miR-30e* is an independent subtype-specific prognostic marker in breast cancer.

BACKGROUND: Breast cancer clinical outcome is affected by tumor molecular features, and the identification of subtype-specific prognostic biomarkers is relevant for breast cancer translational research. Gene expression signatures proved to be able to complement prognostic information provided by classical clinico-pathological features. Recently, microRNAs (miRNAs) have been causally linked to tumorigenesis and cancer progression and have been associated with patient outcome, also in breast cancer. METHODS: MicroRNAs associated with the development of distant metastasis were identified in a cohort of 92 ESR1+/ERBB2- lymph node-negative breast cancers from patients not receiving adjuvant treatment. Results were confirmed and further investigated in a total of 1246 miRNA and gene expression profiles of the Molecular Taxonomy of Breast Cancer International Consortium data set. Moderated t-test, univariable and multivariable Cox regression models were used for statistical analyses. RESULTS: miR-30e* was identified as independent protective prognostic factor in lymph node-negative untreated patients with ESR1+/ERBB2- tumours and retained a significant association with a good prognosis in treated patients with the same tumor subtype as well as in the ERBB2+ subtype, but not in ESR1-/ERBB2- tumours. CONCLUSIONS: We highlighted a relevant and subtype-specific role in breast cancer for miR-30e* and demonstrated that adding miRNA markers to gene signatures and clinico-pathological features can help for a better prognostication.

DNA-PKcs: a T-cell tumour suppressor encoded at the mouse scid locus.

Severe combined immunodeficiency (SCID) mice are defective in their ability to rearrange their variable (V), diversity (D) and joining (J) genetic elements to generate functional immunoglobulin (Ig) and T-cell receptor (TCR) molecules; as a result, they lack mature B and T cells. These mice are highly sensitive to ionizing radiation, suggesting that the product of the scid gene plays a critical role in both V(D)J recombination and DNA double-strand break repair. Recent studies suggest that the SCID defect lies in the gene encoding the catalytic subunit of DNA-dependent protein kinase (DNA-PK; refs 6-8), a nuclear protein made up of the Ku 70 and Ku 86 subunits as well as the large catalytic subunit, DNA-PKcs. Other reports have implied that the SCID phenotype correlates with nonsense mutations at the extreme 3' end of Prkdc, the DNA-PKcs gene. The identity of the gene remains in doubt, however, because the consequences of genetic inactivation of Prkdc have not been determined. This study shows that complete inactivation of Prkdc in a novel insertional mouse mutant recapitulates the SCID phenotype and that Prkdc and scid are alleic. Significantly, DNA-PKcs null mice demonstrate complete penetrance of thymic lymphoblastic lymphomas, strongly suggesting that Prkdc functions in mice as a T-cell tumour suppressor and, by virtue of its association with DNA repair and recombination, belongs to the 'caretaker' class of tumour-suppressor genes that includes ATM, BRCA1 and BRCA2 (ref. 15).

Insomnia and daytime sleepiness predict 20-year mortality in older male adults: data from a population-based study.

Data regarding the possible relationship of insomnia and EDS with mortality are inconclusive. The aim of this study was to investigate the association between these sleep complaints and the risk of long-term (20 years) all-cause mortality in older adults. Between April 2000 and March 2001, 750 subjects aged 65 years and older, who resided in the seventh district of Udine, were recruited. Data on sociodemographic characteristics, past medical history, and pharmacological treatment were collected. Dementia was diagnosed using a comprehensive neurological and neuroradiological assessment. Older adults were interviewed by neuropsychologists trained in sleep disturbances in order to assess the presence of sleep complaints. Vital status was followed over 20 years until March 2020. Older male adults affected by insomnia and EDS were significantly more likely to die over the follow-up period. Indeed, males reporting poor sleep and daytime somnolence had a 60% and 48% higher chance of dying than subjects who were not affected by these sleep complaints, respectively. The HR was attenuated after adjusting for confounding variables among insomniacs, whereas that of somnolent men strengthened. Differently from men, insomnia and EDS did not have any impact on mortality in older women. In conclusion, older male adults affected by insomnia and EDS had a significant increased risk of mortality, which is independent of cancer, depression, dementia, cardiovascular diseases, and sleeping pill use.

Promoter region of the human Harvey ras proto-oncogene: similarity to the EGF receptor proto-oncogene promoter.

