THEMES
Lessons From Genetically Engineered Animal Models
V. Knocking out genes to study liver regeneration: present and future*

Nelson Fausto

Department of Pathology, University of Washington School of Medicine, Seattle, Washington 98195-7470


    ABSTRACT
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Studies utilizing knockout mice have contributed important new knowledge about the mechanisms that initiate liver regeneration. New mouse lines need to be established to address major questions about these mechanisms, targeting genes for which there is experimental evidence of their involvement in important pathways. Development of conditional, liver-specific knockout mice would be of great value for these studies.

hepatocytes; cytokines; growth factors; transcription factors


    INTRODUCTION
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IT HAS BEEN ESTIMATED THAT ~70 genes increase their expression in the immediate early phase response after partial hepatectomy (18). This number is bound to grow when DNA arrays that can measure the expression of 5,000-25,000 genes come into use for the analysis of liver regeneration. However, only a fraction, perhaps a very small fraction, of these genes may be directly related to hepatocyte proliferation. During liver regeneration, hepatocytes retain their differentiated phenotype and compensate for the loss of cells or whole tissue. Thus differential expression of genes at the start of the process may be mostly related to the maintenance and enhancement of hepatic metabolic functions needed for homeostasis and survival. How then is it possible to learn which of these many genes plays a crucial role in hepatocyte replication?

The most direct way to determine whether a particular gene is essential for liver regeneration is to modify its expression: overexpression in transgenic mice and functional inactivation or deletion in knockout animals. Many of the advances of present day biomedical research are due in no small part to the use of gene manipulation in mice and the analysis of morphological and functional alterations created as a consequence of gene expression changes. For pathologists for whom the understanding of normal and abnormal physiology cannot be dissociated from the knowledge of cell, tissue, and organ morphology, the "discovery" of morphology by molecular biologists analyzing gene function is both gratifying and to some extent amusing because of its obvious nature. Transgenic and knockout technologies also have their pitfalls. For transgenics, overexpression of the transgene is generally many times higher than that which occurs in physiological or even pathological conditions. Often the transgene is driven by a potent promoter that may not be normally expressed in the cells that are being examined or the transgene itself is a viral product that may be harmful to the cells. Moreover, overexpression of the transgene throughout the life of the animal may modify the expression of other genes, which may themselves be suppressed or overexpressed to compensate for the high and nonphysiological expression of the transgene. For knockouts, one of the major problems that complicates data analysis is the existence of redundant or interactive pathways, particularly for critical cell functions. Thus the lack of an abnormal phenotype in a knockout mouse does not necessarily mean that the gene in question is not important. Its role might only become apparent by functional inactivation of another gene in a double knockout. Moreover, development and growth can be normal in some knockouts, but they may fail to respond to or survive certain types of stresses such as partial hepatectomy, carcinogen application, or toxic injury.

In any case, gene manipulation in mice will gain even more prominence in the near future, as the physiological roles of thousands of genes sequenced in the human genome project beg identification. New technology that permits conditional and tissue-specific expression of genes will go a long way to minimize many existing problems. Will it then be wise to create knockout models to study every immediate early gene whose expression is altered after partial hepatectomy? Such a quest would be a costly and relatively unrewarding fishing expedition, although admittedly a sophisticated fishing trip, which with some luck could yield a big fish. It would be much more appropriate, however, to limit the development of knockout mouse models for genes that have been well characterized as to function and patterns of expression and that have been shown by other types of experimental evidence (for instance, antibody or antisense blockage, effects on DNA replication in hepatocyte cultures under well controlled conditions, and so forth) to fit into pathways that are important for hepatocyte replication. To perform this task in a systematic rather than haphazard way, it is necessary to develop a model of how liver regeneration works. This model may even be wrong, but, if it is consistently tested, proving it wrong by actual experiments could be as important as demonstrating the correctness of the hypothesis. The main advantage of this type of strategy is that it forces investigators to make a distinction between central and peripheral issues and to focus on important questions about the mechanisms of liver regeneration.


