Name: coordinate mapping of hyolaryngeal mechanics in swallowing
Uploaded: 2014-05-06T15:30:00
Description: Watch this Scientific Journal Video about Coordinate Mapping of Hyolaryngeal Mechanics in Swallowing at JoVE.com

लेखकों Kendrea Focht, CScD, CCC-SLP, और आवाज के लिए एवलिन Trammell संस्थान को स्वीकार करते हैं और एमबीएस इस पद्धति को प्रदर्शित करने के लिए इस्तेमाल किया इमेजिंग फ़ाइलों को साझा करने के लिए, दक्षिण कैरोलिना के चिकित्सा विश्वविद्यालय में निगलना. (पीआई: मार्टिन हैरिस) translational विज्ञान में आगे बढ़ने के लिए और अनुदान संख्या 1K24DC12801 द्वारा राष्ट्रीय केन्द्र से: ये एमबीएस डेटा अनुदान संख्या TL1TR000061 (Focht पीआई) द्वारा वित्त पोषित बाह्य सहायता के माध्यम से एकत्र किए गए थे बहरापन पर राष्ट्रीय संस्थान और अन्य संचार विकार से, और मार्क और एवलिन Trammell ट्रस्ट से अंदर का समर्थन करते हैं. बहरापन और अन्य संचार विकार पर राष्ट्रीय संस्थान से अनुदान संख्या F31DC011705 द्वारा समर्थित है, जबकि इन विधियों मूल रूप से प्रमुख अन्वेषक द्वारा विकसित किए गए. सामग्री केवल लेखकों की ज़िम्मेदारी है और जरूरी बहरापन पर राष्ट्रीय संस्थान और अन्य संचार विकार या राष्ट्रीय संस्थानों के अधिकारी विचार का प्रतिनिधित्व नहीं करतास्वास्थ्य.

1. एक कंप्यूटर का विन्यास <ol><li> ImageJ, MacX वीडियो कनवर्टर नि: शुल्क संस्करण (मैक), और QuickTime (सामग्री / उपकरणों की तालिका देखें): लबादा के लिए निम्न खुला स्रोत या फ्रीवेयर सॉफ्टवेयर डाउनलोड. </li><li> ImageJ, एमपीईजी streamclip (पीसी), ?...

<ol>
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इस अध्ययन निगलने में hyolaryngeal आंदोलन के कई कीनेमेटीक्स माप की गणना करने के लिए संरचनात्मक स्थलों का डेटा समन्वय का उपयोग कर एक विधि की उपयोगिता को दर्शाता है. निर्देशांक और गणना चर के लिए दो नौसिखिए रेटर, स?...

