Epidermal Growth Factor Family Research Articles

The mechanosensory channel PIEZO1 functions upstream of angiopoietin/TIE/FOXO1 signaling in lymphatic development.

Lymphedema is a debilitating disease with no effective cure and affects an estimated 250 million individuals worldwide. Prior studies have identified mutations in piezo-type mechanosensitive ion channel component 1 (PIEZO1), angiopoietin 2 (ANGPT2), and tyrosine kinase with Ig-like and EGF-like domains 1 (TIE1) in patients with primary lymphedema. Here, we identified crosstalk between these molecules and showed that activation of the mechanosensory channel PIEZO1 in lymphatic endothelial cells (LECs) caused rapid exocytosis of the TIE ligand ANGPT2, ectodomain shedding of TIE1 by disintegrin and metalloproteinase domain-containing protein 17 (ADAM17), and increased TIE/PI3K/AKT signaling, followed by nuclear export of the transcription factor FOXO1. These data establish a functional network between lymphedema-associated genes and provide what we believe to be the first molecular mechanism bridging channel function with vascular signaling and intracellular events culminating in transcriptional regulation of genes expressed in LECs. Our study provides insights into the regulation of lymphatic function and molecular pathways involved in human disease.

The promotion of non-small cell lung cancer progression by collagen and calcium binding EGF domain 1 is mediated through the regulation of ERK/JNK/P38 phosphorylation by reactive oxygen species.

Reactive oxygen species (ROS) are metabolic by-products of cells, and abnormal changes in their levels are often associated with tumor development. Our aim was to determine the role of collagen and calcium binding EGF domain 1 (CCBE1) in oxidative stress and tumorigenesis in non-small cell lung cancer cells (NSCLC). We investigated the tumorigenic potential of CCBE1 in NSCLC using in vitro and in vivo models of CCBE1 overexpression and knockdown. Immunohistochemical staining results showed that the expression of CCBE1 in cancer tissues was significantly higher than that in adjacent tissues. Cell counting Kit 8, clonal formation, wound healing, and transwell experiments showed that CCBE1 gene knockdown significantly inhibited the migration, invasion, and proliferation of NSCLC cell lines. In terms of mechanism, the silencing of CCBE1 can significantly promote the morphological abnormalities of mitochondria, significantly increase the intracellular ROS level, and promote cell apoptosis. This change of oxidative stress can affect cell proliferation, migration, and invasion by regulating the phosphorylation level of ERK/JNK/P38 MAPK. Specifically, the downregulation of CCBE1 inhibits the phosphorylation of ERK/P38 and promotes the phosphorylation of JNK in NSCLC, and this regulation can be reversed by the antioxidant NAC. In vivo experiments confirmed that downregulating CCBE1 gene could inhibit the growth of NSCLC in BALB/c nude mice. Taken together, our results confirm the tumorigenic role of CCBE1 in promoting tumor invasion and migration in NSCLC, and reveal the molecular mechanism by which CCBE1 regulates oxidative stress and the ERK/JNK/P38 MAPK pathway.

Functional implication of the homotrimeric multidomain vacuolar sorting receptor 1 (VSR1) from Arabidopsis thaliana

The vacuolar sorting receptors (VSRs) are specific to plants and are responsible for sorting and transporting particular proteins from the trans-Golgi network to the vacuole. This process is critically important for various cellular functions, including storing nutrients during seed development. Despite many years of intense studies on VSRs, a complete relation between function and structure has not yet been revealed. Here, we present the crystal structure of the entire luminal region of glycosylated VSR1 from Arabidopsis thaliana (AtVSR1) for the first time. The structure provides insights into the tertiary and quaternary structures of VSR1, which are composed of an N-terminal protease-associated (PA) domain, a unique central region, and one epidermal growth factor (EGF)-like domain followed by two disordered EGF-like domains. The structure of VSR1 exhibits unique characteristics, the significance of which is yet to be fully understood.

The Expression and Function of Notch Involved in Ovarian Development and Fecundity in Basilepta melanopus.