Regulation of transcription of members of the ras gene family undoubtably plays an important role in controlling cellular growth. Examination of this level of regulation requires identification of the promoter regions of the ras proto-oncogenes. Four major transcriptional start sites were detected in the human Harvey ras 1 proto-oncogene. The promoter region contains neither a TATA box nor a CAAT box in their characteristic upstream positions, has an extremely high G+C content (80 percent), and contains multiple GC boxes including seven CCGCCC repeats and three repeats of the inverted complement, GGGCGG. This region has strong promoter activity when placed upstream from the chloramphenicol acetyl transferase gene and transfected into monkey CV1 cells. In these ways the Harvey ras 1 proto-oncogene promoter resembles the promoter of the gene encoding the epidermal growth factor (EGF) receptor. The similarity between the two proto-oncogene promoters may be relevant to the mechanism by which the expression of such "growth control" genes is regulated.

Binding of the Sp1 transcription factor by the human Harvey ras1 proto-oncogene promoter.

Members of the ras gene family encode proteins that when overproduced or mutated can transform immortalized mammalian cells. It is therefore important to understand the mechanisms by which the ras genes are regulated. The promoter region of the human Harvey ras proto-oncogene c-Ha-ras1 initiates RNA transcription at multiple sites and contains repeated copies of the hexanucleotide GGGCGG and its inverted complement CCGCCC, referred to as GC boxes. These GC boxes consist of sequences identical to those found in the SV40 early promoter, where the human cellular transcriptional factor Sp1 binds. Footprinting analysis with deoxyribonuclease I was used to show that Sp1 binds to six GC box sequences within the c-Ha-ras1 promoter. An in vivo transfection assay showed competition between the 21-base pair repeats of the SV40 promoter and the c-Ha-ras1 promoter for common regulatory factors. In this system the presence of Sp1 is apparently required for c-Ha-ras1 transcription. Analysis of deletions of the c-Ha-ras1 promoter region by means of a transient expression assay revealed that the three Sp1 binding sites closest to the RNA start sites were sufficient for full transcriptional activity.

Amplification and enhanced expression of the epidermal growth factor receptor gene in A431 human carcinoma cells.

The sequence of the human epidermal growth factor (EGF) receptor shows great homology with the avian erythroblastosis virus v-erb B oncogene, raising the possibility that the receptor gene is identical to the c-erb B protooncogene. Human A431 epidermoid carcinoma cells, which have an unusually high number of EGF receptors, were examined to determine whether elevated EGF receptor levels correlate with gene amplification. Southern blots of genomic DNA's from A431 and other human cell lines were probed with either a v-erb B gene fragment or a human EGF receptor complementary DNA clone (pE7), previously isolated from an A431 complementary DNA library. When either probe was used to analyze Eco RI- or Hind III-generated DNA fragments, EGF receptor DNA sequences were amplified about 30-fold in A431. Differences in the banding pattern of A431 DNA fragments relative to normal fibroblast DNA indicate the occurrence of a rearrangement in the region of the receptor gene. Furthermore, A431 cells contain a characteristic, prominent 2.9-kilobase RNA. These results are consistent with the hypothesis that, in A431 cells, gene amplification, possibly associated with a translocation event, may result in the overproduction of EGF receptor protein or the appearance of the transformed phenotype (or both).

Epidermal-growth-factor-dependent transformation by a human EGF receptor proto-oncogene.

The epidermal growth factor (EGF) receptor gene EGFR has been placed in a retrovirus vector to examine the growth properties of cells that experimentally overproduce a full-length EGF receptor. NIH 3T3 cells transfected with the viral DNA or infected with the corresponding rescued retrovirus developed a fully transformed phenotype in vitro that required both functional EGFR expression and the presence of EGF in the growth medium. Cells expressing 4 x 10(5) EGF receptors formed tumors in nude mice, while control cells did not. Therefore, the EGFR retrovirus, which had a titer on NIH 3T3 cells that was greater than 10(7) focus-forming units per milliliter, can efficiently transfer and express this gene, and increased numbers of EGF receptors can contribute to the transformed phenotype.

Prevalence of 'poor sleep' among patients with multiple sclerosis: an independent predictor of mental and physical status.