निगलने की ग्रसनी चरण airway की रक्षा करते हुए घेघा में मौखिक गुहा से एक सांस में स्थानांतरित करने के लिए बीस से अधिक मांसपेशियों और कई कंकाल तत्वों को शामिल एक जटिल प्रक्रिया है. ग्रसनी कसना, hyolaryngeal परिसर के तत्वों (कष्ठिका अस्थि, गला, और ऊपरी esophageal दबानेवाला यंत्र सहित एसोसिएटेड संरचनाओं) पूर्ववर्ती एक पाचन तंत्र में एक सांस की नाली में परिवर्तित करने के लिए विस्थापित कर रहे हैं. गला पूर्व से दूर एक आनेवाला सांस की गति से फिर से स्थित है, और ऊपरी esophageal दबानेवाला यंत्र खुला फैला है. (यह भी एक एमबीएस के रूप में जाना जाता है या बेरियम निगल अध्ययन संशोधित) वीडियो fluoroscopic निगलने पढ़ाई से लिया विज्ञान सम्बन्धी माप hyolaryngeal जटिल 1 के कई आंदोलनों बढ़ाता के लिए प्राथमिक अनुसंधान पद्धति हैं. मात्रात्मक वीडियो fluoroscopic माप निगलने समारोह, संदर्भ के विभिन्न कुल्हाड़ियों मापने के लिए उपयोगी होते हैंऔर विज्ञान सम्बन्धी माप 2 के विभिन्न तरीकों के बीच असंगत हैं कि निष्कर्ष में माप परिणाम की scalars. पुस्तिका चिकित्सक नियंत्रण में रोगी और प्रतिदीप्तिदर्शी की आवाजाही भी इस जटिल शारीरिक प्रक्रिया को मापने की सटीकता घालमेल कर दिया है. इससे भी महत्वपूर्ण बात, विज्ञान सम्बन्धी माप जरूरी अव्यवस्थित निगलने के मूल्यांकन के लिए महत्वपूर्ण संरचना को समारोह रिश्तों को प्रतिबिंबित नहीं करते. विशेष रूप से hyoid की कीनेमेटीक्स कशेरुकाओं के साथ गठबंधन एक संरचनात्मक विमान के संदर्भ में एक पूर्वकाल या बेहतर दिशा में आंदोलन को ट्रैक करने के लिए डिजाइन किया गया है. हालांकि, इस विन्यास hyoid निलंबित है कि मांसपेशियों की कार्रवाई की लाइन का प्रतिनिधित्व नहीं करता. निगलने की ग्रसनी चरण में hyolaryngeal ऊंचाई की एक दो गोफन तंत्र (चित्रा 1) 3,4 पहचान की गई है. suprahyoid मांसपेशियों पूर्वकाल पेशी गोफन शामिल है, और लंबे ग्रसनी मांसपेशियों पी शामिलपेशी गोफन osterior. इन मांसपेशियों hyoid, गला, और ऊपरी esophageal दबानेवाला यंत्र है कि फार्म संरचनाओं सहित hyolaryngeal परिसर के विभिन्न तत्वों को ऊपर उठाना है. Hyolaryngeal यांत्रिकी का समन्वय मानचित्रण पूर्वकाल और कूल्हों पेशी slings (चित्रा 2) के hyolaryngeal परिसर का प्रतिनिधित्व लगाव अंक के तीन कंकाल लीवर और सुविधाओं नक्शा करने के लिए नौ आसानी से पहचाना संरचनात्मक स्थलों का इस्तेमाल करता. निगलने के दौरान, hyolaryngeal परिसर के प्रत्येक कंकाल लीवर और सुविधा गति में है. निर्देशांक एकत्रित करके, प्रणाली किसी भी समय सीमा में कब्जा कर लिया जा सकता है. निर्देशांक के त्रिकोणमितीय रूपांतरण निगलने के दौरान hyolaryngeal आंदोलन के कई कीनेमेटीक्स माप उत्पन्न करने के लिए इस्तेमाल किया जा सकता है. चर साहित्य में रिपोर्ट के निष्कर्षों के साथ तुलना के लिए गणना, या ब्याज की संरचना करने के लिए समारोह रिश्तों का प्रतिनिधित्व नई माप उत्पन्न करने के लिए इस्तेमाल किया जा सकता है. इस कागज का प्राथमिक उद्देश्य संशोधित बेरियम निगल (एमबीएस) के अध्ययन से एकत्र संरचनात्मक मील का पत्थर निर्देशांक का एक सेट से गणना की कई विज्ञान सम्बन्धी माप पैदा करने की एक विधि का प्रदर्शन है. हम एक विशेषज्ञ, अनुभव के साथ तीन रेटर, और दो novices सहित 6 अलग जांचकर्ताओं के अंतर - करदाता विश्वसनीयता निर्धारित करने के लिए Intraclass सहसंबंध गुणांक का उपयोग करके इस विधि की विश्वसनीयता दस्तावेज़. विज्ञान सम्बन्धी परिणामों से, सांस में स्थिरता से यांत्रिकी निगलने में मतभेद मूल्यांकन कर रहे हैं. अंत में, hyoid आंदोलन को मापने में प्रयोग किया संदर्भ के अक्ष के महत्व के विषय में Molefenter और स्टील द्वारा प्रस्तावित सवाल को संबोधित किया है. हम कशेरुकाओं के संदर्भ में और दोनों सांस प्रकार के लिए निर्देशांक का एक ही सेट से गणना जबड़ा, के संदर्भ में hyoid भ्रमण के माप की तुलना में इस सवाल दृष्टिकोण को. Hyoid आंदोलन को मापने के इन दो तरीकों के संबंधों से कार्य करने के लिए एक ही संरचना का प्रतिनिधित्व करते हैंकूल्हे, तो परिणाम जोरदार सहसंबद्ध किया जाना चाहिए. इस अध्ययन के लिए, 40 पार्श्व दृश्य एमबीएस पढ़ाई बेतरतीब ढंग जॉर्जिया राज्य प्रतिनिधियों विश्वविद्यालय संस्थागत समीक्षा बोर्ड और आवाज के लिए एवलिन Trammell संस्थान के साथ अनुसंधान सहयोग में द्वारा अनुमोदित और के चिकित्सा विश्वविद्यालय में निगलना एक शोध प्रोटोकॉल के तहत 139 सामान्य अध्ययन का एक संग्रह से चयन किया गया था दक्षिण कैरोलिना. इस विधि की उपयोगिता का प्रदर्शन, hyolaryngeal कीनेमेटीक्स निस्र्पक दस चर समन्वय डेटा का एक ही सेट (1 टेबल) से गणना की गई. इन गणना की माप के सात पहले सहित साहित्य में इस्तेमाल किया गया है: पूर्वकाल और कशेरुकाओं 5 के संदर्भ में बेहतर hyoid दूरी माप; पूर्वकाल और भी कशेरुकाओं 6 के संदर्भ में सी 2-4 लंबाई के अनुपात के रूप में बेहतर hyoid विस्थापन; कशेरुकाओं 7 के संदर्भ में गला के बेहतर आंदोलन; hyolaryngeal approximatआयन 1; और कशेरुकाओं 1 के संदर्भ में अधिकतम hyoid भ्रमण. इसके अलावा, तीन उपन्यास माप गणना की गई: ग्रसनी छोटा suprahyoid मांसपेशियों 4,8 के संलग्नक approximating palatopharyngeus पेशी के संलग्नक, stylopharyngeus का प्रतिनिधित्व कार्रवाई की एक पंक्ति निम्नलिखित स्वरयंत्रज ऊंचाई, और hyoid भ्रमण approximating. एक विशेषज्ञ सिर और गर्दन anatomist (WP), सीमित अनुभव लेने माप (मुख्य न्यायाधीश, एसआर, टीटी), और दो नौसिखिए जांचकर्ताओं (रुपये, संयुक्त) के साथ तीन जांचकर्ताओं नीचे वर्णित प्रोटोकॉल का उपयोग मानचित्रण डेटा समन्वय प्राप्त की. विशेषज्ञ (WP) के अनुभव के साथ तीन रेटर प्रशिक्षित है, और बदले में ये दो नौसिखिए रेटर प्रशिक्षित किया. विषय से निर्देशांक से गणना समन्वय डेटा और परिणाम के अंतर्राज्यीय करदाता विश्वसनीयता Intraclass सहसंबंध गुणांक 9 द्वारा निर्धारित किया गया था. दो पूंछ टी परीक्षण statisticall निर्धारित करने के लिए प्रत्येक चर पर प्रदर्शन किया गयासांस प्रकार में Y महत्वपूर्ण मतभेद. एक पियर्सन सहसंबंध गुणांक और दृढ़ संकल्प के गुणांक 5 मिलीलीटर पतली तरल निगल और 5 मिलीलीटर के लिए संदर्भ की एक धुरी के रूप में जबड़ा साथ hyoid भ्रमण बनाम संदर्भ की एक धुरी के रूप में कशेरुकाओं के साथ गणना hyoid भ्रमण के परिणामों के बीच समझौते का मूल्यांकन करने के लिए इस्तेमाल किया गया हलवा निगल.