Basilepta melanopus is a pest that severely affects oil tea plants, and the Notch signaling pathway plays a significant role in the early development of insect ovaries. In this study, we explored the function of the notch gene within the Notch signaling pathway in the reproductive system of B. melanopus. The functional domains and expression patterns of Bmnotch were analyzed. Bmnotch contains 45 epidermal growth factor-like (EGF-like) domains, one negative regulatory region, one NODP domain and one repeat-containing domain superfamily. The qPCR reveals heightened expression in early developmental stages and specific tissues like the head and ovaries. The RNA interference (RNAi)-based suppression of notch decreased its expression by 52.1%, exhibiting heightened sensitivity to dsNotch at lower concentrations. Phenotypic and mating experiments have demonstrated that dsNotch significantly impairs ovarian development, leading to reduced mating frequencies and egg production. This decline underscores the Notch pathway's crucial role in fecundity. The findings advocate for RNAi-based, Notch-targeted pest control as an effective and sustainable strategy for managing B. melanopus populations, signifying a significant advancement in forest pest control endeavors.

Role of Epiregulin on Lipopolysaccharide-Induced Hepatocarcinogenesis as a Mediator via EGFR Signaling in the Cancer Microenvironment.

Lipopolysaccharides (LPSs) have been reported to be important factors in promoting the progression of hepatocellular carcinoma (HCC), but the corresponding molecular mechanisms remain to be elucidated. We hypothesize that epiregulin (EREG), an epidermal growth factor (EGF) family member derived from hepatic stellate cells (HSCs) and activated by LPS stimulation, is a crucial mediator of HCC progression with epidermal growth factor receptor (EGFR) expression in the tumor microenvironment. We used a mouse xenograft model of Huh7 cells mixed with half the number of LX-2 cells, with/without intraperitoneal LPS injection, to elucidate the role of EREG in LPS-induced HCC. In the mouse model, LPS administration significantly enlarged the size of xenografted tumors and elevated the expression of EREG in tumor tissues compared with those in negative controls. Moreover, CD34 immunostaining and the gene expressions of angiogenic markers by a reverse transcription polymerase chain reaction revealed higher vascularization, with increased interleukin-8 (IL-8) expression in the tumors of the mice group treated with LPS compared to those without LPS. Our data collectively suggested that EREG plays an important role in the cancer microenvironment under the influence of LPS to increase not only the tumor cell growth and migration/invasion of EGFR-positive HCC cells but also tumor neovascularization via IL-8 signaling.

Abstract 626: EGFL6 induced HER2-HER3 signaling controls accurate centrosome deposition of activated ERK and mitotic spindle formation in ovarian cancer cells

Abstract Epidermal growth factor EGF-like domain multiple-6 (EGFL6) is highly expressed in high grade serous ovarian cancer (HGSOC) cells and ovarian tumor vasculature. EGFL6 acts on tumor cells to drive tumor cell proliferation and migration. To better understand EGFL6 signaling and develop therapeutic strategies to target EGFL6 signaling in HGSOC, we evaluated phosphorylation of putative EGFL6 receptors and their downstream signaling. We demonstrated that EGFL6 induced phosphorylation and activation of ERBB/HER family of receptors; EGFL6 treatment of ovarian cancer cells led to rapid and transient phosphorylation of EGFR while inducing prolonged activation of both HER2 and HER3, with subsequent phosphorylation of ERK. Interestingly, we found that in EGFL6 stimulated ovarian cancer cells undergoing mitosis, pERK localized to the centrosome and the contractile ring. EGFL6 neutralizing antibodies reduce ERK phosphorylation and resulted in pERK being aberrantly localized. This resulted in altered mitotic spindle alignment and an increased number of cells undergoing mitotic catastrophe. Furthermore, combination anti-EGFL6 therapy with the pan-EGFR receptor inhibitor neratinib, compared to either therapy alone, led to an increase in aberrant pERK localization and cancer cell death in vitro. Consistent with these findings, dual anti-EGFL6 and neratinib therapy significantly restricted tumor growth in vivo, resulting in increased tumor cell death and a reduction in angiogenesis. Combined our data identify an unexplored role for pERK at the centrosome and suggest that dual targeting of the EGFL6/HER signaling axis maybe an effective therapeutic strategy in ovarian cancer. Citation Format: Shoumei Bai, Navneet Gupta, Qi Jiang, Ronald J. Buckanovich. EGFL6 induced HER2-HER3 signaling controls accurate centrosome deposition of activated ERK and mitotic spindle formation in ovarian cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 626.

