ABSTRACT
Vascular endothelial growth factor (VEGF) plays a significant role as a pro-angiogenic and pro-permeability factor within the kidney. Bevacizumab is a pharmaceutical monoclonal anti-VEGF antibody that inhibits the growth of new blood vessels, which blocks blood supply and thereby restricts tumor growth. Thus, we conducted a nationwide study to explore the risk of chronic kidney disease (CKD) development in Taiwan residents after bevacizumab therapy. We drew data from the extensive National Health Insurance Research Database (NHIRD), which encompasses data from >99% of Taiwan's population from 1995 onwards. Individuals who received bevacizumab between 2012-2018 were identified as the bevacizumab cohort, with the index date set at the first usage. We randomly selected dates within the study period for the control group to serve as index dates. We excluded patients with a history of CKD prior to the index date or those <20 years old. In both cohorts, patients' propensity scores matched in a 1:1 ratio based on sex, age, index year, income, urbanization level, comorbidities, and medications. We found patients treated with bevacizumab had a significantly higher risk of contracting CKD than patients without bevacizumab (adjusted hazard ratio = 1.35, 95% confidence interval = 1.35-1.73). The risk of CKD was 1.35-fold higher in participants with bevacizumab treatment than those in the control group. These findings suggest that close monitoring of CKD development after bevacizumab administration is needed.
Subject(s)
Renal Insufficiency, Chronic , Vascular Endothelial Growth Factor A , Humans , Young Adult , Adult , Bevacizumab/adverse effects , Retrospective Studies , Taiwan/epidemiology , Renal Insufficiency, Chronic/epidemiologyABSTRACT
BACKGROUND: The association between exposure to air pollution and sudden sensorineural hearing loss (SSNHL) has not been extensively discussed in the literature. Therefore, we conducted this nationwide study to evaluate the risk of SSNHL in Taiwanese residents with exposure to air pollution. METHODS: We enrolled subjects aged older than 20 years with no history of SSNHL from 1998 to 2010, and followed up until developing SSNHL, withdrawn from the National Health Insurance program, and the end of the database (2011/12/31). The air quality data are managed by Taiwan Environmental Protection Administration. The annual concentrations of PM2.5, SO2, CO, NO, and NO2 from 1998 to 2010 were classified into the three levels according to tertiles. We calculated the annual average of pollutants from baseline until the end of the study, and classified into tertiles. The adjusted hazard ratio (aHR) was estimated by using the multivariate Cox proportional hazard model. RESULTS: When considered continuous air pollutants concentration, subjects who exposed with higher concentration of CO (aHR = 2.16, 95% CI 1.50-3.11), NO (aHR = 1.02, 95% CI 1.01-1.03), and NO2 (aHR = 1.02, 95% CI 1.01-1.04) developing significant higher risk of SSNHL. When classified air pollutants concentration into low, moderate and high level by tertiles, and selected low level as reference, patients exposed with moderate (aHR = 1.56, 95% CI 1.20-2.04) or high level (aHR = 1.33, 95% CI 1.01-1.75) of PM2.5 showed significant higher risk of developing SSNHL. CONCLUSION: This study indicated an increased risk of SSNHL in residents with long-term exposure to air pollution. Nevertheless, further experimental, and clinical studies are needed to validate the study findings.
Subject(s)
Air Pollutants , Air Pollution , Hearing Loss, Sensorineural , Hearing Loss, Sudden , Aged , Air Pollutants/toxicity , Air Pollution/statistics & numerical data , Hearing Loss, Sensorineural/chemically induced , Hearing Loss, Sensorineural/epidemiology , Humans , Proportional Hazards Models , Risk FactorsABSTRACT
Umbilical cord-derived mesenchymal stromal cells (UCMSCs) have potential applications in regenerative medicine. UCMSCs have been demonstrated to repair tissue damage in many inflammatory and degenerative diseases. We have previously shown that UCMSC exosomes reduce nerve injury-induced pain in rats. In this study, we characterized UCMSC exosomes using RNA sequencing and proteomic analyses and investigated their protective effects on cisplatin-induced hearing loss in mice. Two independent experiments were designed to investigate the protective effects on cisplatin-induced hearing loss in mice: (i) chronic intraperitoneal cisplatin administration (4 mg/kg) once per day for 5 consecutive days and intraperitoneal UCMSC exosome (1.2 µg/µL) injection at the same time point; and (ii) UCMSC exosome (1.2 µg/µL) injection through a round window niche 3 days after chronic cisplatin administration. Our data suggest that UCMSC exosomes exert protective effects in vivo. The post-traumatic administration of UCMSC exosomes significantly improved hearing loss and rescued the loss of cochlear hair cells in mice receiving chronic cisplatin injection. Neuropathological gene panel analyses further revealed the UCMSC exosomes treatment led to beneficial changes in the expression levels of many genes in the cochlear tissues of cisplatin-injected mice. In conclusion, UCMSC exosomes exerted protective effects in treating ototoxicity-induced hearing loss by promoting tissue remodeling and repair.
