この研究はNIH（R21 MH090384、R01 EB006494、RO1 EB009666、R01 NS051622）によって運営されている。我々は彼らの技術支援のためにH. SarofinとC. Lacadieに感謝。

<ol>
	<li>Mataix-Cols, D., Cullen, S., Lange, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neural+correlates+of+anxiety+associated+with+obsessive-compulsive+symptom+dimensions+in+normal+volunteers.">Neural correlates of anxiety associated with obsessive-compulsive symptom dimensions in normal volunteers.</a> Biol. Psychiatry. 53, 482-493 (2003).</li><li>Mataix-Cols, D., Wooderson, S., Lawrence, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Distinct+neural+correlates+of+washing,+checking,+and+hoarding+symptom+dimensions+in+obsessive-compulsive+disorder.">Distinct neural correlates of washing, checking, and hoarding symptom dimensions in obsessive-compulsive disorder.</a> Arch. Gen. Psychiatry. 61, 564-576 (2004).</li><li>Menzies, L., Chamberlain, S. R., Laird, A. 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M., Fairneny, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increasing+cortical+activity+in+auditory+areas+through+neurofeedback+functional+magnetic+resonance+imaging.">Increasing cortical activity in auditory areas through neurofeedback functional magnetic resonance imaging.</a> Neuroreport. 17, 1273-1278 (2006).</li><li>Caria, A., Sitaram, R., Veit, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Volitional+control+of+anterior+insula+activity+modulates+the+response+to+aversive+stimuli.+A+real-time+functional+magnetic+resonance+imaging+study.">Volitional control of anterior insula activity modulates the response to aversive stimuli. A real-time functional magnetic resonance imaging study.</a> Biological psychiatry. 68, 425-432 (2010).</li><li>Caria, A., Veit, R., Sitaram, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regulation+of+anterior+insular+cortex+activity+using+real-time+fMRI.">Regulation of anterior insular cortex activity using real-time fMRI.</a> Neuroimage. 35, 1238-1246 (2007).</li><li>Hamilton, J. P., Glover, G. H., Hsu, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modulation+of+subgenual+anterior+cingulate+cortex+activity+with+real-time+neurofeedback.">Modulation of subgenual anterior cingulate cortex activity with real-time neurofeedback.</a> Hum. Brain. Mapp. 32, 22-31 (2011).</li><li>Johnston, S., Linden, D. E., Healy, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Upregulation+of+emotion+areas+through+neurofeedback+with+a+focus+on+positive+mood.">Upregulation of emotion areas through neurofeedback with a focus on positive mood.</a> Cognitive, affective &#38; behavioral neuroscience. 11, 44-51 (2011).</li><li>Johnston, S. J., Boehm, S. G., Healy, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurofeedback:+A+promising+tool+for+the+self-regulation+of+emotion+networks.">Neurofeedback: A promising tool for the self-regulation of emotion networks.</a> NeuroImage. 49, 1066-1072 (2010).</li><li>Zotev, V., Krueger, F., Phillips, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Self-regulation+of+amygdala+activation+using+real-time+fMRI+neurofeedback.">Self-regulation of amygdala activation using real-time fMRI neurofeedback.</a> PLoS One. 6, e24522-e24522 (2011).</li><li>Haller, S., Birbaumer, N., Veit, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Real-time+fMRI+feedback+training+may+improve+chronic+tinnitus.">Real-time fMRI feedback training may improve chronic tinnitus.</a> Eur. Radiol. 20, 696-703 (2010).</li><li>Bloch, M. H., Landeros-Weisenberger, A., Kelmendi, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+systematic+review:+antipsychotic+augmentation+with+treatment+refractory+obsessive-compulsive+disorder.">A systematic review: antipsychotic augmentation with treatment refractory obsessive-compulsive disorder.</a> Mol. Psychiatry. 11, 622-632 (2006).</li><li>Jenike, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+practice.+Obsessive-compulsive+disorder.">Clinical practice. Obsessive-compulsive disorder.</a> N. Engl. J. Med. 350, 259-265 (2004).</li><li>Pallanti, S., Quercioli, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Treatment-refractory+obsessive-compulsive+disorder:+methodological+issues,+operational+definitions+and+therapeutic+lines.">Treatment-refractory obsessive-compulsive disorder: methodological issues, operational definitions and therapeutic lines.</a> Prog. Neuropsychopharmacol. Biol. Psychiatry. 30, 400-412 (2006).</li><li>Mataix-Cols, D., Lawrence, N. S., Wooderson, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Maudsley+Obsessive-Compulsive+Stimuli+Set:+validation+of+a+standardized+paradigm+for+symptom-specific+provocation+in+obsessive-compulsive+disorder.">The Maudsley Obsessive-Compulsive Stimuli Set: validation of a standardized paradigm for symptom-specific provocation in obsessive-compulsive disorder.</a> Psychiatry. Res. 168, 238-241 (2009).</li><li>Lang, P. J., Bradley, M. M., Cuthbert, B. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=International+affective+picture+system+(IAPS):+Affective+ratings+of+pictures+and+instruction+manual.">International affective picture system (IAPS): Affective ratings of pictures and instruction manual.</a> Technical Report A-82008. , (2008).</li><li>Burns, G. L., Keortge, S. G., Formea, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Revision+of+the+Padua+Inventory+of+obsessive+compulsive+disorder+symptoms:+distinctions+between+worry,+obsessions,+and+compulsions.">Revision of the Padua Inventory of obsessive compulsive disorder symptoms: distinctions between worry, obsessions, and compulsions.</a> Behaviour research and therapy. 34, 163-173 (1996).</li><li>Scheinost, D., Hampson, M., Bhawnani, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+GPU+accelerated+motion+correction+algorithm+for+real-time+fMRI.">A GPU accelerated motion correction algorithm for real-time fMRI.</a> Human Brain Mapping. , 639 (2011).</li><li>Hampson, M., Scheinost, D., Qiu, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biofeedback+from+the+supplementary+motor+area+reduces+functional+connectivity+to+subcortical+regions.">Biofeedback from the supplementary motor area reduces functional connectivity to subcortical regions.</a> Brain Connectivity. 1, 91-98 (2011).</li><li>Johnson, K. A., Hartwell, K., Lematty, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intermittent+"Real-time"+fMRI+Feedback+Is+Superior+to+Continuous+Presentation+for+a+Motor+Imagery+Task:+A+Pilot+Study.">Intermittent "Real-time" fMRI Feedback Is Superior to Continuous Presentation for a Motor Imagery Task: A Pilot Study.</a> J. Neuroimaging. , (2011).</li><li>Yoo, S. S., Lee, J. H., O'Leary, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+magnetic+resonance+imaging-mediated+learning+of+increased+activity+in+auditory+areas.">Functional magnetic resonance imaging-mediated learning of increased activity in auditory areas.</a> Neuroreport. 18, 1915-1920 (2007).</li></ol>

