Effect of Chronic Exposur to Sulphur Dioxide Vapour and Sulphur Related Compounds on Serum

EFFECT OF CHRONIC EXPOSUR TO SULPHUR DIOXIDE VAPOUR AND SULPHUR RELATED COMPOUNDS ON SERUM LEVELS OF INSULIN, GLUCOSE, C-PEPTID, AND ZINC

Tahia H. Saleem, Howaida A. Nafady ,Hosnee A Hassan , Mahmoud R. Abed El-Fadeel

From departments of Biochemistry, Internal Medicine and Physiology, faculty of Medicine, Assuit University, Egypt

ABSTRACT

Sulphur dioxide and trioxide vapors are currently emitted from the superphosphate factory in Mankabad, AssuitProvince with the possibility of environmental pollution thought the limits of these vapors in the factory may be within the permissible authorized limits. So the present work aimed to elucidate the effects of exposure of these vapors and other related sulphur compounds on fasting blood glucose, insulin, C-peptide , Zinc levels and serum alkaline phosphatase among the workers joining this factory and as a preliminary study aiming a detailed follow-up program concerning detection of any metabolic disturbances or environmental pollution diseases that could possibly be developed.

Twenty-eight male workers were recruited from the workers among the Superphosphate Factory in Mankabad, AssuitProvince. They were chosen on the basis of being continuously exposed to sulphur dioxide and related sulphur compounds during their daily working hours and representing all the areas of the factory. Their ages ranged between 26 and 55 years with meanSD ( 38.646.2). In addition, 84% of them are exposed for 10 years or more.The control group compromised 14 healthy males that were selected to match as nearly as possible the exposed group in age and socioeconomic class. They have no history of potential exposure to this type of environmental pollution. Their ages ranges between 27 and 50 years with meanSD (39.046.2).

From each participant fresh serum samples were used for estimation of fasting blood glucose. In addition serum insulin , C-peptide , Zinc levels and serum alkaline phosphatase were also estimated.Statistical comparison revealed that the workers of the exposed group had meanSD fasting blood glucose levels that were significantly higher than those of control (P<0.001) (mean SD of exposed group 11.315.99 mg/dl) and that of the control was (73.55.854mg/dl). The mean SD serum insulin concentration for the control group was (12.434.24 uU/ml), while that for the exposed group was (16.360.89 uU/ml) and proved to be significantly higher than that of control (P<0.001) . The mean SD of C-peptide for the control group was (1.0571.308ng/ml), while that for the exposed group was (1.9671.472 uU/ml) and proved to be significantly higher than that of control (P<0.02) . Furthermore, in every individual of the exposed group, the level of serum zinc was raised and the mean SD was (118.932.12 ug/d)l which is significantly higher than that of the control group (mean SD was 84.788.604 ug/dl) and p<0.001.It is noteworthy to emphasize that all the workers studied showed serum alkaline phosphatase levels (meanSD 99.9516.93 Iu/L) higher than that reported for control (meanSD 65.717.7Iu/L) and p<0.001, an index pointing to sort of biochemical hepatic dysfunction. Thus, individuals of the exposed group appear to have a sort of hyperglycemia associated with abnormal hyperinsulinaemia in reference to control, although their insulin levels and C-peptide are within the international official ones (up to 25 and 5 ng/ml).

INTRODUCTION

Man is constantly exposed to chemical substances in the environment which have the potential to produce adverse health effects particularly on continuous exposure to these agents. This occurs as a result of performance of daily activities and/ or during surveillance of food and water. The effects of exposure to sulphur dioxide and trioxide vapors on pulmonary functions even at low concentrations have been reported . Irritant-induced inflammation of the airways may aggravate respiratory allergy induced by chemical respiratory allergens (Bates et al., 1970, Hazucha et al., 1973 and Arts et al., 2010). For lung tissues of male mice, exposure to SO2caused increase of levels of IL-6 and TNF-alpha and IL-6 so that inflammatory reaction could be induced in lung tissue by SO2inhalation and the inflammatory reaction might relate to these cytokines and determination of cytokines in lung may be more valuable than in serum when lung injury caused by SO2(Meng et al., 2005).

