ABSTRACT
OBJECTIVE: Recent studies have revealed that the ability of cancer cells to undergo intermediate state of epithelial-to-mesenchymal transition (EMT), partial EMT (p-EMT), poses a higher metastatic risk rather than complete EMT. Here, we examined the prognostic value of p-EMT-related genes in head and neck squamous cell carcinoma (HNSCC) by bioinformatic approaches. MATERIALS AND METHODS: We used RNA-seq data of 519 primary HNSCC cases obtained from TCGA database. We compared the expression of p-EMT-related genes in HNSCC tissues with normal tissues. We evaluated the prognostic value of p-EMT-related genes in HNSCC cases by log-rank test. We examined the expression of p-EMT-, EMT-, and epithelial differentiation-related genes by qPCR. RESULTS: Among p-EMT-related genes that were highly expressed in HNSCC cases, high expression of SERPINE1, ITGA5, TGFBI, P4HA2, CDH13, and LAMC2 was significantly correlated with poor survival of HNSCC patients. By gene expression pattern, HNSCC cell lines were classified into three groups: epithelial phenotype, EMT phenotype, and p-EMT phenotype. CONCLUSIONS: Our findings suggest that p-EMT program may be involved in poor prognosis of HNSCC. SERPINE1, ITGA5, TGFBI, P4HA2, CDH13, and LAMC2 can be used for a prognostic marker. Moreover, HNSCC cells with p-EMT phenotype can be a useful model for investigating a nature of p-EMT.
ABSTRACT
Aire controls the fate of autoreactive thymocytes (i.e., clonal deletion or development into regulatory T cells [Tregs]) through transcriptional control of the expression of tissue-restricted self-antigens (TRAs) from medullary thymic epithelial cells (mTECs) and bone marrow (BM)-derived cells. Although TRAs expressed by mTECs and BM-derived cells are suggested to complement each other to generate a full spectrum of TRAs, little is known about the relative contribution of TRAs from each component for establishment of self-tolerance. Furthermore, the precise role of Aire in specific types of Aire-expressing APCs remains elusive. We have approached these issues by generating two different types of transgenic mouse (Tg) model, which express a prefixed model self-antigen driven by the insulin promoter or the Aire promoter. In the insulin-promoter Tg model, mTECs alone were insufficient for clonal deletion, and BM-derived APCs were required for this action by utilizing Ag transferred from mTECs. In contrast, mTECs alone were able to induce Tregs, although at a much lower efficiency in the absence of BM-derived APCs. Importantly, lack of Aire in mTECs, but not in BM-derived APCs, impaired both clonal deletion and production of Tregs. In the Aire-promoter Tg model, both mTECs and BM-derived APCs could independently induce clonal deletion without Aire, and production of Tregs was impaired by the lack of Aire in mTECs, but not in BM-derived APCs. These results suggest that the fate of autoreactive thymocytes together with the requirement for Aire depend on the cell types that express self-antigens and the types of APCs involved in tolerance induction.
Subject(s)
Antigen Presentation , Antigen-Presenting Cells/immunology , Autoantigens/immunology , Clonal Deletion/immunology , T-Lymphocytes, Regulatory/immunology , Thymus Gland/immunology , Transcription Factors/immunology , Animals , Autoantigens/biosynthesis , Autoantigens/genetics , Epithelial Cells/immunology , Epithelial Cells/metabolism , Female , Gene Expression Regulation/immunology , Gene Knock-In Techniques , Genes, Synthetic , Insulin/genetics , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Ovalbumin/biosynthesis , Ovalbumin/genetics , Ovalbumin/immunology , Promoter Regions, Genetic , Rats , Specific Pathogen-Free Organisms , Thymus Gland/cytology , Transcription Factors/deficiency , Transcription Factors/genetics , Transgenes , AIRE ProteinABSTRACT
Autoimmunity is prevented by the function of the autoimmune regulator [AIRE (Aire in mice)], which promotes the expression of a wide variety of tissue-restricted antigens (TRAs) from medullary thymic epithelial cells (mTECs) and from a subset of peripheral antigen-presenting cells (APCs). We examined the effect of additive expression of human AIRE (huAIRE) in a model of autoimmune diabetes in NOD mice. Unexpectedly, we observed that mice expressing augmented AIRE/Aire developed muscle-specific autoimmunity associated with incomplete maturation of mTECs together with impaired expression of Aire-dependent TRAs. This led to failure of deletion of autoreactive T cells together with dramatically reduced production of regulatory T cells in the thymus. In peripheral APCs, expression of costimulatory molecules was augmented. We suggest that levels of Aire expression need to be tightly controlled for maintenance of immunological tolerance. Our results also highlight the importance of coordinated action between central tolerance and peripheral tolerance under the common control of Aire.
