ABSTRACT
Solid-state [2+2] photodimerization of eniminium ions oriented in a head-to-tail manner controlled by cation-π interactions produced synHT dimers in high yields. As the resulting dimer is readily converted to 1,3-diacetylcyclobutane, the iminium serves as a removable orientation-controlling group for the conjugated ketones.
ABSTRACT
To identify a biomarker for the early diagnosis of enzootic bovine leukosis (EBL) caused by bovine leukemia virus (BLV), we investigated the expression of a microRNA, bta-miR-375, in cattle serum. Using quantitative reverse-transcriptase PCR analysis, we measured bta-miR-375 levels in 27 samples from cattle with EBL (EBL cattle), 45 samples from animals infected with BLV but showing no clinical signs (NS cattle), and 30 samples from cattle uninfected with BLV (BLV negative cattle). In this study, we also compared the kinetics of bta-miR-375 with those of the conventional biomarkers of proviral load (PVL), lactate dehydrogenase (LDH), and thymidine kinase (TK) from the no-clinical-sign phase until EBL onset in three BLV-infected Japanese black (JB) cattle. Bta-miR-375 expression was higher in NS cattle than in BLV negative cattle (P < 0.05) and greater in EBL cattle than in BLV negative and NS cattle (P < 0.0001 for both comparisons). Receiver operating characteristic curves demonstrated that bta-miR-375 levels distinguished EBL cattle from NS cattle with high sensitivity and specificity. In NS cattle, bta-miR-375 expression was increased as early as at 2 months before EBL onset-earlier than the expression of PVL, TK, or LDH isoenzymes 2 and 3. These results suggest that serum miR-375 is a promising biomarker for the early diagnosis of EBL.
Subject(s)
Biomarkers , Early Diagnosis , Enzootic Bovine Leukosis , Leukemia Virus, Bovine , MicroRNAs , Animals , Cattle , Enzootic Bovine Leukosis/diagnosis , Enzootic Bovine Leukosis/blood , Enzootic Bovine Leukosis/virology , MicroRNAs/blood , MicroRNAs/genetics , Biomarkers/blood , Leukemia Virus, Bovine/genetics , ROC Curve , L-Lactate Dehydrogenase/bloodABSTRACT
Spontaneous tumors are a major cause of death in cats. Treatment of human tumors has progressed dramatically in the past decade, partly due to the success of immunotherapies using immune checkpoint inhibitors, such as anti-programmed death 1 (PD-1) and anti-PD-ligand 1 (PD-L1) antibodies. However, little is known about the PD-1 pathway and its association with tumor disease in cats. This study investigated the applicability of anti-PD-1/PD-L1 therapy in feline tumors. We first determined the complete coding sequence of feline PD-L1 and PD-L2, and found that the deduced amino acid sequences of feline PD-L1/PD-L2 share high sequence identities (66-83%) with orthologs in other mammalian species. We prepared recombinant feline PD-1, PD-L1, and PD-L2 proteins and confirmed receptor-ligand binding between PD-1 and PD-L1/PD-L2 using flow cytometry. Next, we established an anti-feline PD-L1 monoclonal antibody (clone CL1Mab-7) to analyze the expression of PD-L1. Flow cytometry using CL1Mab-7 revealed the cell surface expression of PD-L1 in a feline macrophage (Fcwf-4) and five mammary adenocarcinoma cell lines (FKNp, FMCm, FYMp, FONp, and FONm), and showed that PD-L1 expression was upregulated by interferon-γ stimulation. Finally, immunohistochemistry using CL1Mab-7 also showed PD-L1 expression in feline squamous cell carcinoma (5/5, 100%), mammary adenocarcinoma (4/5, 80%), fibrosarcoma (5/5, 100%), and renal cell carcinoma (2/2, 100%) tissues. Our results strongly encourage further investigations of the PD-1/PD-L1 pathway as a potential therapeutic target for feline tumors.
