결핵성흉막염환자에서흉수내 Matrix Metalloproteinases 및 Tissue Inhibitors of Metalloproteinases 농도와잔여흉막비후와의관계 1 가천의과학대학교의학전문대학원, 2 가천의과학대학교길병원호흡기내과최영권 1, 안창혁 2, 김유진 2, 경선영 2, 이상표 2, 박정웅 2, 정성환 2 The Relation of Residual Pleural Thickening with Matrix Metalloproteinases and Tissue Inhibitors of Metalloproteinases of Pleural Effusion in Patients with Tuberculous Pleuritis Youngkwon Choi 1, Chang Hyeok An, M.D. 2, Yu Jin Kim, M.D. 2, Sun Young Kyung, M.D. 2, Sang Pyo Lee, M.D. 2, Jeong Woong Park, M.D. 2, Sung Hwan Jeong, M.D. 2 1 Graduate School of Medicine, Gachon University of Medicine and Science, 2 Division of Pulmonology, Department of Internal Medicine, Gachon University Gil Medical Center, Incheon, Korea Background: Residual pleural thickening (RPT) is the most frequent complication of tuberculous pleurisy (TP), and this can happen despite of administering adequate anti-tuberculous (TB) therapy. Yet there was no definite relation between RPT and other variables. The aim of this study was to examine matrix metalloproteinases (MMPs) and the inhibitors of metalloproteinases (TIMPs) and to identify the factors that can predict the occurrence of RPT. Methods: The patients with newly-detected pleural effusions were prospectively enrolled in this study from January 2004 to June 2005. The levels of MMP-1, -2, -8 and -9, and TIMP-1 and -2 were determined in the serum and pleural fluid by ELISA. The residual pleural thickness was measured at the completion of treatment and at the point of the final follow-up with the chest X-ray films. Results: The study included 39 patients with pleural fluid (PF). Twenty-three had tuberculous effusion, had parapneumonic effusion, had malignant effusion and 2 had transudates. For the 1 patients who completed the anti-tb treatment among the 23 patients with TP, (41%) had RPT and 10 (59%) did not. The level of PF TIMP-1 in the patients with RPT (41,405.9±9,3.3 ng/ml) was significantly higher than that of those patients without RPT (29,134.9±8,801.8) at the completion of treatment (p=0.032). In 13 patients who were followed-up until a mean of 8±5 months after treatment, 2 (15%) had RPT and 11 (85%) did not. The level of PF TIMP-2 in the patients with RPT (34.4±6.5 ng/ml) was lower than that