The work was supported in part by funding from Alberta Livestock and Meat Agency and Natural Science and Research Council Discovery grant to S. D. Proctor.

video ve yazıda hem anlatılan deney hayvanlarında Tüm işlemler Hayvan Bakım Kanada Konseyi tarafından belirlenen kurallar Kurumsal Hayvan Bakım ve Kullanım Kurulu tarafından onaylanmış ve takip edildi. 1. Cerrahi Anestezi ve Neonatal Domuzlar Cerrahi Hazırlık <ol><li> azaperone (0.3 mg / kg) ketamin hidroklorür (10 mg / kg), deksmedetomidin (15 ug / kg), bir ayrı Hol olarak, 25 kg ihtiva eden bir kas içi sedatif anestetik madde kokteyli ile boyun tabanına yakın domuz premedikasyon. Not: Geliştirilmiş intra-operatif ağrı kontrolü için önceden anestezik ilaç kokteyli buprenorfin (0,005-,02 mg / kg) ekleyin. </li><li> Bir yüz maskesi kullanılarak (500 ml-1000 ml / dk O 2 4-5 izofluran%) solunan izofluran inhalasyon gaz ile domuz uyutmak. Bir veteriner laringeal kapsamı (17-25 cm uzunluğunda düz bıçak) kullanarak ses telleri görselleştirmek ve vokal topikal% 10 lidokain spray uygulamakkordonlar. lidokain sprey vokal kord spazm ve havayolu tıkanıklığına olasılığını azaltmak için entübasyon 30-60 sn önce ses telleri temas etmesine izin. </li><li> Izofluran gaz ile anestezi ses tellerinin arasından bir kelepçeli endotrakeal tüpün (5,0-7,0 mm İç Çap (ID)) geçerek domuz entübe ve bakımını (/ 1,000-2,000 ml 0.5-2.0 izofluran% min O 2) ile bir kapalı ameliyat boyunca anestezi sistemi yeniden soluk devresi. çene sesi ile anestezi düzeyi ve hem pedal ve palpebral refleks tepkilerini nasıl değerlendiriyorsunuz. Süresi Doldu anestezik gaz uzaklaştırılmış ve cerrahi paketi dışında havalandırılır. </li><li> % 70 izopropil alkol durulama ardından% 2.0 klorheksidin cerrahi fırçalama çözeltisi ile kulağın dış yüzeyini temizleyin. Ameliyat sırasında, bir 20 G kateter intravenöz ile intravenöz sıvı (5-10 ml / kg / saat laktatlı Ringer çözeltisi) temin etmek üzere bir kulak daman kateter. Med dil mukozal yüzeyine bir pals oksimetresi elde edinical bant kalp hızı ve periferik kan oksijenlenme doygunluk (SpO2) izlemek için. </li><li> dorsal sırt üstü yatar pozisyonda anestezi domuz yerleştirin ve pubis ventral orta toraks kaudal ventral karın tıraş. İki% 2.0 Klorheksidin cerrahi önlük ve steril su yıkar alternatif bu bölgeyi temizleyin. </li><li> Cerrahi paketine anestezi domuz aktarın ve% 70 izopropil alkol durulama nihai cerrahi fırçalayın uygulamak, bu kurumasını bekleyin ve sonra hayvan asmak. </li><li> vücut ısısını izlemek için rektum içine bir rektal sıcaklık probu yaklaşık 2-4 cm yerleştirin. cerrahi işlem sırasında normal vücut sıcaklığına (38-40 ° C) muhafaza edilmesi için bir su sirkülasyon ısıtma pedi üzerinde domuz yerleştirin. </li><li> karın çevresinde örten kadran desen yerleştirilir dört havlu perdeler ile domuz örtün. karın yaklaşık 5 c lateral boyunca, xiphisternum karşısında ikinci asmak ilk örtüyü yerleştirinKarın orta hat m yanal. (Karşı tarafta olmasına rağmen), yaklaşık 5 cm karın orta hatta yanal karın lateral boyunca yerleştirilir ikinci perde gibi, pelvis ileal kret ve dördüncü drape genelinde üçüncü asmak yerleştirin. </li><li> altta yatan havlu perdeler üzerinde cerrahi sitesine erişim sağlayan bir yarık açılması ile, büyük bir tablo asmak yerleştirin ve domuz ve tüm cerrahi tabloyu kapsamaktadır. Nihai asmak büyük masa asmak üzerine yerleştirilen tek kullanımlık bir steri-drape olduğunu. </li></ol> Bağırsak Lenfatik Trunk 2. Abdominal Cerrahi ve Kateterizasyon <ol><li> Altta yatan karın kasları ortaya çıkarmak için bir neşter bıçağı ile 20 cm&#39;lik cilt kesi olun. parietal periton maruz mono-polar elektrokoter (20 Watt ayarı) ile karın kas katmanları İnsizyon. Karın iç organları ve lenfatik damar erişmek için Metzenbaum makas ile parietal periton bir 20 cm doğrusal segmenti açın. </li><li&gt; Ameliyat süresince açık karın boşluğuna tutmak için cerrahi kesi kafatası yönü de retractor yerleştirin. </li><li> tüm cerrahi işlem için sıcak (37 ° C) steril serum fizyolojik ile tüm dokuların nemlendirin. Yavaşça karın boşluğuna gelen kolon, çekum, ileum ve jejunum da dahil olmak üzere bağırsak büyük bir kesimi kaldırın ve üst karın, karaciğer ve lenf damarı erişmek için domuz soldan onu exteriorize. yavaşça bağırsak desteklemek için bir askı oluşturmak için ek havlu perdeler ile pozisyonunda batın bağırsak sabitleyin. </li><li> Lenfatik damar bulun, yaklaşık 4 cm sağ renal ven, 6 cm kuyruk-medial kaval ustabaşı ventral ve pankreas 22,36,37 yakın karaciğer sağ lobunda visseral yönü altında kranial-medial bırakır. Portal ven 36,37 