腎組織消化手冊(cè)

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Worthington組織消化指南

引言

組織消化酶主要用于組織培養(yǎng)和細(xì)胞生物學(xué)研究的原代細(xì)胞的分離和收集。盡管這些消化酶已經(jīng)普遍應(yīng)用多年，但是對(duì)其組織消化和細(xì)胞收集的作用機(jī)制卻沒有全面的理解。由此導(dǎo)致的結(jié)果便是選擇一種消化方法大多是隨機(jī)選擇或基于過去的經(jīng)驗(yàn)，而不是在*理解消化原理的基礎(chǔ)上選擇的消化方案，以及如何根據(jù)具體實(shí)驗(yàn)?zāi)康男拚桨敢垣@得更好的實(shí)驗(yàn)結(jié)果。

細(xì)胞分離程序的目的是zui大程度的獲得功能活性的單個(gè)細(xì)胞，而影響結(jié)果的原因有很多，包括（不限于）以下：

1、組織類型

2、物種來源

3、動(dòng)物年齡

4、遺傳改造情況（如knockouts）

5、所用的消化培養(yǎng)液

6、所用的酶

7、消化酶中的雜質(zhì)

8、所用消化酶的工作濃度

9、溫度

10、孵育時(shí)間

通常前四個(gè)原因沒辦法選擇，而剩下的原因一般由經(jīng)驗(yàn)決定以獲取zui適實(shí)驗(yàn)結(jié)果。

通過查閱文獻(xiàn)來選擇*組織消化酶和zui適消化條件的研究人員通常被互相矛盾的實(shí)驗(yàn)數(shù)據(jù)搞得一頭霧水。這些巨大差異來源于組織胞外基質(zhì)的復(fù)雜而動(dòng)態(tài)的特性，以及過去常用的粗制、未提純的組織分離酶。另外，組織胞外基質(zhì)由各種蛋白、糖蛋白、脂類和糖脂組成，這些成分的組成比例在不同物種、不同組織、甚至不同發(fā)育（年齡）之間存在明顯的差異，這些因素都極大的影響了消化方案的制定。目前通常使用的粗制酶，如鏈霉蛋白酶、NF1:250、以及含有幾種其他不同濃度蛋白酶的膠原酶、以及各種多糖酶、核酶和脂酶。

本指南總結(jié)了目前常用組織消化酶的zui適消化和細(xì)胞收集知識(shí)和方法，描述了標(biāo)準(zhǔn)的實(shí)驗(yàn)步驟，以便于制定一個(gè)*的細(xì)胞分離方案，同時(shí)列舉了一些相關(guān)的參考文獻(xiàn)。

下表列出多種組織的適宜消化酶（含參考文獻(xiàn)，以組織類型和物種分類，詳見鏈接）

脂肪組織	腎上腺組織	骨組織	腦組織
軟骨組織	結(jié)腸組織	內(nèi)皮組織	上皮組織
眼	心臟	腸	腎
肝	肺	淋巴結(jié)	乳腺
其它組織	其他組織	肌組織	神經(jīng)組織
胰腺	腮腺	垂體	前列腺
生殖	動(dòng)物鱗片	皮膚	脾臟
干細(xì)胞	胸腺	甲狀腺/甲狀旁腺	扁桃體
腫瘤

注：使用Worthington公司消化酶的文獻(xiàn)不限于上表鏈接中所列，一般而言，Worthington公司的組織消化酶可用于大多組織類型的消化。

腎組織消化手冊(cè)

