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Mechanisms of lymphatic regeneration after tissue transfer.

Yan A, Avraham T, Zampell JC, Aschen SZ, Mehrara BJ - PLoS ONE (2011)

Bottom Line: Patterns of VEGF-C expression and macrophage infiltration were temporally and spatially associated with lymphatic regeneration.When compared to mice treated with excision only, there was a 4-fold decrease in tail volumes, 2.5-fold increase in lymphatic transport by lymphoscintigraphy, 40% decrease in dermal thickness, and 54% decrease in scar index in skin-grafted animals, indicating that tissue transfer could bypass damaged lymphatics and promote rapid lymphatic regeneration.This process is temporally and spatially associated with VEGF-C expression and macrophage infiltration.

View Article: PubMed Central - PubMed

Affiliation: The Division of Plastic and Reconstructive Surgery, Department of Surgery, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America.

ABSTRACT

Introduction: Lymphedema is the chronic swelling of an extremity that occurs commonly after lymph node resection for cancer treatment. Recent studies have demonstrated that transfer of healthy tissues can be used as a means of bypassing damaged lymphatics and ameliorating lymphedema. The purpose of these studies was to investigate the mechanisms that regulate lymphatic regeneration after tissue transfer.

Methods: Nude mice (recipients) underwent 2-mm tail skin excisions that were either left open or repaired with full-thickness skin grafts harvested from donor transgenic mice that expressed green fluorescent protein in all tissues or from LYVE-1 knockout mice. Lymphatic regeneration, expression of VEGF-C, macrophage infiltration, and potential for skin grafting to bypass damaged lymphatics were assessed.

Results: Skin grafts healed rapidly and restored lymphatic flow. Lymphatic regeneration occurred beginning at the peripheral edges of the graft, primarily from ingrowth of new lymphatic vessels originating from the recipient mouse. In addition, donor lymphatic vessels appeared to spontaneously re-anastomose with recipient vessels. Patterns of VEGF-C expression and macrophage infiltration were temporally and spatially associated with lymphatic regeneration. When compared to mice treated with excision only, there was a 4-fold decrease in tail volumes, 2.5-fold increase in lymphatic transport by lymphoscintigraphy, 40% decrease in dermal thickness, and 54% decrease in scar index in skin-grafted animals, indicating that tissue transfer could bypass damaged lymphatics and promote rapid lymphatic regeneration.

Conclusions: Our studies suggest that lymphatic regeneration after tissue transfer occurs by ingrowth of lymphatic vessels and spontaneous re-connection of existing lymphatics. This process is temporally and spatially associated with VEGF-C expression and macrophage infiltration. Finally, tissue transfer can be used to bypass damaged lymphatics and promote rapid lymphatic regeneration.