BACKGROUND: Patients with multiple sclerosis (MS) report sleep disturbances more frequently than the general population. Besides specific sleep disturbances, many other conditions could impair nocturnal rest in this population. In addition, information regarding the role of disrupted sleep on quality of life (QoL) in MS patients is lacking. This study was performed to bridge this gap. METHODS: A total of 120 patients with MS were enrolled into the study. Demographic, socioeconomic and clinical characteristics (clinical course and duration of MS, EDSS score, therapeutic information, presence of pain, presence of sexual and/or bladder dysfunction, localization of demyelinating plaques, and presence of anxiety and depression) were collected. The Pittsburgh Sleep Quality Index (PSQI), the Charlson Comorbidity Index (CCI) and the Italian version of the 36-item Short Form (SF-36) were used to assess quality of sleep, comorbidity and QoL, respectively. RESULTS: Nearly half (47.5%) of MS patients were classified as "poor sleepers," having significantly higher EDSS (3.1+/-1.4 vs. 2.3+/-1.4, p=0.009) and CCI scores (0.19+/-0.4 vs. 0.03+/-0.2, p=0.009) than "good sleepers." In addition, pain due to MS was more common among "poor sleepers" (33.3% vs. 17.7%, p=0.05). Scores for each domain of the SF-36, and the mental component summary (MCS) and physical component summary (PCS) scores were significantly lower in poor sleepers than in good sleepers (p<0.001 for each score). Of the different variables associated with MCS, the only independent predictors of mental status were: presence of sexual and/or bladder dysfunction and global PSQI score. The independent predictors for physical status (PCS) were age, EDSS score and global PSQI score. CONCLUSIONS: Poor sleep is common in patients with MS, representing an independent predictor of QoL. Patients with MS who are poor sleepers should receive immediate assessment and treatment, bearing in mind that, in addition to specific sleep disturbances, other clinical conditions (both related and unrelated to MS) can disrupt nocturnal sleep.

Hypertrophic gastropathy resembling Ménétrier's disease in transgenic mice overexpressing transforming growth factor alpha in the stomach.

Transforming growth factor alpha (TGF alpha) is thought to participate in the normal and pathologic processes of numerous tissues, including the gastric mucosa. To explore its role in vivo, transgenic mice were generated overexpressing TGF alpha in the stomach. TGF alpha induced dramatic structural and functional lesions of the glandular stomach that were similar to Ménétrier's disease in humans. Transgenic mice developed severe adenomatous hyperplasia that resulted in a striking nodular thickening or hypertrophy of the gastric mucosa. Secretions obtained from affected stomachs contained no detectable gastric acid, suggesting that parietal cell function had been greatly impaired. These findings demonstrate that overproduction of TGF alpha can stimulate cellular proliferation, suppress acid secretion, and perturb organogenesis of the stomach of transgenic mice. Moreover, TGF alpha may contribute to the pathogenesis of related human hypertrophic gastropathies, such as Ménétrier's disease.

Elevated epidermal growth factor receptor gene copy number and expression in a squamous carcinoma cell line.

The human epidermal growth factor (EGF) receptor is known to be homologous to the v-erb B oncogene protein of the avian erythroblastosis virus. Overexpression of the EGF receptor gene in A431 epidermoid carcinoma cells is due to gene amplification. In this study, a variety of squamous cell carcinomas were examined and one, SCC-15, contained high levels of the EGF receptor as determined by immunoprecipitation via an EGF receptor-specific polyclonal antibody. Using a cloned EGF receptor complementary DNA as a probe, the level of EGF receptor RNA was found to be elevated four-fold in SCC-15 relative to normal cultured keratinocytes. When the same probe was used to identify EGF receptor gene fragments on a genomic DNA blot, the SCC-15 cell line was shown to possess an EGF receptor gene copy number amplified four to five times. Gene amplification results in the enhancement in the level of the EGF receptor in several carcinomas and could be responsible for the appearance of the transformed phenotype in these cells.

Pancreatic gastrin stimulates islet differentiation of transforming growth factor alpha-induced ductular precursor cells.

