This work was supported by a National Imaging Facility Subsidised Access Grant to BJC, a National Health and Medical Research Council of Australia program grant (482800) to KAR and PJB. The authors would like to thank Andrew Arthur, Hasar Hazme and Marie-Claude Gregoire for support in developing this method.

Bu çalışmada açıklanan tüm işlemler Sidney Yerel Sağlık İlçesi ve Sydney Hayvan Etik Kurulların Üniversitesi tarafından onaylanmış ve laboratuvar hayvanlarının bakımı ve kullanımı, Sekizinci baskı (2011) için NIH Kılavuzu izlendi. 1. Hayvan hazırlanması NOT: Bu protokol Erkek db / db fareler (BKS.Cg- Dock7 m + / + Leprdb / J) 6 haftalık kadar grup mamasına erişim mahfaza ve su içinde muhafaza edilmiştir. görüntüleme zamanında, fareler, ~ 30 gramdı. Bu protokolde kullanılan bütün fareler, 10 ile 14 mmol / L arasında insülin seviyesini açlık vardı. <ol><li> Gerekirse, fareler oruç. Bu örnekte, deney prosedüründen önce, 5 saat boyunca fareler hızlı. </li><li> Görüntüleme başlamasından önce arzu edilen maddenin (örneğin, bir ilaç, bir protein, peptid) ile fareler tedavi edin. Bu örnekte, 3. insülin deri altına enjeksiyon (yönetmekU / kg insan insülini) veya eşdeğer bir hacim PBS, 30 dakika görüntüleme başlamasından önce. </li></ol> 2. kurma İş Akışı NOT: Bu protokol, bir PET / CT tarayıcı üzerinde gerçekleştirildi. BT veri alımını takiben ilk PET verileri elde edin. <ol><li> PET ayarları: <ol><li> 18F olarak seçin izotop, 3.600 s tarama süresini ayarlamak ve 350 keV üst ve alt düzey enerji ayrım - 650 keV (varsayılan) 3,432 ns (varsayılan) bir çakışma zamanlama penceresi. İzleyici, enjeksiyondan sonra 60 dakika - süresi 0 16 kare (6 x 10 s, 4 x 60 s, 1 x 300 s, 5 x 600 ler) içine histogram liste modu veri. Bir zoom 1.5 ile 2D-FBP kullanarak sinograms emisyonunu yeniden yapılandırma. Not: yeniden görüntü 128 x 128 x 159 vokseller ile 16 dinamik çerçeveler, her bir oluşuyordu ve 0.52 x 0.52 x 0.796 mm3 bir voksel boyutu. </li></ol></li><li> BT ayarları: <ol><li> İçinbir bütün vücut CT taraması, 50 kV, maruz kalma süresi 500 ms ve 360 ​​dönüş hareketi, 200, uzantılar en, 500 A gerilimi akımı ayarlayın. (Tam bir PET FoV aralığı kapsayacak) 3, 30722048 için yatak pozisyonların sayısı (FOV) detektör alanını alan yatak pozisyonları = 30,234713 ve% 4 detektör kümeleştirme arasındaki örtüşme. NOT: BT rekonstrüksiyonu HU kalibrasyonu, iki-doğrusal enterpolasyon ve Shepp-Logan filtresiyle koni ışın tomografi görüntü rekonstrüksiyon yazılımı kullanılarak yapıldı. </li></ol></li><li> 18F-FDG: <ol><li> Yeterli 18F-FDG için (örneğin 0.5 mL 450 MBq) yerel bir sağlayıcıdan, ilk enjeksiyondan önce ~ 30 dakika gelmesi. Hayvanlar alabilmesi için kısım ve 0.1 mL&#39;lik nihai bir hacimde ~ 18F-FDG 10 MBq 18F-FDG seyreltin. </li></ol></li></ol> 3. Görüntüleme Protokolü <ol><li> aseptik koşullar korumak için% 80 (h / h) etanol ile indüksiyon odası ve görüntüleme yatak silin. PlaBir indüksiyon odasında fare ce ve oksijen içinde% 5 izofluran ile anestezi. </li><li> 1 L / dakikalık bir akış oranında - (% 2 bakım, 1,5) izofluran sağlamak için vücut ısısını korumak için bir elektrikli ısıtma yastığı ve bir hassas buharlaştırıcı ucu koni takılmış bir görüntüleme yatağa fare yerleştirin. anestezi altında iken kuruluğunu önlemek için gözlerin üzerine oftalmik merhem uygulayın. </li><li> nefes izlemek ve anestezi yeterli düzlem korunmasını sağlamak için bir sensör pad üzerinde yüzükoyun pozisyonda fare yerleştirin. </li><li> lateral kuyruk damarı genişletmek için 2 dakika - 1 için bir ısı paketi kullanılarak kuyruk ısıtın. lateral kuyruk damarına bir 30-gauge iğne sokarak lateral kuyruk veni kateter. cerrahi yapıştırıcı ile yerinde iğne sabitleyin ve kateter sabitleyin. </li><li> Yük görüntüleme tarayıcıya yatak ve kateter makinenin arkasına erişilebilir, böylece makinenin içinden yatağa taşımak. </li><li> Bir Syr 18F-FDG şırıngaya kateter takıninge sürücüsü. Enjeksiyon ve tatbik edilecek hacim (&lt;100 uL, 10 s üzerine enjekte) önce şırıngada aktivitesine dayanarak tam 18F-FDG doz (10 MBq) hesaplayın. </li><li> Glukoz