Insulin and heparin-binding epidermal growth factor-like growth factor synergistically promote astrocyte survival and proliferation in serum-free medium

Mei Jia, Zhongfang Shi, Xu Yan, Lixin Xu, Liping Dong, Jiaxin Li, Yujiao Wang, Shaohua Yang, Fang Yuan

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Background: In vitro systems allowing maintenance and experimentation on primary astrocyte cultures have been used for decades. Astrocyte cultures are most maintained in serum-containing medium which has been found to alter the morphology and gene profiles of astrocytes. New method: Here, we reported a new serum-free medium for astrocyte culture, which consisted of DMEM and NB media supplemented with insulin and heparin-binding epidermal growth factor-like growth factor (HB-EGF) (SF-I-H medium). Meanwhile FBS-containing (FBS) medium composed of DMEM medium containing 10% FBS were used for comparison study. Cerebral cortex was harvested from postnatal day 1 Wistar rats and brain cells were isolated and seeded to poly-L-lysine coated culture dishes after 15 min differential velocity adherence. Results: Compared with FBS medium, astrocytes in SF-I-H medium were smaller and exhibited process bearing morphologies. MTT assays showed that cell density and proliferation rate were higher in SF-I-H medium than in FBS medium all the time, and flow cytometry analysis revealed that SF-I-H medium promoted cell mitosis in a manner comparable to FBS medium. Consistently, western blot analysis further revealed that insulin and HB-EGF synergistically activated the PI3K/AKT and MAPK/ERK1/2 signaling cascades as FBS. Comparison with existing method(s): Astrocytes cultured in SF-I-H medium grew faster than FBS medium. Conclusion: Taken together, our results indicated that SF-I-H medium, in which cell morphology was similar with astrocytes in brain, was more effective for astrocyte survival and proliferation than FBS medium, providing a new cell model to study astrocyte functions without the interference of serum.

Original languageEnglish
Pages (from-to)240-247
Number of pages8
JournalJournal of Neuroscience Methods
Volume307
DOIs
StatePublished - 1 Sep 2018

Fingerprint

Serum-Free Culture Media
Epidermal Growth Factor
Astrocytes
Heparin
Intercellular Signaling Peptides and Proteins
Insulin
Far-Western Blotting
Brain
Serum
Phosphatidylinositol 3-Kinases
Mitosis
Cerebral Cortex
Lysine
Wistar Rats
Flow Cytometry
Cell Count
Maintenance
Cell Proliferation

Keywords

  • Astrocytes
  • Cell proliferation
  • Cell survival
  • Heparin-binding epidermal growth factor-like growth factor
  • Insulin
  • Serum-free medium

Cite this

Jia, Mei ; Shi, Zhongfang ; Yan, Xu ; Xu, Lixin ; Dong, Liping ; Li, Jiaxin ; Wang, Yujiao ; Yang, Shaohua ; Yuan, Fang. / Insulin and heparin-binding epidermal growth factor-like growth factor synergistically promote astrocyte survival and proliferation in serum-free medium. In: Journal of Neuroscience Methods. 2018 ; Vol. 307. pp. 240-247.
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abstract = "Background: In vitro systems allowing maintenance and experimentation on primary astrocyte cultures have been used for decades. Astrocyte cultures are most maintained in serum-containing medium which has been found to alter the morphology and gene profiles of astrocytes. New method: Here, we reported a new serum-free medium for astrocyte culture, which consisted of DMEM and NB media supplemented with insulin and heparin-binding epidermal growth factor-like growth factor (HB-EGF) (SF-I-H medium). Meanwhile FBS-containing (FBS) medium composed of DMEM medium containing 10{\%} FBS were used for comparison study. Cerebral cortex was harvested from postnatal day 1 Wistar rats and brain cells were isolated and seeded to poly-L-lysine coated culture dishes after 15 min differential velocity adherence. Results: Compared with FBS medium, astrocytes in SF-I-H medium were smaller and exhibited process bearing morphologies. MTT assays showed that cell density and proliferation rate were higher in SF-I-H medium than in FBS medium all the time, and flow cytometry analysis revealed that SF-I-H medium promoted cell mitosis in a manner comparable to FBS medium. Consistently, western blot analysis further revealed that insulin and HB-EGF synergistically activated the PI3K/AKT and MAPK/ERK1/2 signaling cascades as FBS. Comparison with existing method(s): Astrocytes cultured in SF-I-H medium grew faster than FBS medium. Conclusion: Taken together, our results indicated that SF-I-H medium, in which cell morphology was similar with astrocytes in brain, was more effective for astrocyte survival and proliferation than FBS medium, providing a new cell model to study astrocyte functions without the interference of serum.",
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Insulin and heparin-binding epidermal growth factor-like growth factor synergistically promote astrocyte survival and proliferation in serum-free medium. / Jia, Mei; Shi, Zhongfang; Yan, Xu; Xu, Lixin; Dong, Liping; Li, Jiaxin; Wang, Yujiao; Yang, Shaohua; Yuan, Fang.