物種	詳細(xì)來源	細(xì)胞名稱	消化用酶	培養(yǎng)基	參考文獻(xiàn)
鳥類	鳥類, 麻雀	Tubule segments	Collagenase Type 2: 0.1%	HBSS	Goldstein D., Reddy V., and Plaga K.: Second Messenger Production in Avian Medullary Nephron Segments in Response to Peptide Hormones, Am J Physiol 276, R847, 1999
鳥類	鳥類, 雞	Tubule and glomeruli	Collagenase Type 1: 0.1%	RPMI 1640	Reich, C. and Bonar, R.: Separation of Avian Kidney Tubules and Glomeruli for In Vitro Culture, Meth Cell Sci 7, 97, 1982
牛	牛	Papillary duct	Hyaluronidase: 0.2%	Keri's buffer	Husted, R., Hayashi, M., and Stokes, J.: Characteristics of Papillary Collecting Duct Cells in Primary Culture, Am J Physiol 255, F1160, 1988
牛	牛	Papillary duct	Hyaluronidase: 0.2%	HEPES buffered saline
犬	犬	Renal adipose derived cells	Collagenase Type 1: 0.3%	DMEM	Basu, J., Genheimer, C., Sangha, N., Quinlan, S., Guthrie, K., Kelley, R., Ilagan, R., Jain, D., Bertram, T. and Ludlow, J.: Organ Specific Regenerative Markers in Peri-Organ Adipose: Kidney., Lipids Health Dis 10, 171, 2011
犬	狗, 1-5 個(gè)月	Proximal tubules	Collagenase Type 1: 0.15%	Krebs Ringer bicarbonate buffer	States, B., Reynolds, R., Lee, J., and Segal, S.: Cystine Uptake By Cultured Cells Originating From Dog Proximal Tubule Segments, In Vitro Cell Dev Biol 26, 105, 1990
犬	狗, 雜種狗, 20 公斤	Proximal tubular	Deoxyribonuclease I: 0.0125%	(see reference)	Yau, C., Rao, L., and Silverman, M.: Sugar Uptake Into a Primary Culture of Dog Kidney Proximal Tubular Cells, Can J Physiol Pharmacol 63, 417, 1985
魚	美洲擬鰈 (鈍吻黃蓋鰈)	Renal tubule	Trypsin: 0.2%	CMF solution	Dickman, K. and Renfro, J.: Primary Culture of Flounder Renal Tubule Cells:Transepithelial Transport, Am J Physiol 251, 424, 1986
魚	200 - 500 克	Renal tubule	Trypsin: 0.2%	(see reference)
豚鼠	豚鼠, 雄性, 100 克	Single cells	Trypsin: 0.25%	CF salt solution	Phillips, H.: Dissociation of Single Cells from Lung or Kidney Tissue with Elastase, In Vitro 8, 101, 1972
倉鼠	倉鼠	Papillary duct	Hyaluronidase: 0.2%	Keri's buffer	Husted, R., Hayashi, M., and Stokes, J.: Characteristics of Papillary Collecting Duct Cells in Primary Culture, Am J Physiol 255, F1160, 1988
倉鼠	(包括SD大鼠, Wistar大鼠, 150-225 克, 兔, 牛)	Papillary duct	Hyaluronidase: 0.2%	HEPES buffered saline
人類	人類	Renal adipose derived cells	Collagenase Type 1: 0.3%	DMEM	Basu, J., Genheimer, C., Sangha, N., Quinlan, S., Guthrie, K., Kelley, R., Ilagan, R., Jain, D., Bertram, T. and Ludlow, J.: Organ Specific Regenerative Markers in Peri-Organ Adipose: Kidney., Lipids Health Dis 10, 171, 2011
人類	人類, 成年人	Renal tumor cells and proximal tubular epithelial	Collagenase Type 2: 0.1%	DMEM/F12	Valente, M., Henrique, R., Costa, V., Jeronimo, C., Carvalho, F., Bastos, M., Guedes de Pinho, P. and Carvalho, M.: A Rapid and Simple Procedure for the Establishment of Human Normal and Cancer Renal Primary Cell Cultures from Surgical Specimens., PLoS ONE 6, e19337, 2011
