The authors acknowledge all colleagues who made this work possible; Ann Demarr&#233;, Veerle Laurysens, Jan De Jonge and Erik Quartier for technical assistance. Financial support was from the VUB Research Council, the Institute for the Promotion of Innovation by Science and Technology in Flanders (IWT), the Beta Cell Biology Consortium (BCBC), the Fund for Scientific Research Flanders (FWO), Diabetes Onderzoek Nederland (DON) and Interuniversity Attraction Pole (IAP).

Tüm manipülasyonlar Deneysel ve diğer bilimsel Amaçlı kullanılan Omurgalı Hayvanların Korunmasına İlişkin Avrupa Sözleşmesi tarafından verilen yönergeleri izleyin (ETS 123 ve ve 2010/63 / AB). Çalışma Alanının 1. Hazırlanması <ol><li> Özel bir hazırlık alanı, cerrahi alanı ve kurtarma alanında sağlayın. </li><li> Çevre kirletici maddelerin en aza indirmek için, bir düzgün akış kabininde Cerrahi işlemin tamamı yürütün. </li><li> Hazırlık, cerrahi ve kurtarma alanında ihtiyaç duyulan (Gereç ve Yöntem listelendiği gibi) uygun aseptik teknik kullanılarak, sarf malzemeleri birleştirin. </li><li> Cerrahi aletler cerrahi öncesi otoklavlanırlar emin olun. </li><li> Ameliyat sırasında sıcaklık dengelenmesi için 38 ° C&#39;lik bir sıcaklıkta bir çevrimli su ısıtma pedi sağlar. Steril su geçirmez ped ile ısıtma yastığı örtün. </li><li> En azından 6.3X bir büyütme ile bir işletim mikroskop sağlayın. </li><li> Bir enstrüman Steri kullanınBöyle sıcak bir boncuk sterilizatör olarak lizer, cerrahi işlemler arasında aletleri sterilize etmek. </li><li> Düz kağıt yatak ile kaplı geniş bir kafes oluşan bir kurtarma alanı hazırlayın </li></ol> 2. Cerrahi Hayvanlar hazırlanması <ol><li> PDL ve sham için, 8 haftalık erkek fareler standart kafesler içine yerleştirilir ve muhafaza edilen, 12 saat ışık / 12 saat karanlık çevrimi ve standart kemirgen diyeti ile ad libitum kullanımı. Not: Burada, BALB / cJrJ fareleri kullanabilir ama biz de başarılı bir şekilde diğer suşları ve çeşitli transgenik suşları kullanılmıştır. </li><li> (- 0.1 mg / kg 0.05) ameliyat öncesi 30 dakika önleyici analjezi gibi Buprenophine kullanın. </li><li> 10 mg / ksilazin kg - ketamin 100 mg / kg ve 5 intraperitoneal enjeksiyonu ile farelere anestezi. </li><li> Istemli hareket ve kas gevşeme kademeli kaybı gözlemleyerek doğru anesthetization değerlendirin. Ayak sıkıştırarak reflekslerin kaybı test edin. </li><li> Önceki oftalmik merhem sürüngözlerin ent kuruluk anestezi altında iken. </li></ol> 3. Cerrahi Sitesi Hazırlama <ol><li> Antiseptik Klorheksidin çözümü ile toraks ve karın dezenfekte edin. </li><li> 2,5 cm karın x 1.5 cm lik bir alanı tıraş. </li><li> Daha sonra klorheksidin çözeltisi, alkol çözeltisi ve klorheksidin solüsyonunun, nihai uygulama ile ıslatılmış gazlı bez tıraş alanı dezenfekte. </li><li> Hazırlanan cerrahi alan yukarı cerrahı bakacak şekilde cerrahi alanda hayvan yerleştirin. </li><li> Vücudun geri kalanı kaplayan steril çalışma alanı oluşturmak için ise maruz dezenfekte karın bölgesini terk bir açıklığı olan bir su geçirmez cerrahi örtü kullanarak fare örtün. Anestezi derinliği prosedürü öncesinde fare izleyin. </li></ol> 4. Pankreas Kanal Ligasyonu <ol><li> Laparotomi <ol><li> Steril kullanarak göbek için kılıç şeklinde bir süreç uzanan deri bir üst orta hat kesi yapmakCerrahi bıçak. Altta yatan linea alba ve üst karın kadranı ortaya koymak için steril makas kullanılarak Periton ayırın. </li><li> Steril% 0.9 sodyum klorür ile düzenli yağmurlama ile iç organların kuruma-out önleyin. </li><li> Steril cımbız veya bez kullanarak, dalak ve pankreas dalak lob (kuyruk bölge) açığa superior mide geri çekin. </li><li> Yavaşça visseral periton ile örtülü pankreas baş, boyun ve gövde bölgesini ortaya çıkarmak için sağ üst kadranda üst jejunumun duodenum ve bir kısmını geri çekin. </li></ol></li><li> Ligasyonu <ol><li> Pankreasın boyun bölgesinde pankreas ana kanalının