    LIVER REGENERATION AS A MULTISTEP PROCESS
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A simplified view of liver regeneration is that it is a multistep process that starts with gene expression changes that prime quiescent (G0) hepatocytes to enter the cell cycle (6). This phase, which involves the activation of the transcription factors nuclear factor for kappa -chain in B cells (NFkappa B), signal transducer and activator of transcription-3 (STAT3), activator protein-1, and CCAAT enhancer binding protein (C/EBPbeta ) as well as the expression of immediate early genes, is reversible and is neither sufficient by itself to cause DNA replication nor specific for proliferation. Priming is, however, required to make hepatocytes respond fully to at least two growth factors, hepatocyte growth factor (HGF) and transforming growth factor-alpha (TGF-alpha ), which are highly expressed during liver regeneration (19). Once hepatocytes progress through the cell cycle and go over a G1 restriction point, the cells are irreversibly committed to replication. Expression of cyclin D1 during liver regeneration is a good marker to indicate when hepatocytes become mostly autonomous in their replication capacity and no longer require growth factors (2). From that point on, the cell cycle machinery takes over and the hepatocyte functions as other cell types do when reaching the G1/S stage of cell cycle progression. Thus, to look at liver regeneration as a unique biological process, it is logical to focus on its mechanisms of initiation and the activation of pathways that lead from proliferation competence to replication autonomy. However, liver regeneration as a unique phenomenon does not end there. One of the more fascinating and least understood features of the process is why it stops when it does. There are no descriptions in the literature of livers that, at the end of the regenerative process, become significantly larger than the original size (the regenerated liver mass is ±10% of the original hepatic mass). There must be mechanisms that link the replicative activity of hepatocytes in the regenerating liver with body functional demands, but the nature of these mechanisms is not known.


    KNOCKOUT MOUSE MODELS USED IN STUDIES OF LIVER REGENERATION
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The most productive applications of knockout mice to studies of liver regeneration have been directed at the analysis of genes believed to participate in the initiation of the process (Table 1). Studies with tumor necrosis factor (TNF) receptor type 1 (TNFR-1) and type 2 (TNFR-2) knockout mice (23-25) were based on previous data on the effect of anti-TNF antibodies (1). Data generated from these animals fit well with those obtained with interleukin (IL)-6 and inducible nitric oxide synthase (iNOS) knockouts (3, 14). The combined data from these experiments demonstrated that cytokines participate in the initiation of liver regeneration and that TNF is essential for liver regeneration after partial hepatectomy and necessary for optimal growth after CCl4 injury (23-25). TNF proliferative signaling depends on TNFR-1 and involves the sequential activation of NFkappa B, IL-6, and STAT3. IL-6 may be a key element in this sequence and has both proliferative and anti-apoptotic effects (3). Partial hepatectomy causes liver injury rather than regeneration in iNOS knockout mice, despite the induction of TNF, NFkappa B, IL-6, and STAT3 (14). This suggests that nitric oxide induction after partial hepatectomy protects against cytokine injury.

                              
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Table 1.   Gene knockout mice used in studies of liver regeneration

The TNFR-1-mediated pathway is certainly not the only one involved in the initiation of liver regeneration. C/EBPbeta is a leucine zipper transcription factor that transactivates several genes involved in the acute phase response. C/EBPbeta -deficient mice have defective liver regeneration (7). In these animals, DNA synthesis was 25% of normal and the expression of cyclins B and E but not D1 was reduced. C/EBP effects do not depend on IL-6 because IL-6 knockout mice showed no alteration in C/EBP activity.

The analysis of liver regeneration in urokinase type plasminogen activator (uPA)- and uPA receptor (uPAR)-deficient mice examines another aspect of the initiation of liver regeneration. It tested the hypothesis that uPA plays an essential role in the initiation of liver regeneration by converting HGF into the active double-chain form capable of receptor binding (11). Lack of uPA activity in the knockout mice inhibited liver regeneration (15). However, uPAR deficiency had no effect on the growth process, indicating that whatever role uPA may play in liver regeneration it does not require binding to the receptor. It remains to be determined whether uPA deficiency prevented activation of HGF after partial hepatectomy.

cAMP-responsive promoter element modulator (CREM) is a transcription factor activated by phosphorylation of a serine residue by kinases stimulated by cAMP. Servillo et al. (17) showed that CREM activity is greatly increased after partial hepatectomy. Its deficiency in knockout mice delayed entry of hepatocytes into the S phase and disrupted the synchronization of hepatocyte replication. It has not yet been established whether increased CREM activity after partial hepatectomy is a cytokine-mediated event or by what specific mechanisms synchronization is lost. In any case, the work of Servillo et al. (17) uncovered a cAMP-dependent regulatory pathway that is important for liver regeneration.

