Expression Of Betacellulin Research Articles

Biomimetic Hydrogels Promote Pseudoislet Formation to Improve Glycemic Control in Diabetic Mice.

Islet or β-cell transplantation is currently considered to be the ideal treatment for diabetes, and three-dimensional (3D) bioprinting of a bionic pancreas with physiological stiffness is considered to be promising for the encapsulation and transplantation of β-cells. In this study, a 5%GelMA/2%AlgMA hybrid hydrogel with pancreatic physiological stiffness was constructed and used for β-cell encapsulation, 3D bioprinting, and in vivo transplantation to evaluate glycemic control in diabetic mice. The hybrid hydrogel had good cytocompatibility and could induce insulin-producing cells (IPCs) to form pseudoislet structures and improve insulin secretion. Furthermore, we validated the importance of betacellulin (BTC) in IPCs differentiation and confirmed that IPCs self-regulation was achieved by altering the nuclear and cytoplasmic distributions of BTC expression. In vivo transplantation of diabetic mice quickly restored blood glucose levels. In the future, 3D bioprinting of β-cells using biomimetic hydrogels will provide a promising platform for clinical islet transplantation for the treatment of diabetes.

Experimental and Clinical Evidence Suggests That Treatment with Betacellulin Can Alleviate Th2-Type Cytokine-Mediated Impairment of Skin Barrier Function.

Betacellulin (BTC) is a peptide ligand that belongs to the epidermal growth factor family, the members of which have been implicated in skin morphogenesis, homeostasis, repair, and angiogenesis; however, the role of BTC in the regulation of the skin barrier remains unknown. To examine the role of BTC in skin barrier function, we analyzed atopic dermatitis (AD) transcriptomic data from Gene Expression Omnibus (GEO) datasets, performed BTC immunohistochemistry using human skin tissues, and evaluated the effects of BTC on primary human keratinocytes by real-time PCR, Western blotting, and assay of the transepidermal electrical resistance (TER), a functional parameter to monitor the tight junction barrier. We found that the gene expression of BTC was downregulated in skin lesions from patients with AD, and this downregulated expression recovered following biological treatments. Consistently, the BTC protein levels were downregulated in the lesional skin of AD patients compared with the normal skin of healthy participants, suggesting that the BTC levels in skin might be a biomarker for the diagnosis and therapy of AD. Furthermore, in human keratinocytes, BTC knockdown reduced the levels of skin-derived antimicrobial peptides and skin barrier-related genes, whereas BTC addition enhanced their levels. Importantly, in human skin equivalents, BTC restored the increased tight junction permeability induced by Th2 cytokine IL-4/IL-13 treatment. In addition, specific inhibitors of epidermal growth factor receptor (EGFR) and protein kinase C (PKC) abolished the BTC-mediated improvement in skin barrier-related proteins in keratinocyte monolayers. Collectively, our findings suggest that treatment with BTC might improve the Th2-type cytokine-mediated impairment of skin barrier function through the EGFR/PKC axis and that BTC might be a novel potential biomarker and therapeutic target for the treatment of skin conditions characterized by the overproduction of Th2 cytokines and dysfunctional skin barriers, such as AD.

BTC as a Novel Biomarker Contributing to EMT via the PI3K-AKT Pathway in OSCC.

Purpose: Oral squamous cell carcinoma (OSCC) is one of the most common malignant tumors of the head and neck, while metastasis is the main cause of OSCC-related death. There is an urgent need to explore novel prognostic biomarkers and identify biological targets related to metastasis in OSCC treatment. Methods: Analysis of differential expression was performed using datasets in The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO). Immunohistochemistry (IHC) was conducted to assess the expression of betacellulin (BTC) in OSCC. SCC4 and CAL27 cells were used for in vitro experiments, in which CCK-8, transwell assays, and wounding healing assays were performed to verify the biological functions of BTC. The role of BTC in EMT was analyzed by EMT score and Western blot. Results: Through the analysis of the mRNA expression profile data from TCGA database in OSCC, we found that only low expression of BTC was significantly correlated with a poor prognosis in OSCC patients. The results of IHC assays and TCGA databases showed that the expression level of BTC was related to the tumor stage, histological grade, and metastasis status. In vitro analysis showed that overexpression of BTC significantly suppressed the proliferation and migration of OSCC cells. Furthermore, we confirmed that BTC could affect EMT through the PI3K-AKT signaling pathway. Conclusion: The overexpression of BTC suppresses the proliferation, migration, and EMT of OSCC cells via the PI3K-AKT pathways, leading to a better prognosis in OSCC. BTC may be used as a novel molecular marker to assess the prognosis of OSCC patients.

