detection-exosomal-biomarker-electric-field-induced-release

Detection of Exosomal Biomarker by Electric Field-induced Release and Measurement (EFIRM)

Exosomes are microvesicular structures that play a mediating role in intercellular communication. It is of interest to study the internal cargo of ...

University of California, Los Angeles

The cellular and molecular mechanisms that underlie skeletal abnormalities in defective Recql4-related syndromes are poorly understood. Our objective in this study was to explore the function of Recql4 in osteoblast biology both in vitro and in vivo. Immunohistochemistry on adult mouse bone showed Recql4 protein localization in active osteoblasts around growth plate, but not in fully differentiated osteocytes. Consistent with this finding, Recql4 gene expression was high in proliferating mouse osteoblastic MC3T3.E1 cells and decreased as cells progressively lost their proliferation activity during differentiation. Recql4 overexpression in osteoblastic cells exhibited higher proliferation activity, while its depletion impeded cell growth. In addition, bone marrow stromal cells from male Recql4+/- mice had fewer progenitor cells, including osteoprogenitors, indicated by reduced total fibroblast colony forming units (CFU-f) and alkaline phosphatase-positive CFU-f colonies concomitant with reduced bone mass. These findings provide evidence that Recql4 functions as a regulatory protein during osteoprogenitor proliferation, a critical cellular event during skeleton development.

Recql4 haploinsufficiency in mice leads to defects in osteoblast progenitors: Implications for low bone mass phenotype.

Rothmund-Thomson syndrome (RTS) is a heterogeneous disease, associated with increased prevalence of osteosarcoma in very young patients with a mutated RECQL4 gene. In this study, we tested the ability of RECQL4 deficient fibroblasts, derived from a RTS patient to recover from hydrogen peroxide (H(2)O(2))-induced oxidative stress/damage. Immunoperoxidase staining for 8-oxo-deoxyguanosine (8-oxo-dG) formation in RTS and normal human fibroblasts were compared to assess DNA damage. We determined DNA synthesis, cell growth, cell cycle distribution, and viability in RTS and normal human fibroblasts before and after H(2)O(2) treatment. H(2)O(2) induces 8-oxo-dG formation in both RTS and normal fibroblasts. In normal human fibroblasts, RECQL4 was predominantly localized to cytoplasm; nuclear translocation and foci formation occurred in response to oxidant stimulation. After recovery from oxidant exposure, viable RTS fibroblasts showed irreversible growth arrest compared to normal fibroblasts. DNA synthesis decreased significantly in treated RTS cells, with concomitant reduction of cells in the S-phase. These results suggest that enhanced oxidant sensitivity in RECQL4 deficient fibroblasts derived from RTS patients could be attributed to abnormal DNA metabolism and proliferation failure. The ramifications of these findings on osteosarcoma prevalence and heterogeneity in RTS are discussed.

RECQL4-deficient cells are hypersensitive to oxidative stress/damage: Insights for osteosarcoma prevalence and heterogeneity in Rothmund-Thomson syndrome.

The expression of matrix metalloproteinase-13 (MMP-13), involved in bone turnover, is elevated in stretched MC3T3-E1 osteoblast-like cells. Strain-mediated forces impact bone remodeling due in large part to the movement of fluid through the canalicular-lacunar network. The resulting fluid shear stress (FSS) over the surface membranes of bone cells initiates bone remodeling. Although the nuclear events mediating putative FSS-induced changes in osteoblast MMP-13 transcription are unknown, previous studies with bone cells suggest an overlap between osteoblast FSS- and PTH-induced signal response pathways. MMP-13 PTH response is regulated by a 110 bp 5' regulatory region, conserved across the mouse, rat, and human genes, that supports the binding of numerous transcription factors including Runx2, c-fos/c-jun, Ets-1, and nuclear matrix protein 4/cas interacting zinc finger protein (Nmp4/CIZ) a nucleocytoplasmic shuttling trans-acting protein that attenuates PTH-driven transcription. Nmp4/CIZ also binds p130(cas), an adaptor protein implicated in mechanotransduction. Here we sought to determine whether Nmp4/CIZ contributes to FSS-induced changes in MMP-13 transcription. FSS (12 dynes/cm(2), 3-5 h) increased MMP-13 promoter-reporter activity approximately two-fold in MC3T3-E1 osteoblast-like cells attended by a comparable increase in mRNA expression. This was accompanied by a decrease in Nmp4/CIZ binding to its cis-element within the PTH response region, the mutation of which abrogated the MMP-13 response to FSS. Interestingly, FSS enhanced Nmp4/CIZ promoter activity and induced p130(cas) nuclear translocation. We conclude that the PTH regulatory region of MMP-13 also contributes to FSS response and that Nmp4/CIZ plays similar but distinct roles in mediating hormone- and FSS-driven induction of MMP-13 in bone cells.

Nmp4/CIZ contributes to fluid shear stress induced MMP-13 gene induction in osteoblasts.

