Limits...
Angiogenic potential of endothelial progenitor cells and embryonic stem cells.

Rae PC, Kelly RD, Egginton S, St John JC - (2011)

Bottom Line: We also described the production of highly angiogenic EPC-comparable cells from pluripotent embryonic stem cells (ESCs) by direct differentiation using EC-conditioned medium (ECCM).ECCM-treated ESC-derived progenitor cells exhibited angiogenic potential, demonstrated by in vitro tubule formation, and endothelial-specific gene expression equivalent to natural EPCs.We concluded the effect of EPCs is cumulative and beneficial, relying on upregulation of the angiogenic activity of transplanted cells combined with an increase in proliferative cell number to produce significant effects upon transplantation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Clinical Sciences Research Institute, Warwick Medical School, University of Warwick, UK. justin.stjohn@monash.edu.

ABSTRACT

Background: Endothelial progenitor cells (EPCs) are implicated in a range of pathological conditions, suggesting a natural therapeutic role for EPCs in angiogenesis. However, current angiogenic therapies involving EPC transplantation are inefficient due to rejection of donor EPCs. One solution is to derive an expanded population of EPCs from stem cells in vitro, to be re-introduced as a therapeutic transplant. To demonstrate the therapeutic potential of EPCs we performed in vitro transplantation of EPCs into endothelial cell (EC) tubules using a gel-based tubule formation assay. We also described the production of highly angiogenic EPC-comparable cells from pluripotent embryonic stem cells (ESCs) by direct differentiation using EC-conditioned medium (ECCM).

Results: The effect on tubule complexity and longevity varied with transplantation quantity: significant effects were observed when tubules were transplanted with a quantity of EPCs equivalent to 50% of the number of ECs originally seeded on to the assay gel but not with 10% EPC transplantation. Gene expression of the endothelial markers VEGFR2, VE-cadherin and CD31, determined by qPCR, also changed dynamically during transplantation. ECCM-treated ESC-derived progenitor cells exhibited angiogenic potential, demonstrated by in vitro tubule formation, and endothelial-specific gene expression equivalent to natural EPCs.

Conclusions: We concluded the effect of EPCs is cumulative and beneficial, relying on upregulation of the angiogenic activity of transplanted cells combined with an increase in proliferative cell number to produce significant effects upon transplantation. Furthermore, EPCs derived from ESCs may be developed for use as a rapidly-expandable alternative for angiogenic transplantation therapy.

No MeSH data available.


Related in: MedlinePlus

Endothelial differentiation of stem cells. (A) VEGFR2, (B) VE-cadherin and (C) CD31 expression in ESCs, relative to 60% confluent ECs (± SEM; n = 3). Dotted line indicates expression in 60% confluent EPCs. Directed differentiation with ECCM, significant differences between treatments indicated (*P < 0.05, **P < 0.01). (D): VEGFR2, CD133 and CD34 proteins in ECCM-treated ESCs. Scale bars = 10 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3108917&req=5

Figure 4: Endothelial differentiation of stem cells. (A) VEGFR2, (B) VE-cadherin and (C) CD31 expression in ESCs, relative to 60% confluent ECs (± SEM; n = 3). Dotted line indicates expression in 60% confluent EPCs. Directed differentiation with ECCM, significant differences between treatments indicated (*P < 0.05, **P < 0.01). (D): VEGFR2, CD133 and CD34 proteins in ECCM-treated ESCs. Scale bars = 10 μm.

Mentions: To determine whether ESCs could give rise to endothelial-like cells, we differentiated D3 ESCs over 28 days using spontaneous and directed methods employing the hanging droplet technique. VEGFR2 was detected in ESCs from day (D)1 onwards in both approaches. VEGFR2 expression exceeded levels observed in ECs after D4 in spontaneously-differentiated ESCs (P < 0.01) and D2 in directed-differentiated ESCs (P < 0.05; Figure 4a). Expression of VEGFR2 in ESCs was significantly different between differentiation treatments from D1 to D6. At D7 and D14, directed- and spontaneously-differentiated ESCs, respectively, expressed levels of VEGFR2 similar to EPCs and this continued for the remainder of the culture period. VE-cadherin expression was low throughout differentiation of ESCs, with expression not reaching a level comparable to either EPCs or ECs in either treatment before D28 (P > 0.05; Figure 4b). There was no significant difference between differentiation treatments. CD31 transcripts were present in spontaneously- and directed-differentiated ESCs from D1 with expression exceeding EC-equivalent levels at D7 (P < 0.05) and D5 (P < 0.01), respectively (Figure 4c). CD31 expression in D21 and D28 ESCs (using both treatments) was similar to that in EPCs. Furthermore, CD31 expression was significantly different between ESC treatments from D2 to D14 (P < 0.05). ICC confirmed that protein was present for all three genes throughout differentiation for both treatments (Figure 4d).


