Rat Epidermal Growth Factor Research Articles

Delayed recanalization reduced neuronal apoptosis and neurological deficits by enhancing liver-derived trefoil factor 3-mediated neuroprotection via LINGO2/EGFR/Src signaling pathway after middle cerebral artery occlusion in rats

Delayed recanalization at days or weeks beyond the therapeutic window was shown to improve functional outcomes in acute ischemic stroke (AIS) patients. However, the underlying mechanisms remain unclear. Previous preclinical study reported that trefoil factor 3 (TFF3) was secreted by liver after cerebral ischemia and acted a distant neuroprotective factor. Here, we investigated the liver-derived TFF3-mediated neuroprotective mechanism enhanced by delayed recanalization after AIS. A total of 327 male Sprague-Dawley rats and the model of middle cerebral artery occlusion (MCAO) with permanent occlusion (pMCAO) or with delayed recanalization at 3 d post-occlusion (rMCAO) were used. Partial hepatectomy was performed within 5 min after MCAO. Leucine-rich repeat and immunoglobulin-like domain-containing nogo receptor-interacting protein 2 (LINGO2) siRNA was administered intracerebroventricularly at 48 h after MCAO. Recombinant rat TFF3 (rr-TFF3, 30 μg/Kg) or recombinant rat epidermal growth factor (rr-EGF, 100 μg/Kg) was administered intranasally at 1 h after recanalization, and EGFR inhibitor Gefitinib (75 mg/Kg) was administered intranasally at 30 min before recanalization. The evaluation of outcomes included neurobehavior, ELISA, western blot and immunofluorescence staining. TFF3 in hepatocytes and serum were upregulated in a similar time-dependent manner after MCAO. Compared to pMCAO, delayed recanalization increased brain TFF3 levels and attenuated brain damage with the reduction in neuronal apoptosis, infarct volume and neurological deficits. Partial hepatectomy reduced TFF3 levels in serum and ipsilateral brain hemisphere, and abolished the benefits of delayed recanalization on neuronal apoptosis and neurobehavioral deficits in rMCAO rats. Intranasal rrTFF3 treatment reversed the changes associated with partial hepatectomy. Delayed recanalization after MCAO increased the co-immunoprecipitation of TFF3 and LINGO2, as well as expressions of p-EGFR, p-Src and Bcl-2 in the brain. LINGO2 siRNA knockdown or EGFR inhibitor reversed the effects of delayed recanalization on apoptosis and brain expressions of LINGO2, p-EGFR, p-Src and Bcl-2 in rMCAO rats. EGFR activator abolished the deleterious effects of LINGO2 siRNA. In conclusion, our investigation demonstrated for the first time that delayed recanalization may enhance the entry of liver-derived TFF3 into ischemic brain upon restoring blood flow after MCAO, which attenuated neuronal apoptosis and neurological deficits at least in part via activating LINGO2/EGFR/Src pathway.

Dual-energy spectral CT characteristics in surgically resected lung adenocarcinoma: comparison between Kirsten rat sarcoma viral oncogene mutations and epidermal growth factor receptor mutations

BackgroundKirsten rat sarcoma viral oncogene homolog (KRAS) and epidermal growth factor receptor (EGFR) are the two most frequent and well-known oncogene of lung adenocarcinoma. The purpose of this study is to compare the characteristics measured with dual-energy spectral computed tomography (DESCT) in lung adenocarcinoma patients who have KRAS and EGFR gene mutations.MethodsPatients with surgically resected lung adenocarcinoma (n = 72) were enrolled, including 12 patients with KRAS mutations and 60 patients with EGFR mutations. DESCT quantitative parameters, including the CT number at 70 keV, the slopes of the spectral attenuation curves (slope λ HU), normalized iodine concentration (NIC), normalized water concentration (NWC), and effective atomic number (effective Z), were analyzed. A multiple logistic regression model was applied to discriminate clinical and DESCT characteristics between the types of mutations.ResultsThe KRAS mutation was more common in people who smoked than the EGFR mutation. Nodule type differed significantly between the KRAS and EGFR groups (P = 0.035), and all KRAS mutation adenocarcinomas were solid nodules. Most DESCT quantitative parameters differed significantly between solid nodules and subsolid nodules. CT number at 70 keV, slope λ HU, NIC, and effective Z differed significantly between the KRAS and EGFR groups (P = 0.006, 0.017, 0.013 and 0.010) with solid lung adenocarcinoma. Multivariate logistic analysis of DESCT and clinical features indicated that besides smoking history, the CT value at 70 keV (OR = 0.938, P = 0.009) was significant independent factor that could be used to differentiate KRAS and EGFR mutations in solid lung adenocarcinoma.ConclusionsDESCT would be a potential tool to differentiate lung adenocarcinoma patients with a KRAS mutation from those with an EGFR mutation.