alımının değişkenliğine anestezi etkisini en aza indirmek için, anestezinin indüksiyonu ve (örneğin, 30 dakika) 18F-FDG enjeksiyon arasında sabit bir zaman sağlamak. </li><li> PET 18F-FDG enjeksiyonundan hemen önce tarama başlar. PET taraması (3.600 ler) bittikten sonra, dokular ile Radyoaktif alımının birlikte kaydedilmesi için izin vermek için bir CT taraması (~ 10 dakika) gerçekleştirmek. </li><li> , Başlangıç ​​pozisyonuna görüntüleme yatak taşı yatakta hayvan kaldırmak. </li><li> Bu noktada hayvan euthanize veya kurtarmak için izin: <ol><li> Ötenazi için, anestezi altında hala iken servikal dislokasyon gerçekleştirmek ve sonraki analiz için ilgi organları toplamak. </li><li> Fare kurtarmak için izin varsa, ısıtıcı bir ped üzerine tek bir gövde içinde fare koyun veyabir ısıtma lambası önünde. sternal yatma korumak için yeterli kendine geldi kadar fare izleyin. Fare grubu konut dönmeden önce 1 saat geri kazanılması için izin verin. </li></ol></li></ol> 4. PET görüntü işleme NOT: Görüntü rekonstrüksiyon araştırma işyeri yazılım v4.2 içinde edinim işyeri yazılım v1.5.0.28 ve analizi kullanılarak yapıldı. <ol><li> BT ve PET görüntüleri Co-kayıt ve hizalama 3-boyutlu doğru olduğundan emin olun. <ol><li> &#39;Dosya&#39; menüsünden, &#39;Klasör Arama / Al&#39; ı seçin ve verileri içeren klasörü seçin. İstenilen PET ve BT verileri seçin ve &#39;Genel Analiz&#39; sekmesine tıklayın. </li><li> Sıralama CR Kaynak &quot;olarak adlandırılır ve PET &#39;Hedefi&#39; olarak adlandırılır, böylece veriler. &#39;İş Akışı&#39; menüsünde, &#39;Kayıt&#39; seçin. görüntüler ayar gerektiriyorsa th araçları kullanmak doğru kayıtlı eş olmake &#39;Kayıt&#39; menüsü. </li></ol></li><li> &#39;İş Akışı&#39; menüsünde &#39;ROI miktarının belirlenmesi&#39; seçin. <ol><li> İstediğiniz bölgeyi bulmak için &#39;Resim&#39; sekmesinde &#39;Pan&#39; ve &#39;Zoom&#39; işlevlerini kullanın. &#39;Araçlar&#39; menüsünden &#39;Oluştur&#39; sekmesini seçin ve fırça simgesini tıklayın. Görüntünün YG&#39;sine çizin </li></ol></li><li> &#39;Kaydet&#39; menüsünden &#39;Kaydet ROI miktarının belirlenmesi&#39; seçerek zaman-aktivite eğrileri Özü. CSV dosyası olarak verileri kaydedin. </li><li> Doku cm3 başına Bq olarak radyoaktivite alımını ölçmek. CSV bir elektronik tabloya dosyasını yükleyerek cm3 (% ID / cm3) başına enjekte edilen dozun yüzdesi olarak değerleri dönüştürür. </li></ol> 5. Girdi Fonksiyonu <ol><li> Sistem nokta dağılım fonksiyonu için düzeltmek için, daha önce tarif edildiği gibi 9 reblurred Van Cittert ters evrişim yöntemi kullanılarak 5 iterasyon için tahmin edilen sistem PSF deconvolve. NOT: Bunun nedeni fare vena kava küçük boyutu için gereklidir. </li><li> Yukarıda tarif edildiği gibi, bir kan giriş fonksiyonu zaman-aktivite eğrisi oluşturmak için sonrası ters evrişim görüntüleri kullanın. </li></ol> 6. Kinetik modelleme Not: FDG, iki doku bölümünde modeli (Şekil 1), plazma giriş fonksiyonu gerektirir. <ol><li> CSV kan giriş fonksiyonu aşağıdaki denklem 10 kullanılarak plazma giriş fonksiyonuna dosya dönüştürme: Input_plasma = Input_blood x (0.386 E - 0.191t + 1.165). </li><li> kinetik modelleme aracında &#39;Kinetik&#39; butonuna tıklayın. Doku ve plazma toplam etkinlik &#39;Menü&#39; den &#39;Yükleme Süresi Etkinlik Curve&#39; seçerek kinetik modelleme aracı haline CSV dosyalarını saymak içe aktarın. </li><li> &#39;Model&#39; menüsünde 2 doku bölmeleri seçin. Bir sonraki k4 için kutusunu işaretlemeden olduğundan emin olunve ilk kez yerleştirilmesi için 0 değeri girmek vB için kutu (kan hacmi fraksiyonu) işaretini ve% 2 arasında bir değere girin. </li><li> &#39;Fit akım bölgesini tıklayın. Dispersiyon 11, 12 için ilgi ekstre bölge düzeltin. farklı dağılım kez FDG model ki-kare değeri en aza indirerek bu elde edin. </li><li> (K 1 -k 3) bölgesel oran sabitlerini hesaplamak için yüzen vB değeri (sonraki vB için kutuya onay kutusunu) ve optimize edilmiş dağılım değeri kullanılarak ikinci bir uyum gerçekleştirin. I = (k, 1 x k3) K gibi sabit bölge akını / (k 2 + k3) hesaplayın. </li></ol>