Subject(s)
Diabetes Mellitus, Type 1/immunology , Muscles/immunology , Polymyositis/immunology , Thymus Gland/immunology , Transcription Factors/metabolism , Animals , Autoantigens/metabolism , Autoimmunity , Disease Models, Animal , Humans , Immune Tolerance , Mice , Mice, Inbred NOD , Mice, Transgenic , Organ Specificity , Transcription Factors/genetics , AIRE ProteinABSTRACT
Aire in medullary thymic epithelial cells (mTECs) plays an important role in the establishment of self-tolerance. Because Aire(+) mTECs appear to be a limited subset, they may constitute a unique lineage(s) among mTECs. An alternative possibility is that all mTECs are committed to express Aire in principle, but Aire expression by individual mTECs is conditional. To investigate this issue, we established a novel Aire reporter strain in which endogenous Aire is replaced by the human AIRE-GFP-Flag tag (Aire/hAGF-knockin) fusion gene. The hAGF reporter protein was produced and retained very efficiently within mTECs as authentic Aire nuclear dot protein. Remarkably, snapshot analysis revealed that mTECs expressing hAGF accounted for >95% of mature mTECs, suggesting that Aire expression does not represent a particular mTEC lineage(s). We confirmed this by generating Aire/diphtheria toxin receptor-knockin mice in which long-term ablation of Aire(+) mTECs by diphtheria toxin treatment resulted in the loss of most mature mTECs beyond the proportion of those apparently expressing Aire. These results suggest that Aire expression is inherent to all mTECs but may occur at particular stage(s) and/or cellular states during their differentiation, thus accounting for the broad impact of Aire on the promiscuous gene expression of mTECs.
Subject(s)
Epithelial Cells/metabolism , Thymus Gland/metabolism , Transcription Factors/biosynthesis , Animals , Cell Differentiation , Diphtheria Toxin/pharmacology , Epithelial Cells/cytology , Gene Expression , Gene Expression Regulation , Gene Knock-In Techniques , Green Fluorescent Proteins/genetics , Heparin-binding EGF-like Growth Factor/genetics , Humans , Mice , Mice, Inbred C57BL , Mice, Inbred NOD , Mice, Transgenic , Thymus Gland/cytology , Transcription Factors/genetics , AIRE ProteinABSTRACT
Cortical thymic epithelial cells (cTECs) and medullary thymic epithelial cells (mTECs) play essential roles in the positive and negative selection of developing thymocytes, respectively. Aire in mTECs plays an essential role in the latter process through expression of broad arrays of tissue-restricted Ags. To determine whether the location of Aire within the medulla is absolutely essential or whether Aire could also function within the cortex for establishment of self-tolerance, we used bacterial artificial chromosome technology to establish a semiknockin strain of NOD-background (ß5t/Aire-transgenic) mice expressing Aire under control of the promoter of ß5t, a thymoproteasome expressed exclusively in the cortex. Although Aire was expressed in cTECs as typical nuclear dot protein in ß5t/Aire-Tg mice, cTECs expressing Aire ectopically did not confer transcriptional expression of either Aire-dependent or Aire-independent tissue-restricted Ag genes. We then crossed ß5t/Aire-Tg mice with Aire-deficient NOD mice, generating a strain in which Aire expression was confined to cTECs. Despite the presence of Aire(+) cTECs, these mice succumbed to autoimmunity, as did Aire-deficient NOD mice. The thymic microenvironment harboring Aire(+) cTECs, within which many Aire-activated genes were present, also showed no obvious alteration of positive selection, suggesting that Aire's unique property of generating a self-tolerant T cell repertoire is functional only in mTECs.
