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Dermal fibroblasts display similar phenotypic and differentiation capacity to fat-derived mesenchymal stem cells, but differ in anti-inflammatory and angiogenic potential.

Blasi A, Martino C, Balducci L, Saldarelli M, Soleti A, Navone SE, Canzi L, Cristini S, Invernici G, Parati EA, Alessandri G - (2011)

Bottom Line: AD-MSCs released significant amounts of VEGF, HGF and Angiopoietins and their conditioned medium (CM) stimulated ECs proliferation and tube formations.These results underline the importance of evaluating the angiogenic and anti-inflammatory features of MSCs preparation.Their priming with inflammatory cytokines prior to transplantation may improve their efficacy in cell-based therapies for tissue regeneration.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cellular Neurobiology Laboratory, Department of Cerebrovascular Diseases, Fondazione IRCCS Neurological Institute "Carlo Besta", 20133 Milan, Italy. cisiamo2@yahoo.com.

ABSTRACT

Background: Mesenchymal stem cells (MSCs) are multipotent stem cells able to differentiate into different cell lineages. However, MSCs represent a subpopulation of a more complex cell composition of stroma cells contained in mesenchymal tissue. Due to a lack of specific markers, it is difficult to distinguish MSCs from other more mature stromal cells such as fibroblasts, which, conversely, are abundant in mesenchymal tissue. In order to find more distinguishing features between MSCs and fibroblasts, we studied the phenotypic and functional features of human adipose-derived MSCs (AD-MSCs) side by side with normal human dermal fibroblasts (HNDFs) in vitro

Methods: AD-MSCs and HNDFs were cultured, expanded and phenotypically characterized by flow cytometry (FC). Immunofluorescence was used to investigate cell differentiation. ELISA assay was used to quantify angiogenic factors and chemokines release. Cultures of endothelial cells (ECs) and a monocyte cell line, U937, were used to test angiogenic and anti-inflammatory properties.

Results: Cultured AD-MSCs and HNDFs display similar morphological appearance, growth rate, and phenotypic profile. They both expressed typical mesenchymal markers-CD90, CD29, CD44, CD105 and to a minor extent, the adhesion molecules CD54, CD56, CD106 and CD166. They were negative for the stem cell markers CD34, CD146, CD133, CD117. Only aldehyde dehydrogenase (ALDH) was expressed. Neither AD-MSCs nor HNDFs differed in their multi-lineage differentiation capacity; they both differentiated into osteoblast, adipocyte, and also into cardiomyocyte-like cells. In contrast, AD-MSCs, but not HNDFs, displayed strong angiogenic and anti-inflammatory activity. AD-MSCs released significant amounts of VEGF, HGF and Angiopoietins and their conditioned medium (CM) stimulated ECs proliferation and tube formations. In addition, CM-derived AD-MSCs (AD-MSCs-CM) inhibited adhesion molecules expression on U937 and release of RANTES and MCP-1. Finally, after priming with TNFα, AD-MSCs enhanced their anti-inflammatory potential; while HNDFs acquired pro-inflammatory activity.

Conclusions: AD-MSCs cannot be distinguished from HNDFs in vitro by evaluating their phenotypic profile or differentiation potential, but only through the analysis of their anti-inflammatory and angiogenic properties. These results underline the importance of evaluating the angiogenic and anti-inflammatory features of MSCs preparation. Their priming with inflammatory cytokines prior to transplantation may improve their efficacy in cell-based therapies for tissue regeneration.

No MeSH data available.