of those patients without RPT (44.4±15.5) at the point of the final follow-up (p=0.038). Conclusion: The residual pleural thickening in TP might be related to the TIMP-1 and TIMP-2 levels in the pleural fluid. (Tuberc Respir Dis 2008;65:-14) Key Words: Matrix metalloproteinase, Pleural effusion, Residual pleural thickening, Tissue inhibitor of metalloproteinase, Tuberculous pleuritis 서 론 Address for correspondence: Chang Hyeok An, M.D. Division of Pulmonology, Department of Internal Medicine, Gachon University Gil Medical Center, 1198, Guwol-dong, Namdong-gu, Incheon 405-60, Korea Phone: 82-32-460-3200, Fax: 82-32-469-4320 E-mail: anch@gilhospital.com Received: Feb. 1, 2008 Accepted: Jul., 2008 기질단백분해효소인 matrix metalloproteinase (MMP) 와이들의길항체인 tissue inhibitor of metalloproteinase (TIMP) 의불균형이나발현이삼출성흉수를발생시키는흉막질환들에따라차이가있다는연구가최근발표되고있다 1-. 여출성흉수에서보다결핵성흉수에서 MMP-9과 TIMP-1의단백질에대한비가더높다는보고가있었고 2, 폐암에의해발생한흉수내의 MMP-9 농도가결핵성흉
YK Choi et al: Relationship between residual pleural thickening and TIMPs 수에서보다높다는보고도있었으나 3, 다른연구에서는결핵성흉수내의 MMP-9 농도가폐암에의해발생한흉수에서보다높다는보고도있었다 5. 또한농흉, 부폐렴성흉수, 결핵성흉수, 악성흉수, 여출성흉수내의 MMP-1, -2, -8, -9와 TIMP-1, -2의농도차이를비교한연구들도있었다 6. 하지만지금까지의연구들은초기진단시흉수내의 MMP들과 TIMP들의농도를측정함으로써흉수를동반한질환들의감별진단에만초점을둔연구들이며, 진단시 MMP와 TIMP 농도가향후잔여흉막비후와예후와연관되는지관찰한연구는없었다. 잔여흉막비후는결핵성흉막염또는부폐렴성흉수후나타날수있는합병증이다. 특히결핵성흉막염에서는치료후에잔여흉막비후를예측할수있는임상적, 검사실적지표들에대한연구는많으나, 확립된예측인자나발병기전, 치료등은아직까지정립되지못한상황이다. 이에본연구자들은삼출성흉수를발생시키는질환의진단시흉수내의 MMP들과 TIMP 들의농도가각질환의감별진단과초기흉수의양과연관관계가있는지알아보는데에그치지않고, 특히결핵성흉막염에서치료후잔여흉막비후가지속되는예측인자가될수있는지알아보고자하였다. 대상및방법 1. 대상흉수를동반한 18세이상의성인을대상으로, 2004년 1월부터 2005 년 6월사이에가천의대길병원호흡기내과에입원한환자중흉수가발견된환자를대상으로전향적연구를시행하였다. 흉수는 Light 기준에따라삼출액과여출액으로구분하여다음과같이네개의군으로분류하였다 8. 결핵성흉수는다음의기준중하나이상을만족하는경우진단하였다 : (1) 흉수에서결핵균이배양된경우, (2) 흉막조직검사에서건락육아종이관찰되는경우, (3) 림프구-우세삼출액이고흉수의 adenosine deaminase>45 IU/L이고항결핵제에치료반응이좋은경우 9. 농흉을포함한부폐렴성흉수는다음의기준중하나이상을만족할경우진단하였다 : (1) 육안적으로농인경우, (2) 그람염색또는배양에서균이검출된경우, (3) 임상및방사선학적으로폐렴이면서흉수가동반된경우 10. 악성흉수는흉수나흉막조직검사에서악성세포가발견되는경우진단하였다. 여출액은 Light 기준상삼출액에해당되지않으면서타 흉막질환의증거가없으며심부전등여출액이발생할수있는질환이동반된경우진단하였다 8. 이전에한번이라도동일한또는다른흉막질환을앓았거나흉강천자를한환자, 흉부수술을받은환자는제외하여, 입원후첫흉강천자로채취한흉수를그분석대상으로하였다. 또한상기네군에해당하지않는환자들도본연구에서제외하였다. 본연구는가천의대길병원임상시험위원회의심의를거쳐승인을받았다. 2. 방법 1) 방사선학적측정내원당시흉수의양은최초의단순흉부방사선촬영에서흉수정점부의위치를기준으로 grade 1에서 6까지 6단계로나누었다 11. 