Cellular and molecular changes in the early stages of chemical hepatocarcinogenesis in the rat.

The early cellular and molecular changes in the Solt-Farber model of hepatocarcinogenesis with and without initiation was studied by using histochemical, immunohistochemical, and in situ hybridization techniques. Increased cellularity was observed in the periductal space in both models 32 to 56 h after partial hepatectomy. These periductal cells and Ito cells were the only cells that became labeled with tritiated thymidine in the uninitiated liver model. Forty-five to 60% of the labeled periductal cells were positive for gamma-glutamyltranspeptidase. From the periductal area the cells that were positive for antibody raised against oval cells (OV-6) infiltrated into liver parenchyma and were followed by desmin-positive Ito cells. The number of Ito cells in the uninitiated model 6 days after partial hepatectomy was 3.5 times higher in the area occupied by oval cells than elsewhere in the liver. The first alpha-fetoprotein (AFP)-positive cells appeared either as individual cells or as pseudoductal formations 32 or 56 h after partial hepatectomy at the periphery of the periductal space in both initiated and uninitiated animals. A combination of in situ and immunohistochemistry revealed that the OV-6-positive cells were AFP positive, whereas desmin-positive cells were AFP negative. Glutathione S-transferase P (GST-P) transcripts could be found mainly in OV-6-positive oval cells. Bile duct cells were positive for GST-P and negative for transforming growth factor beta 1, whereas cells in the periductal space were positive for both of these transcripts. The GST-P-positive early preneoplastic lesions showed a similar distribution pattern as that of oval cells; the preexisting hepatocytes became trapped between small basophilic hepatocytes that showed either irregular or pseudoalveolar arrangement. This raises the question as to whether cells which are stem cell-like are among the target cells in the Solt-Farber model of hepatocarcinogenesis. Proliferation of transforming growth factor beta 1-producing, desmin-positive cells (Ito cells) and multipotent oval cells in a close proximity to each other indicates an intricate relationship between Ito cells and oval cells in liver that warrants further investigation.

Experimental intraocular malignancy: the effect of intracameral perfusion.

Transplantable Brown-Pearce carcinoma was adapted successfully in the rabbit anterior chamber. Regression of tumor growth was attained on tri-weekly perfusion of the AC with 10 micromolar of methotrexate. Tumor cyclic nucleotide phosphodiesterase (PDE) and protein activator were found to be markedly depressed during the course of chemotherapy and the PDE cAMP/cGMP ratio was similarly altered. Corroborative light and electron-microscopic studies showed specific alterations of intracellular organelles in relation to MTX and tumor cell death. These findings suggest that metabolic pathways of cyclic nucleotides are important biochemical modulators of neoplastic cells. The method of intraocular perfusion precludes systemic toxic effects and avoids compromising the animals' immunocompetence.

Modulation of asialoglycoprotein receptor levels in rat liver by phenobarbital treatment.

The effect of the liver tumor promoter, phenobarbital, on the level of the asialoglycoprotein receptor (ASGP-R) was examined in adult rat liver. ASGP-R, a liver-specific cell surface membrane protein, was studied using antibody against this receptor together with immunofluorescence techniques and radioreceptor assay with asialofetuin as the ligand. Both acute and chronic phenobarbital administration decreased the number of receptors per cell; partial hepatectomy had a similar effect on the number of receptors per cell. However, after phenobarbital administration, the receptor-deficient areas were centrilobular, whereas after partial hepatectomy, ASGP-R positive and negative areas were intermingled throughout the liver lobule but were most pronounced in the periportal area. Phenobarbital treatment, in contrast to its effect on the ASGP-R level, did not change the cell surface binding of concanavalin A on rat hepatocytes. Four days after birth the number of hepatocytes with surface receptors was 50% of that in the adult rats. At 10 days after birth the number of ASGP-R positive cells was the same as in adult rats, although the receptor density was significantly lower than in adults. Treatment with a single dose of chemical carcinogen one day after birth combined with promotion by phenobarbital resulted in a significant reduction of ASGP-Rs in pre-neoplastic and neoplastic areas of livers. Whereas the pre-neoplastic and neoplastic areas displayed uniform reduction in the ASGP-R, normal parts of the liver showed receptor-deficiency primarily in the centrilobular areas.

Occurrence of oval-type cells in hepatitis B virus-associated human hepatocarcinogenesis.