Tumor-derived mesenchymal progenitor cell-related genes in the regulation of breast cancer proliferation.

Breast cancer (BC) is one of the most common malignancies worldwide, and its development is affected in various ways by the tumor microenvironment (TME). Tumor-derived mesenchymal progenitor cells (MPCs), as the most important components of the TME, participate in the proliferation and metastasis of BC in several ways. In this study, we aimed to characterize the genes associated with tumor-derived MPCs and determine their effects on BC cells. Tumor-derived MPCs and normal breast tissue-derived mesenchymal stem cells (MSCs) were isolated from tissues specimens of patients with BC. We conducted culture and passage, phenotype identification, proliferation and migration detection, inflammatory factor release detection, and other experiments on isolated MPCs from tumors and MSCs from normal breast tissues. Three paired tumor-derived MPCs and normal breast tissue-derived MSCs were then subjected to transcriptome analysis to determine the expression profiles of the relevant genes, and quantitative real-time polymerase chain reaction (qRT-PCR) was used to further confirm gene expression. Subsequently, the overexpression plasmids were transfected into tumor-derived MPCs, and the expression of various inflammatory factors of tumor-derived MPCs and their proliferation were characterized with a cell viability test reagent (Cell Counting Kit 8). Subsequently, the transfected tumor-derived MPCs were cocultured with BC cells using a conditioned medium coculture method to clarify the role of tumor-derived MSCs in BC. Tumor-derived MPCs expressed stem cell characteristics including CD105, CD90, and CD73 and exhibited adipogenic and osteogenic differentiation in vitro. The proliferation of tumor-derived MPCs was significantly lower than that of normal breast tissue-derived MSCs, and the invasive metastatic ability was comparable; however, MPCs were found to release inflammatory factors such as interleukin 6 (IL-6) and transforming growth factor β (TGF-β). Transcriptome analysis showed that stomatin (STOM), collagen and calcium binding EGF domains 1 (CCBE1), and laminin subunit alpha 5 (LAMA5) were significantly upregulated in tumor-derived MPCs. Among them, STOM was highly expressed in tumor-derived MPCs, which mediated the slow proliferation of MPCs and promoted the proliferation of BC cells. STOM, CCBE1, and LAMA5 were highly expressed in tumor-derived MPCs, with STOM being found to retard the proliferation of MPCs but promote the proliferation of BC cells. There findings present new possibilities in targeted microenvironmental therapy for BC.

Isolation and characterization of rabbit limbal niche cells

Limbal niche cells (LNCs) are one of the most important supporting cells for corneal epithelial stem cells (CES), however, research on LNCs has been mostly limited to humans and rats previously. To expand the research work into the rabbit animal model, one of the most often used animals in stem cell study, this study was carried out for the in vitro isolation and identification of rabbit LNCs.Rabbit LNCs were isolated by collagenase A digestion method and single cells were obtained, the cells were then seeded on 5% Matrigel-coated plastic surface and cultured in modified embryonic stem cell medium (MESCM). Three biological replicates of the isolating and characterization were recorded from New Zealand White rabbits aged from 2.5 months to 5 months. LNC markers (VIM/CD90/CD105/SCF/PDGFRβ) were analyzed using tyramide signal amplification (TSA) staining, immunohistochemical staining (IHC), western blotting (WB), and real-time reverse transcription polymerase chain reaction (qPCR). TSA staining suggested that VIM was highly expressed in rabbit limbus stroma, which was confirmed by WB, and P63α was expressed in the basal limbus epithelium. Pan-CK and CK12 were highly expressed in the central corneal epithelium but lightly expressed in the limbal epithelium. The WB result indicated that PDGFRβ and VIM expressions in rabbit-LNCs P4 were higher than in P1 and P7. In addition, rabbit corneal epithelium highly expressed Paired Box 6 (PAX6) and Epidermal growth factor-like domain 6(EGFL6). For the three repeat experiments, the cell expansion activity of rabbit-LNC was highest at P4. Rabbit-LNCs were passaged from P0 to P7, and the number of cell doublings (NCD) of P4 for the three repeat experiments was 2.816, 2.737, and 2.849. qPCR showed that high mRNA expression levels of VIM, CD90, CD105, SCF, and PDGFRβ in rabbit-LNCs P4.In conclusion, rabbit-LNCs could be successfully isolated by the collagenase A digestion method as used in human tissue. There were similar characteristics between rabbit and human LNCs (VIM+/CD90+/CD105+/SCF+/PAX6+/PDGFRβ+).

