The role of matrix metalloproteinases 2 and 9 in obstructive sleep apnea

Patients and methods Th e present study was conducted during the period from May 2011 to May 2012; it included 30 obese patients who had been referred to the Chest Department of Kasr Al-Aini Hospital for clinical suspicion of OSA to perform polysomnography. Th e included patients were classifi ed into two groups: (1) Cases: it consisted of 20 obese patients (BMI>30) who were diagnosed with OSA on the basis of both clinical and polysomnographic criteria (AHI≥5 events/h). (2) Controls: it consisted of 10 obese (BMI>30) healthy individuals free from any known diseases without OSA (Apnoea Hypopnea Index (AHI)<5 events/h).


Introduction
In recent years, obstructive sleep apnea (OSA) has emerged as a major public health problem because of its profound impact on patients' health and quality of life [1].
Obesity is one of the most important risk factors for sleep-disordered breathing [2].
Production of matrix metalloproteinase (MMP) 9 is stimulated by hypoxia and by several cytokines, such as IL-6 and TNF-a. Th ese cytokines are increased and hypoxia is induced by apnea and hypopnea during sleep in patients with OSA [3].

Aim
Th e aim of the study was to evaluate serum levels of MMP-2 and MMP-9 as markers of oxidative stress in obese patients with OSA.

Patients and methods
Th e present study was conducted during the period from May 2011 to May 2012; it included 30 obese patients who had been referred to the Chest Department of Kasr Al-Aini Hospital for clinical suspicion of OSA to perform polysomnography.
Th e included patients were classifi ed into two groups: (1) Cases: it consisted of 20 obese patients (BMI>30) who were diagnosed with OSA on the basis of both clinical and polysomnographic criteria (AHI≥5 events/h). (2) Controls: it consisted of 10 obese (BMI>30) healthy individuals free from any known diseases without OSA (Apnoea Hypopnea Index (AHI)<5 events/h).
All study participants were subjected to the following: (7) Measurement of serum levels of MMP-2 and 9: (a) All participants went to sleep at 9:00 p.m. and were awakened at 5:00 a.m.; samples of peripheral venous blood were collected at 5:00 a.m. and were stored at -80°C until assay. (8) Quantitation of human MMP-2 levels in serum: (a) Human MMP-2 levels were measured in serum using Quantikine ELISA Kit (catalog no. DMP2F0; R&D System Inc., Minneapolis, Minnesota, USA). (9) Quantitation of human MMP-9 levels in serum: Quantitation of MMP-9 levels in serum was performed using Quantikine ELISA Kit (catalog no. DMP900; R&D System Inc.).

Results
Th e results are shown in Tables 1-13.  Table 11 shows a positive correlation between AHI, ESS, BMI, and MMP-2 and MMP-9, but it was found to be statistically insignifi cant. Table 12 shows positive signifi cant correlation between MMP-2 and MMP-9 among patients with mild Obstructive Sleep Apnoea Syndrome (OSAS). In addition, there was a positive correlation between AHI, ESS, BMI, and MMP-2 and MMP-9, but it was found to be statistically insignifi cant. Table 13 shows positive signifi cant correlation between MMP-2 and MMP-9 among patients with moderate to severe OSAS. In addition, there was a positive correlation between AHI, ESS, BMI, and MMP-2 and MMP-9, but it was found to be statistically insignifi cant.

Discussion
In recent years, OSA has emerged as a major public health problem because of its profound impact on patients' health and quality of life [1].
Obesity is one of the most important risk factors for sleep-disordered breathing [2].
In addition, Nagayoshi et al. [7] reported that several research studies had repeatedly and consistently confi rmed that OSA is more common in male patients than in female patients and that male-to-female ratio is estimated to be ∼2 : 1 in the general population, supporting our result.
In the present study, the mean age of patients was 51.40 ± 11.68 years, which was higher than that of controls (mean age 43.30 ± 14.96 years) ( Table 2). Th e diff erence between the two groups was statistically insignifi cant.
Th is is in agreement with the study by Peppard et al. [8] who reported an increase in the prevalence of OSA with age, which could not be explained by other risk factors such as obesity.   reduced pharyngeal size, changes in upper airway muscle function, and respiratory instability. Among postmenopausal women, the reductions in the circulating levels of sex hormones and pharyngeal lengthening contribute to increased risk for OSA [9].
In the current study, the BMI mean value among patients was 43.45 ± 9.24, denoting that most our patients were considered class III obesity (Table 3).
Th is is in agreement with the study by Shelton et al. [10] who stated that obesity is believed to predispose to OSA because of mass loading to the upper airway of the neck.
In addition, Strohi and Redline [11] stated that excess body weight is a major risk factor for snoring and Several mechanisms have been proposed to account for the increasing prevalence of OSA with increasing age, such as changes in pharyngeal mechanics,    report morning headache. However, McNicholas and Bonsignore [17] stated that no systematic study has been undertaken to analyze whether morning headache has the potential to predict the presence or absence of OSA. Th erefore, the utility of this symptom in the objective clinical assessment of patients with suspected OSA remains uncertain.
In the present study, there was statistically highly signifi cant increase in AHI, desaturation index, duration of desaturation less than 90%, and minimal O 2 sat% among patients compared with controls. However, average O 2 sat% was lower in patients than in controls, and this reduction was statistically signifi cant ( Table 7).
Th is is in agreement with results obtained by Kaynak et al. [18] who demonstrated that the minimal oxygen saturation point was statistically lower in patients with OSA than in controls.
In addition, Nakagawa et al. [19] studied 93 patients with OSA and 18 controls; they found that there was statistically signifi cant diff erence between patients and controls in AHI, desaturation index, and duration of O 2 desaturation less than 90%.
Th e mean value of FEF25-75% pred among patients was 56.28 ± 23.52, and among controls the mean value was 56.60 ± 20.42. Th e diff erence between patients and controls regarding all spirometric data was statistically insignifi cant. Th is may be attributed to the fact that both groups were obese and they were matched regarding BMI (Table 8).
Th is is in agreement with the study by Biring et al. [20] who found that obesity leads to limitations in airfl ow, with reduction in both FEV 1 and FVC with the FEV 1 / FVC ratio remaining unchanged. Some authors have found a restrictive model in obese patients with an increased FEV 1 /FVC ratio.
In addition, Canoy et al. [21] reported that obese patients are prone to have reduced FEV 1 , FVC, and total lung capacity in lung function tests and they usually present with restrictive lung patterns, and they found that the possible mechanisms for the abnormal lung function tests in obese patients were reduced chest wall compliance and increased peripheral airway resistance.
Comparison between patients and controls regarding arterial blood gases revealed that the mean value of PO 2 among patients was 71.40 ± 14.81, which was lower compared with controls (mean value was 79.30 ± 12.89). For O 2 sat%, the mean value among patients was 93.70 ± 2.53, which was also lower as compared with controls (95.80 ± 2.53). Th e diff erence in PO 2 and sleep apnea and that 70% of patients with OSAS are overweight.
However, the mean value of BMI among controls was 45.12 ± 4.09, which was higher than in patients, but it did not reach any statistical signifi cance (Table 3).
ESS is a validated tool for the systematic assessment of impaired daytime alertness. ESS is applied frequently because of its simplicity in routine practice, especially to describe sleepiness of patients with OSA. ESS was constructed to measure the patient's ability to remain awake or the propensity to doze off in typical daily situations [4].
In this study, it was found that all 20 (100%) patients had daytime sleepiness compared with fi ve (50%) controls who had daytime sleepiness and fi ve (50%) controls who had not (Table 4).
Th e mean value of ESS among patients was 19.80 ± 5.24 compared with controls who had the mean value of 7.60 ± 2.07. Th is increase in daytime sleepiness and ESS among patients compared with controls was statistically highly signifi cant (Table 5).
Th is is in agreement with the study by Banamah [12] who studied 27 patients with OSA and 26 obese individuals without OSA as a control group; he found highly signifi cant increase in ESS among patients with OSA in comparison with controls.
In the current study, we found that, among patients 18 (90%) had snoring, whereas among controls nine (90%) had snoring, with no statistically signifi cant diff erence (Table 10).
Th is is in agreement with the study by Gottlieb et al. [13]; they found that snoring has poor predictive value for OSA owing to a high prevalence in the general population.
However, Viner et al. [14] stated that snoring is a hallmark of OSA, and in its absence the diagnosis of OSA is unlikely.
Morning headache is a less common manifestation of sleep apnea. If reported, one must consider the possibility of hypercapnia secondary to obesity hypoventilation syndrome [15].
In the present study it was found that among patients 13 (65%) had morning headache and among controls four (40%) had morning headache. Th is diff erence was not statistically signifi cant (Table 6).
Th is is not in agreement with the study by Kiely et al. [16] who stated that patients with OSA often Th is is not in agreement with the study by Toshiyuki et al. [24] who stated that in patients with OSAS the levels and activity of MMP-9 were positively correlated with BMI.
In the present study, MMP-2 also was higher in patients than in controls. However, the diff erences were statistically insignifi cant.