The knockout animal models described so far do not involve deficiency of hepatocyte structural components. Hepatocytes contain cytokeratins (CK) 8 and 18 as their only cytokeratins (bile duct cells have CK7 and CK19). Mice deficient in CK8 develop colon hyperplasia and inflammation, but their liver structure and function are apparently normal (10). Nevertheless, all mice died shortly after partial hepatectomy if anesthetized with phenobarbital. Animals survive 1-2 days after the operation if ether is the anesthetic. This rather intriguing finding shows that CK8 is essential for hepatocyte structural integrity but its deficiency is harmful only when hepatocytes are preparing for replication. Presumably, the cell shape changes required for replication do not take place in these animals and prevent the cells from dividing.

Two other animal models have been used for studying events of liver regeneration other than its initiation. One of these involved TGF-alpha deficiency. TGF-alpha knockout mice grew normally and had no deficit in liver regeneration (16). At face value, the results suggest that this growth factor plays no role in liver regeneration, despite the strong correlative evidence linking TGF-alpha expression and hepatocyte proliferation. However, TGF-alpha is a member of the epidermal growth factor (EGF) family of growth factors (which include EGF and heparin-binding EGF as hepatocyte mitogens) that signals through the EGF receptor. Thus a definitive answer for the role of TGF-alpha in liver regeneration will not be forthcoming until mouse lines are established in which more than one of the EGF family of ligands is deleted. Another alternative is to produce lines in which the EGF receptor can be conditionally inactivated because EGF knockout mice die during embryonic development or during the neonatal period.

The introduction of a dominant negative gene (superrepressor) to block NFkappa B binding in rats is an excellent example of transient inhibition of gene expression. This type of system can be adapted to permit "knocking out" a gene in a regulatable manner. Iimuro et al. (8) developed methods to inhibit NFkappa B activation after partial hepatectomy by infecting livers of rats with an adenovirus expressing a truncated form of Ikappa B. This gene functions as a superrepressor of the Ikappa B dissociation from NFkappa B, thus preventing migration of the heterodimer to the cell nucleus. Partially hepatectomized rats infected with the adenovirus vector containing the gene had normal levels of DNA replication but developed marked apoptosis after the peak of DNA synthesis. Clearly, blockage of NFkappa B activity had a major apoptotic effect in the regenerating liver. However, it is surprising that the effect takes place after DNA replication, presumably in G2 cells and not at the start of liver regeneration when NFkappa B is transiently activated (5). It is a good assumption that in the absence of NFkappa B activity TNF may cause apoptosis rather than hepatocyte proliferation in the regenerating liver. This notion is supported by data from cell cultures as well as recent experiments with double transgenic mice that are deficient in both the relA (p65) component of NFkappa B as well as TNFR-1. In contrast to relA-deficient mice, which die during days 14-16 of development with massive liver apoptosis, double knockouts survive development and are born with apparently normal livers (Ref. 4 and M. Rosenfeld and N. Fausto, unpublished observations). Thus, in the developing liver, deficiency in TNFR-1 signaling removes an apoptotic signal that is normally blocked by NFkappa B activity. If a similar mechanism exists in the regenerating liver, it is even more puzzling that hepatocytes expressing the Ikappa B superrepressor did not die in the first few hours after partial hepatectomy. At that time, TNF concentrations in blood and liver are high and NFkappa B has maximal binding activity.

It is possible but unlikely that the protective effect of NFkappa B in the regenerating liver depends on a gene that is transcribed only after the first peak of DNA replication has taken place (24 h or later after partial hepatectomy). Another possibility is that mitogenesis is a critical time for NFkappa B protective effects (8). Other issues need also to be considered in explaining these results. The adenovirus used as vector for the Ikappa B gene can cause liver damage. Although control experiments were performed, it would still be possible that the adenovirus itself may have modified the expression of cytokines after partial hepatectomy. Another question that bears directly into the interpretation of these results is whether blockage of NFkappa B activity in the infected livers occurred in both hepatocytes and nonparenchymal cells. One interesting possibility is that inhibition of NFkappa B activation in nonparenchymal cells after partial hepatectomy causes effects that are earlier than and different from those caused by NFkappa B blockage in hepatocytes.


    WHERE ARE WE GOING? MORE QUESTIONS AND NEW MODELS
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INTRODUCTION
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There are some "big" questions about the mechanisms of liver regeneration that can best be approached by using knockout mice. This list of questions is not comprehensive and is biased toward the model for liver regeneration outlined in this article.