Hepatocyte Growth Factor and Betacellulin Gene Expression for Treating Diabetes: In vitro analysis

Background: Diabetes mellitus is a fast spreading metabolic disease in both developed and developing countries owing to genetic predisposition, lifestyle and or environmental conditions. Recombinant insulin used for managing diabetes has been approved in 1985, but till now there has been no cure. Insulin cannot help the patients who become resistant to it. Islet transplantation has been developed in 2001 and can become a possible cure for Diabetes. Gene therapy can be beneficial in ameliorating the functionality and survival of islets post-transplantation. Plasmid vectors pcDNA3.1 BTC and pcDNA3.1 HGF can help with proliferation and protection of islet cells from apoptosis. Methods: Plasmid vectors encoding genes Betacellulin (BTC) and Hepatocyte growth factor (HGF) were constructed and the gene expression, caspase -3 and cell viability assays were done to assess the activity of these vectors. Results: pcDNA3.1 BTC and pcDNA3.1 HGF showed time dependent increase in expression of BTC and HGF following transfection in rat insulinoma (RIN-5mF) cells by forming complexes with lipofectamine-3000. There was no toxicity associated with the use of lipofectamine-3000 and the expression of BTC and HGF seemed to protect them from cytokine mediated apoptosis. Conclusion: The use of viral vectors is fraught with challenges especially with regards to their safety. Hence, we used plasmid vectors to express therapeutic genes Betacellulin and Hepatocyte growth factor and the use of these vectors on RIN-m5F cells showed that they could protect them from apoptosis mediated cell death and can enhance their function and survival.

Unraveling ERBB network dynamics upon betacellulin signaling in pancreatic ductal adenocarcinoma in mice.

Pancreatic ductal adenocarcinoma (PDAC) will soon belong to the top three cancer killers. The only approved specific PDAC therapy targets the epidermal growth factor receptor (EGFR). Although EGFR is a crucial player in PDAC development, EGFR‐based therapy is disappointing. In this study, we evaluated the role of the EGFR ligand betacellulin (BTC) in PDAC. The expression of BTC was investigated in human pancreatic cancer specimen. Then, we generated a BTC knockout mouse model by CRISPR/Cas9 technology and a BTC overexpression model. Both models were crossed with the Ptf1aCre/+;KRASG12D/+ (KC) mouse model (B−/−KC or BKC, respectively). In addition, EGFR, ERBB2, and ERBB4 were investigated by the pancreas‐specific deletion of each receptor using the Cre‐loxP system. Tumor initiation and progression were analyzed in all mouse lines, and the underlying molecular biology of PDAC was investigated at different time points. BTC is expressed in human and murine PDAC. B−/−KC mice showed a decelerated PDAC progression, associated with decreased EGFR activation. BKC mice developed severe PDAC with a poor survival rate. The dramatically increased BTC‐mediated tumor burden was EGFR‐dependent, but also ERBB4 and ERBB2 were involved in PDAC development or progression, as depletion of EGFR, ERBB2, or ERBB4 significantly improved the survival rate of BTC‐mediated PDAC. BTC increases PDAC tumor burden dramatically by enhanced RAS activation. EGFR signaling, ERBB2 signaling, and ERBB4 signaling are involved in accelerated PDAC development mediated by BTC indicating that targeting the whole ERBB family, instead of a single receptor, is a promising strategy for the development of future PDAC therapies.

Betacellulin regulates the proliferation and differentiation of retinal progenitor cells in vitro.

