Prevention of Skin Carcinogenesis by the Non-ß-blocking R-carvedilol Enantiomer.

Skin cancer is the most common malignancy worldwide and is rapidly rising in incidence, representing a significant public health challenge. The ß-blocker, carvedilol, has shown promising effects in preventing skin cancer. However, as a potent ß-blocker, repurposing carvedilol to an anticancer agent is limited by cardiovascular effects. Carvedilol is a racemic mixture consisting of equimolar S- and R-carvedilol, whereas the R-carvedilol enantiomer does not possess ß-blocking activity. Because previous studies suggest that carvedilol's cancer preventive activity is independent of ß-blockade, we examined the skin cancer preventive activity of R-carvedilol compared with S-carvedilol and the racemic carvedilol. R- and S-carvedilol were equally effective in preventing EGF-induced neoplastic transformation of the mouse epidermal JB6 Cl 41-5a (JB6 P+) cells and displayed similar attenuation of EGF-induced ELK-1 activity. R-carvedilol appeared slightly better than S-carvedilol against UV-induced intracellular oxidative stress and release of prostaglandin E2 from the JB6 P+ cells. In an acute UV-induced skin damage and inflammation mouse model using a single irradiation of 300 mJ/cm2 UV, topical treatment with R-carvedilol dose dependently attenuated skin edema and reduced epidermal thickening, Ki-67 staining, COX-2 protein, and IL6 and IL1ß mRNA levels similar to carvedilol. In a chronic UV (50-150 mJ/cm2) induced skin carcinogenesis model in mice with pretreatment of test agents, topical treatment with R-carvedilol, but not racemic carvedilol, significantly delayed and reduced skin squamous cell carcinoma development. Therefore, as an enantiomer present in an FDA-approved agent, R-carvedilol may be a better option for developing a safer and more effective preventive agent for skin carcinogenesis. PREVENTION RELEVANCE: In this study, we demonstrated the skin cancer preventive activity of R-carvedilol, the non-ß-blocking enantiomer present in the racemic ß-blocker, carvedilol. As R-carvedilol does not have ß-blocking activity, such a preventive treatment would not lead to common cardiovascular side effects of ß-blockers.

Dermal toxicity, dermal irritation, and delayed contact sensitization evaluation of oil body linked oleosin-hEGF microgel emulsion via transdermal drug delivery for wound healing.

Objective: The expression of therapeutic proteins in plant oil body bioreactors has attracted much attention. But its safety is not yet clear. This article determines the risk of safety after using the drug. Methods: The oil body-linked oleosin-hEGF microgel emulsion (OBEME) was prepared by mixing the xanthan gum with suitable concentrations in an appropriate proportion. Skin irritation and sensitization reaction were investigated in rats and guinea pigs using OBEME as test article.Results: The OBEME did not produce dermal erythema/eschar or oedema responses. The dermal subacute and subchronic toxicity of OBEME were evaluated in accordance with OECD guidelines. Compared with the control group, the basic physical signs, such as weight, feed, drinking, excretion, and behaviour of experimental animals, were not abnormal. In addition, no abnormality was found in haematological parameters, biochemical indexes, relative organ weight, and histopathological observation of organs, and there was no significant difference compared with normal saline treatment group. Therefore, we conclude that OBEME has no toxic effects and is safe and reliable to be used for topical application.

FR - Inhibidores del factor de crecimiento epidérmico y toxicidad cutánea: en busca de un protocolo de manejo / RF - Epidermal Growth Factor Receptor Inhibitors and Skin Toxicity: The Search for a Management Protocol

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EGF-GH Axis in Rat Steatotic and Non-steatotic Liver Transplantation From Brain-dead Donors.