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coordinate mapping of hyolaryngeal mechanics in swallowing

Hyolaryngeal आंदोलन निस्र्पक निगलने में कठिनाई अनुसंधान के लिए महत्वपूर्ण है. पिछले विधियों हित के कई चर की गणना करने के लिए निर्देशांक का एक सेट का उपयोग करता संशोधित बेरियम निगल (एमबीएस) का उपयोग hyolaryngeal यांत्रिकी के मानचित्रण समन्वय जबकि एक विज्ञान सम्बन्धी माप प्राप्त करने के लिए कई मापन की आवश्यकता होती है. प्रदर्शन प्रयोजनों के लिए, दस विज्ञान सम्बन्धी माप दो अलग सांस प्रकार निगलने में मतभेद निर्धारित करने के लिए निर्देशांक का एक सेट से उत्पन्न किया गया. कशेरुकाओं और जबड़ा के खिलाफ hyoid भ्रमण की गणना संदर्भ की कुल्हाड़ियों के महत्व को निर्धारित करने के लिए सहसंबद्ध होते हैं. मैपिंग पद्धति समन्वय का प्रदर्शन, 40 एमबीएस पढ़ाई बेतरतीब ढंग से कोई ज्ञात निगलने हानि के साथ स्वस्थ सामान्य विषयों की एक डाटासेट से चयन किया गया था. एक 5 मिलीलीटर पतली तरल सांस और एक 5 मिलीलीटर हलवा निगल प्रत्येक विषय से मापा गया. कपाल आधार, जबड़ा, कशेरुकाओं और Hyo के तत्वों मानचित्रण नौ निर्देशांक,स्वरयंत्रज जटिल, न्यूनतम और अधिकतम hyolaryngeal भ्रमण के तख्ते पर दर्ज किए गए. निर्देशांक गणितीय hyolaryngeal यांत्रिकी के दस चर में परिवर्तित किया गया. इंटर करदाता विश्वसनीयता Intraclass सहसंबंध गुणांक (आईसीसी) द्वारा मूल्यांकन किया गया था. दो पूंछ टी परीक्षण सांस में चिपचिपाहट से कीनेमेटीक्स में मतभेद का मूल्यांकन किया गया. संदर्भ के विभिन्न कुल्हाड़ियों के खिलाफ Hyoid भ्रमण माप सहसंबद्ध थे. 0.97 - 18 निर्देशांक के लिए छह रेटर के बीच अंतर - करदाता विश्वसनीयता आईसीसी से = 0.90 बताया गया. दस विज्ञान सम्बन्धी माप की एक स्लेट छह रेटर के बीच इस विषय से तुलना की थी. एक ग़ैर खारिज कर दिया, और शेष विश्वसनीयता के स्कोर का मतलब आईसीसी = 0.91, 0.84 था - 0.96, 95% सीआई. दस कीनेमेटीक्स चर (5 मिलीलीटर पतली तरल बनाम 5 एमएल हलवा निगल) की तुलना Bonferroni सुधार के साथ दो पूंछ टी परीक्षण hyoid भ्रमण, बेहतर laryngeal आंदोलन, और ग्रसनी shortenin में सांख्यिकीय महत्वपूर्ण अंतर दिखायाजी (पी &lt;0.005). आर = 0.62, 2 आर = 0.38, (पतली तरल);: संदर्भ के दो विभिन्न कुल्हाड़ियों से hyoid भ्रमण माप के पियर्सन सहसंबंध थे आर = 0.52, 2 आर = 0.27, (हलवा). मील का पत्थर निर्देशांक प्राप्त निगलने में कठिनाई अनुसंधान में उपयोगी वीडियो fluoroscopic छवियों से कई कीनेमेटीक्स चर उत्पन्न करने के लिए एक विश्वसनीय तरीका है.

