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Biology
Yetişkin Murine lacrimal ve submandibular bezlerden Myoepitelyal hücrelerin izolasyonu
Yetişkin Murine lacrimal ve submandibular bezlerden Myoepitelyal hücrelerin izolasyonu
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Biology
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JoVE Journal Biology
Isolation of Myoepithelial Cells from Adult Murine Lacrimal and Submandibular Glands

Yetişkin Murine lacrimal ve submandibular bezlerden Myoepitelyal hücrelerin izolasyonu

Full Text
8,562 Views
07:15 min
June 11, 2019

DOI: 10.3791/59602-v

Tatiana Zyrianova1, Liana V. Basova1, Helen Makarenkova1

1Department of Molecular Medicine,The Scripps Research Institute

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Please note that some of the translations on this page are AI generated. Click here for the English version.

Overview

This study presents a protocol for the isolation and separation of two smooth muscle cell types from the lacrimal gland: myoepithelial cells (MECs) and pericytes. Using genetic labeling, the method enables purification for comparative analysis of healthy and diseased cells, thereby contributing to our understanding of cell function and regenerative abilities.

Key Study Components

Research Area

  • Cell biology
  • Regenerative medicine
  • Exocrine gland research

Background

  • MECs and pericytes play crucial roles in glandular function and vascular stability.
  • This protocol is adaptable for isolating similar cell types from other exocrine glands.
  • Understanding their functions can provide insights into various biological processes and diseases.

Methods Used

  • Genetic labeling and purification of myoepithelial cells and pericytes.
  • Murine model with tamoxifen-inducible alpha SMA driven reporter mice.
  • Fluorescence-activated Cell Sorting (FACS) for cell analysis.

Main Results

  • Successfully isolated and labeled MECs and pericytes from murine lacrimal glands.
  • Facilitated downstream applications including cell culture and gene expression studies.
  • Demonstrated differences in cell morphology and labeling efficiency.

Conclusions

  • The study provides a valuable protocol for the isolation of specific cell types within the lacrimal gland.
  • This advancement may enhance future research related to exocrine glands and their cellular functions.

Frequently Asked Questions

What are the key benefits of this isolation protocol?
It allows the purification of specific cell types for detailed analysis of their functions and regenerative abilities.
Can this method be used for other tissues?
Yes, the protocol can be adapted for isolation from other exocrine glands.
What is the role of myoepithelial cells?
Myoepithelial cells assist in glandular secretion and contractile functions.
What are pericytes and their significance?
Pericytes are essential for vascular stability and regulation of blood flow in capillaries.
What applications can the isolated cells be used for?
The cells can be utilized for in vivo/in vitro experiments, including cell culture and molecular studies.
How is cell labeling achieved in this protocol?
Cell labeling is accomplished through injection of tamoxifen to activate the desired reporting mechanism.
What technologies were critical for this study?
Fluorescence microscopy and FACS were essential for cell analysis and sorting.

Lakrimal bezi (LG), α-pürüzsüz Kas aksini (αSMA) ifade eden iki hücre türüne sahiptir: miyoepitelyal hücreler (MECs) ve perikayt. MECS ektodermal orijinli, birçok glandüler dokularda bulunan, perikayt endodermal kökeni vasküler pürüzsüz Kas hücreleri iken. Bu protokol, murine LGs 'den MECs ve perikaytları yalıtır.

Bu protokol, iki düz kas hücre hattı, miyoepitelyal hücreler ve perisitlerin ayrılmasına ve izole edilmesine olanak sağladığı için önemlidir. Bu yöntem, miyoepitelyal hücre ve perisit popülasyonlarını arındırmak için hücre yüzeyi belirteçleri aracılığıyla düz kas hücrelerinin genetik etiketlemesini birleştirir. Bu protokol, sağlıklı ve hastalıklı miyoepitelyal hücrelerin fonksiyon ve rejeneratif yeteneklerinin karşılaştırılmasını ve gen ekspresyonu çalışmaları için bu hücrelerin işlenmesine olanak sağlar.

Miyoepitelyal hücreler meme gibi diğer ekzokrin bezleri var, tükürük, ve pankreas, hangi bu protokol miyoepitelyal hücreleri ve perisitler izolasyon tarafından adapte sağlar diğer dokulardan. Alfa düz kas aktin veya SMA ifade hücreleri etiketlemek için, iki gün boyunca günde bir kez vücut ağırlığı intraperitoneally 20 gram tamoksifen 100 mikrolitre ile üç ila dört haftalık tamoksifen-indüklemez alfa SMA tahrik muhabir fareler enjekte. Lacrymal bezi toplama için, son enjeksiyondan iki ila üç gün sonra, aynı zamanda bezi serbest etmek için küçük makas keskin ucu ile bezi etrafında bağ dokusu çizerek ise yavaşça bir bezi çekmek için cımbız kullanın.

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