Retinal progenitor cells (RPCs) hold great potential for the treatment of retinal degenerative diseases. However, their proliferation capacity and differentiation potential towards specific retinal neurons are limited, which limit their future clinical applications. Thus, it is important to improve the RPCs’ ability to proliferate and differentiate. Currently, epidermal growth factor (EGF) is commonly used to stimulate RPC growth in vitro. In this study, we find that betacellulin (BTC), a member of the EGF family, plays important roles in the proliferation and differentiation of RPCs. Our results showed that BTC can significantly promote the proliferation of RPCs more efficiently than EGF. EGF stimulated RPC proliferation through the EGFR/ErbB2‐Erk pathway, while BTC stimulated RPC proliferation more powerfully through the EGFR/ErbB2/ErbB4‐Akt/Erk pathway. Meanwhile, under differentiated conditions, the BTC‐pre‐treated RPCs were preferentially differentiated into retinal neurons, including photoreceptors, one of the most important types of cells for retinal cell replacement therapy, compared to the EGF‐pre‐treated RPCs. In addition, knockdown of endogenous BTC expression can also obviously promote RPC differentiation into retinal neuronal cells. This data demonstrate that BTC plays important roles in promoting RPC proliferation and differentiation into retinal neurons. This study may provide new insights into the study of RPC proliferation and differentiation and make a step towards the application of RPCs in the treatment of retinal degenerative diseases.

The impact of co-expression of wild-type EGFR and its ligands determined by immunohistochemistry for response to treatment with cetuximab in patients with metastatic colorectal cancer.

Anti-EGFR mAbs cetuximab and panitumumab are routinely used for the treatment of patients with KRAS-wild type metastatic colorectal cancer (mCRC). However, in some patients their efficacy remains modest and with no clear association between the EGFR protein expression determined by PharmDx™ kit, and response to anti-EGFR therapies. Therefore, we investigated the relative expression and predictive value of wild-type EGFR (wtEGFR), mutated EGFRvIII and EGFR ligand proteins in mCRC patients treated with cetuximab. The expression levels of wtEGFR, EGFRvIII, and EGFR ligand were determined by immunohistochemistry (IHC) in 60 tumour specimens using specific antibodies. Sections were scored according to the percentage of positive tumour cells, intensity and cellular location of staining, and these were associated with response, overall survival (OS) and progression-free survival (PFS). At cut-off value > 5%, wtEGFR, and EGFRvIII were present in 44%, and 41%, betacellulin (BTC) in 72%, followed by epigen (67%), TGFα (58%), amphiregulin (34%), EGF (31%) of the cases, respectively and 96% of the wtEGFR positive cases had co-expression of at least one ligand. We found a significant association between the expression of wtEGFR and poor response to cetuximab. In addition, the co-expression of wtEGFR with one ligand at a cut-off value of > 5% and > 10% was associated with worse response to cetuximab (P = 0.021, and P = 0.005 respectively). We found a 3-fold and 5-fold increased risk of shorter OS with expression of BTC and epigen. Interestingly, the expression of wtEGFR and its co-expression with one or two ligands was associated with shorter PFS but not with OS. The relative expression of wtEGFR and its competing ligands, which is the target for therapeutic interventions with anti-EGFR antibodies, could serve as a more reliable predictive biomarker of response to therapy with anti-EGFR mAbs in mCRC patients and warrants further investigation in large prospective studies.

Betacellulin in Chronic Periodontitis Patients With and Without Type 2 Diabetes Mellitus: An Immunohistochemical Study.

The host immune response to bacterial dental plaque determines periodontal disease susceptibility by increasing the secretion of inflammatory cytokines. The Epidermal Growth Factor family cytokines stimulate proliferation and keratinization of cells in dermis and oral epithelium. Epidermal Growth Factor family consists of Amphiregulin, Betacellulin, Epiregulin, Epigen, Heparin binding Epidermal Growth Factor like growth factor and transforming Growth Factor-alpha. The current study aimed to investigate expression of Betacellulin in chronic periodontitis patients with and without type 2 diabetes mellitus and thereby assessing role of betacellulin in periodontal health and disease. Present study comprised of 90 participants, age ranges from 18 to 60-year-old, for the period of March 2010 to May 2011. Participants were categorized into three groups based Gingival index (GI), probing depth (PD) and clinical attachment loss (CA Loss). Group 1 consisted 30 individuals with clinically healthy periodontium, Group-2 consisted 30 individuals with GI>1, PD≥5 mm, and CA Loss>3 mm. Group-3 (Chronic Periodontitis with type 2 diabetes mellitus) consisted 30 with GI >1, PD≥5 mm, and CA Loss>3 mm. Immunohistochemical localization and quantification of Betacellulin was done in gingival tissue samples from all groups. Data showed expression of Betacellulin were higher in chronic periodontitis as compared to healthy. A positive correlation found in Betacellulin expression and Probing Depth in chronic periodontitis. This footmark study impacts the role of Betacellulin in pathogenesis and progression of periodontal disease which will help in exploration of novel immunotherapeutic strategies and immunological research activity in this field.