sağ ventral kesimine bitişik bir saydam bir yapı olarak lenfatik damar belirlemek. </li><li> lenfatik vess ayırınEl nazikçe Q-tip uygulayıcılar ile bağlı doku uzak alay ile çevredeki fasya gelen. kabın yan taraflarından çevreleyen dokudan ayrılır sonra, ince bir küt uçlu forseps teknenin altında bir &quot;tüneli&quot; açıklığı oluşturur. </li><li> ince forseps ile lenfatik altında üç 2-0 ipek sütür geçirin. , Tıkamak dilate ve lenf ile gemi doldurmak için ilk olarak en kaudal dikiş Arter. Kasıtlı lenfatik kateteri sabitlemek için nispeten uzun bu sütür (4 cm) uçlarını bırakın. diğer iki dikiş birbirinden 1,0 cm ayrılmış ve güvenli kaudal sütür kranial 1.0-1.5 cm yaklaşık olarak yerleştirin. damar içine kateter hızlı emniyete için izin vermek için bir &#39;tek gevşek bağ kravat&#39; bu iki sütür bırakın. Not: En kaudal lige sütür ve (iki kafatası sütür) orta sütür arasında bulunan lenfatik segment kateterizasyon için sitedir. SU ile ilgiliGerekirse Ture malzemesi, 2-0 polyglactin sütür 2-0 ipek dikişlerle yerine geçebilir. </li><li> iris makas ile damar içine küçük bir delik kesin ve ince künt forseps ile gemi dilate. kabına bir eğimli ucu ile özel kateter tüp (4.06 Dış Çap (OD) x 2.31 mm ID) yaklaşık 1.0-1.5 cm yerleştirin ve yerine kateteri sabitlemek için iki kranial sütürler kravat. Geminin kateteri sabitlemek için kaudal dikiş uzun dikiş uçlarını kullanın. </li><li> ılık serum fizyolojik ile bol miktarda batın bağırsak yıkayın ve yavaşça bağırsak doğru anatomik konumlandırılmasını sağlayan karın boşluğuna geri getirin. </li><li> (Paralumbar fossa 5-10 cm ventral) Sol orta kanadında kateter exteriorize. bir neşter ile bir cilt kesi yapmak ve kateterin eksteriorizasyon için bir açıklık oluşturmak için deri yüzeyine karın boşluğuna bir Noktalayıcıyı geçmektedir. Karın cav kateteri exteriorize büyük Kelly forseps kullanabilirtrokar deliğinden Sığ. </li><li> yuvarlak (konik) iğne ile 2-0 polyglactin dikiş basit sürekli dikiş deseni ile parietal periton kapatın. yuvarlak iğne 2-0 polyglactin sütür ile basit bir kesintiye dikiş deseni ile karın kas katmanları kapatın. </li><li> Bir kesim iğne 2-0 polyglactin sütür ile bir subkütiküler desen cilt kapatın. Bir çanta-string dikiş deseni ile deri ve kesme iğne 2-0 naylon sütür ile batın kateter sabitleyin. </li><li> hala yerleştirme kolaylığı ve kurtarma sırasında domuz stresi azaltmak için anestezi sırasında domuz üzerinde uzmanlaşmış bir ceket yerleştirin. </li></ol> 3. Post-cerrahi Kurtarma ve Lenf Koleksiyonu <ol><li> önce inhalasyon anestezi kesilmesi yaklaşık 10 dakika, hemen cerrahi sonrası analjezi sağlamak için kas içi bupenorphine (0.1 mg / kg) yönetmek. bupenorphine devam (0.1 mg / kg) 24-48 saat koruması için her 12 saatameliyat sonrası analjezi. </li><li> ameliyat sonrası komplikasyonlar için 7 gün boyunca her 8-12 saat domuz izleyin. </li><li> etilendiamin tetraasetik asit (EDTA) ile kaplanmış ve antibiyotikler ile takviye edilmiş 500 ml polipropilen yıkama şişeleri lenf, toplamak; penisilin (6000 IU), streptomisin (6 mg) ve amfoterisin B (3 mg), 7 günlük bir süre boyunca her 12 saat süredir. </li></ol> Lipoprotein ApoB48, Trigliserid, Kolesterol ve toplam Protein 4. Niceleme Lenf Toplanan <ol><li> 4 ° C&#39;de 5 dakika boyunca 1800 x g&#39;de lenf santrifüjleyin. Süpernatant toplayın ve trigliserid, kolesterol ve toplam proteinin miktar tayini için kullanın. </li><li> seyreltilmemiş bir numunesidir, örnek bir 1:20 seyreltilmiş damıtılmış su ve 1 seyreltilmiş bir son örneği: damıtılmış su ile 100, üç numune halinde süpernatant bölün. </li><li> Bir ticari olarak temin edilebilir kit ile, kolesterol seviyelerini ölçmek için seyreltilmemiş süpernatant kullanın. </li><li> 1:20 ve 1:10 kullanarak0 seyreltilmiş numuneler ticari olarak temin edilebilir kit sırasıyla bisinkoninik asit toplam protein tahlili ile, trigliserid ve toplam protein düzeylerini ölçmek için. </li></ol> Lenf 38 Toplanan Lipoprotein ApoB48 5. miktarının belirlenmesi, <ol><li> Uyarlanmış bağışıklık Western Blot yöntemiyle 38 ile lipoprotein ApoB48 konsantrasyonunu belirleyin. % 3-8 tris-asetat- sodyum dodesil sülfat poliakrilamid jel elektroforezi (SDS-PAGE) ile ayırın toplam lenf. </li><li> bir anti-keçi sekonder antikor ile birleştirmiş, sonra: Bir polivinil florür membran (0,45 mikron) ayrılmış proteinler aktarın ve ApoB (4.000 1) bir keçi poliklonal antikor ile kuluçkaya yatmaktadır. </li><li> saflaştırılmış kemirgen ApoB48 protein standardı ile doğrusal dansitometrik karşılaştırma kullanılarak kemilüminesans ile lipoprotein ApoB48 niceliğini. </li></ol>

