Amount Of Epidermal Growth Factor Research Articles

Effects of Cream Containing Three-Dimensional Human Adipose-Derived Mesenchymal Stem Cell Conditioned Medium Secreting Growth Factors on Skin Elasticity

Purpose: In this study, the effect on skin elasticity of a cream containing three-dimensional human adipose tissue derived mesenchymal stem cell conditioned medium (3D ADMSC-CM) was examined.Methods: The clinical efficacy of the cream was evaluated by examining the skin for skin lifting, elasticity, and resilience after examining the cell proliferative capacity, expression levels of epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF) in 3D ADMSC-CM, and collagen expression in skin cells due to the 3D ADMSC-CM cream.Results: Skin cell viability was well maintained, with no cytotoxicity from the 3D ADMSC-CM. The amount of EGF and VEGF contained in the 3D ADMSC-CM was 2.692 pg/mL and 27.89 pg/mL, respectively. In addition, skin cells treated with the 3D ADMSC-CM cream experienced a collagen expression increase of 183%. In clinical evaluation of the 3D ADMSC-CM cream, skin lifting and skin elasticity improvement showed significant effects at both two and four weeks. After the 3D ADMSC-CM cream was used, the improved skin elasticity and resilience was analyzed by variables, and an improvement was observed in both after two and four weeks of use.Conclusion: A cream containing 3D ADMSC-CM derived from a 3D culture environment, similar to that of an in vivo environment, is highly effective in enhancing skin elasticity, and 3D ADMSC-CM is expected to lead the field of cosmeceuticals.

Can Free Floating SARS-CoV-2 Viral Material Diminish the Amount of Epidermal Growth Factor and Promote Alzheimer Disease in Women with COVID-19 in Menopause Transition?

The question is raised if there are protein sequences of SARS-CoV-2 that stand-alone can cause neuro-gynecological damage. We found that free floating nucleocapsid can capture EGF and that likely SARS-CoV-2 spike can competitively inhibit TGFβ. Women in menopause transition are vulnerable to lack of EGF-to-EGFR. Therefore, in e.g. aftercare of COVID-19, it is wise to pay attention to cognitive functioning of those women.

The investigation of the amounts and expressions of epidermal growth factor, epidermal growth factor receptor, and epidermal growth factor receptor gene polymorphisms in acne vulgaris.

Epidermal growth factor receptor inhibitors (EGFRI) used in cancer chemotherapy cause acneiform folliculitis in 70%-100% of patients in a dose-dependent manner. Acneiform folliculitis is considered to be caused by an inflammatory process due to follicular hyperkeratosis and subsequently a set of changes both in epidermis and hair follicles as a result of epidermal growth factor receptor (EGFR) blockade. Both acne vulgaris and acneiform folliculitis due to EGFRIs show similar changes in the pilosebaceous unit. Furthermore, in both groups of patients, topical application of recombinant human epidermal growth factor (EGF) has been reported to improve the disease. In this study, it was aimed to investigate the role of EGF and EGFR amount, expression, and EGFR gene polymorphisms in the etiopathogenesis of acne vulgaris. 156 acne vulgaris patients, within 18-25years of age, who had 15 or more inflammatory acne lesions on dermatologic evaluation were included in this study. The absence of any known systemic or genetic disease or cancer and any systemic or topical treatment for the last 1month were prerequisites. In the control group, 154 volunteers in the same age range who were examined at the outpatient clinic with diagnoses of melanocytic nevus, ephelid, cherry angioma, and callus and who had no more than 3 inflammatory acne lesions were recruited. The amounts of EGF and EGFR were determined by sandwich ELISA, expressions of EGF and EGFR by reverse transcriptase polymerase chain reaction; EGFR polymorphisms were examined by restriction enzyme digestion, Sanger, and high-resolution melting methods. The patient and control groups were compared in terms of EGFR gene polymorphisms in addition to the amount and expressions of EGF and EGFR. The amount of EGF in the serum was found to be significantly higher in the acne group. (P=.0012). There was no significant difference in other parameters studied. The results of our study showed a significant increase in the amount of EGF in the acne group. Though EGF may be incriminated in the etiopathogenesis of AV, the most likely explanation about its role may be controlling inflammation from the very first stage.

