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1.
Int J Mol Sci ; 22(5)2021 Mar 09.
Artigo em Inglês | MEDLINE | ID: mdl-33803085

RESUMO

Exosomes are extracellular vesicles, enriched in biomolecular cargo consisting of nucleic acids, proteins, and lipids, which take part in intercellular communication and play a crucial role in both physiologic functions and oncogenesis. Bladder cancer is the most common urinary malignancy and its incidence is steadily rising in developed countries. Despite the high five-year survival in patients diagnosed at early disease stage, survival substantially drops in patients with muscle-invasive or metastatic disease. Therefore, early detection of primary disease as well as recurrence is of paramount importance. The role that exosomal biomarkers could play in bladder cancer patient diagnosis and surveillance, as well as their potential therapeutic applications, has not been extensively studied in this malignancy. In the present review, we summarize all relevant data obtained so far from cell lines, animal models, and patient biofluids and tissues. Current literature suggests that urine is a rich source of extracellular vesicle-derived biomarkers, compared with blood and bladder tissue samples, with potential applications in bladder cancer management. Further studies improving sample collection procedures and optimizing purification and analytical methods should augment bladder cancer diagnostic, prognostic, and therapeutic input of extracellular vesicles biomarkers in the future.


Assuntos
Biomarcadores Tumorais/metabolismo , Exossomos/metabolismo , Neoplasias da Bexiga Urinária/diagnóstico , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/terapia , Exossomos/patologia , Humanos , Metástase Neoplásica , Neoplasias da Bexiga Urinária/patologia
2.
Int J Mol Sci ; 22(4)2021 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-33668652

RESUMO

Leukemia is a hematological malignancy that originates from hematopoietic stem cells in the bone marrow. Significant progress has made in understanding its pathogensis and in establishing chemotherapy and hematopoietic stem cell transplantation therapy (HSCT). However, while the successive development of new therapies, such as molecular-targeted therapy and immunotherapy, have resulted in remarkable advances, the fact remains that some patients still cannot be saved, and resistance to treatment and relapse are still problems that need to be solved in leukemia patients. The bone marrow (BM) niche is a microenvironment that includes hematopoietic stem cells and their supporting cells. Leukemia cells interact with bone marrow niches and modulate them, not only inducing molecular and functional changes but also switching to niches favored by leukemia cells. The latter are closely associated with leukemia progression, suppression of normal hematopoiesis, and chemotherapy resistance, which is precisely the area of ongoing study. Exosomes play an important role in cell-to-cell communication, not only with cells in close proximity but also with those more distant due to the nature of exosomal circulation via body fluids. In leukemia, exosomes play important roles in leukemogenesis, disease progression, and organ invasion, and their usefulness in the diagnosis and treatment of leukemia has recently been reported. The interaction between leukemia cell-derived exosomes and the BM microenvironment has received particular attention. Their interaction is believed to play a very important role; in addition to their diagnostic value, exosomes could serve as a marker for monitoring treatment efficacy and as an aid in overcoming drug resistance, among the many problems in leukemia patients that have yet to be overcome. In this paper, we will review bone marrow niches in leukemia, findings on leukemia-derived exosomes, and exosome-induced changes in bone marrow niches.


Assuntos
Medula Óssea/metabolismo , Comunicação Celular , Exossomos/metabolismo , Leucemia/metabolismo , Microambiente Tumoral , Medula Óssea/patologia , Exossomos/patologia , Humanos , Leucemia/patologia
3.
Methods Mol Biol ; 2292: 3-15, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33651347

RESUMO

Recent reports suggest that urine is a useful noninvasive tool for the identification of urogenital tumors, including bladder, prostate, kidney, and other nonurological cancers. As a liquid biopsy, urine represents an important source for the improvement of new promising biomarkers, a suitable tool to identify indolent cancer and avoid overtreatment. Urine is enriched with DNAs, RNAs, proteins, circulating tumor cells, exosomes, and other small molecules which can be detected with several diagnostic methodologies.We provide an overview of the ongoing state of urinary biomarkers underlying both their potential utilities to improve cancer prognosis, diagnosis, and therapeutic strategy and their limitations.


Assuntos
Biomarcadores Tumorais/urina , Neoplasias/urina , Animais , Ácidos Nucleicos Livres/urina , Exossomos/patologia , Humanos , Biópsia Líquida/métodos , Neoplasias/diagnóstico , Neoplasias/patologia , Ácidos Nucleicos/urina , Proteínas , Proteinúria/diagnóstico , Proteinúria/patologia , Proteinúria/urina , Urinálise/métodos
4.
Methods Mol Biol ; 2292: 115-120, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33651356

RESUMO

The analysis of liquid biopsy as a source of diagnostic, prognostic, and predictive biomarkers is still object of the main research in the prostate cancer field. Many advantages, such as less invasiveness compared to plasma or serum analysis and the rich content, confer to urine a role as an interesting fluid to be analysed especially in urological diseases. Here we report a workflow focused on profile, concentration, and protein surface characterization of EVs from urinary supernatant.


