COMPARISON OF CYTOKINE EFFECTS ON MOUSE PANCREATIC ALPHA-CELL AND BETA-CELL LINES - VIABILITY, SECRETORY FUNCTION, AND MHC ANTIGEN EXPRESSION
COMPARISON OF CYTOKINE EFFECTS ON MOUSE PANCREATIC ALPHA-CELL AND BETA-CELL LINES - VIABILITY, SECRETORY FUNCTION, AND MHC ANTIGEN EXPRESSION
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DOI：
10.2337/diabetes.39.4.415
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发表时间：
1990-04-01
期刊：
影响因子：
7.7
通讯作者：
LEITER, EH
中科院分区：
文献类型：
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作者：
HAMAGUCHI, K;LEITER, EH
细胞因子对转化的小鼠胰腺α-IFN-γ的永久细胞系的作用和β-细胞进行了比较。β-肿瘤细胞1（β TC 1）系（来自在大鼠胰岛素II启动子控制下表达SV 40大T抗原癌基因的转基因小鼠中产生的腺瘤）主要产生胰岛素，尽管通过放射免疫测定可检测到少量的细胞内胰高血糖素（100：1胰岛素比胰高血糖素）。α TC 1系（来自在大鼠前胰高血糖素原启动子控制下表达SV 40大T抗原癌基因的转基因小鼠中产生的腺瘤）不仅产生胰高血糖素，而且产生相当大量的胰岛素（胰高血糖素与胰岛素之比为4：1）和前胰岛素原mRNA。因此，我们克隆了α TC 1细胞，并获得了12种产生胰高血糖素的克隆细胞系，其不产生可通过放射免疫测定检测的胰岛素水平。通过对来自两个株系，α TC 1克隆6和9的总RNA的北方印迹分析，证实了前胰岛素原mRNA的缺失。在α TC 1克隆6或9或β TC 1细胞中通过免疫组织化学染色未检测到生长抑素或胰多肽。大鼠重组γ-干扰素（IFN-γ; 5-250 U/ml）或小鼠重组白细胞介素1（IL-1; 1-25 U/ml）单独抑制β TC 1细胞中的DNA合成。这两种细胞因子联合作用，进一步抑制DNA合成，增加细胞毒性。相反，α TC 1克隆9细胞对每种细胞因子单独抑制DNA合成不敏感，尽管胰高血糖素合成受到抑制。这些细胞因子的组合引起α TC 1克隆9细胞中DNA和胰高血糖素合成的显著抑制。α TC 1克隆9细胞比β TC 1细胞对组合的细胞因子的细胞毒性作用具有更大的抗性。用50 U/ml IFN-γ孵育在β TC 1、未克隆的α TC 1和α TC 1克隆6和9的细胞表面上诱导裂解II类MHC分子（I-Ab、I-Ad和I-Ed）并增强I类分子（H-2Kb和H-2Kd）的组成型表达。因此，这些细胞系对于衍生自用于构建转基因小鼠的两种亲本品系[C57 BL/6 J（H-2b）和DBA/2 J（H-2d）]的MHC等位基因是杂合的。IFN-γ诱导的II类基因转录在β TC 1和α TC 1克隆9细胞中通过用A α-，A β-，E α-，和E β- DNA探针。对细胞因子的细胞毒性作用的敏感性的差异可能与β-CD的优先破坏有关。胰岛素依赖型糖尿病的早期阶段。因为I类和II类MHC表达被IFN-γ增强或诱导。在α-和β-细胞系，诱导MHC分子的能力本身不足以解释β-与胰岛素依赖型糖尿病相关的细胞特异性自身免疫识别。
Cytokine effects on permanent cell lines of transformed mouse pancreatic .alpha.- and .beta.-cells were compared. The .beta.-tumor cell 1 (.beta.TC1) line (From an adenoma created in transgenic mice expressing the SV40 large T-antigen oncogene under control of the rat insulin II promoter) produced insulin predominantly, although small quantities of intracellular glucagon (100:1 insulin to glucagon) were detectable by radioimmunoassay. The .alpha.TC1 line (from an adenoma created in transgenic mice expressing the SV40 large T-antigen oncogene under control of the rat preproglucagon promoter) produced not only glucagon but also considerable quantities of insulin (4:1 glucagon to insulin) and preoproinsulin mRNA. We therefore cloned .alpha.TC1 cells and obtained 12 glucagon-producing clonal cell lines that did not produce levels of insulin detectable by radioimmunoassay. Analysis by Northern blotting of total RNA from two lines, .alpha.TC1 clones 6 and 9, confirmed the absence of preproinsulin mRNA. No somatostatin or pancreatic polypeptide was detected by immunohistochemical staining in .alpha.TC1 clones 6 or 9 or .beta.TC1 cells. Rat recombinant .gamma.-interferon (IFN-.gamma.; 5-250 U/ml) or mouse recombinant interleukin 1 (IL-1; 1-25 U/ml) individually inhibited DNA synthesis in .beta.TC1 cells after 3 days of treatment. The two cytokines in combination acted synergistically to further depress DNA synthesis and increase cytotoxicity. In contrast, .alpha.TC1 clone 9 cells were not sensitive to inhibition of DNA synthesis by each cytokine individually, although glucagon synthesis was inhibited. The combination of these cytokines caused marked inhibition of DNA and glucagon syntheses in .alpha.TC1 clone 9 cells. .alpha.TC1 clone 9 cells were somewhat more resistant to the cytotoxic action of the combined cytokines than were .beta.TC1 cells. Incubation with 50 U/ml IFN-.gamma. induced cleass II MHC molecules (I-Ab, I-Ad, and I-Ed) and enhanced the constitutive expression of class I molecules (H-2Kb and H-2Kd) on the cell surface of .beta.TC1, uncloned .alpha.TC1, and .alpha.TC1 clones 6 and 9. Thus, these cell lines are heterozygous for MHC alleles derived from both parental strains used in the construction of the transgenic mice [C57BL/6J (H-2b) and DBA/2J (H-2d)]. Class II gene transcription induced by IFN-.gamma. was confirmed in .beta.TC1 and .alpha.TC1 clone 9 cells by Northen blot anlaysis with A.alpha.-, A.beta.-, E.alpha.-, and E.beta.-DNA probes. The differences in sensitivity to the cytotoxic action of cytokines may relate to the preferential destruction of .beta.-cells in the early stage of insulin-dependent diabetes. Because class I and II MHC expression was enhanced or induced by IFN-.gamma. in both the .alpha.- and .beta.-cell lines, the ability to induce MHC molecules in itself is not sufficient to explain .beta.-cell-specific autoimmune recognition associated with insulin-dependent diabetes.