ABSTRACT
Regular monitoring of Norovirus presence in environmental and food samples is crucial due to its high transmission rates and outbreak potential. For detecting Norovirus GI, reverse transcription qPCR method is commonly used, but its sensitivity can be affected by assay performance. This study shows significantly reduced assay performance in digital PCR or qPCR when using primers targeting Norovirus GI genome 5291-5319 (NC_001959), located on the hairpin of the predicted RNA structure. It is highly recommended to avoid this region in commercial kit development or diagnosis to minimizing potential risk of false negatives.
Subject(s)
Norovirus , Reverse Transcriptase Polymerase Chain Reaction , Norovirus/genetics , Norovirus/isolation & purification , Reverse Transcriptase Polymerase Chain Reaction/methods , RNA, Viral/genetics , RNA, Viral/analysis , Humans , Caliciviridae Infections/diagnosis , Caliciviridae Infections/virologyABSTRACT
Cancer research studies using next-generation sequencing have revealed a number of genes of which aberrant expression is associated with various cancers. Recently, long non-coding RNA (lncRNA) has been highlighted due to its tissue-specific expression and cell cancerization functions, such as the regulation of key tumor suppressors. In this study, we suggest a very efficient approach to survey lncRNAs putatively associated with breast cancer. We targeted lncRNAs linked with breast cancer associated genes (BCAGs) and analyzed their expression pattern in human breast cancer cell lines. A total of 337 BCAGs were retrieved from literature review and the existence of 121 lncRNAs were identified from the 15â¯kb up- and downstream regions of the list of genes. Twenty lncRNAs' expression were detectable in human breast cancer cell lines with different expression patterns. Interestingly, the expression of three lncRNAs, two up-regulated (RAD51C v.4, LOC105371849) and one down-regulated (LOC102724064), were closely correlated with adjacent BCAGs (RAD51C, HEATR6 and BRMS1) in breast cancer cell lines. We thus demonstrated association between the lncRNA and its adjacent BCAG using LOC105371849-HEATR6, of which the function and regulation in breast cancer are still unknown. Knockdown of LOC105371849 by siRNA decreased the expression of HEATR6 mRNA in the MCF7 human breast cancer cell line. In conclusion, this study provides a better understanding about the biological roles of lncRNAs in breast cancer and may be useful in the investigation of proper targets for diagnostic and/or therapeutic breast cancer markers using public databases.
Subject(s)
Breast Neoplasms/genetics , Gene Expression Profiling/methods , RNA, Long Noncoding/genetics , Breast/metabolism , Cell Line, Tumor , Early Detection of Cancer/methods , Female , Gene Expression Regulation, Neoplastic/genetics , High-Throughput Nucleotide Sequencing , Humans , MCF-7 Cells , RNA, Long Noncoding/metabolism , RNA, Messenger/genetics , Sequence Analysis, RNA/methods , Transcriptome/geneticsABSTRACT
Metastases originate from primary tumors and reach distant organs. Growing evidence suggests that metastases are under the control of primary tumors even outside the primary site; however, the mechanisms by which primary tumors remotely control metastases remain unclear. Here, we discovered a molecular mechanism by which primary tumors suppress metastatic growth. Interestingly, we found that extracellular vesicles (EVs) derived from the primary tumor can inhibit the growth of metastases both in vitro and in vivo. miR-1 was particularly enriched in primary tumor-derived EVs (pTDEs) and was found to be responsible for the suppression of metastatic growth. Mechanistically, intracellular reactive oxygen species (ROS) production and DNA damage were induced, which led to cell cycle arrest. Collectively, our data demonstrate that primary tumors restrict the growth of distant metastases via miR-1 in pTDEs and that miR-1 could potentially be used as an antimetastatic agent.
Subject(s)
Extracellular Vesicles , MicroRNAs , Neoplasms , Humans , MicroRNAs/genetics , MicroRNAs/metabolism , Extracellular Vesicles/metabolism , Neoplasms/metabolismABSTRACT
Periprosthetic tissue inflammation is a challenging complication arising in joint replacement surgeries, which is often caused by wear debris from polyethylene (PE) components. In this study, we examined the potential biological effects of grafting a [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium hydroxide (MEDSAH) polymer onto the surface of PE through a solvent-evaporation technique. J774A.1 macrophage-like cells and primary cultured mouse osteoblasts were treated with PE powder with or without the MEDSAH coating. MEDSAH grafting on PE substantially reduced the expression of pro-inflammatory cytokines and other mediators in primary cultured mouse osteoblasts, but did not significantly impact macrophage-mediated inflammation. Our findings suggest that a MEDSAH coating on PE-based materials has potential utility in mitigating periprosthetic tissue inflammation and osteolysis and preventing aseptic loosening in total joint replacements. Further research, including large-scale clinical trials and biomechanical analyses, is needed to assess the long-term performance and clinical implications of MEDSAH-coated PE-based materials in total joint arthroplasty.