The source of TNF to initiate liver regeneration. A reasonable but unproven hypothesis is that it is generated from lipopolysaccharides (LPS; endotoxin). Even if TNF originates from LPS, why is there a surge in blood and liver TNF after partial hepatectomy? LPS is continuously released by intestinal bacteria, but it remains to be established how gut LPS may influence hepatic TNF levels. In mice strains of low LPS sensitivity, liver regeneration is delayed for several hours, suggesting that knockout mice with functional deficiency of LPS receptors would be extremely useful for exploring these questions. A different but related issue is whether both diffusible and membrane-anchored TNF participate in signal transduction at the start of liver regeneration. Knockout mice deficient in TACE (the protease that cleaves anchored TNF) could be a good model for studying this issue. However, the enzyme is not specific for TNF and also cleaves other membrane-bound factors such as TGF-alpha . TACE knockouts are embryonic lethals with pathological changes that cannot be ascribed to lack of cleavage of anchored TNF. Thus other types of strategies are needed to separately analyze the role of anchored and diffusible TNF in liver regeneration.

Multiple TNF signaling pathways. TNF signaling involves activation not only of NFkappa B and STAT3 but also of p38, c-Jun NH2-terminal kinase (JNK), and ERK-1 and ERK-2 (22). Data on NFkappa B and STAT3 are available, but less is known about other TNF-activated pathways in liver regeneration. JNK activity increases after partial hepatectomy, and, in contrast to c-fos (knockout animals deficient in this gene have a normal regenerative response), c-jun is likely to be important for hepatocyte proliferation. However, c-jun knockouts are embryonic lethals exhibiting a complex phenotype that includes hepatoblast apoptosis and tissue destruction as well as hypoplastic hepatic cord formation. A very worthwhile goal would be to develop mice in which c-jun expression is normal but could be shut off in a regulatable manner in the adult liver. The model would be even more valuable if regulated c-jun expression were liver specific.

Cytokine-independent pathways. Cytokines play a prominent role in the initiation of liver regeneration, but there are other pathways that are not cytokine dependent. Not only do these pathways need to be clearly identified, but it is necessary to determine how these multiple pathways interact. Most importantly, it is essential to determine whether growth factors, particularly HGF, act in parallel with cytokines to initiate liver regeneration (12). To start with, there is still disagreement in the literature on whether HGF present in blood after partial hepatectomy is in the active (double chain) form or is an inactive single-chain molecule that does not bind to receptors (13). New strategies need to be devised to study the role of HGF in liver regeneration using mouse knockout models because HGF knockouts are embryonic lethals. Mice in which HGF expression or c-met activity is conditionally regulated in the liver would provide an exciting system for analysis of the role of HGF at the initiation of liver regeneration. They could be used to sort out growth factor-mediated cytokine-independent pathways and to study the mechanisms of HGF activation after partial hepatectomy. Other strategies may include developing lines of mice that carry HGF deletions specific for blockage of mitogenic activity, leaving intact HGF morphogenic and motogenic capacity.

Growth factor redundancy. Many growth factors stimulate DNA synthesis in cultured hepatocytes, and at least two of these, HGF and TGF-alpha , participate in liver regeneration (5, 6). It has not been established whether this multiplicity of factors is a protective mechanism to ensure that regeneration will not fail because of the redundancy of the effects of the various factors or whether HGF and TGF-alpha have complementary activities. In primary cultures of hepatocytes, these two factors act at least partially in a complementary manner, since their combination achieves a greater level of DNA replication than when either HGF or TGF-alpha is exposed as a single agent (20). Despite compelling data associating TGF-alpha expression with hepatocyte proliferation in liver development, regeneration, and carcinogenesis as well as in cultured cells (5, 6), TGF-alpha knockout mice have normal liver regeneration (16). As mentioned, double knockout strategies (deficiency of TGF-alpha and EGF for instance) may provide information on the requirement for EGF family growth factors in liver regeneration. More definitive data to answer this question could be obtained in conditional expressors of EGF receptors, particularly if regulation of EGF receptor activity was restricted to the liver.