The authors have nothing to disclose.

Bağırsak lenf Toplama çeşitli deneysel hayvan modellerinde lipid 8-12 ve ilaç 13-15 metabolizması, kanser metastazı 16,17, hücre kaçakçılığı ve bağışıklık fonksiyonu 18-26 katılan mekanizmaları araştırmak için mükemmel bir yöntemdir. Nitekim, uzun bir süre boyunca ya periferik (afferent) ve merkezi (efferent ve büyük gövde damarlar) lenf büyük miktarlarda hasat yeteneği immün modülasyon ajanları 18-22 ile zorluklar aşağıdaki h?...

Lenf sistemi fizyolojisi ilgili yeterli bir alandır. Lenfatik kateterizasyon Klinik öncesi modeller farklı hayvan türleri 1-8 oluşur ve lipid 8-12 ve ilaç metabolizması 13-15, deneysel tedavi 17 kanser metastazı 16 katılan mekanizmaların araştırılması ilaç sanayi ve araştırma kurumları tarafından kullanılan ve bağışıklık fonksiyonu 18 -26. Bu çalışma, lipoprotein metabolizma bileşenlerini ölçmek için bir iç domuz modelinde bağırsak lenf gövde kateterizasyon kullanımını araştırıyor. Lipoprotein metabolizması üretimi ve salgılanması şilomikronlar bölgesinin ve ilişkili lipid ve toplam proteinin değişiklikler yer almaktadır. Yaygın olarak kullanılan kemirgen modellerinde ve insanlar arasında ve gibi lipid metabolizmasında önemli farklılıklar vardır Bunlar beni lipid incelemek için daha karşılaştırılabilir bilgiler verebilir bağırsak lenf toplamak için domuz modelleri kullanılarak, önemli hususlar vardırinsanlar 27-31 yılında tabolism. Çeşitli cerrahi teknikler büyük hayvan türlerinde bağırsak lenf toplamak için kullanılır: kranial omuz yaklaşımı (yani, göğüs kanal kateterizasyon) 5, lateral üst kanadını yaklaşımı 32-34 ve ventral orta hat veya paramediyan yaklaşım 22,35. Bu video ayrıntılı olarak bağırsak lenf gövde kateterizasyon için ventral orta hat cerrahi yaklaşım kullanılarak domuzlarda cerrahi prosedürü tanımlamaktadır. Dikkatli cerrahi teknik izin lenfatik kateterizasyon bu yöntem, uzun zaman süreleri boyunca büyük lenf miktarlarda ve kendisine hedef toplamak için. Bu teknik, çeşitli fizyolojik fonksiyonları inceleyerek birçok disiplinlerle uygulamalar sayısız açar. Uygulamalar içerir, ancak tüm vücut lipoprotein ve lipid metabolizması, immüno-tümör oluşumu ve metastaz, bağırsak fonksiyonu ve developm, bunlarla sınırlı değildirent ve bağırsak inflamatuvar hastalığın ilerlemesi.