Interdependence of JAK-STAT and MAPK signaling pathways during EGF-mediated HTR-8/SVneo cell invasion.

Invasion of trophoblast cells is spatio-temporally regulated by various cytokines and growth factors. In pregnancy, complications like preeclampsia, shallow invasion of trophoblast cells and low amounts of epidermal growth factor (EGF) have been reported. In the present study, regulatory mechanisms associated with EGF-mediated invasion in HTR-8/SVneo trophoblastic cells have been delineated. Treatment of HTR-8/SVneo cells with EGF (10 ng/ml) led to eight fold increase (p < 0.05) in invasion. Increased invasion of HTR-8/SVneo cells by EGF was associated with an increase in phosphorylation of ERK½. In addition, significant phosphorylation of STAT1 (ser 727) and STAT3 (both tyr 705 and ser 727 residues) was also observed, accompanied by a decrease in total STAT1. Inhibition of ERK½ phosphorylation by U0126 (10 μM) led to a significant decrease in EGF-mediated invasion with simultaneous decrease in the phosphorylated forms of STAT3 and STAT1. Decrease in total STAT1 was also reversed on inhibition of ERK½. Interestingly, inhibition of STAT3 by siRNA led to a significant decrease in EGF-mediated invasion of HTR-8/SVneo cells and phosphorylation of STAT1, but it did not have any effect on the activation of ERK½. On the other hand, inhibition of STAT1 by siRNA, also led to a significant decrease in the EGF-mediated invasion of HTR-8/SVneo cells, showed concomitant decrease in ERK½ phosphorylation and STAT3 phosphorylation at ser 727 residue. These results suggest cross-communication between ERK½ and JAK-STAT pathways during EGF-mediated increase in invasion of trophoblast cells; phosphorylation at ser 727 residue of both STAT3 and STAT1 appears to be critical.

Absence of TGF-β Receptor Activation by Highly Purified hCG Preparations.

Recently, several LH/human chorionic gonadotropin (hCG) receptor-independent activities for hCG have been described, including activation of the TGF-β receptor (TGFβR) by hyperglycosylated hCG and stimulation of trophoblast invasion. Because the hCG concentrations used in these studies have been rather high, reflecting physiological hCG levels in pregnancy, even a minor contamination with growth factors, which act at very low concentrations, may be significant. Several commercial hCG preparations have been found to contain significant amounts of epidermal growth factor (EGF), which we also confirmed here. Furthermore, we found that some hCG preparations also contain significant amounts of TGF-β1. These hCG preparations were able to activate ERK1/2 in JEG-3 choriocarcinoma cells or TGFβR in mink lung epithelial cells transfected with a reporter gene for TGFβR activation. No such activation was found with highly purified hCG or its free β-subunit (hCGβ), irrespective of whether they were hyperglycosylated or not. Taken together, our results suggest that the growth factor contaminations in the hCG preparations can cause activation of TGFβR and, at least in JEG-3 cells, MAPK signaling. This highlights the importance to carefully control for potential contaminations and that highly purified hCG preparations have to be used for biological studies.

Epidermal growth factor is a potential biomarker for poor cetuximab response in tongue cancer cells.

