福馬林固定後石蠟包埋標本內結核桿菌的蜂巢式聚合酶連鎖反應測試與其組織學分析

Detection of Mycobacterium Tuberculosis by Nested PCR and Histological Analysis in Formalin-Fixed Paraffin-Embedded Tissue

結核病在台灣是常見的疾病，以傳統acid-fast stain （AFB）或細菌培養，不是診斷率太低（約15%）就是時間太長（約2個月），尤其在石蠟包埋肉芽腫的診斷史是困難。我們以蜂巢式聚合酶連鎖反應（nested PCR）來偵測福馬林固定石蠟包埋肉芽切片組識中結核桿菌之存在，配合組織學上的表現來進行分析。結果發現以單一步驟放大反應在52個病人中有19人（36.5%）呈陽性反應，而在連續性第二步驟反應，陽性率可提高至63.5%（33人）。結核桿菌蜂巢式聚合酶連鎖反應的陽性組織學型態分析，有下列4種：典型肉芽腫（19/33）、類肉瘤肉芽腫（4/33）、類淋巴上皮肉芽腫（9/33）、星狀肉芽腫（2/33）。另外在33件組織中14件即具微小囊腫的變化。病理學上，典型性藍格罕士巨細胞（Langhan's giant cell）占54.5%（18/33），同時出現典型肉芽腫和類肉瘤肉芽腫的有2例，類淋巴上皮肉芽腫和典型肉芽腫同時出現的有1例。結論：1.以蜂巢式聚合酶連鎖反應診斷結核病是一快速、敏感之方法。2.不同型態的肉芽腫可能是因結核病不同階段，或者不同宿主免疫力所引起。

A simplified protocol based on the nested polymerase chain reaction (PCR) was developed for detection of Mycobacterium tuberculosis in the formalist-fixed paraffin-embedded chronic granulomatous inflammation tissue. By DNA amplification, PCR was positive in l9 (36.5%) of the 52 patients in the one-step amplification, whereas PCR was positive in 33 (63.5%) of the 52 patients following the two-step amplification (P<0.001). Histologically, there were four types of granulomas identified: (1) classical M. tuberculosis granulomas (19/33); (2) sarcoidosis-like granulomas (4/33); (3) Iympho-epithelioma-like granulomas (LELG) (9/33); (4) satellite granulomas (2/33). In addition, the microabscesses were present in 14/33. The typical Langhan's giant cells were present in 54.5% cases (18/33). Interestingly, the classical M. tuberculosis granulomas and sarcoidosis-like granulomas were present simultaneously in two cases. The LELG and classical granulomas were present simultaneously in seven cases. LELG and sarcoidosis-like granulomas were present simultaneously in one case. In summary, the nested PCR is useful for rapid diagnosis of M. tuberculosis. The variety of granulomas may be due to different stages of tuberculosis and/or the difference of individual host immunity.