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Related in: MedlinePlus

Both reconnection and infiltration of LYVE-1+ lymphatic vessels contribute to lymphatic regeneration after tissue transfer.A. Bar graph depicting source of lymphatic vessels in various regions of the skin-grafted tissues 2 or 6 weeks after surgery. Origin of lymphatic vessels in various regions of the skin graft 2 or 6 weeks after surgery was determined using podoplanin/LYVE-1 co-localization in skin grafts harvested from LYVE-1 knockout mice and transferred to nude mice. Recipient lymphatic vessels were identified as podoplanin+/LYVE-1+ (white bars) while donor lymphatics were identified as podoplanin+/LYVE-1- (black bars). Recipient-derived lymphatic vessels were noted primarily in the distal and proximal portions of the graft at the 2-week time point and became more uniform in distribution by 6 weeks. Increased number of recipient vessels in the peripheral regions of the graft at the 6-week time point resulted in a relative decrease in the percentage of donor-derived lymphatics at this time. B, C. Representative images (10x) of LYVE-1 (red) and podoplanin (green) staining of the distal portion of the graft 2 (B) and 6 (C) weeks after surgery. Yellow line marks the junction of the skin graft (located to the right) and recipient (located to the left) tissues. D. Recipient lymphatic vessels (podoplanin+/LYVE-1+) in various regions of the skin graft 2 and 6 weeks after skin grafting. Note that the numbers of recipient vessels in the distal (D) and proximal (P) portions of the tail are significantly greater than the number of recipient lymphatics in the middle (M) portion of the graft at both the 2- and 6-week time points indicating ingrowth of vessels from the periphery (*p<0.05). In addition, the number of recipient lymphatics in various regions of the skin graft increased significantly when comparing 2- and 6-week time points indicating ingrowth of recipient vessels. E. Donor lymphatic vessels (podoplanin+/LYVE-1-) in various regions of the skin graft 2 and 6 weeks after skin grafting (*p<0.05). The number of donor vessels remained essentially unchanged at both time points indicating that donor lymphatics persist over time and do not proliferate after tissue transfer.
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pone-0017201-g003: Both reconnection and infiltration of LYVE-1+ lymphatic vessels contribute to lymphatic regeneration after tissue transfer.A. Bar graph depicting source of lymphatic vessels in various regions of the skin-grafted tissues 2 or 6 weeks after surgery. Origin of lymphatic vessels in various regions of the skin graft 2 or 6 weeks after surgery was determined using podoplanin/LYVE-1 co-localization in skin grafts harvested from LYVE-1 knockout mice and transferred to nude mice. Recipient lymphatic vessels were identified as podoplanin+/LYVE-1+ (white bars) while donor lymphatics were identified as podoplanin+/LYVE-1- (black bars). Recipient-derived lymphatic vessels were noted primarily in the distal and proximal portions of the graft at the 2-week time point and became more uniform in distribution by 6 weeks. Increased number of recipient vessels in the peripheral regions of the graft at the 6-week time point resulted in a relative decrease in the percentage of donor-derived lymphatics at this time. B, C. Representative images (10x) of LYVE-1 (red) and podoplanin (green) staining of the distal portion of the graft 2 (B) and 6 (C) weeks after surgery. Yellow line marks the junction of the skin graft (located to the right) and recipient (located to the left) tissues. D. Recipient lymphatic vessels (podoplanin+/LYVE-1+) in various regions of the skin graft 2 and 6 weeks after skin grafting. Note that the numbers of recipient vessels in the distal (D) and proximal (P) portions of the tail are significantly greater than the number of recipient lymphatics in the middle (M) portion of the graft at both the 2- and 6-week time points indicating ingrowth of vessels from the periphery (*p<0.05). In addition, the number of recipient lymphatics in various regions of the skin graft increased significantly when comparing 2- and 6-week time points indicating ingrowth of recipient vessels. E. Donor lymphatic vessels (podoplanin+/LYVE-1-) in various regions of the skin graft 2 and 6 weeks after skin grafting (*p<0.05). The number of donor vessels remained essentially unchanged at both time points indicating that donor lymphatics persist over time and do not proliferate after tissue transfer.

Mentions: Quantification of donor and recipient lymphatic vessels also suggested that both reconnection and infiltration of recipient-derived lymphatics contribute to lymphatic regeneration in the transplanted skin. Two weeks following tissue transfer, 52% of the vessels in the distal margin and 37% of those at the proximal margin were donor-derived (i.e. podoplanin+/LYVE-1-). In contrast, 92% of the lymphatic vessels in the middle portion were donor-derived. By 6 weeks, the vast majority (>90%) of lymphatic vessels present in the distal and proximal margins were of recipient origin. Similarly, the number of recipient vessels in the middle portion also increased such that the distribution of these vessels with donor-derived lymphatics was 1∶1. These differences in the distribution of the lymphatic vessels were primarily due to the ingrowth of new lymphatics since the number of recipient vessels in the later time points increased substantially while the number of donor vessels (similar to our qualitative observations in the GFP experiment) decreased slightly, though not statistically significant (Figures 3D–E). There were significantly more recipient-derived vessels at the distal and proximal margins as compared to the central aspect of the graft suggesting ingrowth of vessels from the periphery of the transferred tissues (8.2 and 6.0 vs. 3.1; p<0.05). Taken together these findings suggest that donor-derived lymphatic vessels persist over time and that transferred tissues become further infiltrated by lymphatic vessels originating from the recipient beginning at the distal and proximal edges and infiltrating towards the center of the graft.