Assuntos
Exossomos/patologia , Neoplasias da Próstata/diagnóstico , Biomarcadores Tumorais/análise , Biomarcadores Tumorais/urina , Humanos , Biópsia Líquida/métodos , Masculino , Neoplasias da Próstata/patologia , Neoplasias da Próstata/urina , Proteínas/análise , Proteinúria/diagnóstico , Proteinúria/patologia , Proteinúria/urina , Coleta de Urina/métodos , Fluxo de Trabalho
5.
Methods Mol Biol ; 2265: 305-321, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33704724

RESUMO

Tumor-derived exosomes (TEX), a subset of small extracellular vesicles (EVs) which originate from the endocytic compartment of tumor cells, are emerging as key players in cancer progression. TEX circulate freely in patients' body fluids and transfer bioactive cargos from tumor to various recipient cells. The molecular cargo of melanoma cell-derived exosomes (MTEX) mimics that of the tumor, and MTEX serve as a liquid biopsy that provides potentially useful information for cancer diagnosis, prognosis, or responses to therapy. Plasma of melanoma patients contains a mix of MTEX and exosomes produced by nonmalignant cells (NMTEX). Isolation of these exosome subtypes from the bulk of plasma exosomes is necessary to evaluate contributions of each as potential biomarkers of melanoma progression and outcome. Here, methods for separation of MTEX from T cell-derived exosomes from a single small volume of plasma and their subsequent molecular and functional characterization are described. Following size exclusion chromatography (SEC) to isolate total plasma exosomes, immune affinity-based capture of MTEX with anti-CSPG4 antibody and then of exosomes produced by T cells with anti-CD3 antibody is used to sequentially isolate the two subsets. This immune capture method enables the recovery of MTEX and CD3+ exosomes in quantities sufficient both for molecular profiling by flow cytometry or western blotting and for functional analyses.


Assuntos
Biomarcadores Tumorais/sangue , Western Blotting , Exossomos/metabolismo , Citometria de Fluxo , Melanoma/sangue , Linfócitos T/metabolismo , Cromatografia em Gel , Exossomos/patologia , Humanos , Biópsia Líquida , Plasma/metabolismo , Linfócitos T/patologia
6.
Am J Physiol Heart Circ Physiol ; 320(4): H1213-H1234, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33513083

RESUMO

Exosomes are a subgroup of extracellular bilayer membrane nanovesicles that are enriched in a variety of bioactive lipids, receptors, transcription factors, surface proteins, DNA, and noncoding RNAs. They have been well recognized to play essential roles in mediating intercellular signaling by delivering bioactive molecules from host cells to regulate the physiological processes of recipient cells. In the context of heart diseases, accumulating studies have indicated that exosome-carried cellular proteins and noncoding RNA derived from different types of cardiac cells, including cardiomyocytes, fibroblasts, endothelial cells, immune cells, adipocytes, and resident stem cells, have pivotal roles in cardiac remodeling under disease conditions such as cardiac hypertrophy, diabetic cardiomyopathy, and myocardial infarction. In addition, exosomal contents derived from stem cells have been shown to be beneficial for regenerative potential of the heart. In this review, we discuss current understanding of the role of exosomes in cardiac communication, with a focus on cardiovascular pathophysiology and perspectives for their potential uses as cardiac therapies.


Assuntos
Comunicação Celular , Exossomos/metabolismo , Cardiopatias/metabolismo , Miocárdio/metabolismo , Transdução de Sinais , Transplante de Células-Tronco , Animais , Exossomos/patologia , Exossomos/transplante , Cardiopatias/patologia , Cardiopatias/fisiopatologia , Cardiopatias/cirurgia , Humanos , Miocárdio/patologia
7.
J Cancer Res Clin Oncol ; 147(3): 637-648, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33511427

RESUMO

BACKGROUND: Exosomes are extracellular nanometric vesicles used by cells to communicate with each other. They are responsible for many pathological conditions, including tumors by transferring regulatory biomolecules that impact target cell activity. Because of their high concentration in exosomes compared with parental cells and the rest of exosomal content, specificity to the cell of origin, and their well-organized sorting mechanism, microRNAs (miRNAs) are thought to be the most potent exosomes cargo and used by scientists to track exosomes and to detect cell activity changes and prognosis in cancer early. PURPOSE: In this review, the results of studies examining the role of exosomes in cancer pathophysiology and their clinical potential are discussed in detail. Tumor-derived exosomes (TDEs) mediate the dynamic changes of cancer growth and invasion, including local microenvironment remodeling, distance metastasis, angiogenesis, and tumor-associated immunosuppression. They also contribute to hypoxia-induced tumor progression and cancer cell drug resistance. As a result of exosomes being present in all body fluids, it is possible to have early accessible and less-invasive diagnostic and prognostic measures by forming a table for each cancer type and its matched specific miRNAs. Under testing, available therapeutic uses of exosomes include interference of exosomes biogenesis, secretion, or uptake, and recruitment of exosomes as target-specific drug delivery vehicles, and immunostimulatory agents for both cancer patients and healthy population to avoid cancer development from the start. CONCLUSION: These data suggest that exosomes and exosomal microRNA are directly related to cancer progression mechanisms, and could be used in cancer early diagnosis, prognosis, and therapy.