Related in: MedlinePlus

AD-MSCs, but not HNDFs, reduced expression of AMs on U937. AD-MSCs-CM and HNDFs-CM were tested on the adhesion molecules expression of the U937 monocyte cell line stimulated or not with TNFα (25 ng/ml × 12 hrs). Mean Fluorescent Intensity (MFI) of each marker expression was evaluated by FACS and values are the mean ± SD of independent experiments performed with 3 different preparations of AD-MSCs-CM and HNDFs-CM. In (A) the expression of CD54/ICAM-1, (B) the expression of CD44, (C) the expression of CD42L/L-selectin and in (D) the expression of CD49d/VLA-4. Note that, both AD-MSCs-CM and HNDFs-CM did not affect basal adhesion molecules expression on U937. However, AD-MSCs-CM, but not HNDFs-CM (at 1:1), inhibited CD54/ICAM-1, CD44 and CD42L/L-selectin up-modulation produced by TNFα stimuli. CD49d expression was not affected by either AD-MSCs-CM or HNDFs-CM. * p < 0.05 and **p < 0.01 versus CTRL.
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Figure 4: AD-MSCs, but not HNDFs, reduced expression of AMs on U937. AD-MSCs-CM and HNDFs-CM were tested on the adhesion molecules expression of the U937 monocyte cell line stimulated or not with TNFα (25 ng/ml × 12 hrs). Mean Fluorescent Intensity (MFI) of each marker expression was evaluated by FACS and values are the mean ± SD of independent experiments performed with 3 different preparations of AD-MSCs-CM and HNDFs-CM. In (A) the expression of CD54/ICAM-1, (B) the expression of CD44, (C) the expression of CD42L/L-selectin and in (D) the expression of CD49d/VLA-4. Note that, both AD-MSCs-CM and HNDFs-CM did not affect basal adhesion molecules expression on U937. However, AD-MSCs-CM, but not HNDFs-CM (at 1:1), inhibited CD54/ICAM-1, CD44 and CD42L/L-selectin up-modulation produced by TNFα stimuli. CD49d expression was not affected by either AD-MSCs-CM or HNDFs-CM. * p < 0.05 and **p < 0.01 versus CTRL.

Mentions: It has been shown that MSCs display anti-inflammatory capacity [15,23,24]. We thus investigated whether AD-MSCs and HNDFs may produce molecules with anti-inflammatory activity by testing their CM on U937, a human monocyte cell line, under basal culture conditions or in the presence of TNFα inflammatory stimuli. As shown in Figure 4, the expression of the adhesion molecules CD54, CD44, CD62L, CD49d on U937 was not significantly affected by the addition of both AD-MSCs-CM and HNDFs-CM to the basal control medium. After stimulation of U937 with TNFα (25 ng/ml), expression of CD54 (Figure 4A), CD44 (Figure 4B) and CD62L (Figure 4C) was increased and the addition of AD-MSCs-CM (at 1:1 dilution) was able to antagonize the increment (Figure 4A, B and 4C). In contrast, the addition of HNDFs-CM did not affect adhesion molecules increment induced by TNFα; only CD54 expression was slightly reduced (Figure 4A). Neither AD-MSCs-CM nor HNDFs-CM had any affect on CD49d expression (Figure 4D).


Dermal fibroblasts display similar phenotypic and differentiation capacity to fat-derived mesenchymal stem cells, but differ in anti-inflammatory and angiogenic potential.

Blasi A, Martino C, Balducci L, Saldarelli M, Soleti A, Navone SE, Canzi L, Cristini S, Invernici G, Parati EA, Alessandri G - (2011)

AD-MSCs, but not HNDFs, reduced expression of AMs on U937. AD-MSCs-CM and HNDFs-CM were tested on the adhesion molecules expression of the U937 monocyte cell line stimulated or not with TNFα (25 ng/ml × 12 hrs). Mean Fluorescent Intensity (MFI) of each marker expression was evaluated by FACS and values are the mean ± SD of independent experiments performed with 3 different preparations of AD-MSCs-CM and HNDFs-CM. In (A) the expression of CD54/ICAM-1, (B) the expression of CD44, (C) the expression of CD42L/L-selectin and in (D) the expression of CD49d/VLA-4. Note that, both AD-MSCs-CM and HNDFs-CM did not affect basal adhesion molecules expression on U937. However, AD-MSCs-CM, but not HNDFs-CM (at 1:1), inhibited CD54/ICAM-1, CD44 and CD42L/L-selectin up-modulation produced by TNFα stimuli. CD49d expression was not affected by either AD-MSCs-CM or HNDFs-CM. * p < 0.05 and **p < 0.01 versus CTRL.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 4: AD-MSCs, but not HNDFs, reduced expression of AMs on U937. AD-MSCs-CM and HNDFs-CM were tested on the adhesion molecules expression of the U937 monocyte cell line stimulated or not with TNFα (25 ng/ml × 12 hrs). Mean Fluorescent Intensity (MFI) of each marker expression was evaluated by FACS and values are the mean ± SD of independent experiments performed with 3 different preparations of AD-MSCs-CM and HNDFs-CM. In (A) the expression of CD54/ICAM-1, (B) the expression of CD44, (C) the expression of CD42L/L-selectin and in (D) the expression of CD49d/VLA-4. Note that, both AD-MSCs-CM and HNDFs-CM did not affect basal adhesion molecules expression on U937. However, AD-MSCs-CM, but not HNDFs-CM (at 1:1), inhibited CD54/ICAM-1, CD44 and CD42L/L-selectin up-modulation produced by TNFα stimuli. CD49d expression was not affected by either AD-MSCs-CM or HNDFs-CM. * p < 0.05 and **p < 0.01 versus CTRL.
Mentions: It has been shown that MSCs display anti-inflammatory capacity [15,23,24]. We thus investigated whether AD-MSCs and HNDFs may produce molecules with anti-inflammatory activity by testing their CM on U937, a human monocyte cell line, under basal culture conditions or in the presence of TNFα inflammatory stimuli. As shown in Figure 4, the expression of the adhesion molecules CD54, CD44, CD62L, CD49d on U937 was not significantly affected by the addition of both AD-MSCs-CM and HNDFs-CM to the basal control medium. After stimulation of U937 with TNFα (25 ng/ml), expression of CD54 (Figure 4A), CD44 (Figure 4B) and CD62L (Figure 4C) was increased and the addition of AD-MSCs-CM (at 1:1 dilution) was able to antagonize the increment (Figure 4A, B and 4C). In contrast, the addition of HNDFs-CM did not affect adhesion molecules increment induced by TNFα; only CD54 expression was slightly reduced (Figure 4A). Neither AD-MSCs-CM nor HNDFs-CM had any affect on CD49d expression (Figure 4D).

