (30 oscillation/min). Aliquots were

taken at different time interval for viability test and

cytotoxic marker assay.

M. Ameen et al. / Toxicology in Vitro 18 (2004) 419–425 421

2.6. Cell viability and cytosolic enzyme release

Plasma membrane integrity was evaluated by the

cellular exclusion of trypan blue. Cytotoxicity was as-sessed by measuring the release of lactate dehydrogenase

(LDH) in the medium by the method of Wotton (1964).

Protein concentration was determined by the method of

Lowry et al. (1951) using bovine serum albumin as the

standard.

20

40

60

80

100

% V

iab

ility

**

**

2.7. Assay of thiobarbituric acid reactive substances

Thiobarbituric acid reactive substances (TBARS)

produced in the hepatocyte suspension during incuba-

tion of carpet dust with and without Vit-E were assayed

according to Buege and Aust (1978). Briefly, a part ofhepatocyte suspension (0.5 ml) was added to 0.5 ml of

15% trichloroacetic acid (TCA). Thiobarbituric acid

stock reagent containing 15% TCA, 0.375% TBA and

0.25 N HCl (2 ml) were added to the reaction mixture,

followed by boiling for 15 min. The reaction mixture

was then cooled and centrifuged at 1000g for 15 min.The absorbance of the supernatant at 535 nm was

determined with malonaldehyde (MDA) as the stan-dard.

0100 500 1000 5000

Concentration (µg)

Fig. 1. Effect of carpet dust on isolated rat hepatocytes viability in

primary culture at different dust concentration (100–5000 lg) of (r)knotted or (j) tuffted for 3 h million cells�1. Each value represents the

mean±SE of four different observations. Differences were considered

significant at �p < 0:05 values against its corresponding control.

60

80

100

120

Via

bili

ty **

2.8. Estimation of intracellular glutathione

Intracellular GSH content was estimated in the cell

pellets of control and dust treated hepatocytes by the

method of Sedlack and Lindsay (1968). Cells were lysed

with 0.1% EDTA solution and precipitating reagent

which contains 0.16% metaphosphoric acid, 0.02%

EDTA and 3% NaCl. After mixing, the solution was

allowed to stand for 5 min before being filtered. Two

milliliter of filtrate was added to 4 ml of disodiumhydrogen phosphate (0.1 M, pH 8.0) and 1 ml of DTNB

reagent. A blank was prepared from 1.2 ml of precipi-

tating reagent, 0.8 ml of EDTA solution, 4 ml of diso-

dium hydrogen phosphate and 1 ml of DTNB reagent.

The color was immediately read at 412 nm with the help

of spectrophotometer.

0

20

40

0 30 60 180Incubation time (min)

%

Fig. 2. Effect of carpet dust on isolated rat hepatocytes viability in

primary culture at different time incubation (0–180 min) at a concen-

tration 1000 lg of carpet dust (r) knotted or (j) tuffted millioncells�1. Each value represents the mean±SE of four different obser-

vations. Differences were considered significant at �p < 0:05 values

against its corresponding control.

2.9. Statistical analysis

The data were expressed as the mean of four experi-

ments ± SE. The statistical differences among thegroups were determined by one-way analysis of variance

(ANOVA) to discriminate where the differences were.

The statistical comparison between knotted and tuffted

carpet dust at each time intervals or at each dust con-

centrations were performed. Data were considered

with statistical differences at a significance level of

p < 0:05.

3. Results

In order to determine the cytotoxicity, hepatocytes

were incubated with various concentration of carpetdust (100–5000 lg), and at different incubation times(30–180 min) shown in Figs. 1 and 2, respectively. The

effect of carpet dusts i.e., knotted, tuffted on the rat

hepatocytes viability. Trypan blue exclusion test was

used to determine the cell viability during 3 h of incu-

bation. Carpet dust caused a remarkable decline in cell

viability as compared to control. However, tuffted car-

pet dust showed a significant (p < 0:05) decline in thecell viability evaluated after 60 min (68%±2.1%), 180

min (60%±2.2%) of incubation as compared to control,

0

10

20

30

40

100 500 1000 5000Concentration (µg)

nm

ol G

SH

106 c

ells

-1 Control

Knotted

Tuffted

*a*a

a

Fig. 4. Effect of carpet dust on isolated rat hepatocytes intracellular

GSH in primary culture at different dust concentration (100–5000 lg)for 3 h million cells�1. Each value represents the mean±SE of four

different observations. Differences were considered significant at�;ap < 0:05 values against its corresponding control, knotted carpet

dust, respectively.


as well as to knotted carpet dust. Knotted carpet dust

showed a significant decline in cell viability only at 180

min (68%±2.4%) of incubation as compared to the

control.In accordance with the results of trypan blue exclu-

sion assay, carpet dust also caused a concentration and

time dependent release of LDH from rat hepatocytes.