High mobility group box 1 (HMGB1) is a chromatin protein that acts as an immunomodulatory cytokine upon active release from myeloid cells. HMGB1 is also an alarmin, an endogenous molecule released by dying cells that acts to initiate tissue repair. We have previously reported that osteoclasts and osteoblasts release HMGB1 and release by the latter is regulated by parathyroid hormone (PTH), an agent of bone remodeling. A recent study suggests that HMGB1 acts as a chemotactic agent to osteoclasts and osteoblasts during endochondral ossification. To explore the potential impact of HMGB1 in the bone microenvironment and its mechanism of release by osseous cells, we characterized the effects of recombinant protein (rHMGB1) on multiple murine bone cell preparations that together exhibit the various cell phenotypes present in bone. We also inquired whether apoptotic bone cells release HMGB1. rHMGB1 enhanced the RANKL/OPG steady state mRNA ratio and dramatically augmented the release of tumor necrosis factor-alpha (TNFalpha) and interleukin-6 (IL6) in osteoblastogenic bone marrow stromal cell (BMSC) cultures but not in the calvarial-derived MC3T3-E1 cells. Interestingly, rHMGB1 promoted GSK-3beta phosphorylation in MC3T3-E1 cells but not in BMSCs. Apoptotic bone cells released HMGB1, including MLO-Y4 osteocyte-like cells. MLO-Y4 release of HMGB1 was coincident with caspase-3 cleavage. Furthermore, the anti-apoptotic action of PTH on MC3T3-E1 cells correlated with the observed decrease in HMGB1 release. Our data suggest that apoptotic bone cells release HMGB1, that within the marrow HMGB1 is a bone resorption signal, and that intramembraneous and endochondral osteoblasts exhibit differential responses to this cytokine.

HMGB1 is a bone-active cytokine.

The death of osteocytes, the terminally differentiated cells of the osteoblast lineage that are embedded in bone and regulate remodeling, is significant to both normal and pathological bone resorption. Apoptotic osteocytes putatively release a clarion signal that enhances the development of the bone-resorbing osteoclasts and targets their migration to the breach in the osteocyte network. This phenomenon is thought to underlie normal repair of bone microdamage and contribute to the etiologies of inflammatory bone loss. The chromatin protein high mobility group box 1 protein (HMGB1) has been identified as an "alarmin" in other tissues. An alarmin is an endogenous molecule released by dead and dying cells that alert the innate immune system to damage and the need for tissue repair. Wang and colleagues presented evidence in a landmark 1999 study showing that released HMGB1 is a lethal mediator of sepsis. Extracellular HMGB1 is a ligand for the toll-like receptors (TLRs) and for the receptor for advanced glycation end products (RAGE) all of which amplify inflammation. Recent studies by our lab and others have shown that HMGB1 is a bone-active cytokine. It is released by apoptotic osteoblasts in vitro, including the MLO-Y4 osteocyte-like cells. Extracellular HMGB1 enhances the expression of RANKL, TNFalpha, and IL6 in osteoblastogenic bone marrow stromal cell cultures, and it is chemotactic to osteoclasts. In this prospectus we will review HMGB1 activity at the immune-bone interface and propose a role for HMGB1 as an osteocyte alarmin and mediator of normal remodeling and inflammatory bone loss.

Is HMGB1 an osteocyte alarmin?

Glioblastoma multiforme is a deadly primary brain cancer. Because the tumor kills due to recurrences, we tested the hypothesis that a new treatment would lead to immunological memory in a rat model of recurrent glioblastoma multiforme.

Antiglioma immunological memory in response to conditional cytotoxic/immune-stimulatory gene therapy: humoral and cellular immunity lead to tumor regression.

Soluble antigens diffuse out of the brain and can thus stimulate a systemic immune response, whereas particulate antigens (from infectious agents or tumor cells) remain within brain tissue, thus failing to stimulate a systemic immune response. Immune privilege describes how the immune system responds to particulate antigens localized selectively within the brain parenchyma. We believe this immune privilege is caused by the absence of antigen presenting dendritic cells from the brain. We tested the prediction that expression of fms-like tyrosine kinase ligand 3 (Flt3L) in the brain will recruit dendritic cells and induce a systemic immune response against exogenous influenza hemagglutinin in BALB/c mice. Coexpression of Flt3L with HA in the brain parenchyma induced a robust systemic anti-HA immune response, and a small response against myelin basic protein and proteolipid protein epitopes. Depletion of CD4(+)CD25+ regulatory T cells (Tregs) enhanced both responses. To investigate the autoimmune impact of these immune responses, we characterized the neuropathological and behavioral consequences of intraparenchymal injections of Flt3L and HA in BALB/c and C57BL/6 mice. T cell infiltration in the forebrain was time and strain dependent, and increased in animals treated with Flt3L and depleted of Tregs; however, we failed to detect widespread defects in myelination throughout the forebrain or spinal cord. Results of behavioral tests were all normal. These results demonstrate that Flt3L overcomes the brain's immune privilege, and supports the clinical development of Flt3L as an adjuvant to stimulate clinically effective immune responses against brain neo-antigens, for example, those associated with brain tumors.

Exogenous fms-like tyrosine kinase 3 ligand overrides brain immune privilege and facilitates recognition of a neo-antigen without causing autoimmune neuropathology.