Assuntos
Exossomos/patologia , MicroRNAs/genética , MicroRNAs/metabolismo , Neoplasias/patologia , Animais , Progressão da Doença , Exossomos/genética , Exossomos/metabolismo , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/terapia
8.
Methods Mol Biol ; 2174: 171-191, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32813250

RESUMO

The study of tumor exosomes has gained relevance in the last decades due to their potential use for therapeutic and diagnostic application. Although there is extensive knowledge of exosome biology, some biological samples like tumor-derived exosomes have been difficult to characterize due to their complexity and heterogeneity. This distinctive feature makes difficult the identification of specific exosome subpopulations with a shared molecular signature that could allow for targeting of exosomes with therapeutic and diagnostic potential use in cancer patients. Nanoscale flow cytometry has lately emerged as an alternative tool that can be adapted to the study of nanoparticles, such as exosomes. However, the physicochemical properties of these particles are an important issue to consider as nanoparticles need the application of specific settings which differ from those used in conventional flow cytometry of cells. Therefore, in the last few years, one of the main aims has been the optimization of technical and experimental protocols to improve exosome analysis. In this chapter, we discuss several aspects of cytometric systems with a special emphasis in technical considerations of samples and equipment.


Assuntos
Exossomos/química , Exossomos/patologia , Citometria de Fluxo/métodos , Neoplasias/patologia , Calibragem , Centrifugação com Gradiente de Concentração/métodos , Cromatografia em Gel/métodos , Citometria de Fluxo/instrumentação , Humanos , Nanotecnologia/instrumentação , Nanotecnologia/métodos , Prognóstico , Ultracentrifugação/métodos
9.
Methods Mol Biol ; 2174: 143-170, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32813249

RESUMO

Extracellular vesicles (EVs) produced by cancer cells function as a unique form of intercellular communication that can promote cell growth and survival, help shape the tumor microenvironment, and increase invasive and metastatic activity. There are two major classes of EVs, microvesicles (MVs) and exosomes, and they differ in how they are formed. MVs are generated by the outward budding and fission of the plasma membrane. On the other hand, exosomes are derived as multivesicular bodies (MVBs) fuse with the plasma membrane and release their contents. What makes EVs especially interesting is how they mediate their effects. Both MVs and exosomes have been shown to contain a wide-variety of bioactive cargo, including cell surface, cytosolic, and nuclear proteins, as well as RNA transcripts, micro-RNAs (miRNAs), and even fragments of DNA. EVs, and their associated cargo, can be transferred to other cancer cells, as well as to normal cell types, causing the recipient cells to undergo phenotypic changes that promote different aspects of cancer progression. These findings, combined with those demonstrating that the amounts and contents of EVs produced by cancer cells can vary depending on their cell of origin, stage of development, or response to therapies, have raised the exciting possibility that EVs can be used for diagnostic purposes. Moreover, the pharmaceutical community is aggressively pursuing the use of EVs as a potential drug delivery platform. Here, in this chapter, we will highlight what is currently known about how EVs are generated, how they impact cancer progression, and the different ways they are being exploited for clinical applications.


Assuntos
Sistemas de Liberação de Medicamentos/métodos , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/patologia , Biópsia Líquida/métodos , Neoplasias/patologia , Membrana Celular/metabolismo , Membrana Celular/patologia , Exossomos/metabolismo , Exossomos/patologia , Vesículas Extracelulares/classificação , Humanos , Neoplasias/irrigação sanguínea , Neovascularização Patológica/patologia , Microambiente Tumoral/imunologia
10.
Aging (Albany NY) ; 12(23): 23598-23608, 2020 12 13.
Artigo em Inglês | MEDLINE | ID: mdl-33310972

RESUMO

The expression of Hic-5 was detected in osteosarcoma patients and osteosarcoma cell lines by RT-PCR. Then RFP-sh-Hic-5 was transfected into osteosarcoma cell lines. The effect of Hic-5 on cell viability, proliferation and apoptosis were assessed by MTT, EdU kit and Flow cytometry. The exosomes were isolated from MG-63 cell supernatant by an Exosome Isolation Kit. The exosome-Hic-5 was confirmed by transmission electron microscope, particle size detection and RT-PCR. Next, exosome-Hic-5 treated cells were explored the cell viability, proliferation and apoptosis. Further, Co-IP assay was employed for identifying the relationship between Hic-5 and smad4. TCF/LEF and the protein level of components of wnt/ß-catenin signals were detected by TOP luciferase assay and western blot. Hic-5 was upregulated in osteosarcoma tissues and cell. Forced decreased expression Hic-5 inhibited the proliferation of osteosarcoma cell lines, and induced apoptosis of MG-63 and HOS. In vivo, silencing Hic-5 remitted the tumor progression. Further, we isolated the exosomes from MG-63 supernatant, exosomes concluding Hic-5 would regulated the proliferation and apoptosis level of MG-63 and HOS cells. Further, Hic-5 interacted with smad4 and regulated Wnt/ß-catenin signal by decreasing TCF/LEF activity. Silencing Hic-5 inhibited the proliferation and induced apoptosis of osteosarcoma cell via inactivating Wnt/ß-catenin signal by exosome pathway.