<table border="0" cellpadding="0" cellspacing="0" width="567">
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		<tr height="60">
			<td height="60" style="height:60px;width:211px;">3M Surgical steri- drape</td>
			<td style="width:71px;">3M Health Care</td>
			<td style="width:119px;">1040</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:211px;">SDS page gel</td>
			<td style="width:71px;">Invitrogen</td>
			<td style="width:119px;">EA0375BOX</td>
			<td style="width:167px;">3-8 % tris acetate</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:211px;">Polyvinylidene fluoride membrane</td>
			<td style="width:71px;">Millipore</td>
			<td style="width:119px;">IPVH00010</td>
			<td style="width:167px;">0.45 &#956;m pore size</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:211px;">ApoB antibody&#160;</td>
			<td style="width:71px;">EMD Millipore</td>
			<td style="width:119px;">AB742</td>
			<td style="width:167px;">1:4000 dilution</td>
		</tr>
		<tr height="80">
			<td height="80" style="height:80px;width:211px;">Donkey anti-goat IgG-HRP</td>
			<td style="width:71px;">Santa Cruz Biotechnology</td>
			<td style="width:119px;">Sc-2304</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="100">
			<td height="100" style="height:100px;width:211px;">ECL Prime Western Blotting Reagent</td>
			<td style="width:71px;">GE Healthcare LifeSciences</td>
			<td style="width:119px;">RPN2232&#160;&#160;&#160;</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="80">
			<td height="80" style="height:80px;width:211px;">Triglyceride Kit</td>
			<td style="width:71px;">Wako Pure Chemicals</td>
			<td style="width:119px;">998-40391/994-40491</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="80">
			<td height="80" style="height:80px;width:211px;">Total Cholesterol Kit</td>
			<td style="width:71px;">Wako Pure Chemicals</td>
			<td style="width:119px;">439-17501</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:211px;">Total Protein</td>
			<td style="width:71px;">&#160;Pierce&#160;</td>
			<td style="width:119px;">23225</td>
			<td style="width:167px;">Bicinchoninic Acid Assay</td>
		</tr>
	</tbody>
</table>

long-term catheterization of the intestinal lymph trunk and collection of lymph in neonatal pigs

Catheterization of the intestinal lymph trunk in neonatal pigs is a technique allowing for the long-term collection of large quantities of intestinal (central) efferent lymph. Importantly, the collection of central lymph from the intestine enables researchers to study both the mechanisms and lipid constitutes associated with lipid metabolism, intestinal inflammation and cancer metastasis, as well as cells involved in immune function and immunosurveillance. A ventral mid-line surgical approach permits excellent surgical exposure to the cranial abdomen and relatively easy access to the intestinal lymph trunk vessel that lies near the pancreas and the right ventral segment of the portal vein underneath the visceral aspect of the right liver lobe. The vessel is meticulously dissected and released from the surrounding fascia and then dilated with sutures allowing for insertion and subsequent securing of the catheter into the vessel. The catheter is exteriorized and approximately 1 L/24 hr of lymph is collected over a 7 day period. While this technique enables the collection of large quantities of central lymph over an extended period of time, the success depends on careful surgical dissection, tissue handling and close attention to proper surgical technique. This is particularly important with surgeries in young animals as the lymph vessels can easily tear, potentially leading to surgical and experimental failure. The video demonstrates an excellent surgical technique for the collection of intestinal lymph.

<html> Yenidoğan domuz bağırsak lenfatik gövde lenfatik kateterizasyon 7 günlük süre içinde yaklaşık 1 L merkezi lenf / 24 saat toplanmasını sağlar. Bu deneylerde elde lenf lipid metabolizması, yani toplam lenf protein, ApoB48 kolesterol, trigliserid, toplam protein ve kolesterol bileşenlerini ihtiva etmiştir. Tablo 1 vurgulamaktadır üç domuz toplanmış lenf örneklerinden bu lipit bileşenleri temsil miktarları. Özellikle, lenf akışı ve lipid bileşenleri in-line...

Watch this Scientific Journal Video about Long-Term Catheterization of the Intestinal Lymph Trunk and Collection of Lymph in Neonatal Pigs at JoVE.com

Long-Term Catheterization of the Intestinal Lymph Trunk and Collection of Lymph in Neonatal Pigs

We present a surgical procedure to catheterize the intestinal lymph trunk in neonatal pigs to collect large quantities of lipid metabolism components from efferent lymph.

