Localization Of Epidermal Growth Factor Research Articles

Effect of vitamin B17 (amygdalin) found in apricot kernel on the irradiated salivary glands of albino rats.

Radiotherapy is used as a treatment for head and neck cancers but increases the risk of salivary gland hypofunction. The management strategies include pharmacotherapies such as salivary substitutes and sialagogues which are largely temporary. In this study, we examine the regenerative potential of vitamin B17 to improve salivary gland function. The present investigation aims to identify the effect of vitamin B17 (amygdaline) on the irradiated parotid salivary gland of albino rats. Twenty-eight adult male albino rats were randomly divided into two groups subjected to irradiation procedure. Fourteen were in the control group, receiving a daily 5 mL saline by oral gavage (7 rats for 14 days and 7 rats for 30 days) while the other fourteen were treated with a daily dose of vitamin B17 (grounded apricot kernel; GAK) at 400 mg/kg in 5 mL of saline by oral gavage (7 rats for 14 days and 7 rats for 30 days). The parotid glands were dissected from the two groups at 14 and 30 days from the day of exposure to irradiation. The parotid gland sections were subjected to H&E stain, immunohistochemical localization of epidermal growth factor (EGF) and PCR using transforming growth factor beta 2 (TGF-β2). The histological abnormalities corroborate with the immunohistochemical localization of EGF and the PCR results of TGF β2, as their up-regulation in the control group demonstrate oxidative stresses and inflammation. The Treatment with GAK decreased oxidative stress and inflammation while promoting tissue regeneration. Vitamin B17 is a promising anti-inflammatory agent that boosts immunity, as the experimental group showed better histological architecture of the parotid gland than the other one.

Localisation of epidermal growth factor (EGF), its specific receptor (EGF-R) and aromatase at the materno-fetal interface during placentation in the pregnant mare

IntroductionImplantation and placentation in the mare does not commence until as late as day 40 after ovulation. The reasons for this and the growth factors and/or hormones which drive placentation when it does finally occur are of considerable academic and practical interest. MethodPlacental interface tissues recovered from 11 accurately aged and perfused-fixed horse uteri between 20 and 68 days of gestation were stained immunocytochemically for Epidermal Growth Factor (EGF), its specific receptor (EGF-R) and for the steroid hormone enzyme, aromatase. ResultsEGF was present in endometrial gland and lumenal epithelia from day 20 but staining intensity increased noticeably for the protein between days 30 and 40, coincidentally with the commencing secretion of equine Chorionic Gonadotrophin (eCG) from the endometrial cups and immediately prior to attachment and commencing interdigitation between the allantochorion and endometrium. EGF-R, on the other hand, was expressed strongly on the cell surface membrane of both non-invasive and invasive trophoblast and it similarly increased in staining intensity between days 30 and 40. Aromatase, the enzyme necessary for conversion of C-19 androgens to C-18 oestrogens, was expressed strongly and constantly from as early as day 12 in the non-invasive trophoblast of the allantochorion, but not the invasive trophoblast of the chorionic girdle, the progenitor tissue of the endometrial cups. DiscussionThe findings support the hypothesis that, in equine pregnancy, the maternal growth factor EGF synergises with maternally and fetally secreted oestrogens to drive the rapid growth and extensive vascularisation of the non-invasive, epitheliochorial, microcotyledonary placenta which results in the birth of the precocious foal after only 11 months gestation.

Expression and localization of epidermal growth factor, transforming growth factor-α and epidermal growth factor receptor in the canine testis.

Gene expression of epidermal growth factor (EGF), transforming growth factor-α (TGF-α) and EGF receptor (EGF-R) and the localization of the corresponding proteins in the canine testis were studied. Levels of mRNA expressions were determined by semiquantitative reverse transcription polymerase chain reaction in the testes of the peripubertal (4–6 months), young adult (3–4 years), advanced adult (7–8 years) and senescent (11–16 years) groups. The EGF-R mRNA level in the testes of the peripubertal group was significantly higher than those in the other groups, whereas there was no difference in EGF and TGF-α mRNA levels among groups. Immunohistochemical stainings for EGF, TGF-α and EGF-R in the testis revealed that immunoreactivity in the seminiferous epithelium and Sertoli cell was weak and nonspecific for the stage of spermatogenesis, and distinct staining was found in Leydig cells. These results suggest that the EGF family of growth factors may be involved in testicular maturation and function in the dog.

