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์ง€๋ฐฉ์ค„๊ธฐ์„ธํฌ: ๋ถ„๋ฆฌ, ๋ฐฐ์–‘, ๋ถ„ํ™”

— 2015-05-29

 

Adipose derived stem cells(ADSCs): Isolation, expansion and differentiation ์ง€๋ฐฉ์œ ๋ž˜์ค„๊ธฐ์„ธํฌ : ย ๋ถ„๋ฆฌ, ํ™•์žฅ(๋ฐฐ์–‘), ๋ถ„ํ™”(2008)

 

์ด ์—ฐ๊ตฌ ๋…ผ๋ฌธ์€ ์ง€๋ฐฉ์œ ๋ž˜ ์ค„๊ธฐ์„ธํฌ์˜ ๋ฌดํ•œํ•œ ๊ฐ€๋Šฅ์„ฑ์„ ๋ณด์—ฌ์ค๋‹ˆ๋‹ค. ๊ทธ๋ž˜์„œ ์ „์„ธ๊ณ„์˜ ์žฌ์ƒ์˜ํ•™ํšŒ๋“ค์ด ์ง€๋ฐฉ์ค„๊ธฐ์„ธํฌ๋ฅผ ์—ฐ๊ตฌํ•˜๊ณ  ์žˆ์Šต๋‹ˆ๋‹ค.

 

1. ์†Œ๊ฐœ

 

์ •์˜ํ•˜์ž๋ฉด, ์ค„๊ธฐ์„ธํฌ๋Š” ์ž๊ฐ€์žฌ์ƒ๋Šฅ๋ ฅ๊ณผ ๋‹ค๋ถ„ํ™”๊ฐ€ ๊ฐ€๋Šฅํ•˜๊ณ , ์ตœ์ข…์˜ ์„ธํฌ๋กœ๋„ ํ˜•์„ฑ์ด ๊ฐ€๋Šฅํ•ฉ๋‹ˆ๋‹ค. ์ด์ƒ์ ์œผ๋กœ ์žฌ์ƒ์˜ํ•™์˜ ์ ์šฉ์„ ์œ„ํ•œ ์ค„๊ธฐ์„ธํฌ๋Š” ๋‹ค์Œ ๊ธฐ์ค€๋“ค์„ ์ถฉ์กฑํ•ด์•ผ ๊ฐ€๋Šฅํ•ฉ๋‹ˆ๋‹ค. (i) ์ˆ˜๋ฐฑ๋งŒ์—์„œ ์ˆ˜์‹ญ์–ต๊นŒ์ง€์˜ ํ’๋ถ€ํ•œ ์ˆ˜์˜ ์„ธํฌ๊ฐ€ ํ™•๋ณด๋˜์–ด์•ผ ํ•ฉ๋‹ˆ๋‹ค. (ii) ์ตœ์†Œ์˜ ์นจ์Šต ๊ณผ์ •์— ์˜ํ•ด ์ˆ˜์ง‘๋˜๊ณ  ์ˆ˜ํ™•๋  ์ˆ˜ ์žˆ์–ด์•ผ ํ•ฉ๋‹ˆ๋‹ค. (iii) ์žฌํ˜„๋ฐฉ์‹์— ๋”ฐ๋ผ ๋‹ค์–‘ํ•œ ์„ธํฌ์ฃผ ๊ฒฝ๋กœ์— ๋”ฐ๋ผ ๋ถ„ํ™”๋  ์ˆ˜ ์žˆ์–ด์•ผ ํ•ฉ๋‹ˆ๋‹ค. (iv) ์ž๊ฐ€ ๋˜๋Š” ๋™์ข…์— ์•ˆ์ „ํ•˜๊ณ  ํšจ๊ณผ์ ์œผ๋กœ ์ด์‹์ด ๊ฐ€๋Šฅํ•ด์•ผ ํ•ฉ๋‹ˆ๋‹ค.

 

2. ์ง€๋ฐฉ์„ธํฌ์—์„œ ์ค‘๊ฐ„์—ฝ์ค„๊ธฐ์„ธํฌ์˜ ๋ถ„๋ฆฌ ๋ฐฉ๋ฒ•

 