Yenidoğan Domuzların Lenf Bağırsak Lenf Trunk ve Koleksiyonu Uzun Vadeli Kateterizasyon

Catheterization of the intestinal lymph trunk in neonatal pigs is a technique allowing for the long-term collection of large quantities of intestinal ...

long-term-catheterization-intestinal-lymph-trunk-collection-lymph

Research

JoVE Journal

Medicine

8.6K Görüntüleme.  University of Alberta. We present a surgical procedure to catheterize the intestinal lymph trunk in neonatal pigs to collect large quantities of lipid metabolism components from efferent lymph.

Video: Long-Term Catheterization of the Intestinal Lymph Trunk and Collection of Lymph in Neonatal Pigs

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

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

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

Jinekolojik Onkoloji Sentinel lenf nodu intraoperatif Algılama için Multispektral Gerçek zamanlı Floresans Görüntüleme

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

Tıp

Basic Surgical Techniques in the G&ouml;ttingen Minipig: Intubation, Bladder Catheterization, Femoral Vessel Catheterization, and Transcardial Perfusion

The emergence of the G&ouml;ttingen minipig in research of topics such as neuroscience, toxicology, diabetes, obesity, and experimental surgery reflects the close resemblance of these animals to human anatomy and physiology 1-6.The size of the G&ouml;ttingen minipig permits the use of surgical equipment and advanced imaging modalities similar to those used in humans 6-8. The aim of this instructional video is to increase the awareness on the value of minipigs in biomedical research, by demonstrating how to perform tracheal intubation, transurethral bladder catheterization, femoral artery and vein catheterization, as well as transcardial perfusion. 
Endotracheal Intubation should be performed whenever a minipig undergoes general anesthesia, because it maintains a patent airway, permits assisted ventilation and protects the airways from aspirates. Transurethral bladder catheterization can provide useful information about about hydration state as well as renal and cardiovascular function during long surgical procedures. Furthermore, urinary catheterization can prevent contamination of delicate medico-technical equipment and painful bladder extension which may harm the animal and unnecessarily influence the experiment due to increased vagal tone and altered physiological parameters. Arterial and venous catheterization is useful for obtaining repeated blood samples and monitoring various physiological parameters. Catheterization of femoral vessels is preferable to catheterization of the neck vessels for ease of access, when performing experiments involving frame-based stereotaxic neurosurgery and brain imaging. When performing vessel catheterization in survival studies, strict aseptic technique must be employed to avoid infections6. Transcardial perfusion is the most effective fixation method, and yields preeminent results when preparing minipig organs for histology and histochemistry2,9. For more information about anesthesia, surgery and experimental techniques in swine in general we refer to Swindle 2007. Supplementary information about premedication and induction of anesthesia, assisted ventilation, analgesia, pre- and postoperative care of G&ouml;ttingen minipigs are available via the internet at <a href="http://www.minipigs.com">http://www.minipigs.com</a>10. For extensive information about porcine anatomy we refer to Nickel et al. Vol. 1-511.

Basic Surgical Techniques in the G&#246;ttingen Minipig: Intubation, Bladder Catheterization, Femoral Vessel Catheterization, and Transcardial Perfusion

The use of domestic and miniature pigs in science has increased significantly in recent years. By demonstrating how to perform intubation, transurethral bladder catheterization, femoral artery and vein catheterization, as well as transcardial perfusion, we aim to further increase the value of G&ouml;ttingen minipigs in biomedical research.

Göttingen Minipig Cerrahi Temel Teknikleri: Entübasyon, Mesane Kateterizasyon, Femoral Gemi Kateterizasyon, ve Transcardial Perfüzyon

The emergence of the G&ouml;ttingen minipig in research of topics such as neuroscience, toxicology, diabetes, obesity, and experimental surgery reflects ...

Myocardial Infarction and Functional Outcome Assessment in Pigs

Introduction of newly discovered cardiovascular therapeutics into first-in-man trials depends on a strictly regulated ethical and legal roadmap. One important prerequisite is a good understanding of all safety and efficacy aspects obtained in a large animal model that validly reflect the human scenario of myocardial infarction (MI). Pigs are widely used in this regard since their cardiac size, hemodynamics, and coronary anatomy are close to that of humans. Here, we present an effective protocol for using the porcine MI model using a closed-chest coronary balloon occlusion of the left anterior descending artery (LAD), followed by reperfusion. This approach is based on 90 min of myocardial ischemia, inducing large left ventricle infarction of the anterior, septal and inferoseptal walls. Furthermore, we present protocols for various measures of outcome that provide a wide range of information on the heart, such as cardiac systolic and diastolic function, hemodynamics, coronary flow velocity, microvascular resistance, and infarct size. This protocol can be easily tailored to meet study specific requirements for the validation of novel cardioregenerative biologics at different stages (i.e. directly after the acute ischemic insult, in the subacute setting or even in the chronic MI once scar formation has been completed). This model therefore provides a useful translational tool to study MI, subsequent adverse remodeling, and the potential of novel cardioregenerative agents.