The results of cytoplasmic enzyme (LDH) release after

treatment with different concentrations (0–5000 lg) andvarious time incubations (0–180 min) are shown in Fig.

3. Significant (p < 0:05) elevation in LDH leakage aftertreatment with 500, 1000 and 5000 lg quantifies of tu-ffted carpet dust were noted at 180 min as compared to

control as well as knotted carpet dust treated group,

while, significant effects were observed 60 and 180 min

incubation. Knotted carpet dust has caused significant

LDH leakage at 60 and 180 min of incubation as com-

pared to the control.

The intracellular GSH depletion in the rat hepato-

cytes by the effect of carpet dust is illustrated in Fig. 4.Tuffted carpet dust caused a significant decline in GSH

level (500–5000 lg) at 180 min of incubation, as com-pared to the control and as knotted carpet dust; knotted

carpet dust has showed a significant decline only at 180

min with 1000 and 5000 lg as compared to the respec-tive control. Significant decline in the GSH level was

noted after 30 min exposure to 1000 lg of tuffted carpetdust as compared to control and knotted carpet dust.However, significant decline was noted at 60, 180 min of

incubation with knotted carpet dust as compared to the

control as shown in Fig. 5.

Isolated rat hepatocyte suspension has showed that

TBARS production to be significantly increased (p <0:05) in heptocytes incubated (60, 180 min) with boththe dust (1000 lg) as compared to the controls (Fig. 6).However, tuffted carpet dust showed significant increase(p < 0:05) in TBARS formation in hepatocytes incu-bated at 60, 180 min with 1000 lg as compared tocontrol and knotted carpet dust treated hepatocytes.

100

120

140

160

180

0 100 500 1000 5000Concentration (µg)

LD

H (

% c

on

tro

l)

Knotted

Tuffted*

*

a

a

a

Fig. 3. Cytotoxic effect of carpet dust on isolated rat hepatocytes primary cu

time incubation (0–180 min) with carpet dust at a concentration of 1000 lgobservations. Differences were considered significant at �;ap < 0:05 values ag

The effect of exogenous antioxidants and the sup-

pression of reactive oxygen species (ROS) production

mediated by different concentration of carpet dusts, as-

says through LDH release and TBARS formation were

documented in Fig. 6 and Table 1, respectively. Effect of

vit-E (10, 15 lM) on LDH release in isolated rat he-

patocytes co-incubated with carpet dusts were exam-

ined. The LDH release mediated by carpet dusts wereconsiderably controlled by 10 lM vit-E coincubation

attenuates the appreciable level of LDH release which is

statistically significant (p < 0:05). Similarly, TBARSproductions were also significantly controlled by 15 lMVit-E coincubated with isolated hepatocyte treated with

carpet dust. This result further suggests that carpet dust

has the capability to produce reactive oxygen species

these can be diminished by exogenous treatment of an-tioxidants (Fig. 7).

Knotted

Tuffted

100

120

140

160

180


LD

H (

% c

on

tro

l)

**

aa

lture at different dust concentration (100–5000 lg) for 3 h and differentmillion cells�1. Each value represents the mean±SE of four different

ainst its corresponding control, knotted carpet dust, respectively.

nm

ol G

SH

106 c

ells

-1

Control

Knotted

Tuffted

0

10

20

30

40


* a

*a

a

Fig. 5. Effect of carpet dust on isolated rat hepatocytes intracellular

GSH in primary culture at different time incubation (0–180 min) with

carpet dust at a concentration of 1000 lg million cells�1. Each valuerepresents the mean±SE of four different observations. Differences

were considered significant at �;ap < 0:05 values against its corre-

sponding control, knotted carpet dust, respectively.