Termination of liver regeneration. The cessation of liver regeneration might be dependent on the extinction of positive signals or on an independent system of negative regulators and repressors. Regardless of its mediators, hepatocyte growth after partial hepatectomy is sensitive to metabolic demands imposed on the liver, and there might be signals that sense when liver mass has been restored. The key to explaining the precise regulation of liver growth after partial hepatectomy may depend on the identification of interactions between metabolic and mitogenic pathways. Oxidative stress generated at the start of regeneration (9), which could cause Ikappa B degradation and activation of NFkappa B, might be a signal that links these processes. However, it is quite challenging to design a mouse knockout model that would be useful for the analysis of the termination of liver regeneration. The model needs to take into account that in liver regeneration both positive (i.e., growth factors, cyclin D1, mdm2, and so forth) and negative (p53, TGF-beta , activin) cell cycle effectors are stimulated. Furthermore, even in transgenic mice that overexpress TGF-alpha , a strong hepatocyte mitogen, regeneration is more efficient but it is still perfectly regulated (21). The major challenge is then to disrupt a mechanism that operates under strong selective pressure and is precisely regulated.

Studies using knockout mouse models have provided some of the most important advances in understanding the mechanisms of liver regeneration. Many questions need to be answered, but new and more sophisticated gene knockout strategies will have to be developed to provide definitive answers. The establishment of conditional, liver-specific knockout mouse lines would represent a notable advance. These lines should be constructed with genes that are presumed to have key functions in the multistep process of liver regeneration.


    FOOTNOTES

* Fifth in a series of invited articles on Lessons From Genetically Engineered Animal Models.

Address for reprint requests and other correspondence: N. Fausto, Dept. of Pathology, Box 357470, Univ. of Washington School of Medicine, C-516 Health Sciences Bldg., Seattle WA 98195-7470 (E-mail: nfausto{at}u.washington.edu).


    REFERENCES
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ABSTRACT
INTRODUCTION
LIVER REGENERATION AS A...
KNOCKOUT MOUSE MODELS USED...
WHERE ARE WE GOING?...
REFERENCES

1.   Akerman, P., P. Cote, S. Q. Yang, C. McClain, S. Nelson, G. J. Bagby, and A. M. Diehl. Antibodies to tumor necrosis factor-alpha inhibit liver regeneration after partial hepatectomy. Am. J. Physiol. 263 (Gastrointest. Liver Physiol. 26): G579-G585, 1992[Abstract/Free Full Text].

2.   Albrecht, J. H., and L. K. Hansen. Cyclin D1 promotes mitogen-independent cell cycle progression in hepatocytes. Cell Growth Differ. 10: 397-404, 1999[Abstract/Free Full Text].

3.   Cressman, D. E., L. E. Greenbaum, R. A. DeAngelis, G. Ciliberto, E. E. Furth, V. Poli, and R. Taub. Liver failure and defective hepatocyte regeneration in interleukin-6-deficient mice. Science 274: 1379-1383, 1996[Abstract/Free Full Text].

4.   Doi, T. S., M. W. Marino, T. Takahashi, T. Yoshida, T. Sakakura, L. J. Old, and Y. Obata. Absence of tumor necrosis factor rescues RelA-deficient mice from embryonic lethality. Proc. Natl. Acad. Sci. USA 96: 2994-2999, 1999[Abstract/Free Full Text].

5.   Fausto, N., A. D. Laird, and E. M. Webber. Role of growth factors and cytokines in hepatic regeneration. FASEB J. 9: 1527-1536, 1995[Abstract/Free Full Text].

6.   Fausto, N., and E. M. Webber. Liver regeneration. In: The Liver: Biology and Pathobiology, edited by I. Arias, J. Boyer, N. Fausto, W. Jakoby, D. Schachter, and D. Shafritz. New York: Raven, 1994, p. 1059-1084.

7.   Greenbaum, L. E., W. Li, D. E. Cressman, Y. Peng, G. Ciliberto, V. Poli, and R. Taub. CCAAT enhancer-binding protein beta  is required for normal hepatocyte proliferation in mice after partial hepatectomy. J. Clin. Invest. 102: 996-1007, 1998[Abstract/Free Full Text].

8.   Iimuro, Y., T. Nishiura, C. Hellerbrand, K. E. Behrns, R. Schoonhoven, J. W. Grisham, and D. A. Brenner. NFkappa B prevents apoptosis and liver dysfunction during liver regeneration . J. Clin. Invest. 101: 802-811, 1998[Abstract/Free Full Text]. [Corrigenda. J. Clin. Invest. 101: April 1998, p. 1541.]

9.   Lee, F. Y., Y. Li, H. Zhu, S. Yang, H. Z. Lin, M. Trush, and A. M. Diehl. Tumor necrosis factor increases mitochondrial oxidant production and induces expression of uncoupling protein-2 in the regenerating mouse liver. Hepatology 29: 677-687, 1999[Medline].