Role of Epiregulin in Lung Tumorigenesis and Therapeutic Resistance.

Epidermal growth factor (EGF) signaling regulates multiple cellular processes and plays an essential role in tumorigenesis. Epiregulin (EREG), a member of the EGF family, binds to the epidermal growth factor receptor (EGFR) and ErbB4, and it stimulates EGFR-related downstream pathways. Increasing evidence indicates that both the aberrant expression and oncogenic function of EREG play pivotal roles in tumor development in many human cancers, including non-small cell lung cancer (NSCLC). EREG overexpression is induced by activating mutations in the EGFR, KRAS, and BRAF and contributes to the aggressive phenotypes of NSCLC with oncogenic drivers. Recent studies have elucidated the roles of EREG in a tumor microenvironment, including the epithelial-mesenchymal transition, angiogenesis, immune evasion, and resistance to anticancer therapy. In this review, we summarized the current understanding of EREG as an oncogene and discussed its oncogenic role in lung tumorigenesis and therapeutic resistance.

Host protein EPCAM interacting with EtMIC8-EGF is essential for attachment and invasion of Eimeria tenella in chickens

The five epidermal growth factor-like domains (EGF) of Eimeria tenella microneme protein 8 (EtMIC8) (EtMIC8-EGF) plays a vital role in host cell attachment and invasion. These processes require interactions between parasite proteins and receptors on the surface of host cells. In this study, five chicken membrane proteins potentially interacting with EtMIC8-EGF were identified using the GST pull-down assay and mass spectrometry analysis, and only chicken (Gallus gallus) epithelial cell adhesion molecule (EPCAM) could bind to EtMIC8-EGF. EPCAM-specific antibody and recombinant EPCAM protein (rEPCAM) inhibited the EtMIC8-EGF binding to host cells in a concentration-dependent manner. Furthermore, the rEPCAM protein showed a binding activity to sporozoites in vitro, and a significant reduction of E. tenella invasion in DF-1 cells was further observed after pre-incubation of sporozoites with rEPCAM. The specific anti-EPCAM antibody further significantly decreased weight loss, lesion score and oocyst output during E. tenella infection, displaying partial inhibition of E. tenella infection. These results indicate that chicken EPCAM is an important EtMIC8-interacting host protein involved in E. tenella-host cell adhesion and invasion. The findings will contribute to a better understanding of the role of adhesion-associated microneme proteins in E. tenella.

Epigen enhances the developmental potential of in vitro fertilized embryos by improving cytoplasmic maturation

Numerous growth factors contribute to oocyte maturation and embryonic development in vivo; however, only a few are understood. One such factor is epigen, a new member of the epidermal growth factor (EGF) family that is secreted by the granulosa cells of immature oocytes. We hypothesized that epigen may play a role in oocyte maturation, specifically in the nuclear and cytoplasmic aspects. This study aimed to investigate the effects of epigen on porcine oocyte maturation and embryo development in vitro. In this study, three different concentrations of epigen (3, 6, and 30 ng/mL) were added to tissue culture medium-199 (TCM-199) during in vitro maturation of porcine oocytes. A control group that did not receive epigen supplementation was also included. Mature porcine oocytes were fertilized, and the resulting zygotes were cultured until day 7. The levels of intracellular glutathione (GSH) and reactive oxygen species (ROS) were measured in the in vitro matured oocytes. At the same time, the expression patterns of genes related to apoptosis were detected in day 7 blastocysts (BLs) using real-time quantitative PCR Apoptosis was detected by annexin-V assays in mature oocytes. Data were analyzed using ANOVA and Duncan's test on SPSS, and results are presented as mean ± SEM. The group that received 6 ng/mL epigen had a significantly lower rate of germinal vesicle breakdown (GVBD) than the control group without affecting the nuclear maturation among the experimental groups. Among the treatment groups, the 6 ng/mL epigen group showed significantly higher levels of intracellular GSH and lower ROS production. Supplementation with 6 ng/mL epigen significantly improved blastocyst (BL) formation rates compared to those in the control and 3 ng/mL groups. Additionally, the blastocyst expansion rate was significantly higher with epigen supplementation (6 ng/mL). In the fertilization experiment, the group supplemented with 6 ng/mL epigen exhibited significantly higher levels of monospermy and fertilization efficiency and lower levels of polyspermy than the control group. This study indicated that adding epigen at a concentration of 6 ng/mL can significantly enhance the developmental potential of porcine oocytes fertilized in vitro. Specifically, the study found that epigen improves cytoplasmic maturation, which helps prevent polyspermy and emulates monospermic penetration.