Bottom Line: AD-MSCs released significant amounts of VEGF, HGF and Angiopoietins and their conditioned medium (CM) stimulated ECs proliferation and tube formations.These results underline the importance of evaluating the angiogenic and anti-inflammatory features of MSCs preparation.Their priming with inflammatory cytokines prior to transplantation may improve their efficacy in cell-based therapies for tissue regeneration.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cellular Neurobiology Laboratory, Department of Cerebrovascular Diseases, Fondazione IRCCS Neurological Institute "Carlo Besta", 20133 Milan, Italy. cisiamo2@yahoo.com.

ABSTRACT

Background: Mesenchymal stem cells (MSCs) are multipotent stem cells able to differentiate into different cell lineages. However, MSCs represent a subpopulation of a more complex cell composition of stroma cells contained in mesenchymal tissue. Due to a lack of specific markers, it is difficult to distinguish MSCs from other more mature stromal cells such as fibroblasts, which, conversely, are abundant in mesenchymal tissue. In order to find more distinguishing features between MSCs and fibroblasts, we studied the phenotypic and functional features of human adipose-derived MSCs (AD-MSCs) side by side with normal human dermal fibroblasts (HNDFs) in vitro

Methods: AD-MSCs and HNDFs were cultured, expanded and phenotypically characterized by flow cytometry (FC). Immunofluorescence was used to investigate cell differentiation. ELISA assay was used to quantify angiogenic factors and chemokines release. Cultures of endothelial cells (ECs) and a monocyte cell line, U937, were used to test angiogenic and anti-inflammatory properties.

Results: Cultured AD-MSCs and HNDFs display similar morphological appearance, growth rate, and phenotypic profile. They both expressed typical mesenchymal markers-CD90, CD29, CD44, CD105 and to a minor extent, the adhesion molecules CD54, CD56, CD106 and CD166. They were negative for the stem cell markers CD34, CD146, CD133, CD117. Only aldehyde dehydrogenase (ALDH) was expressed. Neither AD-MSCs nor HNDFs differed in their multi-lineage differentiation capacity; they both differentiated into osteoblast, adipocyte, and also into cardiomyocyte-like cells. In contrast, AD-MSCs, but not HNDFs, displayed strong angiogenic and anti-inflammatory activity. AD-MSCs released significant amounts of VEGF, HGF and Angiopoietins and their conditioned medium (CM) stimulated ECs proliferation and tube formations. In addition, CM-derived AD-MSCs (AD-MSCs-CM) inhibited adhesion molecules expression on U937 and release of RANTES and MCP-1. Finally, after priming with TNFα, AD-MSCs enhanced their anti-inflammatory potential; while HNDFs acquired pro-inflammatory activity.

Conclusions: AD-MSCs cannot be distinguished from HNDFs in vitro by evaluating their phenotypic profile or differentiation potential, but only through the analysis of their anti-inflammatory and angiogenic properties. These results underline the importance of evaluating the angiogenic and anti-inflammatory features of MSCs preparation. Their priming with inflammatory cytokines prior to transplantation may improve their efficacy in cell-based therapies for tissue regeneration.

No MeSH data available.


Related in: MedlinePlus