등급 1은흉수가횡격막수준이하인경우로정의하였다. 단순흉부후전위촬영상다음의두조건중하나라도만족하는경우에잔여흉막비후로정의하였다 : (1) 흉골안쪽연에서 2 mm를초과하여흉막이두꺼워진경우, (2) 흉골횡경막각둔화 (costophrenic angle blunting) 가있을때흉골안쪽연에서횡격막까지의두께가 2 mm를초과한경우 12. 흉막의두께는흉골안쪽연에서가장두꺼운부분을측정하였다. 잔여흉막비후의측정은치료종결시와마지막으로추적관찰이가능한시점의측정치를비교분석하였다. 2) 흉수및혈청의분석모든환자에서내원당시각질환의진단및치료에필요한혈액검사와진단적흉강천자, 그리고필요시흉막조직검사를시행하였다. 환자의혈액과흉수는즉시원심분리기를이용하여상청액 (supernatant) 을채취한후 MMPs와 TIMPs 의분석전까지 0 o C에보관하였다. 환자의혈청과흉수내의 MMP-1, -2, -8, -9와 TIMP-1, -2를효소면역측정 (ELISA) 방법 (Amersham Bioscience, Little Chalfont Buckinghamshire, England) 을이용하여측정하였다. 또한흉수의 MMP-2와 MMP-9의경우에는 gelatin zymography 를시행하여그의발현을알아보았는데, 이전연구에서사용한방법을응용하여다음과같이시행하였다 13. 흉수를 10% gelatin 을기질로하여단백질전기영동 (SDS-PAGE) 을시행하였다. 전기영동후 gel을 2.5% Triton X-100에 30분간담가두었다가, 다시 50 mm Tris-HCl, 0.2 M NaCl, 5 mm CaCl 2, 1% Triton X-100을조성으로하는완충제에서 24시간동안 3 o C에서배양하였다. 배양후 gel은 Coo-massie Brilliant Blue R250 (Sigma) 으로염색하였고, 5% methanol 과.5% acetic 8
Table 1. Clinical characteristics of patients with pleural effusions Tuberculosis and Respiratory Diseases Vol. 65. No. 1, Jul. 2008 Groups Tuberculous Parapneumonic Malignant Transudates p-value* Subjects (n) Age (year old) Male:Female PF WBC ( 10 3 /L) Neutrophils (%) Lymphocytes (%) Protein P/S ratio PF LDH (IU/L) PF ADA (U/L) Blood ESR (mm/hr) Serum CRP (mg/dl) 23 40±18 11:12 2,888±2,452 10±14 86±15 0.8±0.1 1,30±653 86.8±18.0 50±21.0±5.3 5±1 5:2 8,81±8,936 68±30 2±33 0.±0.1 2,446±2,349 34.9±20.3 86±19 20.3±14.0 62±15 4:3 1,531±2,098 13±22 81±22 0.±0.1 93±1,046 20.1±14.4 44±1 2.2±1.4 2 66±21 1:1 3±46 13±18 65±32 0.3±0.0 266±14 6.4±1.6 23±2 1.4±0.0 0.010 0.41 0.054 0.00 0.011 0.36 0.046 0.000 0.002 0.006 Data are presented as mean±standard deviation. WBC: white blood cells; PF: pleural fluid; P/S ratio: pleural fluid to serum ratio; LDH: lactate dehydrogenase: ADA; adenosine deaminase; ESR: erythrocyte sedimentation rate; CRP: c-reactive protein. *Kruskal-Wallis test were performed for difference among groups except male:female ratio. acid, 증류수로조성된용액으로탈염색하였다. 그리고 gelatin 가수분해에의하여나타나는 band를관찰하였는데, active MMP-2 는 66 kda, pro-mmp-2 는 2 kda, active MMP-9은 85 kda, pro-mmp-9 은 92 kda의분자량에해당하는 band를찾아관찰하였다. 3. 통계분석통계처리는 SPSS for Windows 12.0 (SPSS Inc., chicago, IL, USA) 을이용하였다. 자료는평균 ± 표준편차로표시하였다. 