Subject(s)
Autoimmunity/genetics , Self Tolerance/genetics , Thymocytes/immunology , Thymus Gland/immunology , Transcription Factors/genetics , Animals , Autoimmunity/immunology , Cell Differentiation/immunology , Chromosomes, Artificial, Bacterial/genetics , Epithelial Cells/cytology , Epithelial Cells/immunology , Gene Knock-In Techniques , Mice , Mice, Inbred NOD , Mice, Knockout , Self Tolerance/immunology , T-Lymphocytes/immunology , Thymocytes/cytology , Thymus Gland/cytology , Transcription Factors/metabolism , AIRE ProteinABSTRACT
Essential roles of NF-κB-inducing kinase (NIK) for the development of medullary thymic epithelial cells (mTECs) and regulatory T cells have been highlighted by studies using a strain of mouse bearing a natural mutation of the NIK gene (aly mice). However, the exact mechanisms underlying the defect in thymic cross-talk leading to the breakdown of self-tolerance in aly mice remain elusive. In this study, we demonstrated that production of regulatory T cells and the final maturation process of positively selected conventional αß T cells are impaired in aly mice, partly because of a lack of mature mTECs. Of note, numbers of thymic dendritic cells and their expression of costimulatory molecules were also affected in aly mice in a thymic stroma-dependent manner. The results suggest a pivotal role of NIK in the thymic stroma in establishing self-tolerance by orchestrating cross-talk between mTECs and dendritic cells as well as thymocytes. In addition, we showed that negative selection was impaired in aly mice as a result of the stromal defect, which accounts for the development of organ-specific autoimmunity through a lack of normal NIK.
Subject(s)
Cell Communication/immunology , Dendritic Cells/immunology , Protein Serine-Threonine Kinases/metabolism , Self Tolerance/immunology , Thymocytes/immunology , Animals , B7-1 Antigen/metabolism , Cell Differentiation , Dendritic Cells/metabolism , Epithelial Cells/metabolism , Gene Expression , Immunophenotyping , Male , Mice , Mice, Transgenic , Models, Immunological , Mutation , Phenotype , Protein Serine-Threonine Kinases/genetics , Receptors, Antigen, T-Cell, alpha-beta/metabolism , Stromal Cells/metabolism , T-Lymphocytes, Regulatory/cytology , T-Lymphocytes, Regulatory/immunology , T-Lymphocytes, Regulatory/metabolism , Thymocytes/metabolism , Thymus Gland/immunology , Thymus Gland/metabolism , NF-kappaB-Inducing KinaseABSTRACT
Understanding the cellular dynamics of Aire-expressing lineage(s) among medullary thymic epithelial cells (AEL-mTECs) is essential for gaining insight into the roles of Aire in establishment of self-tolerance. In this study, we monitored the maturation program of AEL-mTECs by temporal lineage tracing, in which bacterial artificial chromosome transgenic mice expressing tamoxifen-inducible Cre recombinase under control of the Aire regulatory element were crossed with reporter strains. We estimated that the half-life of AEL-mTECs subsequent to Aire expression was â¼7-8 d, which was much longer than that reported previously, owing to the existence of a post-Aire stage. We found that loss of Aire did not alter the overall lifespan of AEL-mTECs, inconsistent with the previous notion that Aire expression in medullary thymic epithelial cells (mTECs) might result in their apoptosis for efficient cross-presentation of self-antigens expressed by AEL-mTECs. In contrast, Aire was required for the full maturation program of AEL-mTECs, as exemplified by the lack of physiological downregulation of CD80 during the post-Aire stage in Aire-deficient mice, thus accounting for the abnormally increased CD80(high) mTECs seen in such mice. Of interest, increased CD80(high) mTECs in Aire-deficient mice were not mTEC autonomous and were dependent on cross-talk with thymocytes. These results further support the roles of Aire in the differentiation program of AEL-mTECs.
Subject(s)
Cell Differentiation/immunology , Cell Lineage/immunology , Epithelial Cells/immunology , Transcription Factors/immunology , Animals , Apoptosis/genetics , Apoptosis/immunology , Autoantigens/immunology , Autoantigens/metabolism , B7-1 Antigen/immunology , B7-1 Antigen/metabolism , Cell Differentiation/genetics , Cell Lineage/genetics , Cells, Cultured , Cross-Priming/genetics , Cross-Priming/immunology , Epithelial Cells/metabolism , Flow Cytometry , Immunohistochemistry , Kinetics , Luminescent Proteins/genetics , Luminescent Proteins/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Thymocytes/cytology , Thymocytes/immunology , Thymocytes/metabolism , Thymus Gland/cytology , Time Factors , Transcription Factors/genetics , Transcription Factors/metabolism , AIRE ProteinABSTRACT
BACKGROUND: Hypoandrogenemia is associated with an increased risk of ischemic diseases. Because actions of androgens are exerted through androgen receptor (AR) activation, we studied hind-limb ischemia in AR knockout mice to elucidate the role of AR in response to ischemia. METHODS AND RESULTS: Both male and female AR knockout mice exhibited impaired blood flow recovery, more cellular apoptosis, and a higher incidence of autoamputation after ischemia. In ex vivo and in vivo angiogenesis studies, AR-deficient vascular endothelial cells showed reduced angiogenic capability. In ischemic limbs of AR knockout mice, reductions in the phosphorylation of the Akt protein kinase and endothelial nitric oxide synthase were observed despite a robust increase in hypoxia-inducible factor 1α and vascular endothelial cell growth factor (VEGF) gene expression. In in vitro studies, siRNA-mediated ablation of AR in vascular endothelial cells blunted VEGF-stimulated phosphorylation of Akt and endothelial nitric oxide synthase. Immunoprecipitation experiments documented an association between AR and kinase insert domain protein receptor that promoted the recruitment of downstream signaling components. CONCLUSIONS: These results document a physiological role of AR in sex-independent angiogenic potency and provide evidence of novel cross-talk between the androgen/AR signaling and VEGF/kinase insert domain protein receptor signaling pathways.
