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Tendon proper- and peritenon-derived progenitor cells have unique tenogenic properties.

Mienaltowski MJ, Adams SM, Birk DE - Stem Cell Res Ther (2014)

Bottom Line: Tendon construct ultrastructure was also compared after 45 days.It also was found that peritenon-derived progenitors secrete factor(s) stimulatory to tenocytes and tendon proper progenitors.Data demonstrate that, relative to peritenon-derived progenitors, tendon proper progenitors have greater potential for forming functional tendon-like tissue.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: Multipotent progenitor populations exist within the tendon proper and peritenon of the Achilles tendon. Progenitor populations derived from the tendon proper and peritenon are enriched with distinct cell types that are distinguished by expression of markers of tendon and vascular or pericyte origins, respectively. The objective of this study was to discern the unique tenogenic properties of tendon proper- and peritenon-derived progenitors within an in vitro model. We hypothesized that progenitors from each region contribute differently to tendon formation; thus, when incorporated into a regenerative model, progenitors from each region will respond uniquely. Moreover, we hypothesized that cell populations like progenitors were capable of stimulating tenogenic differentiation, so we generated conditioned media from these cell types to analyze their stimulatory potentials.

Methods: Isolated progenitors were seeded within fibrinogen/thrombin gel-based constructs with or without supplementation with recombinant growth/differentiation factor-5 (GDF5). Early and late in culture, gene expression of differentiation markers and matrix assembly genes was analyzed. Tendon construct ultrastructure was also compared after 45 days. Moreover, conditioned media from tendon proper-derived progenitors, peritenon-derived progenitors, or tenocytes was applied to each of the three cell types to determine paracrine stimulatory effects of the factors secreted from each of the respective cell types.

Results: The cell orientation, extracellular domain and fibril organization of constructs were comparable to embryonic tendon. The tendon proper-derived progenitors produced a more tendon-like construct than the peritenon-derived progenitors. Seeded tendon proper-derived progenitors expressed greater levels of tenogenic markers and matrix assembly genes, relative to peritenon-derived progenitors. However, GDF5 supplementation improved expression of matrix assembly genes in peritenon progenitors and structurally led to increased mean fibril diameters. It also was found that peritenon-derived progenitors secrete factor(s) stimulatory to tenocytes and tendon proper progenitors.

Conclusions: Data demonstrate that, relative to peritenon-derived progenitors, tendon proper progenitors have greater potential for forming functional tendon-like tissue. Furthermore, factors secreted by peritenon-derived progenitors suggest a trophic role for this cell type as well. Thus, these findings highlight the synergistic potential of including these progenitor populations in restorative tendon engineering strategies.

No MeSH data available.


GDF5 supplementation causes improved expression matrix assembly genes in peritenon progenitors seeded in constructs. After r-mGDF5 supplementation, expression levels for the tendon markers Scx(A) and Tnmd(B) and the vascular marker Emcn(C) still differed. GDF supplementation led to improved expression of matrix assembly genes for peritenon progenitors seeded in tendon constructs - comparing PERI + versus TP at Day 7 (early) and Day 45 (late). Bolstered expression is noted for SLRPs Bgn(D) and early for Dcn(E), for FACIT collagens Col12a1(F) and Col14a1(G), and for fibril-forming collagens Col1a1(H), Col3a1(I) and Col5a1(J). Expression of Col11a1(K) is still greater for tendon proper-derived progenitors. (Biological replicates, n = 5 to 8; Mann–Whitney-Wilcoxon test – P <0.01, *; P <0.05, #; fold changes and statistical significance further reported in Table 3 and expanded results are reported in Additional file 1: Figure S1). GDF5, growth differentiation factor 5; PERI, peritenon-derived progenitors; SLRP, small leucine-rich proteoglycans; TP, tendon proper-derived progenitors.
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Figure 5: GDF5 supplementation causes improved expression matrix assembly genes in peritenon progenitors seeded in constructs. After r-mGDF5 supplementation, expression levels for the tendon markers Scx(A) and Tnmd(B) and the vascular marker Emcn(C) still differed. GDF supplementation led to improved expression of matrix assembly genes for peritenon progenitors seeded in tendon constructs - comparing PERI + versus TP at Day 7 (early) and Day 45 (late). Bolstered expression is noted for SLRPs Bgn(D) and early for Dcn(E), for FACIT collagens Col12a1(F) and Col14a1(G), and for fibril-forming collagens Col1a1(H), Col3a1(I) and Col5a1(J). Expression of Col11a1(K) is still greater for tendon proper-derived progenitors. (Biological replicates, n = 5 to 8; Mann–Whitney-Wilcoxon test – P <0.01, *; P <0.05, #; fold changes and statistical significance further reported in Table 3 and expanded results are reported in Additional file 1: Figure S1). GDF5, growth differentiation factor 5; PERI, peritenon-derived progenitors; SLRP, small leucine-rich proteoglycans; TP, tendon proper-derived progenitors.