In: Journal of Neuroscience Methods, Vol. 307, 01.09.2018, p. 240-247.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Insulin and heparin-binding epidermal growth factor-like growth factor synergistically promote astrocyte survival and proliferation in serum-free medium

AU - Jia, Mei

AU - Shi, Zhongfang

AU - Yan, Xu

AU - Xu, Lixin

AU - Dong, Liping

AU - Li, Jiaxin

AU - Wang, Yujiao

AU - Yang, Shaohua

AU - Yuan, Fang

PY - 2018/9/1

Y1 - 2018/9/1

N2 - Background: In vitro systems allowing maintenance and experimentation on primary astrocyte cultures have been used for decades. Astrocyte cultures are most maintained in serum-containing medium which has been found to alter the morphology and gene profiles of astrocytes. New method: Here, we reported a new serum-free medium for astrocyte culture, which consisted of DMEM and NB media supplemented with insulin and heparin-binding epidermal growth factor-like growth factor (HB-EGF) (SF-I-H medium). Meanwhile FBS-containing (FBS) medium composed of DMEM medium containing 10% FBS were used for comparison study. Cerebral cortex was harvested from postnatal day 1 Wistar rats and brain cells were isolated and seeded to poly-L-lysine coated culture dishes after 15 min differential velocity adherence. Results: Compared with FBS medium, astrocytes in SF-I-H medium were smaller and exhibited process bearing morphologies. MTT assays showed that cell density and proliferation rate were higher in SF-I-H medium than in FBS medium all the time, and flow cytometry analysis revealed that SF-I-H medium promoted cell mitosis in a manner comparable to FBS medium. Consistently, western blot analysis further revealed that insulin and HB-EGF synergistically activated the PI3K/AKT and MAPK/ERK1/2 signaling cascades as FBS. Comparison with existing method(s): Astrocytes cultured in SF-I-H medium grew faster than FBS medium. Conclusion: Taken together, our results indicated that SF-I-H medium, in which cell morphology was similar with astrocytes in brain, was more effective for astrocyte survival and proliferation than FBS medium, providing a new cell model to study astrocyte functions without the interference of serum.

AB - Background: In vitro systems allowing maintenance and experimentation on primary astrocyte cultures have been used for decades. Astrocyte cultures are most maintained in serum-containing medium which has been found to alter the morphology and gene profiles of astrocytes. New method: Here, we reported a new serum-free medium for astrocyte culture, which consisted of DMEM and NB media supplemented with insulin and heparin-binding epidermal growth factor-like growth factor (HB-EGF) (SF-I-H medium). Meanwhile FBS-containing (FBS) medium composed of DMEM medium containing 10% FBS were used for comparison study. Cerebral cortex was harvested from postnatal day 1 Wistar rats and brain cells were isolated and seeded to poly-L-lysine coated culture dishes after 15 min differential velocity adherence. Results: Compared with FBS medium, astrocytes in SF-I-H medium were smaller and exhibited process bearing morphologies. MTT assays showed that cell density and proliferation rate were higher in SF-I-H medium than in FBS medium all the time, and flow cytometry analysis revealed that SF-I-H medium promoted cell mitosis in a manner comparable to FBS medium. Consistently, western blot analysis further revealed that insulin and HB-EGF synergistically activated the PI3K/AKT and MAPK/ERK1/2 signaling cascades as FBS. Comparison with existing method(s): Astrocytes cultured in SF-I-H medium grew faster than FBS medium. Conclusion: Taken together, our results indicated that SF-I-H medium, in which cell morphology was similar with astrocytes in brain, was more effective for astrocyte survival and proliferation than FBS medium, providing a new cell model to study astrocyte functions without the interference of serum.

KW - Astrocytes

KW - Cell proliferation

KW - Cell survival

KW - Heparin-binding epidermal growth factor-like growth factor

KW - Insulin

KW - Serum-free medium

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