The effects of exposure to these pollutants on liver functions and blood glucose levels in human have not been reported. Meanwhile, fewer studies were included but concerned with experimental animals. Exposure of rats to different doses of dioxide lead to significant decrease in the serum albumin/globulin ratio as well as the development of dystrophy in their livers. Total liver microsomal cytochrome P-450 (CYP) contents were diminished in SO2-exposed rats, SO2 exposure can suppress CYP2B1/2 and CYP2E1 in lungs and CYP2B1/2 in livers of rats, thus modifying the liver and lung toxication/detoxication potential, and the total liver microsomal CYP contents were diminished, although the activity and mRNA expression of CYP2E1 in rat livers were not affected by SO2 exposure(Bushtueva, 1966 and Qin Meng 2006).

David et al., 1966 and Qin Meng 2005described the toxicity of PonceauS (2-(5-sulfo-2,4-xylylazo)-1-Naphthol-4-sulfonic acid) administered to rats, mice and dogs in their diets. They reported the development of small haemosidric foci in livers of these animals after three years toleration.SO2 is a systemic oxidative damage agent. It results in a significant increase in the lipid perioxidation process in all organs tested of mice of both sexes, which is accompanied by changes of antioxidant status in these organs.SO2may cause toxicological damage to multiple organs of animals, and it is suggested that the oxidative damage produced by SO2inhalation may influence or promote the progression or occurrence of some disease states of various organs, not only to respiratory system. Further work is required to understand the toxicological role of SO2on multiple or even all organs in mammals (Meng ,2003).

Sulphur dioxide and trioxide vapors are currently emitted from the superphosphate factory in Mankabad, AssuitProvincewith the possibility of environmental pollution thought the limits of these vapors in the factory may be within the permissible authorized limits. So the present work aimed to elucidate the effects of exposure of these vapors and other related sulphur compounds on fasting blood glucose, insulin, C-peptide, Zinc levels and serum alkaline phosphatase among the workers joining this factory and as a preliminary study aiming a detailed follow-up program concerning detection of any metabolic disturbances or environmental pollution diseases that could possibly be developed.

MATERIAL AND METHODS

Twenty-eight male workers were recruited from the workers among the Superphosphate Factory in Mankabad, AssuitProvince. They were chosen on the basis of being continuously exposed to sulphur dioxide and related sulphur compounds during their daily working hours and representing all the areas of the factory.Their ages ranged between 26 and 55 years; meanSD (38.646.2). In addition, 84% of them are exposed for 10 years or more.The control group compromised 14 healthy males that were selected to match as nearly as possible the exposed group in age and socioeconomic class. They have no history of potential exposure to this type of environmental pollution. Their ages ranges between 27 and 50 years; meanSD (39.046.2).

From each participant fresh serum samples were used for estimation of fasting blood glucose. In addition serum insulin , C-peptide , Zinc levels and serum alkaline phosphatase were also estimated. Fasting blood glucose concentrations were estimated by Trinder glucose-oxidase method (1969) using the kits provided by biomerieux products and laboratory reagents, Marcy-1 Etoile 69260 charbonnieres-less Bairs/France. Serum zinc concentrations were measured by Shimadzu Atomic Absorption Flame Spectrophotometer-AA-630-02 at 213.9 A using air- acetylene fuel, the hollow cathode lamp the slit width was 1.9 A and lamp current was 10 m A.

Insulin concentrations of all the sera samples were estimated by radioimmunoassay technique using the human serum insulin kits purchase from Immuno-Nuclear Corporation Cat. No,-06-L2 post No. xp0297,5-ILWATER; MINNESOTA, USA and C-peptide was determined according to the method described by Eashan by ELIZA kits product No. 2725-300, produced by Monobind Inc., Lake forest, USA. Serum alkaline phosphatase was assayed by the method reported by King and King 1954.

Statistical analysis : results are expressed as meanSD unpaired Student's test was used for comparisons between the groups. p-value less than 0.05 were considered to be significant.

RESULTS

Table 1 shows the individual mean, SD and SE values of fasting blood glucose (mg/dl), sera insulin (uU/ml) , C-peptide (ng/ml) and zinc (ug/100 ml) as obtained from the control group (n=14).