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The authors have nothing to disclose.

Burada açıklanan protokol farelerdeki kan dokusu içine akışına ve daha sonra metabolizmasından glukoz alımının kinetiklerini belirlemek için sağlam, invazif olmayan bir yöntem gösterir. Db / db fare insülin direnci ve ilgili müdahaleler incelemek için yaygın olarak kullanılmış olan, tip 2 diyabet 14 köklü bir hayvan modeli olduğu. Fakat daha önceki çalışmalarda sadece kalp 15 ve kalp ve iskelet kasında <sup class="x...

Bu çalışmanın amacı, bir pozitron emisyonu tomografisi / bilgisayarlı tomografi (PET / BT) farelerde spesifik dokular içine kan akımından glikoz, in vivo, gerçek zamanlı alımı ölçmek için temel yöntem geliştirmektir. Bu hücre içi alan, plazma, hücre içi boşluktan taşıma hızı ve oranı ile plazmadan 18F-FDG tutulumunun oranlarını tahmin etmek için glikoz alımını ve kinetik modelleme ölçmek için 18F-işaretli florodeoksiglükoz (FDG) kullanılarak elde edildi 18F-FDG fosforilasyonu. Kemirgenlerde, 18F-FDG çok sayıda kanser tedavileri 1, tümör ilerlemesi 2 ve tümör metabolizması 3 çalışmaları yanı sıra kahverengi yağ depoları 4, nöro-inflamasyonun 5 ve beyin metabolizmasının 6 görüntüleme klinik öncesi değerlendirme kullanılmıştır </&gt; Sup. Genellikle her doku 7 ötenazi, doku toplama ve radyoaktivite ölçümü ile takip edilen 3 H ya da 14 C ile 2-deoksiglukoz radyoaktif ile tedavi edilmesini fareler (ve farelerde) glikoz dokuya özgü alımını incelemek için kullanılan geleneksel yöntemler. PET / BT kullanımı canlı hayvanlarda aynı anda birden fazla organ ve bölgelerde glukoz alımı ve metabolizmanın noninvaziv belirlenmesi için izin verir. ötenazi bir gereklilik değildir Buna ek olarak, bu teknik, uzunlamasına çalışmalarda kullanım için uygundur. Tip 2 diabetes mellitus (T2DM) bozulmuş glikoz metabolizması ve insülin indirgenmiş doku yanıt sekonder hiperglisemi (insülin direnci) ve insülin 8 yeterli miktarda üretilmesi için pankreas-hücrelerinin yetersizlik ile karakterize edilir. glikoz alımının ve metabolizmanın Kinetik analiz içine önemli bilgiler sunabilireylem ve terapötik tedavi başarısının mekanizması olarak hastalığın ilerlemesi ileri izleme sağlar.