Assuntos
Apoptose , Neoplasias Ósseas/metabolismo , Proliferação de Células , Exossomos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas com Domínio LIM/metabolismo , Osteossarcoma/metabolismo , Via de Sinalização Wnt , Adolescente , Adulto , Animais , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Criança , Exossomos/genética , Exossomos/patologia , Exossomos/transplante , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas com Domínio LIM/genética , Masculino , Camundongos Nus , Osteossarcoma/genética , Osteossarcoma/patologia , Carga Tumoral , Adulto Jovem
11.
Int J Nanomedicine ; 15: 8019-8036, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33116515

RESUMO

Exosomes are a subset of tiny extracellular vesicles manufactured by all cells and are present in all body fluids. They are produced actively in tumor cells, which are released and utilized to facilitate tumor growth. Their characteristics enable them to assist major cancer hallmarks, leveraged by cancer cells in fostering cancer growth and spread while implementing ways to escape elimination from the host environment. This review updates on the latest progress on the roles of cancer-derived exosomes, of 30-100 nm in size, in deregulating paracrine trafficking in the tumor microenvironment and circulation. Thus, exosomes are being exploited in diagnostic biomarker development, with its potential in clinical applications as therapeutic targets utilized in exosome-based nanoparticle drug delivery strategies for cancer therapy. Ongoing studies were retrieved from PubMed® and Scopus database and ClinicalTrials.gov registry for review, highlighting how cancer cells from entirely different cell lines rely on genetic information carried by their exosomes for homotypic and heterotypic intercellular communications in the microenvironment to favor proliferation and invasion, while establishing a pre-metastatic niche in welcoming cancer cells' arrival. We will elaborate on the trafficking of tumor-derived exosomes in fostering cancer proliferation, invasion, and metastasis in hematopoietic (leukemia and myeloma), epithelial (breast cancer), and mesenchymal (soft tissue sarcoma and osteosarcoma) cancers. Cancer-derived exosomal trafficking is observed in several types of liquid or solid tumors, confirming their role as cancer hallmark enabler. Their enriched genetic signals arising from their characteristic DNA, RNA, microRNA, and lncRNA, along with specific gene expression profiles, protein, or lipid composition carried by the exosomal cargo shed into blood, saliva, urine, ascites, and cervicovaginal lavage, are being studied as a diagnostic, prognostic, or predictive cancer biomarker. We reveal the latest research efforts in exploiting the use of nanoparticles to improve the overall cancer diagnostic capability in the clinic.


Assuntos
Biomarcadores Tumorais/metabolismo , Exossomos/patologia , Neoplasias/metabolismo , Neoplasias/patologia , Humanos , Neoplasias/tratamento farmacológico
12.
Yonsei Med J ; 61(9): 750-761, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32882759

RESUMO

PURPOSE: Gastric cancer (GC) is a malignant tumor with a high mortality rate. Drug resistance is a major obstacle to GC therapy. This study aimed to investigate the role and mechanism of exosomal circPRRX1 in doxorubicin resistance in GC. MATERIALS AND METHODS: HGC-27 and AGS cells were exposed to different doses of doxorubicin to construct doxorubicin-resistant cell lines. Levels of circPRRX1, miR-3064-5p, and nonreceptor tyrosine phosphatase 14 (PTPN14) were detected by quantitative real-time PCR or Western blot assay. Then, 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide, transwell, and Western blot assays were used to explore the function of circPRRX1 in GC cells. Interactions among circPRRX1, miR-3064-5p, and PTPN14 were confirmed by dual-luciferase reporter assay. The in vivo function of circPRRX1 was analyzed in a xenograft tumor model. RESULTS: CircPRRX1 was highly expressed in doxorubicin-resistant GC cell lines. Knockdown of circPRRX1 reversed doxorubicin resistance in doxorubicin-resistant GC cells. Additionally, extracellular circPRRX1 was carried by exosomes to spread doxorubicin resistance. CircPRRX1 silencing reduced doxorubicin resistance by targeting miR-3064-5p or regulating PTPN14. In GC patients, high levels of circPRRX1 in serum exosomes were associated with poor responses to doxorubicin treatment. Moreover, depletion of circPRRX1 reduced doxorubicin resistance in vivo. CONCLUSION: CircPRRX1 strengthened doxorubicin resistance by modulating miR-3064-5p/PTPN14 signaling and might be a therapeutic target for GC patients.