Protein localization of epidermal growth factor in sheep ovaries and improvement of follicle survival and antrum formation in vitro.

The aims of this study were to characterize EGF protein expression in ovine ovaries and to verify the effect of EGF on the in vitro development of isolated pre-antral follicles. After collection, ovarian tissue was fixed for immunohistochemical analysis. Additional pairs of ovaries were collected, and secondary follicles were cultured for 18days in α-MEM(+) (control) alone or supplemented with EGF (1, 10 or 50ng/ml). The immunostaining for EGF was observed in oocytes from pre-antral and antral follicles, in granulosa cells of primary and secondary follicles, as well as in cumulus and mural cells of antral follicles. After 18days, the results showed that treatment with 50ng/ml EGF significantly increased the percentage of morphologically normal follicles compared with the control group (α-MEM(+) ) and significantly reduced the precocious extrusion of oocytes and increased the percentage of antral follicles compared with the control and 1ng/ml EGF. All the treatments induced a progressive and significant increase of the follicular diameter throughout the period of culture. However, there were no significant differences in follicular diameter or in the daily growth rate among treatments. In conclusion, this study demonstrated the presence of EGF in ovine ovaries. Moreover, 50ng/ml EGF increased the percentage of normal follicles and improved antrum formation in isolated ovine follicles after 18days of in vitro culture.

Epidermal growth factor, transforming growth factor-α, and epidermal growth factor receptor expression and localization in the canine endometrium during the estrous cycle and in bitches with pyometra

Gene expression and immunohistochemical localization of epidermal growth factor (EGF), transforming growth factor-α (TGF-α), and epidermal growth factor receptor (EGF-R) were compared between the endometrium of bitches ( Canis familiaris) with pyometra accompanied by cystic endometrial hyperplasia (CEH) and that of healthy bitches at similar stages of the estrous cycle. In normal bitches, endometrial TGF-α mRNA levels were highest at proestrus and gradually decreased as the cycle progressed to anestrus. Epidermal growth factor receptor mRNA levels were not significantly affected by the stage of the estrous cycle. Epidermal growth factor mRNA levels were higher at Day 35 of diestrus than at other stages of the estrous cycle (P < 0.05). In bitches with pyometra, endometrial TGF-α and EGF-R mRNA levels did not differ significantly from those at diestrus in normal bitches, but EGF mRNA levels were lower than those at Day 35 of diestrus in normal bitches (P < 0.05). In normal bitches, positive immunohistochemical staining for TGF-α, EGF, and EGF-R was mainly present in the glandular and luminal epithelial cells of the endometrium. In contrast, in bitches with pyometra, immunoreactivity for EGF was clearly present in endometrial stromal cells. Inflammatory cells that had infiltrated the endometrial stroma stained strongly for TGF-α and EGF-R. Luminal and glandular epithelial cells also stained positive for EGF-R. In conclusion, expression of TGF-α by inflammatory cells and a low level of expression and differential localization of EGF may be involved in aberrant growth of endometrial glands and development of CEH.

Localization of epidermal growth factor (EGF) and its receptor (EGFR) during postnatal testis development in the alpaca ( Lama pacos)

The objective of the present studies was to determine the localization of epidermal growth factor (EGF) and the epidermal growth factor receptor (EGFR) in testicular tissue collected from male alpacas at 12 and 24 months of age. In the testes of 12-month-old alpacas, positive staining for EGF was not detected. EGFR was localized to Leydig cells within the 12-month-old alpaca testis, but staining was absent within seminiferous tubules. At 24 months of age, EGF was localized to Leydig cells, peritubular myoid cells, Sertoli cells and germ cells of the alpaca testis, with a preferential adluminal compartment staining within the seminiferous tubules. EGFR was also localized to the Leydig cells, peritubular myoid cells, Sertoli cells and germ cells within the 24-month-old alpaca testis, but staining within the tubules was primarily within the basal compartment. Results indicate distinct temporal and spatial regulation of EGF and EGFR in the alpaca testis and support a potential role for EGF and its related ligands in alpaca testis development and spermatogenesis.

