Separation and purification of the bovine milk fat globule membrane protein and its effect on improvement of C<sub>2</sub>C<sub>12</sub>mouse skeletal muscle cell proliferation

Li, He; Xu, Weili; Ma, Ying; Zhou, Shaobo

doi:10.1039/c7nj00560a

Cited by 9 publications

(6 citation statements)

References 47 publications

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“…Mitochondrial function is directly involved in both cellular differentiation and apoptotic cell death [17]. These results were consistent with our previous research, which was found that MFG-E8 can increase the quantity of mitochondrion in C2C12 cells [7] .…”

Section: The Up-regulation Of Myoblast C2c12 Cell Proteinsupporting

confidence: 93%

“…The MFG-E8 protein was extracted according to the method reported previously [7].Briefly, a fresh milk sample (100 mL) containing sucrose (5 g) was centrifuged using an automatic cream separator, at 3500g at 38 °C for 30 min. The buttermilk was initially adjusted to pH 4.8 using 0.01 mol L −1 HCl in order to allow the MFGM to precipitate, and then neutralized with 0.01 mol L −1 NaOH.…”

Section: Preparation Of Mfg-e8mentioning

confidence: 99%

“…In vivo animal and human experiments have been conducted to study the effect and mechanism of MFGM supplementation. Our previous work has shown that MFG-E8 was isolated from MFGM and purified by an ion exchange column [7], LC-MS/MS was used to identify MFG-E8 as the main components in MFGM P2 (82.4%). MFG-E8 can promote C2C12 cell proliferation and differentiation based on the results of flow cytometry, laser confocal microscopy and transmission electron microscopy analyses.…”

Section: Introductionmentioning

confidence: 99%

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MFG-E8 induced differences in proteomic profiles in mouse C2C12 cells and its effect on PI3K/Akt and ERK signal pathways

Guan

et al. 2019

International Journal of Biological Macromolecules

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Milk fat globule-EGF factor 8 (MFG-E8) is one of the major proteins in milk fat globule membrane. In this study, mouse-derived C2C12 myoblast cells were served as an experimentally tractable model system for investigating the molecular basis of skeletal muscle cell specification and development. To examine the biochemical adaptations associated with myocytes formation comprehensively, a liquid chromatography coupled with tandem mass spectrometry label-free semi-quantitative approach was used to analyse the myogenic C2C12 proliferation program. Over 1987 proteins were identified in C2C12 cells. The MFG-E8 (200 g/mL) and MFG-E8 (500 g/mL) with significant differences were compared based on the relative abundance. The result profiles of regulation of MFG-E8 to the expression of proteins in C2C12 cells revealed that differential waves of expression of proteins linked to intracellular signaling, transcription, cytoarchitecture, adhesion, metabolism, and muscle contraction across during the C2C12 cell proliferation process. Based on the analysis of KEGG and STRING database, further to verification the expression of PI3K and ERK phosphorylation levels by Western blot. This study found that the data of proteomic was complementary to recent MFG-E8 studies of protein expression patterns in developing myotubes and provided a holistic framework for understanding how diverse biochemical processes are coordinated at the cellular level during skeletal muscle development.

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Section: The Up-regulation Of Myoblast C2c12 Cell Proteinsupporting

confidence: 93%

Section: Preparation Of Mfg-e8mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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MFG-E8 induced differences in proteomic profiles in mouse C2C12 cells and its effect on PI3K/Akt and ERK signal pathways

Guan

et al. 2019

International Journal of Biological Macromolecules

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“…MFGM was prepared as described in our previous study [20,21] with a slight modification. MFGM (0.2 g) was dissolved in PBS buffer (10 ml, pH 8.0), and the supernatant was applied to a DEAE Cellulose DE-52 (2.6×60 cm), followed by a…”

Section: Separation and Identification Of The Mfgm Proteinmentioning

confidence: 99%

Milk fat globule membrane protein promotes C2C12 cell proliferation through the PI3K/Akt signaling pathway