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Lung cancer is the leading cause of cancer death for both men and women worldwide. Since most of the symptoms found for lung cancer are nonspecific, diagnosis is mostly done at late and progressed stage with the consecutive poor therapy outcome. Effective early detection techniques are sorely needed. The emerging field of salivary diagnostics could provide scientifically credible, easy-to-use, non-invasive and cost-effective detection methods. Recent advances have allowed us to develop discriminatory salivary biomarkers for a variety of diseases from oral to systematic diseases. In this study, salivary transcriptomes of lung cancer patients were profiled and led to the discovery and pre-validation of seven highly discriminatory transcriptomic salivary biomarkers (BRAF, CCNI, EGRF, FGF19, FRS2, GREB1, and LZTS1). The logistic regression model combining five of the mRNA biomarkers (CCNI, EGFR, FGF19, FRS2, and GREB1) could differentiate lung cancer patients from normal control subjects, yielding AUC value of 0.925 with 93.75Â % sensitivity and 82.81Â % specificity in the pre-validation sample set. These salivary mRNA biomarkers possess the discriminatory power for the detection of lung cancer. This report provides the proof of concept of salivary biomarkers for the non-invasive detection of the systematic disease. These results poised the salivary biomarkers for the initiation of a multi-center validation in a definitive clinical context.

Development of transcriptomic biomarker signature in human saliva to detect lung cancer.

Cancer du poumon est la principale cause de d&eacute;c&egrave;s par cancer chez les hommes et les femmes dans le monde. Puisque la plupart des sympt&ocirc;mes trouv&eacute;s pour le cancer du poumon sont non sp&eacute;cifique, diagnostic est fait principalement au stade tardif et ont progress&eacute; avec le r&eacute;sultat de la th&eacute;rapie pauvre cons&eacute;cutives. Techniques de d&eacute;tection pr&eacute;coce efficaces font cruellement d&eacute;faut. Le domaine &eacute;mergent du diagnostic salivaire pourrait fournir des m&eacute;thodes de d&eacute;tection scientifiquement cr&eacute;dible, facile &agrave; utiliser, non invasif et rentable. Les progr&egrave;s r&eacute;cents nous ont permis de d&eacute;velopper des biomarqueurs salivaires discriminatoires pour une vari&eacute;t&eacute; de maladies de l&#39;oral aux maladies syst&eacute;matiques. Dans cette &eacute;tude, transcriptomes salivaires de patients atteints de cancer du poumon ont &eacute;t&eacute; profil&eacute;s et conduits &agrave; la d&eacute;couverte et de pr&eacute;-validation de sept transcriptomique hautement discriminatoire de biomarqueurs salivaires (BRAF, CCNI, VUF, FGF19, FRS2, GREB1 et LZTS1). Le mod&egrave;le de r&eacute;gression logistique combinant cinq des biomarqueurs ARNm (CCNI, EGFR, FGF19, FRS2 et GREB1) peut diff&eacute;rencier les patients atteints de cancer de poumon de sujets de contr&ocirc;le normaux, ce qui donne valeur AUC de 0,925 93,75 % 82,81 % de sensibilit&eacute; et de sp&eacute;cificit&eacute; dans l&#39;ensemble de l&#39;&eacute;chantillon de pr&eacute;-validation. Ces biomarqueurs d&#39;ARNm salivaire poss&egrave;dent le pouvoir discriminatoire pour la d&eacute;tection du cancer du poumon. Ce rapport fournit la preuve de concept de biomarqueurs salivaires de d&eacute;tection non invasive de la maladie syst&eacute;matique. Ces r&eacute;sultats s&#39;appr&ecirc;te les biomarqueurs salivaires pour justifier l&#39;ouverture d&#39;une validation multicentrique dans un contexte clinique d&eacute;finitif.

D&eacute;veloppement de transcriptomique signature de biomarqueurs dans la salive humaine pour d&eacute;tecter le cancer du poumon.

Lungenkrebs ist die h&auml;ufigste krebsbedingte Todesursache f&uuml;r M&auml;nner und Frauen weltweit. Da die meisten der gefunden f&uuml;r Lungenkrebs Symptome unspezifisch sind, erfolgt die Diagnose meist bei sp&auml;t und fortgeschrittener Stufe mit dem aufeinanderfolgenden Armen Therapie-Ergebnis. Effektive fr&uuml;he Abfragung Techniken werden dringend gebraucht. Dem aufstrebenden Gebiet der Speicheldr&uuml;sen Diagnose k&ouml;nnte wissenschaftlich glaubw&uuml;rdige, einfach zu bedienende, nicht-invasive und kosteng&uuml;nstige Nachweismethoden bieten. J&uuml;ngste Fortschritte konnten wir diskriminierende Speicheldr&uuml;sen Biomarker f&uuml;r eine Vielzahl von Krankheiten von Oral systematische Krankheiten zu entwickeln. In dieser Studie wurden die Speicheldr&uuml;sen Transcriptomes von Lungenkrebspatienten profiliert und f&uuml;hrte zur Entdeckung und Vorvalidierung von sieben hochgradig diskriminierend Transcriptomic Speicheldr&uuml;sen Biomarkern (BRAF, CCNI, EGRF, FGF19, FRS2, GREB1 und LZTS1). Das logistische Regressionsmodell kombiniert f&uuml;nf von der mRNA-Biomarker (CCNI, EGFR, FGF19, FRS2 und GREB1) konnte Lungenkrebspatienten von normalen Kontrollpersonen, woraus sich AUC-Wert von 0.925 mit 93,75 % Sensitivit&auml;t und 82.81 % Spezifit&auml;t in das pre-validation Sample Set unterscheiden. Diese Speicheldr&uuml;sen mRNA-Biomarkern besitzen die Trennsch&auml;rfe f&uuml;r die Erkennung von Lungenkrebs. Dieser Bericht enth&auml;lt der Proof of Concept Speicheldr&uuml;sen Biomarker f&uuml;r die nicht-invasive Erkennung der systematische Krankheit. Diese Ergebnisse bereit die Speicheldr&uuml;sen Biomarker f&uuml;r die Einleitung eine multizentrische Validierung in einem endg&uuml;ltigen klinischen Kontext.

