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1.
Sophocarpine attenuates toll‐like receptor 4 in steatotic hepatocytes to suppress pro‐inflammatory cytokines synthesis 下载免费PDF全文
2.
A rapid and sensitive method for measuring monooxygenase activities in hepatocytes cultured in 96-well plates 总被引:3,自引:0,他引:3
Summary Measurement of biotransformation activities in cells is of great importance for drug metabolism and toxicologic studies. It is currently done by measuring the enzymatic activities in partially purified microsomes. In the present work we report on a rapid, easy, sensitive, and reproducible fluorimetric assay for quantifying cytochrome P450-dependent monooxygenase activities (P450IA1, P450IIB1) in hepatocytes cultured in 96-well plates. The procedure involves the direct determination of enzymatic activities in intact hepatocytes while avoiding cell homogenization, thereby permitting use of a the reduced number of cells and allowing cultured cells to be used in later experiments. Substrates (7-ethoxyresorufin, 7-pentoxyresorufin) are added to culture medium and metabolized by hepatocytes. After enzymatic deconjugation, the fluorescent resorufin present in culture medium is quantified by means of a microplate fluorimetric reader. Major advantages of this technique, as compared to other available methods, are: a) no cell disruption is required; b) activity can be measured with a very small number of cells; c) rapid processing time; and d) possibility of performing repeated assays with the same cell monolayer. 相似文献
3.
4.
BACKGROUND/AIMS: Hepatocytes (HEP) have been the major target for structural quantification in the liver, but an estimation of their total number (N), their percentage in relation to the global number of liver cells and the evaluation of the percentage of binucleated hepatocytes (BnHEPs) have never been performed with modern design-based stereological techniques. The establishment of sound technical guidelines and baseline quantitative data in non-pathological conditions are relevant to properly evaluate HEP hyperplasia and BnHEP responses. METHODS: In this study, we combined immunocytochemistry with sound design-based stereology for estimating the N of HEP and the N of non-hepatocytic cells (NHCs). For obtaining systematic uniform random sections (30 microm thick), a smooth fractionator sampling scheme was applied to the liver of five male Wistar rats (3 month old). Those sections were immunostained with polyclonal antibodies against carcinoembryonic antigen. Because biliary canaliculi were then marked, an unequivocal counting of mononucleated hepatocytes (MnHEP) and BnHEP was allowed. RESULTS: The N of HEP was estimated to be 1.93 x 10(9), with a coefficient of error (CE) of 0.02, corresponding to 129 x 10(6) HEP/g of liver. BnHEP represented 26% of total HEP number. The N of NHC was estimated as 1.31 x 10(9) (CE=0.02). CONCLUSION: The strategy here presented provides a reliable method for accessing the N of HEP (distinguishing MnHEP from BnHEP) in situations in which these parameters are relevant, namely for evaluating the magnitude of an hyperplastic liver response from its very early onset. 相似文献
5.
用高速离心、分段盐析、亲和层析、超滤和阴离子交换层析法,从牛胎盘中分离纯化出一种肝细胞生长因子。结果表明,此因子的最终得率为0.005mg/g牛胎盘,其分子量约为99000u,它能强烈刺激原代培养大鼠肝细胞的DNA合成。提示利用自制的亲和层析凝胶,通过此提纯流程可以得到一种肝细胞生长因子。 相似文献
6.
Takahito Adachi Makoto Takeuchi Satomi Uno Motohisa Kato Kiichi Miya Shigetoyo Saji 《Surgery today》1997,27(12):1144-1149
Although tumor cryosurgery would be expected to produce beneficial immunological effects from the enhancement of anti-tumor
activity, under certain conditions the tumor may become enlarged and metastases promoted due to increased immunosuppressive
activity and a high zone tolerance. In the present study, we examined whether hepatocyteproliferating factors were produced
by the inoculation of freezing-thawing hepatic tissue (FTHT). Serum obtained from rats inoculated with FTHT increased DNA
synthesis, according to measurement by [3H]thymidine incorporation in primary cultured rat hepatocytes. This increase was dependent on the serum concentration, with
serum obtained on day 14 after the inoculation being the most potent for hepatocyte proliferation. The sensitized serum promoted
DNA synthesis nearly as much as serum obtained from a 70% hepatectomized rat, but slightly less than 10ng/ml hepatocyte growth
factor. The sensitized serum also protected hepatocytes from carbon tetrachloride (CCI4)-induced hepatotoxicity. Optical density measured by the 3-(4,5-dimethyl-thiazole-2-yl)-2,5-diphenyl tetrozolium bromide
(MTT) cytotoxicity assay was increased, and the release of aspartate aminotransferase, alanine aminotransferase, and lactate
dehydrogenase in medium was decreased by treating hepatocytes damaged by CCI4 with the sensitized serum. These results suggest that certain hepatocyte-proliferating and protective factors are induced
in serum by the inoculation of freezing-thawing hepatic tissue, and that the sensitized serum may be useful in the treatment
of liver failure. 相似文献
7.
