HIBADH Plays an Important Role in the Course of Liver Cell Necrosis

Objective To observe the biological function of human 3-hydroxyisobutyrate dehydrogenase(HIBADH).Methods Human 3-hydroxyisobutyrate dehydrogenase(HIBADH, 3-hydroxy-2-methyl propanoate: NAD+ oxidoreductase) recombinant protein was expressed in E. coli BL21, and purified by Ni+ column. The special antisera was obtained from rabbits immunized by this purified antigen. On the distribution of HIBADH, it was found that HIBADH over-expressed in the injured liver cells when serious hepatitis occurred. The phenomenon was confirmed in the animal models of SD rats with acute liver cell injury induced by CCl4, but this phenomenon did not exist in the models induced by endotoxin combined with galactosamine. Further more, HIBADH's overexpression in liver cells will induce cell necrosis through the pathway of oxidative stress.Results When the liver cells injured by drug or other chemical materials, HIBADH will be compensationally over-expressed for the deficiency of energy, so liver cells can make enough ATP through brand-chain amino acid catabolism. However, the overexpression of HIBADH will be harmful for liver cells through the product of much more active oxygens which will induce the cell necrosis. Conclusions HIBADH over-expression is a signal of the liver cell metabolism injury, and it can aggravate the liver cell injury through oxidative stress.

Comparative study on radiosensitivity of various tumor cells and human normal liver cells

AIM: To investigate the radiation response of various human tumor cells and normal liver cells.METHODS: Cell lines of human hepatoma cells (SMMC-7721),liver cells (L02), melanoma cells (A375) and cervical tumor (HeLa) were irradiated with 60Co γ-rays. Cell survive was documented by a colony assay. Chromatid breaks were measured by counting the number of chromatid breaks and isochromatid breaks immediately after prematurely chromosome condensed by Calyculin-A.RESULTS: Linear quadratic survival curve was observed in all of four cell lines, and dose-dependent increase in radiation-induced chromatid and isochromatid breaks were observed in GB2B phase. Among these four cell lines,A375 was most sensitive to radiation, while, L02 had the lowest radiosensitivity. For normal liver cells, chromatid breaks were easy to be repaired, isochromatid breaks were difficult to be repaired.CONCLUSION: The results suggest that the γ-rays induced chromatid breaks can be possibly used as a good predictor of radiosensitivity, also, unrejoined isochromatid breaks probably tightly related with cell cancerization.

Hepatocytic differentiation of mesenchymal stem cells in cocultures with fetal liver cells

瞄准：为了与胎儿的肝细胞(FLC ) 和可能性在合作文化调查间充质的干细胞(MSC ) 的 hepatocytic 区别膨胀，区分了 hepatocytic 房间。方法：MSC 被制动火箭与绿荧光灯的蛋白质(GFP ) 标记病毒的基因转导变异。同种细胞的显著 MSC 在用与干细胞补充的fibronectin涂的培养皿和媒介刺激条件的肝下面是也有教养的因素( SCF )， hepatocyte 生长因素( HGF )，表皮的生长因素( EGF )，和成纤维细胞生长因素 4 ( FGF-4 )独自一个，或在刚孤立的 FLC 的存在。在合作文化的房间被收获，并且 GFP+ 或 GFP- 房间用荧光被分开激活的房间排序。为肝 specific 标记 cytokeratin-18 (CK-18 ) 的反向的抄写聚合酶链反应(RT-PCR )( 法新社) ，白朊，和 alpha-fetoprotein 在不同房间人口被执行。结果：在指定文化条件下面，与 FLC co 有教养的老鼠 MSC 超过二个星期表示了白朊， CK-18，和 AFP-RNA。在 wk 3， MSC 失去了 hepatocytic 基因表示，可能由于 cocultured FLC 的增生。FLC 也在合作文化和一个很高的生长潜力显示出稳定的肝 specific 基因表情。结论：从骨髓的老鼠 MSC 能面对 FLC 在试管内区分 hepatocytic 房间，在合作文化的 MSC 的存在也为 FLC 的扩大和区别提供有益的环境。