Entwicklung der Transcriptomic-Biomarker-Signatur im menschlichen Speichel, Lungenkrebs zu erkennen.

Il cancro del polmone &egrave; la principale causa di morte per cancro per gli uomini e le donne in tutto il mondo. Poich&eacute; la maggior parte dei sintomi trovati per cancro al polmone sono aspecifica, la diagnosi &egrave; fatto principalmente in fase tardiva e progredito con l&#39;esito della terapia poveri consecutivi. Efficaci tecniche di individuazione precoce sono assolutamente necessarie. Il campo emergente della diagnostica salivare potrebbe fornire metodi di rilevamento scientificamente credibile, facile da usare, non invasivo e conveniente. I recenti progressi ci hanno permesso di sviluppare discriminatorio salivari biomarkers per una variet&agrave; di malattie da orale a malattie sistematiche. In questo studio, transcriptomes salivari di pazienti con cancro del polmone sono state profilate e port&ograve; alla scoperta e pre-validazione di sette transcrittomica altamente discriminatorio salivari biomarcatori (BRAF, CCNI, EGRF, FGF19, FRS2, GREB1 e LZTS1). Il modello di regressione logistica che unisce cinque i biomarcatori di mRNA (CCNI, EGFR, FGF19, FRS2 e GREB1) potrebbe differenziare pazienti con cancro del polmone da soggetti di controllo normali, producendo valore AUC di 0,925 con 93,75% specificit&agrave;, sensibilit&agrave; e 82.81% nel gruppo campione pre-convalida. Questi biomarcatori salivari mRNA possiedono il potere discriminatorio per la rilevazione del cancro del polmone. Questo report fornisce la prova del concetto di biomarcatori salivare per la rilevazione non invasiva della malattia sistematica. Questi risultati in bilico i salivare biomarcatori per l&#39;avvio di una validazione multicentrica in un contesto clinico definitivo.

Sviluppo della firma biomarker transcrittomica nella saliva umana per rilevare il cancro ai polmoni.