Subject(s)
Ischemia/physiopathology , Neovascularization, Physiologic/physiology , Receptors, Androgen/genetics , Receptors, Androgen/metabolism , Vascular Endothelial Growth Factor Receptor-2/metabolism , Amputation Stumps/pathology , Animals , Apoptosis/physiology , Capillaries/physiology , Female , Feminization/genetics , Feminization/metabolism , Hindlimb/blood supply , Hindlimb/pathology , Human Umbilical Vein Endothelial Cells , Humans , Ischemia/metabolism , Ischemia/pathology , Male , Mice , Mice, Knockout , Muscle, Skeletal/blood supply , Muscle, Skeletal/pathology , Nitric Oxide Synthase Type III/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Proto-Oncogene Proteins pp60(c-src)/metabolism , Receptor Cross-Talk/physiology , Signal Transduction/physiologyABSTRACT
OBJECTIVES: Periodontitis, commonly associated with Porphyromonas gingivalis (Pg), involves intricate alterations of oral intercellular interactions, in which extracellular vesicles (EVs) play a pivotal role. The understanding of the miRNA profiles in the EVs derived from Pg-infected cells (Pg-EVs) remains incomplete despite acknowledging their importance in intercellular communication during periodontitis. Therefore, our objective was to identify and characterize the miRNAs enriched in Pg-EVs. METHODS: Microarray analysis was conducted to examine the miRNA profiles in the EVs derived from Pg-infected THP-1 cells. We compared the identified miRNAs with those upregulated in the EVs after stimulation with LPS. Additionally, we explored how inhibiting TLR signaling during Pg infection affects the transcription of specific miRNAs. We investigated the unique sequence motifs specific to the miRNAs concentrated in Pg-EVs. RESULTS: The levels of eleven miRNAs, including miR-155, were increased in Pg-EVs compared with those elevated after LPS stimulation. The Pg-induced miR-155 upregulation via TLR2 but not TLR4 signaling suggests the influence of TLR signaling on the miRNA composition of EVs. Furthermore, the miRNAs upregulated in Pg-EVs contained AGAGGG and GRGGSGC sequence motifs. CONCLUSIONS: Our findings demonstrate that Pg-induced alterations in EV-containing miRNA composition occur in a TLR4-independent manner. Notably, the concentrated miRNAs in Pg-EVs harbor specific motifs with a high G + C content within their sequences. The upregulation of specific miRNAs in EVs under infectious conditions suggests the influence of both innate immune receptor signals and miRNA sequence characteristics.