Angiogenic potential of endothelial progenitor cells and embryonic stem cells.

Rae PC, Kelly RD, Egginton S, St John JC - (2011)

Endothelial differentiation of stem cells. (A) VEGFR2, (B) VE-cadherin and (C) CD31 expression in ESCs, relative to 60% confluent ECs (± SEM; n = 3). Dotted line indicates expression in 60% confluent EPCs. Directed differentiation with ECCM, significant differences between treatments indicated (*P < 0.05, **P < 0.01). (D): VEGFR2, CD133 and CD34 proteins in ECCM-treated ESCs. Scale bars = 10 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3108917&req=5

Figure 4: Endothelial differentiation of stem cells. (A) VEGFR2, (B) VE-cadherin and (C) CD31 expression in ESCs, relative to 60% confluent ECs (± SEM; n = 3). Dotted line indicates expression in 60% confluent EPCs. Directed differentiation with ECCM, significant differences between treatments indicated (*P < 0.05, **P < 0.01). (D): VEGFR2, CD133 and CD34 proteins in ECCM-treated ESCs. Scale bars = 10 μm.
Mentions: To determine whether ESCs could give rise to endothelial-like cells, we differentiated D3 ESCs over 28 days using spontaneous and directed methods employing the hanging droplet technique. VEGFR2 was detected in ESCs from day (D)1 onwards in both approaches. VEGFR2 expression exceeded levels observed in ECs after D4 in spontaneously-differentiated ESCs (P < 0.01) and D2 in directed-differentiated ESCs (P < 0.05; Figure 4a). Expression of VEGFR2 in ESCs was significantly different between differentiation treatments from D1 to D6. At D7 and D14, directed- and spontaneously-differentiated ESCs, respectively, expressed levels of VEGFR2 similar to EPCs and this continued for the remainder of the culture period. VE-cadherin expression was low throughout differentiation of ESCs, with expression not reaching a level comparable to either EPCs or ECs in either treatment before D28 (P > 0.05; Figure 4b). There was no significant difference between differentiation treatments. CD31 transcripts were present in spontaneously- and directed-differentiated ESCs from D1 with expression exceeding EC-equivalent levels at D7 (P < 0.05) and D5 (P < 0.01), respectively (Figure 4c). CD31 expression in D21 and D28 ESCs (using both treatments) was similar to that in EPCs. Furthermore, CD31 expression was significantly different between ESC treatments from D2 to D14 (P < 0.05). ICC confirmed that protein was present for all three genes throughout differentiation for both treatments (Figure 4d).

Bottom Line: We also described the production of highly angiogenic EPC-comparable cells from pluripotent embryonic stem cells (ESCs) by direct differentiation using EC-conditioned medium (ECCM).ECCM-treated ESC-derived progenitor cells exhibited angiogenic potential, demonstrated by in vitro tubule formation, and endothelial-specific gene expression equivalent to natural EPCs.We concluded the effect of EPCs is cumulative and beneficial, relying on upregulation of the angiogenic activity of transplanted cells combined with an increase in proliferative cell number to produce significant effects upon transplantation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Clinical Sciences Research Institute, Warwick Medical School, University of Warwick, UK. justin.stjohn@monash.edu.

ABSTRACT

Background: Endothelial progenitor cells (EPCs) are implicated in a range of pathological conditions, suggesting a natural therapeutic role for EPCs in angiogenesis. However, current angiogenic therapies involving EPC transplantation are inefficient due to rejection of donor EPCs. One solution is to derive an expanded population of EPCs from stem cells in vitro, to be re-introduced as a therapeutic transplant. To demonstrate the therapeutic potential of EPCs we performed in vitro transplantation of EPCs into endothelial cell (EC) tubules using a gel-based tubule formation assay. We also described the production of highly angiogenic EPC-comparable cells from pluripotent embryonic stem cells (ESCs) by direct differentiation using EC-conditioned medium (ECCM).

Results: The effect on tubule complexity and longevity varied with transplantation quantity: significant effects were observed when tubules were transplanted with a quantity of EPCs equivalent to 50% of the number of ECs originally seeded on to the assay gel but not with 10% EPC transplantation. Gene expression of the endothelial markers VEGFR2, VE-cadherin and CD31, determined by qPCR, also changed dynamically during transplantation. ECCM-treated ESC-derived progenitor cells exhibited angiogenic potential, demonstrated by in vitro tubule formation, and endothelial-specific gene expression equivalent to natural EPCs.

Conclusions: We concluded the effect of EPCs is cumulative and beneficial, relying on upregulation of the angiogenic activity of transplanted cells combined with an increase in proliferative cell number to produce significant effects upon transplantation. Furthermore, EPCs derived from ESCs may be developed for use as a rapidly-expandable alternative for angiogenic transplantation therapy.

No MeSH data available.


Related in: MedlinePlus