BACKGROUND: We evaluated the potential dysfunction caused by changes in growth hormone (GH) levels after brain death (BD), and the effects of modulating GH through exogenous epidermal growth factor (EGF) in steatotic and nonsteatotic grafts. METHODS: Steatotic and nonsteatotic grafts from non-BD and BD rat donors were cold stored for 6 hours and transplanted to live rats. Administration of GH and EGF and their underlying mechanisms were characterized in recipients of steatotic and nonsteatotic grafts from BD donors maintained normotensive during the 6 hours before donation. Circulating and hepatic GH and EGF levels, hepatic damage, and regeneration parameters were evaluated. Recipient survival was monitored for 14 days. Somatostatin, ghrelin, and GH-releasing hormones that regulate GH secretion from the anterior pituitary were determined. The survival signaling pathway phosphoinositide-3-kinase/protein kinase B that regulates inflammation (suppressors of cytokine signaling, high-mobility group protein B1, oxidative stress, and neutrophil accumulation) was evaluated. RESULTS: BD reduced circulating GH and increased GH levels only in steatotic livers. GH administration exacerbated adverse BD-associated effects in both types of graft. Exogenous EGF reduced GH in steatotic livers, thus activating cell proliferation and survival signaling pathways, ultimately reducing injury and inflammation. However, EGF increased GH in nonsteatotic grafts, which exacerbated damage. The benefits of EGF for steatotic grafts were associated with increased levels of somatostatin, a GH inhibitor, whereas the deleterious effect on nonsteatotic grafts was exerted through increased amounts of ghrelin, a GH stimulator. CONCLUSIONS: GH treatment is not appropriate in rat liver transplant from BD donors, whereas EGF (throughout GH inhibition) protects only in steatotic grafts.

Neonatal exposure to an inflammatory cytokine, epidermal growth factor, results in the deficits of mismatch negativity in rats.

Perinatal exposure to epidermal growth factor (EGF) induces various cognitive and behavioral abnormalities after maturation in non-human animals, and is used for animal models of schizophrenia. Patients with schizophrenia often display a reduction of mismatch negativity (MMN), which is a stimulus-change specific event-related brain potential. Do the EGF model animals also exhibit the MMN reduction as schizophrenic patients do? This study addressed this question to verify the pathophysiological validity of this model. Neonatal rats received repeated administration of EGF or saline and were grown until adulthood. Employing the odd-ball paradigm of distinct tone pitches, tone-evoked electroencephalogram (EEG) components were recorded from electrodes on the auditory and frontal cortices of awake rats, referencing an electrode on the frontal sinus. The amplitude of the MMN-like potential was significantly reduced in EGF-treated rats compared with saline-injected control rats. The wavelet analysis of the EEG during a near period of tone stimulation revealed that synchronization of EEG activity, especially with beta and gamma bands, was reduced in EGF-treated rats. Results suggest that animals exposed to EGF during a perinatal period serve as a promising neurodevelopmental model of schizophrenia.

A dual deformable liposomal ointment functionalized with retinoic acid and epidermal growth factor for enhanced burn wound healing therapy.

An ointment containing retinoic acid deformable liposomes (TRA DLs) and epidermal growth factor cationic deformable liposomes (EGF CDLs) was prepared for the treatment of deep partial-thickness burns. The characterization tests confirmed both liposomes featured small particle sizes, high drug entrapment efficiencies and sustained drug release behavior. Compared with the free drug, TRA DLs and EGF CDLs exhibited superior skin permeation and remarkably increased drug deposition by 2.9 and 18.8 folds, respectively. Results on HaCaT cells indicated the combined application of two liposomes exerted a synergistic effect and prominently promoted cell proliferation and migration. Application of the dual liposomal ointment on a deep partial-thickness burn model stimulated wound closure (p < 0.001), promoted skin appendage formation and increased collagen production, thus improving healing quality. Finally, it was demonstrated that TRA significantly up-regulated the expression of EGFR and HB-EGF to enhance the therapeutic effect of EGF. Therefore, the dual liposomal ointment is a promising topical therapeutic for burn treatment.

Rapamycin mediates mTOR signaling in reactive astrocytes and reduces retinal ganglion cell loss.