Subject(s)
Inflammation , Osteoblasts , Polyethylene , Animals , Mice , Inflammation/pathology , Osteoblasts/metabolism , Osteoblasts/drug effects , Macrophages/metabolism , Cell Line , Cytokines/metabolism , Osteolysis/etiology , Osteolysis/pathology , Coated Materials, Biocompatible/chemistry , Methacrylates/chemistry , Arthroplasty, Replacement/adverse effectsABSTRACT
Most medical implants are made of titanium. When titanium is exposed to air for a long time, hydrocarbons are deposited and the surface becomes hydrophobic. Cell attachment is important for bone ingrowth to occur on the implant surface, and hydrophilicity can enhance this. We examined whether non-thermal atmospheric pressure plasma treatment could increase the hydrophilicity of the titanium surface. Samples coated with four widely used coating types [grit blasting, micro arc oxidation (MAO), titanium plasma spray (TPS), and direct metal fabrication (DMF)] were treated with plasma. Each of the four surface-treated samples was divided into groups with and without plasma treatment. We analysed wettability by surface analysis and evaluation of contact angles, cell proliferation, and adhesion using scanning electron microscopy (SEM), confocal laser scanning microscopy, absorbance tests, and alkaline phosphatase (ALP) activity assay; four different Ti6Al4V surface types were compared. After plasma treatment, the contact angle was reduced on all surfaces, and the carbon content was reduced on all surfaces based on X-ray photoelectron spectroscopy (XPS) surface analysis. Under confocal laser scanning, the cell layer was thicker on the plasma-treated samples, especially in groups TPS and DMF. Cell proliferation was 41.8%, 17.7%, 54.9%, and 83.8% greater for the plasma- than non-plasma-treated grit blasting, MAO, TPS, and DMF samples, respectively. Hydrophilicity increased significantly under plasma treatment, and biological responsivity was also improved.
Subject(s)
Alloys , Plasma Gases , Titanium/pharmacology , Plasma Gases/pharmacology , Wettability , Cell ProliferationABSTRACT
Epigenetic dysregulation is an important feature for cancer initiation and progression. Long non-coding RNAs (lncRNAs) are transcripts that stably present as RNA forms with no translated protein and have lengths larger than 200 nucleotides. LncRNA can epigenetically regulate either oncogenes or tumor suppressor genes. Nowadays, the combined research of lncRNA plus protein analysis is gaining more attention. LncRNA controls gene expression directly by binding to transcription factors of target genes and indirectly by complexing with other proteins to bind to target proteins and cause protein degradation, reduced protein stability, or interference with the binding of other proteins. Various studies have indicated that lncRNA contributes to cancer development by modulating genes epigenetically and studies have been done to determine which proteins are combined with lncRNA and contribute to cancer development. In this review, we look in depth at the epigenetic regulatory function of lncRNAs that are capable of complexing with other proteins in cancer development.
ABSTRACT
Stem cell-based therapy is one of the most attractive approaches to ischemic heart diseases, such as myocardial infarction (MI). We evaluated the cardio-protective effects of the human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) stably expressing lymphoid enhancer-binding factor 1 (LEF1; LEF1/hUCB-MSCs) in a rat model of MI. LEF1 overexpression in hUCB-MSCs promoted cell-proliferation and anti-apoptotic effects in hypoxic conditions. For the application of its therapeutic effects in vivo, the LEF1 gene was introduced into an adeno-associated virus integration site 1 (AAVS1) locus, known as a safe harbor site on chromosome 19 by CRISPR/Cas9-mediated gene integration in hUCB-MSCs. Transplantation of LEF1/hUCB-MSCs onto the infarction region in the rat model significantly improved overall survival. The cardio-protective effect of LEF1/hUCB-MSCs was proven by echocardiogram parameters, including greatly improved left-ventricle ejection fraction (EF) and fractional shortening (FS). Moreover, histology and immunohistochemistry successfully presented reduced MI region and fibrosis by LEF1/hUCB-MSCs. We found that these overall positive effects of LEF1/hUCB-MSCs are attributed by increased proliferation and survival of stem cells in oxidative stress conditions and by the secretion of various growth factors by LEF1. In conclusion, this study suggests that the stem cell-based therapy, conjugated with genome editing of transcription factor LEF1, which promotes cell survival, could be an effective therapeutic strategy for cardiovascular disease.
