口腔鳞癌TIL和MLNL的增殖能力及表型特征的比较研究

目的 :比较口腔鳞癌同一患者的肿瘤浸润淋巴细胞 (tumjorinfiltratinglymphocyte,TIL)和转移性淋巴结细胞(metestaticlymphnodelymphocyte,MLNL)的增殖能力及表型特征。方法 :从 7例口腔癌原发灶及其转移灶中分离TIL和MLNL ,与rIL - 2共培养 ,观察两种来源的淋巴细胞的增殖能力 ,用FACS检测细胞表型。结果 :6 0 %的淋巴细胞在培养的 4～ 5周可扩增 10 - 10 0倍 ,6例 >10 0倍。MLNL较TIL扩增高峰延迟 1周 ,MLNL培养 4周后仍显示其强有力的扩增潜力。新鲜分离MLNL和TIL以CD 3 、CD 4 、CD 8为主。MLNL中NK较TIL中少 ,CD 4 CD 8分别为 1.2 4和 1.19。培养 4周后 ,CD 3 、CD 4 、CD 8均增多 ,但CD 8增加比例较CD 4 大 ,CD 4 CD 8比分别为 :0 .95和 1.0 8。结论 :口腔鳞癌MLNL较TIL具有更强的增殖潜力。     

口腔鳞癌术前放化疗和未治疗患者TIL增殖及特异性杀伤活性的比较

目的 探讨术前做过放化疗口腔鳞癌患者肿瘤浸润淋巴细胞（tumor infiltrating lymphocyte,TIL）在体外能否继续增殖及恢复特异性杀伤活性。方法 从15例口腔鳞癌原发灶中分离TIL，比较术前放化疗和未治疗患者TIL体外增殖能力及对自体肿瘤细胞的杀伤活性。结果 术未治疗组比治疗组提前一周达增殖高峰，但4周以后术前治疗组也显示出继续扩增的趋势。培养3周时治疗组TIL对自体肿瘤的杀伤活性＜未治疗组（P＜0.05），分别为26％和35％；5周时治疗组TIL对自体肿瘤的杀伤活性与未治疗组无显著性差异（P＞0.05），分别为41％和45％。结论 从术前放化疗患者肿瘤组织中分离的TIL在体外扩增后转输体内是可行的。     

口腔鳞癌TIL和TDLNL杀伤活性的比较研究

目的　比较口腔鳞癌肿瘤浸润淋巴细胞 (tum or infiltrating lym phocyte,TIL)和肿瘤引流淋巴结细胞(tum or draining lym ph node lymphocyte,TDL NL)体外杀伤活性。方法　从 5例口腔癌原发灶及其转移灶中分离TIL和 TDL NL ,用 3H - Td R4h释放法对 TDL NL和 TIL对自体肿瘤的杀伤活性进行比较研究。结果　 3周时 TIL对自体肿瘤的杀伤活性较 TDL NL强 (P<0 .0 2 ) ,分别为 35 %和 2 3% ;4周时 TDL NL对自体肿瘤的杀伤活性大于TIL (P<0 .0 2 ) ,分别为 5 3%和 40 % ;5周时 TDL NL对自体肿瘤的杀伤活性仍显著强于 TIL (P<0 .0 2 ) ,分别为6 2 %和 44 %。结论　在口腔鳞癌过继免疫治疗中 ,TDL NL可能较 TIL具有更大的优势     

口腔癌TIL的提取、培养、表型分析及体外抗瘤活性研究

本文对９例口腔癌手术切除的原发灶肿瘤标本的ＴＩＬ进行了体外分离、培养、扩增、表型分析及ＮＫ和ＬＡＫ活性测定。其中８例扩增获得成功，扩增倍数为１０～１２００（平均４７２．２）倍。表型分析表明，扩增后的ＴＩＬ中ＣＤ，ＣＤ和ＣＤ均有增加，但以ＣＤ、ＣＤ为主。抗瘤活性在培养４周左右达最高峰；ＮＫ和ＬＡＫ活性分别为５２％和４４％。全部病例经肉汤培养基、沙氏培养基、支原体检测药盒及家兔发热试验未检测到细菌、真菌、支原体及热原质。本研究提示，口腔癌ＴＩＬ的临床应用是可行的。     

术前放化疗和未治疗口腔鳞癌患者TIL增殖力的比较

目的 探讨术前做过放化疗口腔鳞癌患者TIL在体外与rIL-2共培养是否具有继续增殖的能力。方法 从15例口腔鳞癌原发灶中分离TIL,用1000u/ml rIL-2与其培养,比较术前放化疗和未治疗患者TIL的增殖能力。结果 术前未治疗组比治疗组扩增快,提前1周达增殖高峰,但4周以后术前治疗组也显示出继续扩增的趋势。结论 从术前放化疗患者肿瘤组织中分离的TIL,在体外经rIL-2刺激后仍能继续扩增。     