人類	人類	Renal cells	Collagenase Type 4: 300 u/ml	HBSS	Presnell, S., Bruce, A., Wallace, S., Choudhury, S., Genheimer, C., Cox, B., Guthrie, K., Werdin, E., Tatsumi-Ficht, P., Ilagan, R., Kelley, R., Rivera, E., Ludlow, J., Wagner, B., Jayo, M. and Bertram, T.: Isolation, Characterization, and Expansion Methods for Defined Primary Renal Cell Populations from Rodent, Canine, and Human Normal and Diseased Kidneys., Tiss Eng 17, 261, 2011
人類	人類	Renal cells	Neutral Protease: see reference	HBSS
人類	人類, 成年人	Renal proximal tubule and cortical fibroblasts	Collagenase Type 2: 383 u/ml	DMEM/F-12	Johnson, D., Saunders, H., Johnson, F., Huq, S., Field, M., and Pollock, C.: Cyclosporin exerts a direct fibrogenic effect on human tubulointerstitial cells: roles of insulin-like growth factor I, transforming growth factor beta1, and plaet-derived growth factor, J Pharmacol Exp Ther 289, 535-42, 1999
人類	人類	Renal cortex	Trypsin: 0.1%	Tissue Culture Grade Water	McAteer, J, Kempson, S., and Evan, A: Culture of Human Renal Cortex Epithelial Cells, J Tiss Cul Meth 13, 143, 1991
人類	人類	Papillary duct	Collagenase: 400 u/ml	Eagle's MEM-HEPES buffer w/L-glutamine	Trifillis, A. and Kahng, M.: Characterization of an In Vitro System of Human Renal Papillary Collecting Duct Cells, In Vitro Cell Dev Biol 26, 441, 1990
人類	人類	Mesangial	Trypsin: 0.25%	DMEM/Ham's F-12	Heieren, M., van der Woude, F., and Balfour Jr., H.: Cytomegalovirus Replicates Efficiently in Human Kidney Mesangial Cells, Proc Natl Acad Sci U S A 85, 1642, 1988
人類	人類, 成年人	Tubular	Collagenase: 250 u/ml	PBS	Yang, A., Gould-Kostka, J., and Oberley, T.: In Vitro Growth and Differentiation of Human Kidney Tubular Cells on a Basement Membrane Substrate, In Vitro Cell Dev Biol 23 (1), 34, 1987
人類	人類, 成年人, 14-66 歲	Tubular	Collagenase: 100 u/ml	Joklik's MEM	Trifillis, A., Regec, A., and Trump, B.: Isolation, Culture, and Charaterization of Human Renal Tubular Cells, J Urol 133, 324, 1985
人類	人類	Malignant	Papain: 0.009%	Sacks solution	Hemstreet, G., Enoch, P., and Pretlow, T.: Tissue Disaggregation of Human Renal Cell Carcinoma with Further Isopyknic and Isokinetic Gradient Purification, Cancer Res 40, 1043, 1980
人類	人類	Stromal	Papain: 0.009%	Sacks solution
人類	人類, 24 初生和夭折的	Renal	Trypsin: 0.25%	See reference	De Oca, H., and Malinin, T.: Dispersion and C*tion of Renal Cells After Short-Term Storage of Kidneys, J Clin Microbiol 2, 243, 1975
人類	(包括兔)	Renal	Trypsin: 0.25%	See reference
猴	猴	Kidney	Trypsin: 0.01% - 0.00001%	Eagle's MEM	Melnick, J., and Wallis, C.: Problems Related to the Use of Serum and Trypsin in the Growth of Monkey Kidney Cells, Dev Biol 37, 77, 1976
小鼠	小鼠	Collecting duct epithelial	Collagenase Type 2: 0.1%	DMEM	DiRocco, D., Kobayashi, A., Taketo, M., McMahon, A. and Humphreys, B.: Wnt4/尾-catenin Signaling in Medullary Kidney Myofibroblasts., J Am Soc Nephrol 24, 1399, 2013