Genetic deletion of the EGFR ligand epigen does not affect mouse embryonic development and tissue homeostasis

The epidermal growth factor receptor (EGFR) is a tyrosine kinase receptor with manifold functions during development, tissue homeostasis and disease. EGFR activation, the formation of homodimers or heterodimers (with the related ERBB2-4 receptors) and downstream signaling is initiated by the binding of a family of structurally related growth factors, the EGFR ligands. Genetic deletion experiments clarified the biological function of all family members except for the last characterized ligand, epigen. We employed gene targeting in mouse embryonic stem cells to generate mice lacking epigen expression. Loss of epigen did not affect mouse development, fertility, or organ physiology. Quantitative RT-PCR analysis revealed increased expression of betacellulin and EGF in a few organs of epigen-deficient mice, suggesting a functional compensation by these ligands. In conclusion, we completed the genetic analysis of EGFR ligands and show that epigen has non-essential functions or functions that can be compensated by other EGFR ligands during growth and tissue homeostasis.

High Cortical Bone Mass Phenotype in Betacellulin Transgenic Mice Is EGFR Dependent

Signaling through the epidermal growth factor receptor (EGFR) by ligands such as epidermal growth factor (EGF), transforming growth factor alpha (TGFA), and amphiregulin (AREG) has been reported to have effects on skeletal growth. The role of betacellulin (BTC), another EGFR ligand, in skeletal development and bone metabolism is unknown. In previous experiments, transgenic mice overexpressing BTC ubiquitously under the control of the chicken beta-actin promoter (BTC-tg) exhibited stunted growth and disproportionately sized long bones. In this study, we performed a detailed phenotypic analysis of BTC-tg mice at 3, 6, and 9 wk of age. Osteoblastic cells from transgenic mice showed strong expression of BTC as determined by Western blots and by immunohistochemistry on bone sections. In femurs of male and female BTC-tg mice, we found reduced longitudinal bone growth and a pronounced increase in total volumetric BMD. The increased femoral BMD was mainly caused by augmented endocortical bone apposition and subsequent cortical bone thickening. In contrast, vertebral BMD was reduced in BTC-tg mice of both sexes. An overall similar phenotype was found in 6-mo-old BTC-tg mice. The increase in cortical bone mass in the appendicular skeleton of BTC-tg mice was largely blocked when they were crossed into the Egfr (Wa5) background characterized by a dominant negative EGFR. Our study showed that overexpression of BTC results in an EGFR-dependent upregulation of cortical bone mass in the appendicular skeleton of mice, uncovering a potential novel anabolic pathway for cortical bone.

Betacellulin overexpression in transgenic mice improves glucose tolerance and enhances insulin secretion by isolated islets in vitro

Betacellulin (BTC), a ligand of the epidermal growth factor receptor, has been shown to promote growth and differentiation of pancreatic beta-cells and to improve glucose metabolism in experimental diabetic rodent models. We employed transgenic mice (BTC-tg) to investigate the effects of long-term BTC overabundance on islet structure and glucose metabolism. Expression of BTC is increased in transgenic islets, which show normal structure and distribution of the different endocrine cell types, without pathological alterations. BTC-tg mice exhibit lower fasted glucose levels and improved glucose tolerance associated with increased glucose-induced insulin secretion. Surprisingly, quantitative stereological analyses revealed that, in spite of increased cell proliferation, the islet and beta-cell volumes were unchanged in BTC-tg mice, suggesting enhanced cell turnover. Insulin secretion in vitro was significantly higher in transgenic islets in medium containing high glucose (11.2 or 16.7mM) as compared to control islets. Our results demonstrate that long-term BTC overabundance does not alter pancreatic islet structure and beta-cell mass, but enhances glucose-induced insulin secretion in vivo as well as in vitro.