Keratinocyte growth factor receptor (KGF-R) in cholesteatoma tissue

Conclusion. Distribution of the receptor for epidermal growth factor (EGF-R) and of the receptor for the keratinocyte growth factor (KGF-R) in cholesteatoma was found to differ in analogy with other epithelial tissues and accordingly to epidermal differentiation and intensity of paracrine stimulation. Moreover, both EGF-R and KGF-R expression was increased, suggesting a fair correlation with aggressiveness and recurrence rate of this pathology. Objectives. To obtain information on the biological behaviour of cholesteatoma by assessing the expression and localization of EGF-R and KGF-R and correlating their tissue distribution with that of cytokeratins as a marker of differentiation. Materials and methods. Cholesteatoma tissue was taken during tympanoplasty surgery and processed for indirect immunofluorescence. Murine monoclonal antibodies were tested for the different growth factor receptors and pancytokeratins analysed. Fluorescence intensity signal was measured on randomly captured digital images, using FISH 2000/HI software, with a pseudocolours generation module. Results. EGF-R was mostly expressed at the level of keratinocytes of the basal layer, while KGF-R signal was mainly distributed on the spinous and granular suprabasal layers that were also highly positive for cytokeratins. Significant correlation between the immunofluorescence signals was found for KGF-R and cytokeratins only, demonstrating that KGF-R expression is increased in more differentiated areas of the cholesteatoma tissue, while EGF-R is associated with proliferative and migratory portions of the lesion.

Localization of the epidermal growth factor (EGF) and epidermal growth factor receptor (EGFR) in the bovine testis

In the last few decades, several growth factors were identified in the testis of various mammalian species. Growth factors are shown to promote cell proliferation, regulate tissue differentiation, and modulate organogenesis. In the present investigation we have studied the localization of EGF and EGFR in the adult bovine testis by means of immunohistochemical method. Our results demonstrated that EGF and EGFR were localized solely to the bovine testicular germ cells (spermatogonia, spermatocytes, and round spermatids). In contrast, the somatic testicular cells (i.e., Sertoli, Leydig, and myofibroblast cells) exhibited no staining affinity. EGF and EGFR were additionally detected in the epithelial lining of straight tubules and rete testis. Interestingly, the distribution of EGF and EGFR in the germ cells was mainly dependent upon the cycle of the seminiferous epithelium since their localization appeared to be preponderant during the spermatogonia proliferation and during the meiotic and spermiogenic processes. In conclusion, such findings may suggest that EGF and EGFR are important paracrine and/or autocrine regulators of spermatogenesis in bovine.

Application of EGFP-EGF fusions to explore mechanism of endocytosis of epidermal growth factor

To develop a simple method for monitoring protein localization of epidermal growth factor (EGF) in living cells. Enhanced green fluorescent protein (EGFP) was used as an autofluorescent tag to label EGF ligands. SDS-PAGE and Western blot analysis were used to detect the expression of the EGFP-tagged EGF (EGFP-EGF) protein. The cell-binding and internalization activity of EGFP-EGF were analyzed by fluorescence-activated cell sorting (FACS) and confocal microscopy. EGFP-EGF protein was expressed in Escherichia coli and purified. A cell-binding assay demonstrated that the EGFP-EGF protein could bind efficiently to the cells expressing EGFR. The binding and internalization of EGFP-EGF can be visualized even at a very low concentration under confocal microscopy. The FACS-based assay for internalization activity indicated the accumulation of internalized EGFP-EGF over time. Furthermore, the results of the competition assay indicated its EGFR binding specificity. Using such a method, it does not need to label EGF with chemicals and avoid light in the experimental process. The fusion protein EGFP-EGF has several characters including high sensitivity, stability and convenience for manipulation, and is a powerful tool for the study of EGF endocytosis.