Milk-borne epidermal growth factor modulates bilirubin levels in neonatal rats

Summary Objectives Breast milk jaundice (BMJ) is commonly of undetermined etiology. Recently, it has been shown that the degree of BMJ was associated with the increased levels of milk-borne EGF (Epidermal Growth Factor). The aim of the present study was to test the effects of enteral administration of EGF on the development of hyperbilirubinemia in neonatal rats. Methods Pups were divided into three groups: Group 1; artificially fed with formula, Group 2; artificially fed with formula supplemented with 500 ng/ml rat EGF, Group 3; fed by their mothers. Hyperbilirubinemia was induced by an intraperitoneal injection of unconjugated bilirubin suspension once daily for 5 consecutive days. Bilirubin levels, length of ideal villi, and EGF-R expression in the intestine was determined at postnatal day 6. Results In the groups of 2 and 3, serum bilirubin levels were significantly higher when compared to animals in the group 1. There was no significant difference in terms of serum bilirubin levels between the Group 2 and Group 3. Length of ileal villi was significantly longer in the Group 2 and Group 3 than the Group 1 animals. The pattern of EGF-R expression in the Group 2 was similar to that seen in Group 3 animals. In contrast, animals given formula without EGF had markedly decreased expression of the EGF-R. Conclusion Results suggest that EGF supplementation in newborn rats leads to a significant increase in intestinal mucosal proliferation and a significant decrease in bilirubin elimination. These data suggest that EGF possibly increases intestinal bilirubin absorption and may have a role in development of breast milk jaundice. Further studies are needed to confirm this hypothesis.

DNA methylation dynamics in the rat EGF gene promoter after partial hepatectomy.

Epidermal growth factor (EGF), a multifunctional growth factor, is a regulator in a wide variety of physiological processes. EGF plays an important role in the regulation of liver regeneration. This study was aimed at investigating the methylation level of EGF gene throughout liver regeneration. DNA of liver tissue from control rats and partial hepatectomy (PH) rats at 10 time points was extracted and a 354 bp fragment including 10 CpG sites from the transcription start was amplified after DNA was modified by sodium bisulfate. The result of sequencing suggested that methylation ratio of four CpG sites was found to be significantly changed when PH group was compared to control group, in particular two of them were extremely striking. mRNA expression of EGF was down-regulated in total during liver regeneration. We think that the rat EGF promoter region is regulated by variation in DNA methylation during liver regeneration.

Induction of specific human cytotoxic T cells using dendritic cells transduced with an adenovector encoding rat epidermal growth factor receptor 2

This study demonstrates the ability to generate antigen-specific cytotoxic T cells (CTLs) against HER2 using a xenoantigenic immune stimulation strategy. Dendritic cells (DCs) were transduced with an adenovirus vector incorporating full-length cDNA for rat (xenoantigen) epidermal growth factor receptor 2 (Adv-HER2). Stimulation of autologous T cells with Adv-HER2 infected DCs led to enhanced HER2-specific reactivity as assessed by quantitative real-time polymerase chain reaction (qRT-PCR) for T cell IFN-γ mRNA. In ELISPOT and intracellular cytokine staining (ICS) assays, CD8+ CTLs induced by Adv-HER2 transduced DCs released IFN-γ following stimulation with irradiated autologous DCs infected with Adv-HER2 or loaded with a human prostate cancer cell line (LNCaP) lysate. DCs pulsed with HER2 peptides were less stimulatory than Adv-HER2 transduced DCs. HER2 DC induced CTL lysed HER2+ HLA-A2+ tumor cells (MCF-7); significantly reduced lysis occurred in HER2+ HLA-A2- tumor cells (SKOV-3), and the NK cell sensitive cell line K-562.