利害の衝突は宣言されません。

リアルタイムfMRIのデータのバイオフィードバックは、新しい技術であり、より多くの仕事は、被験者の学習を最大化するようにこの方法を最適化するために必要とされる。最近の研究では、実行またはスキャンセッション、14、18、27の異なる番号を持つ方法を学ぶの変更を検討されているフィードバックのパラダイムは、28の学習に影響を及ぼし、どのようにかどうか終了後...

real-time fmri biofeedback targeting the orbitofrontal cortex for contamination anxiety

我々は、リアルタイム機能的磁気共鳴画像法（RT - fMRI）をデータのバイオフィードバックを用いて汚染の不安に関連付けられている彼らの眼窩前頭皮質の領域で活性を制御するための訓練科目のための手法を提案する。この地域の活性の増加は、対照群1と強迫性障害（OCD）、2汚染の不安を伴う比較的一般的で衰弱精神疾患を持つ個人の両方の汚染の不安との関係に見られる。多くの脳領域がOCDに関与しているが、眼窩前頭皮質の異常（OFC）が最も一貫性のある結果の一つである。3、さらに4、OFCの多動は、OCDの症状の重大度5と相関することが発見さで多動で減少しているこの地域は減少した症状の重症度と相関することが報告されている6したがって、この脳の領域を制御する能力は、CLに変換する場合があります汚染の不安を含む強迫症状でinical改善。 RT - fMRIデータのバイオフィードバックは、被験者の脳内の特定の地域における活動の時間パターン（または脳活動の特定の分散パターンに関連付けられている）被験者へのフィードバック信号として提供されている新技術です。最近の報告では、RT - fMRIのバイオフィードバックを提供するとき人々は特定の脳領域の活動の制御を開発することができることを示す。特に、7-12、感情処理に関わる脳の領域を対象にこの手法を用いていくつかの研究では、訓練科目の成功を報告しているこれらの領域を制御する。いくつかのケースでは13-18に、RT - fMRIのバイオフィードバック訓練が対象で、認知的、感情的、または臨床的変化を誘導することが報告されている。8、9、13、19はここでの治療に適用されるように我々はこの手法を説明健常者における汚染の不安。このバイオフィードバックの介入は、貴重な浅なります。IC研究ツール：それは、摂動の脳機能への研究者は、脳のダイナミクスの結果としての変化を測定し、汚染の不安やその他の行動措置の変化にそれらを関連付けることができます。さらに、この方法の確立は、OCDの治療的介入としてのfMRIをベースバイオフィードバックの検討の第一歩としての役割を果たします。約OCDの患者の4分の1が治療、20-22の現在利用可能なフォームから、給付を行なう人はほとんど完全に回復しない、この集団を治療するための新たなアプローチが緊急に必要とされることはほとんど恩恵を受けることを考える。