Application of EGFP-EGF fusions to explore mechanism of endocytosis of epidermal growth factor

To develop a simple method for monitoring protein localization of epidermal growth factor (EGF) in living cells. Enhanced green fluorescent protein (EGFP) was used as an autofluorescent tag to label EGF ligands. SDS-PAGE and Western blot analysis were used to detect the expression of the EGFP-tagged EGF (EGFP-EGF) protein. The cell-binding and internalization activity of EGFP-EGF were analyzed by fluorescence-activated cell sorting (FACS) and confocal microscopy. EGFP-EGF protein was expressed in Escherichia coli and purified. A cell-binding assay demonstrated that the EGFP-EGF protein could bind efficiently to the cells expressing EGFR. The binding and internalization of EGFP-EGF can be visualized even at a very low concentration under confocal microscopy. The FACS-based assay for internalization activity indicated the accumulation of internalized EGFP-EGF over time. Furthermore, the results of the competition assay indicated its EGFR binding specificity. Using such a method, it does not need to label EGF with chemicals and avoid light in the experimental process. The fusion protein EGFP-EGF has several characters including high sensitivity, stability and convenience for manipulation, and is a powerful tool for the study of EGF endocytosis.

Expression of mRNA and protein localization of epidermal growth factor and its receptor in goat ovaries

To examine the possibility that epidermal growth factor (EGF) and its receptor (EGF-R) are expressed throughout folliculogenesis, we studied the presence and distribution of EGF and EGF-R in goat ovaries. Ovaries of goats were collected and either fixed in paraformaldehyde for immunohistochemical localization of proteins, or used for the isolation of follicles, luteal cells and ovarian surface epithelium to study mRNA expression for EGF and EGF-R, using the reverse transcriptase polymerase chain reaction. EGF protein and mRNA were found in primordial, primary and secondary follicles as well as in small and large antral follicles and in surface epithelium, but in corpora lutea only the protein could be detected. Antral follicles expressed EGF mRNA in oocyte, cumulus, mural granulosa and theca cells. For EGF-R, both protein and mRNA were present at all stages of follicular development and in all antral follicular compartments. EGF-R protein and mRNA were also found in corpora lutea and surface epithelium. It is concluded that EGF and its receptor are expressed in goat ovarian follicles at all stages of follicle development, in corpora lutea, and in ovarian surface epithelium.

Epidermal growth factor (EGF) expression in lateral line system and in taste buds of adult zebrafish ( Brachidanio rerio)

The mechano and chemosensory organs of adult teleosts undergoes a continuous cell renewal and turnover which is regulated in part by growth factors. Here, we investigated the occurrence and the cell localization of epidermal growth factor (EGF) in the lateral line system and taste bud of adult zebrafish, using Western blot and immunohistochemistry associated to a polyclonal antibody against mammalian EGF. Furthermore, the distribution of S100 protein was studied in parallel to label hair sensory cells in the lateral line system. Western blot revealed one unique protein band with an estimated molecular weight of about 13 kDa, equivalent to the EGF of mammals. Specific immunoreactivity for EGF was observed in the epithelial basal and/or supporting cells of the neuromasts of the lateral line system and taste buds. Conversely, the sensory cells in both sensory structures were devoid of immunostaining. Present results demonstrate the occurrence of EGF in mechano and sensory system of adult zebrafish, suggesting a role for this molecule in the cell renewal and turnover of these structures.

Presence and localization of epidermal growth factor (EGF)- and EGF-receptor-like immunoreactivity in Tetrahymena.

The presence of EGF- and EGF-receptor-like immunoreactivity in Tetrahymena was studied with the help of FITC-labelled monoclonal antibodies, using flow cytometry and confocal microscopy. Tetrahymena has endogeneous EGF and treatment with it significantly increases the hormone content of the cells. The hormone is diffusely localized, particularly in the cytopharynx, where it forms a structure larger than 10 microm. EGF-receptors are also demonstrable, particularly in the cortical region in connection with cilia, and EGF treatment significantly enhances cortical fluorescence. The relationships between these observations and literary data on the effects of EGF in Tetrahymena are discussed.