Subject(s)
Extracellular Vesicles , MicroRNAs , Porphyromonas gingivalis , Extracellular Vesicles/metabolism , Extracellular Vesicles/genetics , MicroRNAs/metabolism , MicroRNAs/genetics , Porphyromonas gingivalis/metabolism , Porphyromonas gingivalis/genetics , Humans , Signal Transduction , Bacteroidaceae Infections/microbiology , Bacteroidaceae Infections/metabolism , Bacteroidaceae Infections/genetics , Lipopolysaccharides/pharmacology , Periodontitis/microbiology , Periodontitis/metabolism , Periodontitis/genetics , Up-Regulation , Microarray AnalysisABSTRACT
Head and neck squamous cell carcinoma (HNSCC) is a significant healthcare burden globally. Previous research using single-cell transcriptome analysis identified TGFBI as a crucial marker for the partial-epithelial-mesenchymal transition (partial-EMT) program. However, the precise role of TGFBI in HNSCC progression remains unclear. Therefore, our study aimed to clarify the impact of TGFBI on the malignant behavior of HNSCC cells. Through RNA-sequencing data from the TCGA database, we validated that increased TGFBI expression correlates with a higher occurrence of lymph node metastasis and unfavorable prognosis in HNSCC cases. Functional experiments demonstrated that TGFBI overexpression enhances the ability of sphere formation, indicating stem-cell-like properties. Conversely, TGFBI depletion reduces sphere formation and suppresses the expression of cancer stem cell (CSC) markers. RNA-sequencing analysis of TGFBI-overexpressing and control HNSCC cells revealed TAGLN as a downstream effector mediating TGFBI-induced sphere formation. Remarkably, TAGLN depletion abolished TGFBI-induced sphere formation, while its overexpression rescued the suppressed sphere formation caused by TGFBI depletion. Moreover, elevated TAGLN expression showed correlations with the expression of TGFBI and partial-EMT-related genes in HNSCC cases. In conclusion, our findings suggest that TGFBI may promote CSC properties through the upregulation of TAGLN. These novel insights shed light on the involvement of the TGFBI-TAGLN axis in HNSCC progression and hold implications for the development of targeted therapies.
Subject(s)
Carcinoma, Squamous Cell , Head and Neck Neoplasms , Humans , Squamous Cell Carcinoma of Head and Neck/pathology , Carcinoma, Squamous Cell/pathology , Head and Neck Neoplasms/pathology , Cell Line, Tumor , Neoplastic Stem Cells/metabolism , Epithelial-Mesenchymal Transition/genetics , RNA/metabolism , Gene Expression Regulation, NeoplasticABSTRACT
It has recently become clear that signals mediated by members of the TNFR superfamily, including lymphotoxin-ß receptor (LTßR), receptor activator for NF-κB (RANK), and CD40, play essential roles in organizing the integrity of medullary thymic epithelial cells (mTECs) required for the establishment of self-tolerance. However, details of the mechanism responsible for the unique and cooperative action of individual and multiple TNFR superfamily members during mTEC differentiation still remain enigmatic. In this study, we show that the LTßR signal upregulates expression of RANK in the thymic stroma, thereby promoting accessibility to the RANK ligand necessary for mTEC differentiation. Cooperation between the LTßR and RANK signals for optimal mTEC differentiation was underscored by the exaggerated defect of thymic organogenesis observed in mice doubly deficient for these signals. In contrast, we observed little cooperation between the LTßR and CD40 signals. Thus, the LTßR signal exhibits a novel and unique function in promoting RANK activity for mTEC organization, indicating a link between thymic organogenesis mediated by multiple cytokine signals and the control of autoimmunity.
Subject(s)
Cell Differentiation/immunology , Epithelial Cells/cytology , Lymphotoxin beta Receptor/metabolism , Organogenesis , Receptor Activator of Nuclear Factor-kappa B/biosynthesis , Signal Transduction , Thymus Gland/embryology , Animals , CD40 Antigens/metabolism , Embryo, Mammalian , Flow Cytometry , Gene Expression , Gene Expression Regulation , Immunohistochemistry , Mice , Mice, Knockout , Reverse Transcriptase Polymerase Chain Reaction , Self Tolerance/immunology , Thymus Gland/cytology , Thymus Gland/metabolismABSTRACT
It recently has been reported that partial-epithelial-mesenchymal transition (p-EMT) program is associated with metastasis in head and neck squamous cell carcinoma (HNSCC). We previously have identified POSTN (which encodes periostin) as an invasion-promoting molecule in HNSCC. Interestingly, POSTN expression is frequently observed in cancer cells with higher p-EMT score by using a previous single-cell transcriptomic data of HNSCC cases. Although it is known that POSTN has 11 splicing variants, the role of them has not been determined in HNSCC. Here, we found that HNSCC cells with EMT features expressed POSTN isoforms, Iso3 (lacking exon 17 and 21) and Iso5 (lacking exon 17), whereas fibroblast expressed Iso3 and Iso4 (lacking exon 17, 18, and 21). The expression of POSTN Iso3 and Iso4 are known to be widely observed in various cell types including stromal cells. Therefore, we focused on the role of novel cancer cell-derived POSTN isoform, Iso5, in HNSCC. Single overexpression of POSTN Iso5 as well as Iso3 promoted invasion. Surprisingly, Iso5 synergistically promoted invasion together with Iso3. Notably, Iso5 as well as Iso3 upregulated p-EMT-related genes. We suggest that a novel cancer-specific POSTN isoform lacking exon 17 (Iso5) can be a useful marker for detecting cancer cells undergoing p-EMT. Moreover, a POSTN Iso5 can be a novel target for diagnosis and therapy in HNSCC.