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			<td height="21" style="height:21px;width:216px;">PET/CT Scanner</td>
			<td style="width:152px;">Siemens</td>
			<td style="width:119px;">Inveon&#160;</td>
			<td style="width:167px;"></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:216px;">18F-FDG</td>
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			<td style="width:119px;"></td>
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			<td height="21" style="height:21px;width:216px;">Isoflurane</td>
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			<td height="21" style="height:21px;width:216px;">30 guage needle</td>
			<td style="width:152px;">BD</td>
			<td align="right" style="width:119px;">305106</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">PMOD modelling software</td>
			<td style="width:152px;">PMOD Technologies</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">BKS.Cg-Dock7m&#160;+/+&#160;Leprdb/J&#160; mice</td>
			<td style="width:152px;">Jackson Laboratory</td>
			<td style="width:119px;">000642</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Human insulin</td>
			<td style="width:152px;">Sigma-Aldrich</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

determining glucose metabolism kinetics using 18f-fdg micro-pet/ct

This paper describes the use of 18F-FDG and micro-PET/CT imaging to determine in vivo glucose metabolism kinetics in mice (and is transferable to rats). Impaired uptake and metabolism of glucose in multiple organ systems due to insulin resistance is a hallmark of type 2 diabetes. The ability of this technique to extract an image-derived input function from the vena cava using an iterative deconvolution method eliminates the requirement of the collection of arterial blood samples. Fitting of tissue and vena cava time activity curves to a two-tissue, three compartment model permits the estimation of kinetic micro-parameters related to the 18F-FDG uptake from the plasma to the intracellular space, the rate of transport from intracellular space to plasma and the rate of 18F-FDG phosphorylation. This methodology allows for multiple measures of glucose uptake and metabolism kinetics in the context of longitudinal studies and also provides insights into the efficacy of therapeutic interventions.

Daha önce glikoz alımının ve metabolizma 13 kinetikleri üzerinde plazma apoA-I düzeylerini arttırma etkisini araştırmak için, db / db fare modelini kullandık. Bu çalışmada gerçek zamanlı olarak gastroknemius kas içine plazmadan 18F-FDG alımını izlemek için, PET / BT görüntüleme yararını gösteren insülin ile muamele edilen db / db fareler kullanılmaktadır. Altı h...

Watch this Scientific Journal Video about Determining Glucose Metabolism Kinetics Using 18F-FDG Micro-PET/CT at JoVE.com

Determining Glucose Metabolism Kinetics Using 18F-FDG Micro-PET/CT

This study describes a protocol that uses 18F-FDG and positron emission tomography/computed tomography (PET/CT) imaging, together with kinetic modelling, to quantify the in vivo, real-time uptake of 18F-FDG into tissues.

Kullanımı Glukoz Metabolizma Kinetiği Belirlenmesi<sup&gt; 18</sup&gt; F-FDG Mikro PET / BT

This paper describes the use of 18F-FDG and micro-PET/CT imaging to determine in vivo glucose metabolism kinetics in mice (and is transferable to rats). ...

determining-glucose-metabolism-kinetics-using-18f-fdg-micro-petct

Research

JoVE Journal

Medicine

Determining Glucose Metabolism Kinetics Using 18F-FDG Micro-PET/CT

13.8K Görüntüleme.  UNSW Australia. This study describes a protocol that uses 18F-FDG and positron emission tomography/computed tomography (PET/CT) imaging, together with kinetic modelling, to quantify the in vivo, real-time uptake of 18F-FDG into tissues.

Video: Determining Glucose Metabolism Kinetics Using 18F-FDG Micro-PET/CT

Non-invasive Imaging of Acute Allograft Rejection after Rat Renal Transplantation Using 18F-FDG PET

The number of patients with end-stage renal disease, and the number of kidney allograft recipients continuously increases. Episodes of acute cellular allograft rejection (AR) are a negative prognostic factor for long-term allograft survival, and its timely diagnosis is crucial for allograft function 1. At present, AR can only be definitely diagnosed by core-needle biopsy, which, as an invasive method, bares significant risk of graft injury or even loss. Moreover, biopsies are not feasible in patients taking anticoagulant drugs and the limited sampling site of this technique may result in false negative results if the AR is focal or patchy. As a consequence, this gave rise to an ongoing search for new AR detection methods, which often has to be done in animals including the use of various transplantation models.
Since the early 60s rat renal transplantation is a well-established experimental method for the examination and analysis of AR 2. We herein present in addition small animal positron emission tomography (PET) using 18F-fluorodeoxyglucose (FDG) to assess AR in an allogeneic uninephrectomized rat renal transplantation model and propose graft FDG-PET imaging as a new option for a non-invasive, specific and early diagnosis of AR also for the human situation 3. Further, this method can be applied for follow-up to improve monitoring of transplant rejection 4.