The role of cyclic AMP response element-binding protein in testosterone-induced differentiation of granular convoluted tubule cells in the rat submandibular gland

The postnatal development of granular convoluted tubules (GCT) in the duct system of the rodent submandibular gland is known to be androgen-dependent, but the underlying molecular mechanism is unclear. To test the possible role of the transcription factor, cyclic AMP response element-binding protein (CREB), in the androgen-induced differentiation of GCT, the effect of testosterone on the expression and localization of epidermal growth factor (EGF), a marker of GCT cells, and of CREB was examined in the submandibular glands of immature 3-week-old rats. Northern blotting demonstrated increases in both EGF and CREB mRNA 1–4 days after testosterone administration. Immunoprecipitation also indicated that CREB protein was increased in amount with testosterone administration, and that induced CREB was phosphorylated at the serine residue as in the active form of CREB. In situ hybridization demonstrated that cells with CREB mRNA signal first appeared in the distal portions of striated ducts at 1 day and had increased in number by 4 days after giving testosterone, when cells with EGF mRNA signal became evident in the same duct portions. Immunohistochemistry also showed the occurrence of CREB protein in the nuclei of duct epithelial cells before their differentiation into EGF-positive GCT cells. Finally, pieces of submandibular gland from immature rats were cultured in vitro and their expression of EGF mRNA analysed by the reverse transcriptase–polymerase chain reaction. Testosterone in the medium caused a marked enhancement of EGF expression in the gland in 1–4 days, which was attenuated by simultaneous administration of the antisense oligonucleotide for CREB as well as that for the androgen receptor. These results suggest the CREB is upregulated by androgen and has a crucial role in androgen-induced differentiation of GCT in the duct system of the rat submandibular gland.

Nuclear localization of epidermal growth factor and epidermal growth factor receptors in human thyroid tissues.

Epidermal growth factor (EGF) has widespread growth effects, and in some tissues proliferation is associated with the nuclear localization of EGF and epidermal growth factor receptor (EGFR). In the thyroid, EGF promotes growth but differs from thyrotropin (TSH) in inhibiting rather than stimulating functional parameters. We have therefore studied the occurrence and cellular distribution of EGF and EGFR in normal thyroid, in Graves' disease, where growth is mediated through the thyrotropin receptor (TSHR), and in a variety of human thyroid tumors. In the normal gland the staining was variable, but largely cytoplasmic, for both EGF and EGFR. In Graves' disease there was strong cytoplasmic staining for both EGF and EGFR, with frequent positive nuclei. Nuclear positivity for EGF and particularly for EGFR was also a feature of both follicular adenomas and follicular carcinomas. Interestingly, nuclear staining was almost absent in papillary carcinomas. These findings document for the first time the presence of nuclear EGF and EGFR in thyroid. Their predominant occurrence in tissues with increased growth (Graves' disease, follicular adenoma, and carcinoma) may indicate that nuclear EGF and EGFR play a role in growth regulation in these conditions. The absence of nuclear EGF and EGFR in papillary carcinomas would suggest that the role played by EGF in growth control differs between papillary carcinoma and follicular adenomas/carcinomas of the thyroid.

Subcellular localization of epidermal growth factor in human parotid gland.

The intracellular distribution of epidermal growth factor was investigated in human parotid gland by immunogold cytochemistry at the electron-microscopy level. Epidermal growth factor immunoreactivity was demonstrated in both acini and ducts. In acinar cells, secretory granules appeared moderately stained, clearly indicating that parotid gland contributes to salivary epidermal growth factor through granule exocytosis. In ductal cells, gold particles were found to decorate numerous cytoplasmic vesicles, particularly abundant in striated duct cells. Since epidermal growth factor reactive vesicles were seen not only at the cellular apex, but nearby lateral plasma membranes as well, it leads to the hypothesis that epidermal growth factor may be discharged both apically into the saliva, and basally into the interstitium.