Subject(s)
Cochlear Diseases/etiology , Cochlear Diseases/therapy , Exosomes/metabolism , Hair Cells, Auditory, Outer/drug effects , Hair Cells, Auditory, Outer/metabolism , Mesenchymal Stem Cells/metabolism , Umbilical Cord/cytology , Animals , Antineoplastic Agents/adverse effects , Biological Therapy , Biomarkers , Cisplatin/adverse effects , Cochlear Diseases/pathology , Disease Models, Animal , Exosomes/transplantation , Hair Cells, Auditory, Outer/pathology , Hearing Loss/etiology , Hearing Loss/metabolism , Hearing Loss/therapy , Immunophenotyping , Mice , MicroRNAs/genetics , Proteomics/methods , Treatment OutcomeABSTRACT
The Ca2+-mediated S100 family protein S100A6 has a crucial task in various intracellular and extracellular activities thereby demonstrating a possible involvement in the advancement and development of malignant tumors. S100A6 has been found to associate with receptor for advanced glycation end products, RAGE, through its extracellular extension. This extension is famously identified as a prominent receptor for many S100 family associates. Additionally, S100A6 binds to S100B protein and forms a heterodimer. Thus, we consider the S100B protein to be a prospective drug molecule to obstruct the interacting regions amongst S100A6 and RAGE V domain. We applied the NMR spectroscopy method to locate the binding area amid the S100A6m (mutant S100A6, cysteine at 3rd position of S100A6 is replaced with serine, C3S) and S100B proteins. The 1H-15N HSQC NMR titrations revealed the probable requisite dynamics of S100A6m and S100B interfaces. Utilizing data from the NMR titrations as input parameters, we ran the HADDOCK program and created a S100A6m-S100B heterodimer complex. The obtained complex was then superimposed with the reported complex of S100A6m-RAGE V domain. This superimposition displayed the possibility of S100B to be a potential antagonist that can block the interface area of the S100A6m and the RAGE V domain. Moreover, an in vitro cancer model using SW480 cells in water-soluble tetrazolium-1 assay (WST-1) showed a noticeable change in the cell proliferation as an effect of these proteins. Our study indicates the possibility to develop a S100B-like competitor that could play a key role in the treatment of S100- and RAGE-mediated human diseases.
Subject(s)
Cell Cycle Proteins/chemistry , Gene Expression Regulation, Neoplastic , Receptor for Advanced Glycation End Products/chemistry , S100 Calcium Binding Protein A6/chemistry , S100 Calcium Binding Protein beta Subunit/chemistry , Binding Sites , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Cell Cycle Proteins/pharmacology , Cell Line, Tumor , Cell Proliferation/drug effects , Cloning, Molecular , Epithelial Cells/drug effects , Epithelial Cells/metabolism , Epithelial Cells/pathology , Escherichia coli/genetics , Escherichia coli/metabolism , Genetic Vectors/chemistry , Genetic Vectors/metabolism , Humans , Molecular Docking Simulation , Nuclear Magnetic Resonance, Biomolecular , Protein Binding , Protein Conformation, alpha-Helical , Protein Conformation, beta-Strand , Protein Interaction Domains and Motifs , Protein Multimerization , Receptor for Advanced Glycation End Products/genetics , Receptor for Advanced Glycation End Products/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Recombinant Proteins/pharmacology , S100 Calcium Binding Protein A6/genetics , S100 Calcium Binding Protein A6/metabolism , S100 Calcium Binding Protein A6/pharmacology , S100 Calcium Binding Protein beta Subunit/genetics , S100 Calcium Binding Protein beta Subunit/metabolism , S100 Calcium Binding Protein beta Subunit/pharmacologyABSTRACT
Metastasis-associated S100A4 protein is a small calcium-binding protein typically overexpressed in several tumor forms, and it is widely accepted that S100A4 plays a significant role in the metastasis of cancer. Tumor suppressor p53 is one of the S100A4's main targets. Previous reports show that through p53, S100A4 regulates collagen expression and cell proliferation. When S100A4 interacts with p53, the S100A4 destabilizes wild type p53. In the current study, based on 1H-15N HSQC NMR experiments and HADDOCK results, S100A4 interacts with the intrinsically unstructured transactivation domain (TAD) of the protein p53 and the pentamidine molecules in the presence of calcium ions. Our results suggest that the p53 TAD and pentamidine molecules share similar binding sites on the S100A4 protein. This observation indicates that a competitive binding mechanism can interfere with the binding of S100A4-p53 and increase the level of p53. Also, we compare different aspects of p53 activity in the WST-1 test using MCF 7 cells. We found that the presence of a pentamidine molecule results in higher p53 activity, which is also reflected in less cell proliferation. Collectively, our results indicate that disrupting the S100A4-p53 interaction would prevent cancer progression, and thus S100A4-p53 inhibitors provide a new avenue for cancer therapy.
