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KMID : 0354719920160040299
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Journal of Korean Diabetes Association
1992 Volume.16 No. 4 p.299 ~ p.307
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Three Dimensional Long-term Culture of Rat Islets using Gelatin Gel
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김덕윤
서광식/고은미/우정택/김성운/양인명/김진우/김영설/김광원
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Abstract
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Background:
@EN To observe the long-term effect of dynamic modulation of insulin secretion, establishment of long-term culture of islets must be an essential step. It has been known that monolayer culture of islets can maintain their function only for 3~6
weeks. In
1986, Freeman reported in vivo-like growth of human tissues in vitro by gel supported culture using specialized collagen gel manufactured from pigskin. With this culture method, diverse human tumors directly from surgery or biopsy can grow at
high
frequency in vitro for long periods of time and still maintain their in vivo properties including three-dimensional growth, maintenance of tissue organization and structure. Therefore, we tested the possibility of the long-term culture of islets
by
three-dimensional culture system using cuppporting material.
@ES Method:
@EN Thirty islets isolated from male Sprague-Dawley rat by collagenase method were put on gelatin sponge (Spongostan, Ferrosan, Denmark) per well and RPMI 1640 media with 22.2 mM glucose were changed every third day. Morphologic changes of islets
were
examined by inverted microsope and insulin secretion of islets was assayed by RIA method using rat RIA kit.
Perifusion study was also performed at 4 weeks after three-dimensional culture with RPMI 1640 containing 11.1 mM glucose to evaluate the viability and response of beta cell to glucose stimulation.
@ES Results:
@EN Initial spherical morphology of islets was well maintained after 60 days of culture and most of the observed islets were intact on inverted microscope. Insulin secretion of three-dimensional culture had maintained constantly until 4 weeks,
however
insulin secretion of monolayered islets were substantially diminished with time. Insulin secretion of islets in the sponge had maintained up to 47 days, thereafter. Perifusion study of islets showed that insulin secretion of islets was intact in
response to glucose stimulation.
@ES Conclusion:
@EN Long-term culture of islets seems to be possible by using a gelatin sponge ad supporting material for the islets and it is expected to be an improved model for the long-term investigation of islet function in vitro.
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