비모수적통계방법으로, 연속변수의각군간의차이여부는 Kruskal-Wallis 검사를시행하였고, 양군간비교에는 Mann-Whitney U 검사를시행하였으며, 상관관계는 Spearman coefficient of correlation 을이용하여비교분석하였다. 각군간의명목식자료는 Chi-square 검사를시행하였다. 결과 1. 임상적특징가천의대길병원호흡기내과에입원한환자중흉수가발견된환자중제외기준에해당하는환자를제외하고총 39명의환자가대상이되었다. 이중결핵성흉막염은 23명, 부폐렴성흉수 명, 악성흉수 명, 여출액 2명이었고, 각군의환자의임상적특징은다음과같았다 (Table 1). 결핵성흉막염환자들의연령 (40±18세 ) 이부폐렴성흉수 (5±1) 나악성흉수 (62±15) 의환자들의연령보다의 미있게낮았다 (p=0.010). 부폐렴성흉수내의중성구분획 (68±30%) 이결핵성흉수 (10±14) 와악성흉수 (13±22) 보다의미있게높았으나 (p=0.00), 림프구분획 (2±33) 은결핵성흉수 (86±15) 와악성흉수 (81±22) 보다의미있게낮았다 (p=0.011). 결핵성흉막염에서의흉수내 adenosine deaminase (ADA)(86.8±18.0 U/L) 가다른세군 ( 부폐렴성 34.9±20.3; 악성 20.1±14.4; 여출액 6.4±1.6) 에서보다의미있게높았다 (p=0.0001). 혈액검사상적혈구침강속도 (erythrocyte sedimentation rate, ESR) 와 c-reactive protein (CRP) 은부폐렴성흉수를가진환자군이다른세군의환자군보다의미있게높았다 (Table 1). 2. 혈청과흉수내의 MMPs와 TIMPs의 level (Table 2) 흉수내의 MMP-8 은부폐렴성흉수 (8,20.2±12,594.4 ng/ml) 가결핵성 (199.0±443.6) 이나악성 (0.2±0.2), 여출액 (0.0±0.0) 보다의미있게높았다 ( 각각 p=0.004; p=0.002; p=0.040). 결핵성흉수내의 MMP-8 은부폐렴성흉수의 MMP-8 보다낮았으나악성이나여출액흉수내의 MMP-8 보다는의미있게높았다 ( 각각 p=0.0001; p=0.021). 결핵성흉수의 MMP-8 은결핵성흉수내백혈구의다형핵백혈구분획과의미있는양의상관관계를보였다 (p=0.021, r=0.49). 흉수 MMP-1 과 MMP-2 는각군간에의미있는차이는보이지않았으며, gelatin zymography 상에서도흉수의 MMP-2 band 는각군간에유사한관찰소견을보였다 (Figure 1). 흉수 MMP-9는부폐렴성흉수 (383.6±410.0 ng/ml) 가결핵성 (4.0±68.), 악성 (62.1±61.0), 여출성 9
YK Choi et al: Relationship between residual pleural thickening and TIMPs Table 2. Levels of metalloproteinases (MMPs) and tissue inhibitors of metalloproteinase (TIMPs) in serum and pleural effusions Groups Tuberculous Parapneumonic Malignant Transudates p-value* Subjects (n) Serum MMP-1 (ng/ml) MMP-2 (ng/ml) MMP-8 (ng/ml) MMP-9 (ng/ml) TIMP-1 (ng/ml) TIMP-2 (ng/ml) Pleural Effusion MMP-1 (ng/ml) MMP-2 (ng/ml) MMP-8 (ng/ml) MMP-9 (ng/ml) TIMP-1 (ng/ml) TIMP-2 (ng/ml) MMP-1/TIMP-1 ( 10 5 ) MMP-8/TIMP-1 ( 10 3 ) MMP-8/TIMP-2 MMP-9/TIMP-1 ( 10 3 ) MMP-9/MMP-2 ( 10 3 ) 23 5.9±5.8 1,299.6±268. 4.5±114.0 358.8±162.4 935.9±810.8 33.2±3.9 3.9±2.3 12,389.9±5,360.5 199.0±443.6 4.0±68. 35,04.1±12,53.8 41.9±18.8 11.0±5.9 5.±13.1 5.0±11.8 2.2±2.2 6.8±.1.2±8. 1,353.4±19.0 3.1±4.4 811.8±254.2 1,342.2±525. 38.0±8.0 9.6±8.5 8,352.0±2,926.4 8,20.2±12,594.4 383.6±410.0 36,53.9±12,291.1 53.3±23.6 25.5±22.3 22.2±332.6 163.