Subject(s)
Cell Proliferation/drug effects , Pentamidine/pharmacology , Protein Multimerization/drug effects , S100 Calcium-Binding Protein A4/metabolism , Tumor Suppressor Protein p53/metabolism , Antineoplastic Agents/metabolism , Antineoplastic Agents/pharmacology , Binding Sites , Drug Screening Assays, Antitumor , Humans , MCF-7 Cells , Pentamidine/metabolism , Protein Binding , S100 Calcium-Binding Protein A4/chemistry , Tumor Suppressor Protein p53/chemistryABSTRACT
BACKGROUND: Bipolar disorder (BD), a type of psychiatric mood disorder, is manifested by chronic and recurrent mood fluctuations. This study aims to determine whether hepatitis B virus (HBV) or hepatitis C virus (HCV) infection is a risk factor for BD. METHODS: A total of 48,215 patients with newly diagnosed viral hepatitis from 2000 to 2010 were identified and frequency-matched with 192,860 people without hepatitis. Both groups were followed until diagnosis with BD, withdrawal from the national health insurance program, or the end of 2011. Patients with viral hepatitis were grouped into 3 cohorts: HBV infection, HCV infection, and HBV/HCV coinfection. The association between viral hepatitis and BD were examined using Cox proportional hazards regression models. RESULTS: The incidence of BD was higher in HBV/HCV coinfection than in the control group, with an adjusted hazard ratio of 2.16 (95% confidence interval 1.06-4.41) when adjusted for sex, age, and comorbidity. After further adjustment, we noted that an age more than 65 years and female may be associated with an increased risk of BD in patients with chronic hepatitis B and C. CONCLUSION: Viral hepatitis may be associated with increased risk of subsequent BD.
Subject(s)
Bipolar Disorder/complications , Hepatitis B/virology , Hepatitis C/complications , Hepatitis C/virology , Adult , Aged , Bipolar Disorder/epidemiology , Comorbidity , Female , Hepatitis B/complications , Humans , Incidence , Male , Middle Aged , Proportional Hazards ModelsABSTRACT
BACKGROUND: Hepatitis B virus (HBV) infection is strongly associated with hepatocellular carcinoma due to the main pathogenic X protein of HBV (HBx). Whether HBV infection and the HBx protein could result in macular degeneration (MD) is not known. The aim of this study is to assess the association and underlying mechanisms between HBV infection and MD. METHODS: The National Health Research Institutes in Taiwan built a large database, the National Health Insurance Research Database (NHIRD), which includes the claims data from the Taiwan National Health Insurance (NHI) program. The Taiwan NHI is a single-payer, compulsory health insurance program for Taiwan citizens. The data for the present study were derived from the Longitudinal Health Insurance Database, which contains the claims data of 1 million insured people within the NHIRD, including beneficiary registration, inpatient and outpatient files, drug use, and other medical services. In this study, we first investigated the association of HBV infection and the risk of MD by a population-based cohorts study enrolling 39,796 HBV-infected patients and 159,184 non-HBV-infected patients. RESULTS: After adjustment of age, sex, and comorbidities, the risk of MD was significantly higher in the HBV-infected cohort than in the non-HBV-infected cohort (adjusted HR = 1.31; 95% CI = 1.17-1.46). In vitro, we provided evidence to demonstrate that overexpression of HBx in the human retinal pigment epithelial (RPE) cell line, ARPE19, significantly reduced cell viability and clonogenic survival upon UV and blue light irradiation. By gene microarray analysis, we further showed that almost all genes in DNA repair pathways including base excision repair, nucleotide excision repair, mismatch repair, and homologous recombination were significantly down-regulated in the UV-induced cell death of HBx-transfected ARPE19 cells. CONCLUSIONS: The HBx protein may sensitize RPE cells to UV and blue light irradiation and increase the risk of HBV-infection-associated MD through down-regulation of multiple DNA repair pathways.
Subject(s)
Epithelial Cells/radiation effects , Hepatitis B virus/physiology , Hepatitis B/virology , Macular Degeneration/pathology , Macular Degeneration/virology , Retinal Pigment Epithelium/pathology , Trans-Activators/metabolism , Ultraviolet Rays , Adult , Cell Death/radiation effects , Cell Line , Cell Proliferation/radiation effects , Cell Shape/radiation effects , Cohort Studies , Epithelial Cells/pathology , Female , Gene Ontology , Gene Regulatory Networks , Hepatitis B/genetics , Humans , Macular Degeneration/genetics , Male , Middle Aged , Molecular Sequence Annotation , Proportional Hazards Models , Risk Factors , Transcription, Genetic/radiation effects , Viral Regulatory and Accessory ProteinsABSTRACT
PURPOSE: Laparoscopic distal pancreatectomy has proven to be feasible and safe. Moreover, robotic surgery provides unique advantages for pancreatic procedures, although single-incision robotic pancreatic surgery is rarely discussed. We applied the single-port modified platform to accomplish robotic distal pancreatectomy in a series of patients. METHODS: The subjects of this study were ten patients who underwent robotic distal pancreatectomy in our hospital between July 1, 2015 and Dec 31, 2016. All patients were placed supine in the reverse Trendelenburg position with the legs abducted. Surgery was performed via a trans-umbilical 5.0-cm incision, using a modified single-port platform (LAGIPORT®) combined with the da Vinci Si Surgical System. The three arms and scope (30-degree up) were inserted through the LAGIPORT® and positioned in a triangle. Endoscopic ultrasound was used to localize the tumor and plan the resection margin. We recorded the surgical time, operation time, blood loss, postoperative pain score, hospital stay, and complications. RESULTS: The surgical time was 236 ± 32 min, the operation time was 172 ± 30 min, and the blood loss was 149 ± 65 ml. All patients underwent robot-assisted distal pancreatectomy without conversion. The average pain score on postoperative day (POD) 3 was 4.5 ± 1. Complications included subsplenic hematoma (n = 1) and minor pancreatic leakage (n = 2). There was no surgical mortality. CONCLUSIONS: Our results demonstrate the safety and efficiency of robotic single-incision distal pancreatectomy via the modified platform (LAGIPORT®).