几种免疫调节剂对口腔鳞癌浸润淋巴细胞的体外增殖、杀瘤活性的影响研究

目的:研究IL-4、GM-CSF和香菇多糖对口腔鳞癌浸润淋巴细胞(TIL)的体外培养和细胞毒效应的影响。方法:将手术切取的标本通过酶消化加机械分离法提取TIL,分成4组。以单纯加IL-2培养的TIL为对照组,另3组在加IL-2的基础上分别加入IL-4、GM-CSF和香菇多糖进行培养。每3d计数1次并绘制细胞生长曲线图。于对数生长期用MTT法检测TIL的杀瘤活性并进行统计学比较。结果:添加IL-4和GM-CSF的TIL增殖速度明显大于对照组和香菇多糖组,对数生长期出现早且持续时间较长,细胞增殖量也较对照组和香菇多糖组多。MTT法检测TIL杀瘤活性显示,IL-4和GM-CSF组的杀瘤活性强于对照组和香菇多糖组,差异均显著。结论:GM-CSF、IL-4协同IL-2能够提高TIL的增殖速度,促进TIL的增殖,且能在一定程度上解除TIL的免疫抑制,提高其杀瘤活性。     

舌癌瘤浸润性淋巴细胞与外周血淋巴细胞抗癌活性的比较

肿瘤过继免疫治疗是近年研究的热门课题。如何获取有效足量,而且来源容易的抗瘤活性细胞是一关键性问题。由口腔内癌瘤制取TIL细胞存在两个问题:一是肿瘤体积较小,该得到治疗数量级的TIL较困难;二是口腔内肿瘤处在一个非无菌环境,制取的TIL细胞易污染。本文旨在通过实验研究,比较同一患者的肿瘤引流区的淋巴结细胞与LAK细胞体外抗瘤活性,为TIL过继免疫治疗提供理论根据。结果表明TIL对Tca8113细胞表现明显的杀伤作用,TIL与LAK细胞组比较,两组间存在显著性差别(P<0.05)。     

经皮下埋植泵诱导化疗治疗口腔癌TIL表型研究

目的 :通过观察诱导化疗前后N0期口腔癌局部肿瘤浸润淋巴细胞表型的改变 ,初步探讨经皮下埋植泵诱导化疗对口腔癌局部免疫功能的影响。方法 :47例N0期口腔癌术前分别经皮下埋植泵、静脉给予甲氨喋呤 卡铂 平阳霉素。采用免疫组化方法对诱导化疗前后癌灶TIL进行识别观察。结果 :免疫组化检测表明 ,经皮下埋植泵诱导化疗后TIL中CD3、CD4 、CD8、CD4 /CD8比值无明显变化 (P >0 .0 5 ) ,6例治疗无效者CD3、CD4 阳性细胞显著下降 ,CD8阳性细胞无变化。而单纯静脉诱导化疗后 ,TIL中CD4 、CD4 /CD8比值显著降低 (P <0 .0 5 )。结论 :通过皮下埋植泵诱导化疗 ,可以减轻化疗药物对口腔癌肿瘤微环境免疫状态的抑制作用 ,治疗无效者局部细胞免疫功能处于抑制状态 ,其中主要是CD4 细胞亚群的形成受到抑制 ,说明局部免疫状况是影响化疗疗效的重要因素     

口腔癌肿瘤浸润淋巴细胞对人舌癌裸鼠移植瘤的生长抑制作用

目的探讨在体外具有强大细胞毒活性的口腔癌浸润淋巴细胞在体内的抑瘤效果以及化疗药物环磷酰胺（ｃｙｃｌｏｐｈｏｓｐｈａｍｉｄｅ，Ｃｙ）与肿瘤浸润淋巴细胞（ｔｕｍｏｒｉｎｆｉｌｔｒａｔｉｏｎｌｙｍｐｈｏｃｙｔｅ，ＴＩＬ）联合应用治疗口腔癌的可能性。方法将舌鳞癌细胞系Ｔｃａ８１１３注入裸鼠背部皮下建立移植瘤模型，取培养４周的ＴＩＬ联合低剂量Ｃｙ局部注射，观察肿瘤体积变化及肿瘤生长情况。结果①ＴＩＬ＋ｒＩＬ２和ＴＩＬ＋ｒＩＬ２＋Ｃｙ组在３周内抑瘤率较高（分别为：８６１％±０４％和９７７％±０６％），３周后抑瘤作用减弱；②ＴＩＬ＋ｒＩＬ２＋Ｃｙ组抑瘤率较ＴＩＬ＋ｒＩＬ２组高（第８周抑瘤率分别为：２００％±１４％和７５５％±２５％，Ｐ＜００１）。结论口腔癌ＴＩＬ在应用后３周内具较强抑瘤作用，３周后减弱；联合应用Ｃｙ可获得更佳抑瘤效果     