Assuntos
Doxorrubicina/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , MicroRNAs/genética , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/metabolismo , Linhagem Celular Tumoral , Exossomos/genética , Exossomos/metabolismo , Exossomos/patologia , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio , Humanos , MicroRNAs/metabolismo , Proteínas Tirosina Fosfatases não Receptoras/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/genética , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia
13.
Am J Physiol Renal Physiol ; 319(4): F664-F673, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32715764

RESUMO

Tubular changes contribute to the development of renal pathologies in diabetic kidney disease (DKD), including interstitial fibrosis. It is unclear how tubular cells relay signals to interstitial fibroblasts. Recently, exosomes have been recognized as crucial mediators of intercellular communication. We hypothesized that exosomes secreted from tubular cells may stimulate fibroblasts for interstitial fibrosis in DKD. In this study, we isolated and purified exosomes from the renal cortex of DKD mice and high glucose-treated mouse proximal tubular cells. Compared with nondiabetic mice, exosome secretion in kidney tissues decreased in DKD mice. Likewise, high glucose incubation reduced exosome secretion in mouse kidney proximal tubular BUMPT cells. To study the effect of tubular cell exosomes on fibroblasts, exosomes from BUMPT cells were added to renal fibroblast NRK-49F cell cultures. Notably, exosomes from high glucose conditioned BUMPT cells induced higher proliferation, significant morphological change, and substantial production of fibronectin, α-smooth muscle actin, and collagen type Ι in NRK-49F fibroblasts. Proteomics analysis was further performed to profile the proteins within tubular cell exosomes. Interestingly, 22 proteins were found to be differentially expressed between tubular exosomes derived from high glucose conditioned cells and those from normal glucose conditioned cells. Cytoscape analysis suggested the existence of two protein-protein interaction networks in these exosomal differentially expressed proteins. While one of the protein-protein interaction networks comprised enolase 1 (Eno1), heat shock protein family A member 8 (Hspa8), thioredoxin 1 (Txn1), peptidylprolyl isomerase A (Ppia), phosphoglycerate kinase 1 (Pgk1), DNA topoisomerase II-ß (Top2b), and ß-actin (Actb), the other had the family proteins of human leucocyte antigen F (Ywhag), a component of the ND10 nuclear body (Ywhae), interferon regulatory factor-8 (Ywhaq), and human leucocyte antigen A (Ywhaz). Gene expression analysis via Nephroseq showed a correlation of Eno1 expression with DKD clinical manifestation. In conclusion, DKD is associated with a decrease in exosome secretion in renal tubular cells. Exosomes from high glucose conditioned tubular cells may regulate the proliferation and activation of fibroblasts, contributing to the paracrine signaling mechanism responsible for the pathological onset of renal interstitial fibrosis in DKD.


Assuntos
Proliferação de Células , Nefropatias Diabéticas/metabolismo , Exossomos/metabolismo , Fibroblastos/metabolismo , Túbulos Renais Proximais/metabolismo , Comunicação Parácrina , Animais , Linhagem Celular , Técnicas de Cocultura , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/patologia , Modelos Animais de Doenças , Progressão da Doença , Exossomos/genética , Exossomos/patologia , Fibroblastos/patologia , Fibrose , Túbulos Renais Proximais/patologia , Masculino , Camundongos Endogâmicos C57BL , Fosfopiruvato Hidratase/metabolismo , Mapas de Interação de Proteínas , Via Secretória , Transdução de Sinais
14.
Toxicol Appl Pharmacol ; 401: 115109, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32544403

RESUMO

Bladder cancer (BCa) is the fourth leading cause of cancer deaths worldwide due to its aggressiveness and resistance against therapies. Intricate interactions between cancer cells and the tumor microenvironment (TME) are essential for both disease progression and regression. Thus, interrupting molecular communications within the TME could potentially provide improved therapeutic efficacies. M2-polarized tumor-associated macrophages (M2 TAMs) were shown to contribute to BCa progression and drug resistance. We attempted to provide evidence for ovatodiolide (OV) as a potential therapeutic agent that targets both TME and BCa cells. First, tumor-suppressing functions of OV were determined by cell viability, colony, and tumor-sphere formation assays using a coculture system composed of M2 TAMs/BCa cells. Subsequently, we demonstrated that extracellular vesicles (EVs) isolated from M2 TAMs containing oncomiR-21 and mRNAs, including Akt, STAT3, mTOR, and ß-catenin, promoted cisplatin (CDDP) resistance, migration, and tumor-sphere generation in BCa cells, through increasing CDK6, mTOR, STAT3, and ß-catenin expression. OV treatment also prevented M2 polarization and reduced EV cargos from M2 TAMs. Finally, in vivo data demonstrated that OV treatment overcame CDDP resistance. OV only and the OV + CDDP combination both resulted in significant reductions in mTOR, ß-catenin, CDK6, and miR-21 expression in tumor samples and EVs isolated from serum. Collectively, we demonstrated that M2 TAMs induced malignant properties in BCa cells, in part via oncogenic EVs. OV treatment prevented M2 TAM polarization, reduced EV cargos derived from M2 TAMs, and suppressed ß-catenin/mTOR/CDK6 signaling. These findings provide preclinical evidence for OV as a single or adjuvant agent for treating drug-resistant BCa.


