摘要 目的探讨转化生长因子β1(TGF-β1)对角膜碱烧伤机体巨噬细胞和淋巴细胞功能的影响。
方法Balb／c鼠单眼角膜Ⅱ度碱烧伤，治疗组于模型制作后1 h内球后注射TGF-β1 50 ng联合腹腔注射TGF-β1 30 ng。动态观察腹腔巨噬细胞吞噬功能和循环淋巴细胞对促分裂原刺激的转化功能，并与对照组进行对比分析。
结果经TGF-β1治疗后，腹腔巨噬细胞吞噬能力在5周内明显低于对照组（P＜0.05)；淋巴细胞转化在植物血凝素(PHA)刺激时，3周内显著降低（P＜0.05)，但在烧伤角膜蛋白刺激时，差别无显著性（P＞0.05)。
结论微量TGF-β1对角膜碱烧伤机体免疫活性细胞的功能具有抑制作用。
分类号 R772.2 R446.6
Effect of transforming growth factor β1 on the immune cells following corneal
alkali burns
Liu Chunmin，Xu Jintang．
（First Affiliated Hospital of Medical College，Jinan University，Guangzhou 510632）
Abstract ObjectiveTo observe the effect of transforming growth factor β1（TGF-β1)on the immune cells following corneal alkali burns．MethodsBalb／c mice were given mild corneal alkali burns．In treatment group，the mice were injected TGF-β1 50 ng postbularly combined with 30 ng abdomino-injection．The control mice were given corneal burns without TGF-β1 injection．In both groups，the macrophages were obtained by washing the abdominal cavity of mice using normal saline，and the lymphocytes in the blood were acquired at the same time by density gradient centrifugation．The normal corneas（control group）and burned corneas were excised and homogenized in PBS．The homogenate was then centrifuged and the supernatant was used as a source of corneal protein．Every other week in a period of 7 weeks，the macrophages were used for tests of phagocytosis while the lymphocytes were cultured in complete RPMI 1 640 culture medium with or without phytoagglutinin（PHA）or mice corneal protein for 54 h followed by incubation with 3 H-TdR for 18 h．Counts per minute（cpm），relative plastid-transformation index（RPI）and stimulation index（SI）were analyzed．ResultsFor mice with TGF-β1 injection，the phagocytosing rate within 5 weeks and phagocytosing index during 3～5 weeks were obviously lower than those in controls（P＜0.05)；and the △cpm，RPI or SI in blastogenesis tests induced by PHA before the 3rd week were markedly lower than those of control mice（P＜0.05)． There was no difference between the two groups in the △cpm，RPI or SI in blastogenesis tests induced by mice corneal proteins during the period of observation（P＞0.05)．ConclusionTGF-β1 can suppress the phagocytic function of macrophages and blastogenesis of lymphocytes induced by PHA．
Key word scornea alkali-burn transforming growth factor-β1
转化生长因子β1(transforming growth factor beta 1，TGF-β1)是一种具有多向生物效应的细胞调节因子，它可作为促进细胞增殖的信号，又能抑制 细胞的增殖，在传递生物信息、创伤修复、多方位控制细胞功能及免疫调节等方面起重要的调控作用［1，2］。
TGF-β1对细胞生[＝F（长和免疫调节的效果取决于其所作用的细胞类型、细胞的分化状态、机体免疫状态，以及环境中存在的其它因子的作用，同时与其剂量有一定的关系［2，3］。多数研究是在体外条件下进行的，活体状态下TGF-β1的作用因受机体复杂因素的影响而难与离体实验相一致。目前尚未见到有关外源性TGF-β1对角膜碱烧伤机体的影响的文献报道。我们以Balb／c鼠作为实验对象，对此进行了初步探讨。
1 材料与方法
1．1 实验试剂
TGF-β1(human，natural，ED50＝0.235 ng／ml，promega）2 μg加入灭菌的0.01 mol／L，pH 7.4 PBS 2 ml，临用前配制。
1．2 实验动物与分组
雄性Balb／c鼠116只，体重约20 g。随机分为：A组(15只)，用于分离代表正常群体标本的淋巴细胞；B组(20只)，用于制备碱烧伤角膜蛋白提取液；C组(9只)，作为正常对照；治疗组(36只)，再分为1，3，5，7周组，各9只；对照组(36只)，再分为1，3，5，7周组，各9只。
1．3 实验方法
1．3．1 制作单眼角膜Ⅱ度碱烧伤模型：麻醉动物后，将单层直径3 mm定量滤纸浸入1 mol／L NaOH溶液中10 s，取出放在右眼角膜中央部20 s后，除去滤纸，用生理盐水冲洗右眼10 min，造成Ⅱ度角膜碱烧伤模型［4］。对治疗组实验鼠，用生理盐水冲洗后，在烧伤眼进行眼球后注射TGF-β1 50 ng，腹腔注射30 ng。
1．3．2 B组实验鼠模型制作后3天，断颈处死，摘取伤侧角膜20只，用0.01 mol／L，pH 7．4 PBS，碾碎、离心，获取上清液；将沉淀物再次离心后，合并两次所得的上清液，最终制得烧伤角膜蛋白提取液约1 ml。
1．3．3 对C组、对照组和治疗组实验鼠，在取材前72 h，腹腔注射灭菌石蜡油约0.3 ml／只。实验当日，摘取眼球放血(加肝素抗凝)，分离淋巴细胞，并用RPMI 1640完全培养液调整细胞数至1×106个／50 μl；按设定板孔加样于96孔培养板内，每样本均设对照、PHA刺激孔、烧伤角膜蛋白刺激孔，各12个复孔，以3H-TdR掺入法进行淋巴细胞转化试验，结果以△cpm，RPI，SI表示［5］。同时，对上述实验鼠进行腹腔冲洗，获取巨噬细胞，以落片法进行巨噬细胞吞噬鸡红细胞试验，结果以吞噬百分率和吞噬指数表示，同时观察巨噬细胞对鸡红细胞的消化程度［5］。烧伤眼球作病理检查。
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