Intraclass सहसंबंध गुणांक स्वतंत्र रूप से 80 वीडियो fluoroscopic फाइलें (40 विषयों से दो सांस परीक्षण) का विश्लेषण किया जो छह जांचकर्ताओं द्वारा एकत्र निर्देशांक (आईसीसी) लेकर आईसीसी से = 0.90-0.97. के रूप में निम्नानुसार समू?...

Watch this Scientific Journal Video about Coordinate Mapping of Hyolaryngeal Mechanics in Swallowing at JoVE.com

Coordinate Mapping of Hyolaryngeal Mechanics in Swallowing

कोआर्डिनेट मानचित्रण निगलने की ग्रसनी चरण में hyolaryngeal बायोमैकेनिक्स की मुख्य विशेषताओं का दस्तावेजीकरण करने की एक विधि है. इस पद्धति संरचनात्मक स्थलों के निर्देशांक रिकॉर्ड करने के लिए छवि विश्लेषण सॉफ्टवेयर का उपयोग करता है. ये निर्देशांक एक एक्सेल स्थूल में आयात और निगलने में कठिनाई अनुसंधान में उपयोगी ब्याज की कीनेमेटीक्स चर में अनुवाद कर रहे हैं.

निगलने में Hyolaryngeal मैकेनिक्स की मैपिंग कोआर्डिनेट

Characterizing hyolaryngeal movement is important to dysphagia research. Prior methods require multiple measurements to obtain one kinematic measurement ...

coordinate-mapping-of-hyolaryngeal-mechanics-in-swallowing

Research

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Medicine

Guide Wire Assisted Catheterization and Colored Dye Injection for Vascular Mapping of Monochorionic Twin Placentas

Monochorionic (MC) twin pregnancies are associated with significantly higher morbidity and mortality rates than dichorionic twins. Approximately 50% of MC twin pregnancies develop complications arising from the shared placenta and associated vascular connections1. Severe twin-to-twin syndrome (TTTS) is reported to account for approximately 20% of these complications2,3. Inter-twin vascular connections occur in almost all MC placentas and are related to the prognosis and outcome of these high-risk twin pregnancies. The number, size and type of connections have been implicated in the development of TTTS and other MC twin conditions. Three types of inter-twin vascular connections occur: 1) artery to vein connections (AVs) in which a branch artery carrying deoxygenated blood from one twin courses along the fetal surface of the placenta and dives into a placental cotyledon. Blood flows via a deep intraparenchymal capillary network into a draining vein that emerges at the fetal surface of the placenta and brings oxygenated blood toward the other twin. There is unidirectional flow from the twin supplying the afferent artery toward the twin receiving the efferent vein; 2) artery to artery connections (AAs) in which a branch artery from each twin meets directly on the superficial placental surface resulting in a vessel with pulsatile bidirectional flow, and 3) vein to vein connections (VVs) in which a branch vein from each twin meets directly on the superficial placental surface allowing low pressure bidirectional flow. In utero obstetric sonography with targeted Doppler interrogation has been used to identify the presence of AV and AA connections4. Prenatally detected AAs that have been confirmed by postnatal placental injection studies have been shown to be associated with an improved prognosis for both twins5. Furthermore, fetoscopic laser ablation of inter-twin vascular connections on the fetal surface of the shared placenta is now the preferred treatment for early, severe TTTS. 
Postnatal placental injection studies provide a valuable method to confirm the accuracy of prenatal Doppler ultrasound findings and the efficacy of fetal laser therapy6. Using colored dyes separately hand-injected into the arterial and venous circulations of each twin, the technique highlights and delineates AVs, AAs, and VVs. This definitive demonstration of MC placental vascular anatomy may then be correlated with Doppler ultrasound findings and neonatal outcome to enhance our understanding of the pathophysiology of MC twinning and its sequelae. Here we demonstrate our placental injection technique.

Vascular mapping of monochorionic (MC) twin placentas after birth provides a means for detailed demonstration of vascular connections between the twins&#x2019; circulations. 
Imbalance of these connections is thought to play a pivotal role in the development of complications of MC twinning including twin-to-twin transfusion syndrome.

गाइड वायर असिस्टेड और Monochorionic जुड़वा placentas के संवहनी मानचित्रण के लिए रंग डाई इंजेक्शन कैथीटेराइजेशन

Monochorionic (MC) twin pregnancies are associated with significantly higher morbidity and mortality rates than dichorionic twins. Approximately 50% of MC ...