Subject(s)
Adenocarcinoma , Carcinoma, Squamous Cell , Animals , Cats , Humans , Adenocarcinoma/pathology , Adenocarcinoma/veterinary , Antibodies, Monoclonal , B7-H1 Antigen/metabolism , Carcinoma, Squamous Cell/pathology , Carcinoma, Squamous Cell/veterinary , Immune Checkpoint Proteins , Immunohistochemistry , Ligands , Programmed Cell Death 1 Ligand 2 Protein/genetics , Cat DiseasesABSTRACT
Bovine leukemia virus (BLV), a retrovirus, infects B cells of ruminants and is integrated into the host genome as a provirus for lifelong infection. After a long latent period, 1% to 5% of BLV-infected cattle develop aggressive lymphoma, enzootic bovine leukosis (EBL). Since the clonal expansion of BLV-infected cells is essential for the development of EBL, the clonality of proviral integration sites could be a molecular marker for diagnosis and early prediction of EBL. Recently, we developed Rapid Amplification of the Integration Site without Interference by Genomic DNA Contamination (RAISING) and an analysis software of clonality value (CLOVA) to analyze the clonality of transgene-integrated cells. RAISING-CLOVA is capable of assessing the risk of adult T-cell leukemia/lymphoma development in human T-cell leukemia virus-I-infected individuals through the clonality analysis of proviral integration sites. Thus, we herein examined the performance of RAISING-CLOVA for the clonality analysis of BLV-infected cells and conducted a comprehensive clonality analysis by RAISING-CLOVA in EBL and non-EBL cattle. RAISING-CLOVA targeting BLV was a highly accurate and reproducible method for measuring the clonality value. The comprehensive clonality analysis successfully distinguished EBL from non-EBL specimens with high sensitivity and specificity. A longitudinal clonality analysis in BLV-infected sheep, an experimental model of lymphoma, also confirmed the effectiveness of RAISING-CLOVA for early detection of EBL development. Therefore, our study emphasizes the usefulness of RAISING-CLOVA as a routine clinical test for monitoring virus-related cancers. IMPORTANCE Bovine leukemia virus (BLV) infection causes aggressive B-cell lymphoma in cattle and sheep. The virus has spread to farms around the world, causing significant economic damage to the livestock industry. Thus, the identification of high-risk asymptomatic cattle before they develop lymphoma can be effective in reducing the economic damage. Clonal expansion of BLV-infected cells is a promising marker for the development of lymphoma. Recently, we have developed a high-throughput method to amplify random integration sites of transgenes in host genomes and analyze their clonality, named as RAISING-CLOVA. As a new application of our technology, in this study, we demonstrate the value of the RAISING-CLOVA method for the diagnosis and early prediction of lymphoma development by BLV infection in cattle. RAISING-CLOVA is a reliable technology for monitoring the clonality of BLV-infected cells and would contribute to reduce the economic losses by EBL development.
Subject(s)
Enzootic Bovine Leukosis , Leukemia Virus, Bovine , Lymphoma , Cattle , Humans , Animals , Sheep , Leukemia Virus, Bovine/genetics , Enzootic Bovine Leukosis/diagnosis , Proviruses/genetics , Virus IntegrationABSTRACT
Bovine viral diarrhea virus (BVDV) causes substantial economic losses in the livestock industry worldwide. Plasmids encoding the BVDV E2 protein are potential DNA vaccines against BVDV, but their immunogenicity has been insufficient. Here, we investigated the adjuvant effect of CD40 and CD63 plasmids on the immune responses to a BVDV E2 DNA vaccine in mice. We constructed pUMVC4a-based plasmids encoding the BVDV E2 protein (pE2), mouse CD40 (pCD40), or mouse CD63 (pCD63). Protein expression by each plasmid was confirmed through Western blot analysis and immunofluorescence staining of cultured cell lines. BALB/c mice were immunized intradermally twice with pE2 in combination with, or without, pCD40 or pCD63, with 3 weeks between the two doses. pE2 with pCD40 induced significantly higher neutralizing antibody titers against BVDV than pE2 alone. pE2 with pCD63 induced significantly higher anti-E2 IgG2a antibody titers than pE2 alone. Furthermore, pE2 with pCD40 or pCD63 induced significantly increased lymphocyte proliferation and interferon (IFN)-γ production in response to BVDV, compared with E2 alone. These results suggest that a plasmid encoding CD40 or CD63 can be used as an adjuvant to enhance immune responses to DNA vaccines against BVDV.