±233.1 10.5±10.4 61.2±58.8 Data are presented as mean±standard deviation. *Kruskal-Wallis test were performed for difference among groups. 4.1±2.8 1,491.9±455.8 0.0±0.0 251.5±100.1 853.0±403.8 32.1±5.9 2.±1. 10,354.6±4,635.6 0.2±0.2 62.1±61.0 21,63.2±9,452. 43.9±20.4 15.4±11.9 0.009±0.012 0.005±0.00 2.±2.6 8.9±11.8 2 5.3±4.9 36.5±0.0 0.0±0.0 282.5±265.2 1,55.9±0.0 50.4±0.0 1.8±0.2 8,323.8±6,205.4 0.0±0.0 40.0±42.4 12,52.±2,658.0 52.3±.3 14.9±4.9 0.002±0.002 0.000±0.0003 2.9±2.8 4.0±2.1 0.941 0.308 0.001 0.003 0.155 0.144 0.100 0.238 0.000 0.10 0.008 0.411 0.26 0.000 0.000 0.131 0.094 Figure 1. Gelatin zymograms of pleural effusions from parapneumonic effusions, tuberculous pleuritis, malignanteffusions and transudates. Gelatinolytic bands of 92 kd (prommp-9), 85 kd (active MMP-9), 2 kd (prommp-2), 66 kd (active MMP-2) were detected. Intense lytic bands corresponding to the MMP-9 were visible in parapneumonic effusion. MMP: metalloproteinase. (40.0±42.4) 보다높은경향을보였으나통계적의미는보이지않았다 (p=0.10). 하지만 gelatin zymography 상에서는부폐렴성흉수에서의 MMP-9 band 가다른세군에서보다두드러지게관찰되었다 (Figure 1). 흉수 MMP-9/ MMP-2 비 ( 10 3 ) 는부폐렴성흉수 (61.2±58.8) 가결핵성 (6.8±.1), 악성 (8.9±11.8), 여출액 (4.0±2.1) 보다높은경 향을보였으나통계적의미는보이지않았다 (p=0.094). 흉수 TIMP-1은결핵성흉수 (35,04.1±12,53.8 ng/ml) 가악성 (21,63.2±9,452.) 이나여출액 (12,52.±2,658±0) 보다의미있게높았다 ( 각각 p=0.006; p=0.021). 흉수 TIMP-2 는각군간에의미있는차이를보이지않았다. 흉수 MMP-8/TIMP-1 비 ( 10 3 ) 는부폐렴성흉수 (22.2±332.6) 가 10
Tuberculosis and Respiratory Diseases Vol. 65. No. 1, Jul. 2008 Figure 2. Pleural fluid TIMP-1 levels according to the RPT in tuberculous pleuritis. The level of TIMP-1 in patients with RPT was significantly higher than that of those without RPT at the completion of treatment. Data are presented as box-plots, where the horizontal line represents the mean. TIMP: tissue inhibitor of metalloproteinase; RPT: residual pleural thickening. Figure 3. Pleural fluid TIMP-2 levels according to the RPT in tuberculous pleuritis. The level of TIMP-2 in patients with RPT was lower than that of those without RPT at the point of final follow-up. The mean follow-up period was 8±5 months after treatment. Data are presented as box-plots, where the horizontal line represents the mean. TIMP: tissue inhibitor of metalloproteinase; RPT: residual pleural thickening. 결핵성 (5.