This protocol describes the porcine myocardial infarction (MI) model using a 90 min closed-chest coronary balloon occlusion of the left anterior descending artery (LAD), followed by reperfusion. Furthermore, the protocol for several outcome parameters, such as cardiac function, hemodynamics, microvascular resistance, and infarct size, are also presented.

Domuzlar miyokard enfarktüsü ve Fonksiyonel Sonuç Değerlendirmesi

Introduction of newly discovered cardiovascular therapeutics into first-in-man trials depends on a strictly regulated ethical and legal roadmap. One ...

Collection and Extraction of Saliva DNA for Next Generation Sequencing

The preferred source of DNA in human genetics research is blood, or cell lines derived from blood, as these sources yield large quantities of high quality DNA. However, DNA extraction from saliva can yield high quality DNA with little to no degradation/fragmentation that is suitable for a variety of DNA assays without the expense of a phlebotomist and can even be acquired through the mail. However, at present, no saliva DNA collection/extraction protocols for next generation sequencing have been presented in the literature. This protocol optimizes parameters of saliva collection/storage and DNA extraction to be of sufficient quality and quantity for DNA assays with the highest standards, including microarray genotyping and next generation sequencing.

DNA extraction from saliva can provide a readily available source of high molecular weight DNA, with little to no degradation/fragmentation. This protocol provides optimized parameters for saliva collection/storage and DNA extraction to be of sufficient quality and quantity for downstream DNA assays with high quality requirements.

Yeni Nesil dizinleri için Toplama ve Tükrük DNA çıkarımı

The preferred source of DNA in human genetics research is blood, or cell lines derived from blood, as these sources yield large quantities of high quality ...

High-frequency Ultrasound Imaging of Mouse Cervical Lymph Nodes

High-frequency ultrasound (HFUS) is widely employed as a non-invasive method for imaging internal anatomic structures in experimental small animal systems. HFUS has the ability to detect structures as small as 30 &#181;m, a property that has been utilized for visualizing superficial lymph nodes in rodents in brightness (B)-mode. Combining power Doppler with B-mode imaging allows for measuring circulatory blood flow within lymph nodes and other organs. While HFUS has been utilized for lymph node imaging in a number of mouse&#160; model systems, a detailed protocol describing HFUS imaging and characterization of the cervical lymph nodes in mice has not been reported. Here, we show that HFUS can be adapted to detect and characterize cervical lymph nodes in mice. Combined B-mode and power Doppler imaging can be used to detect increases in blood flow in immunologically-enlarged cervical nodes. We also describe the use of B-mode imaging to conduct fine needle biopsies of cervical lymph nodes to retrieve lymph tissue for histological&#160; analysis. Finally, software-aided steps are described to calculate changes in lymph node volume and to visualize changes in lymph node morphology following image reconstruction. The ability to visually monitor changes in cervical lymph node biology over time provides a simple and powerful technique for the non-invasive monitoring of cervical lymph node alterations in preclinical mouse models of oral cavity disease.

This protocol describes the application of high-frequency ultrasound (HFUS) for imaging mouse cervical lymph nodes. This technique optimizes visualization and quantification of cervical lymph node morphology, volume and blood flow. Image-guided biopsy of cervical lymph nodes and processing of lymph tissue for histological evaluation is also demonstrated.

Fare Servikal lenf nodu Yüksek frekanslı Ultrasonik Görüntüleme

High-frequency ultrasound (HFUS) is widely employed as a non-invasive method for imaging internal anatomic structures in experimental small animal ...

Surgical Placement of Catheters for Long-term Cardiovascular Exercise Testing in Swine

This protocol describes the surgical procedure to chronically instrument swine and the procedure to exercise swine on a motor-driven treadmill. Early cardiopulmonary dysfunction is difficult to diagnose, particularly in animal models, as cardiopulmonary function is often measured invasively, requiring anesthesia. As many anesthetic agents are cardiodepressive, subtle changes in cardiovascular function may be masked. In contrast, chronic instrumentation allows for measurement of cardiopulmonary function in the awake state, so that measurements can be obtained under quiet resting conditions, without the effects of anesthesia and acute surgical trauma. Furthermore, when animals are properly trained, measurements can also be obtained during graded treadmill exercise.
Flow probes are placed around the aorta or pulmonary artery for measurement of cardiac output and around the left anterior descending coronary artery for measurement of coronary blood flow. Fluid-filled catheters are implanted in the aorta, pulmonary artery, left atrium, left ventricle and right ventricle for pressure measurement and blood sampling. In addition, a 20 G&#160;catheter is positioned in the anterior interventricular vein to allow coronary venous blood sampling.
After a week of recovery, swine are placed on a motor-driven treadmill, the catheters are connected to pressure and flow meters, and swine are subjected to a five-stage progressive exercise protocol, with each stage lasting 3 min. Hemodynamic signals are continuously recorded and blood samples are taken during the last 30 sec of each exercise stage.
The major advantage of studying chronically instrumented animals is that it allows serial assessment of cardiopulmonary function, not only at rest but also during physical stress such as exercise. Moreover, cardiopulmonary function can be assessed repeatedly during disease development and during chronic treatment, thereby increasing statistical power and hence limiting the number of animals required for a study.