Liver cell adenoma:A case report with clonal analysis and literature review

我们向它的 clonality 地位，病原的因素和鉴别诊断与特殊尊重在一个 33 岁的女病人汇报肝房间腺瘤(LCA ) 的一个案例。案例被组织病理学说，免疫组织化学和 clonality 试金在雄激素受体焦点在女体的纸巾和多型性在激活马赛克主义基于 X-chromosomal 检验。从中国和另外的国家的报导案例的 clinicopathological 特征被比较。损害是球形的，缩放在它的最大的尺寸的 2 厘米。组织学地，它由在绳索，其大多数是 two-cell-thick 并且由窦状隙分开了安排的房间组成。焦点的丰满的变化和过多的糖原贮积被观察。肿瘤房间在形状圆或多角形，类似于包围实质的房间。有丝分裂没被发现。没有门道，中央静脉或小导管在损害以内被发现。肿瘤组织为 cytokeratin (CK ) 显示出阳性反应 18，然而并非为 CK19， vimentin，雌激素和孕酮受体。Monoclonality 为损害被表明，证实 LCA 的诊断。Clonality 分析对它从焦点的榴状的增生的区别有用。

YKL40 expression in CD14^+ liver cells in acute and chronic injury

AIM:To demonstrate that CD14 + cells are an important source of the growth factor YKL40 in acute and chronic liver damage.METHODS:Rats were inoculated with one dose of CCl4 to induce acute damage.Liver biopsies were obtained at 0,6,12,24,48 and 72 h.For chronic damage,CCl4 was administered three days per week for 6 or 8 wk.Tissue samples were collected,and cellular populations were isolated by liver digestion and purified by cell sorting.YKL40 mRNA and protein expression were evaluated by realtime polymerase chain reaction and western blot.RESULTS:Acute liver damage induced a rapid increase of YKL40 mRNA beginning at 12 h.Expression peaked at 24 h,with a 26fold increase over basal levels.By 72 h however,YKL40 expression levels had nearly returned to control levels.On the other hand,chronic damage induced a sustained increase in YKL40 expression,with 7and 9fold higher levels at 6 and 8 wk,respectively.The pattern of YKL40 expression in different subpopulations showed that CD14+cells,which include Kupffer cells,are a source of YKL40 after acute damage at 72 h[0.09 relative expression units(REU)]as well as after chronic injury at 6 wk(0.11 REU).Hepatocytes,in turn,accounted for 0.06 and 0.01 REU after 72 h(acute)or 6 wk(chronic),respectively.The rest of the CD14cells(including T lymphocytes,B lymphocytes,natural killer and natural killer T cells) yielded 0.07 and 0.15 REU at 72 h and 6 wk,respectively.YKL40 protein expression in liver was detected at 72 h as well as 6 and 8 wk,with the highest expression relative to controls(11fold;P≤0.05)seen at 6 wk.Macrophages were stimulated by lipopolysaccharide.We demonstrate that under these conditions,these cells showed maximum expression of YKL40 at 12 h,with P<0.05 compared with controls.CONCLUSION:Hepatic CD14 + cells are an YKL40 mRNA and protein source in acute and chronic liver injury,with expression patterns similar to growth factors implicated in inflammationfibrogenesis.

Liver-specific gene expression in mesenchymal stem cells is induced by liver cells