Mechanisms of lymphatic regeneration after tissue transfer.

Yan A, Avraham T, Zampell JC, Aschen SZ, Mehrara BJ - PLoS ONE (2011)

Both reconnection and infiltration of LYVE-1+ lymphatic vessels contribute to lymphatic regeneration after tissue transfer.A. Bar graph depicting source of lymphatic vessels in various regions of the skin-grafted tissues 2 or 6 weeks after surgery. Origin of lymphatic vessels in various regions of the skin graft 2 or 6 weeks after surgery was determined using podoplanin/LYVE-1 co-localization in skin grafts harvested from LYVE-1 knockout mice and transferred to nude mice. Recipient lymphatic vessels were identified as podoplanin+/LYVE-1+ (white bars) while donor lymphatics were identified as podoplanin+/LYVE-1- (black bars). Recipient-derived lymphatic vessels were noted primarily in the distal and proximal portions of the graft at the 2-week time point and became more uniform in distribution by 6 weeks. Increased number of recipient vessels in the peripheral regions of the graft at the 6-week time point resulted in a relative decrease in the percentage of donor-derived lymphatics at this time. B, C. Representative images (10x) of LYVE-1 (red) and podoplanin (green) staining of the distal portion of the graft 2 (B) and 6 (C) weeks after surgery. Yellow line marks the junction of the skin graft (located to the right) and recipient (located to the left) tissues. D. Recipient lymphatic vessels (podoplanin+/LYVE-1+) in various regions of the skin graft 2 and 6 weeks after skin grafting. Note that the numbers of recipient vessels in the distal (D) and proximal (P) portions of the tail are significantly greater than the number of recipient lymphatics in the middle (M) portion of the graft at both the 2- and 6-week time points indicating ingrowth of vessels from the periphery (*p<0.05). In addition, the number of recipient lymphatics in various regions of the skin graft increased significantly when comparing 2- and 6-week time points indicating ingrowth of recipient vessels. E. Donor lymphatic vessels (podoplanin+/LYVE-1-) in various regions of the skin graft 2 and 6 weeks after skin grafting (*p<0.05). The number of donor vessels remained essentially unchanged at both time points indicating that donor lymphatics persist over time and do not proliferate after tissue transfer.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3040774&req=5

pone-0017201-g003: Both reconnection and infiltration of LYVE-1+ lymphatic vessels contribute to lymphatic regeneration after tissue transfer.A. Bar graph depicting source of lymphatic vessels in various regions of the skin-grafted tissues 2 or 6 weeks after surgery. Origin of lymphatic vessels in various regions of the skin graft 2 or 6 weeks after surgery was determined using podoplanin/LYVE-1 co-localization in skin grafts harvested from LYVE-1 knockout mice and transferred to nude mice. Recipient lymphatic vessels were identified as podoplanin+/LYVE-1+ (white bars) while donor lymphatics were identified as podoplanin+/LYVE-1- (black bars). Recipient-derived lymphatic vessels were noted primarily in the distal and proximal portions of the graft at the 2-week time point and became more uniform in distribution by 6 weeks. Increased number of recipient vessels in the peripheral regions of the graft at the 6-week time point resulted in a relative decrease in the percentage of donor-derived lymphatics at this time. B, C. Representative images (10x) of LYVE-1 (red) and podoplanin (green) staining of the distal portion of the graft 2 (B) and 6 (C) weeks after surgery. Yellow line marks the junction of the skin graft (located to the right) and recipient (located to the left) tissues. D. Recipient lymphatic vessels (podoplanin+/LYVE-1+) in various regions of the skin graft 2 and 6 weeks after skin grafting. Note that the numbers of recipient vessels in the distal (D) and proximal (P) portions of the tail are significantly greater than the number of recipient lymphatics in the middle (M) portion of the graft at both the 2- and 6-week time points indicating ingrowth of vessels from the periphery (*p<0.05). In addition, the number of recipient lymphatics in various regions of the skin graft increased significantly when comparing 2- and 6-week time points indicating ingrowth of recipient vessels. E. Donor lymphatic vessels (podoplanin+/LYVE-1-) in various regions of the skin graft 2 and 6 weeks after skin grafting (*p<0.05). The number of donor vessels remained essentially unchanged at both time points indicating that donor lymphatics persist over time and do not proliferate after tissue transfer.
Mentions: Quantification of donor and recipient lymphatic vessels also suggested that both reconnection and infiltration of recipient-derived lymphatics contribute to lymphatic regeneration in the transplanted skin. Two weeks following tissue transfer, 52% of the vessels in the distal margin and 37% of those at the proximal margin were donor-derived (i.e. podoplanin+/LYVE-1-). In contrast, 92% of the lymphatic vessels in the middle portion were donor-derived. By 6 weeks, the vast majority (>90%) of lymphatic vessels present in the distal and proximal margins were of recipient origin. Similarly, the number of recipient vessels in the middle portion also increased such that the distribution of these vessels with donor-derived lymphatics was 1∶1. These differences in the distribution of the lymphatic vessels were primarily due to the ingrowth of new lymphatics since the number of recipient vessels in the later time points increased substantially while the number of donor vessels (similar to our qualitative observations in the GFP experiment) decreased slightly, though not statistically significant (Figures 3D–E). There were significantly more recipient-derived vessels at the distal and proximal margins as compared to the central aspect of the graft suggesting ingrowth of vessels from the periphery of the transferred tissues (8.2 and 6.0 vs. 3.1; p<0.05). Taken together these findings suggest that donor-derived lymphatic vessels persist over time and that transferred tissues become further infiltrated by lymphatic vessels originating from the recipient beginning at the distal and proximal edges and infiltrating towards the center of the graft.