anatomik pozisyonunu bulun. </li><li> Visseral periton ve gastrokolik ligament, retroperitoneal erişim izni pankreas gövde ve kuyruk bölgesini açığa İnsizyon. Boyun bölgesini açığa çıkarmak amacıyla, bir Kocher manevrası gerçekleştirin. Ve duodenum ve baş kaldırınretroperitoneal kapalı pankreas inferior vena kava ve aşağıdaki aort onları yükseltme. </li><li> Dikkatle boyun bölgesine altına spatula yerleştirin. Gastro-duodenal ve dalak lobu ayıran portal ven sol tarafında 6-0 Prolen iplik ile pankreatik kanal Arter. </li><li> Loblar yeterince ayrılmasını temin ilk ligasyon yakın ikinci bir ligasyonu gerçekleştirin. </li><li> Altta yatan kan damarları, yani üstün pancreaticoduodenal arter, alt pancreaticoduodenal arter ve splenik arter pankreas kısmını zarar görmemesi için çok dikkatli Arter. </li><li> Karın boşluğuna geri organları yerleştirin. </li><li> Kas / periton katman için ve cilt için kesintili sütür desen sürekli dikiş desen 4-0 poliglikol ipliksi parçacığı kullanarak kesi kapatın. </li></ol></li></ol> 5. Sham Cerrahisi <ol><li> De tüm adımları uygulayın4.1.4 adımda 1 kayıtsız kalamayız. Pankreas maruz iken, pankreatik kanal ligasyonu yapmazlar. </li><li> Aşama 4.1.4 sonunda geri karın boşluğuna iç organları yerleştirin. </li><li> 4.2.7 açıklandığı gibi kesi kapatın. </li></ol> 6. Post-operasyonel Bakım ve İzleme <ol><li> Cerrahi işlem tamamlandıktan sonra, kurtarma alanında fareyi yerleştirin. Bu, normal vücut sıcaklığının muhafaza edilmesi amacıyla bir ısıtma pedi üzerine yerleştirilir ve düz kağıt yatak ile kaplı bir kafes içerir. </li><li> Kafes altına yerleştirilen nemlendirilmiş gıda post-operatif hipoglisemi önlemek için beslenme desteği sağlamak. Yumuşayana kadar bu amaçla, su ile ıslatılmış standart kemirgen diyeti granül kullanılır. </li><li> Nemlendirilmiş gıda sıvı desteği sağla ve su ad libitum sağlar. </li><li> Ameliyat sonrası 2 gün boyunca günde iki kez - (0.1 mg / kg 0.05) analjezi gibi Buprenophine kullanın. Tüm deneysel sırasında, Gözlem takibie yarasından veya davranış ve aktivite düzeyi damat azalma ile karakterize fiziksel bozulma, düşük iştah ve vücut ağırlığı kaybı sıvı veya irin salgılanması da dahil enfeksiyon olası belirtileri, oluşumu için fareler. </li></ol> Başarılı PDL ve PDL Tail Hasat ve Sham Tail Pankreas 7. Değerlendirme <ol><li> Servikal dislokasyon ile fareler Euthanize. </li><li> Kürk taşınmasını önlemek için karın bölgesini yeniden tıraş. </li><li> Karın deri ve kas tabakası açın ve pankreas iyi erişim elde etmek için deri ve kas geniş bir alanı kaldırmak. </li><li> Steril cımbız veya bez kullanarak, mide, dalak ve pankreas dalak lob (kuyruk bölge) açığa superior çekilir. </li><li> Yavaşça pankreas gastro-duodenal lob (baş bölgesi) ortaya çıkarmak için üst jejunumun duodenum ve bir kısmını dışarı çekin. Not: PDL pankreasın bağlandı bölümünün artık küçültülür ve neredeyse haline gelmiştirsaydam adacıklar küçük beyaz noktalar olarak görünür böylece. Pankreas baş kısmı opak pembe ve görülebilir lobuli farklı exocrine olduğunu. </li><li> Steril makas kullanarak, dalak boyunca keserek, dalak gelen pankreas kuyruk kısmını lige ayırmak. İç organlara pankreas kuyruk bölgesini bağlayan bağ dokusu gevşek kesin. </li><li> Bitişik harfleri ve hasat edilen dokudan buna hemen bitişik doku dışında, hemen ligasyon önünde PDL kuyruk bölgesini kesin. </li><li> Sahte kuyruk doku izole etmek için, adımları 7.5 ile 7.1 izleyin. Kullanımı Steril makas dalak boyunca keserek dalak sahte pankreas kuyruk bölgesini ayrı. İç organlara kuyruk pankreas bağlayan pankreas boyun bölgesi içine kesme ve bağ dokusu gevşek keserek kuyruk bölgesini izole edin Not:, sahte pankreas kuyruk ve baş bölgesi Hem opak pembe ayrı iki parçayı ayırmak için,boyun bölgesinde pankreas kesti. </li></ol>

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The authors have nothing to disclose.