AIM: The origin of putative liver cells from distinct bone marrow stem cells, e.g. hematopoietic stem cells or multipotent adult progenitor cells was found in recent in vitro studies. Cell culture experiments revealed a key role of growth factors for the induction of liver-specific genes in stem cell cultures. We investigated the potential of rat mesenchymal stem cells (MSC) from bone marrow to differentiate into hepatocytic cells in vitro. Furthermore,we assessed the influence of cocultured liver cells on induction of liver-specific gene expression.METHODS: Mesenchymal stem cells were marked with green fluorescent protein (GFP) by retroviral gene transduction. Clonal marked MSC were either cultured under liver stimulating conditions using fibronectin-coated culture dishes and medium supplemented with SCF, HGF,EGF, and FGF-4 alone, or in presence of freshly isolated rat liver cells. Cells in cocultures were harvested and GFP+ or GFP- cells were separated using fluorescence activated cell sorting. RT-PCR analysis for the stem cell marker Thy1 and the hepatocytic markers CK-18, albumin, CK-19,and AFP was performed in the different cell populations.RESULTS: Under the specified culture conditions, rat MSC cocultured with liver cells expressed albumin-, CK-18,CK-19, and AFP-RNA over 3 weeks, whereas MSC cultured alone did not show liver specific gene expression.CONCLUSION: The results indicate that (1) rat MSC from bone marrow can differentiate towards hepatocytic lineage in vitro, and (2) that the microenvironment plays a decisive role for the induction of hepatic differentiation of rMSC.

Hepatitis C virus core proteins derived from different quasispecies of genotype 1b inhibit the growth of Chang liver cells

AIM: To investigate the influence of different quasispecies of hepatitis C virus (HCV) genotype 1b core protein on growth of Chang liver cells. METHODS: Three eukaryotic expression plasmids (pEGFP-N1/core) that contained different quasispecies truncated core proteins of HCV genotype 1b were constructed. These were derived from tumor (T) and non- tumor (NT) tissues of a patient infected with HCV and C191 (HCV-J6). The core protein expression plasmids were transiently transfected into Chang liver cells. At different times, the cell cycle and apoptosis was assayed by flow cytometry, and cell proliferation was assayed by methyl thiazolyl tetrazolium (MTT) assay. RESULTS: The proportion of S-phase Chang liver cells transfected with pEGFP-N1/core was significantly lower than that of cells transfected with blank plasmid at three different times after transfection (all P < 0.05). The proliferation ratio of cells transfected with pEGFP-N1/corewas significantly lower than that of cells transfected with blank plasmid. Among three different quasispecies, T, NT and C191 core expression cells, there was no significant difference in the proportion of S- and G0/G1-phase cells. The percentage of apoptotic cells was highest for T (T > NT > C191), and apoptosis was increased in cells transfected with pEGFP-N1/core as the transfection time increased (72 h > 48 h > 24 h). CONCLUSION: These results suggest that HCV genotype 1b core protein induces apoptosis, and inhibits cell- cycle progression and proliferation of Chang liver cells. Different quasispecies core proteins of HCV genotype 1b might have some differences in the pathogenesis of HCV persistent infection and hepatocellular carcinoma.

Metabolism of Mequindox in Isolated Rat Liver Cells

Mequindox(MEQ),3-methyl-2-quinoxalinacetyl-1,4-dioxide,is widely used in Chinese veterinary medicine as an antimicrobial agent and feed additive.Its toxicity has been reported to be closely related to its metabolism.To understand the pathways underlying MEQ's metabolism more clearly,we studied its metabolism in isolated rat liver cells by using liquid chromatography coupled with electrospray ionization hybrid linear trap quadrupole orbitrap(LC-LTQ-Orbitrap) mass spectrometry.The structures of MEQ metabolites and their product ions were readily and reliably characterized on the basis of accurate MS2 spectra and known structure of MEQ.Eleven metabolites were detected in isolated rat liver cells,two of which were detected for the first time in vitro.The major metabolic pathways reported previously for in vitro metabolism of MEQ in rat microsomes were confirmed in this study,including N → O group reduction,carbonyl reduction,and methyl monohydroxylation.In addition,we found that acetyl hydroxylation was an important pathway of MEQ metabolism.The results also demonstrate that cellular systems more closely simulate in vivo conditions than do other in vitro systems such as microsomes.Taken together,these data contribute to our understanding of the in vivo metabolism of MEQ.