Table I :Fasting blood glucose (mg/dl), serum insulin (uU/ml). C-peptide (ng/ml) and serum zinc(ug/100 ml) in the control group

No. / Glucose / Insulin * / C-peptide / Zinc
1
2
3
4
5
6
7
8
9
10
11
12
13
14 / 70
65
65
75
75
80
75
80
70
75
78
83
65
73 / 10
11
19
11
7
12
18
22
10
12
8
12
12
10 / 0.8
1.5
3.1
0.9
2.5
1.9
2.6
3.1
1.2
0.9
1.1
4
2.5
3.5 / 80
81
90
80
74
85
100
104
80
84
74
88
85
82
Mean
S.D.
S.E. / 73.5
5.854
1.565 / 12.429
4.274
1.142 / 1.057
1.308
0.247 / 84.786
8.604
2.2995

The individual , mean, SD and SE values of fasting blood glucose , sera insulin, C- peptide and zinc that obtained from the exposed group of workers are shown in table II

Table II :Fasting blood glucose (mg/dl), serum insulin (uU/ml). C-peptide (ng/ml) and serum zinc (ug/100 ml) in the exposed group of workers

No. / Glucose / Insulin * / C-peptide / Zinc
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28 / 112
118.7
150
118.7
131.2
131.2
118
100
112
106.2
150
100
100
118
125
131
88
100
106
88
118
100
94
112
125
118
110
118 / 11
16
11.5
9
27
13
14
14
18
15
11.5
21
16
17
14
16
13
21
20
13
24
13
16
21
14
11
24
24 / 0.8
1
0.9
0.7
5
1.1
1.2
1.6
2.1
1.5
0.7
4.1
1.9
1.2
1
0.8
0.7
3.1
4.2
0.9
4.9
0.7
1.6
3.1
1.1
0.7
4
4.5 / 115
117
130
115
112
115
125
113
116
140
119
110
142
110
119
112
132
136
113
96
100
112
110
113
119
133
125
131
Mean
S.D.
S.E. / 113.893
15.990
3.022 / 16.357
4.706
0.889 / 1.967
1.472
0.278 / 118.929
11.238
2.124

Statistical comparison (Table III , figure 1) revealed that the workers of the exposed group had meanSD fasting blood glucose levels that were significantly higher than those of control (P<0.001) (mean SD of exposed group (113.89315.99) mg/dl and that of the control was (73.55.854mg/dl).

Table III Comparison between the control group and the exposed group in all parameters studied

Group / Glucose
Mg/dl / Insulin
uU/ml / C-peptide
Ng/ml / Zinc
Ug/dl / Alkaine phosphatase Iu/L
Control (N=14)
MeanS.E. / 73.51.565 / 12.4291.142 / 1.0570.247 / 84.7862.299 / 65.717.7Iu/L
Exposed(N=28)
MeanS.E. / 113.8933.022 / 16.3570.889 / 1.9670.278 / 118.9292.124 / 99.9516.93Iu/L
P / <0.001 / <0.001 / <0.02 / <0.001 / <0.001

Figure 1: Comparison between the control group and the exposed workers for fasting blood glucose level (mg/dl)

The mean SD serum insulin concentration for the control group was 12.434.24(uU/ml), while that for the exposed group was 16.360.89 (uU/ml) and proved to be significantly higher than that of control (P<0.001) (Table III and Figure 2).

Figure 2: Comparison between the control group and the exposed workers for Serum insulin level (uU/ml)

The mean SD of C-peptide for the control group was 1.0571.308(ng/ml), while that for the exposed group was 1.9671.472 (uU/ml) and proved to be significantly higher than that of control (P<0.02) (Table III and Figure 3).

Figure 3: Comparison between the control group and the exposed workers for C-C-peptide level (ng/ml)

Furthermore, in every individual of the exposed group, the level of serum zinc was raised and the mean SD was 118.932.12 ug/dl, a level that proved to be significantly higher than that of the control group (mean SD was 84.788.604 ug/dl) and p<0.001 (Table III and figure 4).