Mentions: When the culture was supplemented with recombinant mouse GDF5, expression for matrix assembly genes in seeded peritenon progenitors increased to levels seen with the seeded tendon proper-derived progenitors (Table 3). However, after GDF5 supplementation, expression levels for the tendon markers Scx (Figure 5A) and Tnmd (Figure 5B) were unchanged when comparing GDF5-supplemented peritenon progenitor expression (PERI+) to that of tendon proper-derived progenitors not supplemented with GDF5 (TP). Expression of vascular marker Emcn (Figure 5C) also still differed between PERI + and TP progenitors. GDF5 supplementation did result in improved expression of matrix assembly genes for peritenon progenitors seeded in tendon constructs comparing PERI + progenitors and TP progenitors. Bolstered expression is noted for the small leucine-rich proteoglycans (SLRPs) Bgn early (Figure 5D) and Dcn (Figure 5E). Expression of FACIT collagens Col12a1 (Figure 5F) and Col14a1 (Figure 5G) as well as fibril-forming collagens Col1a1 (Figure 5H), Col3a1 (Figure 5I), and Col5a1 (Figure 5J) for GDF5-supplemented PERI + progenitors also became comparable to TP progenitors, yet expression of Col11a1 (Figure 5K) was still greater for TP progenitors. Addition of GDF5 did not lead to significant expression changes among TP progenitors for the tendon markers and matrix assembly genes analyzed, except for Bgn at 45 days; Additional file 1: Figure S1 exhibits expression results for constructs generated by cell type, GDF5 supplementation and culture time.


Tendon proper- and peritenon-derived progenitor cells have unique tenogenic properties.

Mienaltowski MJ, Adams SM, Birk DE - Stem Cell Res Ther (2014)

GDF5 supplementation causes improved expression matrix assembly genes in peritenon progenitors seeded in constructs. After r-mGDF5 supplementation, expression levels for the tendon markers Scx(A) and Tnmd(B) and the vascular marker Emcn(C) still differed. GDF supplementation led to improved expression of matrix assembly genes for peritenon progenitors seeded in tendon constructs - comparing PERI + versus TP at Day 7 (early) and Day 45 (late). Bolstered expression is noted for SLRPs Bgn(D) and early for Dcn(E), for FACIT collagens Col12a1(F) and Col14a1(G), and for fibril-forming collagens Col1a1(H), Col3a1(I) and Col5a1(J). Expression of Col11a1(K) is still greater for tendon proper-derived progenitors. (Biological replicates, n = 5 to 8; Mann–Whitney-Wilcoxon test – P <0.01, *; P <0.05, #; fold changes and statistical significance further reported in Table 3 and expanded results are reported in Additional file 1: Figure S1). GDF5, growth differentiation factor 5; PERI, peritenon-derived progenitors; SLRP, small leucine-rich proteoglycans; TP, tendon proper-derived progenitors.
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Related In: Results  -  Collection