Subject(s)
Bovine Virus Diarrhea-Mucosal Disease , Cattle Diseases , Diarrhea Virus 1, Bovine Viral , Diarrhea Viruses, Bovine Viral , Rodent Diseases , Vaccines, DNA , Viral Vaccines , Adjuvants, Immunologic , Animals , Antibodies, Viral , Bovine Virus Diarrhea-Mucosal Disease/prevention & control , Cattle , Diarrhea/veterinary , Diarrhea Virus 1, Bovine Viral/genetics , Immunity , Mice , Mice, Inbred BALB C , Plasmids/genetics , Viral Envelope ProteinsABSTRACT
Bovine leukemia virus (BLV) is an oncogenic virus belonging to the genus Deltaretrovirus and is the causative agent of enzootic bovine leukosis. Proviral load (PVL) determined by real-time quantitative PCR (qPCR) is now widely used as an indicator of not only BLV infection, but also BLV disease progression. To interpret PVLs determined by different qPCRs used in Japan, we compared a chimeric cycling probe-based qPCR, CY415, targeting the BLV tax region; a TaqMan probe-based qPCR, RC202, targeting the BLV pol region; and a TaqMan probe-based qPCR, CoCoMo, targeting the BLV long terminal repeat (LTR) region. Whole-blood samples collected from 317 naturally BLV-infected cattle (165 Holstein-Friesian and 152 Japanese Black) and tumor tissue samples collected from 32 cattle at a meat inspection center were used. The PVLs determined by each qPCR were strongly correlated. However, the PVL and the proportion of BLV-infected cells determined by RC202 or CoCoMo were significantly higher than those determined by CY415. Genetic analysis of three tumor tissue samples revealed that LTR region mutations or a deletion affected the PVL determined by CoCoMo. These results suggest that the TaqMan-based RC202 or CoCoMo qPCR is better than CY415 for BLV PVL analysis. However, qPCR target region mutations were not rare in tumors and could hamper PVL analysis by using qPCR.
Subject(s)
Enzootic Bovine Leukosis , Leukemia Virus, Bovine , Animals , Cattle , Japan , Leukemia Virus, Bovine/genetics , Proviruses/genetics , Real-Time Polymerase Chain Reaction/methods , Viral Load/methodsABSTRACT
Borrelia miyamotoi infection is an emerging tick-borne disease that causes hard tick-borne relapsing fever. B. miyamotoi is transmitted through the bite of ticks, including Ixodes persulcatus. Although accumulating evidence suggests that tick salivary proteins enhance the infectivity of other tick-borne pathogens, the association of B. miyamotoi with tick-derived proteins remains unknown. In this study, the effect of I. persulcatus sialostatin L2 (Ip-sL2), a tick-derived cystatin, on specific immunity to B. miyamotoi was preliminarily investigated in vitro. Mice were immunized with heat-killed B. miyamotoi and in vitro analyses of the splenocytes of the immunized mice indicated that the expression levels of the activation markers of CD11c+ and CD3+ cells were significantly upregulated by B. miyamotoi stimulation. Spleen cells from B. miyamotoi-immunized mice were used to determine whether Ip-sL2 regulates murine immune responses against B. miyamotoi. Treatment with Ip-sL2 in vitro inhibited the activation of CD11c+ and CD3+ cells as well as inflammatory cytokine production by cultured splenocytes. These findings show that Ip-sL2 has modulatory effects on murine immune responses to B. miyamotoi. Therefore, it is necessary to clarify in the future whether Ip-sL2 is involved in the enhanced infectivity of B. miyamotoi.