Subject(s)
Pancreatectomy/instrumentation , Pancreatic Neoplasms/surgery , Robotic Surgical Procedures/instrumentation , Adult , Aged , Anastomotic Leak/epidemiology , Blood Loss, Surgical/statistics & numerical data , Female , Hematoma/epidemiology , Humans , Length of Stay , Male , Middle Aged , Operative Time , Pain, Postoperative/epidemiology , Pancreatectomy/methods , Postoperative Complications/epidemiology , Robotic Surgical Procedures/methods , Safety , Splenic Diseases/epidemiology , Supine Position , Treatment OutcomeABSTRACT
The human S100 calcium-binding protein A11 (S100A11) is a member of the S100 protein family. Once S100A11 proteins bind to calcium ions at EF-hand motifs, S100A11 changes its conformation, promoting interaction with target proteins. The receptor for advanced glycation end products (RAGE) consists of three extracellular domains, including the V domain, C1 domain, and C2 domain. In this case, the V domain is the target for mutant S100A11 (mS100A11) binding. RAGE binds to the ligands, resulting in cell proliferation, cell growth, and several signal transduction cascades. We used NMR and fluorescence spectroscopy to demonstrate the interactions between mS100A11and RAGE V domain. The tranilast molecule is a drug used for treating allergic disorders. We discovered that the RAGE V domain and tranilast would interact with mS100A11 by using (1)H-(15)N HSQC NMR titrations. According to the results, we obtained two binary complex models from the HADDOCK program, S100A11-RAGE V domain and S100A11-tranilast, respectively. We overlapped two binary complex models with the same orientation of S100A11 homodimer and demonstrated that tranilast would block the binding site between S100A11 and the RAGE V domain. We further utilized a water-soluble tetrazolium-1 assay to confirm this result. We think that the results will be potentially useful in the development of new anti-cancer drugs.
Subject(s)
Cell Proliferation/drug effects , Receptor for Advanced Glycation End Products/metabolism , S100 Proteins/metabolism , ortho-Aminobenzoates/pharmacology , Fluorescence , Humans , Models, Molecular , Nuclear Magnetic Resonance, Biomolecular , Protein Binding , S100 Proteins/chemistry , Tryptophan/metabolismABSTRACT
Human S100A9 (Calgranulin B) is a Ca(2+)-binding protein, from the S100 family, that often presents as a homodimer in myeloid cells. It becomes an important mediator during inflammation once calcium binds to its EF-hand motifs. Human RAGE protein (receptor for advanced glycation end products) is one of the target-proteins. RAGE binds to a hydrophobic surface on S100A9. Interactions between these proteins trigger signal transduction cascades, promoting cell growth, proliferation, and tumorigenesis. Here, we present the solution structure of mutant S100A9 (C3S) homodimer, determined by multi-dimensional NMR experiments. We further characterize the solution interactions between mS100A9 and the RAGE V domain via NMR spectroscopy. CHAPS is a zwitterionic and non-denaturing molecule widely used for protein solubilizing and stabilization. We found out that CHAPS and RAGE V domain would interact with mS100A9 by using (1)H-(15)N HSQC NMR titrations. Therefore, using the HADDOCK program, we superimpose two binary complex models mS100A9-RAGE V domain and mS100A9-CHAPS and demonstrate that CHAPS molecules could play a crucial role in blocking the interaction between mS100A9 and the RAGE V domain. WST-1 assay results also support the conclusion that CHAPS inhibits the bioactivity of mS100A9. This report will help to inform new drug development against cell proliferation.