Watch this Scientific Journal Video about Real-time fMRI Biofeedback Targeting the Orbitofrontal Cortex for Contamination Anxiety at JoVE.com

Real-time fMRI Biofeedback Targeting the Orbitofrontal Cortex for Contamination Anxiety

ここでは、汚染の不安に関与する脳の領域を制御するための訓練の人々のためと汚染の不安と脳の接続性パターンとの関係をプロービングするための手法を提案する。

汚染の不安のために眼窩前頭皮質をターゲットに、リアルタイムfMRIのバイオフィードバック

We present a method for training subjects to control activity in a region of their orbitofrontal cortex associated with contamination anxiety using ...

real-time-fmri-biofeedback-targeting-orbitofrontal-cortex-for

Research

JoVE Journal

Medicine

20.7K ビュー.  Yale University School of Medicine. ここでは、汚染の不安に関与する脳の領域を制御するための訓練の人々のためと汚染の不安と脳の接続性パターンとの関係をプロービングするための手法を提案する。

ビデオ: Real-time fMRI Biofeedback Targeting the Orbitofrontal Cortex for Contamination Anxiety

Multispectral Real-time Fluorescence Imaging for Intraoperative Detection of the Sentinel Lymph Node in Gynecologic Oncology

The prognosis in virtually all solid tumors depends on the presence or absence of lymph node metastases.1-3 Surgical treatment most often combines radical excision of the tumor with a full lymphadenectomy in the drainage area of the tumor. However, removal of lymph nodes is associated with increased morbidity due to infection, wound breakdown and lymphedema.4,5 As an alternative, the sentinel lymph node procedure (SLN) was developed several decades ago to detect the first draining lymph node from the tumor.6 In case of lymphogenic dissemination, the SLN is the first lymph node that is affected (Figure 1). Hence, if the SLN does not contain metastases, downstream lymph nodes will also be free from tumor metastases and need not to be removed. The SLN procedure is part of the treatment for many tumor types, like breast cancer and melanoma, but also for cancer of the vulva and cervix.7 The current standard methodology for SLN-detection is by peritumoral injection of radiocolloid one day prior to surgery, and a colored dye intraoperatively. Disadvantages of the procedure in cervical and vulvar cancer are multiple injections in the genital area, leading to increased psychological distress for the patient, and the use of radioactive colloid.
Multispectral fluorescence imaging is an emerging imaging modality that can be applied intraoperatively without the need for injection of radiocolloid. For intraoperative fluorescence imaging, two components are needed: a fluorescent agent and a quantitative optical system for intraoperative imaging. As a fluorophore we have used indocyanine green (ICG). ICG has been used for many decades to assess cardiac function, cerebral perfusion and liver perfusion.8 It is an inert drug with a safe pharmaco-biological profile. When excited at around 750 nm, it emits light in the near-infrared spectrum around 800 nm. A custom-made multispectral fluorescence imaging camera system was used.9.
The aim of this video article is to demonstrate the detection of the SLN using intraoperative fluorescence imaging in patients with cervical and vulvar cancer. Fluorescence imaging is used in conjunction with the standard procedure, consisting of radiocolloid and a blue dye. In the future, intraoperative fluorescence imaging might replace the current method and is also easily transferable to other indications like breast cancer and melanoma.