adsc_differentiation_1
Fig. 1. Scheme for processing of adipose tissue and isolation of adipose derived stem cells.
(SVF). ์ดˆ๊ธฐ ๋ฐฉ๋ฒ•์€ ์ง€๋ฐฉ ์กฐ์ง์œผ๋กœ๋ถ€ํ„ฐ ์„ธํฌ๋ฅผ ๋ถ„๋ฆฌํ•˜๋Š” ๋ฐฉ๋ฒ•์œผ๋กœ 1960๋…„๋Œ€์— Rodbell๊ณผ ๊ทธ ๋™๋ฃŒ๋“ค์— ์˜ํ•ด ๋ฐœ๊ฒฌ๋˜์—ˆ์Šต๋‹ˆ๋‹ค. ๊ทธ๋“ค์€ ์ฅ์˜ ์ง€๋ฐฉ์ฒด๋ฅผ ๋ถ„๋ฆฌํ•˜์—ฌ, ์˜ค์—ผ๋œ ์กฐํ˜ˆ ์„ธํฌ๋ฅผ ์ œ์–ดํ•˜๊ธฐ์œ„ํ•ด ๊ด‘๋ฒ”์œ„ํ•˜๊ฒŒ ์›Œ์‹ฑ์„ ํ•˜์˜€์Šต๋‹ˆ๋‹ค. ๊ทธ๋ฆฌ๊ณ  collagenase(๋ถ„ํ•ด์งˆ ํšจ์†Œ)๋กœ ์กฐ์ง์˜ ํŒŒํŽธ๋“ค์„ ๋ฐฐ์–‘ํ•˜์—ฌ ํ•ต์‹ฌ๋งŒ์„ ์–ป๊ธฐ์œ„ํ•ด ์›์‹ฌ๋ถ„๋ฆฌ๊ธฐ๋ฅผ ํ–‰ํ•œ๋‹ค. ๊ทธ๋กœ์จ ๋ญ‰์ณ์ง„ SVF( stromal vascular fraction :ย ๊ธฐ์งˆ ํ˜ˆ๊ด€ ์ผ๋ถ€)๋กœ ๋ถ€ํ„ฐ ์„ฑ์ˆ™ํ•œ ์ง€๋ฐฉ์„ธํฌ๋ฅผ ์œ ๋™์ ์œผ๋กœ ๋ถ„๋ฆฌํ•ฉ๋‹ˆ๋‹ค.



 

 

3. ์ง€๋ฐฉ์œ ๋ž˜ ์ค„๊ธฐ์„ธํฌ์˜ ๋ฐฐ์–‘๊ณผ ํ™•์žฅ

adsc_differentiation_3

Fig. 2. Multilineage differentiation potential for adipose derived stem cells.

4. ์ง€๋ฐฉ์œ ๋ž˜์ค„๊ธฐ์„ธํฌ์˜ ๋ƒ‰๋™๊ณผ ์žฅ๊ธฐ๋ณด๊ด€

 

ADSCs๋Š” ๋™๊ฒฐ์‹œํ‚ค๊ธฐ ์œ„ํ•ด 80 %์˜ ํ•ฉ๋ฅ˜์ ์— ์˜ํ•ด ์ˆ˜ํ™•๋˜์–ด์•ผ ํ•ฉ๋‹ˆ๋‹ค. ์„ธํฌ๋ฅผ ์ˆ˜์ง‘ํ•˜๊ธฐ ์œ„ํ•ด, ๋ฐฐ์–‘ ๋ฐฐ์ง€๋ฅผ ์ œ๊ฑฐํ•˜๊ณ  ๋ฉธ๊ท  ๋”ฐ๋œปํ•œ PBS์˜ ์†Œ๋Ÿ‰์œผ๋กœ ๋Œ€์ฒด.PBS๋ฅผ ์ œ๊ฑฐํ•˜๊ณ  ํŠธ๋ฆฝ์‹  EDTA ์šฉ์•ก์œผ๋กœ ๊ต์ฒดํ•ฉ๋‹ˆ๋‹ค. ……… ๋ƒ‰๋™๋œ Vials์„ ์žฅ๊ธฐ ์ €์žฅํ•˜๊ธฐ ์œ„ํ•ด ์•ก์ฒด์งˆ์†Œ์šฉ๊ธฐ๋กœ ์ด๋™์‹œํ‚ต๋‹ˆ๋‹ค. ์„ธํฌ ํŠœ๋ธŒ๋Š” ํ•ญ์ƒ LN2 ํƒฑํฌ์˜ ์ฆ๊ธฐ์ƒ์— ์ €์žฅ๋˜์–ด์•ผ ํ•ฉ๋‹ˆ๋‹ค.

 

5. ์ง€๋ฐฉ์œ ๋ž˜์ค„๊ธฐ์„ธํฌ์˜ ๊ฐ„์—ฝ์„ฑ ๋ถ„ํ™” ๋ถ„์„

 

5.1. ์ง€๋ฐฉ์„ธํฌํ™” ๋ถ„ํ™”(=์ง€๋ฐฉ์„ธํฌ๋กœ์˜ ๋ถ„ํ™”๊ฐ€๋Šฅ)
5.2. ๊ณจํ˜•์„ฑ์˜ ๋ถ„ํ™”(=๋ผˆ๋กœ์˜ ๋ถ„ํ™”๊ฐ€๋Šฅ)
5.3. ์—ฐ๊ณจ์›์„ฑ์˜ ๋ถ„ํ™”(=์—ฐ๊ณจ์กฐ์ง์œผ๋กœ์˜ ๋ถ„ํ™”๊ฐ€๋Šฅ)

adsc_differentiation_2
Fig. 3. Mesenchymal lineage differentiation assays for adipose-derived stem cells.