Immunological aspects of liver cell transplantation

Within the field of regenerative medicine, the liver is of major interest for adoption of regenerative strategies due to its well-known and unique regenerative capacity. Whereas therapeutic strategies such as liver resection and orthotopic liver transplantation(OLT) can be considered standards of care for the treatment of a variety of liver diseases, the concept of liver cell transplantation(LCTx) still awaits clinical breakthrough. Success of LCTx is hampered by insufficient engraftment/long-term acceptance of cellular allografts mainly due to rejection of transplanted cells. This is in contrast to the results achieved for OLT where longterm graft survival is observed on a regular basis and, hence, the liver has been deemed an immuneprivileged organ. Immune responses induced by isolated hepatocytes apparently differ considerably from those observed following transplantation of solid organs and, thus, LCTx requires refined immunological strategies to improve its clinical outcome. In addition, clinical usage of LCTx but also related basic research efforts are hindered by the limited availability of high quality liver cells, strongly emphasizing the need for alternative cell sources. This review focuses on the various immunological aspects of LCTx summarizing data available not only for hepatocyte transplantation but also for transplantation of non-parenchymal liver cells and liver stem cells.

Liver cell adenoma showing sequential alteration of radiological findings suggestive of well-differentiated hepatocellular carcinoma

A liver tumor 35 mm in diameter was found incidentally in a 40-year-old woman who had no history of liver diseases or the use of oral contraceptives.Radiological diagnostics showed the typical findings of liver cell adenoma(LCA).Dynamic computed tomography revealed that the tumor showed a homogenous enhancement in the arterial phase and almost the same enhancement as the surrounding liver parenchyma in the delayed phase.The tumor was found to contain fat on magnetic resonance imaging.A benign fat containing liver tumor was suggested.However,radiological findings altered,which caused us to suspect that a welldifferentiated hepatocellular carcinoma(HCC)containing fat was becoming dedifferentiated.Partial hepatectomy was performed and the pathological findings showed the typical findings of LCA.This case was an extremely rare LCA,which had no background of risk for LCA and developed the sequential alteration of the radiological findings to suspect well-differentiated HCC.

A New Hematopoietic Stimulating Activity Produced by Fetal Liver Cells

Human fetal liver cells were cultured in vitro for 12h and the supernatant(Fetal liver cell conditioned medium,FLCM)was collected.The effects of FLCM ongranulopoiesis were studied.The results show that when combined with colonystimulating factor(CSF),FLCM could significantly stimulate the proliferation of normalmyctoid progenitor cells(CFU-e),and increase ~3H-TdR incorporation into bone mar-row cells.The data suggest that FLCM contains a CSF synergistic activity.

Differential Proteomics in Malignant and Normal Liver Cell Lines

Objective: To detect differential protein expression in malignant and normal liver cell lines in vitro using the SELDI ProteinChip platform, for investigating the pathogenesis of liver cancer. Methods: Two cell lines, human normal liver cell line L02 and hepatoma cell line SMMC-7721 were cultured routinely, harvested in good condition and lysed. After quantification, the supernatant of the lysate was tested by IMAC3 (Immobilized Mental Affinity Capture) and WCX2 (Weak Cation Exchange) chips on the SELDI-TOF-MS ProteinChip reader. Results: Protein expression differed between the malignant and normal liver cell lines. A total of 20 differentially expressed proteins were found, among which, 7 were captured by the IMAC3 chip and 14 by the WCX2 chip. Peaks at 5,419, 7,979 and 11,265 Da were higher and at 8,103, 8,492, 10,160 and 11,304 Da lower in SMMC-7721 cells by the IMAC3 chip; peaks at 7,517, 7,945 and 7,979 Da were higher and at 5,061, 5,551, 5,818, 7,439, 9,401,10,100, 10,312, 11,621, 11,662, 11,830 and 12,772 Da lower in SMMC-7721 cells by the WCX2 chip. Interestingly, both chips captured the 7,979 Da peak. In addition, the 11,081 Da peak corresponded precisely with the molecular mass of the calcium binding protein S100A10, which may participate in the formation of liver cancer in association with p36. Conclusion: Detecting differential protein expression in malignant and normal liver cell lines using the SELDI ProteinChip platform was simple, sensitive and repeatable. The results we obtained can serve as a basis for investigating the pathogenesis of liver cancer and aid the discovery of new therapeutic targets.