ABSTRACT
Breast cancer (BC) is one of the most common cancers in both women and female dogs. Methylation changes of LINE-1 have been reported in human cancers. The aim of this study was to determine the hypomethylation of canine LINE-1 in liquid biopsies for canine mammary tumors (CMT) and to assess its diagnostic performance in human plasma. BC associated LINE-1 methylation was measured by methylation sensitive (HpaII) and insensitive (MspI) restriction enzyme digestion followed by real-time PCR using the cfDNA isolated from 300 µl of plasma. The relative level of methylated canine LINE-1 was less than 0.4 in the benign and malignant CMTs (0.29 ± 0.061 and 0.39 ± 0.066, respectively) when it was 0.92 ± 0.067 in the healthy controls. The area under the ROC curve (AUC) was significantly high in both benign and malignant tumors (0.97 and 0.93). Furthermore, this approach was also successfully implemented in a set of 26 human BCs with 10 healthy controls (AUC = 0.78). Altogether, our data suggest that the comparative approach using a dog model might be helpful to rapidly develop a new diagnostic biomarker and that the methylation of LINE-1 in cfDNA may be a good target as a diagnostic marker of both human BC and CMT.
Subject(s)
Biomarkers, Tumor/blood , Breast Neoplasms/pathology , Cell-Free Nucleic Acids/blood , DNA, Neoplasm/blood , Deoxyribonuclease I , Mammary Neoplasms, Animal/pathology , Adult , Animals , Breast/pathology , Cell Line, Tumor , Deoxyribonuclease I/blood , Deoxyribonuclease I/genetics , Dogs , Female , Humans , Liquid Biopsy , Mammary Glands, Animal/pathology , Middle Aged , Young AdultABSTRACT
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
ABSTRACT
Breast cancer is one of the most frequently diagnosed cancers in both women and female dogs. Genome-wide association studies in human breast cancer (HBC) have identified hundreds of genetic variations and somatic driver mutations. However, only a handful of variants have been studied for rare HBC and their associations remain inconclusive. Spontaneous canine mammary tumor (CMT) is a great model for HBC, with clinical similarity. We thus performed whole-exome sequencing in 20 pairs of CMT and normal tissues in dogs. We newly found that PIK3CA was the most frequently mutated gene in CMT (45%). Furthermore, canine PIK3CA A3140G (H1047R), at what is known as the mutational hotspot of HBC, is also a hotspot in CMT. Targeted sequencing confirmed that 29% of CMTs had the same PIK3CA A3140G mutation. Integration of the transcriptome suggests that the PIK3CA (H1047R) induced cell metabolism and cell cycle via an increase of PCK2 and a decrease of CDKN1B but had no effect on cell apoptosis. We identified additional significantly mutated genes, including SCRN1 and CLHC1, which have not been reported in HBC. Our study recapitulated some known HBC-associated genes and human cancer signatures in CMT, and identified novel genes that may be relevant to HBC. This study may allow us to better understand both HBC and CMT and lend new insights into the development of biomarkers.
ABSTRACT
Breast cancer (BC)/mammary gland carcinoma (MGC) is the most frequently diagnosed and leading cause of cancer-related mortality in both women and canines. To better understand both canine MGC and human BC-specific genes, we sequenced RNAs obtained from eight pairs of carcinomas and adjacent normal tissues in dogs. By comprehensive transcriptome analysis, 351 differentially expressed genes (DEGs) were identified in overall canine MGCs. Based on the DEGs, comparative analysis revealed correlation existing among the three histological subtypes of canine MGC (ductal, simple, and complex) and four molecular subtypes of human BC (HER2+, ER+, ER&HER2+, and TNBC). Eight DEGs shared by all three subtypes of canine MGCs had been previously reported as cancer-associated genes in human studies. Gene ontology and pathway analyses using the identified DEGs revealed that the biological processes of cell proliferation, adhesion, and inflammatory responses are enriched in up-regulated MGC DEGs. In contrast, fatty acid homeostasis and transcription regulation involved in cell fate commitment were down-regulated in MGC DEGs. Moreover, correlations are demonstrated between upstream promoter transcripts and DEGs. Canine MGC- and subtype-enriched gene expression allows us to better understand both human BC and canine MGC, yielding new insight into the development of biomarkers and targets for both diseases.
ABSTRACT
Periacetabular osteotomy (PAO) is a complex surgical procedure to restore acetabular coverage in the dysplastic hip, and the amount of acetabular rotation during PAO plays a key role. Using computational simulations, this study assessed the optimal direction and amount of the acetabular rotation in three dimensions for a patient undergoing PAO. Anatomy-specific finite element (FE) models of the hip were constructed based on clinical CT images. The calculated acetabular rotation during PAO were 9.7°, 18°, and 4.3° in sagittal, coronal, and transverse planes, respectively. Based on the actual acetabular rotations, twelve postoperative FE models were generated. An optimal position was found by gradually varying the amount of the acetabular rotations in each anatomical plane. The coronal plane was found to be the principal rotational plane, which showed the strongest effects on joint contact pressure compared to other planes. It is suggested that rotation in the coronal plane of the osteotomized acetabulum is one of the primary surgical parameters to achieve the optimal clinical outcome for a given patient.