Non-invasive Imaging of Acute Allograft Rejection after Rat Renal Transplantation Using 18F-FDG PET

We herein present a rat renal transplantation model to non-invasively assess acute allograft rejection using positron emission tomography with 18F-fluorodeoxyglucose.

Sıçan Böbrek Nakli sonra Akut Allogreft Reddi non-invaziv Görüntüleme Kullanımı<sup&gt; 18</sup&gt; F-FDG PET

The number of patients with end-stage renal  disease, and the number of kidney allograft recipients continuously increases.  Episodes of acute cellular ...

Tıp

Non-invasive Optical Measurement of Cerebral Metabolism and Hemodynamics in Infants

Perinatal brain injury remains a significant cause of infant mortality and morbidity, but there is not yet an effective bedside tool that can accurately screen for brain injury, monitor injury evolution, or assess response to therapy. The energy used by neurons is derived largely from tissue oxidative metabolism, and neural hyperactivity and cell death are reflected by corresponding changes in cerebral oxygen metabolism (CMRO2). Thus, measures of CMRO2 are reflective of neuronal viability and provide critical diagnostic information, making CMRO2 an ideal target for bedside measurement of brain health.
Brain-imaging techniques such as positron emission tomography (PET) and single-photon emission computed tomography (SPECT) yield measures of cerebral glucose and oxygen metabolism, but these techniques require the administration of radionucleotides, so they are used in only the most acute cases.
Continuous-wave near-infrared spectroscopy (CWNIRS) provides non-invasive and non-ionizing radiation measures of hemoglobin oxygen saturation (SO2) as a surrogate for cerebral oxygen consumption. However, SO2 is less than ideal as a surrogate for cerebral oxygen metabolism as it is influenced by both oxygen delivery and consumption. Furthermore, measurements of SO2 are not sensitive enough to detect brain injury hours after the insult 1,2, because oxygen consumption and delivery reach equilibrium after acute transients 3. We investigated the possibility of using more sophisticated NIRS optical methods to quantify cerebral oxygen metabolism at the bedside in healthy and brain-injured newborns. More specifically, we combined the frequency-domain NIRS (FDNIRS) measure of SO2 with the diffuse correlation spectroscopy (DCS) measure of blood flow index (CBFi) to yield an index of CMRO2 (CMRO2i) 4,5.
With the combined FDNIRS/DCS system we are able to quantify cerebral metabolism and hemodynamics. This represents an improvement over CWNIRS for detecting brain health, brain development, and response to therapy in neonates. Moreover, this method adheres to all neonatal intensive care unit (NICU) policies on infection control and institutional policies on laser safety. Future work will seek to integrate the two instruments to reduce acquisition time at the bedside and to implement real-time feedback on data quality to reduce the rate of data rejection.

We combined frequency-domain near-infrared spectroscopy measures of cerebral hemoglobin oxygenation with diffuse correlation spectroscopy measures of cerebral blood flow index to estimate an index of oxygen metabolism. We tested the utility of this measure as a bedside screening tool to evaluate the health and development of the newborn brain.

Bebeklerde Serebral Metabolizma ve Hemodinamiğin Noninvaziv Optik Ölçüm

Perinatal brain injury remains a significant cause of infant mortality  and morbidity, but there is not yet an effective bedside tool that can  accurately ...

Noninvasive Intratracheal Intubation to Study the Pathology and Physiology of Mouse Lung

The use of a model that mimics the condition of lung diseases in humans is critical for studying the pathophysiology and/or etiology of a particular disease and for developing therapeutic intervention. With the increasing availability of knockout and transgenic derivatives, together with a vast amount of genetic information, mice provide one of the best models to study the molecular mechanisms underlying the pathology and physiology of lung diseases. Inhalation, intranasal instillation, intratracheal instillation, and intratracheal intubation are the most widely used techniques by a number of investigators to administer materials of interest to mouse lungs. There are pros and cons for each technique depending on the goals of a study. Here a noninvasive intratracheal intubation method that can directly deliver exogenous materials to mouse lungs is presented. This technique was applied to administer bleomycin to mouse lungs as a model to study pulmonary fibrosis.

The use of a model that mimics the condition of lung diseases in humans is critical for studying the pathophysiology and/or etiology of a particular disease and for developing therapeutic intervention. Here a noninvasive intratracheal intubation method that can directly deliver exogenous materials to mouse lungs is presented.