In the present study, we describe in detail the methodology behind PDL, a mouse injury model to study beta cell neogenesis and proliferation and transdifferentiation of pancreatic non-beta cells. Ambiguity in data on PDL among labs stimulates the need for a standardized protocol for PDL surgery.
Critical steps in the PDL protocol include the selection of healthy, young mice for surgery. Preferably male mice should be used since unpublished data from our lab suggest that estrogen receptor signa...

300 milyondan fazla insanlara dünya çapında 1,2 etkileyen diyabet prevalansı artan, yetersiz beta hücre kitlesi doldurmak için, in vitro ve in vivo olarak, hem insülin üreten beta hücrelerinin yeni kaynaklar arama artırmıştır. 3 anahtar belirleme beta hücre çoğalması ve beta hücre neojenezi, örneğin, non-beta hücre veya projenitör hücresi beta hücrelerinin farklılaşmasını düzenleyen mekanizma ve faktörleri, diyabetlerde rejeneratif tedavilerin geliştirilmesi için yeni hedefler sağlayabilir. Gelişmekte olan kemirgen pankreas olarak, endokrin hücre tipleri transkripsiyon faktörü Neurogenin3 (Ngn3) ifade eden, endokrin progenitör hücrelerin geçici popülasyonundan ayırt. 4,5 yetişkin kemirgen pankreas olarak, normal fizyolojik koşullar altında, beta hücre kütlesi optimal sayıda tutulan metabolik ihtiyaçlarını karşılamak için. Beta hücre boyutundaki değişiklikler,beta hücrelerinin potansiyel normal fizyolojik koşullar altında prolifere iken apoptoz ve çoğaltma nüfusun genelinde homojen beta hücre genişlemesi ve dönüş bitti. 6-8 başlıca mekanizmalar oluşturan, 9 onların çoğalması oranı düşüktür ve yeniden çoğaltma sınırlandırıldı yaş ve glukoz metabolizması etkilenir dinamik bir sessizlik dönemi veya refrakter dönem 6,7. 10 endokrin progenitör hücreler kadar normal yetişkin pankreas tespit edilmemiştir beri, neojenezini normal erişkin beta hücre büyümesine katkıda bulunduğu düşünülmektedir. 8 Bu nedenle, genişletilebilir ve bir roman, beta hücrelerinin muhtemelen sınırsız bir kaynak sağlayacak yeni beta hücrelerini verimli yeteneğine sahip yetişkin pankreasta bir fakültatif endokrin progenitör hücre tanımlaması. Kısmi kanalı ligasyonu (PDL) beta hücre neogenesi indüklediği tarif edilmiş bir hayvan yaralanması modeliYetişkin pankreasın s. Bu modelde 11,12, pankreas kuyruk boşaltma ana pankreatik kanal cerrahi bağlanır. ekzokrin drenaj elde edilen tıkanma inflamatuvar ortamında enflamasyon ve ligasyon distal asinar atrofi. 12-14 ile birlikte ana dokuların yeniden indükler, Ngn3 indüklenir endokrin progenitör işaret ve beta hücre hacmi yeniden ekspresyonu, iki kat artar . Embriyonik tip endokrin progenitör hücre ve &quot;sessizlik dönemi&quot; olmadan-çoğaltılması yeniden eğilimli önceden varolan ve yeni kurulan beta hücrelerinin çoğalmasından ifade eden Ngn3 yeni beta hücrelerinin kuşağa beta hücre hacmi sonuçlarında bu iki katına. 11,15 Bu tür pankreatektomi 6,7,16-19 ve beta hücrelerinin 20 seçici ablasyonu gibi yaralanma modellerinde, beta hücre neojenezini ve çoğaltma yoğun tarif edilmiştir. Thes içinde Ancak, rejeneratif sonuçE modelleri hasarın ölçüde etkilenir ve azalmış ilk beta hücre kütlesi 21 ile ilişkilidir. PDL başlangıç ​​beta hücre kütlesi etkilenmez ve beta hücre neojenezini ve proliferasyon sağlam aktive olan bir cerrahi bir modeldir. Gerçekten de, PDL uygulanan farelerin pankreasında, Ngn3 ifade eden hücreler, kanalın epitelial yakınında tespit edilir. Bu hücreler, THP (FACS) kullanılarak Ngn3-GFP transgenik farelerin lige pankreas izole edilir ve E12.5 Ngn3 pankreas ve ex vivo kültür içinde bir nakil aşağıdaki işlevsel p hücrelere karşı ayırt etmek mümkün olabilir - / - . farenin 11 Benzer şekilde, Ngn3 CreERT olarak;. Ngn3 genini aktive hücreler sürekli olarak tamoksifen, enjeksiyon, etiket-pozitif Ngn3 hücresinden türetilmiş beta hücreleri PDL sonra tespit sonra etiketlenir ki burada R26 YFP fareler 15 Ayrıca, yeni oluşan beta hücreleri, önceden seyreltilmiş -existHedef spesifik kısa saç tokası RNA anlamlı beta hücre kitlesi ve beta hücre çoğalmasını sonra azalır kullanarak Ngn3 ifadesini azalmış beri beta hücrelerini ing ve tercihen beta hücrelerinin yüksek çoğalma potansiyeli gösteren, içinde küçük adacıklar bulun. 15 Ngn3 PDL sonra beta hücre genişlemesi için önemli olan PDL. 11 Özellikle, PDL sonra Ngn3 hücresinden türetilmiş beta hücrelerinin fraksiyonu ve beta hücre kütlesi kritik Ngn3 indüksiyonu 15 seviyesine bağlıdır. R26 iDTR fareler;. Bu, Ngn3 Ngn3 CreERT difteri toksin verilmesi ile ifade eden hücrelerin seçici şekilde kesip çıkarılması Ngn3 yüksek ifade seviyesi, pankreas gelişimi sırasında multipotent pankreatik progenitörlerinden endokrin bağlılık için kritik bir adım olduğu gözlemiyle uyumludur ek olarak 22 Özellikle küçük adacıkları azalma insülin içeriğine ve düşük beta hücre çoğalması ile sonuçlanır. 15 <pPDL sonra kanal hücrelerinde Ngn3 ifade indüksiyon birçok 11,15,16,23,24 tarafından onaylandıktan rağmen class = &quot;jove_content&quot;&gt;, PDL sonucuna ilişkin adacıklar hücreleri 24,25 ve tutarsızlıklar içinde Ngn3 ifadesi bizim ilk gözlemler meydan artan beta hücre kitlesi 26,27 arasında, Ngn3 görünümü PDL sonra kanal kaynaklı endokrin atalarıdır 24,26,28,29 ve artan beta hücre çoğalmasını 27 dile getirdi. 30 Bunlar çelişkili sonuçlar, en azından kısmen, en önemlisi fareler, post-operatif fizyolojik ve çevresel koşullar ve analizi, vücut ağırlığı, cinsiyet ve yaş ameliyat sonrası zaman noktalarında varyasyonlar dahil faktörlerin bir kombinasyonu isnat edilebilir Cerrahi teknik farklılıklar. Bizim ellerde 30 beta hücre çoğalması, insülin içeriği, beta hücre hacmi ve küçük adacıklar sayısı sürekli PDL sonra artmıştır. AyrıcaNgn3 mRNA sürekli artar ama biz doğrudan açıklama var olan PDL kuyruk pankreaslarından arasında Ngn3 mRNA ekspresyonu büyük farklılıklar vardır. Biz Ngn3 mRNA seviyesi, beta hücrelerinden 15 gelen beta hücre neojenezi derecesi ile ilişkili olabilir hipotezi, ama, bu da ispat gerektirir. Tüm deneysel varyasyonları kaldırmak olmamasına rağmen, PDL ameliyatı gerçekleştirmek için standart bir yöntem sonuçlarında daha iyi tekdüzelik sağlar ve beta hücre çoğalmasını ve neojenezini okuyan yeni yollar açar.