Subject(s)
Borrelia , Ixodes , Relapsing Fever , Tick-Borne Diseases , Animals , Arthropod Proteins , Ixodes/physiology , MiceABSTRACT
Recent evidence indicates that RNA-dependent RNA polymerase (RdRP) activity of human telomerase reverse transcriptase (hTERT) regulates expression of target genes and is directly involved in tumor formation in a telomere-independent manner. Non-canonical function of hTERT has been considered as a therapeutic target for cancer therapy. We have previously shown that hTERT phosphorylation at threonine 249 (p-hTERT), which promotes RdRP activity, is an indicator of an aggressive phenotype and poor prognosis in liver and pancreatic cancers, using two cohorts with small sample sizes with polyclonal p-hTERT antibody. To clarify the clinical relevance of p-hTERT, we developed a specific monoclonal antibody and determined the diagnostic and prognostic value of p-hTERT in cancer specimens using a large cohort. A monoclonal antibody for phosphorylated hTERT (p-hTERT) at threonine 249 was developed and validated. The antibody was used for the immunohistochemical staining of formalin-fixed, paraffin-embedded specimens from 1523 cases of lung, colon, stomach, pancreatic, liver, breast, and kidney cancers. We detected elevated p-hTERT expression levels in cases with a high mitotic activity, high pathological grade, and high nuclear pleomorphism. Elevated p-hTERT expression was an independent prognostic factor for lung, pancreatic, and liver cancers. Furthermore, p-hTERT expression was associated with immature and aggressive features, such as adenosquamous carcinoma (lung and pancreas), invasive type of cancer (lung), high serum alpha-fetoprotein level (liver), and triple-negative status (breast). In conclusion, RdRP activity indicated by p-hTERT expression predicts aggressive cancer phenotypes in various types of cancer. Thus, p-hTERT is a novel biomarker for the diagnosis of aggressive cancers with a poor prognosis. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
Subject(s)
Neoplasms , Telomerase , Antibodies, Monoclonal , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Humans , Neoplasms/genetics , Neoplasms/pathology , Phosphorylation , Prognosis , RNA-Dependent RNA Polymerase , Telomerase/genetics , Threonine/metabolismABSTRACT
Podoplanin is expressed in various human tumors where it promotes tumor progression, epithelial-mesenchymal transition, and distant metastasis. Podoplanin is also expressed in cancer-associated fibroblasts and induces tumor malignancy. The objective of this study was to evaluate podoplanin expression in various types of feline tumor tissues. Immunohistochemical analysis revealed that podoplanin was expressed in cells of 13/15 (87%) squamous cell carcinomas and 5/19 (26%) fibrosarcomas. Moreover, cancer-associated fibroblasts expressed podoplanin in most tumor types, including 18/21 (86%) mammary adenocarcinoma tissues. Our findings demonstrate that various types of feline tumor tissues expressed podoplanin, indicating the importance of the comparative aspects of podoplanin expression, which may be used as a novel research model for podoplanin biology.
Subject(s)
Carcinoma, Squamous Cell , Cat Diseases , Animals , Biomarkers, Tumor , Carcinoma, Squamous Cell/veterinary , Cats , Epithelial-Mesenchymal Transition , Immunohistochemistry , Membrane GlycoproteinsABSTRACT
Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis (EBL), a malignant B cell lymphoma. However, the mechanisms of BLV-associated lymphomagenesis remain poorly understood. Here, after deep sequencing, we performed comparative analyses of B cell microRNAs (miRNAs) in cattle infected with BLV and those without BLV. In BLV-infected cattle, BLV-derived miRNAs (blv-miRNAs) accounted for 38% of all miRNAs in B cells. Four of these blv-miRNAs (blv-miR-B1-5p, blv-miR-B2-5p, blv-miR-B4-3p, and blv-miR-B5-5p) had highly significant positive correlations with BLV proviral load (PVL). The read counts of 90 host-derived miRNAs (bta-miRNAs) were significantly down-regulated in BLV-infected cattle compared to those in uninfected cattle. Only bta-miR-375 had a positive correlation with PVL in BLV-infected cattle and was highly expressed in the B cell lymphoma tissue of EBL cattle. There were a few bta-miRNAs that correlated with BLV tax/rex gene expression; however, BLV AS1 expression had a significant negative correlation with many of the down-regulated bta-miRNAs that are important for tumor development and/or tumor suppression. These results suggest that BLV promotes lymphomagenesis via AS1 and blv-miRNAs, rather than tax/rex, by down-regulating the expression of bta-miRNAs that have a tumor-suppressing function, and this downregulation is linked to increased PVL.