Assuntos
Quinase 6 Dependente de Ciclina/metabolismo , Diterpenos/uso terapêutico , MicroRNAs/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Neoplasias da Bexiga Urinária/metabolismo , beta Catenina/metabolismo , Animais , Carcinogênese/efeitos dos fármacos , Carcinogênese/metabolismo , Linhagem Celular Tumoral , Técnicas de Cocultura , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Diterpenos/isolamento & purificação , Diterpenos/farmacologia , Relação Dose-Resposta a Droga , Exossomos/efeitos dos fármacos , Exossomos/metabolismo , Exossomos/patologia , Feminino , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , MicroRNAs/antagonistas & inibidores , Plantas Medicinais , Serina-Treonina Quinases TOR/antagonistas & inibidores , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/patologia , beta Catenina/antagonistas & inibidores
15.
Cell Prolif ; 53(8): e12865, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32588948

RESUMO

The tumour microenvironment (TME) plays a pivotal role in tumour fate determination. The TME acts together with the genetic material of tumour cells to determine their initiation, metastasis and drug resistance. Stromal cells in the TME promote the growth and metastasis of tumour cells by secreting soluble molecules or exosomes. The abnormal microenvironment reduces immune surveillance and tumour killing. The TME causes low anti-tumour drug penetration and reactivity and high drug resistance. Tumour angiogenesis and microenvironmental hypoxia limit the drug concentration within the TME and enhance the stemness of tumour cells. Therefore, modifying the TME to effectively attack tumour cells could represent a comprehensive and effective anti-tumour strategy. Normal cells, such as stem cells and immune cells, can penetrate and disrupt the abnormal TME. Reconstruction of the TME with healthy cells is an exciting new direction for tumour treatment. We will elaborate on the mechanism of the TME to support tumours and the current cell therapies for targeting tumours and the TME-such as immune cell therapies, haematopoietic stem cell (HSC) transplantation therapies, mesenchymal stem cell (MSC) transfer and embryonic stem cell-based microenvironment therapies-to provide novel ideas for producing breakthroughs in tumour therapy strategies.


Assuntos
Antineoplásicos/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Neovascularização Patológica/tratamento farmacológico , Microambiente Tumoral/efeitos dos fármacos , Exossomos/efeitos dos fármacos , Exossomos/patologia , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/patologia
16.
Cell Prolif ; 53(7): e12833, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32525231

RESUMO

OBJECTIVES: The current study aimed to investigate the mechanism by which exosomes secreted by CHB patients with PNALT and liver inflammation grade (≥A2) affected the development of liver cancer. MATERIALS AND METHODS: Gene expression was assessed by RT-PCR, Western blotting and immunohistochemistry. CCK-8, colony formation, transwell, scratch-wound and flow cytometry assays were used to detect cell viability, proliferation, apoptosis and metastasis. The interaction of TCF21 and HHIP was assessed by co-immunoprecipitation assay. Luciferase reporter was used to detect the combination of TCF21/HHIP and miR-25-3p. Xenograft studies in nude mice manifested tumour growth ability of miR-25-3p. Bioinformatics analyses were conducted using TargetScan, EVmiRNA, TCGA, GEO, DAVID, COEXPEDIA, UALCAN, UCSC and the Human Protein Atlas databases. RESULTS: CHB-PNALT-Exo (≥A2) promoted the proliferation and metastasis of HepG2.2.15 cells. miR-25-3p was upregulated in CHB-PNALT-Exo (≥A2). miR-25-3p overexpression promoted cell proliferation and metastasis and was related to poor survival in patients with CHB-PNALT (≥A2). The cell proliferation- and metastasis-promoting functions of CHB-PNALT-Exo (≥A2) were abolished by miR-25-3p inhibitors. TCF21 directly interacted with HHIP. Inhibition of TCF21 or HHIP promoted cell proliferation and metastasis. Knockdown of TCF21 or HHIP counteracted the effects of CHB-PNALT-Exo (≥A2) containing miR-25-3p inhibitor on cell proliferation, metastasis and the expression of Ki67, E-cadherin and caspase-3/-9. CONCLUSIONS: Transfer of miR-25-3p by CHB-PNALT-Exo promoted the development of liver cancer by inhibiting the co-expression of TCF21 and HHIP.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas de Transporte/genética , Exossomos/genética , Hepatite B Crônica/genética , Neoplasias Hepáticas/genética , Fígado/patologia , Glicoproteínas de Membrana/genética , MicroRNAs/genética , Adulto , Animais , Apoptose/genética , Proliferação de Células/genética , Sobrevivência Celular , Progressão da Doença , Regulação para Baixo/genética , Exossomos/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Hepatite B Crônica/patologia , Humanos , Inflamação/genética , Inflamação/patologia , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Regulação para Cima/genética , Adulto Jovem
17.
Exp Hematol ; 86: 43-52.e1, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32417302