Figure 4: Comparison between the control group and the exposed workers for serum zinc level (ug/dl)

It is noteworthy to emphasize that all the workers studied showed serum alkaline phosphatase levels (meanSD 99.9516.93 Iu/L) higher than that reported for normal control (meanSD 65.717.7Iu/L) and p<0.001; an index pointing to sort of biochemical hepatic dysfunction .

Thus, individuals of the exposed group appear to have a sort of hyperglycemia associated with abnormal hyperinsulinaemia in reference to control, although their insulin levels and C-peptide are within the international official ones (up to 25 and 5 ng/ml).

DISCUSSION

Sulfur dioxide SO2 is a ubiquitous air pollutant, present in low concentrations in the urban air and in higher concentrations in the working environment. The data obtained from the present work revealed that prolonged exposure to sulphur dioxide and other related sulphur compounds induces metabolic changes in the fasting blood glucose, serum insulin , C –peptide and serum zinc levels that are not associated with any clinical manifestations. Such changes are presented in the form of raised fasting blood glucose and serum zinc levels. Referring to insulin and C-peptide, the obtained serum values despite being higher than control, still they are within the reported official normal levels (up to 25 uU/ml and 5 ng/ml).

Vajo Z et al., 2001 , Wild et al., 2004 and Chandrasekhar et al., 2010 postulated that chronic hepatic disease produces hyperinsulinaemia and endogenous insulin resistance before the glucose tolerance is impaired and that diabetes appears only when B-cells are decompensated. The etiology of this insulin resistance and the eventual B- cell decompensation remains obscure. Ahmad, (2004) and Walsh, (2005) suggested that pancreatic insulin secretion is not increased in cirrhosis, so peripheral hyperinsulinemia is due solely to decreased hepatic insulin degradation secondary to either spontaneous portal systemic shunting or to parenchymal damage. In addition , Kjeldsen et al., (1999) and Ferrer et al., (2009) concluded that peripheral hyperinsulinaemia observed in liver cirrhosis could reflect only diminished hormone metabolism since the insulin secretion in this case is decreased. Therefore , the occurrence of raised fasting blood glucose levels together with the other findings could be considered indicative of existence of glucose intolerance in the exposed group of workers.

WHO 1999 and 2008 stated that in diagnosis of diabetes mellitus, glucose values during a glucose tolerance tests are invariably more helpful than simultaneous insulin levels, as the latter tend to be more variable with logarithmic distribution in normal population. Accordingly the authors stated that, in normal subjects and borderline diabetics the plasma insulin level is largely determined by the prevailing glucose concentration as it is more reproducible and more useful diagnostic discreminant. So, data of this work could be considered indicative that workers of the exposed group are more or less liable to exist diabetic manifestations later on.It is noteworthy to emphasize that all the workers studied showed serum alkaline phosphatase levels (meanSD 99.9516.93 Iu/L) higher than that reported for normal control (meanSD 65.717.7Iu/L) and p<0.001; an index pointing to sort of biochemical hepatic dysfunction . Hence, the interpretation of all the above data could be attributed in the basis of glucose intolerance, most probably secondary to hepatic dysfunction. This in turn, may result from a diminished hormone metabolism, most probably secondary to decrease liver insulinase activity or a decrease in liver insulinase affinity. On the other hand, the possibility of peripheral raised insulin level secondary to maintained hyperglycemia could not be excluded as another contributing factor. Lastly but not finally, exposure to sulphur dioxide and other related sulphur compounds seems to induce a subtle chronic or insidious effect that could not be detected clinically but seems to be biochemically apparent and hence further concerning estimation of glucose tolerance and other related parameters together with a full report of various conventional liver functions tests seems essential before a definite conclusion is settled.

Raised serum zinc levels are contradictory to the other findings, Hence , its interpretation remains obscure. It may possibly result from changes or disturbances affecting its levels in erythrocytes (as a result of exposure) or unavailability to proper utilization by tissues in addition the role of zinc as antioxidant should be considered against lipid perioxidation, So further investigation involving studies on the biochemical changes in RBCs and its utilization by other tissues seems to be essential.