The role of epidermal growth factor in the healing tympanic membrane following perforation in rats

Epidermal Growth Factor (EGF) has been identified as playing a critical role in the wound healing process. The objective of this study is to investigate the role that EGF plays in rat tympanic membrane (TM) wound healing using two techniques, microarray and immunohistochemistry. The tympanic membranes of rats were perforated using a sterile needle and sacrificed at time points during 2weeks following perforation. The normalized signal intensities at the time points for EGF and associated genes are presented. The rat EGF mRNA did not change significantly between time points. Five associated proteins, including heparin-binding EGF-like growth factor were found to be differentially expressed above a two fold threshold at 12 h following perforation. EGF staining was found at low levels in the uninjured TM. Levels of EGF staining increased at 24 h in the basal keratinocyte layer, became diffusely elevated in the specimen at 36 h, before a second peak in staining of the keratinocyte layer at Day 4. The staining of EGF corresponds to its multiple roles in TM wound healing.

Evaluation of folate‐PAMAM for the delivery of antisense oligonucleotides to rat C6 glioma cells in vitro and in vivo

In the current study, we evaluated the efficiency of folate-polyamidoamine dendrimers conjugates (FA-PAMAM) for the in situ delivery of therapeutic antisense oligonucleotides (ASODN) that could inhibit the growth of C6 glioma cells. Folic acid was coupled to the surface amino groups of G5-PAMAM dendrimer (G5D) through a 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide bond, and ASODNs corresponding to rat epidermal growth factor receptor (EGFR) were then complexed with FA-PAMAM. At an ASODN to PAMAM ratio of 16:1, agarose electrophoresis indicated that antisense oligonucleotides were completely complexed with PAMAM or FA-PAMAM. The ASODN transfection rates mediated by FA-PAMAM and PAMAM were superior to oligofectamine, resulting in greater suppression of EGFR expression and glioma cell growth. Stereotactic injection of EGFR ASODN:FA-PAMAM complexes into established rat C6 intracranial gliomas resulted in greater suppression of tumor growth and longer survival time of tumor-bearing rats compared with PAMAM and oligofectamine-mediated EGFR-ASODN therapy. The current study demonstrates the suitability of folate-PAMAM dendrimer conjugates for efficient EGFR ASODN delivery into glioma cells, wherein they release the ASODN from the FA-PAMAM to knock down EGFR expression in C6 glioma cells, both in vitro and in vivo. FA-PAMAM may thus represent a novel delivery system for short oligonucleotides in glioma-targeted therapy.

Epidermal Growth Factor Reduces Hepatic Sequelae in Experimental Necrotizing Enterocolitis

Background and Aim: Neonatal necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of premature infants. We recently demonstrated that the gut/liver axis plays an important role in the pathophysiology of NEC through the release of inflammatory mediators into the intestinal lumen. We have also shown that supplementation of formula with epidermal growth factor (EGF) dramatically decreases ileal pathology associated with experimental NEC. In this study, we examined the effects of EGF on the liver portion of the gut/liver axis in the neonatal rat model of NEC. Methods: Newborn rats were divided into three experimental groups, NEC, hand-fed with growth-factor free formula; NEC + EGF, hand-fed with formula supplemented with 500 ng/ml rat EGF; or DF, dam fed. All animals were exposed to asphyxia and cold stress twice daily for 4 days to develop NEC. Results: EGF receptor expression was significantly (p ≤ 0.01) decreased in the NEC + EGF group compared to the NEC group. EGF supplementation significantly decreased Kupffer cell numbers (p ≤ 0.01) as well as hepatic tumor necrosis factor (TNF)-α and interleukin-18 production (p ≤ 0.05). Further, TNF-α in the intestinal luminal contents of the NEC + EGF group were normalized to levels observed in DF controls compared to the NEC group (p ≤ 0.05). Activated nuclear factor-ĸB was also substantially decreased in the NEC + EGF group versus the NEC group. Conclusion: The results of this study indicate that EGF normalizes cytokine overproduction in the liver of neonatal rats with NEC, which contributes to diminished intestinal damage during the development of experimental NEC. These data suggest that supplementation of formula with EGF can have beneficial effects on the gut/liver axis during NEC pathogenesis.