Subject(s)
Antineoplastic Agents/pharmacology , Calgranulin B/chemistry , Cell Proliferation/drug effects , Cholic Acids/pharmacology , Epithelial Cells/drug effects , Receptor for Advanced Glycation End Products/chemistry , Amino Acid Sequence , Antineoplastic Agents/chemistry , Binding Sites , Calgranulin B/genetics , Calgranulin B/metabolism , Cell Line, Tumor , Cholic Acids/chemistry , Cloning, Molecular , Epithelial Cells/metabolism , Epithelial Cells/pathology , Escherichia coli/genetics , Escherichia coli/metabolism , Gene Expression , Humans , Hydrophobic and Hydrophilic Interactions , Molecular Docking Simulation , Molecular Dynamics Simulation , Mutation , Protein Binding , Protein Domains , Protein Multimerization , Protein Structure, Secondary , Receptor for Advanced Glycation End Products/antagonists & inhibitors , Receptor for Advanced Glycation End Products/genetics , Receptor for Advanced Glycation End Products/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Structure-Activity RelationshipABSTRACT
Enhancer of zeste homolog 2 (EZH2), a histone methyltransferase, catalyzes tri-methylation of histone H3 at Lys 27 (H3K27me3) to regulate gene expression through epigenetic machinery. EZH2 functions as a double-facet molecule in regulation of gene expression via repression or activation mechanisms, depending on the different cellular contexts. EZH2 interacts with both histone and non-histone proteins to modulate diverse physiological functions including cancer progression and malignancy. In this review article, we focused on the updated information regarding microRNAs (miRNAs) and long non coding RNAs (lncRNAs) in regulation of EZH2, the oncogenic and tumor suppressive roles of EZH2 in cancer progression and malignancy, as well as current pre-clinical and clinical trials of EZH2 inhibitors.
Subject(s)
Enhancer of Zeste Homolog 2 Protein/genetics , Gene Expression Regulation, Neoplastic , MicroRNAs/genetics , Neoplasms/genetics , RNA, Long Noncoding/genetics , Disease Progression , Enhancer of Zeste Homolog 2 Protein/antagonists & inhibitors , Enzyme Inhibitors/therapeutic use , Humans , Mutation , Neoplasms/drug therapy , Neoplasms/pathology , Tumor Suppressor Proteins/geneticsABSTRACT
The human S100 protein family contains small, dimeric and acidic proteins that contain two EF-hand motifs and bind calcium. When S100A5 binds calcium, its conformation changes and promotes interaction with the target protein. The extracellular domain of RAGE (Receptor of Advanced Glycation End products) contain three domains: C1, C2 and V. The RAGE V domain is the target protein of S100A5 that promotes cell survival, growth and differentiation by activating several signaling pathways. Pentamidine is an apoptotic and antiparasitic drug that is used to treat or prevent pneumonia. Here, we found that pentamidine interacts with S100A5 using HSQC titration. We elucidated the interactions of S100A5 with RAGE V domain and pentamidine using fluorescence and NMR spectroscopy. We generated two binary models-the S100A5-RAGE V domain and S100A5-Pentamidine complex-and then observed that the pentamidine and RAGE V domain share a similar binding region in mS100A5. We also used the WST-1 assay to investigate the bioactivity of S100A5, RAGE V domain and pentamidine. These results indicated that pentamidine blocks the binding between S100A5 and RAGE V domain. This finding is useful for the development of new anti-proliferation drugs.
Subject(s)
Antigens, Neoplasm/chemistry , Antigens, Neoplasm/metabolism , Colorectal Neoplasms/metabolism , Mitogen-Activated Protein Kinases/chemistry , Mitogen-Activated Protein Kinases/metabolism , Pentamidine/administration & dosage , Pentamidine/chemistry , S100 Proteins/chemistry , S100 Proteins/metabolism , Binding Sites , Cell Line, Tumor , Dose-Response Relationship, Drug , Humans , Models, Chemical , Molecular Docking Simulation , Mutation , Protein Binding , Protein Domains , S100 Proteins/ultrastructure , Signal Transduction/drug effectsABSTRACT
The extracellular portion of the human fibroblast growth factor receptor2 D2 domain (FGFR2 D2) interacts with human fibroblast growth factor 1 (hFGF1) to activate a downstream signaling cascade that ultimately affects mitosis and differentiation. Suramin is an antiparasiticdrug and a potent inhibitor of FGF-induced angiogenesis. Suramin has been shown to bind to hFGF1, and might block the interaction between hFGF1 and FGFR2 D2. Here, we titrated hFGF1 with FGFR2 D2 and suramin to elucidate their interactions using the detection of NMR. The docking results of both hFGF1-FGFR2 D2 domain and hFGF1-suramin complex were superimposed. The results indicate that suramin blocks the interaction between hFGF1 and FGFR2 D2. We used the PyMOL software to show the hydrophobic interaction of hFGF1-suramin. In addition, we used a Water-soluble Tetrazolium salts assay (WST1) to assess hFGF1 bioactivity. The results will be useful for the development of new antimitogenic activity drugs.