Gastrin is transiently expressed in fetal islets during a critical period of their development from protodifferentiated islet precursors in fetal pancreatic ducts. To examine the possible role of gastrin as an islet cell growth factor, postnatal islet growth was studied in transgenic mice which overexpress gastrin and TGF alpha in their pancreas. Overexpression of a TGF alpha transgene causes metaplastic ductules containing numerous insulin expressing cells that resemble protodifferentiated precursors of the fetal pancreas. However, islet mass of the TGF alpha transgenic mice was not increased. Pancreatic overexpression of gastrin from a chimeric insulin/gastrin transgene transcribed from the insulin promoter markedly decreased the TGF alpha-stimulated increase in pancreatic duct mass. Furthermore, pancreatic coexpression of both gastrin and TGF alpha significantly increased islet mass in mice expressing both transgenes. These findings indicate that TGF alpha and gastrin can act synergistically to stimulate islet growth, although neither peptide alone is sufficient. Islet growth may possibly be stimulated through gastrin promoting the differentiation of insulin-positive cells in the TGF alpha-induced metaplastic ducts. This transgenic study suggests that islet neogenesis can be reactivated in the ductular epithelium of the adult pancreas by local expression of two growth factors, gastrin and TGF alpha.

Exocrine pancreatic disorders in transsgenic mice expressing human keratin 8.

Keratins K8 and K18 are the major components of the intermediate-filament cytoskeleton of simple epithelia. Increased levels of these keratins have been correlated with various tumor cell characteristics, including progression to malignancy, invasive behavior, and drug sensitivity, although a role for K8/K18 in tumorigenesis has not yet been demonstrated. To examine the function of these keratins, we generated mice expressing the human K8 (hk8) gene, which leads to a moderate keratin-content increase in their simple epithelia. These mice displayed progressive exocrine pancreas alterations, including dysplasia and loss of acinar architecture, redifferentiation of acinar to ductal cells, inflammation, fibrosis, and substitution of exocrine by adipose tissue, as well as increased cell proliferation and apoptosis. Histological changes were not observed in other simple epithelia, such as the liver. Electron microscopy showed that transgenic acinar cells have keratins organized in abundant filament bundles dispersed throughout the cytoplasm, in contrast to control acinar cells, which have scarce and apically concentrated filaments. The phenotype found was very similar to that reported for transgenic mice expressing a dominant-negative mutant TGF-beta type II receptor (TGFbetaRII mice). We show that these TGFbetaRII mutant mice also have elevated K8/K18 levels. These results indicate that simple epithelial keratins play a relevant role in the regulation of exocrine pancreas homeostasis and support the idea that disruption of mechanisms that normally regulate keratin expression in vivo could be related to inflammatory and neoplastic pancreatic disorders.

Modulation of epidermal growth factor receptor proto-oncogene transcription by a promoter site sensitive to S1 nuclease.

The epidermal growth factor (EGF) receptor is the functional target of the mitogen EGF and the cellular homolog of the avian erythroblastosis virus erbB oncogene product. Regulation of expression of the proto-oncogene encoding the EGF receptor can be elucidated by studying the structure and function of the gene promoter outside the confines of the cell. Previously, we reported the isolation of the human EGF receptor gene promoter. The promoter is highly GC rich, contains no TATA or CAAT box, and has multiple transcription start sites. An S1 nuclease-sensitive site has now been found 80 to 110 base pairs (bp) upstream from the major in vivo transcription initiation site. Two sets of direct repeat sequences were found in this area; both conform to the motif TCCTCCTCC. When deletion mutations were made in this region of the promoter by using either Bal 31 exonuclease or S1 nuclease, we found that in vivo activity dropped three- to fivefold, on the basis of transient-transfection analysis. Examination of nuclear protein binding to normal and mutated promoter DNAs by gel retardation analysis and DNase I footprinting revealed that two specific factors bind to the direct repeat region but cannot bind to the S1 nuclease-mutated promoter. One of the specific factors is the transcription factor Sp1. The results suggest that these nuclear trans-acting factors interact with the S1 nuclease-sensitive region of the EGF receptor gene promoter and either directly or indirectly stimulate transcription.

NK1, a natural splice variant of hepatocyte growth factor/scatter factor, is a partial agonist in vivo.

