Isolation of Myoepithelial Cells from Adult Murine Lacrimal and Submandibular Glands

Tatiana Zyrianova; Liana  V. Basova; Helen Makarenkova

doi:10.3791/59602

Isolamento delle cellule Miepiteliali da adulti murina lacrimale e ghiandole sottomandibolari

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Isolation of Myoepithelial Cells from Adult Murine Lacrimal and Submandibular Glands

Isolamento delle cellule Miepiteliali da adulti murina lacrimale e ghiandole sottomandibolari

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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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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.

La ghiandola lacrimale (LG) ha due tipi di cellule che esprimono α-liscio muscolo actina (αSMA): cellule miepiteliali (MECs) e pericytes. I MECs sono di origine ectodermica, trovati in molti tessuti ghiandolari, mentre i periciti sono cellule muscolari lisce vascolari di origine endodermica. Questo protocollo isola i MECs e i periciti di murine LGs.

Questo protocollo è significativo perché consente la separazione e l'isolamento di due linee cellulari muscolari lisce, le cellule mioepiteliali e i periciti. Questo metodo combina l'etichettatura genetica delle cellule muscolari lisce attraverso marcatori di superficie cellulare per purificare le popolazioni di cellule mioepiteliali e periciti. Questo protocollo consente il confronto della funzione e delle capacità rigenerative delle cellule mioepiteliali sane e masticate e l'elaborazione di queste cellule per studi di espressione genica.

Le cellule mioepiteliali esistono in altre ghiandole esocrine come mammarie, salivari e pancreas, il che consente di adattare questo protocollo isolando cellule mioepiteliali e periciti da qualsiasi altro tessuto. Per etichettare l'actina muscolare liscia alfa o le cellule che esprimono SMA, iniettare topi reporter alfa inducibili alfa SMA da tre a quattro settimane con 100 microlitri di tamoxifene per 20 grammi di peso corporeo intraperitonealmente una volta al giorno per due giorni. Per la raccolta della ghiandola lacrimale, da due a tre giorni dopo l'ultima iniezione, utilizzare una pinzetta per tirare delicatamente una ghiandola e allo stesso tempo graffiare il tessuto connettivo intorno alla ghiandola con la punta affilata di un paio di piccole forbici per liberare la ghiandola.

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