Head and neck squamous cell carcinoma is frequently associated with aberrant epidermal growth factor receptor (EGFR) signaling, which contributes to tumor growth. Here, the functional importance of EGFR ligands in relation to proliferation and sensitivity to the EGFR-targeted therapy cetuximab was investigated in three tongue cancer cell lines. The influence of epidermal growth factor (EGF), amphiregulin (AR), and epiregulin (EPR) on tumor cell proliferation and cetuximab response was evaluated by the addition of recombinant human (rh) proteins or by siRNA-mediated downregulation of the endogenous ligand production. The expression, activation and cellular distribution of EGFR were assessed by Western blot analysis and immunocytochemistry. EGF downregulation suppressed the proliferation of all investigated tumor cell lines, whereas the response to an increased level of EGF differed between EGFR-overexpressing and EGFR-non-overexpressing cell lines. Furthermore, tumor cells consistently displayed increased cetuximab resistance upon the addition of rhEGF, whereas EGF silencing was associated with an improved cetuximab response. The data regarding AR and EPR were inconclusive. Our data suggest that the amount of EGF is a determinant of the tumor cell proliferation rate and the response to cetuximab treatment in tongue cancer. Thus, EGF is a potential predictive biomarker of poor cetuximab response and a possible treatment target.

Cord blood serum-based eye drops: the impact of donor haematological and obstetric factors on the variability of epidermal growth factor levels.

Cord blood serum (CBS)-based eye drops are successfully used in corneal epithelial wound healing and are prepared to supply a known amount of epidermal growth factor (EGF). Product standardisation includes expensive EGF dosage in all cord blood (CB) units. The influence of donor obstetric and haematological characteristics on EGF content was evaluated, to exclude unsuitable CBS and pre-select those CB units able to provide the correct EGF supply for healing corneal wounds. Data were retrospectively collected from 135 donors included in the Emilia Romagna Cord Blood Bank records. Obstetric characteristics, parity and gestational age of the mother, sex, birth weight and Apgar score of the neonate, placental weight, duration of labour and mode of delivery were considered. Haematological characteristics, CD34+ cell number, and total nucleated cell, white blood cell and platelet counts were recorded. EGF content in CB units was estimated by enzyme-linked immunosorbent assay. Statistical evaluation was performed by Mann-Whitney unpaired and Student's t tests. Correlations between variables were evaluated by using Pearson's (r) or Spearman's (ρ) correlation coefficients. EGF content was significantly higher in CBS from donors aged <30 years and after vaginal deliveries as compared with scheduled Caesarean sections (1,386±580 vs 1,106±391 pg/mL; P=0.002). EGF content was significantly correlated with duration of labour (r=0.45; P=0.0001), number of CD34+ cells/mL (r=0.3; P=0.002) particularly in vaginal deliveries (r=0.36; P=0.003), mother's age (-0.25; P=0.005), neonate's birth weight (r=0.27; P=0.005), and total nucleated cell (r=0.25; P=0.006), white cell (r=0.29; P=0.001) and platelet (r=0.24; P=0.009) counts. No significant correlations were found between EGF content and parity, gestational age, placental weight, neonate's sex or Apgar scores. EGF levels are higher in CB units from younger mothers (<30 years), with longer labour duration (>6 hours), and higher CD34+ cell content (>0.05×10(6)/mL). In order to optimise the preparation and costs of CBS-based eye drops, pre-selection of CB units is recommended.