10.   Loranger, A., S. Duclos, A. Grenier, J. Price, M. Wilson-Heiner, H. Baribault, and N. Marceau. Simple epithelium keratins are required for maintenance of hepatocyte integrity. Am. J. Pathol. 151: 1673-1683, 1997[Abstract].

11.   Mars, W. M., R. Zarnegar, and G. K. Michalopoulos. Activation of hepatocyte growth factor by the plasminogen activators uPA and tPA. Am. J. Pathol. 143: 949-958, 1993[Abstract].

12.   Michalopoulos, G. K., and M. C. DeFrances. Liver regeneration. Science 276: 60-66, 1997[Abstract/Free Full Text].

13.   Miyazawa, K., T. Shimomura, and N. Kitamura. Activation of hepatocyte growth factor in the injured tissues is mediated by hepatocyte growth factor activator. J. Biol. Chem. 271: 3615-3618, 1996[Abstract/Free Full Text].

14.   Rai, R. M., F. Y. J. Lee, A. Rosen, S. Q. Yang, H. Z. Lin, A. Koteish, F. Y. Liew, C. Zaragoza, C. Lowenstein, and A. M. Diehl. Impaired liver regeneration in inducible nitric oxide synthase-deficient mice. Proc. Natl. Acad. Sci. USA 95: 13829-13834, 1998[Abstract/Free Full Text].

15.   Roselli, H. T., M. Su, K. Washington, D. M. Kerins, D. E. Vaughan, and W. E. Russell. Liver regeneration is transiently impaired in urokinase-deficient mice. Am. J. Physiol. 275 (Gastrointest. Liver Physiol. 38): G1472-G1479, 1998[Abstract/Free Full Text].

16.   Russell, W. E., W. K. Kaufman, S. Sitaric, N. C. Luetteke, and D. C. Lee. Liver regeneration and hepatocarcinogenesis in transforming growth factor-alpha -targeted mice. Mol. Carcinog. 15: 183-189, 1996[Medline].

17.   Servillo, G., M. A. Della Fazia, and P. Sassone-Corsi. Transcription factor CREM coordinates the timing of hepatocyte proliferation in the regenerating liver. Genes Dev. 12: 3639-3643, 1998[Abstract/Free Full Text].

18.   Taub, R. Expression and function of growth-induced genes during liver regeneration. In: Liver Regeneration and Carcinogenesis: Molecular and Cellular Mechanisms, edited by R. L. Jirtle. San Diego, CA: Academic, 1995, p. 71-97.

19.   Webber, E. M., J. Bruix, R. H. Pierce, and N. Fausto. Tumor necrosis factor primes hepatocytes for DNA replication in the rat. Hepatology 28: 1226-1234, 1998[Medline].

20.   Webber, E. M., M. J. FitzGerald, P. I. Brown, M. H. Bartlett, and N. Fausto. TGFalpha expression during liver regeneration after partial hepatectomy and toxic injury, and potential interactions between TGFalpha and HGF. Hepatology 18: 1422-1431, 1993[Medline].

21.   Webber, E. M., J. C. Wu, L. Wang, G. Merlino, and N. Fausto. Overexpression of transforming growth factor-alpha causes liver enlargement and increased hepatocyte proliferation in transgenic mice. Am. J. Pathol. 145: 398-408, 1994[Abstract].

22.   Westwick, J. K., C. Weitzel, A. Minden, M. Karin, and D. A. Brenner. Tumor necrosis factor alpha  stimulates AP-1 activity through prolonged activation of the c-Jun kinase. J. Biol. Chem. 269: 26396-26401, 1994[Abstract/Free Full Text].

23.   Yamada, Y., and N. Fausto. Deficient liver regeneration after carbon tetrachloride injury in mice lacking type 1 but not type 2 tumor necrosis factor receptor. Am. J. Pathol. 152: 1577-1589, 1998[Abstract].

24.   Yamada, Y., I. Kirillova, J. J. Peschon, and N. Fausto. Initiation of liver growth by tumor necrosis factor: deficient liver regeneration in mice lacking type I tumor necrosis factor receptor. Proc. Natl. Acad. Sci. USA 94: 1441-1446, 1997[Abstract/Free Full Text].

25.   Yamada, Y., E. M. Webber, I. Kirillova, J. J. Peschon, and N. Fausto. Analysis of liver regeneration in mice lacking type 1 or type 2 tumor necrosis factor receptor: requirement for type 1 but not type 2 receptor. Hepatology 28: 959-970, 1998[Medline].


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