High-Resolution Endocardial and Epicardial Optical Mapping in a Sheep Model of Stretch-Induced Atrial Fibrillation

Atrial fibrillation (AF) is a complex cardiac arrhythmia with high morbidity and mortality.1,2 It is the most common sustained cardiac rhythm disturbance seen in clinical practice and its prevalence is expected to increase in the coming years.3 Increased intra-atrial pressure and dilatation have been long recognized to lead to AF,1,4 which highlights the relevance of using animal models and stretch to study AF dynamics. Understanding the mechanisms underlying AF requires visualization of the cardiac electrical waves with high spatial and temporal resolution. While high-temporal resolution can be achieved by conventional electrical mapping traditionally used in human electrophysiological studies, the small number of intra-atrial electrodes that can be used simultaneously limits the spatial resolution and precludes any detailed tracking of the electrical waves during the arrhythmia. The introduction of optical mapping in the early 90's enabled wide-field characterization of fibrillatory activity together with sub-millimeter spatial resolution in animal models5,6 and led to the identification of rapidly spinning electrical wave patterns (rotors) as the sources of the fibrillatory activity that may occur in the ventricles or the atria.7-9 Using combined time- and frequency-domain analyses of optical mapping it is possible to demonstrate discrete sites of high frequency periodic activity during AF, along with frequency gradients between left and right atrium. The region with fastest rotors activates at the highest frequency and drives the overall arrhythmia.10,11 The waves emanating from such rotor interact with either functional or anatomic obstacles in their path, resulting in the phenomenon of fibrillatory conduction.12 Mapping the endocardial surface of the posterior left atrium (PLA) allows the tracking of AF wave dynamics in the region with the highest rotor frequency. Importantly, the PLA is the region where intracavitary catheter-based ablative procedures are most successful terminating AF in patients,13 which underscores the relevance of studying AF dynamics from the interior of the left atrium. Here we describe a sheep model of acute stretch-induced AF, which resembles some of the characteristics of human paroxysmal AF. Epicardial mapping on the left atrium is complemented with endocardial mapping of the PLA using a dual-channel rigid borescope c-mounted to a CCD camera, which represents the most direct approach to visualize the patterns of activation in the most relevant region for AF maintenance.

This report provides a detailed description of the methodology and results of simultaneous endocardial and epicardial optical mapping of electrical excitation in the intact left atrium of a Langendorff-perfused sheep heart during stretch-induced atrial fibrillation.

उच्च संकल्प endocardial और Epicardial खींचो प्रेरित atrial fibrillation के एक भेड़ मॉडल में ऑप्टिकल मानचित्रण

 Atrial fibrillation (AF) is a complex cardiac arrhythmia with high morbidity and mortality.1,2 It is the most common sustained cardiac rhythm disturbance ...

Heterogeneity Mapping of Protein Expression in Tumors using Quantitative Immunofluorescence

Morphologic heterogeneity within an individual tumor is well-recognized by histopathologists in surgical practice. While this often takes the form of areas of distinct differentiation into recognized histological subtypes, or different pathological grade, often there are more subtle differences in phenotype which defy accurate classification (Figure 1). Ultimately, since morphology is dictated by the underlying molecular phenotype, areas with visible differences are likely to be accompanied by differences in the expression of proteins which orchestrate cellular function and behavior, and therefore, appearance. The significance of visible and invisible (molecular) heterogeneity for prognosis is unknown, but recent evidence suggests that, at least at the genetic level, heterogeneity exists in the primary tumor1,2, and some of these sub-clones give rise to metastatic (and therefore lethal) disease. 
Moreover, some proteins are measured as biomarkers because they are the targets of therapy (for instance ER and HER2 for tamoxifen and trastuzumab (Herceptin), respectively). If these proteins show variable expression within a tumor then therapeutic responses may also be variable. The widely used histopathologic scoring schemes for immunohistochemistry either ignore, or numerically homogenize the quantification of protein expression. Similarly, in destructive techniques, where the tumor samples are homogenized (such as gene expression profiling), quantitative information can be elucidated, but spatial information is lost. Genetic heterogeneity mapping approaches in pancreatic cancer have relied either on generation of a single cell suspension3, or on macrodissection4. A recent study has used quantum dots in order to map morphologic and molecular heterogeneity in prostate cancer tissue5, providing proof of principle that morphology and molecular mapping is feasible, but falling short of quantifying the heterogeneity. Since immunohistochemistry is, at best, only semi-quantitative and subject to intra- and inter-observer bias, more sensitive and quantitative methodologies are required in order to accurately map and quantify tissue heterogeneity in situ.
We have developed and applied an experimental and statistical methodology in order to systematically quantify the heterogeneity of protein expression in whole tissue sections of tumors, based on the Automated QUantitative Analysis (AQUA) system6. Tissue sections are labeled with specific antibodies directed against cytokeratins and targets of interest, coupled to fluorophore-labeled secondary antibodies. Slides are imaged using a whole-slide fluorescence scanner. Images are subdivided into hundreds to thousands of tiles, and each tile is then assigned an AQUA score which is a measure of protein concentration within the epithelial (tumor) component of the tissue. Heatmaps are generated to represent tissue expression of the proteins and a heterogeneity score assigned, using a statistical measure of heterogeneity originally used in ecology, based on the Simpson's biodiversity index7.
To date there have been no attempts to systematically map and quantify this variability in tandem with protein expression, in histological preparations. Here, we illustrate the first use of the method applied to ER and HER2 biomarker expression in ovarian cancer. Using this method paves the way for analyzing heterogeneity as an independent variable in studies of biomarker expression in translational studies, in order to establish the significance of heterogeneity in prognosis and prediction of responses to therapy.