19 FUNCTIONAL UROTHELIAL DIFFERENTIATION OF URINE-DERIVED STEM CELLS FOR POTENTIAL USE IN URETHRAL TISSUE REPAIR

You have accessJournal of UrologyTrauma/Reconstruction: Traum & Reconstructive Surgery (1)1 Apr 201319 FUNCTIONAL UROTHELIAL DIFFERENTIATION OF URINE-DERIVED STEM CELLS FOR POTENTIAL USE IN URETHRAL TISSUE REPAIR Guihua Liu, Shantaram Bharadwaj, Anthony Atala, and Yuanyuan Zhang Guihua LiuGuihua Liu Winston Salem, NC More articles by this author , Shantaram BharadwajShantaram Bharadwaj Winston Salem, NC More articles by this author , Anthony AtalaAnthony Atala Winston Salem, NC More articles by this author , and Yuanyuan ZhangYuanyuan Zhang Winston Salem, NC More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.1393AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Urothelial cells play important and contradictory roles in the function of the urinary system. They act as a permeability barrier and protect underlying muscle tissue from the caustic effects of urine, which prevent formation of scarring in urethral tissue repair. It is a challenge to induce stem cells to differentiate into urothelial cells with barrier function. Our previous studies demonstrated that urine-derived stem cells (USC) possess self-renewal and multipotent capacity, and can give rise to cells expressing urothelial markers. The goal of this study was to determine whether USC can differentiate into urothelial cells with barrier function in vitro, for potential use in urethral tissue engineering. METHODS Urine-derived cells were harvested from 6 healthy individuals and then induced to differentiate into urothelial cells using epithelial differentiation medium containing epidermal growth factor (EGF) for up to 2 weeks. These differentiated cells were compared to non-differentiated USC. Induced cells were assessed for morphologic changes and expression of urothelial markers via RT-PCR, Western blotting, and immunofluorescent staining. Functional tight junction markers were also assessed by real-time PCR, immunofluorescent stain, and transmission electron microscopy (TEM). Finally, differentiated (4.5×105 cells) and non-induced USC (4.5×105 cells) were seeded on collagen-coated inserts for barrier function assessmenton day 7 and 14. RESULTS Morphologically, urothelial differentiated USC formed cobblestone-like cells. These differentiated USCs expressed transcripts and proteins of urothelial-specific (Uroplakin-III, -Ia) and general epithelial cell markers (CK7, CK13, CK20 and AE1/AE3) in a time-dependent manner. Expression of these markers also depended on the amount of EGF in the media. In an assay of barrier function, urothelial differentiated USC expressed tight junction gene and protein markers (ZO-1 and E-cadherin). TEM demonstrated the ultrastructural aspects of urothelial differentiated USC, including tight junction formation between neighboring cells. Using a fluorescent tracer on differentiated cells cultured on inserts, at least a 50% decrease in leakage of the tracer across the insert occurred after 3h in vitro, indicating that these cells could perform a barrier function similar to the normal urothelial cells. CONCLUSIONS USC can be efficiently differentiated into functional urothelial cells with barrier microstructures, and these cells may be a potential cell source for tissue-engineered urethras. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e7-e8 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Guihua Liu Winston Salem, NC More articles by this author Shantaram Bharadwaj Winston Salem, NC More articles by this author Anthony Atala Winston Salem, NC More articles by this author Yuanyuan Zhang Winston Salem, NC More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Epidermal growth factor-expressing Lactococcus lactis enhances growth performance of early-weaned pigs fed diets devoid of blood plasma

The effect of supplementing Lactococcus lactis (L. lactis) that was engineered to express epidermal growth factor (EGF-LL) to early-weaned pigs fed diets with typical levels of blood plasma (5%) or diets without blood plasma [blood plasma was substituted with soybean (Glycine max) meal and fish meal, based on amino acid supply] was examined. A total of 108 weaned piglets (19-26 d of age; mean initial BW 6.58 kg; 9 pigs per pen) were fed ad libitum according to a 2-phase feeding program without growth promoters. Three pens were assigned to each of 4 treatments: i) blood plasma-containing diet with blank bacterial growth medium (BP-Con), ii) blood plasma-containing diet with fermented EGF-LL (BP-EGF), iii) blood plasma-free diet with blank bacterial growth medium (BPF-Con), and iv) blood plasma-free diet with fermented EGF-LL (BPF-EGF). The amount of epidermal growth factor (EGF) was determined in the fermentation product and pigs were allotted 60 μg EGF/kg BW/d for 3 wk postweaning. There were no differences in overall growth performance between BP-Con and BP-EGF pigs and no differences in overall growth performance between LoCon and BPF-EGF pigs. Pigs fed BPF-EGF showed increased daily BW gain (410 vs. 260 g/d; P < 0.01) and gain:feed (0.67 vs. 0.58; P < 0.05) compared to BPF-Con pigs in wk 3 postweaning; this was comparable to values for the BP-Con group (400 g/d and 0.64). These results indicate that supplementation with EGF-LL can be effective in enhancing the performance of early-weaned piglets fed a low complexity diet and reduces the need for feeding high-quality animal proteins and antibiotics.

