前立腺の細胞機能に関する研究 1. ラット前立腺上皮の分離と初代培養
正常 Wistar 系雄ラットの前立腺より, 比較的短期間に上皮細胞を分離し, 初代培養に成功した. ミンチした前立腺組織を0.25%トリプシン, 0.1%コラジネースの酵素処理により, 前立腺組織からの細胞分離が容易であった. さらに10%仔牛胎児血清を加えたMEM培地にて, 32℃の比較的低温に保持することにより, 線維芽細胞を抑制することができた. 培養開始後10～14日で confluent となったが, この時期の細胞について, Acid phosphatase および分泌蛋白である prostatein 染色をおこなった所, いずれも陽性所見を得た. また形態的には培養細胞は多角形...
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| Published in | 日本泌尿器科學會雑誌 Vol. 78; no. 6; pp. 1084 - 1091 |
|---|---|
| Main Authors | , , , , , , , , , , |
| Format | Journal Article |
| Language | Japanese |
| Published |
社団法人 日本泌尿器科学会
20.06.1987
The Japanese Urological Association 社団法人日本泌尿器科学会 |
| Online Access | Get full text |
| ISSN | 0021-5287 1884-7110 |
| DOI | 10.5980/jpnjurol1928.78.6_1084 |
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| Abstract | 正常 Wistar 系雄ラットの前立腺より, 比較的短期間に上皮細胞を分離し, 初代培養に成功した. ミンチした前立腺組織を0.25%トリプシン, 0.1%コラジネースの酵素処理により, 前立腺組織からの細胞分離が容易であった. さらに10%仔牛胎児血清を加えたMEM培地にて, 32℃の比較的低温に保持することにより, 線維芽細胞を抑制することができた. 培養開始後10～14日で confluent となったが, この時期の細胞について, Acid phosphatase および分泌蛋白である prostatein 染色をおこなった所, いずれも陽性所見を得た. また形態的には培養細胞は多角形で比較的薄く, 僅かに微絨毛をみとめ, 伸展した細胞質突起部分および核周囲のゴルジ装置付近に一致して多数の分泌顆粒をみとめた. 透過切片では分泌細胞の特徴である核周辺の脂肪滴, ライソゾーム, 分泌顆粒さらには滑面および粗面小胞体の発育が良好で, 自由表面に接しては空胞状の cisternae の拡張がみとめられた. 以上の成績から, 培養された細胞は前立腺の上皮細胞と思われ, 今後前立腺機能さらには発癌機序へのアプローチとして好材料となるものと考える. |
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| AbstractList | 正常Wistar系雄ラットの前立腺より,比較的短期間に上皮細胞を分離し,初代培養に成功した.ミンチした前立腺組織を0.25%トリプシン,0.1%コラジネースの酵素処理により,前立腺組織からの細胞分離が容易であった.さらに10%仔牛胎児血清を加えたMEM培地にて,32℃の比較的低温に保持することにより,線維芽細胞を抑制することができた.培養開始後10〜14日でconfluentとなったが,この時期の細胞について,Acid phosphataseおよび分泌蛋白であるprostatein染色をおこなった所,いずれも陽性所見を得た.また形態的には培養細胞は多角形で比較的薄く,僅かに徴絨毛をみとめ,伸展した細胞質突起部分および核周囲のゴルジ装置付近に一致して多数の分泌顆粒をみとめた.透過切片では分泌細胞の特徴である核周辺の脂肪滴,ライソゾーム,分泌顆粒さらには滑面および粗面小胞体の発育が良好で,自由表面に接しては空胞状のcisternaeの拡張がみとめられた.以上の成績から,培養された細胞は前立腺の上皮細胞と思われ,今後前立腺機能さらには発癌機序のアプローチとして好材料となるものと考える.