Here we describe a method to quantify molecular heterogeneity in histological sections of tumor material using quantitative immunofluorescence, image analysis, and a statistical measure of heterogeneity. The method is intended for use in clinical biomarker development and analysis.

प्रोटीन अभिव्यक्ति की मात्रात्मक immunofluorescence का उपयोग ट्यूमर में विविधता मानचित्रण

Morphologic heterogeneity within an individual tumor is well-recognized by histopathologists in surgical practice. While this often takes the form of ...

Label-free Isolation and Enrichment of Cells Through Contactless Dielectrophoresis

Dielectrophoresis (DEP) is the phenomenon by which polarized particles in a non-uniform electric field undergo translational motion, and can be used to direct the motion of microparticles in a surface marker-independent manner. Traditionally, DEP devices include planar metallic electrodes patterned in the sample channel. This approach can be expensive and requires a specialized cleanroom environment. Recently, a contact-free approach called contactless dielectrophoresis (cDEP) has been developed. This method utilizes the classic principle of DEP while avoiding direct contact between electrodes and sample by patterning fluidic electrodes and a sample channel from a single polydimethylsiloxane (PDMS) substrate, and has application as a rapid microfluidic strategy designed to sort and enrich microparticles. Unique to this method is that the electric field is generated via fluidic electrode channels containing a highly conductive fluid, which are separated from the sample channel by a thin insulating barrier. Because metal electrodes do not directly contact the sample, electrolysis, electrode delamination, and sample contamination are avoided. Additionally, this enables an inexpensive and simple fabrication process.
cDEP is thus well-suited for manipulating sensitive biological particles. The dielectrophoretic force acting upon the particles depends not only upon spatial gradients of the electric field generated by customizable design of the device geometry, but the intrinsic biophysical properties of the cell. As such, cDEP is a label-free technique that avoids depending upon surface-expressed molecular biomarkers that may be variably expressed within a population, while still allowing characterization, enrichment, and sorting of bioparticles.
Here, we demonstrate the basics of fabrication and experimentation using cDEP. We explain the simple preparation of a cDEP chip using soft lithography techniques. We discuss the experimental procedure for characterizing crossover frequency of a particle or cell, the frequency at which the dielectrophoretic force is zero. Finally, we demonstrate the use of this technique for sorting a mixture of ovarian cancer cells and fluorescing microspheres (beads).

Contactless dielectrophoresis (cDEP) achieves sorting and enrichment of particles via their intrinsic dielectric properties. Fluidic electrode channels replace metallic electrodes traditional to DEP, suiting cDEP to non-damaging sterile characterization and sorting of biological particles. We demonstrate how to prepare a cDEP microdevice and conduct cell characterization and sorting experiments.

Contactless dielectrophoresis के माध्यम से लेबल मुक्त अलगाव और कोशिकाओं के संवर्धन

Dielectrophoresis (DEP) is the phenomenon by which polarized particles in a non-uniform electric field undergo translational motion, and can be used to ...

MRI Mapping of Cerebrovascular Reactivity via Gas Inhalation Challenges

The brain is a spatially heterogeneous and temporally dynamic organ, with different regions requiring different amount of blood supply at different time. Therefore, the ability of the blood vessels to dilate or constrict, known as Cerebral-Vascular-Reactivity (CVR), represents an important domain of vascular function. An imaging marker representing this dynamic property will provide new information of cerebral vessels under normal and diseased conditions such as stroke, dementia, atherosclerosis, small vessel diseases, brain tumor, traumatic brain injury, and multiple sclerosis. In order to perform this type of measurement in humans, it is necessary to deliver a vasoactive stimulus such as CO2 and/or O2 gas mixture while quantitative brain magnetic resonance images (MRI) are being collected. In this work, we presented a MR compatible gas-delivery system and the associated protocol that allow the delivery of special gas mixtures (e.g., O2, CO2, N2, and their combinations) while the subject is lying inside the MRI scanner. This system is relatively simple, economical, and easy to use, and the experimental protocol allows accurate mapping of CVR in both healthy volunteers and patients with neurological disorders. This approach has the potential to be used in broad clinical applications and in better understanding of brain vascular pathophysiology. In the video, we demonstrate how to set up the system inside an MRI suite and how to perform a complete experiment on a human participant.

Non-invasive imaging of the brain vasculature&#8217;s ability to dilate or constrict may allow a better understanding of cerebrovascular pathophysiology in various neurological diseases. The present report describes a reproducible and patient-comfortable protocol to perform vascular reactivity imaging in humans using magnetic resonance imaging (MRI).

गैस साँस लेना चुनौतियां माध्यम Cerebrovascular जेट के एमआरआई मैपिंग

The brain is a spatially heterogeneous and temporally dynamic organ, with different regions requiring different amount of blood supply at different time. ...