±13.1), 악성 (0.009±0.012), 여출액 (0.002±0.002) 보다의미있게높았으며 ( 각각 p=0.001; p=0.002; p=0.04), 결핵성흉수가악성과여출액보다의미있게높았다 ( 각각 p=0.0001; p=0.021). 흉수 MMP-8/TIMP-2 비는부폐렴성흉수 (163.±233.1) 가결핵성 (5.0±11.8), 악성 (0.005±0.00), 여출액 (0.000±0.0003) 보다의미있게높았으며 ( 각각 p=0.003; p=0.002; p=0.04), 결핵성흉수가악성과여출액보다의미있게높았다 ( 각각 p=0.0001; p=0.021). 흉수 MMP-9/TIMP-1 비 ( 10 3 ) 는부폐렴성흉수 (10.5±10.4) 가결핵성 (2.2±2.2), 악성 (2.±2.6), 여출액 (2.9±2.8) 보다높은경향을보였으나통계적의미는보이지않았다 (p=0.131). 3. 결핵성흉막염에서흉수 MMPs 및 TIMPs 와잔여흉막비후의관계 결핵성흉막염환자의초기흉수양은등급 (grade) 이높아질수록흉수 MMP-8 이의미있게낮았고 (p=0.05, r= 0.413), MMP-8/TIMP-1 비도의미있게낮았다 (p=0.008, r= 0.536). 결핵성흉막염환자 23명중타병원으로전원간 5명을제외하고항결핵제치료를종료한환자는 1명이었다. 이중치료종료시촬영한단순흉부후전위촬영에서잔여흉막비후가없는군은 10명 (59%) 이었으며, 잔여흉막비후 가있는군은 명 (41%) 이었다. 잔여흉막비후가있는군은초진시흉수 TIMP-1 (41,405.9±9,3.3 ng/ml) 이잔여흉막비후가없는군 (29,134.9±8,801.8) 보다의미있게높았다 (p=0.032)(figure 2). 잔여흉막비후가있는군의초진시흉수 TIMP-2 (34.4±6.5 ng/ml) 은잔여흉막비후가없는군 (44.4±15.5) 보다낮았으나통계적의미는보이지않았다 (p=0.08). 치료종료한 1명의환자중추적관찰이불가능한 4명을제외하고 8±5개월의추적관찰이가능한 13명의환자들에서, 마지막으로촬영한흉부후전위촬영에서잔여흉막비후가없는군은 11명 (85%) 이었고, 잔여흉막비후가있는군은 2명 (15%) 이었다. 잔여흉막비후가있는군은초진시흉수 TIMP-2 (34.4±6.5 ng/ml) 가잔여흉막비후가없는군 (44.4±15.5) 보다의미있게낮았다 (p=0.038)(figure 3). 잔여흉막비후가있는군의초진시흉수 TIMP-1 (41405.9±934.3 ng/ml) 은잔여흉막비후가없는군 (29134.9±8801.8) 보다높았으나통계적의미는보이지않았다 (p=0.06). 고찰본연구의주요한결과로는, 결핵성흉막염치료후잔 11
YK Choi et al: Relationship between residual pleural thickening and TIMPs 여흉막비후가있는환자들이흉막비후가없는환자들보다흉수 TIMP-1은높은경향을, TIMP-2 는낮은경향을보였다는것이다. 세포외기질은아교질, proteoglycans 및당단백질등의구조단백질로구성되어있으며, 이의분해는조직의흡수와재형성에필수적인현상이다. 특히 matrixins 라고도명명하는 MMPs 는이러한과정에서매우중요한기능을수행하는것으로알려져있다 14. 대부분의 MMPs의발현은세포성장인자, 호르몬, 시토카인및형질전환등에의해전사조절된다. MMP-1 은 MMP-8, -13, -18 과함께결체조직의콜라겐성분은분해하는아교질분해효소이며, 흉막강에서 MMP-1은감염성흉막염에서섬유화반응을유도한다 15. MMP-9 은 MMP-2와함께젤라틴분해효소이며, 결핵균의 lipoarabinomannan 은 MMP-9 의분비를촉진시키고, 대식세포에서 MMP-1과 MMP-9의유전자발현을촉진시킨다 16. MMPs 의단백질분해활성은전구체로부터의활성화된형태로의전환과내인성억제자인 TIMPs 및 α-macroglobulins와의결합에의해조절되며, 이의균형은염증반응에서도중요한역할을한다 15,1. 결핵성흉수에서 MMP-1, -2, -8, 그리고 -9의수치가심부전증흉수보다높고 2, 국소 MMP-9과 TIMP-1의농도평형은단백질분해의순활성도 (net activity) 를결정하는데에중요한역할을한다고보고된바있다 18. TIMP-1의역할에대해서는아직확립되지는않았으나, MMPs 가기질을분해하는과정이활성화하는것을억제하고조절하는역할을하는것으로알려져있다 19. 특히 MMP-1, MMP-3, MMP-9은 TIMP-1 이 TIMP-2 보다더억제시킨다고알려져있으며 1,20-22, prommp-2도 TIMP-2보다는약하지만억제시킨다고알려져있다 20,23. 이전의연구에서와마찬가지로본연구에서도결핵성흉수내의 MMP-1, -2, -8, -9의농도가여출액보다높은경향을보여결핵성흉막염의염증및섬유화반응에이들 MMPs가관여하는것으로생각된다. 