Isolation and primary culture of normal epithelial cells were established from rat ventral prostate. Minced tissue was digested with 0.25% trypsin and 0.1% collagenase in Hank's balanced solution. Thereafter, epithelial cells were easily isolated from prostatic tissue. These cells were cultured into Eagle's MEM plus 10% fetal bovine serum at 32C 5% CO_2 in air phase. After 12hours the isolated cells had attached to the bottom and grown in shape and number with time. The fibroblast cells had diminished at this low temperature after a 5〜7day culture. After 10〜14days, cultured cells had coalesced to form a confluent monolayer, which were positive for the staining with acid phosphatase and prostatein. Electron microscopic examination indicated that these cells showed a polymorphic type with secretory granules and a few microvilli on the elongated cytoplasmic processes, and joined by desmosome and tight junctions. The cells contained rough and smooth endoplasmic reticulum, Golgi apparatus near the nucleus and abundant mitochondria. In addition, intracellular blebs were observed near the surface, giving the cells an epithelial appearance and secretory function. Cultures established by our methods are being used to further study the basic function and clinical diseases of the prostate. 正常 Wistar 系雄ラットの前立腺より, 比較的短期間に上皮細胞を分離し, 初代培養に成功した. ミンチした前立腺組織を0.25%トリプシン, 0.1%コラジネースの酵素処理により, 前立腺組織からの細胞分離が容易であった. さらに10%仔牛胎児血清を加えたMEM培地にて, 32℃の比較的低温に保持することにより, 線維芽細胞を抑制することができた. 培養開始後10～14日で confluent となったが, この時期の細胞について, Acid phosphatase および分泌蛋白である prostatein 染色をおこなった所, いずれも陽性所見を得た. また形態的には培養細胞は多角形で比較的薄く, 僅かに微絨毛をみとめ, 伸展した細胞質突起部分および核周囲のゴルジ装置付近に一致して多数の分泌顆粒をみとめた. 透過切片では分泌細胞の特徴である核周辺の脂肪滴, ライソゾーム, 分泌顆粒さらには滑面および粗面小胞体の発育が良好で, 自由表面に接しては空胞状の cisternae の拡張がみとめられた. 以上の成績から, 培養された細胞は前立腺の上皮細胞と思われ, 今後前立腺機能さらには発癌機序へのアプローチとして好材料となるものと考える. |
| Author | 安川, 明広 国富, 公人 小林, 勲勇 岡, 保紀 松岡, 則良 三宅, 茂樹 安元, 章浩 武田, 祐輔 竹中, 生昌 久米, 隆 武田, 繁雄 |
| Author_FL | Miyake Shigeki Yasumoto Akihiro Kobayashi Isao Takeda Shigeo Yasukawa Akihiro Oka Yasunori Takenaka Ikumasa Kume Takashi Kunitomi Kimito Takeda Yusuke Matsuoka Noriyoshi |
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| References | 20) Chevalier, S., Bleau, G., Roberts, K. D. and Chapdelaine, A.: Characterization of canine prostatic cells from normal and hyperplastic glands. Mol. Cell Endocrinol., 20, 59-70, 1980. 2) Kaighn, M. E., Narayan, K. S. and Osawa, Y.: Androgen metabolism in human prostatic cell cultures. Tissue culture in medical research (II), Richards, R. J., ed., Pergamon Press, (Oxford), 245-252, 1983. 4) Okada, K. and Schroeder, F. H.: Human prostatic carcinoma in cell culture: Preliminary report on the development and characterization of an epithelial cell line (EB33). Urol Res., 2, 111-121, 1974. 12) Franks, L. M., Riddle, P. N., Carboneu, A. W. and Gey, G. O.: A comparative study of the ultrastructure and lack of growth capacity of adult human prostatic epithelium mechanically separated from its stroma. J. Pathol., 100, 113-119, 1970. 19) Pretlow T. G.: Disaggregation of prostates and purification of epithelial cells from normal and cancerous prostates using sedimentation in an isokinetic density gradient of Ficoll in tissue culture medium. Cancer Chemotherapy Reports, 59, 143-145, 1975. 25) 山田正篤: 細胞培養・「組織培養」39-46, 中野準之助編, 朝倉書店, 東京, 1984. 17) Lewis, R. W., Kaack, B., Roth, J. K. and Fussell, E. N.: Prostate epithelial cell culture. Application to a possible nonhuman primate model of prostate disease. Invest. Urol., 18, 251-257, 1981. 23) 山根績: 培養細胞の性格.「組織培養」236-242, 中野準之助編, 朝倉書店, 東京, 1984. 22) Tritsch, G. L. and Moore, G. F.: Spontaneous decomposition of glutamine in cell culture media. Exp. Cell Res., 28, 360-364, 1962. 7) Pontes, J. E., Pierce, J. M. Jr., Choe, B. K. and Rose, N. R.: MA 160 and EB 33 cell lines: HeLa cell contaminants, hybrids or prostatic epithelial cells? In Vitro, 15, 469-472, 1979. 