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In vitro proliferation method for pancreatic islet cell

A technology of islet cell and in vitro proliferation, applied in the field of in vitro culture and proliferation of islet cells, can solve problems such as the effect of beta cell efficiency, safety problems, etc., and achieve obvious effects of promoting proliferation, reducing immunogenicity, and promoting angiogenesis

Inactive Publication Date: 2007-10-17
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the above experiments generally involve a serum-free culture process, and most of them require more complicated treatments to ensure their proliferation and induce their differentiation, so the efficiency of β cell proliferation in vitro is affected to a certain extent.
At the same time, as far as we know, there has been no report so far that transplanted islet cells proliferated in vitro can effectively restore normal blood sugar levels and treat diabetes.
[0009] In the in vitro proliferation of islet cells, some research groups have adopted the method of transferring viral proto-oncogenes into β-cells to maintain the immortalization of β-cells. After transplantation, it can also play a role in treating diabetes, but the program faces great safety problems.

Method used

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  • In vitro proliferation method for pancreatic islet cell
  • In vitro proliferation method for pancreatic islet cell
  • In vitro proliferation method for pancreatic islet cell

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Example 1, bFGF stimulates the proliferation of islet cells cultured in vitro

[0060] This experiment is designed to detect whether islet cells cultured in vitro can achieve massive proliferation under the stimulation of bFGF.

[0061] As shown in Figure 2, the proliferation rate of untreated control cells gradually decreased under common in vitro culture conditions, and finally stopped growing about two weeks after isolation. In contrast, islet cells treated with bFGF can maintain an average proliferation rate of about 48 hours for at least one generation, and continue to proliferate for more than two months. In addition, islet cells stimulated by bFGF showed gradual homogenization of morphology while proliferating, that is, they showed a uniform fibroblast-like state.

[0062] This result indicated that bFGF could stimulate the proliferation of islet cells cultured in vitro, and provided a potential cell source for islet transplantation.

Embodiment 2

[0063] Example 2: Gene expression of proliferating islet cells

[0064] This experiment is designed to detect the expression of related genes in the proliferation of islet cells stimulated by bFGF.

[0065] The genes detected in this experiment include islet endocrine hormones: insulin, glucagon and glucocorticoid; pancreatic exocrine enzymes: amylase; islet progenitor marker molecule: Nestin; and islet cell isolation marker molecule: C-myc .

[0066] The time points tested in this experiment included freshly isolated islets, and days 10, 20, and 40 of in vitro culture.

[0067] As shown in Figure 3, the expression of exocrine enzymes disappeared quickly during in vitro culture, while the expression levels of endocrine hormones also gradually weakened at different rates, and the expression of insulin could still be detected at a relatively high level on the 20th day of culture. The expression level.

[0068] At the same time, the increase of Nestin expression can be detecte...

Embodiment 3

[0070] Example 3: Implantation of proliferating pancreatic islet cells can reduce blood sugar in diabetic model animals to normal levels

[0071] This experiment is designed to determine whether bFGF-treated islet cells cultured in vitro can achieve the purpose of treating diabetes by transplanting them into diabetic model rats.

[0072] The source of transplanted cells was islet cells cultured in vitro for 15 days, stimulated by bFGF to proliferate (n=10); and cultured in vitro for 15 days without treatment in vitro cultured islet cells transplanted (n=7) and phosphate buffer to place the operation (n=10) were controls.

[0073] The model animal receiving the transplantation is a diabetic rat model induced by streptozotocin, and the blood glucose is between 300 and 600 mg / dl one day before receiving the transplantation.

[0074] The transplantation route was hepatic portal vein transplantation.

[0075] The blood glucose and body weight levels of the transplanted diabetic m...

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Abstract

The present invention belongs to cell culture technical field, more specificly a islet cell in vitro proliferation method. The recombinant alkaline fibroblast growth factor is used for processing islet cell cultivated in vitro to make is proliferate for 20 generations. The proliferation time is more than two month. The islet cell keep secretion character during a certain time. The islet cell stimulated by alkaline fibroblast growth factor is transplanted diabetes rat liver via portal vein, and the diabetes symptom of rat disappear in one week. The transplanted rat keep a normal blood sugar level over two months without any immune inhibitor process.The sugar resistance experiment also prove good. The inventive islet cell is transplanted to model animal and then expresses insulin in liver and in other aspect promotes islet regeneration so achieve the aim for treating diabetes.

Description

technical field [0001] The invention belongs to the technical field of cell culture, and in particular relates to an in vitro culture and proliferation method of islet cells. Background technique [0002] Diabetes is a chronic metabolic disease caused by relative and absolute insufficiency of insulin secretion, especially abnormal blood sugar levels, which can lead to lesions of kidneys, fundus, peripheral nervous system, blood vessels and other tissues and organs in the late stage, accompanied by blindness, renal failure, etc. Risk of developing severe symptoms. It is estimated that more than 150 million people worldwide suffer from diabetes, and it is estimated that the number of diabetic patients will reach 300 million in 2025. [0003] There are two main types of diabetes: type 1 diabetes and type 2 diabetes. The former is mainly caused by the absolute deficiency of insulin, while the absolute deficiency of insulin is due to the serious shortage of the only cells in th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/06A61K35/12A61P3/10C12N5/08A61K35/39C12N5/071
Inventor 李戈卢大儒
Owner FUDAN UNIV
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