Astrocytes Excessively Engulf Synapses in a Mouse Model of Alzheimer's Disease.

Synapse loss is one of the most critical features in Alzheimer's disease (AD) and correlates with cognitive decline. Astrocytes mediate synapse elimination through multiple EGF-like domains 10 (MEGF10) pathways in the developing and adult brain to build the precise neural connectivity. However, whether and how astrocytes mediate synapse loss in AD remains unknown. We here find that the phagocytic receptor MEGF10 of astrocytes is significantly increased in vivo and in vitro, which results in excessive engulfment of synapses by astrocytes in APP/PS1 mice. We also observe that the astrocytic lysosomal-associated membrane protein 1 (LAMP1) is significantly elevated, colocalized with the engulfed synaptic puncta in APP/PS1 mice, and astrocytic lysosomes contain more engulfed synaptic puncta in APP/PS1 mice relative to wild type mice. Together, our data provide evidence that astrocytes excessively engulf synapses in APP/PS1 mice, which is mediated by increased MEGF10 and activated lysosomes. The approach targeting synapse engulfment pathway in astrocytes would be a potent therapy for AD.

Cysteine-rich with EGF-like domains 2 (CRELD2) is an endoplasmic reticulum stress-inducible angiogenic growth factor promoting ischemic heart repair

Tissue repair after myocardial infarction (MI) is guided by autocrine and paracrine-acting proteins. Deciphering these signals and their upstream triggers is essential when considering infarct healing as a therapeutic target. Here we perform a bioinformatic secretome analysis in mouse cardiac endothelial cells and identify cysteine-rich with EGF-like domains 2 (CRELD2), an endoplasmic reticulum stress-inducible protein with poorly characterized function. CRELD2 was abundantly expressed and secreted in the heart after MI in mice and patients. Creld2-deficient mice and wild-type mice treated with a CRELD2-neutralizing antibody showed impaired de novo microvessel formation in the infarct border zone and developed severe postinfarction heart failure. CRELD2 protein therapy, conversely, improved heart function after MI. Exposing human coronary artery endothelial cells to recombinant CRELD2 induced angiogenesis, associated with a distinct phosphoproteome signature. These findings identify CRELD2 as an angiogenic growth factor and unravel a link between endoplasmic reticulum stress and ischemic tissue repair.

Structural insight into CD93 recognition by IGFBP7

The CD93/IGFBP7 axis proteins are key factors expressed in endothelial cells (EC) that mediate EC angiogenesis and migration. Their upregulation contributes to tumor vascular abnormality and a blockade of this interaction promotes a favorable tumor microenvironment for therapeutic interventions. However, the interactions of these proteins with each other remain unclear. In this study, we determined a partial structure of the human CD93-IGFBP7 complex comprising the EGF1 domain of CD93 and the IB domain of IGFBP7. Mutagenesis studies confirmed interactions and specificities. Cellular and mouse tumor studies demonstrated the physiological relevance of the CD93-IGFBP7 interaction in EC angiogenesis. Our study provides leads for the development of therapeutic agents to precisely disrupt unwanted CD93-IGFBP7 signaling in the tumor microenvironment. Additionally, analysis of the CD93 full-length architecture provides insights into how CD93 protrudes on the cell surface and forms a flexible platform for binding to IGFBP7 and other ligands.