Proliferation of a new population of epithelial cells with distinct structure, as well as cytokeratin and alpha-fetoprotein expression, was observed in nonneoplastic liver tissues from 14 cases (13 hepatitis B virus-positive) of human hepatocellular carcinoma. These cells were characterized by oval nuclei; scant, pale cytoplasm; small cell size; and cross-reaction with a monoclonal antibody against rat oval cells. These putative human oval cells were strongly positive for cytokeratin 19 and displayed considerable heterogeneity in alpha-fetoprotein and albumin expression. The oval cells were most prominent in actively regenerating nodules and in liver tissue surrounding the cancer. Oval cells and transitional types of cells appear to be the principal producers of alpha-fetoprotein in the regenerating liver. Cancer cells positive for cytokeratins 8, 18 and 19 were observed in half the hepatocellular carcinomas studied. The data suggest that a new cell population structurally similar to oval cells seen in early stages of chemical hepatocarcinogenesis in rats is consistently present in regenerating liver lesions associated with human hepatocellular carcinoma. Furthermore, it is possible that the proliferation of these oval-type cells may partly account for the elevation of serum alpha-fetoprotein frequently seen in precancerous stages of hepatitis B virus-associated human hepatocellular carcinoma.

Expression of hepatocyte growth factor and c-met genes during hepatic differentiation and liver development in the rat.

Hepatocyte growth factor (HGF) is a potent mitogen for mature hepatocytes in vitro. The receptor for HGF has recently been characterized as the product of the proto-oncogene c-met. We have examined the possible involvement of HGF in hepatic growth and differentiation in the rat. The experimental systems used were acetylaminofluorene treatment combined with partial hepatectomy to induce proliferation and differentiation of oval cells in adult liver and the pre- and postnatal liver. In the acetylaminofluorene model, Northern blot analysis showed that level of HGF transcripts increased one day after partial hepatectomy, reached a peak by day 6, were maintained at that level until day 13, and then declined, reaching normal level at 20 days. The expression of c-met also increased gradually, reached a peak around 9 to 13 days after partial hepatectomy, at which time oval cell proliferation was most prominent. In the developing liver, an elevated level of HGF transcripts was found between 4 and 21 days after birth. The expression of c-met also slightly increased at the same time. In situ hybridization showed that the transcripts for HGF were localized in desmin-positive Ito cells, whereas the transcripts for c-met were strongly expressed by oval cells. We have shown earlier that Ito cells and oval cells proliferate simultaneously and exist in close proximity in the acetylaminofluorene model and that Ito cells are a primary source of growth factors such as transforming growth factor-alpha and acidic fibroblast growth factors. The data presented here suggest that HGF is, in combination with other growth factors, involved in the proliferation and differentiation of oval cells via a paracrine mechanism.

Expression of transforming growth factor-alpha in regenerating liver and during hepatic differentiation.

Both the level of expression and cellular distribution of transcripts for transforming growth factor-alpha (TGF-alpha) were studied in adult rat liver after partial hepatectomy and during hepatic differentiation in fetal, neonatal, and adult livers by northern blot analysis and in situ hybridization. A marked increase in the expression of TGF-alpha was observed in neonatal livers and in adult livers after partial hepatectomy and during hepatic regeneration following modification of the Solt-Farber protocol. Quantitation of silver grains after in situ hybridization with a TGF-alpha riboprobe revealed a sixfold to eightfold increase in fetal and neonatal hepatocytes. Moreover, the expression of TGF-alpha in the liver 3 wk after birth was still fourfold higher than that of the adult quiescent liver. Both proliferating oval cells and basophilic foci of hepatocytes generated by modification of the Solt-Farber protocol were positive for TGF-alpha transcripts. The level of TGF-alpha transcripts was sixfold higher in the basophilic foci than in the surrounding liver. High concentrations of TGF-alpha transcripts were observed in the oval cells that lined pseudoducts and in the transitional cells proliferating within the ducts. The combination of in situ hybridization and immunocytochemistry using cell-specific antibodies revealed the presence of TGF-alpha transcripts in both oval cells and in perisinusoidal stellate cells. The observation that TGF-alpha transcripts were found both in primitive liver epithelial cells and perisinusoidal stellate cells suggests that this growth factor, in addition to its mitogenic action, may also have other important functions in the liver.

Activation of hepatic stem cell compartment in the rat: role of transforming growth factor alpha, hepatocyte growth factor, and acidic fibroblast growth factor in early proliferation.

