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Characterization of human mesenchymal stem cell secretome at early steps of adipocyte and osteoblast differentiation.

Chiellini C, Cochet O, Negroni L, Samson M, Poggi M, Ailhaud G, Alessi MC, Dani C, Amri EZ - BMC Mol. Biol. (2008)

Bottom Line: Analysis of identified proteins showed that 52 % corresponded to classical secreted proteins characterized by a signal peptide, that 37 % previously described in the extracellular compartment were devoid of signal peptide and that 11 % neither exhibited a signal peptide nor had been previously described extracellularly.Quantitative analysis has been performed for 8 candidates: PAI-1, PEDF, BIGH3, PTX3, SPARC, ENO1, GRP78 and MMP2.Among them, PAI-1 was detected at day 0 and day 3 of osteoblast differentiation but never in adipocyte secretome.

View Article: PubMed Central - HTML - PubMed

Affiliation: ISBDC, Université de Nice Sophia-Antipolis, CNRS ; 28 avenue de Valrose, 06100 Nice, France. Chiara.Chiellini@gmail.com

ABSTRACT

Background: It is well established that adipose tissue plays a key role in energy storage and release but is also a secretory organ and a source of stem cells. Among different lineages, stem cells are able to differentiate into adipocytes and osteoblasts. As secreted proteins could regulate the balance between both lineages, we aimed at characterizing the secretome of human multipotent adipose-derived stem cell (hMADS) at an early step of commitment to adipocytes and osteoblasts.

Results: A proteomic approach, using mono-dimensional electrophoresis and tandem mass spectrometry, allowed us to identify a total of 73 proteins at day 0 and day 3 of adipocyte and osteoblast differentiation. Analysis of identified proteins showed that 52 % corresponded to classical secreted proteins characterized by a signal peptide, that 37 % previously described in the extracellular compartment were devoid of signal peptide and that 11 % neither exhibited a signal peptide nor had been previously described extracellularly. These proteins were classified into 8 clusters according to their function. Quantitative analysis has been performed for 8 candidates: PAI-1, PEDF, BIGH3, PTX3, SPARC, ENO1, GRP78 and MMP2. Among them, PAI-1 was detected at day 0 and day 3 of osteoblast differentiation but never in adipocyte secretome. Furthermore we showed that PAI-1 mRNA was down-regulated in the bone of ovariectomized mice.

Conclusion: Given its regulation during the early events of hMADS cell differentiation and its status in ovariectomized mice, PAI-1 could play a role in the adipocyte/osteoblast balance and thus in bone diseases such as osteoporosis.

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Related in: MedlinePlus

Analysis of terminal differentiation of hMADS cells into adipocytes and osteoblasts and Coomassie Blue staining of hMADS cell secretome. A. qRT-PCR analysis of mRNA levels of specific adipogenic (adiponectin) and osteogenic (alkaline phosphatase) markers at day 3 and day 14 of differentiation as compared to day 0. The data are representative of three independent experiments. B. Microphotographs of hMADS cells differentiated into adipocytes and osteoblasts at day 14. Bar scale = 50 μm. C. Representative gel of secreted proteins from hMADS cells at day 0 and day 3 of differentiating adipocytes (adipo) and osteoblasts (osteo) after 6 h of incubation. The gel is representative of 3 independent experiments.
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Figure 1: Analysis of terminal differentiation of hMADS cells into adipocytes and osteoblasts and Coomassie Blue staining of hMADS cell secretome. A. qRT-PCR analysis of mRNA levels of specific adipogenic (adiponectin) and osteogenic (alkaline phosphatase) markers at day 3 and day 14 of differentiation as compared to day 0. The data are representative of three independent experiments. B. Microphotographs of hMADS cells differentiated into adipocytes and osteoblasts at day 14. Bar scale = 50 μm. C. Representative gel of secreted proteins from hMADS cells at day 0 and day 3 of differentiating adipocytes (adipo) and osteoblasts (osteo) after 6 h of incubation. The gel is representative of 3 independent experiments.

Mentions: hMADS cells differentiate into adipocytes and osteoblasts, as shown in Figure 1A. Gene expression of representative markers of adipocyte (adiponectin) and osteoblast differentiation (alkaline phosphatase) are reported and in agreement with previously published data [14-17]. The terminal differentiation of hMADS cells into adipocytes and osteoblasts is also illustrated by the typical cellular morphology as shown in Figure 1B.