&#x80BA;&#x304C;&#x3093;&#x306F;&#x3001;&#x7537;&#x6027;&#x3068;&#x5973;&#x6027;&#x306F;&#x4E16;&#x754C;&#x4E2D;&#x306E;&#x764C;&#x6B7B;&#x306E;&#x4E3B;&#x8981;&#x306A;&#x539F;&#x56E0;&#x3067;&#x3059;&#x3002;&#x767A;&#x898B;&#x80BA;&#x304C;&#x3093;&#x306E;&#x75C7;&#x72B6;&#x306E;&#x307B;&#x3068;&#x3093;&#x3069;&#x306F;&#x975E;&#x7279;&#x7570;&#x7684;&#x3067;&#x3042;&#x308B;&#x306E;&#x3067;&#x3001;&#x8A3A;&#x65AD;&#x306F;&#x4E3B;&#x9023;&#x7D9A;&#x8CA7;&#x3057;&#x3044;&#x6CBB;&#x7642;&#x30A2;&#x30A6;&#x30C8;&#x30AB;&#x30E0;&#x3068;&#x5F8C;&#x534A;&#x3068;&#x9032;&#x884C;&#x3057;&#x305F;&#x6BB5;&#x968E;&#x3067;&#x884C;&#x308F;&#x308C;&#x307E;&#x3059;&#x3002;&#x52B9;&#x679C;&#x7684;&#x306A;&#x65E9;&#x671F;&#x691C;&#x51FA;&#x6280;&#x8853;&#x306F;&#x3001;&#x75DB;&#x3093;&#x3067;&#x5FC5;&#x8981;&#x3068;&#x3055;&#x308C;&#x307E;&#x3059;&#x3002;&#x553E;&#x6DB2;&#x8A3A;&#x65AD;&#x306E;&#x65B0;&#x8208;&#x5206;&#x91CE;&#x306E;&#x79D1;&#x5B66;&#x7684;&#x306B;&#x4FE1;&#x3058;&#x3001;&#x7C21;&#x5358;&#x306B;&#x4F7F;&#x7528;&#x3067;&#x304D;&#x308B;&#x3001;&#x975E;&#x4FB5;&#x8972;&#x7684;&#x306A;&#x8CBB;&#x7528;&#x5BFE;&#x52B9;&#x679C;&#x306E;&#x691C;&#x51FA;&#x65B9;&#x6CD5;&#x3092;&#x63D0;&#x4F9B;&#x3067;&#x304D;&#x307E;&#x3059;&#x3002;&#x6700;&#x8FD1;&#x306E;&#x9032;&#x6B69;&#x306F;&#x53E3;&#x8154;&#x304B;&#x3089;&#x75C5;&#x6C17;&#x306B;&#x4F53;&#x7CFB;&#x7684;&#x306A;&#x75C5;&#x6C17;&#x306E;&#x69D8;&#x3005; &#x306A;&#x5DEE;&#x5225;&#x306E;&#x553E;&#x6DB2;&#x30D0;&#x30A4;&#x30AA; &#x30DE;&#x30FC;&#x30AB;&#x30FC;&#x306E;&#x958B;&#x767A;&#x3092;&#x8A31;&#x53EF;&#x3057;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x672C;&#x7814;&#x7A76;&#x3067;&#x306F;, &#x80BA;&#x764C;&#x60A3;&#x8005;&#x306E;&#x553E;&#x6DB2;&#x30C8;&#x30E9;&#x30F3;&#x30B9;&#x30AF;&#x30EA;&#x30D7;&#x30C8;&#x30FC;&#x30E0; &#x30D7;&#x30ED;&#x30D5;&#x30A1;&#x30A4;&#x30EA;&#x30F3;&#x30B0;&#x3057;&#x3001;&#x767A;&#x898B;&#x3068; 7 &#x306E;&#x9AD8;&#x5EA6;&#x306B;&#x5DEE;&#x5225;&#x7684;&#x30C8;&#x30E9;&#x30F3;&#x30B9;&#x30AF;&#x30EA;&#x30D7;&#x30C8;&#x30FC;&#x30E0;&#x553E;&#x6DB2;&#x306E;&#x30DE;&#x30FC;&#x30AB;&#x30FC; (BRAF&#x3001;&#x3068;&#x306F;&#x3044;&#x3048;&#x3001;EGRF&#x3001;FGF19&#x3001;FRS2&#x3001;GREB1&#x3001;&#x304A;&#x3088;&#x3073; LZTS1) &#x306E;&#x4E8B;&#x524D;&#x691C;&#x8A3C;&#x306B;&#x3064;&#x306A;&#x304C;&#x3063;&#x305F;&#x3002;MRNA &#x306E;&#x30D0;&#x30A4;&#x30AA; &#x30DE;&#x30FC;&#x30AB;&#x30FC; (&#x3068;&#x306F;&#x3044;&#x3048;&#x3001;EGFR&#x3001;FGF19&#x3001;FRS2&#x3001;&#x304A;&#x3088;&#x3073; GREB1) &#x306E; 5 &#x3064;&#x3092;&#x7D44;&#x307F;&#x5408;&#x308F;&#x305B;&#x3066;&#x3001;&#x30ED;&#x30B8;&#x30B9;&#x30C6;&#x30A3;&#x30C3;&#x30AF;&#x56DE;&#x5E30;&#x30E2;&#x30C7;&#x30EB;&#x306F;&#x6B63;&#x5E38;&#x5BFE;&#x7167;&#x7FA4;, AUC &#x5024; 0.925 &#x306E; 93.75 &#xFF05; &#x306E;&#x611F;&#x5EA6;&#x3068; 82.81 &#xFF05; &#x306E;&#x7279;&#x7570;&#x6027;&#x3001;&#x4E8B;&#x524D;&#x691C;&#x8A3C;&#x30B5;&#x30F3;&#x30D7;&#x30EB;&#x3067;&#x964D;&#x4F0F;&#x304B;&#x3089;&#x80BA;&#x764C;&#x60A3;&#x8005;&#x3092;&#x533A;&#x5225;&#x3067;&#x304D;&#x307E;&#x3059;&#x3002;&#x3053;&#x308C;&#x3089;&#x553E;&#x6DB2;&#x4E2D;&#x306E; mRNA &#x306E;&#x30D0;&#x30A4;&#x30AA; &#x30DE;&#x30FC;&#x30AB;&#x30FC;&#x80BA;&#x764C;&#x306E;&#x691C;&#x51FA;&#x306E;&#x5DEE;&#x5225;&#x7684;&#x306A;&#x96FB;&#x6E90;&#x3092;&#x6301;&#x3063;&#x3066;&#x3044;&#x307E;&#x3059;&#x3002;&#x3053;&#x306E;&#x30EC;&#x30DD;&#x30FC;&#x30C8;&#x553E;&#x6DB2;&#x30D0;&#x30A4;&#x30AA; &#x30DE;&#x30FC;&#x30AB;&#x30FC;&#x306E;&#x30B3;&#x30F3;&#x30BB;&#x30D7;&#x30C8;&#x306E;&#x8A3C;&#x660E;&#x4F53;&#x7CFB;&#x7684;&#x75BE;&#x60A3;&#x306E;&#x975E;&#x4FB5;&#x8972;&#x7684;&#x691C;&#x51FA;&#x3092;&#x63D0;&#x4F9B;&#x3057;&#x307E;&#x3059;&#x3002;&#x3053;&#x308C;&#x3089;&#x306E;&#x7D50;&#x679C;&#x304B;&#x3089;&#x553E;&#x6DB2;&#x30D0;&#x30A4;&#x30AA; &#x30DE;&#x30FC;&#x30AB;&#x30FC;&#x306B;&#x3088;&#x308B;&#x591A;&#x65BD;&#x8A2D;&#x691C;&#x8A3C;&#x306E;&#x958B;&#x59CB;&#x306E;&#x6C7A;&#x5B9A;&#x7684;&#x306A;&#x81E8;&#x5E8A;&#x30B3;&#x30F3;&#x30C6;&#x30AD;&#x30B9;&#x30C8;&#x3067;&#x69CB;&#x3048;&#x3002;