We have demonstrated previously a pronounced increase in the expression of hepatocyte growth factor (HGF) (Z. Hu, R. P. Evarts, K. Fujio, E. R. Marsden, and S. S. Thorgeirsson, Am. J. Pathol., 142: 1823-1830, 1993), transforming growth factor alpha (TGF-alpha) (R. P. Evarts, H. Nakatsukasa, E. R. Marsden, Z. Hu, and S. S. Thorgeirsson, Mol. Carcinog., 5: 25-31, 1992), and acidic fibroblast growth factor (aFGF) (E. R. Marsden, Z. Hu, K. Fujio, H. Nakatsukasa, S. S. Thorgeirsson, and R. P. Evarts, Lab. Invest., 67: 427-433, 1992) that coincided with the proliferation and differentiation of putative hepatic stem cells and perisinusoidal stellate (Ito) cells. Here, we examine the earliest stages of stem cell activation in rat liver using an experimental model involving treatment with acetylaminofluorene and partial hepatectomy (R. P. Evarts, P. Nagy, E. Marsden, and S. S. Thorgeirsson, Carcinogenesis (Lond.), 8: 1737-1740, 1987). Histochemical identification of stem cell progeny and Ito cells was accomplished by OV6 and desmin antibodies, respectively. Expression of the 2.1-kilobase alpha-fetoprotein transcripts and the concomitant DNA synthesis ([3H]thymidine label) were used as indicators for the activation of the stem cell compartment. Expression of HGF, TGF-alpha, and aFGF was analyzed at the time of partial hepatectomy and 4, 12, 24, 48, 72, and 92 h after the operation. [3H]-Thymidine-labeled OV6- and desmin-positive cells were present in the portal space and in the Glisson capsule 4 h after partial hepatectomy.(ABSTRACT TRUNCATED AT 250 WORDS)

Hepatic regeneration in vitamin A-deficient rats: changes in the expression of transforming growth factor alpha/epidermal growth factor receptor and retinoic acid receptors alpha and beta.

We have studied the effect of vitamin A deficiency on the expression of transforming growth factor alpha (TGF-alpha), hepatocyte growth factor, acidic fibroblast growth factor, and TGF-beta 1 after partial hepatectomy of vitamin A-supplemented and vitamin A-deficient rats. In addition, the expressions of epidermal growth factor receptor and retinoic acid receptors alpha (RAR alpha) and beta (RAR beta) were studied. Partial hepatectomy was performed on the animals from the vitamin A-supplemented and -deficient groups at the age of 10 weeks when the weights of the animals on the deficient diet had reached a plateau. Two animals from each group were sacrificed before the operation and also 12, 24, 48, and 72 h and 5 days after the operation. Partial hepatectomy of the vitamin A-deficient rats leads to a focal necrosis of liver followed by a rapid restoration of liver mass. Expression of the TGF-alpha and epidermal growth factor receptor was highly elevated in the livers of deficient animals after partial hepatectomy. In the vitamin A-supplemented animals, the level of epidermal growth factor receptor was down-regulated following partial hepatectomy. Proliferation of oval cells in vitamin A-deficient livers following partial hepatectomy and subsequent increase in 2.1-kilobase alpha-fetoprotein mRNA was observed, suggesting an activation of the stem cell compartment. Another unexpected result was an inverse relationship between RAR beta and RAR alpha expression, the latter becoming the major species after partial hepatectomy in animals on the vitamin A-deficient regimen.(ABSTRACT TRUNCATED AT 250 WORDS)

Expression of stem cell factor and its receptor, c-kit, during liver regeneration from putative stem cells in adult rat.

