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Effects of Simultaneous Exposure to Formaldehyde Vapor and Noise on Mouse Testicular Tissue and Sperm Parameters


1 Department of Occupational and Environmental Health, School of Medical Sciences, Tarbiat Modares University, Tehran, IR Iran
2 Department of Anatomical Sciences, School of Medical Sciences, Tarbiat Modares University, Tehran, IR Iran
*Corresponding author: Ali Khavanin, Corresponding author: Ali Khavanin, Jalale-Ale-Ahmad Highway, Tarbiat Modares University, Faculty of Medical Sciences, Department of Occupational and Environmental Health, P.O. Box 14115-175, Tehran, IR Iran, Tel.: +98- 2182883825, Fax: +98-2182883825, E-mail:, E-mail: [email protected].
Health Scope. 2012 September; 1(3): 110-117. , DOI: 10.17795/jhealthscope-7973
Article Type: Research Article; Received: Aug 28, 2012; Revised: Sep 1, 2012; Accepted: Sep 7, 2012; epub: Nov 22, 2012; ppub: Sep 23, 2012
Running Title: Effects of Formaldehyde and Noise on Male Mice Reproductive Parameters

Abstract


Background: Workers are often simultaneously exposed to formaldehyde and noise in workplaces, and noise can reinforce the harmful effects of some chemical pollutants.

Objectives: This study aimed to investigate the effects of simultaneous formaldehyde and noise exposure on reproductive parameters in male mice.

Materials and Methods: Forty eight National Medical Research Institute adult male mice were randomly assigned to control and experimental groups which were exposed to formaldehyde (10 ppm) (F), noise (100 dB) (N), and simultaneous formaldehyde with noise (NF), respectively for 10 days (8 hours a day). Animals were killed 24 hours following exposure (short-term effects) and the remainder was killed 35 days after the end of exposure (long-term effects). Sperm was analyzed by a computer aided sperm analysis system and histological changes in the testis were determined. Plasma concentrations of testosterone, LH, FSH were measured.

Results: The results of the short-term analysis showed that serum testosterone in all exposure groups decreased significantly compared to the controls (P < 0.001). In the long-term analysis, the percentages of progressive motile sperm groups N, F, and NF were 33.73 ± 3.4, 26.65 ± 1.61, and 14.78 ± 4.65, respectively, which indicated a significant decrease (P < 0 .05) compared to the control group (44.47 ± 2.88). The progressive motile sperm in the simultaneous exposure group was less than the N and F groups (P < 0.001). Histological study of testes specimens in the experimental groups (F and NF) revealed displacement of germinal cells and degeneration of Leydig cells and seminiferous tubules.

Conclusions: Exposure to formaldehyde vapor can destroy testicular structure and decrease percentages of sperm count and progressive motility. The present study indicated that a reduction in the percentage of progressive motile sperm may be potentiated by noise (700-5700 HZ) in the simultaneous exposure to formaldehyde and noise group. Thus, it can be anticipated that simultaneous exposure to formaldehyde and noise in workplaces can increase the possibility of damage to testicular structures and reproductive functions in humans, if the same effects transpire.

Keywords: Formaldehyde; Noise; Gonadal Steroid Hormones; Mice

1. Background


Formaldehyde (H2CO) is a colorless, flammable, chemical precursor, which is widely used in various industries. The World Health Organization (WHO) has defined 36 industries in which workers are exposed to formaldehyde (1). Formaldehyde is considered to be an economically important substance. The production of this material has increased from 23.21 million tons in 2000, to 36 million tons in 2007, which is a 70% increase during seven years (2). Industries or occupations with significant formaldehyde exposure include; medical specialties (coroners, hospital housekeeping staff, and laboratory workers), embalmers, industrial workers (formaldehyde synthesis, molding compounds, decorative laminates, plastic moldings and photographic films), textile and wood workers (plywood, particle board and furniture) (2).


<p>Impairments resulting from high noise levels have been reported as one of the ten most hazardous occupational factors. Based on research and statistics in the United States, 7.4-10.2 million workers are exposed to noise stress (3). In Europe, more than 26% of the population are exposed to environmental noise levels higher than 65 dB (4). The harmful effects of formaldehyde in the air have been well documented for the respiratory system, these include; nasal squamous cell carcinoma and mutagenicity (5). However, its effects on other systems and organs are still being studied. Chowdhury et al. (6) noted an inhibition of steroidogenesis, disruption of Leydig cells and spermatogenesis arrest, that resulted from intraperitoneal injections of formaldehyde at dosages of 5, 10 and 15 mg/kg in rats. The negative impact of formaldehyde exposure and sperm parameters has been investigated in several studies (7, 89). The results revealed atrophy of the seminiferous tubules, decreased sperm count and sperm progressive motility in the epididymis. It was shown that exposure to formaldehyde increased the secretion of corticotrophin releasing hormone (CRH) from the hypothalamus and caused corticosteroid adrenal release which decreased testosterone hormone by affecting the hypothalamus-pituitary-testis (HPT) axis (10).