Subject(s)
B-Lymphocytes/metabolism , Enzootic Bovine Leukosis/metabolism , Leukemia Virus, Bovine/isolation & purification , MicroRNAs/metabolism , Animals , B-Lymphocytes/cytology , Cattle , Proviruses/isolation & purification , Viral LoadABSTRACT
Maze procedure has achieved high cure rates and become the surgical golden standard for the treatment of atrial fibrillation. But, atrial arrhythmia after maze procedure is often persistent and drug-resistant. In these cases, diagnosis by electrophysiological study (EPS) and treatment by catheter ablation (ABL) are useful. In our hospital, maze procedure has been actively performed for mitral valve surgery with atrial arrhythmia. We examined the cases that required ABL after maze procedure in our hospital. We reported 2 such typical cases where ablation of cavo-tricuspid isthmus line (CTI) in the right atrium and left superior pulmonary vein-left atrial appendage space( LSPV-LAA ridge) in the left atrium was effective.
Subject(s)
Atrial Fibrillation , Catheter Ablation , Pulmonary Veins , Atrial Fibrillation/surgery , Humans , Maze Procedure , Pulmonary Veins/surgery , Treatment OutcomeABSTRACT
Ferritin 2 (FER2) is an iron storage protein, which has been shown to be critical for iron homeostasis during blood feeding and reproduction in ticks and is therefore suitable as a component for anti-tick vaccines. In this study, we identified the FER2 of Ixodes persulcatus, a major vector for zoonotic diseases such as Lyme borreliosis and tick-borne relapsing fever in Japan, and investigated its functions. Ixodes persulcatus-derived ferritin 2 (Ip-FER2) showed concentration-dependent iron-binding ability and high amino acid conservation, consistent with FER2s of other tick species. Vaccines containing the recombinant Ip-FER2 elicited a significant reduction of the engorgement weight of adult I. persulcatus. Interestingly, the reduction of engorgement weight was also observed in Ixodes ovatus, a sympatric species of I. persulcatus. In silico analyses of FER2 sequences of I. persulcatus and other ticks showed a greater similarity with I. scapularis and I. ricinus and lesser similarity with Hyalomma anatolicum, Haemaphysalis longicornis, Rhipicephalus microplus, and R. appendiculatus. Moreover, it was observed that the tick FER2 sequences possess conserved regions within the primary structures, and in silico epitope mapping analysis revealed that antigenic regions were also conserved, particularly among Ixodes spp ticks. In conclusion, the data support further protective tick vaccination applications using the Ip-FER2 antigens identified herein.
Subject(s)
Arthropod Proteins/genetics , Ferritins/genetics , Ixodes/genetics , Vaccines/genetics , Amino Acid Sequence , Animals , Arthropod Proteins/chemistry , Arthropod Proteins/metabolism , Base Sequence , Ferritins/chemistry , Ferritins/metabolism , Ixodes/metabolism , Phylogeny , Sequence Alignment , Vaccines/analysisABSTRACT
CD19 is a type I transmembrane glycoprotein belonging to the immunoglobulin superfamily. It is expressed in normal and neoplastic B cells, and it modulates the threshold of B cell activation for amplifying B cell receptor signaling. Blinatumomab (a CD3-CD19-bispecific T cell-engaging antibody) and tisagenlecleucel (genetically modified T cells that express a CD19 chimeric antigen receptor [CART-19]) provide significant benefits for patients with CD19-positive relapsed or refractory B cell malignancies. In this study, we first employed the Cell-Based Immunization and Screening (CBIS) method to produce anti-CD19 monoclonal antibodies using CD19-overexpressing cells for both immunization and screening. One established clone-C19Mab-1-proved to be useful in flow cytometry assays against lymphoma cell lines, such as BALL-1, P30/OHK, and Raji. Second, the extracellular domain of CD19 was immunized into mice, and enzyme-linked immunosorbent assays were performed for the first screening. One established clone-C19Mab-3-was determined to be useful for Western blotting and immunohistochemical analysis. Due to their complementary utility, a combination of C19Mab-1 (established using CBIS) and C19Mab-3 (established using conventional method) could be useful for the pathological analysis of CD19.