Surgical Models of Gastroesophageal Reflux with Mice

Multiple surgical procedures have been reported to induce gastroesophageal reflux in animals. Herein, we report three surgical models with mice aiming to induce reflux of gastric contents, duodenal contents or mixed contents. Surgical procedures and general principles have been described in detail. A researcher with surgical experience should be able to grasp the technique after a short period of practice. After surgery, most mice can survive and develop reflux esophagitis similar to that in humans. However, it should be noted that histological differences between mouse and human esophagus are the inherent limitations of these surgical models. If used for research on Barrett&#8217;s esophagus and adenocarcinoma, these procedures may need to be combined with genetic modifications.

This article demonstrates surgical procedures of gastroesophageal reflux with mice. These models are useful tools for research on mechanisms and treatment of gastroesophageal reflux disease and potentially Barrett&#8217;s esophagus and esophageal adenocarcinoma.

चूहों के साथ Gastroesophageal भाटा के सर्जिकल मॉडल

Multiple surgical procedures have been reported to induce gastroesophageal reflux in animals. Herein, we report three surgical models with mice aiming to ...

Electrophysiological Assessment of Murine Atria with High-Resolution Optical Mapping

Recent genome-wide association studies targeting atrial fibrillation (AF) have indicated a strong association between the genotype and electrophysiological phenotype in the atria. That encourages us to utilize a genetically-engineered mouse model to elucidate the mechanism of AF. However, it is difficult to evaluate the electrophysiological properties in murine atria due to their small size. This protocol describes the electrophysiological evaluation of atria using an optical mapping system with a high temporal and spatial resolution in Langendorff perfused murine hearts. The optical mapping system is assembled with dual high-speed complementary metal oxide semiconductor cameras and high magnification objective lenses, to detect the fluorescence of a voltage-sensitive dye and Ca2+ indicator. To focus on the assessment of murine atria, optical mapping is performed with an area of 2 mm &#215; 2 mm or 10 mm x 10 mm, with a 100 &#215; 100 resolution (20 &#181;m/pixel or 100 &#181;m/pixel) and sampling rate of up to 10 kHz (0.1 ms) at maximum. A 1-French size quadripolar electrode pacing catheter is placed into the right atrium through the superior vena cava avoiding any mechanical damage to the atrium, and pacing stimulation is delivered through the catheter. An electrophysiological study is performed with programmed stimulation including constant pacing, burst pacing, and up to triple extrastimuli pacing. Under a spontaneous or pacing rhythm, the optical mapping recorded the action potential duration, activation map, conduction velocity, and Ca2+ transient individually in the right and left atria. In addition, the programmed stimulation also determines the inducibility of atrial tachyarrhythmias. Precise activation mapping is performed to identify the propagation of the excitation in the atrium during an induced atrial tachyarrhythmia. Optical mapping with a specialized setting enables a thorough electrophysiological evaluation of the atrium in murine pathological models.

This protocol describes the electrophysiological evaluation of murine atria utilizing an optical mapping system with a high temporal and spatial resolution, including dual recordings of the membrane voltage and Ca2+ transient under programmed stimulation through a specialized electrode catheter.

उच्च-रिज़ॉल्यूशन ऑप्टिकल मैपिंग के साथ Murine Atria का Electrophysiological आकलन

Recent genome-wide association studies targeting atrial fibrillation (AF) have indicated a strong association between the genotype and ...

Quantitative Mapping of Specific Ventilation in the Human Lung using Proton Magnetic Resonance Imaging and Oxygen as a Contrast Agent

Specific ventilation imaging (SVI) is a functional magnetic resonance imaging technique capable of quantifying specific ventilation &#8213; the ratio of the fresh gas entering a lung region divided by the region&#8217;s end-expiratory volume &#8213; in the human lung, using only inhaled oxygen as a contrast agent. Regional quantification of specific ventilation has the potential to help identify areas of pathologic lung function. Oxygen in solution in tissue shortens the tissue&#8217;s longitudinal relaxation time (T1), and thus a change in tissue oxygenation can be detected as a change in T1-weighted signal with an inversion recovery acquired image. Following an abrupt change between two concentrations of inspired oxygen, the rate at which lung tissue within a voxel equilibrates to a new steady-state reflects the rate at which resident gas is being replaced by inhaled gas. This rate is determined by specific ventilation. To elicit this sudden change in oxygenation, subjects alternately breathe 20-breath blocks of air (21% oxygen) and 100% oxygen while in the MRI scanner. A stepwise change in inspired oxygen fraction is achieved through use of a custom three-dimensional (3D)-printed flow bypass system with a manual switch during a short end-expiratory breath hold. To detect the corresponding change in T1, a global inversion pulse followed by a single shot fast spin echo sequence was used to acquire two-dimensional T1-weighted images in a 1.5&#160;T MRI scanner, using an eight-element torso coil. Both single slice and multi-slice imaging are possible, with slightly different imaging parameters. Quantification of specific ventilation is achieved by correlating the time-course of signal intensity for each lung voxel with a library of simulated responses to the air/oxygen stimulus. SVI estimations of specific ventilation heterogeneity have been validated against multiple breath washout and proved to accurately determine the heterogeneity of the specific ventilation distribution.

