Tiggelman A M, Boers W, Linthorst C, Brand H S, Sala M, Chamuleau R A
Academic Medical Center, University of Amsterdam, J. van Gool Laboratory for Experimental Internal Medicine, The Netherlands.
J Hepatol. 1995 Sep;23(3):295-306.
BACKGROUND/AIMS: Interleukin-6 is a major trigger for the synthesis of acute phase proteins by liver parenchymal cells. Acute phase proteins may contribute to the regulation of liver fibrosis by inhibition of proteases (e.g. collagenase) and by binding of cytokines. Since liver (myo)fibroblasts play an important role in the production of extracellular matrix in fibrotic livers, a study was undertaken into whether these cells are able to synthesize interleukin-6, which would give them the opportunity to contribute to regulation of synthesis of acute phase proteins by neighbouring parenchymal cells.
In the present study we investigated interleukin-6 production by two cell types obtained from human liver tissue: human fat-storing cells obtained from 5-15% Percoll fractions, which transformed in culture into myofibroblasts co-expressing alpha-smooth muscle actin and vimentin (VA cells) and fibroblasts obtained from 30-40% Percoll fractions which express vimentin only (V vells). Interleukin-6 production was measured in culture media of these cells using an enzyme-linked immunosorbent assay after incubation with lipopolysaccharide, and mediators like interleukin-1 beta, tumor necrosis factor-alpha, transforming growth factor-beta and interferon-gamma, known to be present in elevated concentrations in fibrotic livers.
Unstimulated human liver (myo)fibroblasts produced considerable amounts of interleukin-6 (287 ng/mg cellular protein (VA cells), and 54 ng/mg cellular protein (V cells), within 48 h). Biological activity of these high concentrations of interleukin-6 measured in the enzyme-linked immuno-sorbent assay was confirmed in the B9-bioassay for interleukin-6 and by stimulation of alpha 2-macroglobulin production in rat liver parenchymal cell cultures. Lipopolysaccharide, interleukin-1 beta and tumor necrosis factor-alpha were potent stimulators of basal interleukin-6 production by VA and V cells, 1 microgram/ml lipopolysaccharide enhanced basal interleukin-6 production 3-fold within 48 h. 100 U/ml interleukin-1 beta and 1000 U/ml tumor necrosis factor-alpha each stimulated basal interleukin-6 production by VA cells 2-5 fold, whereas V cells were stimulated 10-25 fold. These effects were specific since the stimulation by lipopolysaccharide was completely inhibited by polymyxin B and the enhancing effects of interleukin-1 beta and tumor necrosis factor-alpha were neutralized by specific antibodies. Transforming growth factor-beta and interferon gamma did not influence interleukin-6 synthesis by either cell type in culture.
These results indicate that transformed fat-storing cells (VA cells) and fibroblasts (V cells) may function as a local source of interleukin-6 in the human liver. Since interleukin-6 plays a key role in the regulation of the production of acute phase proteins by liver parenchymal cells, we hypothesize that human liver (myo)fibroblasts may stimulate local production of acute phase proteins in the fibrotic liver, thus contributing to local regulation of inflammatory and fibrogenic reactions.
背景/目的：白细胞介素-6是肝实质细胞合成急性期蛋白的主要触发因素。急性期蛋白可能通过抑制蛋白酶（如胶原酶）和结合细胞因子来促进肝纤维化的调节。由于肝（肌）成纤维细胞在纤维化肝脏的细胞外基质产生中起重要作用，因此开展了一项研究，以探讨这些细胞是否能够合成白细胞介素-6，这将使它们有机会促进相邻实质细胞对急性期蛋白合成的调节。
在本研究中，我们调查了从人肝组织获得的两种细胞类型产生白细胞介素-6的情况：从5%-15% Percoll分层中获得的人贮脂细胞，其在培养中转化为共表达α-平滑肌肌动蛋白和波形蛋白的肌成纤维细胞（VA细胞），以及从30%-40% Percoll分层中获得的仅表达波形蛋白的成纤维细胞（V细胞）。在用脂多糖以及已知在纤维化肝脏中浓度升高的介质如白细胞介素-1β、肿瘤坏死因子-α、转化生长因子-β和干扰素-γ孵育后，使用酶联免疫吸附测定法测量这些细胞培养基中的白细胞介素-6产生量。
未受刺激的人肝（肌）成纤维细胞产生大量白细胞介素-6（48小时内，287 ng/mg细胞蛋白（VA细胞）和54 ng/mg细胞蛋白（V细胞））。在白细胞介素-6的B9生物测定中以及通过刺激大鼠肝实质细胞培养物中α2-巨球蛋白的产生，证实了酶联免疫吸附测定中测量的这些高浓度白细胞介素-6的生物学活性。脂多糖、白细胞介素-1β和肿瘤坏死因子-α是VA和V细胞基础白细胞介素-6产生的有效刺激物，1 μg/ml脂多糖在48小时内将基础白细胞介素-6产生量提高了3倍。100 U/ml白细胞介素-1β和1000 U/ml肿瘤坏死因子-α分别将VA细胞的基础白细胞介素-6产生量刺激了2-5倍，而V细胞受到了10-25倍的刺激。这些作用是特异性的，因为脂多糖的刺激被多粘菌素B完全抑制，白细胞介素-1β和肿瘤坏死因子-α的增强作用被特异性抗体中和。转化生长因子-β和干扰素-γ在培养中均不影响两种细胞类型的白细胞介素-6合成。
这些结果表明，转化的贮脂细胞（VA细胞）和成纤维细胞（V细胞）可能是人肝脏中白细胞介素-6的局部来源。由于白细胞介素-6在肝实质细胞急性期蛋白产生的调节中起关键作用，我们推测人肝（肌）成纤维细胞可能刺激纤维化肝脏中急性期蛋白的局部产生，从而有助于局部调节炎症和纤维化反应。