小鼠	小鼠	Glomeruli	Collagenase Type 4: 0.1%	RPMI 1640	Kabgani, N., Grigoleit, T., Schulte, K., Sechi, A., Sauer-Lehnen, S., Tag, C., Boor, P., Kuppe, C., Warsow, G., Schordan, S., Mostertz, J., Chilukoti, R., Homuth, G., Endlich, N., Tacke, F. and Weiskirchen, R.: Primary Cultures of Glomerular Parietal Epithelial Cells or Podocytes with Proven Origin., PLoS ONE 7, e34907, 2012
小鼠	小鼠, 10-14 week	Proximal tubule	Collagenase Type 1: 0.1%	See reference	Park, J., Pasupulati, R., Feldkamp, T., Roeser, N. and Weinberg, J.: Cyclophilin D and the Mitochondrial Permeability Transition in Kidney Proximal Tubules after Hypoxic and Ischemic Injury., Am J Physiol/Renal 301, F134, 2011
小鼠	小鼠, 胚胎	Kidney	Collagenase Type 1: 0.25%	HBSS	Brown, A., Blank, U., Adams, D., Karolak, M., Fetting, J., Hill, B. and Oxburgh, L.:Isolation and Culture of Cells from the Nephrogenic Zone of the Embryonic Mouse Kidney., J Vis Exp 50, e2555, 2011
			Pancreatin: 1.0%
			Deoxyribonuclease I: 1 u/ml
小鼠	小鼠,成年的	Kidney Sphere-Derived	Collagenase Type 4: 0.1%	Iscove's Modified Dulbecco's	Huang, Y., Johnston, P., Zhang, B., Zakari, A., Chowdhry, T., Smith, R., Marban, E., Rabb, H. and Womer, K.: Kidney-Derived Stromal Cells Modulate Dendritic and T Cell Responses., J Am Soc Nephrol 20, 831-41, 2009
小鼠	小鼠, 雄性, 8-16 周	Renal tubular epithelial	Collagenase: 200 u/ml	HBSS	Breggia, A. and Himmelfarb, J.: Primary Mouse Renal Tubular Epithelial Cells have Variable Injury Tolerance to Ischemic and Chemical Mediators of Oxidative Stress., Oxid Med Cell Longev 1, 33, 2008
小鼠	小鼠, 雄性, 8-16 周	Renal tubular epithelial	Soybean Trypsin Inhibitor: see reference	HBSS
小鼠	小鼠	Proximal tubule	Collagenase Type 2: 0.013%	see reference	Wright, J., Morales, M., Sousa-Menzes, J., Ornellas, D., Sipes, J., Cui, Y., Cui, I., Hulamm, P., Cebotaru, V., Cebotaru, L., Guggino, W. and Guggino, S.: Transcriptional Adaptation to Clcn5 Knockout in Proximal Tubules of Mouse Kidney., Physiol Genomics 33, 341, 2008
小鼠	小鼠, 4 周	Endothelial kidney	Collagenase Type 1: 0.1%	DMEM	Kondo, S., Scheef, E., Sheibani, N. and Sorenson, C.: PECAM-1 Isoform-Specific Regulation of Kidney Endothelial Cell Migration and Capillary Morphogenesis., Am J Physiol Cell Physiol 292, C2070, 2007
小鼠	小鼠, 成年的	Cortex, proximal tubule	Collagenase: 0.15%	DMEM	Syal Ashu, Schiavi Susan, Chakravarty Sumana, Dwarakanath Vangipuram, Quigley Raymond, Baum Michel: Fibroblast growth factor-23 increases mouse PGE2 production in vivo and in vitro, Am J Physiol/Renal 290, F450-5, 2006
小鼠	小鼠	Kidney Mesenchymal	Collagenase: 250 u/ml	DMEM/F12	Plotkin, M. and Goligorsky, M.: Mesenchymal Cells from Adult Kidney Support Angiogenesis and Differentiate into Multiple Interstitial Cell Types including Erythropoietin-Producing Fibroblasts., Am J Physiol Renal Physiol 291, F902-12, 2006
小鼠	小鼠	Kidney Mesenchymal	Neutral Protease: 33.3 u/ml	DMEM/F12