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surgical injury to the mouse pancreas through ligation of the pancreatic duct as a model for endocrine and exocrine reprogramming and proliferation

Bir embriyo veya yetişkin kök hücre farklılaşması ile in vivo veya ex vivo veya beta hücrelerinin üretimi pankreatik beta hücrelerinin genişlemesi, diyabet için bir tedavi olarak, insan doku adacığı transplantasyonundan verici kıtlığı hafifletmek için beta hücrelerinin yeni bir genleşebilen kaynaklar sağlar. Son gelişmeler bu amaca yönelik yapılmış olmasına rağmen, bir kök / progenitör hücreden beta hücre genişlemesi ve farklılaşmasını düzenleyen mekanizmalar karakterize edilmesi gerekmektedir. Burada, doku tekrar modellemesinde ve beta hücre proliferasyonu ve farklılaşmasının mekanizmaları incelemek için bir araç olarak işlev görebilir yetişkin fare pankreası bir yaralanma modeli için bir protokol açıklar. Kısmi kanalı ligasyonu (PDL) pankreas kuyruk bölgesi dışına ekzokrin ürünlerinin drenaj tıkanma sonucu ana pankreatik kanal cerrahi ligasyon içeren kemirgen pankreasın bir deneysel yaralanma olduğunu. tahribatını asiner atrofi, bağışıklık hücre infilt uyarmaktadırrasyon ve şiddetli doku yeniden. Biz daha önce endojen progenitör benzeri hücreler ve PDL sonra beta hücre çoğalmasında bir artış ifade Neurogenin (NGN) 3 aktivasyonunu bildirmiştir. Bu nedenle, PDL beta hücre dinamikleri katılan sinyalleri ve yetişkin pankreas endokrin progenitör özelliklerini incelemek için bir temel sağlar. , Hala faktörler ve yollar PDL beta hücre neojenezi ve çoğalmasına katkıda bulunan, büyük ölçüde belirsizdir yana, PDL için standart protokol laboratuvarlar arasında karşılaştırma için izin verecektir.