Subject(s)
Cell Adhesion Molecules , Head and Neck Neoplasms , Humans , Cell Line, Tumor , Epithelial-Mesenchymal Transition , Gene Expression Regulation, Neoplastic , Head and Neck Neoplasms/genetics , Protein Isoforms/genetics , Squamous Cell Carcinoma of Head and Neck/genetics , Cell Adhesion Molecules/metabolismABSTRACT
BACKGROUND: Comprehending the molecular mechanisms underlying head and neck squamous cell carcinoma (HNSCC) is vital for the development of effective treatment strategies. Deubiquitinating enzymes (DUBs), which regulate ubiquitin-dependent pathways, are potential targets for cancer therapy because of their structural advantages. Here we aimed to identify a potential target for HNSCC treatment among DUBs. METHODS: A screening process was conducted using RNA sequencing data and clinical information from HNSCC patients in the TCGA database. A panel of 88 DUBs was analyzed to identify those associated with poor prognosis. Subsequently, HNSCC cells were modified to overexpress specific DUBs, and their effects on cell proliferation and invasion were evaluated. In vivo experiments were performed to validate the findings. RESULTS: In HNSCC patients, USP10, USP14, OTUB1, and STAMBP among the screened DUBs were associated with a poor prognosis. Among them, OTUB1 showed the most aggressive characteristics in both in vitro and in vivo experiments. Additionally, OTUB1 regulated the stability and nuclear localization of YAP1, a substrate involved in cell proliferation and invasion. Notably, OTUB1 expression exhibited a positive correlation with the HNSCC-YAP score in HNSCC cells. CONCLUSIONS: This study highlights the critical role of OTUB1 in HNSCC progression via modulating YAP1. Targeting the OTUB1-YAP1 axis holds promise as a potential therapeutic strategy for HNSCC treatment.
Subject(s)
Deubiquitinating Enzymes , Head and Neck Neoplasms , Squamous Cell Carcinoma of Head and Neck , YAP-Signaling Proteins , Humans , Cell Line, Tumor , Head and Neck Neoplasms/genetics , Squamous Cell Carcinoma of Head and Neck/genetics , Ubiquitin Thiolesterase , Deubiquitinating Enzymes/metabolism , YAP-Signaling Proteins/metabolismABSTRACT
Dental implants made of titanium are commonly used. Although titanium implants succeed by osseointegration with bone, the detailed molecular mechanism of osseointegration is unclear. To clarify the involvement of microRNA (miRNA) in the acquisition of osseointegration on titanium, here we compared the miRNA expression profiles of mouse osteoblast-like cells (MC3T3-E1) cultured on titanium-, gold-, and stainless steel-coating glass dishes by microarray analysis. Three kinds of metals, namely titanium, gold, and stainless steel, were coated on the surface of the glass dishes by sputtering with similar roughness and shape of their surface. After MC3T3-E1 cells were cultured on the dishes without coating or coating with titanium, gold, or stainless steel for 6 h, total RNA was extracted, and miRNA expression was analyzed by microarray. To confirm the expression of the selected miRNA during osteogenic differentiation of MC3T3-E1 cells, real-time PCR analysis was performed. Furthermore, the effects of selected miRNA were examined by ectopic overexpression in MC3T3-E1 cells. The microarray analysis revealed that the expressions of miR-155-5p and miR-7023-3p were significantly increased in MC3T3-E1 cells cultured on titanium-coating glass dishes, compared to non-coating, gold-, and stainless steel-coating glass dishes. Interestingly, miR-155-5p was upregulated during osteogenic differentiation of MC3T3-E1 cells. Furthermore, overexpression of miR-155-5p enhanced the expression of Runx2 and Col1a1. In this study, miR-155-5p may be involved in the acquisition of osseointegration on titanium implant via upregulating osteogenic differentiation-related genes.