Subject(s)
Antibodies, Bispecific/pharmacology , Antigens, CD19/immunology , Lymphoma, B-Cell/drug therapy , Receptors, Antigen, T-Cell/immunology , Animals , Antibodies, Bispecific/immunology , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/pharmacology , Antigens, CD19/genetics , B-Lymphocytes/immunology , B-Lymphocytes/pathology , Flow Cytometry , Humans , Immunoglobulins/immunology , Lymphocyte Activation/drug effects , Lymphoma, B-Cell/immunology , Lymphoma, B-Cell/pathology , Mice , Receptors, Antigen, T-Cell/genetics , T-Lymphocytes/drug effects , T-Lymphocytes/immunologyABSTRACT
Bovine mycoplasmosis caused by Mycoplasma bovis results in pneumonia and mastitis in cattle. We previously demonstrated that the programmed death 1 (PD-1)/PD-ligand 1 (PD-L1) pathway is involved in immune dysfunction during M. bovis infection and that prostaglandin E2 (PGE2) suppressed immune responses and upregulated PD-L1 expression in Johne's disease, a bacterial infection in cattle. In this study, we investigated the role of PGE2 in immune dysfunction and the relationship between PGE2 and the PD-1/PD-L1 pathway in M. bovis infection. In vitro stimulation with M. bovis upregulated the expressions of PGE2 and PD-L1 presumably via Toll-like receptor 2 in bovine peripheral blood mononuclear cells (PBMCs). PGE2 levels of peripheral blood in infected cattle were significantly increased compared with those in uninfected cattle. Remarkably, plasma PGE2 levels were positively correlated with the proportions of PD-L1+ monocytes in M. bovis-infected cattle. Additionally, plasma PGE2 production in infected cattle was negatively correlated with M. bovis-specific interferon (IFN)-γ production from PBMCs. These results suggest that PGE2 could be one of the inducers of PD-L1 expression and could be involved in immunosuppression during M. bovis infection. In vitro blockade assays using anti-bovine PD-L1 antibody and a cyclooxygenase 2 inhibitor significantly upregulated the M. bovis-specific IFN-γ response. Our study findings might contribute to the development of novel therapeutic strategies for bovine mycoplasmosis that target PGE2 and the PD-1/PD-L1 pathway.
ABSTRACT
The telomerase reverse transcriptase is upregulated in the majority of human cancers and contributes directly to cell transformation. Here we report that hTERT is phosphorylated at threonine 249 during mitosis by the serine/threonine kinase CDK1. Clinicopathological analyses reveal that phosphorylation of hTERT at threonine 249 occurs more frequently in aggressive cancers. Using CRISPR/Cas9 genome editing, we introduce substitution mutations at threonine 249 in the endogenous hTERT locus and find that phosphorylation of threonine 249 is necessary for hTERT-mediated RNA dependent RNA polymerase (RdRP) activity but dispensable for reverse transcriptase and terminal transferase activities. Cap Analysis of Gene Expression (CAGE) demonstrates that hTERT phosphorylation at 249 regulates the expression of specific genes that are necessary for cancer cell proliferation and tumor formation. These observations indicate that phosphorylation at threonine 249 regulates hTERT RdRP and contributes to cancer progression in a telomere independent manner.