Noninvaziv Entübasyon Fare Akciğer Patolojisi ve Fizyolojisi Eğitim için

The use of a model that mimics the condition of lung diseases in humans is critical for studying the pathophysiology and/or etiology of a particular ...

Non-invasive Imaging and Analysis of Cerebral Ischemia in Living Rats Using Positron Emission Tomography with 18F-FDG

Stroke is the third leading cause of death among Americans 65 years of age or older1. The quality of life for patients who suffer from a stroke fails to return to normal in a large majority of patients2, which is mainly due to current lack of clinical treatment for acute stroke. This necessitates understanding the physiological effects of cerebral ischemia on brain tissue over time and is a major area of active research. Towards this end, experimental progress has been made using rats as a preclinical model for stroke, particularly, using non-invasive methods such as 18F-fluorodeoxyglucose (FDG) coupled with Positron Emission Tomography (PET) imaging3,10,17. Here we present a strategy for inducing cerebral ischemia in rats by middle cerebral artery occlusion (MCAO) that mimics focal cerebral ischemia in humans, and imaging its effects over 24 hr using FDG-PET coupled with X-ray computed tomography (CT) with an Albira PET-CT instrument. A VOI template atlas was subsequently fused to the cerebral rat data to enable a unbiased analysis of the brain and its sub-regions4. In addition, a method for 3D visualization of the FDG-PET-CT time course is presented. In summary, we present a detailed protocol for initiating, quantifying, and visualizing an induced ischemic stroke event in a living Sprague-Dawley rat in three dimensions using FDG-PET.

Non-invasive Imaging and Analysis of Cerebral Ischemia in Living Rats Using Positron Emission Tomography with 18F-FDG

Brain damage resulting from cerebral ischemia may be non-invasively imaged and studied in rats using pre-clinical positron emission tomography coupled with the injectable radioactive probe, 18F-fluorodeoxyglucose. Further, the use of modern software tools that include volume of interest (VOI) brain templates dramatically increase the quantitative information gleaned from these studies.

Pozitron Emisyon Tomografi ile kullanma Non-invaziv Görüntüleme ve Serebral iskemi Analizi Yaşam Sıçanlarda<sup&gt; 18</sup&gt; F-FDG

Stroke is the third leading cause of death among Americans 65 years of age or older1. The quality of life for patients who suffer from a stroke fails to ...

Characterization of Metabolic Status in Nonhuman Primates with the Intravenous Glucose Tolerance Test

The intravenous glucose tolerance test (IVGTT) plays a key role in the characterization of glucose homeostasis. When taken together with serum biochemical profiles, inclusive of blood glucose levels in both the fed and fasted state, HbA1c, insulin levels, clinical history of diet, body composition, and body weight status, an assessment of normal and abnormal glycemic control can be made. Interpretation of an IVGTT is done through measurement of changes in glucose and insulin levels over time in relation to the dextrose challenge. Critical components to be considered are: peak glucose and insulin levels reached in relation to T0 (end of glucose infusion), the glucose clearance rate K derived from the slope of rapid glucose clearance in the first 20 min (T1 to T20), the time to return to glucose baseline, and the area under the curve (AUC). These IVGTT measures will show characteristic changes as glucose homeostasis moves from a healthy to a diseased metabolic state5. Herein we will describe the characterization of nonhuman primates (Rhesus and Cynomolgus macaques), which are the most relevant animal model of Type II diabetes (T2D) in humans and the IVGTT and clinical profiles of these animals from a lean healthy, to obese dysmetabolic, and T2D state 8, 10, 11.

The goal of this protocol is to present a standard method to perform intravenous glucose tolerance tests (IVGTTs) to assess glycemic control in nonhuman primates and assess their metabolic status from healthy to dysmetabolic.

İntravenöz Glikoz Tolerans Testi ile insan olmayan primatlarda Metabolik Durumu Karakterizasyonu

The intravenous glucose tolerance test (IVGTT) plays a key role in the characterization of glucose homeostasis. When taken together with serum biochemical ...