ICAM-1和LFA-1单抗对口腔鳞癌肿瘤浸润淋巴细胞杀伤活性的影响

目的　研究ICAM-1/LFA-1单抗阻断对口腔鳞癌肿瘤浸润淋巴细胞(TIL)对自体肿瘤细胞杀伤活性及分泌TNF-α的影响。方法　从15例口腔鳞癌组织中分离TIL并培养,用流式细胞术检测新鲜分离和激活的TIL细胞上ICAM-1和LFA-1的表达水平;比较用ICAM-1/LFA-1单抗阻断前后TIL对自体肿瘤细胞杀伤活性及产生TNF-α 水平的影响。结果　激活的TIL较新鲜分离的TIL细胞ICAM-1和LAF-1的表达显著增加(P<0·05);阻断ICAM-1, TIL对自体肿瘤细胞的杀伤活性无显著改变;阻断ICAM-1和LAF-1后,TIL分泌TNF-α无显著差异(P>0·05),但TIL 对自体肿瘤细胞的杀伤活性显著降低(P<0·05)。结论　anti-LFA-1单克隆抗体(McAb)可能通过干扰抗原非特异性的粘附过程而发挥作用。     

The prognostic role of tumour‐infiltrating lymphocytes in oral squamous cell carcinoma: A meta‐analysis

It has been suggested that tumour‐infiltrating lymphocytes (TILs) are associated with the progression of oral squamous cell carcinoma (OSCC). However, the prognostic value of TILs is inconclusive due to the heterogeneity of immune cells within the tumour microenvironment. In this meta‐analysis, we aimed to assess the prognostic value of TILs in OSCC. The PubMed, Cochrane, Embase, Scopus and Web of Science databases were searched up to April 20, 2019, and 33 studies were ultimately included in this meta‐analysis. Our pooled meta‐analysis showed that high infiltration of CD8⁺ TILs, CD45RO⁺ TILs and CD57⁺ TILs favoured better overall survival (OS). However, high infiltration of CD68⁺ macrophages and CD163⁺ macrophages was associated with poor prognosis in OSCC. These findings suggest that CD8⁺ TILs, CD45RO⁺ TILs, CD57⁺ TILs, CD68⁺ macrophages and CD163⁺ macrophages might serve as novel prognostic factors and therapeutic targets in OSCC.     

腺样囊性癌细胞凋亡抗原负载的树突状细胞诱导抗肿瘤效应的体外研究

目的 探讨负载腮腺腺样囊性癌细胞凋亡肿瘤抗原的树突状细胞,诱导淋巴细胞介导的免疫反应,观察其体外杀伤腺样囊性癌细胞的细胞毒性效应.方法 外周血单核细胞,在粒细胞/巨噬细胞集落刺激因子(Granulocyte-macrophage colony-stimulating factor GM-CSF) 白细胞介素-4(Interleukin-4 IL-4)的诱导下体外培养,用凋亡肿瘤细胞抗原冲击后,流式细胞仪检测树突状细胞抗原负载前后CD1a、CD83表达量的变化.采用MTT比色法检测同种混合淋巴细胞反应和诱导细胞毒性T细胞(Cytotoxic T lymphocyte CTL)反应.结果 凋亡肿瘤抗原刺激后,CD83 表达急剧增加(P＜0. 01),而CD1a表达量下调(P＜0.01).负荷凋亡肿瘤抗原树突状细胞体外诱导出明显的细胞毒性反应,并刺激同种T淋巴细胞增殖.结论 GM-CSF IL-4 诱导的单核细胞来源的树突状细胞 ,能在体外摄取凋亡肿瘤抗原而进一步成熟,通过激活淋巴细胞杀伤癌细胞.     

Determination of lymphocyte populations and subpopulations extracted from chronically inflamed human periodontal tissues

The lymphocyte populations and subpopulations extracted from inflamed periodontal tissues of patients with adult periodontitis were determined. 34 patients were grouped according to the gingival index score (GI) of 1, 2 and 3. Gingival tissue from 2 involved teeth was excised, treated with collagenase, and infiltrating cells were isolated and identified using monoclonal antibodies for lymphocyte sets and subsets. The % of CD3+ cells was about 54.5% in all 3 patient groups, but the percentage of CD22+ cells increased from 28.9 +/- 3.3% in the group with GI = 1 to 33 +/- 1.2% in the group with GI = 3. %s of CD4+ cells and activated CD4+ cells increased from 30.2 +/- 2.1% and 4.7 +/- 1.7% in the group with GI = 1 to 38.4 +/- 1.2% and 16.0 +/- 3.4% in the group with GI = 3, respectively, while in the same groups, the % of CD8+ cells decreased from 24.9 +/- 2.0% to 17.7 +/- 1.6%. These data indicate a possible importance of activated CD4+ cells in pathogenetic mechanisms of periodontitis.     