Subject(s)
CDC2 Protein Kinase/metabolism , Neoplasms/metabolism , Neoplasms/pathology , Telomerase/metabolism , Animals , CDC2 Protein Kinase/antagonists & inhibitors , CDC2 Protein Kinase/genetics , Cell Line, Tumor , Cell Proliferation , Cell Transformation, Neoplastic , Gene Expression Regulation, Neoplastic , HeLa Cells , Humans , Mice , Mitosis , Mutation , Neoplasms/genetics , Phosphorylation , RNA-Dependent RNA Polymerase/metabolism , Telomerase/genetics , ThreonineABSTRACT
Irradiation of HX (X=CF3 SO3 or CF3 CO2 ) salts of 1-aryl-4-pyridylbutadienes 1 a-1 c in the solid-state afforded syn head-to-tail dimers in good yields among a number of possible dimers, whereas irradiation of the neutral substrates gave a complex mixture or no products. A comparison of the X-ray crystal structures of the neutral compounds and the HX salts clarified that their orientation modes are head-to-head and head-to-tail, respectively. Moreover, while the distances between the two neighboring double bonds of the neutral compounds are relatively far apart from each other, those of HX salts are close together, satisfying Schmidt's requirement. These findings suggested that cation-π interactions between the pyridinium and aromatic rings are effective for the preorientation of the HX salts of substrates, leading to photodimers in high regio- and stereoselectivities.
ABSTRACT
STUDY DESIGN: Case-control study. OBJECTIVE: We aimed to identify predictors for latent myelopathy and to develop a diagnostic protocol based on these factors. SUMMARY OF BACKGROUND DATA: There is no diagnostic protocol for latent myelopathy to avoid misdiagnosis in patients complaining only of lower extremity symptoms. METHODS: This case-control study identified 791 patients discussed at conferences from April 2006 to August 2012. Overall, 460 patients complaining only of lower extremity symptoms and who underwent spine surgery were included as participants; 54 underwent surgery involving the cervical and thoracic vertebrae and were assigned to the cervical-thoracic group (C-T group); 406 underwent lumbar surgery and were assigned to the lumbar group (L group). RESULTS: By univariate analysis, age ≥67 years, patellar tendon (PT) hyperreflexia, Achilles tendon (AT) hyperreflexia, spastic gait, and gait inability were more common in the C-T group than in the L group. By multivariate analysis, age ≥67 years (OR, 8; Pâ=â0.001), AT hyperreflexia (OR, 20.5; Pâ<â0.001), spastic gait (OR, 225; Pâ<â0.001), and gait inability (OR, 64; Pâ<â0.001) were significant predictive factors. In patients with age ≥67 years, PT hyperreflexia, and/or AT hyperreflexia, the sensitivity for myelopathy diagnosis was 98%. In patients with spastic gait or gait inability, the specificity of myelopathy diagnosis was 96%. CONCLUSIONS: We analyzed factors that predict latent myelopathy in patients complaining only of lower extremity symptoms. We believe a diagnostic protocol based on the predictors shown in this study would contribute to the accurate diagnosis of latent myelopathy. LEVEL OF EVIDENCE: 4.
Subject(s)
Lower Extremity , Spinal Cord Diseases/diagnostic imaging , Adult , Aged , Bone Marrow Diseases , Case-Control Studies , Cervical Vertebrae/surgery , Diagnostic Errors , Female , Gait , Gait Disorders, Neurologic , Humans , Male , Middle Aged , Multivariate Analysis , Prognosis , Spinal Cord Diseases/surgery , Thoracic VertebraeABSTRACT
Pancreatic ductal adenocarcinoma (PDAC) is among the cancers with the poorest prognoses due to its highly malignant features. BTB and CNC homology 1 (BACH1) has been implicated in RAS-driven tumor formation. We focused on the role of BACH1 in PDAC, more than 90% of which have KRAS mutation. Knockdown of BACH1 in PDAC cell lines reduced cell migration and invasion, in part, by increasing E-cadherin expression, whereas its overexpression showed opposite effects. BACH1 directly repressed the expression of FOXA1 that is known to activate the expression of CDH1 encoding E-cadherin and to inhibit epithelial-to-mesenchymal transition. BACH1 also directly repressed the expression of genes important for epithelial cell adhesion including CLDN3 and CLDN4. In a mouse orthotopic implantation model, BACH1 was required for the high metastatic ability of AsPC-1 cells. IHC analysis of clinical specimens with a newly developed anti-BACH1 mAb revealed that high expression of BACH1 is a poor prognostic factor. These results suggest that the gene regulatory network of BACH1 and downstream genes including CDH1 contribute to the malignant features of PDAC by regulating epithelial-to-mesenchymal transition. SIGNIFICANCE: Greater understanding of the gene regulatory network involved in epithelial-to-mesenchymal transition of pancreatic cancer cells will provide novel therapeutic targets and diagnostic markers.