M. M. Rasulov I. G. Kuznetsov R. G. Shakirova A. G. Zabozlaev A. A. Belousov M. G. Voronkov 《Bulletin of experimental biology and medicine》1993,115(1):73-75
Translated fromByulleten' Experimental'noi Biologii i Meditsiny, Vol. 115, No. 1, pp. 69–71, January, 1993 相似文献
8.
Growth factor-dependent activation of the MAPK pathway in human pancreatic cancer: MEK/ERK and p38 MAP kinase interaction in uPA synthesis 总被引:3,自引:0,他引:3
Increased expression of the hepatocyte growth factor (HGF) receptor (c-met) and urokinase type plasminogen (uPA) correlated
with the development and metastasis of cancers. To investigate the role of HGF/c-met signaling on metastasis in cancer cells
stimulated with HGF, we examined the effects of a specific MEK1 inhibitor (PD98059) and a p38 MAP kinase inhibitor (SB203580)
on HGF-induced uPA expression in pancreatic cancer cell lines, L3.6PL and IMIM-PC2. Pretreatment of PD98059 decreased HGF-mediated
phosphorylation of extracellular receptor kinase (ERK), uPA secretion and expression of matrix metalloproteinases (MMP-2 and
MMP-9) in a dose-dependent manner. In contrast, SB203580 pretreatment increased HGF-stimulated ERK phosphorylation, uPA secretion
and expression of MMPs. SB203580 also reversed the inhibition of HGF-mediated ERK activation and uPA secretion in the PD98059-pretreated
cells. These results suggest that ERK activation by HGF might play important roles in the metastasis of pancreatic cancer
and the p38 MAPK pathway also involved in the HGF-mediated uPA secretion and metastasis by regulation of ERK pathway.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
9.
Social isolation stress augments angiogenesis induced by colon 26-L5 carcinoma cells in mice 总被引:3,自引:0,他引:3
Wu W Murata J Murakami K Yamaura T Hayashi K Saiki I 《Clinical & experimental metastasis》2000,18(1):1-10
We have previously shown that tumor necrosis factor-α (TNF-α), which is an important angiogenesis-related factor, was over-secreted in male BALB/c mice under social isolation stress as compared with the control, and closely associated with a remarkable elevation of tumor invasion and metastasis of colon 26-L5 carcinoma cells. In the present study, we explored the effect of isolation stress on the angiogenesis caused by colon 26-L5 carcinoma cells in vivo and in vitro. Social isolation lead to the enhancement of tumor growth after intrahepatic implantation with a fragment of colon 26-L5 tumor. Angiogenic response (number of vessels oriented towards tumor mass) and tumor growth (size) were significantly increased in the socially isolated mouse relative to that in the group-housed mice. Furthermore, higher protein level of hepatic TNF-α was found in the stressed mice than that in the control. Expression of mRNA for vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) were also elevated in the tumor regions and liver tissues of the stressed mice in comparison with that in group-housed mice. On the other hand, hepatic sinusoidal endothelial (HSE) cells treated with TNF-α exhibited a marked promotion of the migration, invasion, expression of mRNA for matrix metalloproteinase (MMP)-9, and tube-like formation, but no cytotoxicity against the cells in vitro. The above data suggest that the social isolation stress augmented the tumor-induced angiogenesis probably by up-regulating the angiogenesis-related factors, including TNF-α, VEGF and HGF, and consequently mediating the functions of endothelial cells such as migration, invasion, and tube-like formation. 相似文献
10.
促肝细胞生长素诱导人肝癌细胞(BEL-7402)凋亡 总被引:1,自引:0,他引:1
本文从细胞学、DNA凝胶电泳、流式细胞术三方面研究促肝细胞生长素(pHGF)在体外对肝癌细胞增殖活性的影响。结果表明:pHGF对肝癌BEL-7402细胞增殖有抑制作用,并存在剂量和时间相关性。其48h的半数抑制浓度(ID50)为0.37mg/ml±0.04mg/ml。而37℃灭活的pHGF对BEL-7402细胞增殖在15h无抑制作用,在24和48h抑制作用很弱(ID50>1.5mg/ml)。DNA凝胶电泳结果表明,pHGF可诱导BEL7402细胞产生细胞凋亡(Apoptosis)。流式细胞术(FCM)结果显示:pHGF抑制BEL-7402细胞增殖过程是先使细胞停留在G0/G1期,继而诱导细胞产生凋亡。后两项结果均显示pHGF对人肝癌细胞凋亡的诱导里时间和剂量相关性。 相似文献