NIT-2細胞
運輸方式： 凍存運輸
數量： 大量
細胞類型： 其他細胞類型
是否是腫瘤細胞： 0
物種來源： 小鼠
年限： 10 week old
ATCC Number： CRL-2364?
相關**： 其他**
生長狀態： 貼壁生長
細胞形態： 上皮樣
器官來源： 胰腺
Designations: NIT-2
Depositors: EH Leiter
NIT-2細胞Biosafety Level: 2 [Cells contain polyomavirus DNA sequences ]
Shipped: frozen
Medium & Serum: See Propagation
Growth Properties: adherent
Organism: Mus musculus, transgenic for SV40 large T antigen deposited as mouse, transgenic for SV40 large T antigen
Morphology: epithelial
Source: Organ: pancreas
Disease: adenoma; carboxypeptidase E defective
Cell Type: beta cell;
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Age: 10 week old
Gender: male
Comments: The NIT-2 cell line was derived from the pancreatic beta cells of Cpe(fat)/Cpe(fat) mice by crossing C57BLKS/J-Cpe(fat)/+ mice with NOD/Lt-Tg(RIPTag)1Lt mice. [38844]
NIT-2細胞NOD/Lt-Tg(RIPTag)1Lt mice are transgenic for the SV40 large T antigen under the control of a rat insulin promoter, and spontaneously develop beta adenomas.
Carboxypeptidase E is required for complete conversion of proinsulin to mature insulin. A spontaneous point mutation in the coding region of the carboxypeptidase E (CPE) gene in Cpe(fat)/Cpe(fat) mice affects proinsulin processing. [38845]
The NIT-2 cell line was cultured from adenomatous islets obtained from a 10 week old F2 male and was compared with the NIT-1 cell line (ATCC -CRL-2055) previously developed from mice with wild-type CPE.
Electron microscopy of the cultured NIT-2 showed increased numbers of enlarged and electron-lucent granules compared with NIT-1 cells.
Pro-CPE, but not the mature form of CPE, is present in NIT-2 cells, and neither pro-CPE nor CPE are secreted into the medium.
Proinsulin is less extensively processed in NIT-2 than in NIT-1 cells, indicating that the Cpe(fat)mutation affects both the endopeptidase and carboxypeptidase reactions.
The secretion of insulin/proinsulin from NIT-2 cells is significantly elevated by secretagogues, indicating that CPE is not required for sorting proinsulin into the regulated pathway.
Propagation: ATCC complete growth medium: Ham's F12K medium with 2 mM L-glutamine adjusted to contain 1.5 g/L sodium bicarbonate, 90%; heat-inactivated dialyzed fetal bovine serum, 10%.
Temperature: 37.0°C
Subculturing: Subcultivation Ratio: A subcultivation ratio of 1:2 to 1:3 is recommended
Medium Renewal: Twice per week
Subcultures are prepared using a cell dissociation buffer NIT-2細胞(an enzyme free Hanks' based solution; Catalog number: 13150-016 available from GIBCO).
Remove the medium from the culture flask, add 2 ml of cell dissociation buffer per 25 sq. cm flask (5 ml per 75 sq. cm. flask and gently rock the flask at room temperature for 1 to 2 minutes to bathe the cells in the buffer.
Aspirate the solution and discard. Allow the flask to sit at room temperature for 3 to 4 additional minutes (total time from initial addition of cell dissociation buffer is approximately 5 minutes).
Firmly tap the flask against the palm of the hand to dislodge cells.
Add 5 ml of fresh medium per 25 sq. cm. flask (10 ml per 75 sq. cm. flask) and triturate up and down directing the stream along the bottom of the flask to dislodge the cells and break up some of the clumps.
Add fresh medium, aspirate and dispense into new flasks.
Preservation: Culture medium, 95%; DMSO, 5%
Related Products: Recommended medium (without the additional supplements or serum described under ATCC Medium):ATCC 30-2004
recommended serum:ATCC 30-2020
References: 38844: Varlamov O, et al. Beta-cell lines derived from transgenic Cpe(fat)/Cpe(fat) mice are defective in carboxypeptidase E and proinsulin processing. Endocrinology 138: 4883-4892, 1997. PubMed: 9348219
38845: Naggert JK, et al. NIT-2細胞Hyperproinsulinaemia in obese fat/fat mice associated with a carboxypeptidase E mutation which reduces enzyme activity. Nat. Genet. 10: 135-142, 1995. PubMed: 7663508