<p>Noise is one of the hazardous factors in the workplace that activates nerve-hormone tracts and consequently increases catecholamine, glucocorticoids and cortisol plasma (11). Decreases in testosterone hormone levels were found in adult male rats exposed to noise (100 dB, 1000 HZ) for a period of 60 days (3 hours a day) (12). Furthermore, Fathollahi, et al. (13) exposed rats to noise (90-130 dB) for 50 days (12 hours a day). They reported a significant decrease in testosterone, FSH, and LH levels, as well as a negative impact on fertility in rats.

2. Objectives


So far, several studies have investigated the effects of formaldehyde and noise on the reproductive system in animal models (14). However, there is inadequate information concerning their toxic effects on reproductive system functioning. Thus, the present research attempts to examine the effects of simultaneous formaldehyde and noise exposure on the means of seminiferous tubular diameters (STD), gonadotropic hormonal changes and sperm parameters in mice.

3. Materials and Methods


3.1. Animals and Experimental Design


<p>Group C: Control group which were in an experimental condition, but were not exposed to formaldehyde or noise (n = 12).


<p>Group N: Animals that were exposed to noise (n = 12).


<p>Group F: Animals that were exposed to formaldehyde (10 ppm) (n = 12).


<p>Group NF: Animals that were exposed to formaldehyde (10 ppm) and noise simultaneously (n = 12).


15) and this was measured and monitored four times an hour, by a photo ionization detector (Photocheck + 5000, Ionscience Co., UK). Precision of the measurement was controlled according to the 3500 method recommended by the National Institute of Occupational Health and Safety (NIOSH) (15, 16).


17, 1819


3.2. Epididymal Sperm Preparation and Sperm Quality Evaluation

202021). Evaluation of live (unstained) and dead (red stained) spermatozoa was carried out using light microscope at 400x magnification.


3.3. Histological Analysis and Morphometric Technique

22). Sections were viewed and photographed by a microscope (Magnum-3, Ceti, England) with an attached camera (Sony-DSC-H50). An estimate of seminiferous tubular diameters (STD) was performed by examining 20 fields in five histological sections from each testis (22), using digitalized microscopic images (10X) with software Image Tools 3.0 (http://compdent.uthscsa.edu/dig/itdesc.html).


3.4. Hormone Assay

23


3.5. Statistical Analysis

4. Results


4.1. Sperm Parameters

The results obtained from the short-term analysis of the physiological parameters of epididymis sperm (24 hours after exposure) did not show any significant differences in the N, F, and NF experimental groups in term of sperm count, sperm motility and percentage of sperm viability in comparison with the control group (Table 1). The results of the study in the long-term analysis indicated that the mean of sperm viability in the control group was



Table 1Table 1).


4.2. Testicular Histopathology

Histological examination of the test showed numerous structural changes in the F and NF experimental groups (long-term analysis) in comparison with the control. The main pathological changes included; seminiferous tubule atrophy, increases in the spaces between germ cells, degeneration of Leydig cells, disintegration of seminiferous epithelial cells and degeneration of a number of seminiferous tubules. No histological changes were seen in the control specimens (Figure 1 A-D). The morphometric findings indicated that the mean of seminiferous tubular diameters (STD) decreased significantly in both the short (P < 0.05) and long-term (P < 0.001) analysis in the experimental groups when compared with the control group (Table 2).




4.3. Gonadotropic Hormonal

The short-term and long-term analysis of mean values for serum levels of testosterone, LH, FSH from all subjects in both the control and exposure groups are shown in Table 3. Our data in the short-term analysis indicated that testosterone declined significantly in all experimental groups, N, F, and NF (P < 0.001), similarly, serum levels of LH, were significantly reduced in the exposure groups F and NF, compared with the control group (P < 0.05, P < 0.001). Also, a comparison of the serum levels of FSH in the exposure and control groups indicated that there was a statistically significant decrease only in the simultaneous exposure group NF (P < 0.05). In the long-term analysis, mean values for testosterone, LH, and FSH hormone levels in the experimental groups increased in comparison to the same groups in the short-term analysis. The means of these hormones with exposure to noise, and exposure to formaldehyde groups, were not statistically significant compared to the control group (P > 0.05). Moreover, while the level of testosterone hormone in the simultaneous exposure group in the short-term analysis had increased, there was a significant difference observed in comparison with the control group (P < 0.05).


5. Discussion


The present study has shown that there is an inhibitory effect of formaldehyde vapor on mouse sperm resulting in decreased sperm motility after formaldehyde vapor exposure is potentiated by simultaneous noise exposure. The effects of formaldehyde on male fertility potential in some mammals has been investigated previously, but to our knowledge there are few reports on the effects of simultaneous formaldehyde and noise exposure on sperm motility in short (one day) and long duration (35 days) exposure times. The results of the long-term analysis revealed that formaldehyde vapor alone, or in combination with noise exposure, decreased progressive motile sperm in mice. In fact, our results involve novel information about the effect that simultaneous formaldehyde and noise exposure has on sperm progressive motility, in regard to the detrimental reinforced effects of noise on sperm motility in mice (24, 25).


A study by Zhou et al. (8) described exposure to formaldehyde vapor (10 mg/m3 for two weeks) that led to a decrease in the epididymis sperm count and motility in rats, this study also showed that the activity of enzymatic antioxidants decreased significantly in the testes of rats exposed to formaldehyde inhalation compared to the control group. One possibility for the reduction in sperm motility is that formaldehyde may cross the blood barrier thus inducing oxidative stress by increasing reactive oxygen species or decreasing antioxidant activity within the luminal surfaces. In a study by Kose et al. (26


The findings obtained from the long-term sperm analysis in our study are compatible with the results of the above studies. The present research showed that formaldehyde vapor decreased sperm progressive motility. According to Mazilli, et al. (27) non-progressive, immotile and abnormal sperm can produce anion superoxidase, which is an oxidative factor in itself and this can decrease sperm quality, including sperm motility. In the current study, animals sacrificed 24 hours after formaldehyde exposure showed slight histopathological changes in the testes tissues. In contrast, the results of a histological examination in the F and NF experimental groups (long-term analysis) revealed that seminiferous epithelial cell degeneration of some seminiferous tubules had occurred, increasing the inter-tubular spaces and reducing the means of the seminiferous tubular diameter (STD). Ozen et al., (7) revealed that sub-chronic exposure to formaldehyde (5-10 ppm) for 91 days caused significant reductions in tubular diameters (P < 0.001). The experimental study of Golalipour et al., (28) also showed that formaldehyde vapor exposure for 18 weeks on rats, can induce histological changes in seminiferous tubules due to a significant decrease in the STD mean.


Zhou et al., (8) described how exposure to formaldehyde vapor (10 mg/m3 for two weeks) led to the atrophication of seminiferous tubules, decreasing numbers of spermatogenesis cells and seminiferous epithelial cell disintegration. With regard to the findings of our study, formaldehyde vapor alone and simultaneous exposure at the concentration levels and duration previously mentioned, are able to cause histological changes in the seminiferous epithelium and decrease the mean of seminiferous tubular diameters (STD) in mice. The morphometric findings obtained from the present study are in line with the findings of Golalipour et al. (28). In a study by Kose et al., (26) about the effect of formaldehyde on rat reproductive systems, experimental animals were exposed to formaldehyde vapor (10 ppm/1h) for 35 days and the detrimental effects of formaldehyde on sperm count, motility and normal morphology were observed. They also reported a relationship between the reductions of the seminiferous tubular diameters and a decrease in the number of Leydig cells in rats. Furthermore, Henkel et al., (29) determined a direct correlation between sperm motility and a decrease in the number of Leydig cells.


According to a study by Tang et al., (8) an intraperitoneal injection of formaldehyde with dosages of 0.2,2 and 20 (mg/Kg) have a negative impact on sperm count, viability and sperm motility in rats. They reported that seminiferous tubules atrophy and degeneration of seminiferous tubules led to a reduction in sperm counts. The findings obtained from the long-term sperm analysis in our study are compatible with the results of the above studies. Furthermore, the results of the long- term analysis (35 days after the end of exposure) revealed that there was a significant difference between the experimental groups F and NF, in terms of progressive motile sperm. Another explanation regarding the findings of our study (decrease in motility and sperm viability), may be that noise and formaldehyde vapor exposure has an effect on the hypothalamus-pituitary-testis (HPT) axis in mice, which leads to a decrease in the levels of sex androgen hormones. In addition, our study showed that a decrease in testosterone hormone levels occurred in all experimental groups compared to the control.