Damage and loss of retinal ganglion cells (RGCs) can cause visual impairment. The underlying molecular mechanisms that mediate RGC death in ischemic retinal diseases are still unclear. In this study, we sought to understand the neuroprotective effect of rapamycin, the selective inhibitor of mTORC1, on RGC survival and the cellular mechanics that mediate this effect. Recent studies have reported that the epidermal growth factor (EGF) receptor shows an increase in expression in astrocytes after injury, and this receptor can promote their transformation into reactive astrocytes. Our results, along with previous works from others, show the colocalization of phosphor-EGF receptors with the astrocyte marker glial fibrillary acidic proteins in reactive astrocytes in the injured retina. In our in vitro studies, using primary astrocyte cultures of the optic nerve head of rats, showed that rapamycin significantly blocked EGF-induced mTOR signaling mainly through the PI3K/Akt pathway in primary astrocytes, but not through the MAPK/Erk pathway. Additionally, rapamycin dramatically inhibited the activation of mTOR signaling in our ratinal ischemia-reperfusion (I/R) injury model in vivo. Astrocyte activation was assessed by immunostaining retinal flat mounts or cross sections with antibody against GFAP, and we also used western blots to detect the expression of GFAP. Taken together, these results revealed that rapamycin decreases the activation of astrocytes after retinal ischemia-reperfusion injury. Furthermore, rapamycin can improve retinal RGC survival in rats during I/R, as detected by FluoroGold labeling. Our data reveals the neuroprotective effects of rapamycin in an experimental retina injury model, possibly through decreasing glial-dependent intracellular signaling mechanisms for suppressing apoptosis of RGCs. Our study also presents an approach to targeting reactive astrocytes for the treatment of optic neurodegenerations.

Human epidermal growth factor coupled to different structural classes of cell penetrating peptides: A comparative study.

Human epidermal growth factor (hEGF) plays important roles in wound healing. Due to large molecular weight and hydrophilic nature, cellular uptake and skin permeation of hEGF are very poor, significantly limiting its efficacy. By using recombinant technology, four structural classes of cell penetrating peptides (CPPs) were fused at the C-terminus of hEGF, expressed and purified into homogeneity. Comparative studies were conducted to evaluate their activity, cytotoxicity, cellular uptake and skin permeation. Cell viability assay and in vitro scratch wound-healing assay showed that all four fusion proteins had similar activities with commercial rhEGF. Obvious cytotoxicity was not detected for EGF-TAT, EGF-Pep-1 and EGF-AA3H. However, EGF-MAP was cytotoxic at both moderate and high concentrations. Confocal microscopy indicated that the cellular uptake of the fusion proteins was markedly improved compared with rhEGF, with EGF-TAT and EGF-Pep-1 showing the most abundant presence within cells at incubation concentration of 25µM. Permeation across the excised mouse skin followed the order of EGF-Pep-1>EGF-TAT>EGF-AA3H>rhEGF. These findings demonstrated that there were great gaps between the abilities of different structural types of CPPs to deliver EGF across cell membrane and the skin. EGF coupled with a well-chosen CPP will become a more promising pharmaceutical agent than rhEGF.

Apobec1 Promotes Neurotoxicity-Induced Dedifferentiation of Müller Glial Cells.

Retinal Müller glial cells in mammals acquire stem and progenitor cell properties after neurotoxic treatment. However, the molecular mechanisms underlying proliferation and dedifferentiation of adult Müller cells in the mammalian retina were unclear. In this study, treatments with N-methyl-D-aspartate (NMDA) plus epidermal growth factor (EGF) led to the proliferation of Müller cells and expression of stem cell markers including Nanog and Nestin in the retina. The increased mRNA for Nanog and Nestin were coincident with reduced methylation of a Nanog promoter and a Nestin enhancer specific in the neural stem cells, respectively. We found that Apolipoprotein B mRNA editing catalytic subunit 1 (Apobec1) was upregulated early in the retina treated with NMDA and EGF. Moreover, overexpression of Apobec1 in primary Müller cells increased expression of Nestin and reduced methylation of the Nestin enhancer. The data suggest that neurotoxicity-induced Apobec1 may promote expression of Nestin and help cell cycle reentry of retinal Müller cells via DNA demethylation. This study provides novel insights into the molecular mechanisms underlying dedifferentiation and proliferation of Müller cells in the mammalian retina.

Increased l1 retrotransposition in the neuronal genome in schizophrenia.

