니코틴이MG63 조골세포주의 오스테오칼신과오스테오프로 테제린의 생성 및 mRNA 발현에 미치는 영향 Effects of nicotine on the formation of osteocalcin and osteoprotegerin and synthesis of its mRNA in MG63 osteoblast-like cell원문보기
흡연이 골형성 세포의 기능을 억제한다는 것은 이미 잘 알려져 있으나 대다수의 연구가 주로 역학조사에 기초하여 이루어져 왔다. 본 연구의 목적은 흠연과 골형성 세포의 상관관계를 실험적으로 평가하기 위한 것이다. 본 연구를 위해 인간 골육종 세포인 MG63 조골제포주를 이용하였으며, 니코틴이 조골세포의 증식, 염기성 인산분해효소의 활성, 오스테오칼신곽 오스테오프로테제린의 생성 그리고 mRNA발현에 미치는 영향을 관찰하였다 MG63조골세포를 니코틴 농도가 각각 $1.0{\mu}M$,\;1.0mM, 2.5mM, 5.0mM, 7.5mM, 그리고 10.0mM,이 함유된 배지에서 1일, 2일, 3일 그리고 6일 동안 배양 실험을 하여 다음과 같은 결과를 얻었다. MG63 조골세포의 증식이 저농도의 니코틴에서는 일시적으로 활성화되었지만 5.0mM 이상의 고농도에서는 억제되었다. 염기성 인산분해효소의 활성은 니코틴 농도가 증가함에 따라 감소하였다. 오스테오칼신 생성양은, 배양 1일 후, 5.0mM 이하의 농도에서 증가하였으나 7.5mM 이상의 고농도에서는 감소하였다. MG63 조골세포를 3일 배양한 경우, 오스테오칼신 생성양은 1.0mM의 저농도에서도 감소하였다 니코틴이 오스테오칼신 단백질 생성과 오스테오칼신 mRNA생성에 미치는 영향은 1일과 3일에서 다소 차이가 있었다. 오스테오프로테제린의 생성양은 모든 실험군에서 니코틴을 함유하지 않은 대조군보다 감소하였다. 하지만 mRNA 수치는 단백질 생성과 상반되게 7.5mM (3일), 5.0mM (6일) 이상의 고농도에서 증가하였다.
흡연이 골형성 세포의 기능을 억제한다는 것은 이미 잘 알려져 있으나 대다수의 연구가 주로 역학조사에 기초하여 이루어져 왔다. 본 연구의 목적은 흠연과 골형성 세포의 상관관계를 실험적으로 평가하기 위한 것이다. 본 연구를 위해 인간 골육종 세포인 MG63 조골제포주를 이용하였으며, 니코틴이 조골세포의 증식, 염기성 인산분해효소의 활성, 오스테오칼신곽 오스테오프로테제린의 생성 그리고 mRNA발현에 미치는 영향을 관찰하였다 MG63조골세포를 니코틴 농도가 각각 $1.0{\mu}M$,\;1.0mM, 2.5mM, 5.0mM, 7.5mM, 그리고 10.0mM,이 함유된 배지에서 1일, 2일, 3일 그리고 6일 동안 배양 실험을 하여 다음과 같은 결과를 얻었다. MG63 조골세포의 증식이 저농도의 니코틴에서는 일시적으로 활성화되었지만 5.0mM 이상의 고농도에서는 억제되었다. 염기성 인산분해효소의 활성은 니코틴 농도가 증가함에 따라 감소하였다. 오스테오칼신 생성양은, 배양 1일 후, 5.0mM 이하의 농도에서 증가하였으나 7.5mM 이상의 고농도에서는 감소하였다. MG63 조골세포를 3일 배양한 경우, 오스테오칼신 생성양은 1.0mM의 저농도에서도 감소하였다 니코틴이 오스테오칼신 단백질 생성과 오스테오칼신 mRNA생성에 미치는 영향은 1일과 3일에서 다소 차이가 있었다. 오스테오프로테제린의 생성양은 모든 실험군에서 니코틴을 함유하지 않은 대조군보다 감소하였다. 하지만 mRNA 수치는 단백질 생성과 상반되게 7.5mM (3일), 5.0mM (6일) 이상의 고농도에서 증가하였다.
The purpose of this study was to evaluate the correlation between nicotine and the activity of bone forming cell. MG63 osteoblast-like cells were used for this study. Several factors were examined including the proliferation of cell, alkaline phosphatase activity, the formation of osteocalcin and os...