&#x30C8;&#x30E9;&#x30F3;&#x30B9;&#x30AF;&#x30EA;&#x30D7;&#x30C8;&#x30FC;&#x30E0; &#x30D0;&#x30A4;&#x30AA; &#x30DE;&#x30FC;&#x30AB;&#x30FC;&#x7F72;&#x540D;&#x80BA;&#x304C;&#x3093;&#x3092;&#x691C;&#x51FA;&#x3059;&#x308B;&#x305F;&#x3081;&#x306B;&#x553E;&#x6DB2;&#x4E2D;&#x306E;&#x958B;&#x767A;&#x3002;

&#xD3D0;&#xC554;&#xC740; &#xB0A8;&#xC131;&#xACFC; &#xC5EC;&#xC131; &#xC804;&#xC138;&#xACC4; &#xC554; &#xC0AC;&#xB9DD;&#xC758; &#xC8FC;&#xC694; &#xC6D0;&#xC778;. &#xB54C;&#xBB38;&#xC5D0; &#xD3D0;&#xC554;&#xC744; &#xBC1C;&#xACAC; &#xD558;&#xB294; &#xC99D;&#xC0C1;&#xC758; &#xB300;&#xBD80;&#xBD84;&#xC740; &#xD2B9;&#xC774; &#xD604;&#xC0C1;&#xC774;, &#xC9C4;&#xB2E8; &#xC5F0;&#xC18D; &#xBD88; &#xC30D; &#xD55C; &#xCE58;&#xB8CC; &#xACB0;&#xACFC;&#xB97C; &#xB2A6;&#xAC8C; &#xD558; &#xACE0; &#xC9C4;&#xD589; &#xB2E8;&#xACC4;&#xC5D0;&#xC11C; &#xC8FC;&#xB85C; &#xC774;&#xB8E8;&#xC5B4;&#xC9D1;&#xB2C8;&#xB2E4;. &#xD6A8;&#xACFC;&#xC801;&#xC778; &#xC870;&#xAE30; &#xD0D0;&#xC9C0; &#xAE30;&#xC220;&#xC740; &#xC815;&#xB9D0;&#xB85C; &#xD544;&#xC694; &#xD569;&#xB2C8;&#xB2E4;. &#xCE68; &#xC9C4;&#xB2E8;&#xC758; &#xC2E0;&#xD765; &#xBD84;&#xC57C; &#xACFC;&#xD559;&#xC801;&#xC73C;&#xB85C; &#xC2E0;&#xC6A9;&#xD560; &#xC218; &#xC788;&#xB294;, &#xC0AC;&#xC6A9;&#xC774; &#xC26C;&#xC6B4;, &#xBE44;-&#xCE68;&#xB7B5; &#xC801; &#xBC0F; &#xBE44;&#xC6A9; &#xD6A8;&#xC728;&#xC801;&#xC778; &#xAC80;&#xC0C9; &#xBC29;&#xBC95;&#xC744; &#xC81C;&#xACF5;&#xD560; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;. &#xCD5C;&#xADFC;&#xC758; &#xC9C4;&#xBCF4;&#xB294; &#xB2E4;&#xC591; &#xD55C; &#xCCB4;&#xACC4;&#xC801;&#xC778; &#xC9C8;&#xBCD1;&#xC5D0; &#xAD6C;&#xAC15;&#xC5D0;&#xC11C; &#xC9C8;&#xBCD1;&#xC5D0; &#xB300; &#xD55C; &#xCC28;&#xBCC4;&#xC801;&#xC778; &#xCE68; &#xBC14;&#xC774;&#xC624; &#xB9C8;&#xCEE4;&#xC758; &#xAC1C;&#xBC1C;&#xC744; &#xD5C8;&#xC6A9; &#xD588;&#xB2E4;. &#xC774; &#xC5F0;&#xAD6C;&#xC5D0;&#xC11C;&#xB294; &#xD3D0; &#xC554; &#xD658;&#xC790;&#xC758; &#xCE68; transcriptomes &#xD504;&#xB85C; &#xD30C;&#xC77C;&#xB9C1; &#xD558; &#xACE0; &#xBC1C;&#xACAC;&#xACFC; 7 &#xB9E4;&#xC6B0; &#xCC28;&#xBCC4;&#xC801;&#xC778; transcriptomic &#xCE68; &#xC0DD;&#xCCB4; (BRAF, CCNI, EGRF, FGF19, FRS2, GREB1, &#xBC0F; LZTS1)&#xC758; &#xC608;&#xC57D; &#xD655;&#xC778;&#xC744; &#xD588;&#xB2E4;. 5 MRNA &#xC0DD;&#xCCB4; (CCNI, EGFR, FGF19, FRS2, &#xBC0F; GREB1)&#xC744; &#xACB0;&#xD569; &#xD558;&#xB294; &#xB85C;&#xC9C0;&#xC2A4;&#xD2F1; &#xD68C;&#xADC0; &#xBAA8;&#xB378; &#xC0AC;&#xC804; &#xC0D8;&#xD50C; &#xC138;&#xD2B8;&#xC5D0;&#xC11C; 93.75% &#xAC10;&#xB3C4; &#xBC0F; 82.81% &#xD2B9;&#xC774;&#xC131; 0.925&#xC758; AUC &#xAC12; &#xC0DD;&#xC131; &#xD558;&#xB294; &#xC77C;&#xBC18;&#xC801;&#xC778; &#xC81C;&#xC5B4; &#xACFC;&#xBAA9;&#xC5D0;&#xC11C; &#xD3D0; &#xC554; &#xD658;&#xC790;&#xB97C; &#xCC28;&#xBCC4;&#xD654; &#xC218; &#xC788;&#xC2B5;&#xB2C8;&#xB2E4;. &#xC774;&#xB7EC;&#xD55C; &#xCE68; mRNA &#xC0DD;&#xCCB4; &#xD3D0;&#xC554;&#xC758; &#xAC80;&#xCD9C;&#xC5D0; &#xB300; &#xD55C; &#xCC28;&#xBCC4;&#xC801;&#xC778; &#xC804;&#xB825; &#xBCF4;&#xC720;. &#xC774; &#xBCF4;&#xACE0;&#xC11C;&#xB294; &#xCCB4;&#xACC4;&#xC801;&#xC778; &#xC9C8;&#xBCD1;&#xC758; &#xBE44;-&#xCE68;&#xB7B5; &#xC801; &#xAC80;&#xCD9C;&#xC744; &#xC704;&#xD55C; &#xCE68; biomarkers&#xC758; &#xAC1C;&#xB150; &#xC99D;&#xBA85;&#xC744; &#xC81C;&#xACF5;&#xD569;&#xB2C8;&#xB2E4;. &#xC774;&#xB7EC;&#xD55C; &#xACB0;&#xACFC; &#xD655;&#xC2E4; &#xD55C; &#xC784;&#xC0C1; &#xCEE8;&#xD14D;&#xC2A4;&#xD2B8;&#xC5D0;&#xC11C; &#xBA40;&#xD2F0; &#xC13C;&#xD130; &#xC720;&#xD6A8;&#xC131; &#xAC80;&#xC0AC;&#xC758; &#xAC1C;&#xC2DC;&#xC5D0; &#xB300; &#xD55C; &#xCE68; biomarkers &#xD0DC;&#xC138; &#xC774;&#xB2E4;.

