Siavoshian S, Segain J P, Kornprobst M, Bonnet C, Cherbut C, Galmiche J P, Blottière H M
Centre de Recherche en Nutrition Humaine de Nantes, INSERM U539, CHU Hôtel-Dieu, Nantes, France.
Gut. 2000 Apr;46(4):507-14. doi: 10.1136/gut.46.4.507.
Sodium butyrate, a product of colonic bacterial fermentation, is able to inhibit cell proliferation and to stimulate cell differentiation of colonic epithelial cell lines. It has been proposed that these cellular effects could be linked to its ability to cause hyperacetylation of histone through the inhibition of histone deacetylase.
To analyse the molecular mechanisms of butyrate action on cell proliferation/differentiation and to compare them with those of trichostatin A, a well known inhibitor of histone deacetylase.
HT-29 cells were grown in the absence or presence of butyrate or trichostatin A. Cell proliferation and cell cycle distribution were studied after DNA staining by crystal violet and propidium iodide respectively. Cell cycle regulatory proteins were studied by western blot and reverse transcription-polymerase chain reaction. Cell differentiation was followed by measuring brush border enzyme activities. Histone acetylation was studied by acid/urea/Triton acrylamide gel electrophoresis.
Butyrate blocked cells mainly in the G(1) phase of the cell cycle, whereas trichostatin A was inhibitory in both G(1) and G(2) phases. Butyrate inhibited the mRNA expression of cyclin D1 without affecting its protein expression and stimulated the protein expression of cyclin D3 without affecting its mRNA expression. Trichostatin A showed similar effects on cyclin D1 and D3. Butyrate and trichostatin A stimulated p21 expression both at the mRNA and protein levels, whereas their effects on the expression of cyclin dependent kinases were slightly different. Moreover, butyrate strongly stimulated the activity of alkaline phosphatase and dipeptidyl peptidase IV, whereas trichostatin A had no effect. Finally, a six hour exposure to butyrate or trichostatin A induced histone H4 hyperacetylation. At 15 and 24 hours, histone H4 remained hyperacetylated in the presence of butyrate, whereas it returned to control levels in the presence of trichostatin A.
The data may explain how butyrate acts on cell proliferation/differentiation, and they show that trichostatin A does not reproduce every effect of butyrate, mainly because of its shorter half life.
丁酸钠是结肠细菌发酵的产物，能够抑制结肠上皮细胞系的细胞增殖并刺激其细胞分化。有人提出，这些细胞效应可能与其通过抑制组蛋白脱乙酰酶导致组蛋白高度乙酰化的能力有关。
分析丁酸盐对细胞增殖/分化作用的分子机制，并将其与著名的组蛋白脱乙酰酶抑制剂曲古抑菌素A的作用机制进行比较。
HT-29细胞在无或有丁酸盐或曲古抑菌素A的情况下培养。分别用结晶紫和碘化丙啶进行DNA染色后，研究细胞增殖和细胞周期分布。通过蛋白质印迹法和逆转录-聚合酶链反应研究细胞周期调节蛋白。通过测量刷状缘酶活性来跟踪细胞分化。通过酸/尿素/ Triton丙烯酰胺凝胶电泳研究组蛋白乙酰化。
丁酸盐主要将细胞阻滞在细胞周期的G(1)期，而曲古抑菌素A在G(1)期和G(2)期均有抑制作用。丁酸盐抑制细胞周期蛋白D1的mRNA表达而不影响其蛋白质表达，并刺激细胞周期蛋白D3的蛋白质表达而不影响其mRNA表达。曲古抑菌素A对细胞周期蛋白D1和D3表现出类似的作用。丁酸盐和曲古抑菌素A在mRNA和蛋白质水平上均刺激p21表达，而它们对细胞周期蛋白依赖性激酶表达的影响略有不同。此外，丁酸盐强烈刺激碱性磷酸酶和二肽基肽酶IV的活性，而曲古抑菌素A则无作用。最后，暴露于丁酸盐或曲古抑菌素A六小时可诱导组蛋白H4高度乙酰化。在15和24小时时，在丁酸盐存在下组蛋白H4仍保持高度乙酰化，而在曲古抑菌素A存在下它恢复到对照水平。
这些数据可以解释丁酸盐如何作用于细胞增殖/分化，并表明曲古抑菌素A不能重现丁酸盐的所有作用，主要是因为其半衰期较短。