Increased expression of epidermal growth factor receptor and betacellulin during the early stage of gastric ulcer healing

Epidermal growth factor receptor (EGFR) is important for the proliferation and differentiation of gastric mucosal cells. Betacellulin (BTC) is a novel ligand for EGFR Since their role is unclear in the ulcer healing process, we investigated their expression. Gastric ulcers in 30 Sprague-Dawley rats were induced by acetic acid. RT-PCR and Western blotting were performed to detect EGFR and BTC. Immunohistochemical studies were performed to detect EGFR, BTC and proliferating cell nuclear antigen (PCNA). The expression of EGFR and the BTC gene was significantly increased at 12 h, 24 h and 3 days after ulcer induction (P<0.05). The expression of EGFR and BTC protein was significantly increased at 24 h and 3 days and at 12 h, 24 h and 3 days after ulcer induction, respectively (P<0.05). The phosphorylation of EGFR also increased significantly and reached a maximum 24 h after ulcer induction (P<0.05). Immunostaining for EGFR and BTC was observed in numerous epithelial cells from the ulcer margin and ulcer bed, and corresponded to the localization of PCNA. To conclude, there was an increase in EGFR and BTC expression in the early stages of ulcer healing, localized in the epithelial cells of the ulcer margins and regenerating glands with proliferating activity. BTC/EGFR may play an important role in the early stages of ulcer healing.

Remission of Diabetes by β-Cell Regeneration in Diabetic Mice Treated With a Recombinant Adenovirus Expressing Betacellulin

Type 1 diabetes results from destruction of the majority of the pancreatic β cells by β cell-specific autoimmune responses; therefore, expansion of the β-cell mass in vivo Is a possible approach to its treatment. Betacellulin (BTC) is known to promote β-cell growth and differentiation. We investigated whether transient, constitutive expression, and secretion of BTC would regenerate sufficient numbers of pancreatic β cells to restore normoglycemia in diabetic animals. We constructed a recombinant adenoviral vector (rAd-BTC) containing the cytomegalovirus promoter/enhancer, β-globin chimeric intron, and albumin leader sequence to facilitate secretion, followed by BTC (1-80) complementary DNA (cDNA) encoding mature BTC. A single intravenous (IV) administration of rAd-BTC resulted in complete remission of streptozotocin (STZ)-induced diabetes within 2 weeks in mice. The mice remained normoglycemic for >100 days; glucose tolerance tests showed kinetics similar to normal, nondiabetic mice. Pancreatic insulin content, β-cell mass, and serum insulin levels in rAd-BTC-treated mice were significantly higher than in the controls. Treatment of autoimmune diabetic mice with rAd-BTC in combination with an immune suppressor resulted in remission of diabetes. We conclude that transient expression of BTC by rAd-BTC administration results in prolonged remission of diabetes in mice, by the regeneration of sufficient numbers of β cells in the pancreas.

Reversal of streptozotocin-induced hyperglycemia by continuous supply of betacellulin in mice

Previous studies have shown the efficacy of betacellulin (BTC) to promote β-cell regeneration. Because of its short half-life, however, the effect of BTC may have been underestimated. This study was conducted to assess the effect of continuous administration of BTC on β-cell regeneration. Adenovirus vectors encoding proBTC (Ad-proBTC) and mature BTC (Ad-mBTC) were prepared, and the efficacy of secretion of BTC was compared in AML12 hepatocytes. When AML12 cells were infected with Ad-proBTC or Ad-mBTC, cells infected with Ad-mBTC secreted considerably larger amount of BTC. We then infused Ad-mBTC into the mouse tail vein. Expression of BTC was detected in the liver for at least 21 days, and serum BTC was maintained at approximately 1 ng/ml for 7 days. When Ad-mBTC was infused immediately after administration of STZ (170 mg/kg), elevation of the plasma glucose induced by STZ was markedly inhibited, and the plasma glucose concentration remained at less than 200 mg/dl for 21 days. The insulin content and the β-cell mass were significantly increased in Ad-mBTC-infused mice. These results indicate that continuous administration of BTC is quite effective in promoting regeneration of β-cells.