Matrix stiffening sensitizes epithelial cells to EGF and enables the loss of contact inhibition of proliferation

Anchorage to a compliant extracellular matrix (ECM) and contact with neighboring cells impose important constraints on the proliferation of epithelial cells. How anchorage and contact dependence are inter-related and how cells weigh these adhesive cues alongside soluble growth factors to make a net cell cycle decision remain unclear. Here, we show that a moderate 4.5-fold stiffening of the matrix reduces the threshold amount of epidermal growth factor (EGF) needed to over-ride contact inhibition by over 100-fold. At EGF doses in the range of the dissociation constant (K(d)) for ligand binding, epithelial cells on soft matrices are contact inhibited with DNA synthesis restricted to the periphery of cell clusters. By contrast, on stiff substrates, even EGF doses at sub-K(d) levels over-ride contact inhibition, leading to proliferation throughout the cluster. Thus, matrix stiffening significantly sensitizes cells to EGF, enabling contact-independent spatially uniform proliferation. Contact inhibition on soft substrates requires E-cadherin, and the loss of contact inhibition upon matrix stiffening is accompanied by the disruption of cell-cell contacts, changes in the localization of the EGF receptor and ZO-1, and selective attenuation of ERK, but not Akt, signaling. We propose a quantitative framework for the epigenetic priming (via ECM stiffening) of a classical oncogenic pathway (EGF) with implications for the regulation of tissue growth during morphogenesis and cancer progression.

Sialoadenectomy enhances hepatic injury induced by lipopolysaccharide/galactosamine in mice

Submandibular salivary glands (SMGs) synthesize, accumulate and secrete a large amount of epidermal growth factor (EGF) in mice. It is known that surgical removal of SMG (sialoadenectomy) alters cell turnover in the liver and exacerbates liver injury induced by lipopolysaccharide/galactosamine (LPS/GalN). Here we show that such increased hepatotoxicity is not the consequence of the lack of EGF production from SMG. On the contrary, it appears to be the consequence of an inadequate cytokine production by the liver of sialoadenectomized mice. Thus, we found that the increase of plasma tumour necrosis factor-alpha and interleukin-6 was slower in sialoadenectomized than in sham-operated mice. This is because of a decreased rate of production of both cytokines by the liver. We found that the increase of plasma corticosterone (CS) concentration is lower in sialoadenectomized than that in sham-operated mice. Adrenalectomy exacerbated liver injury induced by LPS/GalN. In these animals, sialoadenectomy did not further increase the effect of LPS/GalN. Our results suggest that the effect of sialoadenectomy on LPS/GalN-induced liver toxicity may be the consequence of an altered cytokine production by the liver and a reduced CS release from adrenal glands.

A convenient and adjustable surface-modified complex containing poly-L-glutamic acid conjugates as a vector for gene delivery

In order to quantify the amount of ligands or poly(ethylene glycol) (PEG) on each vector, here we developed a system in which poly-L-glutamic acid (PLG) was used as surface modification loading backbone, to which one PEG (MW 5000, 10000, 20000) or epidermal growth factor (EGF) was linked. The PLG conjugates can electro-statically adsorb upon DNA/ polycation complex with positive charge, and, the amount of EGF or PEG on the surface of complexes could be varied. We have made a series of complexes containing the various PLG conjugates and examined their physicochemical properties, and made a comparison of properties and transfection efficiency between these complexes. EGF- and PEG-modified complexes showed 10-25-folds higher cell transfection efficiency than unmodified complexes in medium with or without serum.