et al. 2018

International Journal of Biological Macromolecules

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Milk fat globule membrane (MFGM) protein is known to have several health benefits, including an anti-sarcopenia effect; however, its mechanism is unclear. The aim of this study was to investigate the potential mechanism of action of the MFGM protein. The MFGM protein was extracted and separated into 4 fractions, and Fraction 2 (57% of total MFGM) demonstrated the greatest effect on CC cell proliferation. Milk fat globule-EGF factor 8 (MFG-E8) accounted for 82.35% of the MFGM protein. The effects of whole Fraction 2 (100μg/mL, 200μg/mL and 300μg/mL) on cell proliferation and morphology were measured. Using qRT-PCR or a Western blot assay, several regulatory factors, e.g., PI3K P85α, p-pI3K p85α (Tyr 508), Akt, p-Akt (Ser 473), mTOR and p-mTOR (Ser 2448), were measured in cells incubated with 200μg/mL of Fraction 2 with or without wortmannin. The results demonstrated that Fraction 2 induced CC cell proliferation in a dose-dependent manner, upregulated the mRNA expression of mTOR and p70S6K, and activated PI3K, Akt, mTOR and P70S6K phosphorylation; however, Fraction 2 inhibited FOXO3a and 4E-BP. The results demonstrate that the MFGM protein, predominantly MFG-E8, promotes cell proliferation through the PI3K/Akt/mTOR signaling pathway. This study elucidated the molecular mechanism of the MFGM protein, primarily MFG-E8, in promoting CC cell proliferation via the PI3K/Akt/mTOR/P70S6K signal pathway.

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“…Following incubation, cells were washed with PBS (twice, 5 min each) and examined for condensed/fragmented nuclei under a fluorescence microscope (Leica IX71, Germany) at ×200 magnification. The transmission electron microscopy (TEM) of apoptotic HT-29 cells was performed as described previously 67 . HT-29 cells were respectively treated with each of five MFGM samples (100 μg/mL) for 72 h at 37 °C in 5% CO 2 /95% air.…”

Section: Methodsmentioning

confidence: 99%

Goat and buffalo milk fat globule membranes exhibit better effects at inducing apoptosis and reduction the viability of HT-29 cells

Cui

et al. 2019

Sci Rep

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Bovine milk fat globule membrane (MFGM) has shown many health benefits, however, there has not been much study on non-cattle MFGMs. The purpose of this study was to compare the anti-proliferation effects and investigate the mechanisms of MFGMs from bovine, goat, buffalo, yak and camel milk in HT-29 cells. Results showed that protein content in MFGM of yak milk is the highest among five MFGM. All MFGMs reduced cellular viability which was in agreement with cell morphology and apoptosis. However, the number of cells in S-phase from 24 h to 72 h was increased significantly by treatment with goat, buffalo and bovine MFGMs (100 μg/mL), but not yak and camel. All MFGMs treatment significantly reduced the mitochondrial membrane potential (with an order of goat > buffalo > bovine > camel > yak) and Bcl-2 expression, but increased the expression of both Bax and Caspase-3. Taken together, the results indicate that all MFGMs, especially goat and buffalo MFGMs, showed better effects at inducing apoptosis and reduction the viability of HT-29 cells. The mechanism might be arresting the cell cycle at S phase, depolarization of mitochondrial membrane potential, down-regulation of Bcl-2 expression and increase of Bax and Caspase-3 expression.

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Separation and purification of the bovine milk fat globule membrane protein and its effect on improvement of C₂C₁₂mouse skeletal muscle cell proliferation

Abstract: A novel method to improve the proliferation activity of C2C12cells by the bovine milk fat globule membrane (MFGM) protein was established in this study.

Cited by 9 publications

References 47 publications

MFG-E8 induced differences in proteomic profiles in mouse C2C12 cells and its effect on PI3K/Akt and ERK signal pathways

MFG-E8 induced differences in proteomic profiles in mouse C2C12 cells and its effect on PI3K/Akt and ERK signal pathways

Milk fat globule membrane protein promotes C2C12 cell proliferation through the PI3K/Akt signaling pathway

Goat and buffalo milk fat globule membranes exhibit better effects at inducing apoptosis and reduction the viability of HT-29 cells

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Separation and purification of the bovine milk fat globule membrane protein and its effect on improvement of C2C12mouse skeletal muscle cell proliferation

Abstract: A novel method to improve the proliferation activity of C2C12cells by the bovine milk fat globule membrane (MFGM) protein was established in this study.

Cited by 9 publications

References 47 publications

MFG-E8 induced differences in proteomic profiles in mouse C2C12 cells and its effect on PI3K/Akt and ERK signal pathways

MFG-E8 induced differences in proteomic profiles in mouse C2C12 cells and its effect on PI3K/Akt and ERK signal pathways

Milk fat globule membrane protein promotes C2C12 cell proliferation through the PI3K/Akt signaling pathway

Goat and buffalo milk fat globule membranes exhibit better effects at inducing apoptosis and reduction the viability of HT-29 cells

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Separation and purification of the bovine milk fat globule membrane protein and its effect on improvement of C₂C₁₂mouse skeletal muscle cell proliferation