Hepatocyte growth factor/scatter factor (HGF/SF) is a potent mitogen, motogen, and morphogen for epithelial cells expressing its tyrosine kinase receptor, the c-met proto-oncogene product, and is required for normal development in the mouse. Inappropriate stimulation of Met signal transduction induces aberrant morphogenesis and oncogenesis in mice and has been implicated in human cancer. NK1 is a naturally occurring HGF/SF splice variant composed of only the amino terminus and first kringle domain. While the biological activities of NK1 have been controversial, in vitro data suggest that it may have therapeutic value as an HGF/SF antagonist. Here, we directly test this hypothesis in vivo by expressing mouse NK1 in transgenic mice and comparing the consequent effects with those observed for mice carrying an HGF/SF transgene. Despite robust expression, NK1 did not behave as an HGF/SF antagonist in vivo. Instead, NK1-transgenic mice displayed most of the phenotypic characteristics associated with HGF/SF-transgenic mice, including enlarged livers, ectopic skeletal-muscle formation, progressive renal disease, aberrant pigment cell localization, precocious mammary lobuloalveolar development, and the appearance of mammary, hepatocellular, and melanocytic tumors. And like HGF/SF-transgenic livers, NK1 livers had higher levels of tyrosine-phosphorylated complexes associated with Met, suggesting that the mechanistic basis for the effects of NK1 overexpression in vivo was autocrine activation of Met. We conclude that NK1 acts in vivo as a partial agonist. As such, the efficacy of NK1 as a therapeutic HGF/SF antagonist must be seriously questioned.

The p53 response to DNA damage in vivo is independent of DNA-dependent protein kinase.

Ionizing radiation (IR) exposure causes mammalian cells to undergo p53-dependent cell cycle arrest and/or apoptosis. The in vivo role of DNA-dependent protein kinase (DNA-PK) in the transduction of the DNA damage signal to p53 remains unresolved. To determine the relationship between DNA-PK and p53, we studied the cell cycle and apoptotic responses to IR in mice deficient in DNA-PK. Using the slip mouse, which harbors an inactivating mutation of the DNA-PK catalytic subunit (DNA-PKcs), we demonstrated not only that these DNA-PKcs null mutants were highly radiosensitive but also that upon IR treatment, p53 accumulated in their cultured cells and tissue. Induced p53 was transcriptionally active and mediated the induction of p21 and Bax in slip cells. Examination of the thymic cell cycle response to IR treatment indicated that the slip G(1)/S-phase cell cycle checkpoint function was intact. We further show that slip mice exhibited a higher level of spontaneous thymic apoptosis as well as a more robust apoptotic response to IR than wild-type mice. Together, these data demonstrate that the p53-mediated response to DNA damage is intact in cells devoid of DNA-PK activity and suggest that other kinases, such as the product of the gene (ATM) mutated in ataxia telangiectasia, are better candidates for regulating IR-induced phosphorylation and accumulation of p53.

Disassociation of met-mediated biological responses in vivo: the natural hepatocyte growth factor/scatter factor splice variant NK2 antagonizes growth but facilitates metastasis.

Hepatocyte growth factor/scatter factor (HGF/SF) stimulates numerous cellular activities capable of contributing to the metastatic phenotype, including growth, motility, invasiveness, and morphogenetic transformation. When inappropriately expressed in vivo, an HGF/SF transgene induces numerous hyperplastic and neoplastic lesions. NK1 and NK2 are natural splice variants of HGF/SF; all interact with a common receptor, Met. Although both agonistic and antagonistic properties have been ascribed to each isoform in vitro, NK1 retains the full spectrum of HGF/SF-like activities when expressed as a transgene in vivo. Here we report that transgenic mice broadly expressing NK2 exhibit none of the phenotypes characteristic of HGF/SF or NK1 transgenic mice. Instead, when coexpressed in NK2-HGF/SF bitransgenic mice, NK2 antagonizes the pathological consequences of HGF/SF and discourages the subcutaneous growth of transplanted Met-containing melanoma cells. Remarkably, the metastatic efficiency of these same melanoma cells is dramatically enhanced in NK2 transgenic host mice relative to wild-type recipients, rivaling levels achieved in HGF/SF and NK1 transgenic hosts. Considered in conjunction with reports that in vitro NK2 induces scatter, but not other activities, these data strongly suggest that cellular motility is a critical determinant of metastasis. Moreover, our results demonstrate how alternatively structured ligands can be exploited in vivo to functionally dissociate Met-mediated activities and their downstream pathways.

Expression of a human multidrug resistance cDNA (MDR1) in the bone marrow of transgenic mice: resistance to daunomycin-induced leukopenia.