In vitro cytotoxic response to lithium disilicate dental ceramics.

OBJECTIVES: The use of lithium disilicate dental ceramics is increasing in dentistry and previous reports have suggested that they may have greater biological risks than previously thought. We tested a hypothesis that composition and processing influence the biological properties of these ceramics. METHODS: The cytotoxicity of two machined and three pressed lithium disilicate materials (n=6) were tested in vitro using mouse fibroblasts in direct contact with the materials for 72h. Cellular response was estimated by mitochondrial succinate dehydrogenase activity (MTT method). Mitochondrial activity was expressed as a percentage of Teflon controls, then compared to Teflon using 2-sided t-tests (alpha=0.05). Polished materials were aged in artificial saliva and tested for cytotoxicity periodically over 6 weeks, then were repolished (320grit SiC paper), aged and tested again for 4 weeks. RESULTS: All materials significantly (50-70%) suppressed cellular mitochondrial activity in the initial week, but suppression decreased by 25-30% over the next 2 weeks. In weeks 4 and 6 some materials exhibited a cytotoxic 'relapse' of 10-20%. The cytotoxic response was no different for machined or pressed materials, but the presence of ZnO had at least an association with longer-term cytotoxicity and relapse. Repolishing to 320grit did not increase cytotoxicity significantly. SIGNIFICANCE: Our results suggest that lithium disilicates are not biologically inert, and that many have a similar cytotoxicity dynamic regardless of small differences in composition or processing.     

Cytotoxicity of amalgams, alloys, and their elements and phases

The purpose of this study was to compare the relative cytotoxicity of amalgams, alloys, and their constituent elements and phases, by means of a rapid and sensitive in vitro cell culture test. Pure copper and zinc showed intensive cytotoxicity, significantly greater than that of pure silver and mercury. Pure tin was non-cytotoxic. The gamma-one phase (Ag2Hg3) revealed moderate cytotoxicity, which was significantly decreased by the addition of 1.5% and 5% Sn. However, the addition of 1.5% Zn to gamma 1 containing 1.5% Sn dramatically increased the cytotoxicity of gamma 1 to the same level as that of pure zinc. Whenever zinc was present in amalgams, higher cytotoxicity was revealed. High-copper amalgams showed the same cytotoxicity as a zinc-free low-copper amalgam. The addition of selenium did not reduce the cytotoxicity of amalgam. The cytotoxicity of amalgams was reduced after 24 h. The results of this study suggest that the major contributor to the cytotoxicity of alloy for amalgam is probably copper, while that for amalgam is zinc.     

新型双层骨引导再生壳聚糖膜修复大鼠颅骨缺损的实验研究

目的: 通过比较新型双层壳聚糖膜和Bio-Gide膜对大鼠颅骨缺损的修复效果,评估其作为引导骨再生膜的可行性。方法: 双层壳聚糖膜与MC3T3-E1细胞共培养,应用扫描电子显微镜（scanning electron microscope,SEM）观察细胞对膜的黏附效果;采用cell counting kit-8（CCK-8）法在3、7和10 d检测膜的细胞毒性;选择18只Sprague-Dawley（SD）大鼠,随机分为A、B、C组,在其颅骨中线两侧制作5 mm的极量骨缺损,左侧为实验侧,覆盖双层壳聚糖膜,右侧为对照侧覆盖Bio-Gide膜,分别于术后第2、4及8周时处死各组大鼠,通过大体观察、微计算机体层扫描技术（micro-computed tomography,micro-CT）及组织学方法,评价2种膜修复骨缺损的效果。采用SPSS20.0软件包对结果进行统计学分析。结果: 通过MC3T3-E1细胞在双层壳聚糖膜上培养2 d后的扫描电镜照片,观察到细胞较好地粘附于多孔层上。第3、7及10天时,实验组细胞相对增殖率分别为114.49%、107.17%和98.73%,细胞毒性级别为0或1级,表明膜的细胞毒性轻微。2周时,Bio-Gide膜组的骨形成量（BV）和骨体积分数（BVF、BV/TV）与实验组差异显著（P<0.05）;但在第4和8周时,2组的BV和BVF无显著差异（P>0.05）。结论: 新型双层壳聚糖的体外和体内生物学性能符合GBR技术的要求,具有作为GBR膜的潜力。