Subject(s)
Dental Implants , MicroRNAs , Animals , Cell Differentiation/genetics , Core Binding Factor Alpha 1 Subunit/genetics , Gold/pharmacology , Mice , MicroRNAs/genetics , MicroRNAs/metabolism , Osseointegration , Osteoblasts , Osteocalcin/genetics , Osteogenesis/genetics , Stainless Steel/pharmacology , Titanium/pharmacologyABSTRACT
BACKGROUND: It is well-established that cervical lymph node metastasis is the most important prognostic factor in head and neck squamous cell carcinoma (HNSCC). Cancer cells invade the underlying stroma during metastasis by breaching the basement membrane. HIGHLIGHT: The ability to metastasize is a key hallmark of cancer progression and this characteristic can be attained by undergoing epithelial-to-mesenchymal transition (EMT). EMT is a biological process in which epithelial cells lose their epithelial features and gain mesenchymal features. Recent evidence reports the intermediate state in the induction of EMT and partial-EMT. Notably, the partial-EMT phenotype is more aggressive than the complete EMT phenotype. However, the role of partial-EMT is not fully understood. CONCLUSION: In this review, we highlight the features of partial-EMT in HNSCC by summarizing previous studies. Moreover, we discuss the therapeutic potential for targeting partial-EMT.
Subject(s)
Carcinoma, Squamous Cell , Head and Neck Neoplasms , Carcinoma, Squamous Cell/genetics , Epithelial-Mesenchymal Transition/genetics , Humans , Lymphatic Metastasis , Squamous Cell Carcinoma of Head and NeckABSTRACT
Controlled proteolytic degradation of specialized junctional structures, corneodesmosomes, by epidermal proteases is an essential process for physiological desquamation of the skin. Corneodesmosin (CDSN) is an extracellular component of corneodesmosomes and, although considerable debate still exists, genetic studies have suggested that the CDSN gene in the major psoriasis-susceptibility locus (PSORS1) may be responsible for susceptibility to psoriasis, a human skin disorder characterized by excessive growth and aberrant differentiation of keratinocytes. CDSN is also expressed in the inner root sheath of hair follicles, and a heterozygous nonsense mutation of the CDSN gene in humans is associated with scalp-specific hair loss of poorly defined etiology. Here, we have investigated the pathogenetic roles of CDSN loss of function in the development of skin diseases by generating a mouse strain with targeted deletion of the Cdsn gene. Cdsn-deficient mouse skin showed detachment of the stratum corneum from the underlying granular layer and/or detachment within the upper granular layers due to the disrupted integrity of the corneodesmosomes. When grafted onto immunodeficient mice, Cdsn-deficient skin showed rapid hair loss together with epidermal abnormalities resembling psoriasis. These results underscore the essential roles of CDSN in hair physiology and suggest functional relevance of CDSN gene polymorphisms to psoriasis susceptibility.
Subject(s)
Gene Deletion , Gene Targeting , Glycoproteins/genetics , Hair/physiology , Skin Physiological Phenomena/genetics , Animals , Desmosomes/metabolism , Desmosomes/pathology , Glycoproteins/deficiency , Hair/growth & development , Mice , Phenotype , Psoriasis/pathology , Skin Abnormalities/genetics , Skin Abnormalities/ultrastructure , Skin TransplantationABSTRACT
(1) Background: OK-432 is a penicillin-killed, lyophilized formulation of a low-toxicity strain (Su) of Streptococcus pyogenes (Group A). It is a potent immunotherapy agent for several types of cancer, including oral cancer. We previously showed that (i) OK-432 treatment induces a high amount of IFN-? production from peripheral blood mononuclear cells (PBMCs), and (ii) conditioned medium (CM) from oral cancer cells suppresses both the IFN-? production and cytotoxic activity of PBMCs driven by OK-432. The aim of this study was to determine the inhibitory mechanism of OK-432-induced IFN-? production from PBMCs by CM. (2) Methods: We performed cDNA microarray analysis, quantitative RT-PCR, and ELISA to reveal the inhibitory mechanism of CM. (3) Results: We found that CD40 plays a key role in IFN-? production via IL-12 production. Although OK-432 treatment upregulated the expression levels of the IL-12p40, p35, and CD40 genes, CM from oral cancer cells downregulate these genes. The amount of IFN-? production by OK-432 treatment was decreased by an anti-CD40 neutralizing antibody. (4) Conclusions: Our study suggests that uncertain soluble factor(s) produced from oral cancer cells may inhibit IFN-? production from PBMCs via suppressing the CD40/CD40L-IL-12 axis.