Conclusion
Obesity is considered a major risk for OSA, and it is associated with local adipose tissue hypoxia and adipose tissue dysfunction.
OSA is associated with chronic intermittent hypoxia resulting in hypoxia, oxidative stress, and production of oxygen free radicals.
Th e current study highlights the eff ect of OSA on levels of certain oxidative markers.
It was concluded that serum MMP-9 was signifi cantly higher in obese patients with OSA than in obese individuals without OSA.
Th ese markers could be useful as prognostic factors to assess the response following CPAP treatment or Bariatric surgery; however, further studies still needed to confi rm this fact.
O 2 sat% may be related to age diff erences between the two groups (Table 9).
Th is is in agreement with the study by Zhang et al. [22]; they demonstrated that aging is associated with both hypoxia and increases in reactive oxygen species in aging men.
Several studies indicated that OSAS-induced hypoxic stress activates the production of infl ammatory mediators by monocytes such as MMP-9 and TNF-a, and this phenomenon may contribute to the development of atherosclerosis. Th erefore, it is suggested that Continuous Positive Airway Pressure (CPAP) treatment could play a role in the prevention of atherosclerosis in OSAS patients [23].
In a comparison between patients and controls regarding serum levels of MMP-9, it was found that the mean value of MMP-9 among patients was 169.57 ± 135.22, which was higher than in controls, as mean value was 87.29 ± 43.01, and the diff erence was statistically signifi cant (Table 10).
Th ese results are in agreement with those of Toshiyuki et al. [24] who demonstrated that serum levels of MMP-9 were signifi cantly higher in all patients with OSAS than in obese controls and that the serum levels of MMP-9 were signifi cantly higher in patients with moderate to severe OSAS than in patients with mild OSAS.
Th is result is in agreement with the result of Jin et al. [25] who reported that serum concentration of MMP-9 was signifi cantly higher in patients with OSAS than in obese controls. Levels of MMP-9 were also signifi cantly higher in moderate to severe OSAS than in the mild OSAS group or obese controls.
It was found to be in agreement also with the study by Shinji et al. [23] who reported that, in OSAS patients, the production of MMP-9 by monocytes was signifi cantly elevated after sleep in the early morning than in controls, and was decreased after long-term CPAP treatment. In addition, the production of MMP-9 by monocytes is attributed to OSAS-induced hypoxic stress.
In the present study, there was a positive correlation between AHI and MMP-9, but it was statistically insignifi cant and this may be explained by the small number of patients.
Jin et al. [25] reported that serum concentration of MMP-9 was positively correlated with BMI in OSAS patients.
In our study, there was a positive correlation between MMP-9 and BMI but was statistically insignifi cant (Table 11).