이전의연구들에서는결핵성흉수내의 TIMP-1 이부폐렴성이나악성, 여출액내보다높은경향이있는가하면 6, 이와달리여출액보다낮았다는보고도있었으나 2, 잔여흉막비후와비교되지는않았다. 본연구에서는초기결핵성흉막염진단시흉수 TIMP-1의농도가높은경우에는치료후에잔여흉막비후가남는경향을보여, TIMP-1이 MMP-1, -2, -9의활성화를억제시킴으로써염증반응으로발생된흉막비후의흡수를저해하는것으로추측하게한다. TIMP-2는 TIMP-1과마찬가지로 MMP-1, MMP-2, MMP-3, MMP-9 를억제시킨다고알려져있다 1,20-23. 하지 만 TIMP-2는 basic fibroblast growth factor 유도성혈관내피세포의성장억제등, MMPs의억제작용과는독립적인생물학적활성을보이기도한다 24. 본연구에서는흉수 TIMP-1에서의결과와는반대로, 초기결핵성흉막염진단시흉수 TIMP-2 의농도가낮은경우에치료후잔여흉막비후가남는경향을보였다. 이같은결과는, 상기와같은 MMPs의억제작용과는독립적인 TIMP-2 의생물학적활성때문인경우 ; TIMP-2 는높은농도에서는 MMP-2를억제시키지만 prommp-2가활성화되기위해서는오히려어느정도농도의 TIMP-2를필요로하므로 25, MMP-2의활성화에관련된경우 ; 또는아직밝혀지지않은다른기전과의상호작용에의한경우등을추정해볼수있다. 염증성흉수, 특히결핵성흉막염에서흉수내의 MMPs 와 TIMPs의농도는흉강내의국소적염증반응, 즉 tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin(il)-6, IL-8 림프구들이관여하여흉강내농도가증가된다 2. 증가된 MMPs와 TIMPs 는흉강에서체순환으로수동확산이되어혈중 MMPs와 TIMPs 의농도가증가된다 2. 결핵에의한염증반응으로흉막의투과성이증가되면반대로혈액내의 MMPs와 TIMPs 가흉강내로수동확산이되기도한다 2. 본연구에서는 MMP-2, MMP-8, TIMP-1, TIMP-2 의흉수내농도가혈청내농도보다높았고, 특히혈청 MMP-8 농도는다른군에서보다의미있게높았다. 이는국소적인흉수 MMP-8가많이생산된후혈액내로수동확산이된것으로추정되고, 결핵성흉막염과타질환과의감별에도움이될수있는가능성을기대하게했다. 하지만혈중 MMP 농도는다른질환이나신체상태에따라다른기원에서발생되어증가될수있으며, 본연구에서는다른기저질환이나신체상태에대해서는연구대상에서제외하였기때문에명확한연관관계를알수는없었다. 본연구의제한점으로는결핵성흉막염군외의대상자수가적어비모수적통계를사용했다는것이다. 기존의연구들과비교적유사한결과를보인것도있었으나, 통계적의미를보는데에는제한이따를수밖에없었다. 향후에는연구대상자수를늘려타질환과의비교분석을하는것이요할것으로사료된다. 더욱이 MMPs는 TIMPs 에의해서만생산이나활성화에영향을받는것이아니라다른염증매개체나섬유소용해인자에도영향을받으므로 6,15, 다른 MMPs, TIMPs 와이에영향을주는인자들의추가연구를한다면결핵성흉막염의잔여흉막비후의기전을밝히는데에도움이될것으로생각된다. ELISA 방법으로 12
Tuberculosis and Respiratory Diseases Vol. 65. No. 1, Jul. 2008 측정하는 MMPs와 TIMPs 는유리형과복합형이함께측정되는한계가있다. 연구자들에따라젤라틴분해효소인 MMP-2와 MMP-9은 gelatin zymography를통해 pro form 과 active form의농도를측정하여비교하기도한다. 본연구에서는결핵성흉수의 MMP-2와 MMP-9의 gelatin zymography 를통해관찰된 band 가다른군들에비해두드러지지않아직접 densitometry로측정하여비교하지는않고 ELISA 방법에의한것만비교분석하였다. 요약하면결핵성흉막염에서, 진단시흉수 TIMP-1 농도가높고흉수 TIMP-2 농도가낮은경우에치료후잔여흉막비후가지속된경우가더높아, 이들사이에연관관계가있을것으로추정된다. 흉부후전위촬영에서잔여흉막비후가없는군은 11명 (85%) 이었고, 잔여흉막비후가있는군은 2명 (15%) 이었다. 잔여흉막비후가있는군은흉수 TIMP-2 (34.4±6.5 ng/ml) 가잔여흉막비후가없는군 (44.4±15.5) 보다의미있게낮았다 (p=0.038). 결론 : 결핵성흉막염의잔여흉막비후의발생에 TIMP-1 과 TIMP-2이관여될수도있을것으로추정된다. 감사의글본연구에아낌없는도움을주신윤진영연구원께감사를드립니다 요 약 참고문헌 연구배경 : 잔여흉막비후는결핵성흉막염치료후흔히나타날수있는합병증중의하나이며, 이로인해호흡기능에지장을주는경우가있다. 이에결핵성흉막염진단시흉수내의 metalloproteinase (MMP)s 와 tissue inhibitor of metalloproteinase (TIMP)s 의농도가치료후잔여흉막비후가지속되는지예측할수있는인자가될수있는지알아보고자하였다. 