The Effects of Sialoadenectomy and Epidermal Growth Factor on Gingival Tissue in Rats: An Ultrastructural Study

ABSTRACTSubmandibular salivary glands (SMGs) synthesize, accumulate and secrete a large amount of epidermal growth factor (EGF) in rats. EGF stimulates cell proliferation and differentiation by binding to its reseptors (EGFR).The aim of the present study was to mimic an endogenous epidermal growth factor deficiency through sialoadenectomy in paralel to exogenous EGF administration and to observe the ultrastructural changes in rat gingival tissue, employing electron microscopy.Thirty adult female Wistar albino rats were divided randomly into three groups: a control group (n:10), a sialoadenectomy group (n:10) and a sialoadenectomy group to which an EGF was administered (n:10).The experimental groups of rats were subjected to sialoadenectomy in order to create EGF deficiency. After 27 days both control and experimental rat groups were euthanized by pentobarbital and their gingival tissue was removed. Tissue samples from gingiva were processed for ultrastructural study.Electron microscopic evaluation indicated that while in the control group gingival tissues showed a normal appearance changes were observed in the experimental groups. We observed a partial degeneration of the epithelial cell junctions. Widespread crystolisis was also observed in a group of mitochondria.It is concluded that epidermal growth factor deficiency achieved by sialoadenectomy caused ultrastructural changes in gingival epithelium.

Effect of electro-acupuncture at Foot-Yangming Meridian on somatostatin and expression of somatostatin receptor genes in rabbits with gastric ulcer

To discuss the protective effect of electroacupuncture at the Foot-Yangming Meridian on gastric mucosal lesion, somatostatin (SS) and the expression of SS receptor genes (SSR(1)mRNA ) in rabbits with gastric ulcer and to further explore the relative specificity of meridians and viscera at gene expression level. Forty rabbits were randomly divided into control group (A), gastric ulcer model group (B), Foot-Yangming Meridian group (C), Foot-Shaoyang Meridian group (D) and Foot-Taiyang Meridian group (E). The gastric ulcer model was prepared by infusing alcohol into stomach. Groups C-E were treated with electro-acupuncture at points along the above meridians using meridian stimulating instruments for 7 d respectively. By the end of treatment, the index of gastric ulcer was determined, the amount of epidermal growth factor(EGF) and somatostatin was measured by radioimmunoassay (RIA). SS-R(1)mRNA expression in gastric mucosa was determined by RT-PCR. The value of EGF in model group was obviously lower (73.6+/-14.8 vs 91.3+/-14.9 pg/mL, P<0.01) than that in control group. The index of gastric ulcer, content of SS and expression of SSR1mRNA in gastric mucosa were significantly higher than those in control group (24.88+/-6.29 vs 8.50+/-2.98 scores, P<0.01; 2978.6+/-587.6 vs 1852.4+/-361.7 mIU/mL, P<0.01; 2.56+/-0.25 vs 1.04+/-0.36, P<0.01). The value of EGF in Foot-Yangming Meridian group was higher than that in model group (92.2+/-6.7 vs 73.6+/-14.8 pg/mL, P<0.01). The index of gastric ulcer, content of SS and expression of SS-R(1)mRNA in gastric mucosa were significantly lower than those in control group (10.88+/-3.23 vs 24.88+/-6.29 scores, P<0.01; 1800.2+/-488 vs 2978.6+/-587.6 mIU/mL, P<0.01; 1.07+/-0.08 vs 2.56+/-0.25 mIU/mL, P<0.01). Compared to the model group, the content of SS and expression of SSR1mRNA in gastric mucosa in Foot-Shaoyang Meridian group decreased (2441.0+/-488.vs 2978.6+/-587.6 mIU/mL, P<0.05;1.73+/-0.16 vs 2.56+/-0.25 mIU/mL, P<0.01). But the above parameters in Foot-Taiyang Meridian group did not improve and were significantly different from those in Foot-Yangming Meridian group (P<0.05). Electro-acupuncture at Foot-Yangming Meridian can protect gastric mucosa against injury. The mechanism may be related to the regulation of brain-gut peptides and the expression of SSR(1)mRNA.