It was shown that chemical and physical stress could increase the level of glucocorticoid hormones such as cortisol (30, 31), which directly or indirectly inhibit secretion of sex hormones and gonadotropin releasing hormones (10-13). The findings of the researchers obtained from the experiments performed on different animal models have indicated that increasing cortisol caused a decrease in secretion pulses GnRH and LH in addition to decreases in testis endocrine function (32). On the other hand, formaldehyde and noise may, via two specific mechanisms, have an effect on spermatogenesis and sperm parameters; first by changing the composition and structure of seminiferous tubules and dysfunction in the spermatogenesis process, and second by influencing the hypothalamic-hypophyseal-gonadal axis. In this regard Sari et al. (10) revealed that chronic exposure to low concentrations of formaldehyde (82-1874 ppb) affected the hypothalamus-hypophyseal-adrenal (HPA) axis and increased the secretion of corticotrophin releasing hormones (CRH) consequently this increased adrenal corticosteroid. Moreover, in the study by Sorg et al, (31) it was confirmed that corticosterone levels increased in rats, due to the effects of frequent exposure to low concentrations of formaldehyde. Furthermore in agreement with our results, Swami et al. (12) reported changes in testis tissue and a decrease in serum testosterone in rats exposed to 100 dB noise. Fathollahi et al. (13) observed that testosterone hormones decreased in rats which were exposed to 90-130 dB noise. As a result of the close relationship between the sub-cortical structure of the central nervous system and the auditory system in mice, noise could have an influence on the hypothalamus-hypophyseal-adrenal system through activation of this axis by the corticotrophin releasing hormone (30).


With respect to the findings of our study, sub-acute exposure to high concentrations of formaldehyde vapor in the workplace, may lead to a decrease in testosterone and LH hormone levels and noise can potentiate these reduction levels. Our study also shows that simultaneous exposure to formaldehyde and noise can decrease the percentage of progressive motile sperms, along with the percentage of sperm viability. However, further investigation needs to be conducted in order to understand the mechanisms underlying the effects of gonadotropin secretion suppression on sperm velocity parameters and sperm swimming patterns.


Acknowledgments

This research was supported by financial aid from Tarbiat Modares University, Tehran, Iran. The authors would like to gratefully thank the Vice President of the Research Department of Tarbiat Modares University for their Contribution to this research.

Footnotes

Implication for health policy/practice/research/medical education: This article has focused on investigating the effects of simultaneous formaldehyde and noise exposure on reproductive parameters in male mice.
Please cite this paper as: Vosoughi S, Khavanin A, Salehnia M, Asilian Mahabadi H, Soleimanian A. Effects of Simultaneous Exposure to Formaldehyde Vapour and Noise on Mouse Testicular Tissue and Sperm Parameters. Health Scope. 2012; 1 (3): 110-7.
Authors’ Contribution: All authors have been worked equally.
Financial Disclosure : None declared.
Funding/Support: None declared.

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Table 1


Experimenta Sperm Count, 106/ml Progressive, Motility (%) Non-Progressive, Motility (%) Immotile, % Sperm Viability, %
Short-term analysis
Control noise
Formaldehyde
Noise-formaldehyde
Long-term analysis
Control noise
a, e a, d a, d a, e a, d
Formaldehyde b, e b, e b, e b, e
Noise-formaldehyde c, e c, e c, e c, d
a Difference between experimental group (N) and control group
c Difference between experimental group (NF) and control group
c Difference between experimental group (NF) and control group
b Difference between experimental group (F) and control group
c Difference between experimental group (NF) and control group

Table 2


P-Value, Versus With control group P-Value, Versus With control group
C
N 0.354 < 0.05
F < 0.05 < 0.001
NF < 0.05 < 0.001

Table 3


P-Value, (Versus With control group) P-Value, Versus With control group
Testosterone Testosterone
C
N < 0.001 0.337
F < 0.001 0.134
NF < 0.001 < 0.05
Luteinizing hormone (LH) Luteinizing hormone (LH)
C
N 0.795 0.984
F < 0.05 0.584
NF < 0.001 0.346
Follicle stimulation hormone(FSH) Follicle stimulation hormone(FSH)
C
N 1.000 0.999
F 0.603 0.793
NF < 0.05 0.490