 

5.4. ์‹ ๊ฒฝ์œผ๋กœ์„œ ๋ถ„ํ™”(=์‹ ๊ฒฝ์„ธํฌ๋กœ์˜ ๋ถ„ํ™”๊ฐ€๋Šฅ)

 

7. ๊ฒฐ๋ก 

 

์ง€๋ฐฉ์œ ๋ž˜์ค„๊ธฐ์„ธํฌ(ADSCs)๋Š” ์œ ์ „์  ์งˆ๋ณ‘ ๋˜๋Š” ํš๋“ ์งˆ๋ณ‘๋“ค์˜ ๊ด‘๋Œ€ํ•œ ๋ฒ”์œ„์— ๋Œ€ํ•ด, ์ƒˆ๋กญ๊ฒŒ ์น˜๋ฃŒํ•  ์ˆ˜ ์žˆ๋Š” ์—ฐ๊ตฌ ๊ธฐํšŒ๋ฅผ ์ œ๊ณตํ•ฉ๋‹ˆ๋‹ค. ๋˜ํ•œ ADSCs๋Š” ์•ฝ๋ฌผ ๋ฐœ๊ฒฌ์„ ์œ„ํ•œ ์ƒˆ๋กœ์šด ๋ถ„์ž ํ‘œ์ ์„ ์‹๋ณ„ํ•  ์ˆ˜ ์žˆ๋Š” ๊ธฐํšŒ๋ฅผ ์ œ๊ณตํ•ฉ๋‹ˆ๋‹ค. ์ด ์—ฐ๊ตฌ์—์„œ, ์ง€๋ฐฉ์œ ๋ž˜์ค„๊ธฐ์„ธํฌ์˜ ๋ถ„๋ฆฌ, ํŠน์„ฑ, ํ™•์žฅ์„ ์ฐจ๋ณ„ํ™” ํ•  ์ˆ˜ ์žˆ๋Š” ํ”„๋กœํ† ์ฝœ์„ ์ œ๊ณตํ•ฉ๋‹ˆ๋‹ค. ์—ฐ๊ตฌ์ž๋ผ๋ฉด ๋ณด๋‹ค ๋…ผ๋ฌธ์„ ์ž์„ธํžˆ ๋ณด์‹œ๊ธฐ ๋ฐ”๋ž๋‹ˆ๋‹ค. ์ด๋Ÿฌํ•œ ํ”„๋กœํ† ์ฝœ๋“ค์€ ์ง€๋ฐฉ ์กฐ์ง ์ƒ˜ํ”Œ์˜ ํฌ๊ธฐ์˜ ์ฐจ์ด๋ฅผ ์šฉ์ดํ•˜๊ฒŒ ์กฐ์ •ํ•˜๋„๋ก ๊ตฌ์„ฑ๋  ์ˆ˜ ์žˆ์Šต๋‹ˆ๋‹ค. ์ง€๋ฐฉ์ค„๊ธฐ์„ธํฌ๋ฅผ ์ธ๊ฐ„์—๊ฒŒ ์ž„์ƒ์ ์šฉํ•˜๊ธฐ ์ „์— ๋งŽ์€ ์ค‘์š”ํ•œ ๊ณผํ•™๊ณผ ์˜๋ฃŒ์˜ ์งˆ๋ฌธ๋“ค์ด ์žˆ์Šต๋‹ˆ๋‹ค. ์—ฌ๊ธฐ์—๋Š” ํ˜„์žฌ์˜ GMP๋ฅผ ์ค€์ˆ˜ํ•˜์—ฌ ์„ธํฌ๋ฅผ ์ƒ์„ฑํ•˜๊ธฐ ์œ„ํ•ด ์ ์ ˆํ•œ ํ’ˆ์งˆ ๊ด€๋ฆฌ ๋ฐ ๋Œ€๊ทœ๋ชจ์˜ ์ œ์กฐ ๋ฐฉ๋ฒ•์˜ ๊ฐœ๋ฐœ์„ ํฌํ•จํ•ฉ๋‹ˆ๋‹ค.

 

Source : www.ncbi.nlm.nih.gov/pubmed/18593609
www.elsevier.com/locate/ymeth
Adipose-derived stem cells: Isolation, expansion and differentiation(2008)



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