Fluorescence imaging is a promising innovative modality for image-guided surgery in surgical oncology. In this video we describe the technical procedure for detection of the sentinel lymph node using fluorescence imaging as showcased in gynecologic oncologicy. A multispectral fluorescence camera system, together with the fluorescent agent indocyanine green, is applied.

婦人科腫瘍におけるセンチネルリンパ節の術中検出のためのマルチスペクトルのリアルタイム蛍光イメージング

The prognosis in virtually all solid tumors depends on the presence or absence of lymph node metastases.1-3 Surgical treatment most often combines radical ...

医学

Remote Magnetic Navigation for Accurate, Real-time Catheter Positioning and Ablation in Cardiac Electrophysiology Procedures

New remote navigation systems have been developed to improve current limitations of conventional manually guided catheter ablation in complex cardiac substrates such as left atrial flutter. This protocol describes all the clinical and invasive interventional steps performed during a human electrophysiological study and ablation to assess the accuracy, safety and real-time navigation of the Catheter Guidance, Control and Imaging (CGCI) system. Patients who underwent ablation of a right or left atrium flutter substrate were included. Specifically, data from three left atrial flutter and two counterclockwise right atrial flutter procedures are shown in this report. One representative left atrial flutter procedure is shown in the movie. This system is based on eight coil-core electromagnets, which generate a dynamic magnetic field focused on the heart. Remote navigation by rapid changes (msec) in the magnetic field magnitude and a very flexible magnetized catheter allow real-time closed-loop integration and accurate, stable positioning and ablation of the arrhythmogenic substrate.

This report provides a detailed description of a new remote navigation system based on magnetic driven forces, which has been recently introduced as a new robotic tool for human cardiac electrophysiology procedures.

心臓電気生理学プロシージャでの正確なリアルタイムカテーテルポジショニングとアブレーション用のリモート磁気ナビゲーション

New remote navigation systems have been developed to improve current limitations of conventional manually guided catheter ablation in complex cardiac ...

MicroRNA Detection in Prostate Tumors by Quantitative Real-time PCR (qPCR)

MicroRNAs (miRNAs) are single-stranded, 18&#x2013;24 nucleotide long, non-coding RNA molecules. They are involved in virtually every cellular process including development1, apoptosis2, and cell cycle regulation3. MiRNAs are estimated to regulate the expression of 30% to 90% of human genes4 by binding to their target messenger RNAs (mRNAs)5. Widespread dysregulation of miRNAs has been reported in various diseases and cancer subtypes6. Due to their prevalence and unique structure, these small molecules are likely to be the next generation of biomarkers, therapeutic agents and/or targets.
Methods used to investigate miRNA expression include SYBR green I dye- based as well as Taqman-probe based qPCR. If miRNAs are to be effectively used in the clinical setting, it is imperative that their detection in fresh and/or archived clinical samples be accurate, reproducible, and specific. qPCR has been widely used for validating expression of miRNAs in whole genome analyses such as microarray studies7. The samples used in this protocol were from patients who underwent radical prostatectomy for clinically localized prostate cancer; however other tissues and cell lines can be substituted in. Prostate specimens were snap-frozen in liquid nitrogen after resection. Clinical variables and follow-up information for each patient were collected for subsequent analysis8.
Quantification of miRNA levels in prostate tumor samples. The main steps in qPCR analysis of tumors are: Total RNA extraction, cDNA synthesis, and detection of qPCR products using miRNA-specific primers. Total RNA, which includes mRNA, miRNA, and other small RNAs were extracted from specimens using TRIzol reagent. Qiagen's miScript System was used to synthesize cDNA and perform qPCR (Figure 1). Endogenous miRNAs are not polyadenylated, therefore during the reverse transcription process, a poly(A) polymerase polyadenylates the miRNA. The miRNA is used as a template to synthesize cDNA using oligo-dT and Reverse Transcriptase. A universal tag sequence on the 5' end of oligo-dT primers facilitates the amplification of cDNA in the PCR step. PCR product amplification is detected by the level of fluorescence emitted by SYBR Green, a dye which intercalates into double stranded DNA. Specific miRNA primers, along with a Universal Primer that binds to the universal tag sequence will amplify specific miRNA sequences. 
The miScript Primer Assays are available for over a thousand human-specific miRNAs, and hundreds of murine-specific miRNAs. Relative quantification method was used here to quantify the expression of miRNAs. To correct for variability amongst different samples, expression levels of a target miRNA is normalized to the expression levels of a reference gene. The choice of a gene on which to normalize the expression of targets is critical in relative quantification method of analysis. Examples of reference genes typically used in this capacity are the small RNAs RNU6B, RNU44, and RNU48 as they are considered to be stably expressed across most samples. In this protocol, RNU6B is used as the reference gene.