Expression of betacellulin and epidermal growth factor receptor in hepatocellular carcinoma: implications for angiogenesis

Hepatocellular carcinoma (HCC) is becoming one of the common malignant tumors worldwide and is characterized by high vascularity. Angiogenesis (formation of new microvessels) is critical for growth and progression of various human solid tumors. Betacellulin (BTC) is a member of the epidermal growth factor (EGF) family, and its signal action is mediated through EGF receptors (EGFR). In this study, to understand the role of BTC in relation to EGFR in HCC, we examined localization, expression, and involvement in angiogenesis of BTC and EGFR. The results revealed that expression of BTC, EGFR, and tumor growth factor-alpha messenger RNA in HCC was increased by 80%, 60%, and 40%, respectively, as compared with those in the nontumorous tissues. Increased expression of BTC protein was observed in 31 (61%) of 51 HCC specimens, and the level of tumor growth factor-alpha protein was higher in 17 (33%) of 51 HCC specimens than in nonmalignant hepatocytes. Betacellulin was predominantly expressed in HCC cells, whereas EGFR was observed in sinusoidal endothelial cells of HCC in 25 tumors (49%). Betacellulin was secreted in all 4 examined HCC cell lines. The HCC specimens showing positive EGFR expression in tumor endothelial cells had a significantly higher microvessel density than those without EGFR expression (P < .005). A strong correlation was found between BTC expression in cancer cells and EGFR expression in tumor endothelial cells (P < .001). These findings suggest that overexpression of BTC by HCC cells and EGFR by tumor endothelial cells enhance vascularity in a paracrine manner.

Changes in the expression and cytological localization of betacellulin and its receptors (ErbB-1 and ErbB-4) in the trophoblasts in human placenta over the course of pregnancy.

Betacellulin (BTC), purified and cloned from mouse beta cell tumor (BTC-JC10), is regarded as a new member of the epidermal growth factor family. The present study was conducted to clarify the expression of BTC and its receptors, ErbB-1 and ErbB-4, in the trophoblasts in the human placenta over the course of pregnancy. Human placental tissues were obtained from 4 pregnant women at the 4th to 5th week of pregnancy (very early placentas), 10 women at the 6th to 12th week (early placentas), 5 women at the 18th to 21st week (mid placentas) and 8 women at the 38th and 40th week (term placentas). The mRNA expressions of BTC, erbB-1 and erbB-4 were evaluated by quantitative RT-PCR with Southern blotting and the expression of the soluble form of BTC was determined by western immunoblot with a specific antibody to BTC protein. Immunohistochemical staining of BTC, ErbB-1 and ErbB-4 was also performed. The levels of BTC mRNA expression in early and mid placentas were significantly higher than those in term placentas. The soluble form of BTC protein with an estimated molecular mass of 9.5 kDa was expressed in early and mid placentas, whereas the soluble form was not detected in term placentas. BTC from very early placentas until mid placentas was immunolocalized in syncytiotrophoblasts (S-cells), and was most abundant in early placentas. In contrast, BTC was immunolocalized in extravillous trophoblasts (EVTs), but not in villous trophoblasts in term placentas. The levels of erbB-1 mRNA in the early and mid placentas were significantly higher than those in term placentas, whereas the levels of erbB-4 mRNA in early placentas were significantly lower than those in mid and term placentas. ErbB-1 was immunolocalized in cytotrophoblasts in very early placentas, whereas it was immunolocalized in S-cells from early until term placentas. ErbB-4 from very early placentas until mid placentas was immunolocalized in S-cells, whereas ErbB-4 in the term placentas was detected in EVTs, but not in villous trophoblasts. These findings provide evidence for changes in expression and cytological localization of BTC and its receptors in the trophoblasts in human placenta over the course of pregnancy. BTC may play a pivotal role as a local growth factor in promoting the differentiated villous trophoblastic function via ErbB-1 in early placentas and in contributing to placental growth through the maintenance of EVT cell function via ErbB-4 in term placentas.