Recent studies indicate that long interspersed nuclear element-1 (L1) are mobilized in the genome of human neural progenitor cells and enhanced in Rett syndrome and ataxia telangiectasia. However, whether aberrant L1 retrotransposition occurs in mental disorders is unknown. Here, we report high L1 copy number in schizophrenia. Increased L1 was demonstrated in neurons from prefrontal cortex of patients and in induced pluripotent stem (iPS) cell-derived neurons containing 22q11 deletions. Whole-genome sequencing revealed brain-specific L1 insertion in patients localized preferentially to synapse- and schizophrenia-related genes. To study the mechanism of L1 transposition, we examined perinatal environmental risk factors for schizophrenia in animal models and observed an increased L1 copy number after immune activation by poly-I:C or epidermal growth factor. These findings suggest that hyperactive retrotransposition of L1 in neurons triggered by environmental and/or genetic risk factors may contribute to the susceptibility and pathophysiology of schizophrenia.

Blockade of the intermediate-conductance Ca(2+)-activated K+ channel inhibits the angiogenesis induced by epidermal growth factor in the treatment of corneal alkali burn.

Epidermal growth factor (EGF) is used to treat alkali-burned corneas. However, EGF-induced corneal angiogenesis, which is currently untreatable, is a side effect of this therapy. We therefore explored the role of the intermediate-conductance Ca(2+)-activated K(+) channel (KCa3.1) in EGF-induced angiogenesis and tested whether KCa3.1 blockade can suppress EGF-induced corneal angiogenesis. The proliferation, migration and tube formation of HUVECs (human umbilical vein endothelial cells) in response to EGF, the MEK inhibitor PD98059 and the KCa3.1 inhibitor TRAM-34 were analyzed in vitro via MTT, cell counting, scratch and tube formation assays. The protein and mRNA levels of KCa3.1, phosphorylated-ERK (P-ERK), total-ERK (T-ERK), cyclin-dependent kinase 4 (CDK4), vimentin and MMP-2 were assessed via western blotting and RT-PCR. KCa3.1 and vimentin expression were also detected through immunofluorescence staining. Flow cytometry was performed to examine the cell cycle. Further, an in vivo murine alkali-burned cornea model was developed and treated with EGF and TRAM-34 eye drops to analyze the effect of these treatments on corneal healing and angiogenesis. The corneas were also analyzed by histological staining. The in vitro results showed that EGF induces the upregulation of KCa3.1 and P-ERK in HUVECs and that this upregulation is suppressed by PD98059. EGF stimulates proliferation, migration and tube formation in HUVECs, and this effect can be suppressed by TRAM-34. TRAM-34 also arrests HUVECs in the G1 phase of the cell cycle and downregulates CDK4, vimentin and MMP-2 in these cells. The in vivo results indicated that TRAM-34 suppresses EGF-induced corneal angiogenesis without affecting EGF-induced corneal wound healing. In summary, the upregulation of KCa3.1 may be crucial for EGF-induced angiogenesis through the MAPK/ERK signaling pathway. Thus, KCa3.1 may be a potential target for the treatment of EGF-induced corneal angiogenesis.

Chitosan nanoparticles as a dual growth factor delivery system for tissue engineering applications.

Growth factors are essential in cellular signaling for migration, proliferation, differentiation and maturation. Sustainable delivery of therapeutic as well as functional proteins is largely required in the pharmacological and regenerative medicine. Here we have prepared chitosan nanoparticles (CNP) and incorporated growth factors such as epidermal growth factor (EGF) and fibroblast growth factor (FGF), either individually or in combination, which could ultimately be impregnated into engineered tissue construct. CNP was characterized by Fourier transform infrared (FTIR) spectroscopy, Zeta sizer and high resolution transmission electron microscope (HRTEM). The particles were in the size range of 50-100 nm with round and flat shape. The release kinetics of both EGF and FGF incorporated CNP showed the release of growth factors in a sustained manner. Growth factors incorporated nanoparticles did not show any toxicity against fibroblasts up to 4 mg/ml culture medium. Increased proliferation of fibroblasts in vitro evidenced the delivery of growth factors from CNP for cellular signaling. Western blotting results also revealed the poor inflammatory response showing less expression of proinflammatory cytokines such as IL-6 and TNFα in the macrophage cell line J774 A-1.