Control

Knotted

Tuffted

0

3

6

9

12

15


TB

AR

S

(nm

ol f

orm

ed 1

06 c

ells

-1

*

a

*

a

Fig. 6. Effect of carpet dust on isolated rat hepatocytes TBARS pro-

duction in primary at different time incubation (0–180 min) with carpet

dust at a concentration of 1000 lg million cells�1. Each value repre-sents the mean±SE of four different observations. Differences were

considered significant at �;ap < 0:05 values against its corresponding

control, knotted carpet dust, respectively.

100

110

120

130

140

150

160

170


LD

H (

% c

on

tro

l) Knotted

Tuffted

Knot+10µM

Knot+15µM

Tuff +10µM

Tuff+15µM

*

a

a

*

*a*

*

cb

Fig. 7. Alteration of exogeneous antioxidant (Vit-E; 10, 15 lM) incytotoxic effect of carpet dust on isolated rat hepatocytes primary

culture at different dust concentration (100–5000 lg) for 3 h millioncells�1. Each value represents the mean±SE of four different obser-

vations. Differences were considered significant at �;ap < 0:05 values

against its corresponding control, knotted carpet dust, respectively.b;cp < 0:05 corresponds significance of Vit-E (15 lM) treatment againstknotted carpet dust alone treated group, tuffted carpet dust alone

treated group, respectively.

Table 1

Effect of Vit-E on carpet dust mediated TBARS formation on isolated

rat hepatocytes

Parameter TBARS

(nmol MDA formed 106 cells�1)

Control 5.71±0.35

Knotted 8.32±0.45�

Knott ± 10 lM Vit-E 7.65±0.32�

Knott ± 15 lM Vit-E 6.85±0.42b

Tuffted 14.36±0.36�;a

Tuffted± 10 lM Vit-E 10.67±0.52a

Tuffted± 15 lM Vit-E 6.32±0.42�;c

The isolated rat hepatocyte suspension was incubated at 37 �C withand without 10 lM Vit-E, 15 lM Vit-E with 1000 lg of dust millioncells�1 for 3 h. Each values represents the mean±SE of four different

observations. Differences were considered significant at �;ap < 0:05

values against its corresponding control, knotted carpet dust, respec-

tively. b;cp < 0:05 corresponds significance of Vit-E (15 lM) treatmentagainst knotted carpet dust alone treated group, tuffted carpet dust

alone treated group, respectively.


4. Discussion

Since last decade, there has been several vital debates

and discussions in pharmaco-toxicology on the use ofanimal models for human safety assessment studies

(Kocarek et al., 1995). In toxicology, it is generally ac-

cepted that some differences among species in the

intensity and duration of the effects following exposure

to xenobiotics exist (Nebbia, 2001). Hepatocytes have

proven to be a useful model for studies of xenobiotic

transformation (Crosis and Williams, 1985), carcino-

genesis, mutagenesis (Mitchell et al., 1983) and cyto-toxicity (Chao et al., 1988; Sandy et al., 1988). However,

in vitro models for toxicity assessment are required to

study the toxic effects of air-borne carpet dust. In our

present investigation the assessment of cytotoxic and

oxidative tone and antioxidant level have been investi-

gated upon exposure of carpet dusts to rat hepatocyte

primary culture and an attempt was made to study themechanism of toxic potential. With the present obser-

vations, the results of cell viability and cytotoxic effects

were found to be in accordance with our previous results

observed for alveolar macrophages in vitro (Ameen

et al., 2003a,b).

Cell membrane poses primary role in cell viability as

well as enzyme leakage, cellular damage caused by toxic

substances is frequently accompanied by an increase inthe cell membrane permeability (Smith and Orrenius,

1984). Usually, the cell viability is estimated by an


uptake of trypan blue and the leakage of cytoplasmic

enzyme (LDH), where LDH serves as an indicator of the

cellular integrity (Kim et al., 2001). The viability of the

cell coincides more with LDH leakage in a time and dosedependent manner, where in the tuffted carpet dust has

been found to have a greater effect than knotted carpet

dust. Furthermore, Russel et al. (1982) and Donaldson

(1990) supported the observation that cotton dust and

cotton bracts produces disruption of bronchial epithe-

lium. Our recent study, Ameen et al. (2003a) also re-

vealed that single intratracheal instillation of carpet dust

causes significant elevation in alkaline phosphatase,gamma glutamyl transferase in rat BALF at post expo-