PDL asiner atrofi ve inflamasyon indükler ama vücut ağırlığı ve glisemiyi etkilemez Mide ve duodenum bitişiğinde bulunan organın başkanı etkilenmemiş kalırken 8 haftalık erkek BALB / c farelerde, pankreas kuyruk ekzokrin enzimleri drenaj kanalı bağlanır. Yaş, cinsiyet ve kilo eşleştirilmiş erkek BALB / c fareler pankreatik kanal ligasyonu dışında kısmi kanal ligasyonu cerrahisi tüm adımları yansıtan sahte cerrahi, geçmesi. Pankreas dokusu 14, 3...

Watch this Scientific Journal Video about Surgical Injury to the Mouse Pancreas through Ligation of the Pancreatic Duct as a Model for Endocrine and Exocrine Reprogramming and Proliferation at JoVE.co

Surgical Injury to the Mouse Pancreas through Ligation of the Pancreatic Duct as a Model for Endocrine and Exocrine Reprogramming and Proliferation

This protocol describes an injury model involving the surgical ligation of the pancreatic duct in the adult mouse pancreas, resulting in severe injury that establishes an environment that allows beta cell neogenesis and proliferation. This model can be used as a tool to study mechanisms involved in beta cell formation.

Endokrin ve Ekzokrin yeniden programlanması ve Silahların Yayılmasını Önleme Modeli Olarak Pankreas kanalının bağlanmasını yoluyla Fare Pankreas Cerrahi Sakatlık

Expansion of pancreatic beta cells in vivo or ex vivo, or generation of beta cells by differentiation from an embryonic or adult stem cell, can provide ...

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Research

JoVE Journal

Medicine

16.0K Görüntüleme.  Vrije Universiteit Brussel. This protocol describes an injury model involving the surgical ligation of the pancreatic duct in the adult mouse pancreas, resulting in severe injury that establishes an environment that allows beta cell neogenesis and proliferation. This model can be used as a tool to study mechanisms involved in beta cell formation. 

Video: Surgical Injury to the Mouse Pancreas through Ligation of the Pancreatic Duct as a Model for Endocrine and Exocrine Reprogramming and Proliferation

A Murine Model of Myocardial Ischemia-reperfusion Injury through Ligation of the Left Anterior Descending Artery

Acute or chronic myocardial infarction (MI) are cardiovascular events resulting in high morbidity and mortality. Establishing the pathological mechanisms at work during MI and developing effective therapeutic approaches requires methodology to reproducibly simulate the clinical incidence and reflect the pathophysiological changes associated with MI. Here, we describe a surgical method to induce MI in mouse models that can be used for short-term ischemia-reperfusion (I/R) injury as well as permanent ligation. The major advantage of this method is to facilitate location of the left anterior descending artery (LAD) to allow for accurate ligation of this artery to induce ischemia in the left ventricle of the mouse heart. Accurate positioning of the ligature on the LAD increases reproducibility of infarct size and thus produces more reliable results. Greater precision in placement of the ligature will improve the standard surgical approaches to simulate MI in mice, thus reducing the number of experimental animals necessary for statistically relevant studies and improving our understanding of the mechanisms producing cardiac dysfunction following MI. This mouse model of MI is also useful for the preclinical testing of treatments targeting myocardial damage following MI.

We introduce a surgical method to induce experimental ischemia/reperfusion (I/R) injury to simulate myocardial infarction (MI) in mouse models that allows for more clarity in positioning of the ligation on the left anterior descending artery (LAD) to increase the reproducibility of MI experiments in mice.

Sol ön inen arter ligasyonu ile Miyokard İskemi-reperfüzyon Hasarı bir fare modeli

Acute or chronic myocardial infarction (MI) are cardiovascular events resulting in high morbidity and mortality. Establishing the pathological mechanisms ...

Tıp

Bile Duct Ligation in Mice: Induction of Inflammatory Liver Injury and Fibrosis by Obstructive Cholestasis

In most vertebrates, the liver produces bile that is necessary to emulsify absorbed fats and enable the digestion of lipids in the small intestine as well as to excrete bilirubin and other metabolic products. In the liver, the experimental obstruction of the extrahepatic biliary system initiates a complex cascade of pathological events that leads to cholestasis and inflammation resulting in a strong fibrotic reaction originating from the periportal fields. Therefore, surgical ligation of the common bile duct has become the most commonly used model to induce obstructive cholestatic injury in rodents and to study the molecular and cellular events that underlie these pathophysiological mechanisms induced by inappropriate bile flow. In recent years, different surgical techniques have been described that either allow reconnection or reanastomosis after bile duct ligation (BDL), e.g., partial BDL, or other microsurgical methods for specific research questions. However, the most frequently used model is the complete obstruction of the common bile duct that induces a strong fibrotic response after 21 to 28 days. The mortality rate can be high due to infectious complications or technical inaccuracies. Here we provide a detailed surgical procedure for the BDL model in mice that induce a highly reproducible fibrotic response in accordance to the 3R rule for animal welfare postulated by Russel and Burch in 1959.