MicroRNA Detection in Prostate Tumors by Quantitative Real-time PCR qPCR

Quantitative Real Time polymerase chain reaction (qPCR) is a rapid and sensitive method to investigate the expression levels of various microRNA (miRNA) molecules in tumor samples. Using this method expression of hundreds of different miRNA molecules can be amplified, quantified, and analyzed from the same cDNA template.

定量的リアルタイムPCR（定量PCR）による前立腺腫瘍におけるマイクロRNAの検出

MicroRNAs (miRNAs) are single-stranded, 18&#x2013;24 nucleotide long, non-coding RNA molecules. They are involved in virtually every cellular process ...

Movement Retraining using Real-time Feedback of Performance

Any modification of movement - especially movement patterns that have been honed over a number of years - requires re-organization of the neuromuscular patterns responsible for governing the movement performance. This motor learning can be enhanced through a number of methods that are utilized in research and clinical settings alike. In general, verbal feedback of performance in real-time or knowledge of results following movement is commonly used clinically as a preliminary means of instilling motor learning. Depending on patient preference and learning style, visual feedback (e.g. through use of a mirror or different types of video) or proprioceptive guidance utilizing therapist touch, are used to supplement verbal instructions from the therapist. Indeed, a combination of these forms of feedback is commonplace in the clinical setting to facilitate motor learning and optimize outcomes.
Laboratory-based, quantitative motion analysis has been a mainstay in research settings to provide accurate and objective analysis of a variety of movements in healthy and injured populations. While the actual mechanisms of capturing the movements may differ, all current motion analysis systems rely on the ability to track the movement of body segments and joints and to use established equations of motion to quantify key movement patterns. Due to limitations in acquisition and processing speed, analysis and description of the movements has traditionally occurred offline after completion of a given testing session.
This paper will highlight a new supplement to standard motion analysis techniques that relies on the near instantaneous assessment and quantification of movement patterns and the display of specific movement characteristics to the patient during a movement analysis session. As a result, this novel technique can provide a new method of feedback delivery that has advantages over currently used feedback methods.

Retraining abnormal movement patterns following injury or disease is a key component of physical rehabilitation. Recent advances in technology have permitted accurate assessment of movement during a variety of tasks, with near instantaneous quantification of results. This provides new opportunities for modification of faulty movement patterns in real time.

パフォーマンスのリアルタイムのフィードバックを使用して再教育運動

Any modification of movement - especially movement patterns that have been honed over a number of years - requires re-organization of the neuromuscular ...

Transplantation into the Anterior Chamber of the Eye for Longitudinal, Non-invasive In vivo Imaging with Single-cell Resolution in Real-time

Intravital imaging has emerged as an indispensable tool in biological research. In the process, many imaging techniques have been developed to study different biological processes in animals non-invasively. However, a major technical limitation in existing intravital imaging modalities is the inability to combine non-invasive, longitudinal imaging with single-cell resolution capabilities. We show here how transplantation into the anterior chamber of the eye circumvents such significant limitation offering a versatile experimental platform that enables non-invasive, longitudinal imaging with cellular resolution in vivo. We demonstrate the transplantation procedure in the mouse and provide representative results using a model with clinical relevance, namely pancreatic islet transplantation. In addition to enabling direct visualization in a variety of tissues transplanted into the anterior chamber of the eye, this approach provides a platform to screen drugs by performing long-term follow up and monitoring in target tissues. Because of its versatility, tissue/cell transplantation into the anterior chamber of the eye not only benefits transplantation therapies, it extends to other in vivo applications to study physiological and pathophysiological processes such as signal transduction and cancer or autoimmune disease development.

Transplantation into the Anterior Chamber of the Eye for Longitudinal, Non-invasive In vivo Imaging with Single-cell Resolution in Real-time

A new approach combining intraocular transplantation and confocal microscopy enables longitudinal, non-invasive real-time imaging with single-cell resolution within grafted tissues in vivo. We demonstrate how to transplant pancreatic islets into the anterior chamber of the mouse eye.