An Organotypic High Throughput System for Characterization of Drug Sensitivity of Primary Multiple Myeloma Cells

In this work we describe a novel approach that combines ex vivo drug sensitivity assays and digital image analysis to estimate chemosensitivity and heterogeneity of patient-derived multiple myeloma (MM) cells. This approach consists in seeding primary MM cells freshly extracted from bone marrow aspirates into microfluidic chambers implemented in multi-well plates, each consisting of a reconstruction of the bone marrow microenvironment, including extracellular matrix (collagen or basement membrane matrix) and stroma (patient-derived mesenchymal stem cells) or human-derived endothelial cells (HUVECs). The chambers are drugged with different agents and concentrations, and are imaged sequentially for 96 hr through bright field microscopy, in a motorized microscope equipped with a digital camera. Digital image analysis software detects live and dead cells from presence or absence of membrane motion, and generates curves of change in viability as a function of drug concentration and exposure time. We use a computational model to determine the parameters of chemosensitivity of the tumor population to each drug, as well as the number of sub-populations present as a measure of tumor heterogeneity. These patient-tailored models can then be used to simulate therapeutic regimens and estimate clinical response.

We describe a high-throughput drug sensitivity assay for primary multiple myeloma cells. It consists of a reconstruction of the bone marrow microenvironment (including extracellular matrix and stroma) in multi-well plates, and a non-invasive method for longitudinal quantification of cell viability.

Primer Multipl Miyelom Hücrelerinin İlaç Duyarlılık Karakterizasyonu için bir Organotipik Yüksek Verimli Sistemi

In this work we describe a novel approach that combines ex vivo drug sensitivity assays and digital image analysis to estimate chemosensitivity and ...

Study of In Vivo Glucose Metabolism in High-fat Diet-fed Mice Using Oral Glucose Tolerance Test (OGTT) and Insulin Tolerance Test (ITT)

Obesity represents the most important single risk factor in the pathogenesis of type 2 diabetes, a disease which is characterized by a resistance to insulin-stimulated glucose uptake and a gross decompensation of systemic glucose metabolism. Despite considerable progress in the understanding of glucose metabolism, the molecular mechanisms of its regulation in health and disease remain under-investigated, while novel approaches to prevent and treat diabetes are urgently needed. Diet derived glucose stimulates the pancreatic secretion of insulin, which serves as the principal regulator of cellular anabolic processes during the fed-state and thus balances blood glucose levels to maintain systemic energy status. Chronic overfeeding triggers meta-inflammation, which leads to alterations in peripheral insulin receptor-associated signaling and thus reduces the sensitivity to insulin-mediated glucose disposal. These events ultimately result in elevated fasting glucose and insulin levels as well as a reduction in glucose tolerance, which in turn serve as important indicators of insulin resistance. Here, we present a protocol for the generation and metabolic characterization of high-fat diet (HFD)-fed mice as a frequently used model of diet-induced insulin resistance. We illustrate in detail the oral glucose tolerance test (OGTT), which monitors the peripheral disposal of an orally administered glucose load and insulin secretion over time. Additionally, we present a protocol for the insulin tolerance test (ITT) to monitor whole-body insulin action. Together, these methods and their downstream applications represent powerful tools to characterize the general metabolic phenotype of mice as well as to specifically assess alterations in glucose metabolism. They may be especially useful in the broad research field of insulin resistance, diabetes and obesity to provide a better understanding of pathogenesis as well as to test the effects of therapeutic interventions.

Study of In Vivo Glucose Metabolism in High-fat Diet-fed Mice Using Oral Glucose Tolerance Test OGTT and Insulin Tolerance Test ITT

The current article describes the generation and metabolic characterization of high-fat diet-fed mice as a model of diet-induced insulin resistance and obesity. It further features detailed protocols to perform the oral glucose tolerance test and the insulin tolerance test, monitoring whole-body alterations of glucose metabolism in vivo.

Vivo glikoz metabolizması farelerde Oral glukoz tolerans testi (OGTT) ve insülin tolerans testi (ITT) kullanarak yüksek yağlı diyet beslenen incelenmesi

Obesity represents the most important single risk factor in the pathogenesis of type 2 diabetes, a disease which is characterized by a resistance to ...

Whole Body and Regional Quantification of Active Human Brown Adipose Tissue Using 18F-FDG PET/CT

In endothermic animals, brown adipose tissue (BAT) is activated to produce heat for defending body temperature in response to cold. BAT&#39;s ability to expend energy has made it a potential target for novel therapies to ameliorate obesity and associated metabolic disorders in humans. Though this tissue has been well studied in small animals, BAT&#39;s thermogenic capacity in humans remains largely unknown due to the difficulties of measuring its volume, activity, and distribution. Identifying and quantifying active human BAT is commonly performed using 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography and computed tomography (PET/CT) scans following cold-exposure or pharmacological activation. Here we describe a detailed image-analysis approach to quantify total-body human BAT from 18F-FDG PET/CT scans using an open-source software. We demonstrate the drawing of user-specified regions of interest to identify metabolically active adipose tissue while avoiding common non-BAT tissues, to measure BAT volume and activity, and to further characterize its anatomical distribution. Although this rigorous approach is time-consuming, we believe it will ultimately provide a foundation to develop future automated BAT quantification algorithms.