The anthraquinone derivative Emodin ameliorates neurobehavioral deficits of a rodent model for schizophrenia.

Abnormality in cytokine signaling is implicated in the neuropathology of schizophrenia. Previously, we established an animal model for schizophrenia by administering epidermal growth factor (EGF) to neonatal rats. Here we investigated effects of the anthraquinone derivatives emodin (3-methyl-1,6,8-trihydroxyanthraquinone) and sennoside (bis-[D: -glucopyranosyl-oxy]-tetrahydro-4,4'-dihydroxy-dioxo[bianthracene]-2,2'-dicarboxylic acid) on behaviors of this model and EGF signaling. Subchronic oral administration of emodin (50 mg/kg) suppressed acoustic startle responses and abolished prepulse inhibition (PPI) deficits in this rodent model. ANCOVA revealed that emodin had distinct effects on PPI and startle responses. In contrast, sennoside (50 mg/kg) had no effects. Emodin attenuated weight gain initially during treatment but had no apparent effect on weight gain and locomotor activity thereafter. Application of emodin to neocortical cultures attenuated the phosphorylation of ErbB1 and ErbB2. We conclude that emodin can both attenuate EGF receptor signaling and ameliorate behavioral deficits. Therefore, emodin might be a novel class of a pro-drug for anti-psychotic medication.

Fyn is a novel target of (-)-epigallocatechin gallate in the inhibition of JB6 Cl41 cell transformation.

The cancer preventive action of (-)-epigallocatechin gallate (EGCG), found in green tea, is strongly supported by epidemiology and laboratory research data. However, the mechanism by which EGCG inhibits carcinogenesis and cell transformation is not clear. In this study, we report that EGCG suppressed epidermal growth factor (EGF)-induced cell transformation in JB6 cells. We also found that EGCG inhibited EGF-induced Fyn kinase activity and phosphorylation in vitro and in vivo. Fyn was implicated in the process because EGF-induced JB6 cell transformation was inhibited by small interfering RNA (siRNA)-Fyn-JB6 cells. With an in vitro protein-binding assay, we found that EGCG directly bound with the GST-Fyn-SH2 domain but not the GST-Fyn-SH3 domain. The K(d) value for EGCG binding to the Fyn SH2 domain was 0.367 +/- 0.122 microM and B(max) was 1.35 +/- 0.128 nmol/mg. Compared with control JB6 Cl41 cells, EGF-induced phosphorylation of p38 MAP kinase (p38 MAPK) (Thr180/Tyr182), ATF-2 (Thr71) and signal transducer and activator of transcription 1 (STAT1) (Thr727) was decreased in siRNA-Fyn-JB6 cells. EGCG could inhibit the phosphorylation of p38 MAPK, ATF-2, and STAT1. The DNA binding ability of AP-1, STAT1, and ATF-2 was also decreased in siRNA-Fyn-JB6 cells. Overall, these results demonstrated that EGCG interacted with Fyn and inhibited Fyn kinase activity and thereby regulated EGF-induced cell transformation. Inhibition of Fyn kinase activity is a novel and important mechanism that may be involved in EGCG-induced inhibition of cell transformation.

Inhibition of gap junctional intercellular communication by noncoplanar polychlorinated biphenyls: inhibitory potencies and screening for potential mode(s) of action.