縦用の眼の前房に移植、非侵襲<emインビボ&gt;</emリアルタイムでのシングルセル分解能&gt;イメージング

Intravital imaging has emerged as an indispensable tool in biological research.  In the process, many imaging techniques have been developed to study ...

Cerebrospinal Fluid MicroRNA Profiling Using Quantitative Real Time PCR

MicroRNAs (miRNAs) constitute a potent layer of gene regulation by guiding RISC to target sites located on mRNAs and, consequently, by modulating their translational repression. Changes in miRNA expression have been shown to be involved in the development of all major complex diseases. Furthermore, recent findings showed that miRNAs can be secreted to the extracellular environment and enter the bloodstream and other body fluids where they can circulate with high stability. The function of such circulating miRNAs remains largely elusive, but systematic high throughput approaches, such as miRNA profiling arrays, have lead to the identification of miRNA signatures in several pathological conditions, including neurodegenerative disorders and several types of cancers. In this context, the identification of miRNA expression profile in the cerebrospinal fluid, as reported in our recent study, makes miRNAs attractive candidates for biomarker analysis.
There are several tools available for profiling microRNAs, such as microarrays, quantitative real-time PCR (qPCR), and deep sequencing. Here, we describe a sensitive method to profile microRNAs in cerebrospinal fluids by quantitative real-time PCR. We used the Exiqon microRNA ready-to-use PCR human panels I and II V2.R, which allows detection of 742 unique human microRNAs. We performed the arrays in triplicate runs and we processed and analyzed data using the GenEx Professional 5 software.
Using this protocol, we have successfully profiled microRNAs in various types of cell lines and primary cells, CSF, plasma, and formalin-fixed paraffin-embedded tissues.

We describe a protocol of real time PCR to profile microRNAs in the cerebrospinal fluid (CSF). With the exception of RNA extraction protocols, the procedure can be extended to RNA extracted from other body fluids, cultured cells, or tissue specimens.

定量リアルタイムPCRを用いて脳脊髄液マイクロRNAプロファイリング

MicroRNAs (miRNAs) constitute a potent layer of gene regulation by guiding RISC to target sites located on mRNAs and, consequently, by modulating their ...

Real-time Cytotoxicity Assays in Human Whole Blood

A live cell-based whole blood cytotoxicity assay (WCA) that allows access to temporal information of the overall cell cytotoxicity is developed with high-throughput cell positioning technology. The targeted tumor cell populations are first preprogrammed to immobilization into an array format, and labeled with green fluorescent cytosolic dyes. Following the cell array formation, antibody drugs are added in combination with human whole blood. Propidium iodide (PI) is then added to assess cell death. The cell array is analyzed with an automatic imaging system. While cytosolic dye labels the targeted tumor cell populations, PI labels the dead tumor cell populations. Thus, the percentage of target cancer cell killing can be quantified by calculating the number of surviving targeted cells to the number of dead targeted cells. With this method, researchers are able to access time-dependent and dose-dependent cell cytotoxicity information. Remarkably, no hazardous radiochemicals are used. The WCA presented here has been tested with lymphoma, leukemia, and solid tumor cell lines. Therefore, WCA allows researchers to assess drug efficacy in a highly relevant ex&#160;vivo condition.

The whole blood cytotoxicity assay (WCA) is a cytotoxicity assay developed by incorporating high-throughput cell positioning technology with fluorescence microscopy and automated image processing. Here, we describe how lymphoma cells treated with an anti-CD20 antibody can be analyzed real-time in human whole blood to provide quantitative cellular cytotoxicity analysis.

ヒト全血でのリアルタイム細胞毒性アッセイ

A live cell-based whole blood cytotoxicity assay (WCA) that allows access to temporal information of the overall cell cytotoxicity is developed with ...

Extended Time-lapse Intravital Imaging of Real-time Multicellular Dynamics in the Tumor Microenvironment

In the tumor microenvironment, host stromal cells interact with tumor cells to promote tumor progression, angiogenesis, tumor cell dissemination and metastasis. Multicellular interactions in the tumor microenvironment can lead to transient events including directional tumor cell motility and vascular permeability. Quantification of tumor vascular permeability has frequently used end-point experiments to measure extravasation of vascular dyes. However, due to the transient nature of multicellular interactions and vascular permeability, the kinetics of these dynamic events cannot be discerned. By labeling cells and vasculature with injectable dyes or fluorescent proteins, high-resolution time-lapse intravital microscopy has allowed the direct, real-time visualization of transient events in the tumor microenvironment. Here we describe a method for using multiphoton microscopy to perform extended intravital imaging in live mice to directly visualize multicellular dynamics in the tumor microenvironment. This method details cellular labeling strategies, the surgical preparation of a mammary skin flap, the administration of injectable dyes or proteins by tail vein catheter and the acquisition of time-lapse images. The time-lapse sequences obtained from this method facilitate the visualization and quantitation of the kinetics of cellular events of motility and vascular permeability in the tumor microenvironment.