ABSTRACT
The ability of cancer cells to undergo partial-epithelial mesenchymal transition (p-EMT), rather than complete EMT, poses a higher metastatic risk. Although Fusobacterium nucleatum mainly inhabits in oral cavity, attention has been focused on the F. nucleatum involvement in colorectal cancer development. Here we examined the p-EMT regulation by F. nucleatum in oral squamous cell carcinoma (OSCC) cells. We cultured OSCC cells with epithelial, p-EMT or EMT phenotype with live or heat-inactivated F. nucleatum. Expression of the genes involved in epithelial differentiation, p-EMT and EMT were examined in OSCC cells after co-culture with F. nucleatum by qPCR. Cell growth and invasion of OSCC cells were also examined. Both live and heat-inactivated F. nucleatum upregulated the expression of p-EMT-related genes in OSCC cells with epithelial phenotype, but not with p-EMT or EMT phenotype. Moreover, F. nucleatum promoted invasion of OSCC cells with epithelial phenotype. Co-culture with other strains of bacteria other than Porphyromonas gingivalis did not alter p-EMT-related genes in OSCC cells with epithelial phenotype. F. nucleatum infection may convert epithelial to p-EMT phenotype via altering gene expression in OSCC. Oral hygiene managements against F. nucleatum infection may contribute to reduce the risk for an increase in metastatic ability of OSCC.
Subject(s)
Biomarkers, Tumor/genetics , Carcinoma, Squamous Cell/virology , Fusobacterium Infections/complications , Fusobacterium nucleatum/pathogenicity , Mouth Neoplasms/virology , Carcinoma, Squamous Cell/genetics , Cell Line, Tumor , Cell Movement , Cell Proliferation , Disease Progression , Epithelial-Mesenchymal Transition , Fusobacterium Infections/genetics , Gene Expression Regulation, Neoplastic , Humans , Mouth Neoplasms/genetics , Neoplasm Metastasis , Oral HygieneABSTRACT
Factors that determine the spectrum of target organs involved in autoimmune destruction are poorly understood. Although loss of function of autoimmune regulator (AIRE) in thymic epithelial cells is responsible for autoimmunity, the pathogenic roles of AIRE in regulating target-organ specificity remain elusive. In order to gain insight into this issue, we have established NOD mice, an animal model of type 1 diabetes caused by autoimmune attack against beta cell islets, in which Aire has been abrogated. Remarkably, acinar cells rather than beta cell islets were the major targets of autoimmune destruction in Aire-deficient NOD mice, and this alteration of intra-pancreatic target-organ specificity was associated with production of autoantibody against pancreas-specific protein disulfide isomerase (PDIp), an antigen expressed predominantly by acinar cells. Consistent with this pathological change, the animals were resistant to the development of diabetes. The results suggest that Aire not only is critical for the control of self-tolerance but is also a strong modifier of target-organ specificity through regulation of T cell repertoire diversification. We also demonstrated that transcriptional expression of PDIp was retained in the Aire-deficient NOD thymus, further supporting the concept that Aire may regulate the survival of autoreactive T cells beyond transcriptional control of self-protein expression in the thymus.
Subject(s)
Pancreas/pathology , Transcription Factors/genetics , Transcription Factors/physiology , Animals , Autoimmune Diseases/genetics , Diabetes Mellitus/genetics , Diabetes Mellitus/prevention & control , Disease Models, Animal , Female , Genetic Predisposition to Disease , Male , Mice , Mice, Inbred NOD , Mice, Transgenic , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Thymus Gland/metabolism , AIRE ProteinABSTRACT
The discovery of Aire-dependent transcriptional control of many tissue-restricted self-antigen (TRA) genes in thymic epithelial cells in the medulla (medullary thymic epithelial cells, mTECs) has raised the intriguing question of how the single Aire gene can influence the transcription of such a large number of TRA genes within mTECs. From a mechanistic viewpoint, there are two possible models to explain the function of Aire in this action. In the first model, TRAs are considered to be the direct target genes of Aire's transcriptional activity. In this scenario, the lack of Aire protein within cells would result in the defective TRA gene expression, while the maturation program of mTECs would be unaffected in principle. The second model hypothesizes that Aire is necessary for the maturation program of mTECs. In this case, we assume that the mTEC compartment does not mature normally in the absence of Aire. If acquisition of the properties of TRA gene expression depends on the maturation status of mTECs, a defect of such an Aire-dependent maturation program in Aire-deficient mTECs can also result in impaired TRA gene expression. In this brief review, we will focus on these two contrasting models for the roles of Aire in controlling the expression of TRAs within mTECs.