&#xD3D0;&#xC554;&#xC744; &#xAC10;&#xC9C0; &#xD558;&#xB294; &#xC778;&#xAC04;&#xC758; &#xD0C0; &#xC561; transcriptomic biomarker &#xC11C;&#xBA85;&#xC758; &#xAC1C;&#xBC1C;.

C&aacute;ncer de pulm&oacute;n es la causa principal de muerte por c&aacute;ncer para hombres y mujeres en todo el mundo. Puesto que la mayor&iacute;a de los s&iacute;ntomas de c&aacute;ncer de pulm&oacute;n son inespec&iacute;fica, diagn&oacute;stico se realiza principalmente en etapa tard&iacute;a y progresada con el resultado de la terapia pobre consecutivos. T&eacute;cnicas efectivas de detecci&oacute;n temprana se necesitan urgentemente. El campo emergente de diagn&oacute;stico salival podr&iacute;a proporcionar m&eacute;todos de detecci&oacute;n cient&iacute;ficamente cre&iacute;ble, f&aacute;cil de usar, no invasiva y rentable. Los avances recientes han permitido desarrollar biomarcadores salivales discriminatorias para una variedad de enfermedades de oral a enfermedades sistem&aacute;ticas. En este estudio, transcriptomas salivales de pacientes con c&aacute;ncer de pulm&oacute;n fueron perfilados y condujo al descubrimiento y prevalidaci&oacute;n de siete biomarcadores salival transcript&oacute;mico altamente discriminatorias (BRAF, CCNI, EGRF, FGF19, FRS2, GREB1 y LZTS1). El modelo de regresi&oacute;n log&iacute;stica combinando cinco de los biomarcadores del mRNA (CCNI, EGFR, FGF19, FRS2 y GREB1) podr&iacute;a diferenciar a pacientes con c&aacute;ncer de pulm&oacute;n de temas normales del control, dando valor AUC de 0.925 con 93.75% especificidad, sensibilidad y 82.81% en el conjunto de la muestra pre-validation. Estos biomarcadores de mRNA salival poseen el poder discriminatorio para la detecci&oacute;n del c&aacute;ncer de pulm&oacute;n. Este informe proporciona la prueba de concepto de salivales biomarcadores para la detecci&oacute;n no invasiva de la enfermedad sistem&aacute;tica. Estos resultados hab&iacute;an preparada los biomarcadores salivales para la iniciaci&oacute;n de una validaci&oacute;n multic&eacute;ntrico en un contexto cl&iacute;nico definitivo.

Desarrollo de la firma de biomarcadores transcript&oacute;mico en saliva humana para detectar c&aacute;ncer de pulm&oacute;n.

To assess the diagnostic value of high-resolution ultrasonography for detecting of anterior disc displacement without reduction (ADDWR) of temporomandibular joint (TMJ).

[The diagnostic value of high-resolution ultrasonography for detecting anterior disc displacement without reduction of temporomandibular joint].