This protocol describes the use of multiphoton microscopy to perform extended time-lapse imaging of multicellular interactions in real time, in vivo at single cell resolution.

腫瘍微小環境におけるリアルタイム多細胞ダイナミクスの拡張タイムラプス生体内イメージング

In the tumor microenvironment, host stromal cells interact with tumor cells to promote tumor progression, angiogenesis, tumor cell dissemination and ...

Real-time Pressure-volume Analysis of Acute Myocardial Infarction in Mice

Acute myocardial infarction can lead to acute heart failure and cardiogenic shock. The evaluation of hemodynamics is critical for the evaluation of any potential therapeutic approach directed against acute left ventricular (LV) dysfunction. Current imaging modalities (e.g., echocardiography and magnetic resonance imaging) have several limitations since data on LV pressure cannot directly be measured. LV catheterization in mice undergoing coronary artery occlusion could serve as a novel method for a real-time evaluation of LV function.
At the beginning of the procedure, mice were anesthetized followed by endotracheal intubation. For LV catheterization, the right carotid artery was exposed via middle-neck incision. The catheter was introduced and placed into the LV cavity. Left thoracotomy was conducted and the left main coronary artery (LCA) was ligated. To induce reperfusion, the suture was released after 45 min. Pressure-volume data was recorded at all times.
Ligation of the LCA caused a decrease in LV systolic function as evidenced by a 30% reduction in stroke volume, LV ejection fraction (EF), and cardiac output. Maximum dP/dt as a parameter for LV contractility was also significantly reduced and diastolic function was severely impaired (minimum dP/dt -40%). Reperfusion over a period of 20 min did not lead to a complete recovery of LV function.
Real-time pressure-volume analysis served as a valid procedure for monitoring cardiac function during acute myocardial infarction in mice. Maintaining stable anesthesia and a standardized surgical approach was crucial to ensure valid results. As the early phase of acute myocardial infarction is critical for morbidity and mortality, the delineated method could be beneficial for preclinical evaluation of new strategies for cardioprotection.

Acute myocardial infarction in mice induces acute but incompletely characterized changes in left ventricular (LV) function. LV catheterization in mice undergoing coronary artery occlusion serves as a novel method for a real-time evaluation of LV function.

マウスにおける急性心筋梗塞のリアルタイム圧解析

Acute myocardial infarction can lead to acute heart failure and cardiogenic shock. The evaluation of hemodynamics is critical for the evaluation of any ...

Bio-energetics Investigation of Candida albicans Using Real-time Extracellular Flux Analysis

Mitochondria are essential organelles for the cellular metabolism and survival. A variety of key events take place in mitochondria, such as cellular respiration, oxidative metabolism, signal transduction, and apoptosis. Consequently, mitochondrial dysfunction is reported to play an important role in the antifungal drug tolerance and virulence of pathogenic fungi. Recent data have also led to the recognition of the importance of the mitochondria as an important contributor to fungal pathogenesis. Despite the importance of the mitochondria in fungal biology, standardized methods to understand its function are poorly developed. Here, we present a procedure to study the basal oxygen consumption rate (OCR), a measure of mitochondrial respiration, and extracellular acidification rates (ECAR), a measure of glycolytic function in C. albicans strains. The method described herein can be applied to any Candidaspp. strains without the need to purify mitochondria from the intact fungal cells. Furthermore, this protocol can also be customized to screen for inhibitors of mitochondrial function in C. albicans strains.

Bio-energetics Investigation of Candida albicans Using Real-time Extracellular Flux Analysis

Here, we present a stepwise protocol to investigate the mitochondrial respiration and glycolytic function in Candida Albicans using an extra flux analyzer.

カンジダを使用してリアルタイム細胞フラックス解析のバイオ エネルギー調査

Mitochondria are essential organelles for the cellular metabolism and survival. A variety of key events take place in mitochondria, such as cellular ...