Disruption of bile flow results in severe inflammatory cholestatic liver injury with a characteristic time-dependent sequence of morphological alterations. Here we present a protocol for the surgical ligation of the common bile duct in mice that allows to induce a strong fibrotic response after 21 to 28 days.

Farelerde Safra Kanalı ligasyonu: Obstrüktif Kolestaz tarafından İnflamatuar Karaciğer Hasarı ve Fibrozis indüksiyonu

In most vertebrates, the liver produces bile that is necessary to emulsify absorbed fats and enable the digestion of lipids in the small intestine as well ...

Pancreatic Duct Infusion: An Effective and Selective Method of Drug and Viral Delivery

The pancreas is a bifunctional organ with both endocrine and exocrine components. A number of pathologies can afflict the pancreas, including diabetes, pancreatitis, and pancreatic cancer. All three of these diseases mark active areas of study, not only to develop immediate therapy, but also to better understand their pathophysiology. There are few tools to further these areas of study. Pancreatic duct infusion is an important technique that can allow for lineage tracing, gene introduction, and cell line-specific targeting. The technique requires the intricate dissection of the second portion of the duodenum and ampulla, followed by the occlusion of the bile duct and the cannulation of the pancreatic duct. Although the technique is technically challenging at first, the applications are myriad. Ambiguity in the specifics of the procedure between groups highlighted the need for a standard protocol. This work describes the expression of a green fluorescent protein (GFP) within the pancreas after the pancreatic duct infusion of a viral vector expressing GFP versus a sham surgery. The infusion and therefore expression is specific to the pancreas, without expression present in any other tissue type.

Pancreatic duct infusion is an important technique that can allow for lineage tracing, gene introduction, and cell line-specific targeting. A pancreatic duct infusion technique for drug and viral delivery to pancreatic cells is presented here.

Pankreas Kanalı infüzyonu: İlaç ve Viral Doğumda Etkili ve Seçici Bir Yöntem

The pancreas is a bifunctional organ with both endocrine and exocrine components. A number of pathologies can afflict the pancreas, including diabetes, ...

Dissection of the Mouse Pancreas for Histological Analysis and Metabolic Profiling

We have been investigating the pancreas specific transcription factor, 1a cre-recombinase; lox-stop-lox- Kristen rat sarcoma, glycine to aspartic acid at the 12 codon (Ptf1acre/+;LSL-KrasG12D/+) mouse strain as a model of human pancreatic cancer. The goal of our current studies is to identify novel metabolic biomarkers of pancreatic cancer progression. We have performed metabolic profiling of urine, feces, blood, and pancreas tissue extracts, as well as histological analyses of the pancreas to stage the cancer progression. The mouse pancreas is not a well-defined solid organ like in humans, but rather is a diffusely distributed soft tissue that is not easily identified by individuals unfamiliar with mouse internal anatomy or by individuals that have little or no experience performing mouse organ dissections. The purpose of this article is to provide a detailed step-wise visual demonstration to guide novices in the removal of the mouse pancreas by dissection. This article should be especially valuable to students and investigators new to research that requires harvesting of the mouse pancreas by dissection for metabolic profiling or histological analyses.

This video article provides a detailed demonstration of the procedures required to successfully remove the pancreas from a mouse by dissection for histological analysis and metabolic profiling.

Fare pankreas diseksiyon histolojik analiz ve metabolik profil oluşturma

We have been investigating the pancreas specific transcription factor, 1a cre-recombinase; lox-stop-lox- Kristen rat sarcoma, glycine to aspartic acid at ...

Robotic Lateral Pancreaticojejunostomy for Chronic Pancreatitis

Lateral pancreaticojejunostomy (LPJ) has shown good postoperative outcomes in patients with painful, morphine dependent, chronic pancreatitis (CP). The recent rise of robotic and laparoscopic pancreatic surgery has found benefits such as reduced time to functional recovery. Few studies have reported on the feasibility, technique and outcome of robotic LPJ, especially including transection of the gastroduodenal artery. The present study describes the main steps for robotic LPJ in a patient with painful chronic pancreatitis with a dilated main pancreatic duct. The patient underwent robotic LPJ. The LPJ anastomosis is performed using a running suture technique in a longitudinal side-to-side manner. Routinely, the gastroduodenal artery is transected to drain the entire length of the main pancreatic duct. The patient is in French position; 7 trocars are placed (4 robotic, 2 laparoscopic assistants, 1 liver-retractor). After docking of the robot system, the omental bursa is opened, and the right gastroepiploic artery and vein are ligated at their base at the lower border of the pancreas. Intraoperative ultrasonography is performed in order to determine trajectory of the dilated main pancreatic duct which is opened for its entire length after the gastroduodenal artery has been suture ligated both cranially and caudally from the main pancreatic duct. A Roux limb is created, and a latero-lateral PJ is fashioned using several 3-0 barbed sutures. A stapled jejuno-jejunostomy is created at sufficient distance from the pancreatic anastomosis, aided by a 50 cm suture. The described technique for robotic LPJ is a complex but feasible operation for patients with treatment refractory CP and a dilated main pancreatic duct. Due to its complexity, implementation in high volume centers with extensive experience with CP surgery may improve outcomes.