小鼠	小鼠, 10-12 周	Kidney	Collagenase Type 4: 0.075%	DMEM	Dekel, B., Zangi, L., Shezen, E., Reich-Zeliger, S., Eventov-Friedman, S., Katchman, H., Jacob-Hirsch, J., Amariglio, N., Rechavi, G., Margalit, R. and Reisner, Y.: Isolation and Characterization of Nontubular sca-1+lin- Multipotent Stem/Progenitor Cells from Adult Mouse Kidney., J Am Soc Nephrol 17, 3300, 2006
小鼠	小鼠, 雄性, 12-16 周	Renal proximal tubule cells	Collagenase Type 2: 0.1%	DMEM/F-12	Cunningham, R., Xiaofei, E., Steplock, D., Shenolikar, S. and Weinman, E.: Defective PTH Regulation of Sodium-Dependent Phosphate Transport in NHERF-1-/- Renal Proximal Tubule Cells and Wild-Type Cells Adapted to Low-Phosphate Media., Am J Physiol Renal Physiol 289, F933, 2005
小鼠	小鼠, 雄性, 12-16 周	Renal proximal tubule cells	Soybean Trypsin Inhibitor: 0.25%	DMEM/F-12
小鼠	小鼠, 雄性, 2 個(gè)月	Cortical collecting duct	Collagenase: 54-178 u/ml	MEM	Sindic, A., Velic, A., Basoglu, C., Hirsch, J., Edemir, B., Kuhn, M. and Schlatter, E.:Uroguanylin and Guanylin Regulate Transport of Mouse Cortical Collecting Duct Independent of Guanylate Cyclase C., Kidney Int 68, 1008, 2005
小鼠	小鼠, 雄性, 2 個(gè)月	Cortical collecting duct	Protease: see reference	MEM
小鼠	小鼠, 6 周	Glomerular mesangial cells	Collagenase Type 4: 0.1%	RPMI 1640	Radeke HH, Janssen-Graalfs I, Sowa EN, Chouchakova N, Skokowa J, Loscher F, Schmidt RE, Heeringa P, Gessner JE.: Opposite regulation of type II and III receptors for immunoglobulin G in mouse glomerular mesangial cells and in the induction of anti-glomerular basement membrane (GBM) nephritis, J Biol Chem 277(30), 27535, 2002
小鼠	小鼠, 3-7 個(gè)月	Fibroblasts, mesangial, smooth muscle	Trypsin: 0.25%	DMEM	Bradshaw AD, Francki A, Motamed K, Howe C, Sage EH: Primary mesenchymal cells isolated from SPARC-null mice exhibit altered morphology and rates of proliferation, Mol Biol Cell 10, 1569-79, 1999
			Collagenase: see reference
			Soybean Trypsin Inhibitor: .05%
小鼠	小鼠	Epithelial	Collagenase Type 4: 1%	DMEM /F-12	Taub, M: Renal Tubule Cells,Cell & Tissue Culture: Laboratory Procedures Vol. 2,Doyle, A., Griffiths, J., and Newell, D., John Wiley & Sons, Ltd., 14B:6.1, 1995
小鼠	小鼠,單純的 SJL (H-2S)	Proximal tubular epithelial	Deoxyribonuclease I: 15 礸/ml	RPMI 1640	Haverty, T., Kelly, C., Hines, W., Amenta, P., Watanabe, M., Harper, R., Kefalides, N., and Neilson, E.: Characterization of a Renal Tubular Epithelial Cell Line Which Secretes the Autologous Target Antigen of Autoimmune Experimental Interstitial Nephritis, J Cell Biol 107, 1359, 1988
豬	豬, 雜種	Kidney	Collagenase Type 1: 0.1%	HBSS	Heussner, A. and Dietrich, D.: Primary Porcine Proximal Tubular Cells as an Alternative to Human Primary Renal Cells in Vitro: an Initial Characterization., BMC Cell Biol 14, 55, 2013
