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Neuroscience
软脑膜淋巴内皮细胞的收获和原代培养
软脑膜淋巴内皮细胞的收获和原代培养
JoVE Journal
Neuroscience
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JoVE Journal Neuroscience
Harvest and Primary Culture of Leptomeningeal Lymphatic Endothelial Cells

软脑膜淋巴内皮细胞的收获和原代培养

Full Text
2,306 Views
06:44 min
September 8, 2023

DOI: 10.3791/65872-v

Hong-Ji Deng1, Kun Wu2, Han-Fu Yu1, Yong-Jin Zhang1,3, Yun-Cong Li1, Chong Li1, Fei Wang1

1Department of Neurosurgery,The First Affiliated Hospital of Kunming Medical University, 2Department of Clinical Laboratory,The First Affiliated Hospital of Kunming Medical University, 3Clinical Medical Research Center,The First Affiliated Hospital of Kunming Medical University

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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 harvesting and culturing leptomeningeal lymphatic endothelial cells (LLECs) from mice, an intracranial cell type with largely unexplored functions. The established in vitro primary cultures of LLECs can facilitate research into their cellular functions and potential clinical implications.

Key Study Components

Area of Science

  • Neuroscience
  • Cell Biology
  • In Vitro Cultures

Background

  • Leptomeningeal lymphatic endothelial cells (LLECs) are a recently identified cell type in the skull.
  • Their functions and characteristics remain poorly understood in the field of neuroscience.
  • Existing protocols for LLEC cultivation are lacking, necessitating this reproducible method.
  • The study aims to enable further investigation into the potential roles and clinical relevance of LLECs.

Purpose of Study

  • To establish a reliable method for harvesting LLECs from mice.
  • To cultivate these cells in vitro for future research.
  • To investigate the biological significance of LLECs in the nervous system context.

Methods Used

  • The study involved cell culture techniques for LLECs derived from mouse brains.
  • Primary LLEC cultures were established with a specified protocol, achieving over 95% purity.
  • Key steps included tissue digestion, centrifugation, and separation via magnetic selection.
  • Cell behavior was observed and verified through immunofluorescence staining.

Main Results

  • The developed protocol successfully yielded LLECs with high purity, revealing their distinct morphology and characteristics.
  • Immunofluorescent staining confirmed LLECs' identity and differentiation from other cell types.
  • Cultured LLECs exhibited typical endothelial-like features over time.

Conclusions

  • This study provides a foundational protocol for LLEC study, facilitating exploration of their roles in health and disease.
  • The in vitro system opens avenues for understanding LLEC functions in the central nervous system.
  • Future investigations may clarify LLECs’ clinical implications and biological significance in neuroscience.

Frequently Asked Questions

What are the advantages of using the established protocol?
The protocol provides a reproducible method for isolating and cultivating LLECs with high purity, enabling reliable experimentation and understanding of these cells.
How are LLECs harvested from mouse brains?
LLECs are harvested by carefully removing leptomeninges from the brain surface, followed by enzymatic digestion and cell culture techniques.
What type of data or outcomes can researchers expect?
Researchers can obtain insights into the cellular characteristics of LLECs, including their morphology and specific marker expressions.
How can the method be adapted for other research purposes?
Variations of the protocol can be applied to study other brain-derived cell types or explore different biochemical environments by modifying digestion enzymes and media conditions.
What are the limitations of the described method?
The method primarily focuses on LLECs from mice, which may not fully represent LLECs in other species, limiting broader applicability without further validation.

软脑膜淋巴内皮细胞 (LEC) 是一种最近发现的颅内细胞类型,其功能知之甚少。本研究提出了一种可重复的方案,用于从小鼠收获 LLEC 并建立 体外 原代培养物。该协议旨在使研究人员能够深入研究 LEC 的细胞功能和潜在临床意义。

该协议是为其他研究人员设计的,只对建立多程序和文化主要区域感兴趣的区域我们的协议最终导致了纯度超过95%的区域CS的初级培养物的建立目前没有现有的收获和培养协议我们的结果为进一步研究LLECS在虚拟中的类似功能铺平了道路。在RUC上,我们最近发现的颅内细胞群,RUC的生物学意义我们将对细胞功能进行进一步的研究,并回顾其临床意义。

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软脑膜淋巴内皮细胞 LLECs 收获 原代培养 颅内细胞群 外周淋巴内皮细胞 功能研究 体外 方案 纤连蛋白包被 软脑膜解剖 酶消化 单细胞悬浮液 血管内皮生长因子-C (VEGF-C) 淋巴管透明质酸受体-1 (LYVE-1) 磁活化细胞分选 (MACS) 原代培养建立 纯度确认 免疫荧光染色 流式细胞术分析

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