Immunohistochemical localization of epidermal growth factor in cat paradental tissues during tooth movement

Epidermal growth factor enhances proliferation and differentiation of cells during growth, maturation, and tissue healing. The objectives were to localize the epidermal growth factor in paradental cells and to determine the effect of orthodontic treatment on its concentrations in periodontal ligament fibroblasts, alveolar bone surface lining cells, and epithelial rests of Malassez. Sixty male cats, 1 year old, were divided into 2 groups: active and sham, and further divided into 10 time groups. In the active group, 1 maxillary canine was retracted by 80 g force; in the sham group, the animals received an inactive appliance. Sagittal sections of each half maxilla were stained for epidermal growth factor; staining intensity was measured microphotometrically in 10 periodontal ligament fibroblasts, alveolar bone surface lining cells, and epithelial rests of Malassez cells in sites of periodontal ligament tension and compression, and in corresponding sites near control and sham canines. The overall mean staining intensity of the cells of the active group animals was 30.47%, whereas that of the sham group was 21.78% (P <.0001). In all 3 types, cells near the actively treated canines stained significantly darker (P <.0001) than cells near the sham or control canines, particularly between 12 hours and 7 days. These results demonstrate that orthodontic forces increase epidermal growth factor concentrations in paradental cells, suggesting that epidermal growth factor participates in the tissue remodeling that facilitates tooth movement. (Am J Orthod Dentofacial Orthop 2000;118:210-9)

Subcellular localization of epidermal growth factor in human submandibular gland.

Epidermal growth factor in human submandibular gland was localized at the subcellular level by means of an immunogold staining method. Labelling was observed in serous acini and ducts. In the acini, gold particles were found within secretory granules, indicating that the growth factor is released into the saliva through granule exocytosis. In the ductal system, the most intense reactivity was revealed in the principal cells of striated ducts. In these cells, an abundant population of small cytoplasmic vesicles was specifically stained. Immunoreactive vesicles were found both apically and basally, suggesting that ductal cells can release their products not only into the saliva but also into the interstitium.

Immunohistochemical localisation of epidermal growth factor, transforming growth factor alpha and EGF receptor during organogenesis of the murine hypophysis in vivo.

In previous study of melanocyte-stimulating hormone (MSH) cell development in the proliferating pars intermedia, which is in close apposition to the presumptive pars nervosa, no direct cell-to-cell contact was found between the boundary neurohypophyseal pituicytes (PIC), adenohypophyseal precursor stem cells (PSC) and the related diencephalic mesenchymal cells. Here, we have used immunohistochemistry to examine cytokine expression in the development of the hypophysis during foetal stages II-IV. Light and confocal laser scanning microscopy indicated diffuse expression of both TGF alpha and EGF in the hypophysis at different foetal stages. While no findings indicative for temporary changes of TGF alpha and EGF patterns were found in the foetal hypophysis, a temporary increment of EGF molecules was distinct in the diencephalic mesenchyme at stages III and IV. On the other hand, light microscopy intensively immuno-localized EGFR in the adenohypophysis and neurohypophysis at different developmental stages. Immunoreactivity of EGFR in the cytoplasm and nucleus suggested active proliferative events in the PIC and PSC of stages II-IV mouse pituitaries.

Immunohistochemical localization of epidermal growth factor, transforming growth factor-α and growth factor-βs in the caprine peri-implantation period

Control over the action of steroid hormones in the uterus and conceptus during the initial period of gestation appears to be regulated locally by growth factors. This study involved immunohistochemical detection of epidermal growth factor (EGF), transforming growth factor-α (TGF-α) and transforming growth factor-βs (TGF-βs), to determine their role in the caprine peri-implantation period. Epidermal growth factor was expressed in the luminal and glandular endometrial epithelium of goats on all days studied (Days 22 to 30 post coitum), but it was not detected in trophoblastic cells or in other embryonic structures. Between Days 22 and 30 post coitum, TGF-α was detected in the epithelial cells and superficial stroma of the uterus and in the trophoendodermic cells of the embryo. Transforming growth factor-βs expression, observed in the endometrium, embryo and extraembryonic membranes on Day 22 post coitum, decreased by Day 24 post coitum and disappeared in the embryo by Day 30 post coitum, while remaining in the other structures. The presence of these growth factors during the periimplantation period in the goat suggests their participation in proliferation and differentiation phenomena which occur during implantation and embryonic development.