RESUMO

Whole transferrin receptor (TfR) is present in reticulocyte exosomes. Soluble transferrin receptor (sTfR) is cleaved from whole TfR in human plasma, with the remnant cytoplasmic domain (cTfR) remaining membrane associated. In humans, sTfR is a biomarker that can detect iron deficiency in the presence of inflammatory disease. This condition is still a diagnostic dilemma in veterinary species. We aimed to (1) confirm the presence of exosomes and exosome-associated TfR in the serum of dogs, cats, and horses; and (2) to assess and compare the proportion of cTfR to total (cTfR + whole) in exosomal membranes of healthy and diseased dogs and cats and in healthy horses to indirectly predict their anticipated levels of circulating sTfR. We used discarded serum and whole blood samples from canine and feline patients, separated into healthy and diseased groups based on the health status of each patient, and healthy equine participants from a previous study. Ultracentrifugation, followed in some experiments by OptiPrep discontinuous density gradient fractionation, was used to isolate exosomes. Exosomes and associated TfR were identified using TEM and Western blot for TfR, respectively. Densitometry tracings of Western blots of serum exosomes were used to measure the proportion of cTfR to total TfR. Extracellular vesicles compatible with exosomes were successfully isolated and expressed TfR. The proportion of cTfR in dogs was greater than 50%, indicating that a majority of the whole TfR was cleaved to produce sTfR (and remnant cTfR). There was significant interindividual variation and no significant difference between healthy and diseased animals. The proportion of cTfR in cats was very low at 11%, indicating that very little sTfR was likely produced. There was a small yet significant difference between healthy and diseased cats. Healthy horses do not appear to cleave exosome-associated TfR. Diagnosis of iron deficiency in the presence of inflammatory disease remains a challenge in veterinary medicine. Our results indicate that TfR is poorly or unpredictably cleaved in veterinary species, revealing that there are species differences in exosomal TfR handling. These data suggest that development of an assay for the detection and quantification of sTfR in the species investigated may not be warranted.


Assuntos
Doenças do Gato/sangue , Doenças do Cão/sangue , Exossomos/metabolismo , Doenças dos Cavalos/sangue , Receptores da Transferrina/sangue , Animais , Doenças do Gato/patologia , Gatos , Doenças do Cão/patologia , Cães , Exossomos/patologia , Doenças dos Cavalos/patologia , Cavalos
18.
Oncogene ; 39(24): 4681-4694, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32398867

RESUMO

We previously identified that the development of early-stage myeloid-derived suppressor cells (eMDSCs) in breast cancer with high IL-6 (IL-6high) expression was correlated with the SOCS3 deficiency-dependent hyperactivation of the JAK/STAT signaling pathway. However, the regulatory mechanisms have not yet been elucidated. In this study, we aimed to investigate how the posttranscriptional regulation mediated by cancer exosome-derived miRNAs affected the JAK/STAT signaling pathway and the development of eMDSCs. Using miRNA microarray, we screened miR-9 and miR-181a which were exclusively upregulated in eMDSCs and inversely associated with SOCS3 expression. We found both miRNAs promoted the amplification of immature eMDSCs with the strong suppression on T-cell immunity in mice and humans. Furthermore, miR-9 and miR-181a promoted 4T1 tumor growth and immune escape via enhancing eMDSCs infiltration in situ. But miR-9 and miR-181a stimulated eMDSCs development by separately inhibiting SOCS3 and PIAS3, two crucial regulators in the negative feedback loop of the JAK/STAT signaling pathway. Elevated miR-9 and miR-181a in eMDSCs was derived from tumor-derived exosomes, and blocking the exosome release could fully attenuate the miRNA-mediated regulation on eMDSCs development. In summary, our findings indicated that tumor exosome-derived miR-9 and miR-181a activated the JAK/STAT signaling pathway via targeting SOCS3 and PIAS3, respectively, and thus promoted the expansion of eMDSCs which might provide potential therapeutic target for IL-6high breast cancer treatment.


Assuntos
Neoplasias da Mama/imunologia , Exossomos/imunologia , Neoplasias Mamárias Experimentais/imunologia , MicroRNAs/imunologia , Chaperonas Moleculares/imunologia , Células Supressoras Mieloides/imunologia , Proteínas Inibidoras de STAT Ativados/imunologia , Proteína 3 Supressora da Sinalização de Citocinas/imunologia , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Exossomos/genética , Exossomos/patologia , Feminino , Humanos , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , MicroRNAs/genética , Chaperonas Moleculares/genética , Células Supressoras Mieloides/patologia , Proteínas Inibidoras de STAT Ativados/genética , Proteína 3 Supressora da Sinalização de Citocinas/genética
19.
Sensors (Basel) ; 20(5)2020 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-32121271