Analysis of expression profiles of islet-associated transcription and growth factors during beta-cell neogenesis from duct cells in partially duct-ligated mice.

Beta-cell neogenesis from pancreatic duct cells has been reported to occur in duct-ligated rat. Nevertheless, detailed process of this phenomenon has not been clarified. To clarify the mechanism of beta-cell neogenesis, a partial pancreatic duct ligation mouse model was created. Proliferation of duct cells, beta-cell neogenesis, and expression of transcription factors and differentiation/growth factors were studied by immunohistochemistry, cDNA array, and RT-PCR methods. In the duct-ligated portion of the pancreas, newly formed islet-like cell clusters (ICCs) were observed arising from the ducts on day 7 and afterward. Transcription factors, such as pancreatic and duodenal homeobox gene-1 (PDX-1), paired box factor 6 (Pax6), islet1 and Nkx2.2-positive cells, and protein gene product 9.5 (PGP9.5) were also induced in duct lining cells. By cDNA microarray analysis, expression of insulin-like growth factor-1 (IGF-1) and transforming growth factor beta1 (TGF-beta1) were above control levels on day 5, and RT-PCR showed an increase from day 5 to day 28. IGF-1 and activin A-positive cells were detected in ducts. In addition, expression of betacellulin (BTC), heparin-binding epidermal growth factor-like growth factor (HB-EGF), and TGF-alpha were also increased from day 3 or 5. These findings suggest that beta-cell or endocrine precursors are localized among duct lining cells. Induction of several islet cell-associated transcription factors and differentiation and/or growth factors may play important roles during beta-cell neogenesis in this model.

Auto-induction and growth stimulatory effect of betacellulin in human pancreatic cancer cells.

Betacellulin (BTC) was identified in mouse pancreatic beta cell tumors as a member of the epidermal growth factor (EGF) family, and was found to bind and activate the EGF receptor. BTC is also expressed in some human malignancies and may have an important role in tumor growth progression. We examined whether BTC and EGF have a growth stimulatory effect on human pancreatic cancer cell lines both in vitro and in vivo. We also investigated the BTC expression and autonomous induction of BTC in pancreatic cancer cells. in vitro, both BTC and EGF had almost the same proliferative effect on Panc-1, MIA PaCa-2 and AsPC-1. in vivo, in a Panc-1 inoculated athymic mice model, BTC-treated tumors grew approximately five times larger than in control. Immunocytochemistry showed that BTC expression occurred in three pancreatic cancer cell lines, with MIA PaCa-2 showing the strongest intensity. Semi-quantitative RT-PCR of MIA Paca-2 showed that mRNA levels of BTC gradually increased after treatment with 1 nM BTC. Immunocytochemistry also demonstrated that the intensity of BTC-like immunoreactivity was increased when treated with 1 nM BTC but was reduced after treatment with 100 nM of AG1478, an EGF receptor tyrosine kinase inhibitor. BTC has thus a significant growth stimulatory effect on pancreatic cancer cells and might function as an autocrine and paracrine growth factor. BTC expression in pancreatic cancer cells is, at least in part, controlled by an auto-induction mechanism.

Expression of Betacellulin and Epiregulin Genes in the Mouse Uterus Temporally by the Blastocyst Solely at the Site of Its Apposition Is Coincident with the “Window” of Implantation

In the mouse, the process of implantation is initiated by the attachment reaction between the blastocyst trophectoderm and uterine luminal epithelium that occurs at 2200–2300 h on day 4 (day 1 = vaginal plug) of pregnancy. Several members of the EGF family are considered important in embryo–uterine interactions during implantation. This investigation demonstrates that the expression of two additions to the family, betacellulin and epiregulin, are exquisitely restricted to the mouse uterine luminal epithelium and underlying stroma adjacent to the implanting blastocyst. These genes are not expressed during progesterone-maintained delayed implantation, but are rapidly switched on in the uterus surrounding the implanting blastocyst following termination of the delay by estrogen. These results provide evidence that expression of betacellulin and epiregulin in the uterus requires the presence of an active blastocyst and suggest an involvement of these growth factors in the process of implantation.