BACKGROUND: Stem cell factor (SCF) and its receptor, c-kit, are known to play important roles in hematopoiesis, melanogenesis, and gametogenesis. The biologic effects of the SCF/c-kit system are believed to involve survival, proliferation, and migration of early stem cell progeny. Although SCF and c-kit receptor are widely expressed during normal embryonic development, their expression in the adult is limited. EXPERIMENTAL DESIGN: The expression of SCF and c-kit genes was examined during liver regeneration via the oval cell compartment utilizing partial hepatectomy (PH) combined with the administration of a noncarcinogenic dose of 2-acetylaminofluorene (AAF) for 8 days (AAF/PH model). RESULTS: Both the ligand and the receptor genes were expressed during the early stages of oval cell proliferation after partial hepatectomy in the AAF/PH model, while neither simple partial hepatectomy nor AAF administration alone induced a noticeable expression of the SCF/c-kit system. The level of SCF mRNA increased within 12 hours after partial hepatectomy and reached a peak around day 4. Thus, the expression of SCF preceded the major expansion of the oval cell compartment. The level of c-kit transcripts gradually increased from the 12-hour time point and stayed elevated until day 11, when a large proportion of the oval cells differentiated into small basophilic hepatocytes. Separation of liver cells at day 3 in the AAF/PH model into parenchymal and nonparenchymal fractions demonstrated that the expression of both SCF and c-kit receptor genes was restricted to the nonparenchymal cells. Furthermore, in situ hybridization revealed that the c-kit transcripts were restricted to oval cells, whereas the SCF transcripts were expressed in both oval cells and Ito cells. CONCLUSIONS: The transcripts for the c-kit receptor are expressed in the early progeny of the hepatic stem cells. The SCF/c-kit system may, possibly in combination with other growth factor/receptor systems, be involved in the early activation of the hepatic stem cells as well as in the expansion and differentiation of oval cells.

Expression of fibroblast growth factor receptors flg and bek during hepatic ontogenesis and regeneration in the rat.

Fibroblast growth factors (FGFs) mediate their cellular responses through specific cell surface receptors. Previous studies have indicated that acidic FGF is involved in liver regeneration and hepatic differentiation via the stem cell compartment, as well as in liver development (Marsden et al., Lab. Invest. 67:427-433, 1992). To further define the role of acidic FGF and its receptor systems in the liver, we examined the expression and cellular localization of FGF receptor-1 (flg) and FGF receptor-2 (bek) in the liver by Northern blot analysis and in situ hybridization techniques during liver regeneration, hepatic differentiation, and ontogenesis. In the normal adult liver, flg is absent in hepatocytes, whereas a low level of bek can be detected. The transcripts for bek increased, while flg exhibited little change during liver regeneration after partial hepatectomy. Both flg and bek were expressed at high levels in the developing liver. flg was expressed at a high level in embryonic liver and further increased after birth, whereas a significant increase of bek occurred at the postnatal stage of liver development. In the 2-acetylaminofluorene/partial hepatectomy model, both flg and bek are expressed at high levels during the period of active proliferation and differentiation of oval cells. In situ hybridization showed that flg was mainly localized in oval cells, whereas bek was highly expressed in both oval and Ito cells. The data suggest that bek is involved in the proliferation of mature hepatocyte proliferation during liver regeneration, while flg is characteristic of primitive hepatic cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Expression of acidic fibroblast growth factor in regenerating liver and during hepatic differentiation.

BACKGROUND: Acidic fibroblast growth factor belongs to a family of growth factors that show a high affinity for heparin sulfate proteoglycans. In vitro, it participates in various cellular functions including proliferation, differentiation, angiogenesis, and cell migration, but in vivo, the physiologic role of this growth factor is still not clearly defined. EXPERIMENTAL DESIGN: The level of expression and also cellular distribution of transcripts for acidic fibroblast growth factor (aFGF) were studied in adult rat liver after partial hepatectomy and during hepatic differentiation in fetal, neonatal, and adult livers by Northern analysis and in situ hybridization techniques. RESULTS: After partial hepatectomy a significant increase in the transcripts for aFGF was observed at 24 hours, whereas at 4 and 12 hours after the operation, the level of transcripts were similar to those of sham-operated animals. In the postnatal liver a high level of aFGF expression was present when the most evident transition from 2 to 3 cell thick hepatic cords to normal hepatic structure is taking place (Ogawa K, Medine A, Farber E. Br J Cancer 1979;40: 782-90). In contrast during the prenatal period, when the liver is still a hemopoietic organ and only a small number of sinusoids are present, low level of aFGF transcripts could be found. Animals treated with 2-acetylaminofluorene and partial hepatectomy (Evarts RP, Nagy P, Marsden E, Thorgeirsson SS. Carcinogenesis 1987;8:1737-40) displayed a marked increase in hepatic aFGF transcripts at the peak of proliferation of primitive liver epithelial cells (oval cells) and perisinusoidal stellate cells (Ito cells) in addition to hepatocytes. In situ hybridization combined with immunocytochemistry using oval and Ito cell specific antibodies revealed the presence of transcripts both in oval cells and Ito cells. Basophilic areas composed of small hepatocytes had a 3-fold increase in the level of transcripts as compared with the surrounding hepatocytes. CONCLUSIONS: These experiments demonstrate that the expression of aFGF is highest during the late stages of hepatic morphogenesis in newborn animals as well as during hepatic differentiation in adult liver.