방법 : 2004년 1월부터 2005년 6월사이에흉수가발견되어입원한환자를대상으로전향적연구를시행하였다. 진단시흉수의분석을통해결핵성흉막염, 부폐렴성흉수, 악성흉수, 여출액군으로나누고, 환자의혈청과흉수에서 ELISA 방법을이용하여 MMP-1, -2, -8, -9와 TIMP-1, -2를측정하였다. 결핵성흉막염의경우흉부엑스선검사로항결핵제치료종결시점과마지막추적관찰시점에잔여흉막비후의두께를측정하여잔여흉막비후가있는군과없는군으로나누었다. 결과 : 흉수가발견되어입원한환자중제외기준에해당하는환자를제외하고총 39명의환자가대상이되었다. 이중결핵성흉막염은 23명, 부폐렴성흉수 명, 악성흉수 명, 여출액 2명이었다. 결핵성흉막염환자 23명중본원에서항결핵제치료를종료한환자는 1명이었으며, 이중잔여흉막비후가없는군은 10명 (59%) 이었으며, 잔여흉막비후가있는군은 명 (41%) 이었다. 잔여흉막비후가있는군은흉수 TIMP-1 (41,405.9±9,3.3 ng/ml) 이잔여흉막비후가없는군 (29,134.9±8,801.8) 보다의미있게높았다 (p=0.032). 치료종료후평균 8±5개월의추적관찰이가능한 13명의환자들에서, 마지막으로촬영한 1. Eickelberg O, Sommerfeld CO, Wyser C, Tamm M, Reichenberger F, Bardin PG, et al. MMP and TIMP expression pattern in pleural effusions of different origins. Am J Respir Crit Care Med 199;156:198-92. 2. Hoheisel G, Sack U, Hui DSC, Huse K, Chan KS, Chan KK, et al. Occurrence of matrix metalloproteinases and tissue inhibitors of metalloproteinases in tuberculous pleuritis. Tuberculosis 2001;81:203-9. 3. Jin HY, Lee KS, Jin SM, Lee YC. Vascular endothelial growth factor correlates with matrix metalloproteinase-9 in the pleural effusion. Respir Med 2004;98: 115-22. 4. Kotyza J, Pesek M, Puzman P, Havel D. Progelatinase B/matrix metalloproteinase-9 proenzyme as a marker of pleural inflammation. Exp Lung Res 2004;30:29-309. 5. Park KJ, Hwang SC, Sheen SS, Oh YJ, Han JH, Lee KB. Expression of matrix metalloproteinase-9 in pleural effusions of tuberculosis and lung cancer. Respiration 2005;2:166-5. 6. Iglesias D, Alegre J, Aleman C, Ruiz E, Soriano T, Armadans LI, et al. Metalloproteinaes and tissue inhibitors of metalloproteinase in exudative pleural effusions. Eur Respir J 2005;25:104-9.. Di Carlo A, Terracciano D, Mariano A, Macchia V. Matrix metalloproteinase-2 and matrix metalloproteinase-9 type IV collagenases in serum of patients with pleural effusions. Int J Oncol 2005;26:1363-8. 8. Light RW, MacGregor MI, Luchsinger PC, Ball WC Jr. Pleural effusion: the diagnostic separation of transudates and exudates. Ann Intern Med 192;:50-13. 9. Ocaña I, Martínez Vázquez JM, Segura RM, Fernandez de Sevilla T, Capdevila JA. Adenosine deaminase in 13
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