3 आयामी राल कास्टिंग और माउस पोर्टल नस या intrahepatic पित्त नली प्रणाली की इमेजिंग
Summary
Visualizing और माउस यकृत पोर्टल शिरा या intrahepatic पित्त नली के 3 आयामी संरचना बढ़ाता के एक विधि का वर्णन है. इस डाली राल तकनीक भी अन्य ductal या संवहनी प्रणालियों के लिए लागू किया जा सकता है और के लिए अनुमति देता है<em> स्वस्थानी में</em> दृश्य या एक सिस्टम बरकरार वास्तुकला संवाद स्थापित करने की मात्रा का ठहराव.
Abstract
In organs, the correct architecture of vascular and ductal structures is indispensable for proper physiological function, and the formation and maintenance of these structures is a highly regulated process. The analysis of these complex, 3-dimensional structures has greatly depended on either 2-dimensional examination in section or on dye injection studies. These techniques, however, are not able to provide a complete and quantifiable representation of the ductal or vascular structures they are intended to elucidate. Alternatively, the nature of 3-dimensional plastic resin casts generates a permanent snapshot of the system and is a novel and widely useful technique for visualizing and quantifying 3-dimensional structures and networks.
A crucial advantage of the resin casting system is the ability to determine the intact and connected, or communicating, structure of a blood vessel or duct. The structure of vascular and ductal networks are crucial for organ function, and this technique has the potential to aid study of vascular and ductal networks in several ways. Resin casting may be used to analyze normal morphology and functional architecture of a luminal structure, identify developmental morphogenetic changes, and uncover morphological differences in tissue architecture between normal and disease states. Previous work has utilized resin casting to study, for example, architectural and functional defects within the mouse intrahepatic bile duct system that were not reflected in 2-dimensional analysis of the structure1,2, alterations in brain vasculature of a Alzheimer’s disease mouse model3, portal vein abnormalities in portal hypertensive and cirrhotic mice4, developmental steps in rat lymphatic maturation between immature and adult lungs5, immediate microvascular changes in the rat liver, pancreas, and kidney in response in to chemical injury6.
Here we present a method of generating a 3-dimensional resin cast of a mouse vascular or ductal network, focusing specifically on the portal vein and intrahepatic bile duct. These casts can be visualized by clearing or macerating the tissue and can then be analyzed. This technique can be applied to virtually any vascular or ductal system and would be directly applicable to any study inquiring into the development, function, maintenance, or injury of a 3-dimensional ductal or vascular structure.
Protocol
Discussion
हम विशिष्ट उदाहरण के लिए कैसे पोर्टल शिरा और जिगर की intrahepatic पित्त नली प्रणाली डाली जा सकती है वर्णित है, लेकिन इस तकनीक मामूली रूपांतरों के साथ लगभग किसी भी अन्य ductal या नाड़ी तंत्र के लिए लागू किया जा सकता ?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
इस काम के स्वास्थ्य (NIH) के राष्ट्रीय संस्थानों से SSH के लिए अनुदान (R01 DK078640) / HHMI Vanderbilt विश्वविद्यालय सर्टिफिकेट प्रोग्राम के माध्यम से हॉवर्ड ह्यूजेस मेडिकल (HHMI) TJW आण्विक चिकित्सा संस्थान (GRDOT56006779), से, द्वारा समर्थित किया गया Vanderbilt मधुमेह अनुसंधान और प्रशिक्षण केन्द्र (P30 DK020593) और Vanderbilt पाचन रोग अनुसंधान केंद्र (P30-DK058404) कोर सेवाओं प्रदान करते हैं.
Materials
| Name of reagent | Company | Catalog number | Comments |
| PE10 polyethylene tubing | Fisher Scientific | 1417012P | |
| 5-0 surgical black braided silk | Roboz Surgical | SUT-15-1 | |
| Steriject 32 G x 13 mm needle | Air-tite | TSK3213 | |
| Spring scissors | Fine Science Tools | 15000-08 | |
| Number 5 forceps | Fine Science Tools | 91150-20 | |
| Mercox II kit | Ladd Research | 21247 | |
| Benzyl alcohol | Fisher Scientific | 1816-04 | Only required for BABB-clearing |
| Benzyl benzoate | MP Biomedicals, LLC | 154839 | Only required for BABB-clearing |
| Phosphate-buffered saline (PBS) | |||
| Modeling clay | |||
| Scale | |||
| Laboratory scissors | |||
| 15 ml cap polypropylene tubes | |||
| 4% paraformaldahyde | |||
| 15% potassium hydroxide (KOH) | Only required for maceration | ||
| Razor blade | |||
| 100% methanol | |||
| 3 ml luer lock syringe |
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