The human multidrug resistance gene (MDR1) encodes a drug efflux pump glycoprotein (P-glycoprotein) responsible for resistance to multiple cytotoxic drugs. A plasmid carrying a human MDR1 cDNA under the control of a chicken beta-actin promoter was used to generate transgenic mice in which the transgene was mainly expressed in bone marrow and spleen. Immunofluorescence localization studies showed that P-glycoprotein was present on bone marrow cells. Furthermore, leukocyte counts of the transgenic mice treated with daunomycin did not fall, indicating that their bone marrow was resistant to the cytotoxic effect of the drug. Since bone marrow suppression is a major limitation to chemotherapy, these transgenic mice should serve as a model to determine whether higher doses of drugs can cure previously unresponsive cancers.

Structure and localization of genes encoding aberrant and normal epidermal growth factor receptor RNAs from A431 human carcinoma cells.

A431 cells have an amplification of the epidermal growth factor (EGF) receptor gene, the cellular homolog of the v-erb B oncogene, and overproduce an aberrant 2.9-kilobase RNA that encodes a portion of the EGF receptor. A cDNA (pE15) for the aberrant RNA was cloned, sequenced, and used to analyze genomic DNA blots from A431 and normal cells. These data indicate that the aberrant RNA is created by a gene rearrangement within chromosome 7, resulting in a fusion of the 5' portion of the EGF receptor gene to an unidentified region of genomic DNA. The unidentified sequences are amplified to about the same degree (20- to 30-fold) as the EGF receptor sequences. In situ hybridization to chromosomes from normal cells and A431 cells show that both the EGF receptor gene and the unidentified DNA are localized to the p14-p12 region of chromosome 7. By using cDNA fragments to probe DNA blots from mouse-A431 somatic cell hybrids, the rearranged receptor gene was shown to be associated with translocation chromosome M4.

Restless legs syndrome: diagnosis, epidemiology, classification and consequences.

Restless legs syndrome (RLS) is a sensorimotor disorder characterised by a complaint of an almost irresistible urge to move the legs. RLS is diagnosed clinically by means of the four essential criteria of the International Restless Legs Syndrome Study Group. In doubtful cases, neurophysiological examinations, such as polysomnography and/or a suggested immobilisation test, can be performed to confirm a clinical suspicion of RLS. Several other conditions may present sensorimotor complaints with features similar to RLS; a careful sleep history is required to avoid a misdiagnosis. Three different scales have been validated to assess the severity of RLS. In the general population, RLS prevalence ranges from 0.1% to 11.5%, with a high number of patients affected by a primary form of the sleep disturbance (70%-80%). However, several clinical conditions have been associated with RLS, such as iron deficiency, uraemia, pregnancy and polyneuropathy. Furthermore, recent studies show that RLS may be associated also to type 2 diabetes mellitus and to multiple sclerosis. RLS has a negative impact on sleep, cognitive functions, quality of life and mental status. Higher awareness of RLS among physicians is required; it remains an underdiagnosed clinical condition.

Development of liver tumors in transforming growth factor alpha transgenic mice.

We studied the development of liver tumors in male transforming growth factor alpha (TGF-alpha) transgenic mice of the CD1 strain and examined the expression of the transgene by immunohistochemistry and in situ hybridization. Livers of 4-5-week-old transgenic mice contained areas of centribobular hypertrophy with low glucose-6-phosphatase activity. These areas progressively expanded, and hypertrophy and dysplasia became generalized in livers of mice at 10-12 months of age. The expression of the transgene, determined by either immunohistochemistry or in situ hybridization, was uneven in animals that were 10 weeks old or older. The positive hepatocytes formed patches with a predominant centrilobular distribution. We studied a total of 23 liver tumors (7 hepatocellular carcinomas and 16 adenomas) obtained from 11 mice at 13-15 months of age and from one 7-month-old animal which received zinc sulfate to induce the transgene. The carcinomas were well differentiated tumors, without glucose-6-phosphatase or gamma-glutamyltranspeptidase activity, that developed from the dysplastic parenchyma and occasionally within an adenoma. In all carcinomas and in 56% of the adenomas there was overexpression of the transgene in relationship to the surrounding tissue. The majority of the tumors that overexpressed TGF-alpha were alpha-fetoprotein positive, while alpha-fetoprotein staining was not detected in tumors (all adenomas) that did not show excessive transgene expression. We conclude that TGF-alpha functions as a promoter of liver carcinogenesis through its effect as an autocrine inducer of hepatocyte proliferation. Further, the data indicate that TGF-alpha overexpression may favor tumor progression.