兔	兔	Renal proximal tubular	Collagenase Type 1: 60 u/ml	DMEM/F12	Nowak, G. and Bakajsova, D.: Assessment of Mitochondrial Functions and Cell Viability in Renal Cells Overexpressing Protein Kinase C Isozymes., J Vis Exp 71, e4301, 2013
兔	兔, 新西蘭	Proximal tubule cells	Collagenase Type 4: 0.005%	DMEM/F-12	Taub, M.: Primary Kidney Proximal Tubule Cells., Methods Mol Biol 290, 231, 2005
兔	兔, 新西蘭	Proximal tubule cells	Soybean Trypsin Inhibitor: 0.00025%	DMEM/F-12
兔	兔, 新西蘭白, 雌性	Nephron segments	Collagenase Type 2: 0.05%	Eagle's MEM	Schafer, J., Watkins, M., Li, L., Herter, P., Haxelmans, S., and Schlatter, E.: A Simplified Method for Isolation of Large Numbers of Defined Nephron Segments, Am J Physiol 273, F650, 1997
兔	兔, 新西蘭白, 雌性	Nephron segments	Deoxyribonuclease I: 50 u/ml	Eagle's MEM
兔	兔, 新西蘭白, 雄性, 1.5 - 2.0 公斤	Duct	Soybean Trypsin Inhibitor: 0.025%	Hank's Solution with calcium and HEPES	Naray-Fejes-Toth, A., and Fejes-Toth, G.: Immunoelection and Culture of Cortical Collecting Duct Cells, J Tiss Cul Meth 13, 179, 1991
兔	兔, 新西蘭掰,雌性, 2-3 公斤	Renal proximal tubules	Deoxyribonuclease I: 70 u/ml	Modified DME-F12	Rodeheaver, D., Aleo, M., and Schnellmann, R.: Differences in Enzymatic and Mechanical Isolated Rabbit Renal Proximal Tubules: Comparison in Long-Term Incubation, In Vitro Cell Dev Biol 26, 898, 1990
兔	兔	Papillary duct	Hyaluronidase: 0.2%	Keri's buffer	Husted, R., Hayashi, M., and Stokes, J.: Characteristics of Papillary Collecting Duct Cells in Primary Culture, Am J Physiol 255, F1160, 1988
兔	(包括, SD 大鼠和Wistar大鼠, 150-225克, 馬, 牛)	Papillary duct	Hyaluronidase: 0.2%	HEPES buffered saline
兔	兔, 新西蘭, 白色, 2-3 公斤	MTALH cells	Collagenase: 0.1%	DMEM	Allen, M., Nakao, A., Sonnenburg, W., Burnatowska-Hledin, M., Speilman, W., and Smith, W.: Immunodissection of Cortical and Medullary Thick Ascending Limb Cells From Rabbit Kidney, Am J Physiol 255, F704, 1988
兔	兔, 新西蘭, 白色, 2-3 公斤	RCCT cells	Collagenase: 0.1%	DMEM
兔	兔, 新西蘭, 白色	Collecting tubule	Trypsin: 0.05%	Kreb's Ringer buffer	Grenier, F., and Smith, W.: Formation of 6-keto-PGF-₁_aby Collecting Tubule Cells Isolated from Rabbit Renal Papillae, Prostaglandins 16, 759, 1978
兔	兔, 新西蘭, 白色, 雌性, 2.5 公斤	Papillary collecting duct	Collagenase: 0.2%	(see reference)	Dworzack, D., and Grantham, J.: Preparation of Renal Papillary Collecting Duct Cells for Study, Nat Neurosci 8, 191, 1975
大鼠	SD大鼠, 胎兒	Fetal kidney	Collagenase Type 4: 0.1%	MEM	Gupta, A., Jadhav, S., Tripathy, N. and Nityanand, S.: Fetal Kidney Cells Can Ameliorate Ischemic Acute Renal Failure in Rats through Their Anti-Inflammatory, Anti-Apoptotic and Anti-Oxidative Effects., PLoS ONE 10, e0131057, 2015
大鼠	Lewis大鼠, 雄性	Renal adipose derived cells	Collagenase Type 1: 0.3%	DMEM	Basu, J., Genheimer, C., Sangha, N., Quinlan, S., Guthrie, K., Kelley, R., Ilagan, R., Jain, D., Bertram, T. and Ludlow, J.: Organ Specific Regenerative Markers in Peri-Organ Adipose: Kidney., Lipids Health Dis 10, 171, 2011