Expression of alpha-fetoprotein and stem cell factor/c-kit system in bile duct ligated young rats.

The existence of a facultative hepatic stem cell compartment in bile ductules that participates in the renewal process of epithelial cell populations in the liver is well documented. The present study was undertaken to determine whether the immature bile epithelium responds differently to growth stimulus induced by bile stasis to that seen in the adult animal. In addition, the possible involvement of the growth factor/receptor systems associated with early activation of hepatic stem cells in bile duct proliferation was also examined. Bile duct ligation was used to induce the proliferation of bile epithelial cells. The expression of full-length alpha-fetoprotein (AFP) was used as an indicator for activation of the stem cell compartment. AFP was highly and selectively expressed in small bile ducts 7 days after bile duct ligation in immature rats up to 5 weeks of age. Although no significant increase in the expression of stem cell factor (SCF) c-kit, hepatocyte growth factor (HGF), and transforming growth factor-alpha (TGF-alpha) was observed 7 days after bile duct ligation in adult rats, the expression of all these growth factors was increased in bile duct ligated rats up to 5 weeks of age. These results suggest that the bile ductular epithelium in the young rats responds to bile stasis in a fashion that is phenotypically similar to that seen during early activation of hepatic stem cells in adult liver.

Precursor-product relationship between oval cells and hepatocytes: comparison between tritiated thymidine and bromodeoxyuridine as tracers.

The expansion and differentiation of oval cells in the acetylaminofluorene (AAF)/partial hepatectomy (PH) model was studied utilizing pulse-chase labeling with both tritiated thymidine ([3H]TdR) and bromodeoxyuridine (BUdR). Animals in which a significant decrease in serum albumin and increase in alanine aminotransferase and bilirubin were observed demonstrated the most prominent differentiation of oval cells into hepatocytes. Administration of [3H]TdR or BUdR, either individually or together, to the animals on day 6 after partial hepatectomy resulted in labeling of the majority of the oval cells by days 7 and 9 after PH. A striking difference in the distribution of [3H]TdR- and BUdR-labeled cells in the double labeling experiments was observed on day 11, at which time the number of [3H]TdR-labeled cells increased 6-fold and that of double labeled cells decreased 2-fold. Furthermore, on day 11 the basophilic foci were weakly positive for BUdR and negative at later time points in animals receiving BUdR alone or together with [3H]TdR. In contrast, the cells in basophilic foci as well as transitional cells were positive for [3H]TdR. Cells heavily labeled with both [3H]TdR and BUdR were present at all time points, indicating an inhibition of the proliferative activity. Pulse labeling of rat liver epithelial cells with BUdR in vitro demonstrated that immunodetection of BUdR was lost after three or more cell divisions. We conclude that the BUdR tagging method is particularly sensitive to label dilution during cell cycling and may not be suitable for establishment of a precursor-product relationship between cell lineages when the progenitor population proliferates more than three times.

Effect of vitamin A deficiency on the integrity of hepatocytes after partial hepatectomy.

The effect of vitamin A deficiency on hepatic regeneration in male and female rats was studied after partial hepatectomy. A fourfold increase in the number of positive dUTP end-labeled nuclei was observed in the deficient animals as early as 30 minutes after partial hepatectomy and their number reached a peak by 8 hours after the operation. The bile duct cells were both morphologically and biochemically intact at all time points. Administration of retinyl palmitate 1 hour before partial hepatectomy significantly reduced the number of positive nuclei, and treatment with retinyl palmitate 24 or 48 hours before the operation reduced the number of positive cells to the level observed in control vitamin A-supplemented rats. The level of transcripts for c-jun, c-fos, c-myc, and transforming growth factor-beta 1 were increased for an extended period of time in livers of deficient animals, whereas the expression of both p53 and max were unchanged. Immunocytochemistry demonstrated the presence of latent transforming growth factor-beta 1 in cells showing evident apoptotic or necrotic changes in their nuclei. This study demonstrates the importance of vitamin A for the survival of hepatocytes both in intact vitamin A-deficient liver and after partial hepatectomy, whereas the ductal cells appear to be less sensitive to vitamin A deficiency.

Expression of transforming growth factor alpha/epidermal growth factor receptor, hepatocyte growth factor/c-met and acidic fibroblast growth factor/fibroblast growth factor receptors during hepatocarcinogenesis.