Characterization of human mesenchymal stem cell secretome at early steps of adipocyte and osteoblast differentiation.

Chiellini C, Cochet O, Negroni L, Samson M, Poggi M, Ailhaud G, Alessi MC, Dani C, Amri EZ - BMC Mol. Biol. (2008)

Analysis of terminal differentiation of hMADS cells into adipocytes and osteoblasts and Coomassie Blue staining of hMADS cell secretome. A. qRT-PCR analysis of mRNA levels of specific adipogenic (adiponectin) and osteogenic (alkaline phosphatase) markers at day 3 and day 14 of differentiation as compared to day 0. The data are representative of three independent experiments. B. Microphotographs of hMADS cells differentiated into adipocytes and osteoblasts at day 14. Bar scale = 50 μm. C. Representative gel of secreted proteins from hMADS cells at day 0 and day 3 of differentiating adipocytes (adipo) and osteoblasts (osteo) after 6 h of incubation. The gel is representative of 3 independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2279142&req=5

Figure 1: Analysis of terminal differentiation of hMADS cells into adipocytes and osteoblasts and Coomassie Blue staining of hMADS cell secretome. A. qRT-PCR analysis of mRNA levels of specific adipogenic (adiponectin) and osteogenic (alkaline phosphatase) markers at day 3 and day 14 of differentiation as compared to day 0. The data are representative of three independent experiments. B. Microphotographs of hMADS cells differentiated into adipocytes and osteoblasts at day 14. Bar scale = 50 μm. C. Representative gel of secreted proteins from hMADS cells at day 0 and day 3 of differentiating adipocytes (adipo) and osteoblasts (osteo) after 6 h of incubation. The gel is representative of 3 independent experiments.
Mentions: hMADS cells differentiate into adipocytes and osteoblasts, as shown in Figure 1A. Gene expression of representative markers of adipocyte (adiponectin) and osteoblast differentiation (alkaline phosphatase) are reported and in agreement with previously published data [14-17]. The terminal differentiation of hMADS cells into adipocytes and osteoblasts is also illustrated by the typical cellular morphology as shown in Figure 1B.

Bottom Line: Analysis of identified proteins showed that 52 % corresponded to classical secreted proteins characterized by a signal peptide, that 37 % previously described in the extracellular compartment were devoid of signal peptide and that 11 % neither exhibited a signal peptide nor had been previously described extracellularly.Quantitative analysis has been performed for 8 candidates: PAI-1, PEDF, BIGH3, PTX3, SPARC, ENO1, GRP78 and MMP2.Among them, PAI-1 was detected at day 0 and day 3 of osteoblast differentiation but never in adipocyte secretome.

View Article: PubMed Central - HTML - PubMed

Affiliation: ISBDC, Université de Nice Sophia-Antipolis, CNRS ; 28 avenue de Valrose, 06100 Nice, France. Chiara.Chiellini@gmail.com

ABSTRACT

Background: It is well established that adipose tissue plays a key role in energy storage and release but is also a secretory organ and a source of stem cells. Among different lineages, stem cells are able to differentiate into adipocytes and osteoblasts. As secreted proteins could regulate the balance between both lineages, we aimed at characterizing the secretome of human multipotent adipose-derived stem cell (hMADS) at an early step of commitment to adipocytes and osteoblasts.

Results: A proteomic approach, using mono-dimensional electrophoresis and tandem mass spectrometry, allowed us to identify a total of 73 proteins at day 0 and day 3 of adipocyte and osteoblast differentiation. Analysis of identified proteins showed that 52 % corresponded to classical secreted proteins characterized by a signal peptide, that 37 % previously described in the extracellular compartment were devoid of signal peptide and that 11 % neither exhibited a signal peptide nor had been previously described extracellularly. These proteins were classified into 8 clusters according to their function. Quantitative analysis has been performed for 8 candidates: PAI-1, PEDF, BIGH3, PTX3, SPARC, ENO1, GRP78 and MMP2. Among them, PAI-1 was detected at day 0 and day 3 of osteoblast differentiation but never in adipocyte secretome. Furthermore we showed that PAI-1 mRNA was down-regulated in the bone of ovariectomized mice.

Conclusion: Given its regulation during the early events of hMADS cell differentiation and its status in ovariectomized mice, PAI-1 could play a role in the adipocyte/osteoblast balance and thus in bone diseases such as osteoporosis.

Show MeSH
Related in: MedlinePlus