Growth factors and their receptors in the middle ear mucosa during otitis media.

The hyperplastic response of the middle ear mucosa during bacterial otitis media is thought to be mediated by the actions of growth factors and their respective receptors. The purpose of the study was to explore the expression of growth factors known to stimulate epithelial cells in other systems, as well as their receptors, in the middle ear mucosa during otitis media. Expression of mRNA growth factors and receptors was measured over time after inoculation of the rat middle ear with bacteria. The middle ears of 12 male Sprague-Dawley rats were injected with 10(5)/mL Haemophilus influenzae strain 3655 (nontypeable, biotype II). Three rats were killed at 6, 24, 48, and 72 hours. Three untreated rats were also killed to serve as negative controls. The middle ear mucosa samples were surgically removed and homogenized. Reverse transcription-polymerase chain reaction was performed on each sample with primers for rat epidermal growth factor, epidermal growth factor receptor (ErbB), heparin binding epidermal-like growth factor, hepatocyte growth factor, hepatocyte growth factor receptor, keratinocyte growth factor, betacellulin, amphiregulin, and neuregulin-alpha. Hepatocyte growth factor and epidermal growth factor receptor primers demonstrated polymerase chain reaction products of the expected size that were not displayed in the normal middle ear mucosa. Keratinocyte growth factor and hepatocyte growth factor receptor demonstrated polymerase chain reaction products at all time points tested. Betacellulin and neuregulin-alpha products were present at all time points except 72 hours after infection. The results of the study support a role for growth factors in the middle ear mucosa during otitis media. These bioactive ingredients contribute to mucosal hyperplasia.

The Wilms' Tumor Gene Product WT1 Mediates the Down-regulation of the Rat Epidermal Growth Factor Receptor by Nerve Growth Factor in PC12 Cells

Recently, we characterized the rat epidermal growth factor receptor (EGFR) promoter and demonstrated that TCC repeat sequences are required for the down-regulation of EGFR by nerve growth factor (NGF) in PC12 cells. In this study, we report that the Wilms' tumor gene product WT1, a zinc finger transcription factor, is able to enhance the activity of the rat EGFR promoter in cotransfection assays. Gel mobility shift assays demonstrate that WT1 binds to the TCC repeat sequences of the rat EGFR promoter. Overexpression of WT1 resulted in up-regulation of the expression levels of endogenous EGFR in PC12 cells. Interestingly, NGF down-regulated the expression levels of WT1 and EGFR in PC12 cells, but not in the p140(trk)-deficient variant PC12nnr5 cells or in cells expressing either dominant-negative Ras or dominant-negative Src. Most importantly, we evaluated the inhibitory effect of antisense WT1 RNA on EGFR expression, and we found that antisense WT1 RNA could substantially reduce EGFR repression in either histochemical staining study or immunoblot analysis. These results indicate that NGF-induced down-regulation of the EGFR in PC12 cells is mediated through WT1 and that WT1 may play an important role in the differentiation of nerve cells.

Regulation of nitric oxide synthase to promote cytostasis in ovarian follicular development.