Studies of the Kinetochore Proteins of the Regenerating Liver and the Liver Cells of Rats at Different Stages of Development

The kinetochore composition of rat liver cells was studied by indirect immunofluorescence andimmunoblotting using human anti-kinetochore/centromere autoantibodies(ACAs).Besides threemajor antigens(50kD,42 kD and 34 kD),ACAs used in this study could also identify those of 32-30 kD and 20 kD in newborn rat liver cells,90 kD in old rat liver cells,37 kD and 32-30 kD inregenerating liver cells.These results indicate that some kinetochore antigen(s)may be related to cellproliferation or specific for different stages of development.

Surgical management of spontaneous rupture of primary liver cell carcinoma:a case report

Primary liver cell carcinoma (PLCC) or Hepatocellular carcinoma (HCC) is the most common primary malignant liver tumor in Nigeria. It is a difficult problem in surgery for the diagnosis and therapy of spontaneous liver rupture. The clinical presentation can be varied owing to its clinical signs being usually not specific; therefore, correct diagnosis and management are very important. Without any treatment, the outcome is poor and survival rate is only 10%. Surgeons operate on those patients who present with ruptured PLCC; consisting of packing, hepatic artery ligation and hepatectomy. However, it is often associated with a high mortality rate; as high as 70%, even for the less invasive procedures like packing, argon beam coagulation or hepatic artery ligation. We present a 24-year old lady who had ligation of hepatic artery at an emergency laparotomy for ruptured primary liver cell carcinoma.

The impact of Hypoglycemic Ziyabiti Tablets (HZT) on diabetes mellitus rats and its activity in cultured rat liver cells

Objective Hypoglycemic Ziyabiti Tablets(HZT)is a traditional multicomponent treatment for diabetes in Xinjiang Uyghur Traditional Medicine.The present study aimed to investigate the effect of HZT in diabetic rats and its activity in cultured liver cells to investigate the relative mechanisms.Methods 10 days high-fat diet fed rats were intraperitoneally injected with alloxan(ALX)at next two subsequent days to induce diabetes mellitus(DM).Then were divided into 5 groups:saline,positive DM control and DM groups treated with different doses of HZT.Fasting blood glucose(FBG),total cholesterol(TC),total triglycerides(TG),high-density lipoprotein(HDL-C),fasting insulin(FI),insulin secretion(IS)and insulin sensitivity index(ISI)were measured.The IC_(50) of HZT in L-02 cells was determined by MTT assay,in intact and in paracetamol-induced liver injury(Par),on lactate dehydrogenase(LDH)activity and on glucose consumption.Results HZT decreased FBG and TC(P <0.05),increased IS(P <0.05)and at 440 mg·kg^(-1)·d^(-1) increased FI(P < 0.01).In vitro,HZT at 0.1,0.2,and 0.4 mg/mL decreased LDH activity and promoted glucose consumption.Conclusion The hypoglycemic mechanism of HZT is possibly related to increased insulin secretion from the pancreas and increased utilization of glucose by the liver.