The purpose of this study was to evaluate the correlation between nicotine and the activity of bone forming cell. MG63 osteoblast-like cells were used for this study. Several factors were examined including the proliferation of cell, alkaline phosphatase activity, the formation of osteocalcin and osteoprotegerin. and the synthesis of its mRNA. MG63 osteoblast-like cells were incubated for 1, 2, 3 and 6 days with nicotine added to the culture medium in 1.0 ${\mu}M$, 1.0mM, 2.5mM, 5.0mM, 7.5mM, and 10.0mM concentrations. The proliferation of MG63 osteoblast-like cells was temporarily activated at the low nicotine concentrations. At high concentrations (>5.0 mM), however. it was suppressed. Alkaline phosphatase activity was suppressed in a dose-dependent manner as the concentration of nicotine increased. Osteocalcin decreased in a dose-dependent manner at high nicotine concentrations of more than 7.5mM and the same result was show when the osteoblasts were treated with low concentrations for longer than 3 days. There was a difference in the influence of nicotine on the synthesis of osteocalcin mRNA and formation of osteocalcin itself at 1 and 3 days. Generally, osteoprotegrin notably declined in all experimental groups. However, the level of its mRNA increased at high nicotine concentrations of more than 7.5mM after 3 days and more than 5.0mM after 6days.
The purpose of this study was to evaluate the correlation between nicotine and the activity of bone forming cell. MG63 osteoblast-like cells were used for this study. Several factors were examined including the proliferation of cell, alkaline phosphatase activity, the formation of osteocalcin and osteoprotegerin. and the synthesis of its mRNA. MG63 osteoblast-like cells were incubated for 1, 2, 3 and 6 days with nicotine added to the culture medium in 1.0 ${\mu}M$, 1.0mM, 2.5mM, 5.0mM, 7.5mM, and 10.0mM concentrations. The proliferation of MG63 osteoblast-like cells was temporarily activated at the low nicotine concentrations. At high concentrations (>5.0 mM), however. it was suppressed. Alkaline phosphatase activity was suppressed in a dose-dependent manner as the concentration of nicotine increased. Osteocalcin decreased in a dose-dependent manner at high nicotine concentrations of more than 7.5mM and the same result was show when the osteoblasts were treated with low concentrations for longer than 3 days. There was a difference in the influence of nicotine on the synthesis of osteocalcin mRNA and formation of osteocalcin itself at 1 and 3 days. Generally, osteoprotegrin notably declined in all experimental groups. However, the level of its mRNA increased at high nicotine concentrations of more than 7.5mM after 3 days and more than 5.0mM after 6days.
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제안 방법
Because ALPase induces the apposition of bone mineral, a decrease in ALPase reduces bone mineralization. In this study, nicotine dose-dependently inhibited the formation of ALPase. Especially when nicotine was administered for more than 2 days at >5.
The control groups which were not treated with nicotine and experimental groups which were treated with each concentrations of nicotine (1.0 zzM, 1.0 mM, 2.5 mM, 5 mM, 7.5 mM, 10 mM) were incubated for 1, 3, and 6 days. The total of 21 samples were prepared.
2). The samples, which each containing distinct concentrations of nicotine, were incubated for 1, 3, and 6 days and stored for up to 24 h이jis at 4℃. A 100 月 substrate solution was added to each well for 15 minutes and, finally a 100stop solution (1N H2SO4) was gently mixed into the well by hand.
대상 데이터
MG63 osteoblast-like cells were obtained from the Korea Cell Line Bank. They were incubated at 37℃ in 5% CO2 in D니becco's Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 100 units/ml penicillin-streptomycin.
5 mM, 10 mM) were incubated for 1, 3, and 6 days. The total of 21 samples were prepared.
데이터처리
were obtained. The changes of cell proliferation, alkaline phosphatase and the formation of osteocalcin and osteoprotegrin between control and experimental group were evaluated by paired t-test.
이론/모형
The effect of nicotine on the cell proliferation was determined by microtiter assay which uses the tetrazolium salt (MTT). A 100 of 3-(4, 5—dimethyl thiaz이-2-yl) — 2, 5-diphenyltetraslum bromide were added to the cells after 1, 2, 3, and 6 days.
성능/효과
2. ALPase was suppressed in a dose-dependent manner as the concentration of nicotine increased. Particularly, the activity of ALPase notably declined when it was exposed to highly concentrated nicotine (>5.
3. Osteocalcin increased one day after treatment with low concentrations of nicotine «5.0 mM), compared with the control group. At the high nicotine concentrations of more than 7.
5. The formation of osteoprotegerin notably declined in all experimental groups, compared with the control group, except that it increased somewat more than the control group at concentrations of 1.0 mM and 2.5 mM after 6 days.
6. The level of osteoprotegerin mRNA increased at high nicotine concentrations of more than 7.5 mM after 3 days and more than 5.0 mM after 6 days. However, there were no significant changes in the other experimental groups.
After 1 day, the concentrations of nicotine of less than 5.0 mM showed no change in ALPase activity and those of more than 7.5 mM showed a decrease. However, after 3 and 6 days, every experimental g「oup showed notable decreases in ALPase activity compared with the control group.
0 mM, the amount of osteocalcin decreased in a dose dependent manner. After 6 days, all the experimental groups showed a decrease in osteocalcin formation, and especially at high nicotine concentrations of more than 5.0 mM, there was a remarkable decreasing tendency (Table 4).
However, after 3 days, there were no remarkable differences between the control gro나p and the experimental groups. After 6 days, the mRNA lev이 of osteocalcin decreased in a dosedependent manner; particularly, when the concentration of nicotine was higher than 7.5 mM, the mRNA of osteocalcin was undetected (Table 6).
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