sure period. Further increased level of clara cell secretory

protein (CC16), a marker of clara cell damage, where

also observed in BALF after post exposure of tuffted and

knotted carpet dusts; among former one shows signifi-

cant effect than latter one. The present study documented

that in presence of carpet dust the content of GSH de-

creased markedly in hepatocytes in a dose and timedependent manner. Toxic potential of workplace dusts

have been shown to have clear cut association between

TBARS formation and total GSH level; earlier studies

provide support for this existence of these changes in

cellular biochemistry associate with cellular injury

(Hogberg and Kristoferson, 1977). Moldeus and Quan-

guan (1987) suggested that GSH depletion may ulti-

mately leads to cell death by impairing the cells defenseagainst oxidative damage. On the other hand, in presence

of carpet dust the levels of reduced glutathione, were

markedly decreased. A decreased level of reduced glu-

tathione content was found to be negatively correlated

with an LDH leakage. This decrease in glutathione levels

may possibly contribute to the carpet dust mediated

toxicity.

Previous studies from our laboratory documentedthat carpet dust has the ability to produce reactive

oxygen species and reactive nitrogen species as seen in

in vivo and in vitro assays (Ameen et al., 2002, 2003a,b).

These findings tempted us to use an exogenous antiox-

idant to find out whether there is any reduction in the

carpet dust mediated toxicity. Among many antioxi-

dants that are available, we chose Vit-E and Vit-C, in

which Vit-E was found to be more significant. As suchVit-E is the most effective antioxidant that stabilizes cell

membranes by interfering with lipid peroxidation

(Tappel, 1970; Andres and Cascales, 2002). Production

of thiobarbituric acid reactive substances was sup-

pressed by Vit-E remarkably, where in a concentration

of 15 lM Vit-E showed a significant effect than a con-

centration of 10 lM. It is documented that Vit-E

inhibits lipid peroxidation by donating a hydrogen atomto the lipid radical thereby terminating the propagative

process (Mc Cay, 1985). By these present findings of

increased, cytosolic enzyme release, TBARS formation

and attenuation of these effects by supplementation of

exogenous antioxidants supports the suspecion of carpet

dust mediated oxidative tone in hepatocytes, which may

be the possible factor in the attack of bio-membranes,

seen in the acute toxicity.Carpet dust caused lipid peroxidation, cytotoxicity

and oxidative stress in isolated rat hepatocytes, which

was correlated with the injury of the hepatocytes. It

seems likely that the oxidative metabolism greatly con-

tributes to lipid peroxidation and that a decrease of

intracellular glutathione is closely related to its toxicity.

Earlier studies further support that fibres such as cellu-

lose have the ability to produce reactive oxygen species(ROS) (Cullen et al., 2000), AMs also showed an in-

creased production of reactive oxygen intermediates

(ROIs) by treatment with O3, endotoxin and silica in

vivo and interferon-alpha, lipopolysaccharides (LPS) in

vitro (Denis, 1994). Earlier observation from our labo-

ratory substantiates that dust produced during weaving

is not inert, but has the ability to produce ROIs, wherein

tuffted carpet dust causes a greater effect than knottedone. Whether cotton, wool dust or other occupational

exposures are responsible for the development of lung

disease in the workers still remains to be determined.

There seems to be a general impression that all fibres/

particulates small enough to reach the distal lung pose a

health threat to humans. As an immediate solution to

the problem, the dust exposure in factories should be

controlled, and also a detailed toxicological evaluationhas to be carried out.

In conclusion, the above experiments prove that a

single in vitro model alone may not predict all potential

aspects of toxicity of the dust. However, the content of

wool may play a vital role in the pathogenicity.

Acknowledgements

The authors are thankful to Dr. P.K. Seth, Director

for his keen interest and support on this study. The

financial grant from the Ministry of Environment &

Forests (Government of India) is thankfully acknowl-

edged. Thanks are also due to Mr. M. Ashquin, Tech-

nical officer for his skillful technical assistance. One of

the authors M. Ameen thankfully acknowledges Prof.

M.A. Baig, Head, Department of Biochemistry, Ham-dard University, New Delhi, India for his invaluable

suggestion and kind guidance for making this manu-

script successfully.

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Cytotoxic effect and role of exogenous antioxidants in carpet dust mediated toxicity in rat hepatocytes in vitro