21) Tung, P. S., Dorrington, J. H. and Fritz, I. B.: Structural changes induced by follicle-stimulating hormone or dibutyryl cyclic AMP on presumptive Sertoli cells in culture. Proc. Nat. Acad. Sci., 72, 1838-1842, 1975. 13) Merchant, D. J.: Prostatic tissue cell growth and assessment. Semin. Oncol., 3, 131-140, 1976. 14) Terracio, L., Douglas, W. H. J., Pennachio, D., Vena, R. L. and Ofner, P.: Primary epithelial cell cultures derived from canine prostate: Isolation, culture, and characterization. Am. J. Anat., 164, 311-332, 1982. 9) Burston, M. S.: Histochemical demonstration of phosphatases in frozen sections with naphthol AS-phosphates. J. Histochem. Cytochem., 9, 146-153, 1961. 10) Lea, O. A., Petrusz, P. and French, F. S.: Prostatein: A major secretory protein of the rat ventral prostate. J. Biol. Chemistry 254, 6169-6202, 1979. 15) Stonington, O. G., Szwec, N. and Webber, M.: Isolation and identification of the human malignant prostatic epithelial cell in pure monolayer culture. J. Urol., 114, 903-908, 1975. 16) Feuchter, F. A., Rowley, D. R. and Heidger, P. M. Jr.: Isolation, in vitro cultivation and electron microscopy of normal and malignant prostatic epithelial cells from the Copenhagen rat. Urol. Res., 8, 139-152, 1980. 1) Bolton, N. J., Lahtonen, R., Hammond, G. L. and Vihko, R.: Distribution and concentrations of androgens in epithelial and stromal compartments of the human benign hypertrophic prostate. J. Endocr., 90, 125-130, 1981. 18) 伊藤善一, 北浦宏一, 井上栄子, 中里洋一: ラット前立腺腹葉細胞株 (DPE) の樹立とその性状. 北関東医学, 31, 481-489, 1981. 11) Petrusz, P., DiMeo, P., Ordroneau, P., Weaver, C. and Keefer, D. A.: Improved immunoglobulin-enzyme bridge method for light microscopic demonstration of hormone-containing cell of the rat adenohypophysis. Histochemistry, 46, 9-26, 1975. 6) Kaighn, M. E., Narayan, K. S., Ohnuki, Y., Lechner, J. F. and Jones, L. W.: Establishment and characterization of a human prostatic carcinoma cell line (PC-3). Invest. Urol., 17, 16-23, 1979. 24) Eagle, H. and Piez, K.: The population dependent reguirement by cultured mammalian cells for metabolite which they can synthesize. J. Exp. Med., 116, 29-43, 1962. 8) Franks, L. M.: Tissue culture in the investigation of prostatic cancer. Urol. Res., 5, 159-162, 1977. 3) Fraley, E. E., Ecker, S. and Vincent, M. M.: Spontaneous in vitro neoplastic transformation of adult human prostatic epithelium. Science, 170, 540-542, 1970. 5) Stone, K. R., Mickey, D. D., Wunderli, H., Mickey, G. H. and Paulson, D. F.: Isolation of human prostate carcinoma cell line (DU 145). Int. J. Cancer, 21, 274-281, 1978. |
| References_xml | – reference: 11) Petrusz, P., DiMeo, P., Ordroneau, P., Weaver, C. and Keefer, D. A.: Improved immunoglobulin-enzyme bridge method for light microscopic demonstration of hormone-containing cell of the rat adenohypophysis. Histochemistry, 46, 9-26, 1975. – reference: 20) Chevalier, S., Bleau, G., Roberts, K. D. and Chapdelaine, A.: Characterization of canine prostatic cells from normal and hyperplastic glands. Mol. Cell Endocrinol., 20, 59-70, 1980. – reference: 18) 伊藤善一, 北浦宏一, 井上栄子, 中里洋一: ラット前立腺腹葉細胞株 (DPE) の樹立とその性状. 北関東医学, 31, 481-489, 1981. – reference: 21) Tung, P. S., Dorrington, J. H. and Fritz, I. B.: Structural changes induced by follicle-stimulating hormone or dibutyryl cyclic AMP on presumptive Sertoli cells in culture. Proc. Nat. Acad. Sci., 72, 1838-1842, 1975. – reference: 9) Burston, M. S.: Histochemical demonstration of phosphatases in frozen sections with naphthol AS-phosphates. J. Histochem. Cytochem., 9, 146-153, 1961. – reference: 7) Pontes, J. E., Pierce, J. M. Jr., Choe, B. K. and Rose, N. R.: MA 160 and EB 33 cell lines: HeLa cell contaminants, hybrids or prostatic epithelial cells? In Vitro, 15, 469-472, 1979. – reference: 10) Lea, O. A., Petrusz, P. and French, F. S.: Prostatein: A major secretory protein of the rat ventral prostate. J. Biol. Chemistry 