Genome-Wide Association Study Identifies Rice Panicle Blast-Resistant Gene Pb4 Encoding a Wall-Associated Kinase.

Rice blast is one of the most devastating diseases, causing a significant reduction in global rice production. Developing and utilizing resistant varieties has proven to be the most efficient and cost-effective approach to control blasts. However, due to environmental pressure and intense pathogenic selection, resistance has rapidly broken down, and more durable resistance genes are being discovered. In this paper, a novel wall-associated kinase (WAK) gene, Pb4, which confers resistance to rice blast, was identified through a genome-wide association study (GWAS) utilizing 249 rice accessions. Pb4 comprises an N-terminal signal peptide, extracellular GUB domain, EGF domain, EGF-Ca2+ domain, and intracellular Ser/Thr protein kinase domain. The extracellular domain (GUB domain, EGF domain, and EGF-Ca2+ domain) of Pb4 can interact with the extracellular domain of CEBiP. Additionally, its expression is induced by chitin and polygalacturonic acid. Furthermore, transgenic plants overexpressing Pb4 enhance resistance to rice blast. In summary, this study identified a novel rice blast-resistant gene, Pb4, and provides a theoretical basis for understanding the role of WAKs in mediating rice resistance against rice blast disease.

TIE2 expression in hypertensive ICH and its therapeutic modulation with AKB-9778: Implications for brain vascular health

Hypertensive intracerebral hemorrhage (ICH) is a devastating condition, the molecular underpinnings of which remain not fully understood. By leveraging high-throughput transcriptome sequencing and network pharmacology analysis, this study unveils the significant role of the tyrosine kinase with immunoglobulin-like and EGF-like domains 2 (TIE2) in ICH pathogenesis. Compared to controls, a conspicuous downregulation of TIE2 was observed in the cerebral blood vessels of hypertensive ICH mice. In vitro assays with human brain microvascular endothelial cells (HBMEC), HBEC-5i revealed that modulation of TIE2 expression significantly influences cellular proliferation, migration, and angiogenesis, mediated via the Rap1/MEK/ERK signaling pathway. Notably, the small molecule AKB-9778 was identified to target and activate TIE2, affecting the functional attributes of HBEC-5i. In vivo experiments further demonstrated that combining AKB-9778 with antihypertensive drugs could mitigate the incidence and volume of bleeding in hypertensive ICH mouse models, suggesting potential therapeutic implications.

[Research progress on HEG1 in cardiovascular generation and tumor development].

Heart development protein with EGF-like domains 1 (HEG1) is a novel mucin-like membrane protein with a long O-glycosylation region and EGF domain. HEG1 plays critical roles in embryo development and cardiogenesis, and is closely related to the occurrence and progression of malignant tumors. Here this article demonstrates the research progress on HEG1 in cardiovascular formation and tumor development in recent years, to inspire new ideas for the pathogenesis, diagnosis and treatment of related diseases.

Research Note: Preliminary functional analysis of EGF-like domains of Eimeria tenella microneme protein 7 identified in sporozoites and merozoites

Eimeria tenella, an obligate intracellular apicomplexan parasite, is the major causative agent of chicken coccidiosis. Some epidermal growth factor (EGF)-like domain-containing proteins of other members of apicomplexan parasites have been reported to contribute to parasite survival. To date, however, EGF-like domain-containing proteins of E. tenella are not well studied. In this study, a gene fragment that encodes 4 EGF-like domains of E. tenella microneme protein 7 (EGF-EtMIC7) was amplified and expressed using an Escherichia coli expression system. Following generation of polyclonal antibodies that recognize recombinant EGF-EtMIC7 (rEGF-EtMIC7), the expression of EtMIC7 in sporozoites and merozoites was examined. Moreover, its roles in cellular regulation were investigated. The native EtMIC7 in E. tenella sporozoites and merozoites was detected by using Western blot and indirect immunofluorescence assays. rEGF-EtMIC7 could activate Akt, whereas blockade of EGF receptor (EGFR) failed to induce Akt phosphorylation. Compared with the control group, LMH cells treated with rEGF-EtMIC7 showed increased cell proliferation and expressed higher levels of B cell leukemia/lymphoma 2 (BCL-2). These findings contribute to the better understanding of parasite-host interactions at the molecular level during E. tenella infection.