Robotic lateral pancreaticojejunostomy (RLPJ) may be used in patients with painful, morphine dependent, chronic pancreatitis and a dilated main pancreatic duct. We describe a standardized and reproducible technique for RLPJ, which includes transection of the gastroduodenal artery.

Kronik Pankreatit için Robotik Lateral Pankreaticojejunostomi

Lateral pancreaticojejunostomy (LPJ) has shown good postoperative outcomes in patients with painful, morphine dependent, chronic pancreatitis (CP). The ...

Laparoscopic Radical Left Pancreatectomy for Pancreatic Cancer: Surgical Strategy and Technique Video

Radical resection margins, resection of Gerota&#8217;s (perirenal) fascia, and adequate lymph node dissection are crucial for an adequate oncological resection of left-sided pancreatic cancer. Several surgical techniques have been described in recent years, but few were specifically designed for minimally invasive approaches. This study describes and demonstrates a standardized and reproducible technique for an adequate oncological resection of pancreatic cancer: laparoscopic radical left pancreatectomy (LRLP).
A 61-year-old woman presented with an incidental finding of a 3 cm mass in the left pancreas suspect for malignancy. Imaging did not reveal distant metastases, central vascular involvement, or morbid obesity, hence the patient was suitable for LRLP. This study describes the main steps of LRLP for pancreatic cancer. First, the lesser sac is opened by transecting the gastrocolic ligament. The splenic flexure of the colon is mobilized and the inferior border of the pancreas including Gerota&#39;s fascia is dissected down to the inferior border of the spleen. The pancreas is tunneled and hung, including Gerota&#8217;s fascia with a vessel loop. At the pancreatic neck, a tunnel is created between the pancreas and the portal vein, likewise a vessel loop is passed. The pancreas is then transected using the graded compression technique with an endostapler. Both the splenic vein and artery are transected before completing the resection. The entire specimen is extracted in a retrieval bag via a small Pfannenstiel incision.
Duration of the surgery was 210 min with 250 mL blood loss. Pathology revealed a R0-resection (&#62;1 mm) of a well-to-moderately differentiated adenocarcinoma originating from an intraductal papillary mucinous neoplasm. A total of 15 tumor-negative lymph nodes were resected. This is a detailed description of LRLP for left-sided pancreatic cancer as is currently being used within the international, multicenter randomized DIPLOMA (Distal Pancreatectomy Minimally Invasive or Open for PDAC) trial.

Oncologically safe left pancreatectomy requires radical resection (R0), Gerota&#8217;s (perirenal) fascia resection, and adequate lymph node dissection. This study describes the technical details of laparoscopic radical left pancreatectomy (LRLP), used in the first international multicenter randomized trial comparing minimally invasive with open left pancreatectomy for pancreatic cancer, the DIPLOMA trial.

Pankreas Kanseri için Laparoskopik Radikal Sol Pankretomi: Cerrahi Strateji ve Teknik Video

Radical resection margins, resection of Gerota&#8217;s (perirenal) fascia, and adequate lymph node dissection are crucial for an adequate oncological ...

A Rat Lung Transplantation Model of Warm Ischemia/Reperfusion Injury: Optimizations to Improve Outcomes

From our experience with rat lung transplantation, we have found several areas for improvement. Information in the existing literature regarding methods for choosing appropriate cuff sizes for the pulmonary vein (PV), pulmonary artery (PA), or bronchus (Br) are varied, thus making the determination of proper cuff size during rat lung transplantation an exercise of trial and error. By standardizing the cuffing technique to use the smallest effective cuff appropriate for the size of the vessel or bronchus, one can make the transplantation procedure safer, faster, and more successful. Since diameters of the PV, PA, and Br are related to the body weight of the rat, we present a strategy to choosing an appropriate size using a weight-based guide. Since lung volume is also related to body weight, we recommend that this relationship should also be considered when choosing the proper volume of air for donor lung inflation during warm ischemia as well as for the proper volume of PBS to be instilled during bronchoalveolar lavage (BAL) fluid collection. We also describe methods for 4th intercostal space dissection, wound closure, and sample collection from both the native and transplanted lobes.

Here, we present optimizations to a rat lung transplantation model that serve to improve outcomes. We provide a size guide for cuffs based on body weight, a measurement strategy to ascertain the 4th intercostal space, and methods of wound closure and BAL (bronchoalveolar lavage) fluid and tissue collection.

Sıcak iskemi / reperfüzyon hasarının bir sıçan akciğer transplantasyon modeli: sonuçları iyileştirmek için optimizasyonlar

From our experience with rat lung transplantation, we have found several areas for improvement. Information in the existing literature regarding methods ...