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Figure 5: GDF5 supplementation causes improved expression matrix assembly genes in peritenon progenitors seeded in constructs. After r-mGDF5 supplementation, expression levels for the tendon markers Scx(A) and Tnmd(B) and the vascular marker Emcn(C) still differed. GDF supplementation led to improved expression of matrix assembly genes for peritenon progenitors seeded in tendon constructs - comparing PERI + versus TP at Day 7 (early) and Day 45 (late). Bolstered expression is noted for SLRPs Bgn(D) and early for Dcn(E), for FACIT collagens Col12a1(F) and Col14a1(G), and for fibril-forming collagens Col1a1(H), Col3a1(I) and Col5a1(J). Expression of Col11a1(K) is still greater for tendon proper-derived progenitors. (Biological replicates, n = 5 to 8; Mann–Whitney-Wilcoxon test – P <0.01, *; P <0.05, #; fold changes and statistical significance further reported in Table 3 and expanded results are reported in Additional file 1: Figure S1). GDF5, growth differentiation factor 5; PERI, peritenon-derived progenitors; SLRP, small leucine-rich proteoglycans; TP, tendon proper-derived progenitors.
Mentions: When the culture was supplemented with recombinant mouse GDF5, expression for matrix assembly genes in seeded peritenon progenitors increased to levels seen with the seeded tendon proper-derived progenitors (Table 3). However, after GDF5 supplementation, expression levels for the tendon markers Scx (Figure 5A) and Tnmd (Figure 5B) were unchanged when comparing GDF5-supplemented peritenon progenitor expression (PERI+) to that of tendon proper-derived progenitors not supplemented with GDF5 (TP). Expression of vascular marker Emcn (Figure 5C) also still differed between PERI + and TP progenitors. GDF5 supplementation did result in improved expression of matrix assembly genes for peritenon progenitors seeded in tendon constructs comparing PERI + progenitors and TP progenitors. Bolstered expression is noted for the small leucine-rich proteoglycans (SLRPs) Bgn early (Figure 5D) and Dcn (Figure 5E). Expression of FACIT collagens Col12a1 (Figure 5F) and Col14a1 (Figure 5G) as well as fibril-forming collagens Col1a1 (Figure 5H), Col3a1 (Figure 5I), and Col5a1 (Figure 5J) for GDF5-supplemented PERI + progenitors also became comparable to TP progenitors, yet expression of Col11a1 (Figure 5K) was still greater for TP progenitors. Addition of GDF5 did not lead to significant expression changes among TP progenitors for the tendon markers and matrix assembly genes analyzed, except for Bgn at 45 days; Additional file 1: Figure S1 exhibits expression results for constructs generated by cell type, GDF5 supplementation and culture time.

Bottom Line: Tendon construct ultrastructure was also compared after 45 days.It also was found that peritenon-derived progenitors secrete factor(s) stimulatory to tenocytes and tendon proper progenitors.Data demonstrate that, relative to peritenon-derived progenitors, tendon proper progenitors have greater potential for forming functional tendon-like tissue.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: Multipotent progenitor populations exist within the tendon proper and peritenon of the Achilles tendon. Progenitor populations derived from the tendon proper and peritenon are enriched with distinct cell types that are distinguished by expression of markers of tendon and vascular or pericyte origins, respectively. The objective of this study was to discern the unique tenogenic properties of tendon proper- and peritenon-derived progenitors within an in vitro model. We hypothesized that progenitors from each region contribute differently to tendon formation; thus, when incorporated into a regenerative model, progenitors from each region will respond uniquely. Moreover, we hypothesized that cell populations like progenitors were capable of stimulating tenogenic differentiation, so we generated conditioned media from these cell types to analyze their stimulatory potentials.

Methods: Isolated progenitors were seeded within fibrinogen/thrombin gel-based constructs with or without supplementation with recombinant growth/differentiation factor-5 (GDF5). Early and late in culture, gene expression of differentiation markers and matrix assembly genes was analyzed. Tendon construct ultrastructure was also compared after 45 days. Moreover, conditioned media from tendon proper-derived progenitors, peritenon-derived progenitors, or tenocytes was applied to each of the three cell types to determine paracrine stimulatory effects of the factors secreted from each of the respective cell types.

Results: The cell orientation, extracellular domain and fibril organization of constructs were comparable to embryonic tendon. The tendon proper-derived progenitors produced a more tendon-like construct than the peritenon-derived progenitors. Seeded tendon proper-derived progenitors expressed greater levels of tenogenic markers and matrix assembly genes, relative to peritenon-derived progenitors. However, GDF5 supplementation improved expression of matrix assembly genes in peritenon progenitors and structurally led to increased mean fibril diameters. It also was found that peritenon-derived progenitors secrete factor(s) stimulatory to tenocytes and tendon proper progenitors.

Conclusions: Data demonstrate that, relative to peritenon-derived progenitors, tendon proper progenitors have greater potential for forming functional tendon-like tissue. Furthermore, factors secreted by peritenon-derived progenitors suggest a trophic role for this cell type as well. Thus, these findings highlight the synergistic potential of including these progenitor populations in restorative tendon engineering strategies.

No MeSH data available.