Subject(s)
Fibroblast Growth Factor 1/chemistry , Fibroblast Growth Factor 1/metabolism , Receptor, Fibroblast Growth Factor, Type 2/chemistry , Receptor, Fibroblast Growth Factor, Type 2/metabolism , Suramin/chemistry , Suramin/pharmacology , Antiparasitic Agents , Binding Sites/drug effects , Down-Regulation , Fibroblast Growth Factor 1/ultrastructure , Humans , MCF-7 Cells , Molecular Docking Simulation , Protein Binding/drug effects , Protein Domains/drug effects , Signal Transduction/drug effects , Signal Transduction/physiologyABSTRACT
S100B is a calcium sensing protein belonging to the S100 protein family with intracellular and extracellular roles. It is one of the EF hand homodimeric proteins, which is known to interact with various protein targets to regulate varied biological functions. Extracellular S100B has been recently reported to interact with FGF2 in a RAGE-independent manner. However, the recognition mechanism of S100B-FGF2 interaction at the molecular level remains unclear. In this study, the critical residues on S100B-FGF2 interface were mapped by combined information derived from NMR spectroscopy and site directed mutagenesis experiments. Utilizing NMR titration data, we generated the structural models of S100B-FGF2 complex from the computational docking program, HADDOCK which were further proved stable during 15ns unrestrained molecular dynamics (MD) simulations. Isothermal titration calorimetry studies indicated S100B interaction with FGF2 is an entropically favored process implying dominant role of hydrophobic contacts at the protein-protein interface. Residue level information of S100B interaction with FGF2 was useful to understand the varied target recognition ability of S100B and further explained its role in effecting extracellular signaling diversity. Mechanistic insights into the S100B-FGF2 complex interface and cell-based assay studies involving mutants led us to conclude the novel role of S100B in FGF2 mediated FGFR1 receptor inactivation.
Subject(s)
Fibroblast Growth Factor 2/metabolism , Molecular Docking Simulation , Molecular Dynamics Simulation , Multiprotein Complexes/metabolism , Receptor, Fibroblast Growth Factor, Type 1/metabolism , S100 Calcium Binding Protein beta Subunit/metabolism , Signal Transduction/physiology , Cell Line, Tumor , Fibroblast Growth Factor 2/chemistry , Fibroblast Growth Factor 2/genetics , Humans , Magnetic Resonance Spectroscopy , Multiprotein Complexes/chemistry , Multiprotein Complexes/genetics , Receptor, Fibroblast Growth Factor, Type 1/chemistry , Receptor, Fibroblast Growth Factor, Type 1/genetics , S100 Calcium Binding Protein beta Subunit/chemistry , S100 Calcium Binding Protein beta Subunit/geneticsABSTRACT
Globo H, a cancer-associated carbohydrate antigen, is highly expressed in various types of cancers. However, the role of Globo H in hepatocellular carcinoma (HCC) remains elusive. In our study, we performed glycan microarray analysis of 134 human serum samples to explore anti-Globo H antibody changes and found that Globo H is upregulated in hepatitis B virus (HBV)-positive HCC. Similarly, immunohistochemistry showed that Globo H expression was higher in tumors compared to normal tissues. In addition, fucosyltransferase 2 (FUT2), the main synthetic enzyme of Globo H, was also increased in HCC cells overexpressing HBV X protein (HBX). HBX plays an important role in promoting cell proliferation and may be related to increased levels of FUT2 and Globo H. Furthermore, using microRNA profiling, we observed that microRNA-15b (miR-15b) was downregulated in patients with HCC and confirmed association of FUT2 expression with expression of its product, Globo H. Therefore, our results suggest that HBX suppressed the expression of miR-15b, which directly targeted FUT2 and then increased levels of Globo H to enhance HCC cell proliferation. Additionally, proliferation of HBX-overexpressing HCC cells was significantly inhibited by treatment with Globo H antibody in vitro. In xenograft animal experiments, we found that overexpression of miR-15b effectively suppressed tumor growth. The newly identified HBX/miR-15b/FUT2/Globo H axis suggests one possible molecular mechanism of HCC cell proliferation and represents a new potential therapeutic target for HCC treatment.
Subject(s)
Antigens, Tumor-Associated, Carbohydrate/biosynthesis , Carcinoma, Hepatocellular/virology , Fucosyltransferases/metabolism , Liver Neoplasms/virology , MicroRNAs/metabolism , Trans-Activators/metabolism , Antigens, Tumor-Associated, Carbohydrate/genetics , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/pathology , Cell Growth Processes/physiology , Cell Line, Tumor , Down-Regulation , Fucosyltransferases/genetics , Hep G2 Cells , Hepatitis B/genetics , Hepatitis B/pathology , Hepatitis B/virology , Hepatitis B virus/genetics , Hepatitis B virus/metabolism , Humans , Liver Neoplasms/genetics , Liver Neoplasms/pathology , MicroRNAs/genetics , Trans-Activators/genetics , Up-Regulation , Viral Regulatory and Accessory Proteins , Galactoside 2-alpha-L-fucosyltransferaseABSTRACT
The S100P protein has been known to mediate cell proliferation by binding the receptor for advanced glycation end products (RAGE) to activate signaling pathways, such as the extracellular regulated kinase (ERK) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathways. S100P/RAGE signaling is involved in a variety of diseases, such as cancer, metastasis, and diabetes. Cromolyn is an anti-allergy drug that binds S100P to block the interaction between S100P and RAGE. In the present study, we characterized the properties of the binding between cromolyn and calcium-bound S100P using various biophysical techniques. The binding affinity for S100P and cromolyn was measured to be in the millimolar range by fluorescence spectroscopy. NMR-HSQC titration experiments and HADDOCK modeling was employed to determine the spatial structure of the proposed heterotetramer model of the S100P-cromolyn complex. Additional MD simulation results revealed the important properties in the complex stability and conformational flexibility of the S100P-cromolyn complex. This proposed model has provided an understanding of the molecular level interactions of S100P-cromolyn complex.
