大鼠肝臟Kupffer細胞的分離、純化_《中國普外基礎與臨床雜志》

作者：

蔡偉 , 李非

首都醫科大學北京宣武醫院普外科（北京100053）;

關鍵詞：

Kupffer細胞細胞分離純化

視頻：

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目的　建立簡單、有效而穩定的大鼠肝臟Kupffer細胞分離、純化的實驗方法。
方法　①大鼠肝臟的膠原酶離體灌注； ②Percoll液不連續梯度離心分離Kupffer細胞； ③選擇性貼壁純化Kupffer細胞； ④吞噬墨汁實驗、DAB染色，光鏡、電鏡下觀察鑒定Kupffer細胞。
結果　最終獲得的大鼠肝臟Kupffer細胞活性 gt;90%，純度為95%，光鏡下Kupffer細胞具有較強的貼壁及吞噬碳粒能力，DAB染色呈黃色煎蛋樣結構；新分離的Kupffer細胞呈圓形，形態大小一致； 2 d后，細胞發生伸展，呈多角形或星形。電鏡下觀察可見典型特點：有偽足及杯口狀改變，有豐富的溶酶體和吞噬體。
結論　本實驗建立的大鼠肝臟Kupffer細胞的分離培養方法效果穩定，同時培養的細胞具有良好的生物學性狀。

引用本文： 蔡偉,李非. 大鼠肝臟Kupffer細胞的分離、純化. 中國普外基礎與臨床雜志, 2007, 14(3): 292-295. doi: 復制

1.	Burt AD, Le Bail B, Balabaud C, et al. Morphologic investigation of sinusoidal cells [J]. Semin Liver Dis, 1993; 13(1)∶21.
2.	Seglen PO. Preparation of rat liver cells. 3. Enzymatic requirements for tissue dispersion [J]. Exp Cell Res, 1973; 82(2)∶391.
3.	Smedsrod B, Pertoft H. Preparation of pure hepatocytes and re-ticuloendothelial cells in high yield from a single rat liver by means of Percoll centrifugation and selective adherence [J]. J Leukoc Biol, 1985; 38(2)∶213.
4.	韓偉，成令忠，顧云娣. 大鼠肝枯否細胞分離培養與免疫活性的研究 [J]. 解剖學雜志, 1988; 11（2）∶71.
5.	Heuff G, Steenbergen JJ, Van de Loosdrecht AA, et al. Isolation of cytotoxic Kupffer cells by a modified enzymatic assay: a methodological study [J]. J Immunol Methods, 1993; 159(1-2)∶115.
6.	Knook DL, Seffelaar AM, de Leeuw AM. Fat-storing cells of the rat liver. Their isolation and purification [J]. Exp Cell Res, 1982; 139(2)∶468.
7.	Ikejima K, Enomoto N, Seabra V, et al. Pronase destroys the lipopolysaccharide receptor CD14 on Kupffer cells [J]. Am J Physiol, 1999; 276(3 Pt 1)∶G591.
8.	Smedsrod B, Pertoft H. Preparation of pure hepatocytes and reticuloendothelial cells in high yield from a single rat liver by means of Percoll centrifugation and selective adherence [J]. J Leukoc Biol, 1985; 38(2)∶213.
9.	高春芳，孔憲濤，范列英. 大鼠肝貯脂細胞、 Kupffer細胞的分離、培養和鑒定 [J]. 中華消化雜志， 1995; 15(3)∶142.
10.	Nagelkerke JF, Barto KP, van Berkel TJ. In vivo and in vitro uptake and degradation of acetylated low density lipoprotein by rat liver endothelial, Kupffer, and parenchymal cells [J]. J Biol Chem, 1983; 258(20)∶12221.
11.	Valatas V, Xidakis C, Roumpaki H, et al. Isolation of rat Kupffer cells: a combined methodology for highly purified primary cultures [J]. Cell Biol Int, 2003; 27(1)∶67.
12.	成令忠主編. 組織學 [M]. 第2版. 北京: 人民衛生出版社, 1993∶1200~1209.

1. Burt AD, Le Bail B, Balabaud C, et al. Morphologic investigation of sinusoidal cells [J]. Semin Liver Dis, 1993; 13(1)∶21.
2. Seglen PO. Preparation of rat liver cells. 3. Enzymatic requirements for tissue dispersion [J]. Exp Cell Res, 1973; 82(2)∶391.
3. Smedsrod B, Pertoft H. Preparation of pure hepatocytes and re-ticuloendothelial cells in high yield from a single rat liver by means of Percoll centrifugation and selective adherence [J]. J Leukoc Biol, 1985; 38(2)∶213.
4. 韓偉，成令忠，顧云娣. 大鼠肝枯否細胞分離培養與免疫活性的研究 [J]. 解剖學雜志, 1988; 11（2）∶71.
5. Heuff G, Steenbergen JJ, Van de Loosdrecht AA, et al. Isolation of cytotoxic Kupffer cells by a modified enzymatic assay: a methodological study [J]. J Immunol Methods, 1993; 159(1-2)∶115.
6. Knook DL, Seffelaar AM, de Leeuw AM. Fat-storing cells of the rat liver. Their isolation and purification [J]. Exp Cell Res, 1982; 139(2)∶468.
7. Ikejima K, Enomoto N, Seabra V, et al. Pronase destroys the lipopolysaccharide receptor CD14 on Kupffer cells [J]. Am J Physiol, 1999; 276(3 Pt 1)∶G591.
8. Smedsrod B, Pertoft H. Preparation of pure hepatocytes and reticuloendothelial cells in high yield from a single rat liver by means of Percoll centrifugation and selective adherence [J]. J Leukoc Biol, 1985; 38(2)∶213.
9. 高春芳，孔憲濤，范列英. 大鼠肝貯脂細胞、 Kupffer細胞的分離、培養和鑒定 [J]. 中華消化雜志， 1995; 15(3)∶142.
10. Nagelkerke JF, Barto KP, van Berkel TJ. In vivo and in vitro uptake and degradation of acetylated low density lipoprotein by rat liver endothelial, Kupffer, and parenchymal cells [J]. J Biol Chem, 1983; 258(20)∶12221.
11. Valatas V, Xidakis C, Roumpaki H, et al. Isolation of rat Kupffer cells: a combined methodology for highly purified primary cultures [J]. Cell Biol Int, 2003; 27(1)∶67.
12. 成令忠主編. 組織學 [M]. 第2版. 北京: 人民衛生出版社, 1993∶1200~1209.

《中國普外基礎與臨床雜志》

大鼠肝臟Kupffer細胞的分離、純化

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《中國普外基礎與臨床雜志》

大鼠肝臟Kupffer細胞的分離、純化

摘要 全文 圖表 視頻 參考文獻 施引文獻 補充材料

Format

Content

摘要全文圖表視頻參考文獻施引文獻補充材料