It is widely believed that abnormal production of polypeptide growth factors, together with other molecular alterations, play an important role in neoplastic development. Transforming growth factor alpha (TGFalpha), hepatocyte growth factor (HGF) and acidic fibroblast growth factor (aFGF) are the three major growth factors that contribute to liver regeneration occurring via both hepatocyte replication and oval cell proliferation. It is not clear, however, whether and to what extent these growth factors are also involved in hepatocarcinogenesis. In the present study, the gene expression of TGFalpha, HGF and aFGF and their corresponding receptors was examined by Northern blotting and in situ hybridization during hepatocarcinogenesis induced by the Solt-Farber protocol. All three growth factor/receptor systems, TGFalpha/epidermal growth factor receptor (EGFR), HGF/c-met and aFGF/FGF receptors (flg and bek) were significantly elevated at early time points when oval cells were proliferating. Their respective expression decreased after 1 month and remained at a low level until the development of liver tumors. In all hepatocellular carcinomas (HCC) examined, the transcripts of TGFalpha and aFGF were highly expressed, while those of HGF were low. With regard to the receptor expression in the tumors, EGFR was present at varying levels, c-met was expressed at higher levels and flg increased significantly, whereas bek remained at low levels. These data suggest that TGFalpha and aFGF are the major growth factors involved in the progression of HCC, and that the signal of aFGF is mainly transduced by the receptor flg in HCC. Furthermore, HCC cells were phenotypically very similar to oval cells with regard to the gene expression of growth factor/receptor systems. These results, along with the finding that all the HCC cells are positive for the oval cell antigen OV6, and that cytokeratin 19 is heavily expressed in both tumor and oval cells, strongly suggest that at least some of the HCC induced by the Solt-Farber protocol may be derived from oval cells.

Coexpression of stem cell factor and c-kit in embryonic and adult liver.

Stem cell factor and its receptor c-kit constitute an important signal transduction system implicated in survival, proliferation, and differentiation of stem cells in hematopoiesis, gametogenesis, and melanogenesis. In the present study we used both immunocytochemical methods and Western analysis to demonstrate the presence of this cytokine/receptor system in both embryonic and adult rat liver. Stem cell factor was present in the ductular cells around the portal vein during the late embryonic stage of the liver. In the adult liver both bile ducts and bile ductules were positive for stem cell factor and c-kit. When the activation of the liver stem cell compartment was induced by combining administration of acetylaminofluorene and partial hepatectomy, both stem cell factor and c-kit were expressed in the infiltrating oval cell population, but absent in the newly formed basophilic hepatocytes. Activation of oval cell proliferation following administration Of D-galactosamine also produced a similar but less prominent increase in the level of the stem cell factor. Our data suggest that the stem cell factor/c-kit signal transduction system is involved in the development of bile ducts and that it may also be an important member of the growth factor/receptor systems associated with the biology of liver stem cells.

Expression of leukemia inhibitory factor and its receptor during liver regeneration in the adult rat.

Leukemia inhibitory factor (LIF) is a polyfunctional cytokine that was discovered in the conditioned medium from Buffalo rat liver cells. In the liver, LIF is known to induce acute phase proteins in the hepatocytes. No comprehensive study has yet been performed on the physiological role of this cytokine during liver regeneration. Thus, we studied the level of expression and cellular distribution of transcripts for LIF, its receptor (LIFR), and signal transducing subunit gp13O during rat liver regeneration after both simple partial hepatectomy (PH) and the oval cell activation induced by the combination of 2-acetylaminofluorene and PH. In addition, the expression of an acute phase protein alpha1-acidglycoprotein was examined. The level of transcripts for LIF and its receptor subunits increased and remained elevated during oval cell expansion. In contrast, after PH, the transcripts were induced only transiently, showing a peak 24 hours after the operation. LIF and receptor subunits were expressed in both parenchymal and nonparenchymal fractions in the 2-acetylaminofluorene/PH model, but the level of expression was most pronounced in the nonparenchymal fraction. In situ hybridization clearly revealed a strong expression of LIF, LIFR, and gp13O in the oval cells and demonstrated only a weak expression in the parenchyma. Interestingly, transcripts of alpha1-acidglycoprotein were exclusively detected in the parenchyma. These results suggest a phenotypic difference between oval cells and hepatocytes in their signaling through gp130. We hypothesize that the LIF/LIFR gp130 system may be involved in the expansion and differentiation of the liver stem cell compartment.