Subject(s)
Anti-Allergic Agents/pharmacology , Calcium-Binding Proteins/metabolism , Cromolyn Sodium/pharmacology , Neoplasm Proteins/metabolism , Calcium/metabolism , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , Protein Binding , Spectrometry, FluorescenceABSTRACT
BACKGROUND: A dual-function phantom designed to quantify the apparent diffusion coefficient (ADC) in different fat contents (FCs) and glass bead densities (GBDs) to simulate the human tissues has not been documented yet. We propose a dual-function phantom to quantify the FC and to measure the ADC at different FCs and different GBDs. METHODS: A fat-containing diffusion phantom comprised by 30 glass-bead-containing fat-water emulsions consisting of six different FCs (0, 10, 20, 30, 40, and 50%) multiplied by five different GBDs (0, 0.1, 0.25, 0.5, and 1.0 g/50 mL). The FC and ADC were measured by the "iterative decomposition of water and fat with echo asymmetry and least squares estimation-IQ," IDEAL-IQ, and single-shot echo-planar diffusion-weighted imaging, SS-EP-DWI, sequences, respectively. Linear regression analysis was used to evaluate the relationship among the fat fraction (FF) measured by IDEAL-IQ, GBD, and ADC. RESULTS: The ADC was significantly, negatively, and linearly associated with the FF (the linear slope ranged from -0.005 to -0.017, R2 = 0.925 to 0.986, all p < 0.001). The slope of the linear relationship between the ADC and the FF, however, varied among different GBDs (the higher the GBD, the lower the slope). ADCs among emulsions across different GBDs and FFs were overlapped. Emulsions with low GBDs plus high FFs shared a same lower ADC range with those with median or high GBDs plus median or lower FFs. CONCLUSIONS: A novel dual-function phantom simulating the human tissues allowed to quantify the influence of FC and GBD on ADC. RELEVANCE STATEMENT: The study developed an innovative dual-function MRI phantom to explore the impact of FC on ADC variation that can affect clinical results. The results revealed the superimposed effect on FF and GBD density on ADC measurements. KEY POINTS: ⢠A dual-function phantom made of glass bead density (GBD) and fat fraction (FF) emulsion has been developed. ⢠Apparent diffusion coefficient (ADC) values are determined by GBD and FF. ⢠The dual-function phantom showed the mutual ADC addition between FF and GBD.
Subject(s)
Diffusion Magnetic Resonance Imaging , Magnetic Resonance Imaging , Humans , Diffusion Magnetic Resonance Imaging/methods , Echo-Planar Imaging , Water , Phantoms, ImagingABSTRACT
Nuclear existence of epidermal growth factor receptor (EGFR) has been documented for more than two decades. Resistance of cancer to radiotherapy is frequently correlated with elevated EGFR expression, activity, and nuclear translocation. However, the role of nuclear EGFR (nEGFR) in radioresistance of cancers remains elusive. In the current study, we identified a novel nEGFR-associated protein, polynucleotide phosphorylase (PNPase), which possesses 3' to 5' exoribonuclease activity toward c-MYC mRNA. Knockdown of PNPase increased radioresistance. Inactivation or knock-down of EGFR enhanced PNPase-mediated c-MYC mRNA degradation in breast cancer cells, and also increased its radiosensitivity. Interestingly, the association of nEGFR with PNPase and DNA-dependent protein kinase (DNAPK) increased significantly in breast cancer cells after exposure to ionizing radiation (IR). We also demonstrated that DNAPK phosphorylates PNPase at Ser-776, which is critical for its ribonuclease activity. The phospho-mimetic S776D mutant of PNPase impaired its ribonuclease activity whereas the nonphosphorylatable S776A mutant effectively degraded c-MYC mRNA. Here, we uncovered a novel role of nEGFR in radioresistance, and that is, upon ionizing radiation, nEGFR inactivates the ribonuclease activity of PNPase toward c-MYC mRNA through DNAPK-mediated Ser-776 phosphorylation, leading to increase of c-MYC mRNA, which contributes to radioresistance of cancer cells.