Here we present a protocol to assess cardiopulmonary function in awake swine, at rest and during graded treadmill exercise. Chronic instrumentation allows for repeated hemodynamic measurements uninfluenced by cardiodepressive anesthetic agents.

Domuz uzun süreli Kardiyovasküler Egzersiz Testi için Kateter Cerrahi Yerleştirme

This protocol describes the surgical procedure to chronically instrument swine and the procedure to exercise swine on a motor-driven treadmill. Early ...

Laparoscopic Technique for Serial Collection of Liver and Mesenteric Lymph Nodes in Macaques

The mesenteric lymph nodes (MLN) and the liver are exposed to microbes and microbial products from the gastrointestinal (GI) tract, making them immunologically unique. The GI tract and associated MLN are sites of early viral replication in human immunodeficiency virus (HIV) infection and the MLN are likely important reservoir sites that harbor latently-infected cells even after prolonged antiretroviral therapy (ART). The liver has been shown to play a significant role in immune responses to lentiviruses and appears to play a significant role in clearance of virus from circulation. Nonhuman primate (NHP) models for HIV and Acquired Immunodeficiency Syndrome (AIDS) closely mimic these aspects of HIV infection and serial longitudinal sampling of primary sites of viral replication and the associated immune responses in this model will help to elucidate critical events in infection, pathogenesis, and the impact of various intervention strategies on these events. Current published techniques to sample liver and MLN together involve major surgery and/or necropsy, which limits the ability to investigate these important sites in a serial fashion in the same animal. We have previously described a laparoscopic technique for collection of MLN. Here, we describe a minimally invasive laparoscopic technique for serial longitudinal sampling of liver and MLN through the same two port locations required for the collection of MLN. The use of the same two ports minimizes the impact to the animals as no additional incisions are required. This technique can be used with increased sampling frequency compared to major abdominal surgery and reduces the potential for surgical complications and associated local and systemic inflammatory responses that could complicate interpretation of results. This procedure has potential to facilitate studies involving NHP models while improving animal welfare.

Here, we describe a minimally invasive laparoscopic technique for serial sampling of liver and mesenteric lymph nodes (MLN) in macaques that allows for increased sampling frequency, and reduces the potential for surgical complications when compared to performing a laparotomy.

Macaques Karaciğer ve Mezenterik lenf nodu Seri Koleksiyonu için Laparoskopik Tekniği

The mesenteric lymph nodes (MLN) and the liver are exposed to microbes and microbial products from the gastrointestinal (GI) tract, making them ...

Flow Cytometry Analysis of Immune Cell Subsets within the Murine Spleen, Bone Marrow, Lymph Nodes and Synovial Tissue in an Osteoarthritis Model

Osteoarthritis (OA) is one of the most prevalent musculoskeletal diseases, affecting patients suffering from pain and physical limitations. Recent evidence indicates a potential inflammatory component of the disease, with both T-cells and monocytes/macrophages potentially associated with the pathogenesis of OA. Further studies postulated an important role for subsets of both inflammatory cell lineages, such as Th1, Th2, Th17, and T-regulatory lymphocytes, and M1, M2, and synovium-tissue-resident macrophages. However, the interaction between the local synovial and systemic inflammatory cellular response and the structural changes in the joint is unknown. To fully understand how T-cells and monocytes/macrophages contribute towards OA, it is important to be able to quantitively identify these cells and their subsets simultaneously in synovial tissue, secondary lymphatic organs and systemically (the spleen and bone marrow). Nowadays, the different inflammatory cell subsets can be identified by a combination of cell-surface markers making multi-color flow cytometry a powerful technique in investigating these cellular processes. In this protocol, we describe detailed steps regarding the harvest of synovial tissue and secondary lymphatic organs as well as generation of single cell suspensions. Furthermore, we present both an extracellular staining assay to identify monocytes/macrophages and their subsets as well as an extra- and intra-cellular staining assay to identify T-cells and their subsets within the murine spleen, bone marrow, lymph nodes and synovial tissue. Each step of this protocol was optimized and tested, resulting in a highly reproducible assay that can be utilized for other surgical and non-surgical OA mouse models.