SO2 is a systemic oxidative damage agent. It results in a significant increase in the lipid perioxidation process in all organs, which is accompanied by changes of antioxidant status in these organs. SO2 may cause toxicological damage to multiple organs of animals, and it is suggested that the oxidative damage produced by SO2inhalation may influence or promote the progression or occurrence of some disease states of various organs, Further work is required to understand the toxicological role of SO2on multiple or even all organs in mammals.

REFERANCES

Ahmad B (2004):Pharmacology of insulin.Br J Diabetes Vasc Dis,4:10-14.Publisher Full Text.

- Arts JH,Jacobs EJ,Kuper CF (2010):Pre-exposure to sulfur dioxide attenuates most allergic reactions upon trimellitic anhydride challenge in sensitized Brown Norway rats. Inhal Toxicol.Feb;22(3):179-91.

-Bates, D.V et al., (1970): Can Med. Ass.J., 103, 833-837. Combined effect of ozone R sulphur dioxide on human pulmonary function. Quoted from nature vol.257, September 4:50-51.

- Bushtueva, K.A. (1966): The toxicity of oxidase of sulphur with prolonged and continuous action. The biological action and hygienic importance of atmospheric contaminants Meditsina: Moscow 142-172.

-Chandrasekhar, Gurramkonda,Sulena Polez,Natasa Skoko,Ahmad Adnan,Thomas Gäbel,Dipti Chugh,Sathyamangalam Swaminathan,Navin Khanna,Sergio TisminetzkyandUrsula Rinas(2010) : UsingPichia pastoriswith subsequent purification and conversion to human insulin. Microbial Cell Factories,9:31doi:10.1186/1475-2859-9-31.

-David, K.J. Nelson, A.A., Z. Wickey, E.R., Hansen, W.H. Fitzhuch , O.G. (1966): Chronic toxicity of ponceau S. 2. (5-sulfo-2,4-xylylazol)-1-napththol-4-sulfonic acid to rats, mice 8 dogs.Toxical Appl Pharmacol 8 (2): 306-317.

- Ferrer-Miralles N, Domingo-Espin J, Corchero JL, Vazquez E, Villaverde A (2009):Microbial factories for recombinant pharmaceuticals.Microb Cell Fact ,8:17.PubMed Abstract|BioMed Central Full Text|PubMed Central Full Text.

-Hazucha, M, Silverman F, Perent C, Field S. Bates D.V. (1973): Pulmonary functionin man after short term exposure to ozone. Archs Environm. Hlth, 27, 183-188.

- King R.N andKing E.J (1954): Estimation of plasma phosphatase by determination of hydrolysed phenol with amino-antipyine. J.Cin. Path., 7:322-326.

-Kjeldsen T, Pettersson AF, Hach M (1999):Secretory expression and characterization of insulin inPichia pastoris.Biotechnol Appl Biochem ,29:79-86.PubMed Abstract|Publisher Full Text.

-Meng Z. (2003): Oxidative damage of sulfur dioxide on various organs of mice: sulfur dioxide is a systemic oxidative damage agent.Inhal Toxicol.15(2):181-95.

-Meng Z,Liu Y,Wu D. (2005):Effect of sulfur dioxide inhalation on cytokine levels in lungs and serum of mice. Inhal Toxicol.17(6):303-7.

-Qin G , and Meng Z( 2005): Effect of sulfur dioxide inhalation on CYP1A1 and CYP1A2 in rat liver and lung. Toxicol Lett. 30; 160(1):34-42.

-Qin G, Meng Z( 2006):The expressions of proto-oncogene and CYP1A in lungs of rats exposed to sulfur dioxide and benzo(a)prune. Regul Toxicol Pharmacol. 2006 Jun; 45(1):36-43.

-Vajo Z, Fawcett J, Duckworth WC (2001):Recombinant DNA technology in the treatment of diabetes: insulin analogs.Endocr Rev,22:706-717.PubMed Abstract|Publisher Full Text.

- Trinder, P.(1969): Ann. Clin. Biochem., 6,24 .

-Walsh G (2005) :Therapeutic insulins and their large-scale manufacture.Appl Microbiol Biotechnol,67:151-159.PubMed Abstract|Publisher Full Text.

-WHO Document-WHO/NCD/NCS/99.2 (1999): Definition, diagnosis and classification of diabetes mellitus and itscomplications[webciteWorld Health Organization.