Specific ventilation imaging is a functional magnetic resonance imaging technique that allows for quantification of regional specific ventilation in the human lung, using inhaled oxygen as a contrast agent. Here, we present a protocol to collect and analyze specific ventilation imaging data.

एक विपरीत एजेंट के रूप में प्रोटोन चुंबकीय अनुनाद इमेजिंग और ऑक्सीजन का उपयोग कर मानव फेफड़ों में विशिष्ट वेंटिलेशन की मात्रात्मक मानचित्रण

Specific ventilation imaging (SVI) is a functional magnetic resonance imaging technique capable of quantifying specific ventilation &#8213; the ratio of ...

High-Throughput Analysis of Optical Mapping Data Using ElectroMap

Optical mapping is an established technique for high spatio-temporal resolution study of cardiac electrophysiology in multi-cellular preparations. Here we present, in a step-by-step guide, the use of ElectroMap for analysis, quantification, and mapping of high-resolution voltage and calcium datasets acquired by optical mapping. ElectroMap analysis options cover a wide variety of key electrophysiological parameters, and the graphical user interface allows straightforward modification of pre-processing and parameter definitions, making ElectroMap applicable to a wide range of experimental models. We show how built-in pacing frequency detection and signal segmentation allows high-throughput analysis of entire experimental recordings, acute responses, and single beat-to-beat variability. Additionally, ElectroMap incorporates automated multi-beat averaging to improve signal quality of noisy datasets, and here we demonstrate how this feature can help elucidate electrophysiological changes that might otherwise go undetected when using single beat analysis. Custom modules are included within the software for detailed investigation of conduction, single file analysis, and alternans, as demonstrated here. This software platform can be used to enable and accelerate the processing, analysis, and mapping of complex cardiac electrophysiology.

This protocol describes the setup and use of ElectroMap, a MATLAB-based open-source software platform for analysis of cardiac optical mapping data. ElectroMap provides a versatile high-throughput tool for analysis of optical mapping voltage and calcium datasets across a wide range of cardiac experimental models.

इलेक्ट्रोमैप का उपयोग करऑप्टिकल मानचित्रण डेटा के उच्च-Throughput विश्लेषण

Optical mapping is an established technique for high spatio-temporal resolution study of cardiac electrophysiology in multi-cellular preparations. Here we ...

Sarcomere Shortening of Pluripotent Stem Cell-Derived Cardiomyocytes using Fluorescent-Tagged Sarcomere Proteins.

Pluripotent stem cell-derived cardiomyocytes (PSC-CMs) can be produced from both embryonic and induced pluripotent stem (ES/iPS) cells. These cells provide promising sources for cardiac disease modeling. For cardiomyopathies, sarcomere shortening is one of the standard physiological assessments that are used with adult cardiomyocytes to examine their disease phenotypes. However, the available methods are not appropriate to assess the contractility of PSC-CMs, as these cells have underdeveloped sarcomeres that are invisible under phase-contrast microscopy. To address this issue and to perform sarcomere shortening with PSC-CMs, fluorescent-tagged sarcomere proteins and fluorescent live-imaging were used. Thin Z-lines and an M-line reside at both ends and the center of a sarcomere, respectively. Z-line proteins &#8212;&#160;&#945;-Actinin (ACTN2), Telethonin (TCAP), and actin-associated LIM protein (PDLIM3) &#8212; and one M-line protein &#8212; Myomesin-2 (Myom2) &#8212;&#160;were tagged with fluorescent proteins. These tagged proteins can be expressed from endogenous alleles as knock-ins or from adeno-associated viruses (AAVs). Here, we introduce the methods to differentiate mouse and human pluripotent stem cells to cardiomyocytes, to produce AAVs, and to perform and analyze live-imaging. We also describe the methods for producing polydimethylsiloxane (PDMS) stamps for a patterned culture of PSC-CMs, which facilitates the analysis of sarcomere shortening with fluorescent-tagged proteins. To assess sarcomere shortening, time-lapse images of the beating cells were recorded at a high framerate (50-100 frames per second) under electrical stimulation (0.5-1 Hz). To analyze sarcomere length over the course of cell contraction, the recorded time-lapse images were subjected to SarcOptiM, a plug-in for ImageJ/Fiji. Our strategy provides a simple platform for investigating cardiac disease phenotypes in PSC-CMs.

This method can be used to examine sarcomere shortening using pluripotent stem cell-derived cardiomyocytes with fluorescent-tagged sarcomere proteins.

सारकोमेरे फ्लोरोसेंट-टैग किए गए सारकोमेरे प्रोटीन का उपयोग करके प्लुरिपोटेंट स्टेम सेल-व्युत्पन्न कार्डियोमायोसाइट्स का छोटा होना।

Pluripotent stem cell-derived cardiomyocytes (PSC-CMs) can be produced from both embryonic and induced pluripotent stem (ES/iPS) cells. These cells ...