RESUMO

Cancer represents one of the conditions with the most causes of death worldwide. Common methods for its diagnosis are based on tissue biopsies-the extraction of tissue from the primary tumor, which is used for its histological analysis. However, this technique represents a risk for the patient, along with being expensive and time-consuming and so it cannot be frequently used to follow the progress of the disease. Liquid biopsy is a new cancer diagnostic alternative, which allows the analysis of the molecular information of the solid tumors via a body fluid draw. This fluid-based diagnostic method displays relevant advantages, including its minimal invasiveness, lower risk, use as often as required, it can be analyzed with the use of microfluidic-based platforms with low consumption of reagent, and it does not require specialized personnel and expensive equipment for the diagnosis. In recent years, the integration of sensors in microfluidics lab-on-a-chip devices was performed for liquid biopsies applications, granting significant advantages in the separation and detection of circulating tumor nucleic acids (ctNAs), circulating tumor cells (CTCs) and exosomes. The improvements in isolation and detection technologies offer increasingly sensitive and selective equipment's, and the integration in microfluidic devices provides a better characterization and analysis of these biomarkers. These fully integrated systems will facilitate the generation of fully automatized platforms at low-cost for compact cancer diagnosis systems at an early stage and for the prediction and prognosis of cancer treatment through the biomarkers for personalized tumor analysis.


Assuntos
Detecção Precoce de Câncer/métodos , Biópsia Líquida/métodos , Microfluídica/métodos , Biomarcadores Tumorais/sangue , Ácidos Nucleicos Livres/sangue , Exossomos/patologia , Humanos , Dispositivos Lab-On-A-Chip , Células Neoplásicas Circulantes/patologia
20.
PLoS One ; 15(3): e0229602, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32126572

RESUMO

AIM: This study analyzed microvesicles and exosomes, called as extracellular vesicles (EVs) excreted in serum and cerebrospinal fluid (CSF) from patients with cerebral or gestational toxoplasmosis. METHODS: Clinical samples from 83 individuals were divided into four groups. Group I, 20 sera from healthy individuals and pregnant women (seronegative for toxoplasmosis); group II, 21 sera from seropositive patients for toxoplasmosis (cerebral or gestational forms); group III, 26 CSF samples from patients with cerebral toxoplasmosis/HIV co-infection (CT/HIV) (seropositive for toxoplasmosis); and group IV, 16 CSF samples from seronegative patients for toxoplasmosis, but with HIV infection and other opportunistic infections (OI/HIV). Serum and CSF samples were ultracentrifuged to recover EVs. Next, vesicle size and concentration were characterized by Nanoparticle Tracking Analysis (NTA). RESULTS: Concentrations of serum-derived EVs from toxoplasmosis patients (mean: 2.4 x 1010 EVs/mL) were statically higher than of non-infected individuals (mean: 5.9 x 109 EVs/mL). Concentrations of CSF-derived EVs were almost similar in both groups. CT/HIV (mean: 2.9 x 109 EVs/mL) and OI/HIV (mean: 4.8 x 109 EVs/mL). Analyses by NTA confirmed that CSF-derived EVs and serum-derived EVs had size and shape similar to microvesicles and exosomes. The mean size of EVs was similar in serum and CSF. Thus, the concentration, and not size was able distinguish patients with toxoplasmosis than healthy individuals. Presence of exosomes was also confirmed by transmission electron microscopy and evidence of tetraspanins CD63 and CD9 in immunoblotting. Relative expressions of miR-146a-5p, miR-155-5p, miR-21-5p, miR-29c-3p and miR-125b-5p were estimated in exosomal miRNA extracted of EVs. Serum-derived EVs from group II (cerebral and gestational toxoplasmosis) up-expressed miR-125b-5p and miR-146a-5p. CSF-derived EVs from CT/HIV patients) up-expressed miR-155-5p and miR-21-5p and were unable to express miR-29c-3p. CONCLUSION: These data suggest the participation of EVs and exosomal miRNAs in unbalance of immune response as elevation of TNF-α, IL-6; and downregulation of IFN-γ in cerebral and gestational forms of toxoplasmosis.


Assuntos
Complicações Parasitárias na Gravidez/sangue , Complicações Parasitárias na Gravidez/líquido cefalorraquidiano , Toxoplasmose Cerebral/sangue , Toxoplasmose Cerebral/líquido cefalorraquidiano , Toxoplasmose/complicações , Micropartículas Derivadas de Células/genética , Micropartículas Derivadas de Células/patologia , Exossomos/genética , Exossomos/patologia , Vesículas Extracelulares/genética , Vesículas Extracelulares/patologia , Feminino , Expressão Gênica , Infecções por HIV/sangue , Infecções por HIV/líquido cefalorraquidiano , Infecções por HIV/complicações , Voluntários Saudáveis , Humanos , MicroRNAs/sangue , MicroRNAs/líquido cefalorraquidiano , MicroRNAs/genética , Microscopia Eletrônica de Transmissão , Gravidez , Complicações Parasitárias na Gravidez/genética , Toxoplasmose/sangue , Toxoplasmose/líquido cefalorraquidiano , Toxoplasmose Cerebral/genética
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