Stabilized Longitudinal In Vivo Cellular-Level Visualization of the Pancreas in a Murine Model with a Pancreatic Intravital Imaging Window

Direct in vivo cellular-resolution imaging of the pancreas in a live small animal model has been technically challenging. A recent intravital imaging study, with an abdominal imaging window, enabled visualization of the cellular dynamics in abdominal organs in vivo. However, due to the soft sheet-like architecture of the mouse pancreas that can be easily influenced by physiologic movement (e.g., peristalsis and respiration), it was difficult to perform stabilized longitudinal in vivo imaging over several weeks at the cellular level to identify, track, and quantify islets or cancer cells in the mouse pancreas. Herein, we describe a method for implanting a novel supporting base, an integrated pancreatic intravital imaging window, that can spatially separate the pancreas from the bowel for longitudinal time-lapse intravital imaging of the pancreas microstructure. Longitudinal in vivo imaging with the imaging window enables stable visualization, allowing for the tracking of islets over a period of 3 weeks and high-resolution three-dimensional imaging of the microstructure, as evidenced here in an orthotopic pancreatic cancer model. With our method, further intravital imaging studies can elucidate the pathophysiology of various diseases involving the pancreas at the cellular level.

Stabilized Longitudinal In Vivo Cellular-Level Visualization of the Pancreas in a Murine Model with a Pancreatic Intravital Imaging Window

In vivo high-resolution imaging of the pancreas was facilitated with the pancreatic intravital imaging window.

Pankreas intravital görüntüleme penceresine sahip bir Murine Modelinde Pankreasın Stabilize Boyuna In Vivo Hücresel Düzey Görselleştirmesi

Direct in vivo cellular-resolution imaging of the pancreas in a live small animal model has been technically challenging. A recent intravital imaging ...

Establishment of a Mouse Severe Acute Pancreatitis Model using Retrograde Injection of Sodium Taurocholate into the Biliopancreatic Duct

The prevalence of acute pancreatitis (AP), especially severe acute pancreatitis (SAP), is increasing in younger age groups annually. However, there is a lack of effective treatments in the current clinical practice. With the easy accessibility of transgenic and knockout strains and their small size, which allows minimal doses of drugs required for in vivo evaluation, a well-established experimental model in mice is preferred for AP research. Moreover, SAP induced through sodium taurocholate (TC) is currently one of the most widely used and best characterized models. This model has been investigated for novel therapies and possible molecular events during the process of AP. Here, we present the generation of an AP mouse model using sodium taurocholate and a simple homemade microsyringe. Moreover, we also provide the methodology for the subsequent histology and serological testing.

A mouse model of severe acute pancreatitis is described herein. The procedure presented here is very rapid, simple, and accessible, thereby potentially allowing the study of the molecular mechanisms and different therapeutic interventions in acute pancreatitis in a convenient way.

Biliopankreatik Kanala Sodyum Taurocholate Retrograd Enjeksiyonu Kullanılarak Fare Şiddetli Akut Pankreatit Modelinin Kurulması

The prevalence of acute pancreatitis (AP), especially severe acute pancreatitis (SAP), is increasing in younger age groups annually. However, there is a ...

Partial Sciatic Nerve Ligation: A Mouse Model of Chronic Neuropathic Pain to Study the Antinociceptive Effect of Novel Therapies

Management of chronic pain remains challenging to this day, and current treatments are associated with adverse effects, including tolerance and addiction. Chronic neuropathic pain results from lesions or diseases in the somatosensory system. To investigate potential therapies with reduced side effects, animal pain models are the gold standard in preclinical studies. Therefore, well-characterized and well-described models are crucial for the development and validation of innovative therapies. 
Partial ligation of the sciatic nerve (pSNL) is a procedure that induces chronic neuropathic pain in mice, characterized by mechanical and thermal hypersensitivity, ongoing pain, and changes in limb temperature, making this model a great fit to study neuropathic pain preclinically. pSNL is an advantageous model to study neuropathic pain as it reproduces many symptoms observed in humans with neuropathic pain. Furthermore, the surgical procedure is relatively fast and straightforward to perform. Unilateral pSNL of one limb allows for comparison between the ipsilateral and contralateral paws, as well as evaluation of central sensitization. 
To induce chronic neuropathic hypersensitivity, a 9-0 non-absorbable nylon thread is used to ligate the dorsal third of the sciatic nerve. This article describes the surgical procedure and characterizes the development of chronic neuropathic pain through multiple commonly used behavioral tests. As a plethora of innovative therapies are now being investigated to treat chronic pain, this article provides crucial concepts for standardization and an accurate description of surgeries required to induce neuropathic pain.

Partial sciatic nerve ligation induces long-lasting chronic neuropathic pain, characterized by exaggerated responses to thermal and mechanical stimuli. This mouse model of neuropathic pain is commonly used to study innovative therapies for pain management. This article describes in detail the surgical procedure to improve standardization and reproducibility.

Kısmi Siyatik Sinir Ligasyonu: Yeni Tedavilerin Antinosiseptif Etkisini İncelemek için Kronik Nöropatik Ağrının Bir Fare Modeli

Management of chronic pain remains challenging to this day, and current treatments are associated with adverse effects, including tolerance and addiction. ...