Whole Body and Regional Quantification of Active Human Brown Adipose Tissue Using 18F-FDG PET/CT

Using free, open-source software, we have developed an analytical approach to quantify total and regional brown adipose tissue (BAT) volume and metabolic activity of BAT using 18F-FDG PET/CT.

18F-FDG PET/BT kullanarak yağ dokusu kahverengi tüm vücut ve bölgesel Quantification etkin insan

In endothermic animals, brown adipose tissue (BAT) is activated to produce heat for defending body temperature in response to cold. BAT&#39;s ability to ...

Semi-quantitative Assessment Using [18F]FDG Tracer in Patients with Severe Brain Injury

Patients with severe traumatic brain injury (sTBI) have difficulty knowing whether they are accurately expressing their thoughts and emotions because of disorders of consciousness, disrupted higher brain function, and verbal disturbances. As a consequence of an insufficient ability to communicate, objective evaluations are needed from family members, medical staff, and caregivers. One such evaluation is the assessment of functioning brain areas. Recently, multimodal brain imaging has been used to explore the function of damaged brain areas. [18F]-fluorodeoxyglucose positron emission tomography-computed tomography ([18F]FDG-PET/CT) is a successful tool for examining brain function. However, the assessment of brain glucose metabolism based on [18F]FDG-PET/CT is not standardized and depends on several varying parameters, as well as the patient&#39;s condition. Here, we describe a series of semiquantitative assessment protocols for a region-of-interest (ROI) image analysis using self-produced [18F]FDG tracers in patients with sTBI. The protocol focuses on screening the participants, preparing the [18F]FDG tracer in the hot lab, scheduling the acquisition of [18F]FDG-PET/CT brain images, and measuring glucose metabolism using the ROI analysis from a targeted brain area.

Semi-quantitative Assessment Using [18F]FDG Tracer in Patients with Severe Brain Injury

[18F]-fluorodeoxyglucose (FDG) positron emission tomography-computed tomography is useful for studying glucose metabolism related to brain function. Here, we present a protocol for an [18F]FDG tracer set-up and semiquantitative assessment of the region-of-interest analysis for targeted brain areas associated with clinical manifestations in patients with severe traumatic brain injury.

Yarı kantitatif değerlendirmesi ağır beyin hasarı olan hastalarda [18F] FDG izleyici kullanarak

Patients with severe traumatic brain injury (sTBI) have difficulty knowing whether they are accurately expressing their thoughts and emotions because of ...

Visualization of Endogenous Mitophagy Complexes In Situ in Human Pancreatic Beta Cells Utilizing Proximity Ligation Assay

Mitophagy is an essential mitochondrial quality control pathway, which is crucial for pancreatic islet beta cell bioenergetics to fuel glucose-stimulated insulin release. Assessment of mitophagy is challenging and often requires genetic reporters or multiple complementary techniques not easily utilized in tissue samples, such as primary human pancreatic islets. Here we demonstrate a robust approach to visualize and quantify formation of key endogenous mitophagy complexes in primary human pancreatic islets. Utilizing the sensitive proximity ligation assay technique to detect interaction of the mitophagy regulators NRDP1 and USP8, we are able to specifically quantify formation of essential mitophagy complexes in situ. By coupling this approach to counterstaining for the transcription factor PDX1, we can quantify mitophagy complexes, and the factors that can impair mitophagy, specifically within beta cells. The methodology we describe overcomes the need for large quantities of cellular extracts required for other protein-protein interaction studies, such as immunoprecipitation (IP) or mass spectrometry, and is ideal for precious human islet&#160;samples generally not available in sufficient quantities for these approaches. Further, this methodology obviates the need for flow sorting techniques to purify beta cells from a heterogeneous islet population for downstream protein applications. Thus, we describe a valuable protocol for visualization of mitophagy highly compatible for use in heterogeneous and limited cell populations.

This protocol outlines a method for quantitative analysis of mitophagy protein complex formation specifically in beta cells from primary human islet samples. This technique thus allows analysis of mitophagy from limited biological material, which are crucial in precious human pancreatic beta cell samples.

Yakınlık ligasyon assay kullanarak ınsan pankreatik Beta hücrelerinde situ Içinde endojen Mitophagy kompleksleri görselleştirme

Mitophagy is an essential mitochondrial quality control pathway, which is crucial for pancreatic islet beta cell bioenergetics to fuel glucose-stimulated ...