Calcium sulfate와 혈소판 유래성장인자의 혼합사용이 치주인대세포에 미치는 영향

Abstract

[한글]

생체 내 골이식재제와 흡수성 차단막재료로 사용할 수 있는 calcium sulfate가 치주인대세포에 미치는 영향과 세포의 성장, 형성기능을 조절하는 성장인자의 Vehicle로서 사용가능성이 있는지를 알아보고자 치주인대세포를 10% FBS가 함유된 α-MEM에서 배양한 군을 대조군으로 하고 동일한 배지조건 하에서 calcium sulfate를 넣어서 배양한 군(calcium sulfate군), calcium sulfate에 15ng/㎖ 농도의 PDGF를 주입하여 배양한 군(calcium sulfate+POGF군), α-MEM에 15ng/㎖ 농도의 PDGF를 주입하여 배양한 군(PDGF군)을 실험군으로 하여 세포배양 1, 2, 3일에 각군의 세포수 산정, MTT assay, 교원질 합성능을 측정하여 다음과 같은 결론을 얻었다.

1. 세포수 산정에 의한 세포증식에 있어서 calcium sulfate군과 calcium sulfate+PDGF군은 대조군과 비교시 1, 2, 3일에 유의성있는 차이가 없었고 PDGF군은 calcium sulfate군에 비해서 1, 2일에 유의성있는 차이가 있었다(P<0.05).

2. 용출액을 이용한 MTT assay에 의한 세포증식에 있어서 calcium sulfate군과 calcium sulfate+PDGF군은 대조군과 비교시 1, 2, 3일에 유의성있는 차이가 없었고 PDGF군은 calcium sulfate군에 비해서 2, 3일에 유의성있는 차이가 있었으며 calcium sulfate+PDGF군은 calcium sulfate군에 비해 2일에 유의성있는 차이가 있었다(P<0.05).

3. Transwell을 이용한 MTT assay에 의한 세포증식에 있어서 calcium sulfate군과 calcium sulfate+PDGF군은 대조군과 PDGF군과 비교시 1일에는 유의성있는 차이가 있었으나 2, 3일에는 유의성있는 차이가 없었다(P<0.05).

4. Immunoblotting assay에 의한 교원질합성능에 있어서 calcium sulfate군은 3일에, calcium sulfate+PDGF군은 1일에, PDGF군은 2일에 높게 나타났다.

이상의 결과에서 볼 때 calcium sulfate는 치주인대세포에 유해한 독성반응을 나타내지 않는 생체적합성이 입증되었으며 PDGF같은 성장인자의 주입시 Vehicle의 역할 가능성을 보여주고 있으므로 치주조직 재생에 있어서 생체재료로서의 가치가 있음을 시사해주고 있

다.

[영문]

It was well known that calcium sulfate was biocompatible, resorbed rapidly in the body, had potential as a good barrier membrane. Platelet-derived growth factor(PDGF) was one of polypeptide growth factor that had been reported as a biological mediator which regulates activities of wound healing process including the cell proliferation, migration and metabolism.

The purpose of this study was to evaluate the effects of a combination of calcium sulfate and PDGF on periodontal ligament cells in vitro to use as a regeneration promoting agent of periodontal tissue. Human periodontal ligament cells were prepared from the premolar tooth extracted for the orthodontic treatment. Cells were cultured in α-MEM contained with 20% FBS, at the 37℃, 100% of humidity, 5% Co^^2 incubator. Cells were inoculated and cultured into 96 well culture plate with 1×10**4cells/well of α-MEM for 1 day.

After discarding the medium, those cells were cultured in α-MEM contained with 10% FBS alone(control group), in calcium sulfate(calcium sulfate group), in calcium sulfate treated with 15ng/㎖ of PDGF-BB(calcium sulfate+PDGF group), in α-MEM contained with 10% FBS treated with 15ng/㎖ of PDGF-BB(PDGF group) for 1, 2, 3 day respectively. And then each group was characterized by examining of the cell counting, MTT assay, collagen synthesis.

The results were as follows.

1. In the analysis of cell proliferation by cell counting, both calcium sulfate group and calcium sulfate plus PDGF group showed no stastically significant difference compared to control group, but there was stastically significant difference between PDGF group and calcium sulfate group at 1, 2 day(P<0.05).

2. In the analysis of cell proliferation by MTT assay in calcium sulfate extracts, both calcium sulfate group and calcium sulfate plus PDGF group showed no stastically significant difference compared to control group, but there was stastically significant difference between PDGF group and calcium sulfate group at 2, 3 day, and between calcium sulfate plus PDGF group and calcium sulfate group at 2 day(P<0.05).

3. In the analysis of cell proliferation by MTT assay in transwell, both control group and PDGF group showed stastically significant difference compared to both calcium sulfate group and calcium sulfate plus PDGF group at 1 day, but there was no stastically significant difference compared to both calcium sulfate group and calcium sulfate plus PDGF group at 2, 3 day(P<0.05).

4. In the analysis of collagen synthesis by immunoblotting assay in calcium sulfate extracts, high level was detected on calcium sulfate group at 3 day, on calcium sulfate plus PDGF group at 1 day, on PDGF group at 2 day.

On the basis of these results, calcium sulfate was biocompatible on the periodontal ligament cells and might have potential possibility as a vehicle of PDGF in the periodontal tissue regeneration.