Here, we describe a detailed and reproducible flow cytometry protocol to identify monocyte/macrophage and T-cell subsets using both extra- and intracellular staining assays within the murine spleen, bone marrow, lymph nodes and synovial tissue, utilizing an established surgical model of murine osteoarthritis.

Bir Osteoartrit Modelinde Minör Dalak, Kemik İliği, Lenf Düğümleri ve Sinovyal DokuDaki İmmün Hücre Alt Kümelerinin Akış Sitometri Analizi

Osteoarthritis (OA) is one of the most prevalent musculoskeletal diseases, affecting patients suffering from pain and physical limitations. Recent ...

Analyzing Long-Term Electrocardiography Recordings to Detect Arrhythmias in Mice

Arrhythmias are common, affecting millions of patients worldwide. Current treatment strategies are associated with significant side effects and remain ineffective in many patients. To improve patient care, novel and innovative therapeutic concepts causally targeting arrhythmia mechanisms are needed. To study the complex pathophysiology of arrhythmias, suitable animal models are necessary, and mice have been proven to be ideal model species to evaluate the genetic impact on arrhythmias, to investigate fundamental molecular and cellular mechanisms, and to identify potential therapeutic targets.
Implantable telemetry devices are among the most powerful tools available to study electrophysiology in mice, allowing continuous ECG recording over a period of several months in freely moving, awake mice. However, due to the huge number of data points (&gt;1 million QRS complexes per day), analysis of telemetry data remains challenging. This article describes a step-by-step approach to analyze ECGs and to detect arrhythmias in long-term telemetry recordings using the software, Ponemah, with its analysis modules, ECG Pro and Data Insights, developed by Data Sciences International (DSI). To analyze basic ECG parameters, such as heart rate, P wave duration, PR interval, QRS interval, or QT duration, an automated attribute analysis was performed using Ponemah to identify P, Q, and T waves within individually adjusted windows around detected R waves. 
Results were then manually reviewed, allowing adjustment of individual annotations. The output from the attribute-based analysis and the pattern recognition analysis was then used by the Data Insights module to detect arrhythmias. This module allows an automatic screening for individually defined arrhythmias within the recording, followed by a manual review of suspected arrhythmia episodes. The article briefly discusses challenges in recording and detecting ECG signals, suggests strategies to improve data quality, and provides representative recordings of arrhythmias detected in mice using the approach described above.

Here we present a step-by-step protocol for a semiautomated approach to analyze murine long-term electrocardiography (ECG) data for basic ECG parameters and common arrhythmias. Data are obtained by implantable telemetry transmitters in living and awake mice and analyzed using Ponemah and its analysis modules.

Farelerde Aritmileri Tespit Etmek için Uzun Süreli Elektrokardiyografi Kayıtlarının Analizi

Arrhythmias are common, affecting millions of patients worldwide. Current treatment strategies are associated with significant side effects and remain ...

Robot-assisted Total Mesorectal Excision and Lateral Pelvic Lymph Node Dissection for Locally Advanced Middle-low Rectal Cancer

Since their approval for clinical use, da Vinci surgical robots have shown great advantages in gastrointestinal surgical operations, especially in complex procedures. The high-quality 3-D visual, multijoint arm and natural tremor filtration allow the surgeon to expose and dissect more accurately with minimal invasion. Total mesorectal excision is the standard surgical technique for the treatment of resectable rectal cancer. To reduce the lateral recurrence rate, lateral pelvic lymph node dissection can be performed, as it is a safe and feasible procedure for locally advanced middle-low rectal cancer with a high possibility of metastasis to the lateral lymph nodes. However, the complexity of the anatomic structures and the high postoperative complication rate limit its application. Recently, several surgeons have increasingly used robotic techniques for total mesorectal excision and lateral pelvic lymph node dissection. Compared with open and laparoscopic surgery, the robotic technique has several advantages, such as less blood loss, fewer blood transfusions, minimal trauma, shorter postoperative hospitalization, and quicker recovery. A robotic approach is generally regarded as a reasonable alternative for complicated procedures such as lateral pelvic lymph node dissection, although there are a limited number of high-quality prospective randomized controlled studies reporting direct evidence. Here, we provide the detailed steps of robot-assisted total mesorectal excision and lateral pelvic lymph node dissection performed at the First Affiliated Hospital of Xi&#39;an Jiaotong University.

The robotic technique described herein aims to detail a stepwise approach to robot-assisted total mesorectal excision and lateral pelvic lymph node dissection for locally advanced (T3/T4) rectal cancer located below the peritoneal reflection.

Lokal İlerlemiş Orta-Düşük Rektum Kanseri için Robot Destekli Total Mezorektal Eksizyon ve Lateral Pelvik Lenf Nodu Diseksiyonu

Since their approval for clinical use, da Vinci surgical robots have shown great advantages in gastrointestinal surgical operations, especially in complex ...