大鼠	大鼠	Proximal tubule	Collagenase Type 2: 0.1%	see reference	Panico, C., Luo, Z., Damiano, S., Artigiano, F., Gill, P. and Welch, W.: Renal Proximal Tubular Reabsorption is Reduced in Adult Spontaneously Hypertensive Rats: Roles of Superoxide and Na+/H+ Exchanger 3., Hypertension 54, 1291, 2009
大鼠	SD大鼠, 雄性, 200-250克	Medullary thick ascending limb	Collagenase: 0.1%	HEPES-saline	Silva, G., Beierwaltes, W., and Garvin, J.: Extracellular ATP stimulates NO production in rat thick ascending limb, Hypertension 47, 563-7, 2006
大鼠	Wistar大鼠, 300克	Renal proximal tubules	Collagenase Type 2: 0.2%	DMEM/F12	Deng Aihua, Miracle Cynthia M, Lortie Mark, Satriano Joseph, Gabbai Francis B, Munger Karen A, Thomson Scott C, Blantz Roland C: Kidney oxygen consumption, carbonic anhydrase, and proton secretion, Am J Physiol/Renal 290, F1009-15, 2006
大鼠	大鼠, Wistar, 雄性, 300-350 克	Renal proximal tubules	Collagenase Type 2: 0.2%	DMEM/F-12	Deng, A., Miracle, C., Suarez, J., Lortie, M., Satriano, J., Thomson, S., Munger, K. and Blantz, R.: Oxygen Consumption in the Kidney: Effects of Nitric Oxide Synthase Isoforms and Angiotensin II., Kidney Int 68, 723, 2005
大鼠	大鼠	Cortical tubule cells	Collagenase: 0.6%	DMEM	Arystarkhova, E., Wetzel R., and Sweadner K.: Distribution and Oligomeric Association of Splice Forms of Na(+)-K(+)-ATPase Regulatory Gamma-subunit in Rat Kidney, Am J Physiol/Renal 282(3), F393, 2002
大鼠	SD大鼠, 雄性 275-300 克	Glomerular mesangial cells	Collagenase Type 1: 250 u/ml	DMEM	Amiri Farhad, Shaw Sean, Wang Xiaodan, Tang Jie, Waller Jennifer L, Eaton Douglas C, Marrero Mario B: Angiotensin II activation of the JAK/STAT pathway in mesangial cells is altered by high glucose, Kidney Int Suppl 61, 1605-16, 2002
大鼠	SD大鼠, 1 周	Cortical collection duct	Collagenase Type 2: 0.15%	DMEM/Ham's F12	Valencia L., Bidet M., Martial S., Sanchez E., Melendez E., Tauc M., Poujeol C., Martin D., Namorado M., Reyes J., and Poujeol P.: Nifedipine-activated Ca(2+) Permeability in Newborn Rat Cortical Collecting Duct Cells in Primary Culture, Am J Physiol Cell Physiol280(5), C1193, 2001
大鼠	SD大鼠, 250-300克	Microvessels	Collagenase Type 2: 0.1%	PSS	Li, N., Teggatz, E., Li, P., Allaire, R., and Zou, A.: Formation and Actions of Cyclic ADP-Ribose in Renal Microvessels, Microvasc Res 60, 149, 2000
大鼠	SD大鼠, 250-300 克	Renal	Collagenase Type 2: 0.2%	see reference	Mattson D., and Wu F.: Nitric Oxide Synthase Activity and Isoforms in Rat Renal Vasculature, Hypertension 35, 337, 2000
大鼠	SD大鼠, 雄性, 5-6 周	Renal tubules	Collagenase: 0.1%	see reference	Miyata, N., Park, F., Li, X., and Cowley, A.: Distribution of Angiotensin AT1 and AT2 Receptor Subtypes in the Rat Kidney, Am J Physiol 277, F437, 1999
大鼠	SD大鼠, 225-250 g	Glomerular mesangial cells	Collagenase Type 1: 250 u/ml	DMEM	Amiri F, Garcia R: Regulation of angiotensin II receptors and PKC isoforms by glucose in rat mesangial cells, Am J Physiol 276, F691-9, 1999