Polychlorinated biphenyls (PCBs), a structurally diverse group of environmental pollutants, are effective promoters in two-stage cancer models, which implies that epigenetic mechanisms are involved. Inhibition of gap junctional intercellular communication (GJIC) belongs among critical epigenetic events of tumor promotion. We determined the relative potencies of a series of environmentally relevant PCB congeners to inhibit GJIC in vitro in a rat liver epithelial cell line with pluripotent oval cell characteristics. The nonplanar PCBs were potent inhibitors of GJIC, whereas the coplanar PCBs did not inhibit GJIC. We then compared the effects of the coplanar PCB 126 (3,3',4,4',5-pentachlorobiphenyl) and the noncoplanar PCB 153 (2,2',4,4',5,5'-hexachlorobiphenyl) with effects of two model GJIC inhibitors, a tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and epidermal growth factor (EGF). In contrast to TPA or EGF, PCB 153 elicited a long-term downregulation of GJIC (up to 48 h). Using Western blot analysis with phospho-specific antibodies, it was found that PCB 153, and not PCB 126, activated mitogen-activated protein kinases ERK1/2; however in contrast to TPA and EGF, this activation was observed at the time points subsequent to GJIC inhibition. Moreover, blocking of ERK1/2 activation did not prevent the GJIC inhibition induced by PCB 153. Therefore, additional intracellular signaling pathways potentially involved in the downregulation of GJIC by PCBs were screened by using specific chemical probes inhibiting serine/threonine kinases, tyrosine kinases, and phospholipases. The inhibition of diacylglycerol lipase partially blocked and the selective inhibition of Src kinases and phosphatidylcholine-specific phospholipase C (PC-PLC) completely blocked the inhibitory effects of the noncoplanar PCB on GJIC, indicating that PC-PLC or sphingomyelinase and Src might be upstream regulators of noncoplanar PCB-induced inhibition of GJIC.

An epidermal growth factor-like toxin and two sodium channel toxins from the sea anemone Stichodactyla gigantea.

Three peptide toxins (gigantoxins I-III) with crab toxicity were isolated from the sea anemone Stichodactyla gigantea by gel filtration on Sephadex G-50 and reverse-phase HPLC on TSKgel ODS-120T and their complete amino acid sequences were determined. Gigantoxins II (44 residues) and III (48 residues) have LD(50) (against crabs) of 70 and 120 microg/kg, respectively, and are analogous to the known type 1 and 2 sea anemone sodium channel toxins, respectively. On the other hand, gigantoxin I (48 residues) is potently paralytic to crabs (ED(50) 215 microg/kg), although its lethality is very weak (LD(50)>1000 microg/kg). Interestingly, gigantoxin I has 31-33% homologies with mammalian epidermal growth factors (EGFs), with the same location of six cysteine residues. In accordance with the sequence similarity, gigantoxin I exhibits EGF activity as evidenced by rounding of A431 cells and tyrosine phosphorylation of the EGF receptor in the cells, although much less potently than human EGF. Gigantoxin I is the first example of EGF-like toxins of natural origin.

A cleavable adapter to reduce nonspecific cytotoxicity of recombinant immunotoxins.

One of the problems associated with the administration of immunotoxins is hypersensitivity reaction such as vascular leak syndrome. This may be prevented by decreasing the plasma half-life. To improve immunotoxins with respect to reduced side effects, we have previously described the development of a cleavable adapter. This adapter links the toxic moiety and ligand that are usually directly coupled. In our study, the cytotoxicity of saporin linked either directly or via the adapter to epidermal growth factor (EGF) was evaluated in vitro. The immunotoxins exhibited similar cytotoxic activity towards A-431 and HER14 cells (IC(50) < 10 nM). The supernatant from 6 hr cultures of HER14 cells incubated in the presence of the adapter-containing immunotoxin exhibited a significantly reduced cytotoxicity as compared to the directly coupled immunotoxin. Western blotting revealed that the adapter was cleaved, thus supporting our proposal that cleavable adapters may reduce nonspecific effects. A similar reduced half-life was detected in platelet-poor plasma. In contrast MCF-7 cells remain unaffected by the immunotoxins. This was shown to be due to the absence of detectable EGF-receptor in comparison to A-431 and HER14 cells as determined by Western blotting. Furthermore, we could show that the adapter does not exert an effect on the N-glycosidase activity of saporin. These results suggest that the use of cleavable adapters may be a useful tool in immunotoxins for reducing the killing of surrounding noncancerous cells due to nonspecific binding.

Systemic proliferative changes and clinical signs in cynomolgus monkeys administered a recombinant derivative of human epidermal growth factor.