Endosulfan Causes Neoplastic Changes in the Liver Cells of Mice

The rapid growth in global population continues to challenge the world’s ability to provide enough food. As one of the most crucial issues for human development, food production must increase to offset hunger and poverty as well as social unrest. To augment the yield of crops a variety of pesticides like Endosulfan, Rogor, Aldrin, Chlorpyrifos, etc. are being used liberally by the farmers. In the present investigation, Endosulfan was administered orally (daily) by gavage method to female Swiss albino mice group for 4 weeks @ 3.0 mg/kg b.w. After that, they were left for 6 months and then sacrificed and liver tissues were fixed for light microscopy and Transmission Electron Microscopic study. The histopathological study of Endosulfan administered group liver showed hepatocytes with congestion in central vein with less dense cytoplasm, haemorrhaged bile duct, degenerated cytoplasm and central vein with vacuolations in sinusoidal spaces. Neoplastic changes in hepatocytes are the major finding of study. The ultrastuctural study revealed dilation in the nuclear pore complex and massive movement of cytoplasmic material from cytoplasm to the nucleus which is major finding which denotes neoplastic changes. Presence of abundant free lying polyribosomes in the cytoplasm, which denotes neoplastic changes in the cellis also one of theimportant finding observed. The present study thus deciphers that Endosulfan toxicity leads to onset of neoplasia thence carcinogenesis in liver cells in Swiss albino mice which is the novel finding in the field of toxicology.

Cultivating human liver cell line (CL-1) on microcarriers as biomaterial of bioartificial liver

objective: To cultivate human liver cell line (CL-1) on microcarriers and study the synthetic and transformational function of this culture system. Methods:CL-1 were cultivated on Cytodex-3 microcarriers. The cell growth was kinetically inspected with light microscope and scanning electronic microscope on the lst, 3rd, 5th, 7th, 9th day, and the amount of diazepam transformation and albumin synthesis were deter mined at the same time. Results:On 7th day after inoculating, the CL-1 cell density could reach 2. 16 ×106/ ml ; the amount of diazepam trans formation was 619 μg and albumin synthesis 78. 23 μg. Conclusion:CL-1 can be cultivated to a high density on microcarriers and has hepatic specific biotransformation and biosynthesis functions. So the culture system may be further studied for being used as the biomaterial of bioartificial liver.

Malignant Transformation of Human Embryonic Liver Cells Induced by Hepatitis B Virus and Aflatoxin B_1

In order to investigate the effect of hepatitis B virus (HBV) and aflatoxin B 1 (AFB 1) on hepatocarcinogenesis, the human embryonic liver cells infected with HBV were transplanted to nude mice by subcutaneous route and the transplanted mice were divided into 4 groups for study, in which the group A of mice was injected with HBV-infected human embryonic liver cells and followed by injections of AFB 1 once a week (HBV+AFB 1); the group B was treated with HBV as group A, but no AFB 1 was given (HBV +); the group C was injected with normal human embryonic liver cells and AFB 1 was used as group (AFB 1 +) and the group D or control group was injected with normal embryonic liver cells without addition of AFB 1. The experimental results showed that the incidences of tumor formation in different groups were 27.3% (6/22) in group A; 0% (0/13) in group B; 13.3% (2/15) in group C and 0% (0/14) in group D respectively. All the tumors formed were proved to be human hepatocellular carcinoma (HCC) by pathological examinations and the tumor tissues were anthrogenetic as demonstrated by EMA monoclonal antibody. The HBV-X and HBV-S genes could be detected in the tumor tissues by means of slot hybridization and PCR amplification, indicating that the HBV-DNA genes had integrated into DNA of host cells. Thus, we have successfully induced the human HCC through HBV infection and introduction of AFB 1 with a synergistic effect between HBV and AFB 1 in hepatocarcinogenesis.

Liver cell transplantation for Crigler-Najjar syndrome type Ⅰ: Update and perspectives

Liver cell transplantation is an attractive technique to treat liver-based inborn errors of metabolism. The feasibility and efficacy of the procedure has been demonstrated, leading to medium term partial metabolic control of various diseases. Crigler-Najjar is the paradigm of such diseases in that the host liver is lacking one function with an otherwise normal parenchyma. The patient is at permanent risk for irreversible brain damage. The goal of liver cell transplantation is to reduce serum bilirubin levels within safe limits and to alleviate phototherapy requirements to improve quality of life. Preliminary data on Gunn rats, the rodent model of the disease, were encouraging and have led to successful clinical trials. Herein we report on two additional patients and describe the current limits of the technique in terms of durability of the response as compared to alternative therapeutic procedures. We discuss the future developments of the technique and new emerging perspectives.