Our own recent studies have demonstrated that inducible nitric oxide synthase (iNOS) is predominantly localized in granulosa cells of healthy immature follicles in the rat ovary, whereas granulosa cells of either healthy mature follicles or follicles destined to be atretic are devoid of iNOS. These findings suggest that iNOS is pivotal for immature follicles to remain dormant. To test this hypothesis, we examined the effects of a GnRH agonist (buserelin), a proapoptotic substance, and epidermal growth factor (EGF), a mitogenic and, consequently, antiapoptotic factor, on the amount of iNOS mRNA in rat granulosa cells. Administration of buserelin in immature female rats transiently diminished iNOS mRNA levels in the ovaries as determined by Northern blot analysis. In cultured rat granulosa cells, buserelin and EGF increased the incidence of apoptosis and DNA synthesis, respectively, whereas both reduced iNOS mRNA levels as determined by reverse transcription-coupled polymerase chain reaction. The concomitant addition of S-nitroso-N-acetyl-DL-penicillamine, an NO donor, together with buserelin or EGF eliminated the observed effects of these substances (i.e., induction of apoptosis and stimulation of DNA synthesis, respectively). These results suggest that the changes in developmental status of immature follicles either into development or atresia are associated with reduced iNOS levels in granulosa cells, thus reinforcing the notion of NO as a cytostatic factor in ovarian follicles.

DNA sequence and functional characterization of the human and rat epidermal growth factor promoter: regulation by cell growth

Epidermal growth factor (EGF) regulates cell growth and differentiation through intracellular transduction networks activated by its tyrosine kinase receptor, EGFR. In this report we describe the structure and DNA sequence of transcriptional control regions from both human and Wistar-rat single copy EGF genes and their functional analysis in epithelial cell cultures. By sequence comparison we show these proximal gene regions have remained conserved in evolution to −640 (relative to the rodent mRNA initiation site), where similarity is interrupted by a rodent interspersed-repeat element (SINE). Transcript mapping reveals complexity in EGF initiation site selection: whereas a single rat liver initiation site (+1) appears 30 bp 3′ to the TTTAA element, an additional upstream site is detected in kidney RNA at −14. In contrast, in human RNA a single initiation is observed, which is displaced 12 bp 3′ to the rodent RNA terminus. Both promoters were defined in transient expression assays. Our results show the human promoter to be at least 20-fold more active than the equivalent rodent sequence, although both are activated during cell proliferation and negatively regulated in contact inhibited and quiescent cultures. The results indicate EGF gene expression and cell division are temporally linked, suggesting its promoter comprises a growth responsive regulatory domain.

Systemic treatment with epidermal growth factor but not insulin-like growth factor I decreases the involution of the prostate in castrated rats.

Epidermal growth factor (EGF) and insulin-like growth factor I (IGF-I) are strong inducers of proliferation to prostate cells cultured in serum-free medium. Accordingly we wanted to study the growth of the prostate gland in castrated rats after treatment with EGF, IGF-I and testosterone. Castrated Wistar rats were treated with growth factors (EGF 35 microg/rat per day; IGF-I 350 microg/rat per day) or testosterone (2 mg/rat per day) for 3 days either immediately after or 10 days after castration. Prostate tissue was examined by stereological and immunohistochemical techniques and by enzyme-linked immunosorbent assay (ELISA). Treatment with EGF inhibited the involution of the prostate (P < 0.05), whereas treatment with IGF-I did not affect the prostate involution as compared to castrated controls. EGF treatment significantly increased the endogenous rat EGF in the ventral prostate, but cellular proliferation was not affected. Testosterone treatment increased the weight of the prostate, by increase of all tissue components of the prostate, and significantly increased cellular proliferation. Systemic administration of EGF but not IGF-I decreased the involution of the rat prostate induced by castration. Compared with testosterone, the effects of EGF treatment on the prostate involution were moderate, and the effects of EGF were not related to cellular proliferation.

Cloning and Characterization of the Promoter Region of the Rat Epidermal Growth Factor Receptor Gene and Its Transcriptional Regulation by Nerve Growth Factor in PC12 Cells