254, 6169-6202, 1979. – reference: 15) Stonington, O. G., Szwec, N. and Webber, M.: Isolation and identification of the human malignant prostatic epithelial cell in pure monolayer culture. J. Urol., 114, 903-908, 1975. – reference: 13) Merchant, D. J.: Prostatic tissue cell growth and assessment. Semin. Oncol., 3, 131-140, 1976. – reference: 5) Stone, K. R., Mickey, D. D., Wunderli, H., Mickey, G. H. and Paulson, D. F.: Isolation of human prostate carcinoma cell line (DU 145). Int. J. Cancer, 21, 274-281, 1978. – reference: 8) Franks, L. M.: Tissue culture in the investigation of prostatic cancer. Urol. Res., 5, 159-162, 1977. – reference: 4) Okada, K. and Schroeder, F. H.: Human prostatic carcinoma in cell culture: Preliminary report on the development and characterization of an epithelial cell line (EB33). Urol Res., 2, 111-121, 1974. – reference: 14) Terracio, L., Douglas, W. H. J., Pennachio, D., Vena, R. L. and Ofner, P.: Primary epithelial cell cultures derived from canine prostate: Isolation, culture, and characterization. Am. J. Anat., 164, 311-332, 1982. – reference: 17) Lewis, R. W., Kaack, B., Roth, J. K. and Fussell, E. N.: Prostate epithelial cell culture. Application to a possible nonhuman primate model of prostate disease. Invest. Urol., 18, 251-257, 1981. – reference: 2) Kaighn, M. E., Narayan, K. S. and Osawa, Y.: Androgen metabolism in human prostatic cell cultures. Tissue culture in medical research (II), Richards, R. J., ed., Pergamon Press, (Oxford), 245-252, 1983. – reference: 19) Pretlow T. G.: Disaggregation of prostates and purification of epithelial cells from normal and cancerous prostates using sedimentation in an isokinetic density gradient of Ficoll in tissue culture medium. Cancer Chemotherapy Reports, 59, 143-145, 1975. – reference: 1) Bolton, N. J., Lahtonen, R., Hammond, G. L. and Vihko, R.: Distribution and concentrations of androgens in epithelial and stromal compartments of the human benign hypertrophic prostate. J. Endocr., 90, 125-130, 1981. – reference: 16) Feuchter, F. A., Rowley, D. R. and Heidger, P. M. Jr.: Isolation, in vitro cultivation and electron microscopy of normal and malignant prostatic epithelial cells from the Copenhagen rat. Urol. Res., 8, 139-152, 1980. – reference: 22) Tritsch, G. L. and Moore, G. F.: Spontaneous decomposition of glutamine in cell culture media. Exp. Cell Res., 28, 360-364, 1962. – reference: 12) Franks, L. M., Riddle, P. N., Carboneu, A. W. and Gey, G. O.: A comparative study of the ultrastructure and lack of growth capacity of adult human prostatic epithelium mechanically separated from its stroma. J. Pathol., 100, 113-119, 1970. – reference: 6) Kaighn, M. E., Narayan, K. S., Ohnuki, Y., Lechner, J. F. and Jones, L. W.: Establishment and characterization of a human prostatic carcinoma cell line (PC-3). Invest. Urol., 17, 16-23, 1979. – reference: 24) Eagle, H. and Piez, K.: The population dependent reguirement by cultured mammalian cells for metabolite which they can synthesize. J. Exp. Med., 116, 29-43, 1962. – reference: 25) 山田正篤: 細胞培養・「組織培養」39-46, 中野準之助編, 朝倉書店, 東京, 1984. – reference: 23) 山根績: 培養細胞の性格.「組織培養」236-242, 中野準之助編, 朝倉書店, 東京, 1984. – reference: 3) Fraley, E. E., Ecker, S. and Vincent, M. M.: Spontaneous in vitro neoplastic transformation of adult human prostatic epithelium. Science, 170, 540-542, 1970. |
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| Snippet | 正常 Wistar 系雄ラットの前立腺より, 比較的短期間に上皮細胞を分離し, 初代培養に成功した. ミンチした前立腺組織を0.25%トリプシン, 0.1%コラジネースの酵素処... 正常Wistar系雄ラットの前立腺より,比較的短期間に上皮細胞を分離し,初代培養に成功した.ミンチした前立腺組織を0.25%トリプシン,0.1%コラジネースの酵素処理により,前立... |
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| Subtitle | 1. ラット前立腺上皮の分離と初代培養 |
| Title | 前立腺の細胞機能に関する研究 |
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| ispartofPNX | 日本泌尿器科學會雑誌, 1987/06/20, Vol.78(6), pp.1084-1091 |
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