Epidermal growth factor (EGF) effects have been explored extensively in vivo in rodents, but little is known about trophic responses in nonhuman primates. A previous publication reports the hyperplastic epithelial/parenchymal changes noted in the digestive tract (tongue, esophagus, stomach, intestine, liver, gallbladder, pancreas, and salivary glands) of adult cynomolgus monkeys treated with recombinant human EGF(1-48) (rhEGF(1-48)). This report documents clinical findings and structural effects in the remaining epithelium-containing tissues of these animals. Two monkeys/sex/dose received rhEGF(1-48) by intravenous bolus at 0 (vehicle), 10, 100, 500 (females only), or 1,000 microg/kg/day (males only) daily for up to 2 weeks. Treatment- and dose-related clinical findings included emesis, fecal alterations (soft feces and diarrhea), lacrimation, nasal discharge, hypoactivity, transient hypotension, and salivation after dosing. Male monkeys administered 1,000 microg/kg became moribund after 5 days of treatment and were necropsied. All other monkeys completed the 2-week treatment period. Necropsy findings in nongastrointestinal tissues were: enlarged, pale kidneys at 100 microg/kg and greater; small thymuses seen sporadically at all doses; and enlarged adrenals and small thyroids in males at 1,000 microqg/kg. Respective organ-to-brain weight ratios at 500 and 1,000 microg/kg for kidneys were 1.5- and 2.6-fold greater and for heart were 1.7- and 1.3-fold greater than controls. Microscopically, pronounced dose-related epithelial hypertrophy and hyperplasia were evident in kidney, urinary bladder, skin (epidermis and adnexa), mammary gland, prostate, seminal vesicles, epididymis, uterus, cervix, vagina, thyroid, thymus, tonsillar crypts, cornea, trachea, and pulmonary airways. Epitheliotrophic effects were conspicuous in many tissues at 100 to 1,000 microg/kg. Changes to renal collecting ducts were present at 10 microg/kg, suggesting that kidneys were a relatively sensitive target. Proliferative alterations were not apparent in testes, intraocular structures, brain ependyma and choroid plexus at any dose. Aside from the noted exceptions, rhEGF(1-48) was a pantrophic epithelial mitogen in cynomolgus monkeys when used intravenously at suprapharmacologic doses.

Growth and development in rats given recombinant human epidermal growth factor(1-48) as neonates.

To assess effects of supraphysiologic doses of human recombinant epidermal growth factor(1-48) (rhEGF(1-48)) on neonatal rats, 10 litters of Wistar rats/treatment group were given 0 (formulated vehicle), 10, 100, or 1000 microg/kg daily by subcutaneous injection on postnatal days (PND) 1 through 6. Clinical signs, body weight, acquisition of developmental landmarks and reflexes, and behavior were monitored during treatment and for 5 weeks thereafter (to PND 42). A subset of animals was euthanized weekly from PND 7-28 and necropsied. Selected tissues were examined microscopically. Body weight gain at 1000 microg/kg during treatment was significantly less than control. Precocious incisor eruption, eye opening, vaginal opening, and preputial separation occurred at 100 and/or 1000 microg/kg. Acquisition of reflexes (negative geotaxis, wire maneuver, acoustic startle reflex, and visual placing) was delayed at 1000 microg/kg. Acquisition of adult locomotion was also delayed at 1000 microg/kg. These effects were transient, as locomotor activity at PND 28 and 42 did not differ from control. Effects on acoustic-startle responding persisted in females to final assessment on PND 42. Habituation to repeated acoustic stimuli was impaired, as well as response inhibition following a prepulse acoustic stimulus. rhEGF(1-48) induced structural changes in the skin, retina, kidney, oral and nasal mucosa, lung, and liver. Many of these changes were consistent with the expected mitogenic activity of rhEGF(1-48) and were transient in nature, as severity and incidence diminished with time. An exception was changes observed in the retina at 1000 microg/kg (rosettes/folds and focal defects in the outer nuclear/photoreceptor layers) that were still present 3 weeks after termination of treatment. Acceleration of developmental landmarks; suppression of reflexes, behavior, and somatic growth; and mitogenic responses in epidermal tissues have been reported in rodents treated with epidermal growth factor (EGF) derived from various mammalian species. These results demonstrate that a 48-amino acid fragment of human EGF produced by recombinant technology also induces such effects.