Synthesis and Physiological Remodeling of CD34 Cells in the Skin following the Reversal of Fibrosis through Intensive Treatment for Lower Limb Lymphedema: A Case Report

Round 1

Reviewer 1 Report

The reviewer wishes to thank the editor and authors for the opportunity to review this manuscript. The article seeks to describe the redistribution of dermal CD34 (+) cells, presumed telocytes, in one patient with primary lymphedema both prior to and after treatment (with an end goal of clinical reversal of fibrosis) via the Godoy Method. In this single patient clinical trial, prior to treatment CD34 (+) cells were relatively restricted to the deeper dermis and after treatment with clinical reversal of fibrosis CD34 (+) cells were present both superficially and deep.  The authors indicate given these changes were induced by lymphatic stimulation, that possibly other fibrotic and inflammatory processes may benefit from similar treatment.

Over all, while the article is well written, it may perhaps be better received as a case report.  In terms of an official study the sample size (n=1) is too low to provide sufficient power to fully draw any definitive conclusions.

That being said, there is a significant difference (236% increase) in numbers of CD34 (+) cells and the patient did clinically improve.  As a biopsy of an unaffected area was not done, the manuscript may be more attractive if the normal/expected distribution and density of CD34 (+) cells in the dermis (both superficial and deep) is discussed.  Specifically if there are any changes known to occur in the lower extremities.  Otherwise, the implication that (line159) the number of TCs has “normalized” cannot be substantiated.

Additionally, given lines 153 to 155, were ultrastructural studies (i.e. electron microscopy) performed to identify 1) presence of ‘telocytes’ and 2) a specific pattern of damage to said ‘telocytes’?  If so the manuscript would be more attractive with some electron microscopy images. 

While for an original article I would want more patients involved with biopsies of uninvolved skin (possibly from the contralateral leg), perhaps for a case report this makes for a nice preliminary finding for additional larger studies.

Author Response

The reviewer wishes to thank the editor and authors for the opportunity to review this manuscript. The article seeks to describe the redistribution of dermal CD34 (+) cells, presumed telocytes, in one patient with primary lymphedema both prior to and after treatment (with an end goal of clinical reversal of fibrosis) via the Godoy Method. In this single patient clinical trial, prior to treatment CD34 (+) cells were relatively restricted to the deeper dermis and after treatment with clinical reversal of fibrosis CD34 (+) cells were present both superficially and deep.  The authors indicate given these changes were induced by lymphatic stimulation, that possibly other fibrotic and inflammatory processes may benefit from similar treatment.

Over all, while the article is well written, it may perhaps be better received as a case report.  In terms of an official study the sample size (n=1) is too low to provide sufficient power to fully draw any definitive conclusions.

Reply: ok;

That being said, there is a significant difference (236% increase) in numbers of CD34 (+) cells and the patient did clinically improve.  As a biopsy of an unaffected area was not done, the manuscript may be more attractive if the normal/expected distribution and density of CD34 (+) cells in the dermis (both superficial and deep) is discussed.  Specifically if there are any changes known to occur in the lower extremities.  Otherwise, the implication that (line159) the number of TCs has “normalized” cannot be substantiated.

Reply: Ok, change in the text.

Additionally, given lines 153 to 155, were ultrastructural studies (i.e. electron microscopy) performed to identify 1) presence of ‘telocytes’ and 2) a specific pattern of damage to said ‘telocytes’?  If so the manuscript would be more attractive with some electron microscopy images. 

Reply: We agree, however, in this study, electron microscopy was not performed, a great suggestion for our future studies, so agree that it is a case report.

While for an original article I would want more patients involved with biopsies of uninvolved skin (possibly from the contralateral leg), perhaps for a case report this makes for a nice preliminary finding for additional larger studies.

Reply: Agree case report because  is a preliminary, other studies  with a larger number of  participants are under development.

Author Response File: Author Response.docx

Reviewer 2 Report

This is an interesting work that shows reversal of distribution and number of CD34 positive cells in the dermis after the treatment of lymphedema. However, I have some major concerns.

Authors’ conclusions are based on only 1 patient/skin sample. Adding more samples would improve the quality of the paper. Their result is weak and tone should be lowered especially in discussion section.

Authors define telocytes in the entire introduction section and then call the cell type in their sample as CD34 positive cells. What do this CD34 positive cells include? Do they mean telocytes? If they mean telocytes only, additional stain to describe telocytes such as PDGFR-a may be needed. As CD34 is not specific for telocytes and other interstitial cells such as dermal dendritic cells express CD34 as well. Currently, CD34/PDGRF-a combination is considered to be the best to highlight the presence of telocytes in the skin (doi: 10.3390/cells11233903). If this CD34 positivity represents a cell group, authors can speculate on other cells that may play role in the process of reversal of fibrosis.

In discussion section (150-157), authors mention ultrastructural abnormalities in telocytes in systemic sclerosis and psoriasis. Without using electron microscopy, they claim that these ultrastructural abnormalities are also seen in lymphedema. They should either confirm it with electron microscopy or correct this statement.

Author Response

Open Review II

Comments and Suggestions for Authors

This is an interesting work that shows reversal of distribution and number of CD34 positive cells in the dermis after the treatment of lymphedema. However, I have some major concerns.

Authors’ conclusions are based on only 1 patient/skin sample. Adding more samples would improve the quality of the paper. Their result is weak and tone should be lowered especially in discussion section.

Reply: This study is the initial report of a histopathological finding and a new study is being carried out with a larger number of patients. But agree case report, change in text and ttle.

Authors define telocytes in the entire introduction section and then call the cell type in their sample as CD34 positive cells. What do this CD34 positive cells include? Do they mean telocytes? If they mean telocytes only, additional stain to describe telocytes such as PDGFR-a may be needed. As CD34 is not specific for telocytes and other interstitial cells such as dermal dendritic cells express CD34 as well. Currently, CD34/PDGRF-a combination is considered to be the best to highlight the presence of telocytes in the skin (doi: 10.3390/cells11233903). If this CD34 positivity represents a cell group, authors can speculate on other cells that may play role in the process of reversal of fibrosis.

Reply:This other evaluation has been identified, but several articles report telocytes only using CD34.

In discussion section (150-157), authors mention ultrastructural abnormalities in telocytes in systemic sclerosis and psoriasis. Without using electron microscopy, they claim that these ultrastructural abnormalities are also seen in lymphedema. They should either confirm it with electron microscopy or correct this statement.

Reply: We agree, however, in this study, electron microscopy was not performed, a great suggestion for our future studies,

Author Response File: Author Response.doc

Round 2

Reviewer 2 Report

Thank you for turning this paper into a case report, as it was a scientifically accurate change. However due to major grammatical and linguistic errors, that are mostly present in the newly written paragraphs, it is very hard to understand the paper.

Especially the discussion section as well as the introduction section repeatedly sway away from the main topic and cause distraction.

Once again, this case report simply shows increase in number of CD34 positive cells (possibly telocytes) especially around the vessels after the treatment. Some conclusions in the discussion such as “The relationship between telocytes and blood vessels and macrophages in the control of cell degradation products is observed in this study” are beyond its power of results.

Author Response

please see attachment

Author Response File: Author Response.pdf