The Inhibitory Effect of ErbB2 on Epidermal Growth Factor-induced Formation of Clathrin-coated Pits Correlates with Retention of Epidermal Growth Factor Receptor-ErbB2 Oligomeric Complexes at the Plasma Membrane

By constructing stably transfected cells harboring the same amount of epidermal growth factor (EGF) receptor (EGFR), but with increasing overexpression of ErbB2, we have demonstrated that ErbB2 efficiently inhibits internalization of ligand-bound EGFR. Apparently, ErbB2 inhibits internalization of EGF-bound EGFR by constitutively driving EGFR-ErbB2 hetero/oligomerization. We have demonstrated that ErbB2 does not inhibit phosphorylation or ubiquitination of the EGFR. Our data further indicate that the endocytosis deficiency of ErbB2 and of EGFR-ErbB2 heterodimers/oligomers cannot be explained by anchoring of ErbB2 to PDZ-containing proteins such as Erbin. Instead, we demonstrate that in contrast to EGFR homodimers, which are capable of inducing new clathrin-coated pits in serum-starved cells upon incubation with EGF, clathrin-coated pits are not induced upon activation of EGFR-ErbB2 heterodimers/oligomers.

Differentiation Status-dependent Regulation of Cyclooxygenase-2 Expression and Prostaglandin E2 Production by Epidermal Growth Factor via Mitogen-activated Protein Kinase in Articular Chondrocytes

Although large amounts of epidermal growth factor (EGF) are found in the synovial fluids of arthritic cartilage, the role of EGF in arthritis is not clearly understood. This study investigated the effect of EGF on differentiation and on inflammatory responses such as cyclooxygenase-2 (COX-2) expression and prostaglandin E(2) (PGE(2)) production in articular chondrocytes. EGF caused a loss of differentiated chondrocyte phenotype as demonstrated by inhibition of type II collagen expression and proteoglycan synthesis. EGF also induced COX-2 expression and PGE(2) production. EGF-induced dedifferentiation was caused by EGF receptor-mediated activation of extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) but not p38 kinase, whereas the activation of both ERK1/2 and p38 kinase was necessary for COX-2 expression and PGE(2) production. Neither the inhibition of COX-2 expression and PGE(2) production nor the addition of exogenous PGE(2) affected EGF-induced dedifferentiation. However, COX-2 expression and PGE(2) production were significantly enhanced in chondrocytes that were dedifferentiated by serial subculture, and EGF also potentiated COX-2 expression and PGE(2) production, although these cells were less sensitive to EGF. Dedifferentiation-induced COX-2 expression and PGE(2) production were mediated by ERK1/2 and p38 kinase signaling. Our results indicate that EGF in articular chondrocytes stimulates COX-2 expression and PGE(2) production via ERK and p38 kinase signaling in association with differentiation status.

Ca2+ Pools and Cell Growth: EVIDENCE FOR SARCOENDOPLASMIC Ca2+-ATPases 2B INVOLVEMENT IN HUMAN PROSTATE CANCER CELL GROWTH CONTROL

The present study demonstrates for the first time that intracellular calcium-ATPases and calcium pool content are closely associated with prostate cancer LNCaP cell growth. Cell growth was modulated by changing the amount of epidermal growth factor, serum, and androgene in culture media. Using the microspectrofluorimetric method with Fura-2 and Mag Fura-2 as probes, we show that in these cells, the growth rate is correlated with intracellular calcium pool content. Indeed, an increased growth rate is correlated with an increase in the calcium pool filling state, whereas growth-inhibited cells show a reduced calcium pool load. Using Western blotting and immunocytochemistry, we show that endoplasmic reticulum calcium pump expression is closely linked to LNCaP cell growth, and are a common target of physiological stimuli that control cell growth. Moreover, we clearly demonstrate that inhibition of these pumps, using thapsigargin, inhibits LNCaP cell growth and prevents growth factor from stimulating cell proliferation. Our results thus provide evidence for the essential role of functional endoplasmic reticulum calcium pumps and calcium pool in control of prostate cancer LNCaP cell growth, raising the prospect of new targets for the treatment of prostate cancer.