Subject(s)
DNA-Activated Protein Kinase/metabolism , ErbB Receptors/metabolism , Exoribonucleases/metabolism , Gamma Rays , Nuclear Proteins/metabolism , Proto-Oncogene Proteins c-myc/metabolism , RNA Stability/radiation effects , RNA, Messenger/metabolism , Amino Acid Substitution , Cell Line, Tumor , DNA-Activated Protein Kinase/genetics , ErbB Receptors/genetics , Exoribonucleases/genetics , Humans , Mutation, Missense , Nuclear Proteins/genetics , Phosphorylation/genetics , Phosphorylation/radiation effects , Proto-Oncogene Proteins c-myc/genetics , RNA Stability/genetics , RNA, Messenger/genetics , Radiation Tolerance/genetics , Radiation Tolerance/radiation effectsABSTRACT
BACKGROUND AND OBJECTIVES: The mechanisms of photodynamic therapy (PDT) have been studied on the cellular and tissue levels. However, the cellular behaviors of cancer cells survived from PDT are still not clear. Previously, we have found that PDT-derived variants A375/3A5 and A375/6A5 have reduced invasion ability. This study attempted to further elucidate the possible molecules associated with the altered invasiveness in the PDT-derived variants and cancer cells treated with PDT. STUDY DESIGN/MATERIALS AND METHODS: Scratch wound healing assay and invasion assay were performed to evaluate the migration and invasion ability of human A375 melanoma and MDA-MB-231 breast adenocarcinoma cells. Single colony selection and microarray analysis were performed to examine the differentially expressed transcripts in parental A375 and PDT-derived variants. RT-PCR and Western blots analysis were performed to examine the expression levels of matrix metalloproteinase 9 (MMP9) and chloride intracellular channel 4 (CLIC4). The MMP9 activity was examined by Zymography assay. CLIC4 expressing construct was used to examine the influence on MMP9 expression and invasion ability of cancer cells treated with PDT. RESULTS: Correlated with the reduced invasiveness, we found that A375/3A5 and A375/6A5 cells have decreased production of MMP9. Microarray analysis and RT-PCR showed CLIC4 was down-regulated in the PDT-derived variants. Furthermore, down-regulation of CLIC4 and MMP9 was found in cancer cells treated with PDT. Transfection of surviving cancer cells with a plasmid vector encoding CLIC4 increased MMP9 expression and cell invasion. Furthermore, overexpression of CLIC4 in A375 and MDA-MB-231 cancer cells constrains PDT-induced suppression of invasiveness. CONCLUSION: Our results showed that the reduced expression of CLIC4 could further down-regulate MMP9 and result in the suppression of invasion in cancer cells treated with PDT. These results provide an insight into a new mechanism by which PDT affects the metastatic potential of cancer cells through down-regulation of MMP9 by CLIC4.
Subject(s)
Adenocarcinoma/pathology , Breast Neoplasms/pathology , Chloride Channels/physiology , Lung Neoplasms/pathology , Melanoma/pathology , Photochemotherapy , Adenocarcinoma/metabolism , Adenocarcinoma/therapy , Breast Neoplasms/metabolism , Breast Neoplasms/therapy , Cell Culture Techniques , Cell Line, Tumor , Cell Movement , Down-Regulation/radiation effects , Humans , Lung Neoplasms/metabolism , Lung Neoplasms/therapy , Matrix Metalloproteinase 9/metabolism , Melanoma/metabolism , Melanoma/therapy , Neoplasm Invasiveness/pathologyABSTRACT
Enhancer of zeste homolog 2 (EZH2) regulates stem cells renewal, maintenance, and differentiation into different cell lineages including neuron. Changes in intracellular Ca(2+) concentration play a critical role in the differentiation of neurons. However, whether EZH2 modulates intracellular Ca(2+) signaling in regulating neuronal differentiation from human mesenchymal stem cells (hMSCs) still remains unclear. When hMSCs were treated with a Ca(2+) chelator or a PLC inhibitor to block IP(3)-mediated Ca(2+) signaling, neuronal differentiation was disrupted. EZH2 bound to the promoter region of PIP5K1C to suppress its transcription in proliferating hMSCs. Interestingly, knockdown of EZH2 enhanced the expression of PIP5K1C, which in turn increased the amount of PI(4,5)P(2), a precursor of IP(3), and resulted in increasing the intracellular Ca(2+) level, suggesting that EZH2 negatively regulates intracellular Ca(2+) through suppression of PIP5K1C. Knockdown of EZH2 also enhanced hMSCs differentiation into functional neuron both in vitro and in vivo. In contrast, knockdown of PIP5K1C significantly reduced PI(4,5)P(2) contents and intracellular Ca(2+) release in EZH2-silenced cells and resulted in the disruption of neuronal differentiation from hMSCs. Here, we provide the first evidence to demonstrate that after induction to neuronal differentiation, decreased EZH2 activates the expression of PIP5K1C to evoke intracellular Ca(2+) signaling, which leads hMSCs to differentiate into functional neuron lineage. Activation of intracellular Ca(2+) signaling by repressing or knocking down EZH2 might be a potential strategy to promote neuronal differentiation from hMSCs for application to neurological dysfunction diseases.