大鼠	SD大鼠, 雄性	Proximal tubule suspensions	Collagenase Type 4: 0.1%	HEPES buffer	Eitle E, Hiranyachattada S, Wang H, Harris PJ: Inhibition of proximal tubular fluid absorption by nitric oxide and atrial natriuretic peptide in rat kidney, Am J Physiol 274, C1075-80, 1998
大鼠	SD大鼠, 雄性	Proximal tubule suspensions	Pronase E: 2 u/ml	HEPES buffer
大鼠	大鼠, SD, 雄性	Nephron segments	Collagenase Type 2: 0.05%	Eagle's MEM	Schafer, J., Watkins, M., Li, L., Herter, P., Haxelmans, S., and Schlatter, E.: A Simplified Method for Isolation of Large Numbers of Defined Nephron Segments, Am J Physiol 273, F650, 1997
大鼠	大鼠, SD, 雄性	Nephron segments	Deoxyribonuclease I: 50 u/ml	Eagle's MEM
大鼠	大鼠, 12-周大	Renal	Collagenase: 0.1%	DMEM	Ishikawa, S., Kusaka, I., Higashiyama, M., Nagasaka, S., Saito, T., Honda, K., and Saito, T.: Cellular Signaling and Proliferative Action of AVP in Mesangium of SHR: Effect of Low Density Lipoprotein, Nat Neurosci 50, 1506, 1996
大鼠	大鼠, 雄性, 150-200 克	Epithelial	Protease: 0.1%	HBSS/CMF	Elliget, K., and Trump, B.: Primary Cultures of Normal Rat Kidney Proximal Tubule Epithelial Cells for Studies of Renal Cell Injury, In Vitro Cell Dev Biol 27, 739, 1991
大鼠	大鼠, 雄性, 150-200 克	Proximal tubule	Protease: 0.1%	HBSS/CMF
大鼠	大鼠, 成年的	Inner medullary collecting duct	Collagenase Type 2: 0.1%	PBS	Brion, L., Schwartz, J., Lachman, H., Zavilowitz, B., and Schwartz, G.: Development of H+ Secretion by Cultured Renal Inner Medullary Collecting Duct Cells, Am J Physiol 257, F486, 1989
大鼠	大鼠, 成年的	Papillae	Collagenase Type 2: 0.1%	PBS
大鼠	Wistar大鼠, 雌性, 100-150 克	Glomerular mesangial	Collagenase: 0.025%	RPMI 1640	Wang, J., Kester, M., and Dunn, M.: The Effects of Endotoxin on Plaet-Activating Factor Synthesis in Cultured Rat Glomerular Mesangila Cells, Biochim Biophys Acta 969, 217, 1988
大鼠	Wistar大鼠, 雄性, 150 - 200 克	Renal target	Collagenase: 1.0% (also 0.7%)	Eagle's MEM	Barlet-Bas, C., Khadouri, C., Marsey, S., and Doucet, A.: Sodium-Independent In VitroInduction of Na+,K+-ATPase by Aldosterone in Renal Target Cells: Permissive Effect of Triiodothyronine, Proc Natl Acad Sci U S A 85, 1707, 1988
大鼠	Wistar大鼠, 雄性, 成年的	Proximal tubules	Collagenase: 0.2%	Krebs-Henseleit buffer	Gesek, F., Wolff, D., and Strandhov, J.: Improved Separation Method for Rat Proximal and Distal Renal Tubules, Am J Physiol 253, F358, 1987
大鼠	Wistar大鼠, 300-400克	Proximal tubules	Collagenase: 0.15 %	Krebs Henseleit solution	Vinay, P., Gougoux, A., and Lemieux, G.: Isolation of a Pure Suspension of Rat Proximal Tubules, Am J Physiol 241, F403, 1981
大鼠	大鼠	Fibroblasts	Trypsin: up to 0.25%	Dulbecco-Vogt MEM	Wallach, D., Anderson, W., and Pastan, I.: Activation of Adenylate Cyclase in Cultured Fibroblasts by Trypsin, J Biol Chem 253, 24, 1978
大鼠	大鼠	Kidney	Trypsin: up to 0.25%	Dulbecco-Vogt MEM

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