Our previous studies have shown that treatment of PC12 cells with nerve growth factor (NGF) causes a profound down-regulation of the epidermal growth factor receptor (EGFR) mRNA and protein. Further, the NGF-induced down-regulation of the EGFR is under transcriptional control. To elucidate the molecular mechanism of this down-regulation we have cloned a 2.7-kilobase sequence from the promoter region of the rat EGFR from a rat P1 library. Six transcriptional start sites were identified by 5'-rapid amplification of cDNA ends and primer extension. Sequence analysis showed a 62% overall homology with the human EGFR promoter region. To investigate its transcription, 1.1 kilobases of the 5'-flanking sequence were fused to a luciferase reporter gene. This sequence exhibited functional promoter activity in transient transfection experiments with PC12, C6, and CV-1 cells. Treatment of PC12 cells with NGF inhibited promoter activity. By transfection of promoter deletion constructs, a silencer element was found between nucleotides -260 and -181, and TCC repeat sequences appeared to be at least partially responsible for the down-regulation of the EGFR by NGF. Supportive evidence for the relevance of this sequence was obtained from gel mobility shift assays and by transfection of TCC mutation constructs. Our results demonstrate that TCC repeat sequences are required for the down-regulation of rat EGFR by NGF in PC12 cells and may lead to the identification of the NGF-responsive transcription factors.

The stability of truncated epidermal growth factor receptor mRNA is higher than that of the full-length receptor mRNA in rat hepatoma cells.

Truncated and full-length epidermal growth factor (EGF) receptors are produced in rat liver cells. The truncated EGF receptor mRNA is almost identical to the full-length EGF receptor mRNA except for the lack of a 3' region of the full-length receptor mRNA. To understand the stability of rat EGF receptor mRNAs, we analyzed the expression of EGF receptor mRNAs in the hepatoma cell line, AH66 and liver cells. Ten, 7 and 5 kb full-length and 2.7 kb truncated EGF receptor mRNAs were detected in both of them. The half-lives of the 10 and 2.7 kb EGF receptor mRNAs were determined in AH66 cells using a transcriptional inhibitor, 5,6-dichlororibofuranosylbenzimidazole. The half-lives of the 10 and 2.7 kb mRNAs were 1.2 and 11 h, respectively. These results indicated that the truncated mRNA is 4 times more stable than the full-length mRNA in rat cells. As for the stability, the role of a sequence of 3'-untranslated region of the EGF receptor mRNA was discussed.

Transforming growth factor-alpha in vivo stimulates epithelial cell proliferation in digestive tissues of suckling rats.

The role that exogenous transforming growth factor-alpha (TGF-alpha) may exert on cell proliferation in vivo is poorly understood. To investigate the effect of rat TGF-alpha on epithelial cell proliferation in all suckling rat digestive tissues and to compare it with that of rat epidermal growth factor (EGF). ANIMAL AND METHODS: TGF-alpha and EGF were given three times daily either subcutaneously (10 or 20 micrograms/kg) or intraperitoneally (100 micrograms/kg) to rats from the ninth postnatal day. Cell proliferation was assessed through 5-bromo- 2-deoxyuridine incorporation and estimation of labelling indices. For both growth factors, the highest dose given for only two days significantly increased stomach and intestinal weights compared with controls (p < 0.05 to p < 0.001). The proliferative responded depended on the dose given, colonic mucosa being the most sensitive whereas oxyntic mucosa remained unresponsive. TGF-alpha was as potent as EGF in stimulating epithelial cell proliferation in antral, duodenal, and colonic mucosae. However, EGF was more active on oesophageal and jejunal cell proliferation whereas TGF-alpha was more active on pancreatic exocrine cell proliferation and the differences between the two growth factor treated groups were significant. These results prove for the first time the stimulating effect in vivo of exogenous rat TGF-alpha on epithelial cell proliferation in rat digestive tissues during the developmental period and support a functional role for TGF-alpha at that time.

Epidermal growth factor regulates adenylate cyclase activity via Gs and Gi1-2 proteins in pancreatic acinar membranes