Subject(s)
Basic-Leucine Zipper Transcription Factors/metabolism , Carcinoma, Pancreatic Ductal/genetics , Epithelial-Mesenchymal Transition/genetics , Gene Expression Regulation, Neoplastic , Pancreatic Neoplasms/genetics , Animals , Antigens, CD/genetics , Basic-Leucine Zipper Transcription Factors/genetics , Cadherins/genetics , Carcinoma, Pancreatic Ductal/mortality , Carcinoma, Pancreatic Ductal/pathology , Cell Movement/genetics , Claudin-3/genetics , Claudin-4/genetics , Female , Gene Knockout Techniques , Gene Regulatory Networks , Hepatocyte Nuclear Factor 3-alpha/genetics , Hepatocyte Nuclear Factor 3-alpha/metabolism , Humans , Kaplan-Meier Estimate , Male , Mice , Middle Aged , Neoplasm Invasiveness/genetics , Neoplasm Invasiveness/pathology , Neoplasm Metastasis/genetics , Pancreatic Neoplasms/mortality , Pancreatic Neoplasms/pathology , Prognosis , RNA-SeqABSTRACT
Tick saliva contains immunosuppressants which are important to obtain a blood meal and enhance the infectivity of tick-borne pathogens. In Japan, Ixodes persulcatus is a major vector for Lyme borreliosis pathogens, such as Borrelia garinii, as well as for those causing relapsing fever, such as B. miyamotoi. To date, little information is available on bioactive salivary molecules, produced by this tick. Thus, in this study, we identified two proteins, I. persulcatus derived sialostatin L1 (Ip-sL1) and sL2 (Ip-sL2), as orthologs of I. scapularis derived sL1 and sL2. cDNA clones of Ip-sL1 and Ip-sL2 shared a high identity with sequences of sL1 and sL2 isolated from the salivary glands of I. scapularis. Semi-quantitative PCR revealed that Ip-sL1 and Ip-sL2 were expressed in the salivary glands throughout the life of the tick. In addition, Ip-sL1 and Ip-sL2 were expressed even before the ticks started feeding, and their expression continued during blood feeding. Recombinant Ip-sL1 and Ip-sL2 were developed to characterize the proteins via biological and immunological analyses. These analyses revealed that both Ip-sL1 and Ip-sL2â¯had inhibitory effects on cathepsins L and S. Ip-sL1 and Ip-sL2 inhibited the production of IP-10, TNFα, and IL-6 by LPS-stimulated bone-marrow-derived dendritic cells (BMDCs). Additionally, Ip-sL1 significantly impaired BMDC maturation. Taken together, these results suggest that Ip-sL1 and Ip-sL2 confer immunosuppressive functions and appear to be involved in the transmission of pathogens by suppressing host immune responses, such as cytokine production and dendritic cell maturation. Therefore, further studies are warranted to investigate the immunosuppressive functions of Ip-sL1 and Ip-sL2 in detail to clarify their involvement in pathogen transmission via I. persulcatus.
Subject(s)
Arthropod Proteins/immunology , Cystatins/immunology , Immunity, Innate/physiology , Ixodes/physiology , Amino Acid Sequence , Animals , Base Sequence , Female , Mice , Mice, Inbred BALB C , Phylogeny , Sequence AlignmentABSTRACT
Peripherally substituted tetradecaazaheptacene (N14Hp) compounds, exhibiting amphiprotism-coupled emission, have been synthesized. X-ray crystallography reveals a planar acene-like chromophore, and electronic absorption and emission occur in the near-infrared biological transparency window (650-900 nm). The compounds exhibit long-wavelength emission with photoluminescence quantum yields ΦPL up to â¼0.61 at 686 nm, with the monodeprotonated state ΦPL ≈ 0.58 at 712 nm. This unprecedented highly nitrogenous chromophore illustrates the stability and utility of the pyrazinacenes for different applications based on their photophysical properties and chemical structures.