Involvement of ErbB-2 in rheumatoid synovial cell growth.

The synovial tissue affected by rheumatoid arthritis (RA) is characterized by hyperproliferation of synovial cells. High amounts of epidermal growth factor (EGF) in the synovial fluid of RA patients contribute to the growth of rheumatoid synovial cells. To characterize the receptor for EGF in rheumatoid synovial cells, the expression and function of ErbB family members were examined. Synovial tissues were obtained from surgical excisions. The expression of ErbB products was examined by immunohistochemistry and immunoblotting by using specific antibodies. Primary cultures were established from the surgical materials. Cell growth was measured using MTT. The levels and phosphorylation state of the ErbB-2 protein were analyzed by immunoprecipitation and immunoblotting. The expression of ErbB-2, but not other ErbB-related products, was detected in synovium with RA as compared with that with osteoarthritis (OA) and ligament injury. Growth of primary synovial cells with RA was inhibited by genistein, a tyrosine kinase inhibitor, and herceptin, a specific monoclonal antibody against ErbB-2. Herceptin showed a small effect on growth of primary synovial cells with OA. EGF stimulated the phosphorylation of ErbB-2 in primary synovial cells with RA. This EGF-stimulated phosphorylation was completely abrogated by genistein and herceptin. ErbB-2 is expressed in rheumatoid synovial cells and may function as the receptor for EGF. Our data suggest that mitotic signals from EGF family members are transduced by ErbB-2 in these cells. Inhibition of ErbB-2 may provide a new approach to the effective treatment for RA.

Comparative image analysis of EGF immunoreaction in rat submandibular gland using 3,3'-diaminobenzidine with metal enhancer substrate

We have shown the efficacy of image analysis using 3,3'-diaminobenzidine (DAB) with a metal enhancer substrate for demonstrating quantitative differences in the amount of epidermal growth factor (EGF) in the submandibular gland from normal, castrated, and testosterone propionate (TP) treated castrated rats. Immunohistochemical determination of EGF visualized by DAB-nickel reagent was performed with image analysis using a computed image analyzer system (ACAS 570). Immunohistochemistry for EGF disclosed positive staining in granular convoluted tubule cells in the tissue sections from each experimental group. Using the tools of a competent data program installed in the ACAS 570 software, we measured quantitative differences among the experimental glands examined. Castration was shown to elicit a significant reduction in the EGF-positive area and staining intensity, and administration of TP to the castrated animals restored these parameters to levels greater than those of normal rats. Our study demonstrates that a simple, inexpensive, commercially available metal enhancer substrate can be applied accurately to the computer assisted quantification of histochemical hormone-induction studies.

ＥＧＦ（Ｅｐｉｄｅｒｍａｌ Ｇｒｏｗｔｈ Ｆａｃｔｏｒ）オートクリン作用によるヒト口腔へん平上皮癌細胞の運動能促進

We studied whether epidermal growth factor (EGF) enhances the motility of human oral squamous cell carcinoma (SCC) cells in an autocrine manner. The motility of five SCC cell lines (SAS, Ca 9-22, HSC-2, -3, -4) was examined with the use of a phagokinetic track assay. The motility of SAS cells and HSC-2 cells was significantly increased without EGF stimulation as compared with the three other cell lines. Amounts of EGF in the culture supernatants of the five cell lines after 48h of culture were examined with the immunoblotting method. EGF was detected in the supernatants of SAS cells and HSC-2 cells (SAS, 8.65±0.45; HSC-2, 2.60±1.13ng/ml). SAS cells, showing the highest levels of motility and EGF production, was used for further study. When SAS cells were treated with anti-EGF antibody, anti-EGF receptor antibody, and erbstatin analog, the motility of SAS cells was significantly inhibited. Furthermore, the culture supernatant of SAS cells enhanced the motility of HSC-3 cells in a concentration-dependent fashion. These results suggest that EGF may stimulate the motility of human oral SCC cells in an autocrine manner.