Exosomes biomarkers mediating important biological process, especially in the systemic disease diagnostics and therapeutics, yet the protective exosomal vesicle structure hinders rapid, simple detection of the harbored molecules. We have established a new method, the electric field-induced release and measurement (EFIRM), which can simultaneously disrupt exosomes to release the contents and on-site monitoring the harbored exosomal RNA/proteins biomarkers. When exposed to a non-uniform electrical field, exosomal RNA and proteins are rapidly released. Bio-recognition of these biomolecules is carried out concurrently. We tested the hypothesis that the lung cancer cell line, H460 stably transfected with hCD63-GFP, would shed hCD63-GFP expressing exosomes that could be detected in serum and saliva. We confirmed in vivo that H460-CD63-GFP shed exosomes were transported to blood and saliva. This result demonstrates for the first time tumor-shed exosomes were detected in saliva, in addition to blood, presenting a new translational utility of exosome-based biomarker detection in saliva.

Detection of exosomal biomarker by electric field-induced release and measurement (EFIRM).

To evaluate the anesthetic effects and safety of Gow-Gates technique of inferior alveolar nerve block in impacted mandibular third molar extraction.

[The anesthetic effects of Gow-Gates technique of inferior alveolar nerve block in impacted mandibular third molar extraction].

Green tea (GT) and caffeine in combination were shown to increase energy expenditure and fat oxidation, but less is known about the effects of black tea (BT) and oolong tea (OT). This study investigated whether decaffeinated polyphenol extracts from GT, BT, and OT decrease body fat and inflammation in male C57BL/6J mice fed high-fat/high-sucrose [HF/HS (32% energy from fat, 25% energy from sucrose)] diets. Mice were fed either an HF/HS diet with 0.25% of polyphenol from GT, OT, or BT or a low-fat/high-sucrose [LF/HS (10.6% energy from fat, 25% energy from sucrose)] diet for 20 wk. Monomeric tea polyphenols were found in the liver and adipose tissue of mice fed the HF/HS diet with GT polyphenols (GTPs) and OT polyphenols (OTPs) but not BT polyphenols (BTPs). Treatment with GTPs, OTPs, BTPs, and an LF/HS diet led to significantly lower body weight, total visceral fat volume by MRI, and liver lipid weight compared with mice in the HF/HS control group. Only GTPs reduced food intake significantly by ∼10%. GTP, BTP, and LF/HS-diet treatments significantly reduced serum monocyte chemotactic protein-1 (MCP-1) compared with HF/HS controls. In mesenteric fat, monocyte chemotactic protein-1 (Mcp1) gene expression was significantly decreased by treatment with GTPs, BTPs, OTPs, and an LF/HS diet and in liver tissue by GTP and BTP treatments. Mcp1 gene expression in epididymal fat was significantly decreased by the BTP and LF/HS diet interventions. In epididymal fat, consistent with an anti-inflammatory effect, adiponectin gene expression was significantly increased by GTPs and OTPs. Angiogenesis during adipose tissue expansion is anti-inflammatory by maintaining adipocyte perfusion. We observed significantly increased gene expression of vascular endothelial growth factor A by GTPs and vascular endothelial growth factor receptor 2 by BTPs and the LF/HS diet and a decrease in pigment epithelium-derived factor gene expression by OTPs and BTPs. In summary, all 3 tea polyphenol extracts induced weight loss and anti-inflammatory and angiogenic effects, although the tissue content of polyphenols differed significantly.

Green tea, black tea, and oolong tea polyphenols reduce visceral fat and inflammation in mice fed high-fat, high-sucrose obesogenic diets.

The discovery of disease-specific biomarkers in oral fluids has revealed a new dimension in molecular diagnostics. Recent studies have reported the mechanistic involvement of tumor cells derived mediators, such as exosomes, in the development of saliva-based mRNA biomarkers. To further our understanding of the origins of disease-induced salivary biomarkers, we here evaluated the hypothesis that tumor-shed secretory lipidic vesicles called exosome-like microvesicles (ELMs) that serve as protective carriers of tissue-specific information, mRNAs, and proteins, throughout the vasculature and bodily fluids. RNA content was analyzed in cell free-saliva and ELM-enriched fractions of saliva. Our data confirmed that the majority of extracellular RNAs (exRNAs) in saliva were encapsulated within ELMs. Nude mice implanted with human lung cancer H460 cells expressing hCD63-GFP were used to follow the circulation of tumor cell specific protein and mRNA in the form of ELMs in vivo. We were able to identify human GAPDH mRNA in ELMs of blood and saliva of tumor bearing mice using nested RT-qPCR. ELMs positive for hCD63-GFP were detected in the saliva and blood of tumor bearing mice as well as using electric field-induced release and measurement (EFIRM). Altogether, our results demonstrate that ELMs carry tumor cell-specific mRNA and protein from blood to saliva in a xenografted mouse model of human lung cancer. These results therefore strengthen the link between distal tumor progression and the biomarker discovery of saliva through the ELMs.

Jieping Yang

School of Medicine,
Clinical Nutrition,
School of Medicine, Clinical Nutrition

University of California, Los Angeles

Detection of Exosomal Biomarker by Electric Field-induced Release and Measurement (EFIRM)

Jieping Yang

School of Medicine,Clinical Nutrition,School of Medicine, Clinical Nutrition

University of California, Los Angeles

Detection of Exosomal Biomarker by Electric Field-induced Release and Measurement (EFIRM)

School of Medicine,
Clinical Nutrition,
School of Medicine, Clinical Nutrition