Bottom Line: Patterns of VEGF-C expression and macrophage infiltration were temporally and spatially associated with lymphatic regeneration.When compared to mice treated with excision only, there was a 4-fold decrease in tail volumes, 2.5-fold increase in lymphatic transport by lymphoscintigraphy, 40% decrease in dermal thickness, and 54% decrease in scar index in skin-grafted animals, indicating that tissue transfer could bypass damaged lymphatics and promote rapid lymphatic regeneration.This process is temporally and spatially associated with VEGF-C expression and macrophage infiltration.

View Article: PubMed Central - PubMed

Affiliation: The Division of Plastic and Reconstructive Surgery, Department of Surgery, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America.

ABSTRACT

Introduction: Lymphedema is the chronic swelling of an extremity that occurs commonly after lymph node resection for cancer treatment. Recent studies have demonstrated that transfer of healthy tissues can be used as a means of bypassing damaged lymphatics and ameliorating lymphedema. The purpose of these studies was to investigate the mechanisms that regulate lymphatic regeneration after tissue transfer.

Methods: Nude mice (recipients) underwent 2-mm tail skin excisions that were either left open or repaired with full-thickness skin grafts harvested from donor transgenic mice that expressed green fluorescent protein in all tissues or from LYVE-1 knockout mice. Lymphatic regeneration, expression of VEGF-C, macrophage infiltration, and potential for skin grafting to bypass damaged lymphatics were assessed.

Results: Skin grafts healed rapidly and restored lymphatic flow. Lymphatic regeneration occurred beginning at the peripheral edges of the graft, primarily from ingrowth of new lymphatic vessels originating from the recipient mouse. In addition, donor lymphatic vessels appeared to spontaneously re-anastomose with recipient vessels. Patterns of VEGF-C expression and macrophage infiltration were temporally and spatially associated with lymphatic regeneration. When compared to mice treated with excision only, there was a 4-fold decrease in tail volumes, 2.5-fold increase in lymphatic transport by lymphoscintigraphy, 40% decrease in dermal thickness, and 54% decrease in scar index in skin-grafted animals, indicating that tissue transfer could bypass damaged lymphatics and promote rapid lymphatic regeneration.

Conclusions: Our studies suggest that lymphatic regeneration after tissue transfer occurs by ingrowth of lymphatic vessels and spontaneous re-connection of existing lymphatics. This process is temporally and spatially associated with VEGF-C expression and macrophage infiltration. Finally, tissue transfer can be used to bypass damaged lymphatics and promote rapid lymphatic regeneration.

Show MeSH
Related in: MedlinePlus