In vitro effects of heparin-binding epidermal growth factor on adhesion stage of implantation.

A member of the epidermal growth factor (EGF) family, the heparin-binding EGF (HB-EGF) is expressed in the uteri of both humans and mice during the implantation process. To study the effects of HB-EGF on adhesion stage, we developed an in vitro implantation model employing Ishikawa cell line and JAR cell line, which may attach to Ishikawa cells. For 1, 6, 12, and 24 hours, co-cultures of JAR spheroids grown on Ishikawa monolayers were treated with 1, 10, and 100 ng∕mL doses of HB-EGF. Using immunocytochemistry and Western blot analysis, the effects of HB-EGF on the protein expressions of E-cadherin, Erb-B2 receptor tyrosine kinase 4 (ErbB4), and integrin ανβ3 in Ishikawa and JAR cells were examined semi-quantitatively and quantitatively. Ultrastructural changes of in vitro implantation model were investigated by transmission electron microscopy. We revealed that HB-EGF influenced trophoblast cell adhesion to endometrial cells by upregulating the expression of the proteins ErbB4 and trophoblastic integrin ανβ3. Decrease in trophoblastic E-cadherin expression and increase in endometrial E-cadherin expression were demonstrated accompanying morphological variations in cells required for the invasion. We discovered ultrastructurally that Ishikawa cells acquired uterodome-like appearance, including the organelles, when 10 and 100 ng∕mL dosages of HB-EGF were administered for 12 and 24 hours. However, following additional hours of adhesion and invasion, their intercellular spaces enlarged. The trafficking of vesicular transport was enhanced by JAR spheroids. We therefore discovered that in this implantation paradigm, HB-EGF may enhance the receptivity of Ishikawa cells and the adherence of JAR cells.

Identification of the Expression of TIE1 and Its Mediated Immunosuppression in Gastric Cancer.

Background: Recently, various evidence has confirmed that Tyrosine Kinase with Immunoglobulin-like and EGF-like domains 1 (TIE1) promotes tumor growth in many cancers. However, the precise mechanism underlying TIE1's involvement in Gastric Cancer (GC) remains elusive. This research aimed to investigate the biological function of TIE1 in regulating GC progression. Methods: The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), GEPIA2.0, Sangerbox3.0 and TIMER databases were used to analyze the TIE1 expression. Immunohistochemistry (IHC) was used to demonstrate the expression of TIE1. TCGA, GEPIA2.0 and Kaplan-Meier were utilized for survival analysis and to explore the association of TIE1 with clinicopathological features. Protein-Protein Interaction (PPI) networks were constructed using Cytoscape. The potential molecular mechanism of TIE1 was investigated by Gene Ontology (GO), Kyoto Encyclopedia of Gene Genomes (KEGG), and Gene Set Enrichment Analysis (GSEA). We studied the relationships between TIE1 and mutations, immune checkpoints (ICs), tumor mutational burden (TMB), as well as microsatellite instability (MSI) to explore the underlying mechanism of immunity in GC. Results: Compared with normal tissue, TIE1 was significantly overexpressed in GC tissues (p = 0.0072) and was associated with poor survival (P < 0.05). According to GO and KEGG enrichment analyses, TIE1 was enriched in signal pathways related to the occurrence, invasion, and migration of malignant tumors (i.e., PI3K-Akt signaling pathway, Calcium signaling pathway, etc.). Immune infiltration analysis suggested that TIE1 is positively correlated with macrophages M2 and negatively correlated with Mast cells, naive B cells and Follicular helper T cells (TFH), which may be a contributing factor to tumor progression. Furthermore, the research on the tumor microenvironment (TME) and tumor purity also proved that TIE1 may be an oncogene. Mutation analysis showed that the high expression group of TIE1 had a higher frequency of mutations in TP53 and ARID1, while the TMB score was lower. Conclusion: TIE1 might be an oncogene via regulating dysregulated immune infiltration to cause immunosuppression in GC and could be identified as a biomarker for prognosis and a therapeutic target for GC.