caa a22 4500 477081827 CHVBK 20180405111452.0 cr unu---uuuuu 170330e19960501xx s 000 0 eng 10.1007/BF02723061 doi (NATIONALLICENCE)springer-10.1007/BF02723061 Normal rat kidney proximal tubule cells in primary and multiple subcultures [Elektronische Daten] [Pankaj Sikka, Kenneth McMartin] Summary: Anin vitro model to establish primary and subcultures of rat kidney proximal tubule (RPT) cells is described. After excising the kidneys and separating the cortex, the cortical tissue is digested with the enzyme DNAse-collagenase (Type I) resulting in a high yield of viable RPT Cells. The isolated RPT cells are then seeded onto rat tail collagen-coated surfaces and grown to confluency in a serum-free, hormonally defined medium. The cell yield can be increased by transfering the conditioned medium on Day 1 to more rat tail collagen-coated surfaces. RPT cell attachment and morphology was better on rat tail collagen-coated surfaces than on bovine collagen Type I coated surfaces. The culture medium was a 1∶1 mixture of Ham's F-12 and Dulbecco's modified Eagle's medium supplemented with bovine serum albumin, insulin, transferrin, selenium, hydrocortisone, triiodothyronine, epidermal growth factor, and glutamine. The RPT cells became confluent in 7-10 d, at which point they could be subcultured by trypsinizing and growth in the same medium. In some studies, 10 ng/ml cholera toxin was added to the culture medium. We could passage the RPT cells up to 14 times in the presence of cholera toxin. The cells were investigated for activity of several markers. The cells were histochemically positive for alkaline phosphatase and γ-glutamyl transpeptidase activity and synthesized the intermediate filament pankeratin. The RPT cells displayed apically directed sodium-dependent active glucose transport in culture. Hence, the RPT cells retain structural and functional characteristics of transporting renal epithelia in culture. This rat cell culture model will be a valuable tool for substrate uptake and nephrotoxicity studies. Society of In Vitro Biology, 1996 proximal tubule nationallicence rat tail collagen nationallicence serum-free medium nationallicence subculture nationallicence Sikka Pankaj Department of Pharmacology and Therapeutics, Louisiana State University Medical Center, 1501 Kings Highway, 71130-3932, Shreveport, Louisiana aut McMartin Kenneth Department of Pharmacology and Therapeutics, Louisiana State University Medical Center, 1501 Kings Highway, 71130-3932, Shreveport, Louisiana aut In Vitro Cellular & Developmental Biology - Animal Springer-Verlag 32/5(1996-05-01), 285-291 1071-2690 32:5<285 1996 32 11626 https://doi.org/10.1007/BF02723061 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/BF02723061 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Sikka Pankaj Department of Pharmacology and Therapeutics, Louisiana State University Medical Center, 1501 Kings Highway, 71130-3932, Shreveport, Louisiana aut NATIONALLICENCE 700 1- McMartin Kenneth Department of Pharmacology and Therapeutics, Louisiana State University Medical Center, 1501 Kings Highway, 71130-3932, Shreveport, Louisiana aut NATIONALLICENCE 773 0- In Vitro Cellular & Developmental Biology - Animal Springer-Verlag 32/5(1996-05-01), 285-291 1071-2690 32:5<285 1996 32 11626 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer