背景:细胞毒性淋巴细胞(CLs)表达强效毒素,包括穿孔素(P)和颗粒酶-B(G),导致目标细胞死亡。本研究的目的是通过P和G分析来评估肿瘤浸润性CLs的杀伤能力。探讨无桥本甲状腺炎(HT)的甲状腺乳头状癌(PTC)与淋巴结转移的关系。
方法:在冰冻切片中观察到PTC中淋巴细胞的浸润。收集新鲜肿瘤组织和有淋巴细胞浸润的癌旁组织并制备成单细胞悬液。流式细胞术检测CD3+P+的百分比,CD3+G+,CD8+P+,和CD8+G+T淋巴细胞(TLs)和CD16-CD56+P+和CD16-CD56+G+自然杀伤(NK)细胞。最后,我们研究了配对肿瘤组织中NK细胞和细胞毒性T淋巴细胞(CTL)中P和G的差异表达(T组,n=44)和癌旁组织(N组,n=44)来自具有BRAFV600E突变的PTC患者。此外,根据是否存在颈中央区淋巴结转移(CCLNM)将患者分为两组:A组(有淋巴结转移,n=27)和B组(无淋巴结转移,n=17)。根据CCLNM阳性总数将患者分为三组:B组,C组(伴有低度淋巴结转移,小于5,n=17)和D组(淋巴结转移高,不小于5,n=10)。
结果:N组CD3+P+CTLs百分比明显高于T组(P<0.05)。T组CD8+G+CTLs百分比明显高于N组(P<0.05)。CD3+G+的百分比,T组和N组CD16-CD56+P+和CD16-CD56+G+NK细胞无明显差异(P>0.05)。A组和C组的CD3+P+CTLs在癌旁组织中的百分比明显高于肿瘤组织(P<0.05)。A组和C组肿瘤组织中CD8+G+CTLs的百分比明显高于癌旁组织(P<0.05)。D组肿瘤组织中CD16-CD56+G+NK细胞的比例明显高于癌旁组织(P<0.05)。
结论:肿瘤组织和癌旁组织对PTC浸润性CLs的杀伤能力不同。在使用CCLNM的情况下,肿瘤组织中CD16-CD56+G+NK细胞的高表达可能与淋巴结转移的高风险有关。
BACKGROUND: Cytotoxic lymphocytes (CLs) express potent toxins, including perforin (P) and granzyme-B (G), which brings about target cell death. The purpose of this study was to evaluate the killing capacity of tumor-infiltrating CLs by means of P and G analysis, and explore the association with lymph node metastasis in papillary carcinoma of thyroid (PTC) without Hashimoto\'s thyroiditis (HT).
METHODS: Infiltration of lymphocytes in PTC was observed in frozen sections. Both fresh tumor tissues and paracancerous tissues with lymphocyte infiltration were collected and prepared into a single cell suspension. Flow cytometry was used to detect the percentages of CD3+P+, CD3+G+, CD8+P+, and CD8+G+ T lymphocytes (TLs) and CD16-CD56+P+ and CD16-CD56+G+ natural killer (NK) cells. Finally, we investigated differential expression of P and G in NK cells and cytotoxic T lymphocytes (CTLs) in paired tumor tissues (group T, n = 44) and paracancerous tissues (group N, n = 44) from patients with PTC with the BRAF V600E mutation. Furthermore, patients were divided into two groups according to whether cervical central lymph node metastasis (CCLNM) existed: group A (with lymph node metastases, n = 27) and group B (with nonlymph node metastases, n = 17). Patients were also divided into three groups according to the total number of positive CCLNM: group B, group C (with low-level lymph node metastases, less than 5, n = 17) and group D (with high-level lymph node metastases, no less than 5, n = 10).
RESULTS: The percentage of CD3+P+ CTLs was significantly higher in group N than in group T (P < 0.05). The percentage of CD8+G+ CTLs was significantly higher in group T than in group N (P < 0.05). The percentages of CD3+G+, CD16-CD56+P+and CD16-CD56+G+ NK cells showed no significant difference in either group T or group N (P > 0.05). The percentages of CD3+P+ CTLs in group A and group C were significantly higher in the paracancerous tissue than in the tumor tissue (P < 0.05). The percentages of CD8+G+ CTLs in group A and group C were significantly higher in the tumor tissues than in the paracancerous tissues (P < 0.05). The percentage of CD16-CD56+G+ NK cells in group D was significantly higher in the tumor tissues than in the paracancerous tissues (P < 0.05).
CONCLUSIONS: The killing capacity of infiltrating CLs in PTC differed between tumor tissues and paracancerous tissues. In cases with CCLNM, higher expression of CD16-CD56+G+ NK cells in tumor tissues may be associated with a high risk of lymph node metastasis.