In the present study, Western-blot and radioreceptor analyses have revealed the presence of the epidermal growth factor (EGF) receptor in pancreatic acinar membranes. Isolated pancreatic acinar membranes, which allow access of functional antibodies to individual components of the signal transduction cascade, were used to examine EGF-induced regulation of adenylate cyclase activity. Forskolin, vasoactive intestinal peptide (VIP) and to a smaller extent EGF increased cAMP production in pancreatic acinar membranes. Preincubation of the membranes with anti-GS alpha antibody abolished EGF- and VIP-induced cAMP production, but had no effect on forskolin-induced cAMP accumulation. In the presence of either VIP or forskolin, EGF inhibited the VIP- and forskolin-induced cAMP production with an IC50 of 5 nM. Anti-G alpha i1-2 protein antibody, but not anti-G alpha i3 antibody, increased basal cAMP production, indicating that Gi proteins exert an inhibitory influence on basal adenylate cyclase activity. Anti-G alpha i1-2 antibody, but not anti-G alpha i3 antibody, abolished the inhibitory effect of EGF on the forskolin- and VIP-induced cAMP accumulation. A peptide corresponding to the juxtamembrane region in the cytosolic domain of the rat EGF receptor increased cAMP production in pancreatic acinar membranes in an anti-G alpha s antibody-sensitive fashion, whereas the EGF receptor peptide did not mimic the inhibitory effect of the native EGF receptor. The tyrosine kinase inhibitors genistein and pp60v-src (137-157) inhibited both the stimulatory and the inhibitory effects of EGF on cAMP production. Thus the data of the present study show that EGF regulates adenylate cyclase via activation of Gs and Gi proteins by a tyrosine phosphorylation-dependent mechanism in pancreatic acinar membranes. This leads to stimulation of basal and inhibition of forskolin- and VIP-induced adenylate cyclase activity respectively.

EGF receptor mRNA and protein in rat lacrimal acinar cells: evidence of its EGF-dependent phosphotyrosilation.

This study was designed to demonstrate the presence of the epidermal growth factor (EGF) receptor in the rat exorbital lacrimal gland. EGF receptor gene transcription was demonstrated 1) by reverse transcription-polymerase chain reaction analysis of lacrimal gland and acinar cells from RNA with a set of specific primers deduced from the rat EGF receptor sequence and 2) by Northern blot analysis of rat lacrimal gland mRNA. Lacrimal acinar cell preparations contain a low but detectable amount of specific 125I-EGF binding sites and efficiently internalize the ligand on binding at 37 degrees C. A sheep polyclonal antibody, directed against the human EGF receptor, detects a protein of 170 kDa by Western blot analysis of membrane proteins of the whole gland. This protein can be immunoprecipitated by the same antibody from whole gland membrane proteins as well as from solubilized acinar cells. Incubations of acinar cells in the presence of EGF results in an increased content of tyrosine-phosphorylated residues in immunoprecipitated 170-kDa protein. Taken together, these results demonstrate for the first time both EGF receptor gene transcription and protein expression in a lacrimal tissue, i.e., the rat exorbital lacrimal gland. These results also suggest a specific cellular location of the EGF receptor in a cell population contained in acinar cell preparations.

Regulation of transcription by the rat EGF gene promoter in normal and ischemic murine kidney cells.

Epidermal growth factor (EGF) is a small polypeptide belonging to a class of molecules that can mediate cell growth, differentiation, and acute phase responses. EGF mRNA is transcribed primarily in cells of the salivary gland and the kidney. We have found that the tissue and cellular specificities of EGF gene expression are controlled by a promoter region located upstream from the start of mRNA transcription. In a variety of experimentally induced forms of acute renal failure, the mRNA and protein levels for kidney EGF fall markedly and remain low for a prolonged period. This decrease was determined by nuclear runoff transcription to be a consequence of diminished transcription of the EGF gene rather than increased instability of the mRNA. Using transgenic mice, we found the effect of renal ischemia on EGF mRNA transcription to be a result of a disease-mediated interruption of the function of this upstream promoter region, presumably from alterations in the activity of one or more cellular trans-acting factors.

Hepatocyte cell density regulates a minimal human and rat epidermal growth factor transcriptional regulatory region: [formula omitted], E. Fodor, A. Feren, A. Jones: Cell Biology, VA Medical Center and Dept. of Medicine, and Liver Center, University of California, San Francisco, CA 94143

Hepatocyte cell density regulates a minimal human and rat epidermal growth factor transcriptional regulatory region: [formula omitted], E. Fodor, A. Feren, A. Jones: Cell Biology, VA Medical Center and Dept. of Medicine, and Liver Center, University of California, San Francisco, CA 94143