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Isolation, culture, purification and ultrastructural investigation of cardiac telocytes.

Li YY, Zhang S, Li YG, Wang Y - Mol Med Rep (2016)

Bottom Line: Furthermore, immunofluorescence demonstrated that almost all the sorted TCs expressed vimentin, a marker of TCs.Moreover, electron micrographs showed typical TCs based on their ultrastructural features.Using this method, we developed a reproducible protocol for the isolation and purification of cardiac TCs from rat hearts, which yielded TCs with typical characteristics.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiovascular Diseases, Xinhua Hospital, Shanghai Jiaotong University, School of Medicine, Shanghai 200092, P.R. China.

ABSTRACT
Telocytes (TCs), a novel type of stromal cells, are crucial to cardiac renovation and regeneration. To dissect the pathophysiological effects of cardiac TCs in heart disease, it is essential to develop an effective method to isolate, culture, purify and characterize these cells. In the present study, cardiac TCs were isolated from the hearts of rats by enzymatic digestion. Histology and CD34/PDGFRα expression by flow cytometric assay were used to characterize the cultured cardiac TCs, which were purified by flow cytometric sorting and confirmed by immunofluorescence and electron microscopy. Typical TCs were observed in primary culture, with these exhibiting typical fusiform cell bodies with long moniliform telopodes. Based on flow cytometric sorting with antibodies to CD34 and PDGFRα, there was a substantial increase in the purity of cardiac TCs. Furthermore, immunofluorescence demonstrated that almost all the sorted TCs expressed vimentin, a marker of TCs. Moreover, electron micrographs showed typical TCs based on their ultrastructural features. Using this method, we developed a reproducible protocol for the isolation and purification of cardiac TCs from rat hearts, which yielded TCs with typical characteristics.

No MeSH data available.


Related in: MedlinePlus

Immunofluorescent staining for vimentin in the sorted cardiac TCs. (A) Fluorescence inverted microscopy showed that nearly all sorted TCs are positive for vimentin (red). Nuclei were counterstained with DAPI (blue). (B) Higher magnification of the telocytes. Staining was concomitant on cell body and thin telopode. Original magnification, ×400. TCs, telocytes.
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f3-mmr-14-02-1194: Immunofluorescent staining for vimentin in the sorted cardiac TCs. (A) Fluorescence inverted microscopy showed that nearly all sorted TCs are positive for vimentin (red). Nuclei were counterstained with DAPI (blue). (B) Higher magnification of the telocytes. Staining was concomitant on cell body and thin telopode. Original magnification, ×400. TCs, telocytes.

Mentions: The percentage of vimentin-positive cells was quantified and the mean was determined from 5 randomly selected magnification, ×200 fields. The results showed that 96.7% of the sorted TCs were vimentin positive. The cells showed red immunofluorescence, indicating that they had been labeled with antibody and thus expressed vimentin and the nucleus was stained blue with DAPI (Fig. 3). Flow cytometric sorting and immunofluorescence of the isolated cells revealed positive expression of CD34, PDGFRα and vimentin, which are markers for TCs.


Isolation, culture, purification and ultrastructural investigation of cardiac telocytes.

Li YY, Zhang S, Li YG, Wang Y - Mol Med Rep (2016)

Immunofluorescent staining for vimentin in the sorted cardiac TCs. (A) Fluorescence inverted microscopy showed that nearly all sorted TCs are positive for vimentin (red). Nuclei were counterstained with DAPI (blue). (B) Higher magnification of the telocytes. Staining was concomitant on cell body and thin telopode. Original magnification, ×400. TCs, telocytes.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4940097&req=5

f3-mmr-14-02-1194: Immunofluorescent staining for vimentin in the sorted cardiac TCs. (A) Fluorescence inverted microscopy showed that nearly all sorted TCs are positive for vimentin (red). Nuclei were counterstained with DAPI (blue). (B) Higher magnification of the telocytes. Staining was concomitant on cell body and thin telopode. Original magnification, ×400. TCs, telocytes.
Mentions: The percentage of vimentin-positive cells was quantified and the mean was determined from 5 randomly selected magnification, ×200 fields. The results showed that 96.7% of the sorted TCs were vimentin positive. The cells showed red immunofluorescence, indicating that they had been labeled with antibody and thus expressed vimentin and the nucleus was stained blue with DAPI (Fig. 3). Flow cytometric sorting and immunofluorescence of the isolated cells revealed positive expression of CD34, PDGFRα and vimentin, which are markers for TCs.

Bottom Line: Furthermore, immunofluorescence demonstrated that almost all the sorted TCs expressed vimentin, a marker of TCs.Moreover, electron micrographs showed typical TCs based on their ultrastructural features.Using this method, we developed a reproducible protocol for the isolation and purification of cardiac TCs from rat hearts, which yielded TCs with typical characteristics.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiovascular Diseases, Xinhua Hospital, Shanghai Jiaotong University, School of Medicine, Shanghai 200092, P.R. China.

ABSTRACT
Telocytes (TCs), a novel type of stromal cells, are crucial to cardiac renovation and regeneration. To dissect the pathophysiological effects of cardiac TCs in heart disease, it is essential to develop an effective method to isolate, culture, purify and characterize these cells. In the present study, cardiac TCs were isolated from the hearts of rats by enzymatic digestion. Histology and CD34/PDGFRα expression by flow cytometric assay were used to characterize the cultured cardiac TCs, which were purified by flow cytometric sorting and confirmed by immunofluorescence and electron microscopy. Typical TCs were observed in primary culture, with these exhibiting typical fusiform cell bodies with long moniliform telopodes. Based on flow cytometric sorting with antibodies to CD34 and PDGFRα, there was a substantial increase in the purity of cardiac TCs. Furthermore, immunofluorescence demonstrated that almost all the sorted TCs expressed vimentin, a marker of TCs. Moreover, electron micrographs showed typical TCs based on their ultrastructural features. Using this method, we developed a reproducible protocol for the isolation and purification of cardiac TCs from rat hearts, which yielded TCs with typical characteristics.

No MeSH data available.


Related in: MedlinePlus