Plasma Levels of Matrix Metalloprotease MMP-9 and Tissue Inhibitor TIMP-1 in Caucasian Patients with Polypoidal Choroidal Vasculopathy

Round 1

Reviewer 1 Report

The authors describe important information on the extracellular matrix in Caucasian polypoidal choroidal vasculopathy (PCV) patients. To date, mainly studies in Asian patients with this disease have been published, which stresses the importance of studies like these, even when negative results are found.

 

 

General comments:

 

• In my opinion, it would be important also to mention – with appropriate references – that some authors describe PCV as ‘aneurysmal type 1 neovascularization’.
• The optimal treatment for PCV is currently unknown, despite the fact that the standard treatment is indeed photodynamic therapy plus intravitreal injections with anti-vascular endothelial growth factor receptor medication. Please provide the information that some patients do only need either photodynamic therapy (and with which settings) or intravitreal injections with anti-vascular endothelial growth factor receptor medication.
• Were optical coherence tomography scanning and fluorescein angiography in patients and optical coherence tomography scanning, fluorescein angiography and indocyanine green angiography in controls also performed? When this is the case: please also include this in the Methods section. When this is not the case: it is important to mention this in your Discussion.
• Were patients and controls also gender-matched, as differences in the levels of the tested enzymes between males and females could have been present? Please explain in the Discussion why it was chosen not to do this on beforehand, despite the fact that adjusting for female gender did not change the conclusion that MMP-9 did not differ between 118 patients with PCV and healthy controls.
• In the Discussion, it would be good to include some more information on the pathogenesis of PCV. In its current form, only the fact that no conclusions on the origin of PCV can be drawn – based on the outcome of this study. However, what are your thoughts on the pathogenesis of PCV?

Author Response

Reviewer comment #1:

The authors describe important information on the extracellular matrix in Caucasian polypoidal choroidal vasculopathy (PCV) patients. To date, mainly studies in Asian patients with this disease have been published, which stresses the importance of studies like these, even when negative results are found.

 

Authors’ response:

Thank you for your time and comments.

 

 

Reviewer comment #2:

 In my opinion, it would be important also to mention – with appropriate references – that some authors describe PCV as ‘aneurysmal type 1 neovascularization’.

 

Authors’ response:

Thank you for this suggestion, which is now added to the manuscript.

 

 

Reviewer comment #3:

The optimal treatment for PCV is currently unknown, despite the fact that the standard treatment is indeed photodynamic therapy plus intravitreal injections with anti-vascular endothelial growth factor receptor medication. Please provide the information that some patients do only need either photodynamic therapy (and with which settings) or intravitreal injections with anti-vascular endothelial growth factor receptor medication.

 

Authors’ response:

Thank you for this suggestion, which is now added to the manuscript.

 

 

Reviewer comment #4:

Were optical coherence tomography scanning and fluorescein angiography in patients and optical coherence tomography scanning, fluorescein angiography and indocyanine green angiography in controls also performed? When this is the case: please also include this in the Methods section. When this is not the case: it is important to mention this in your Discussion.

 

Authors’ response:

For diagnosis of exudative retinopathies, we used both OCT and retinal angiography using both fluorescein and indocyanine green. We used OCT for controls, but not retinal angiography; however, it was a requirement for controls to have no exudative changes and no retinopathy whatsoever. We have provided a comment on this in the discussion as suggested. 

 

 

Reviewer comment #5:

Were patients and controls also gender-matched, as differences in the levels of the tested enzymes between males and females could have been present? Please explain in the Discussion why it was chosen not to do this on beforehand, despite the fact that adjusting for female gender did not change the conclusion that MMP-9 did not differ between 118 patients with PCV and healthy controls.

 

Authors’ response:

We were not aware that gender could influence plasma MMP9 values beforehand, and therefore we did not design the study with a priori gender-matching. However, as the reviewer also comments, as gender seemed to influence MMP9 values, this was included in a multivariate analysis to evaluate its potential influence on the results. We have provided a comment on this in the discussion as suggested. 

 

 

Reviewer comment #6:

In the Discussion, it would be good to include some more information on the pathogenesis of PCV. In its current form, only the fact that no conclusions on the origin of PCV can be drawn – based on the outcome of this study. However, what are your thoughts on the pathogenesis of PCV?

 

Authors’ response:

Thank you for this comment. We have elaborated on the potential pathogenesis of PCV in the discussion as suggested.

Reviewer 2 Report

General comments

Sørensen and co-workers report on MMP-9 and TIMP-1 levels in a Caucasian population with polypoidal choroidal vasculopathy (PCV). Plasma levels of MMP-9 and TIMP-1 were quantified in PCV patients and age-matched controls. A sample size of n = 26 PCV patients is a decent sample size in a Caucasian population.

In a Chinese population serum MMP-9 has been reported to be twice as high in PCV compared to healthy age-matched controls. The authors tested the hypothesis that plasma MMP-9 and inhibitor TIMP-1 levels were higher in PCV in a Caucasian population compared to age-matched Caucasian controls. No difference in MMP-9 levels and TIMP-1 between the groups was identified indicating a different etiology in Caucasian PCV.

Overall, the study is well conducted and the authors provide a good explanation of the rationale behind the study. The study is a valuable contribution to the journal as it brings insights into differences in PCV etiology between Caucasian and Asian populations.

The study is well-suited for publication in Vision after minor corrections. English language must be fine tuned. Parts of the manuscript has rather informal English language that is best avoided in written English. In some sections of the manuscript the authors use general terms that should be replaced with more specific terms – please see specific comments. Also, the authors must revise their description of findings in PCV.

 

Specific comments

Abstract

Please revise the abstract. A better explanation of the background is needed. The reader should be able to understand why it is relevant to quantify MMP-9 and TIMP-1

Introduction

The term “inter-ethnical differences” is used several times in the manuscript. The authors may be more specific. For example, the authors may refer to differences in inter-ethnical etiology or pathogenesis.

Line 34: ”choroidal originated vessels” should be replaced with “vessels originating from the choroid”

Lines 34-37: Please state differences in treatment between neovascular AMD and PCV. It will be useful for the reader.

Lines 38-43: Introducing pulsation on ICGA as a principle finding in PCV may not be correct. Pulsation on ICGA may not be the first lesion to look for. More frequent findings on ICGA may include branching network of inner choroidal vessels, nodular polypoidal aneurysms, dilations at the edge of these abnormal vessel networks, which correspond to orange sub­retinal nodules.

Lines 60-63: Please rephrase. Instead of writing “we wondered” and finishing with a question mark, it would be more elegant to write “we hypothesized” or “we tested the hypothesis that…”.

Lines 60-62: The term ‘interracial differences’ is not sufficiently specific. Differences in etiology may be more correct.

Line 64: please consider replacing “investigate” with “quantifiy” because what is done in the study is do quantify the proteins. Investigate may refer to many ways of characterizing a protein including folding, posttranslational modifications and interactions.

Lines 64-66. This reviewer misses the rationale behind quantification of TIMP-1. Did the study by Zeng et al., which is referred to, also quantify TIMP-1? A couple of lines about how TIMP-1 inhibits MMP-9 would be useful for the reader. In the introduction please state that TIMP-1 is also known by other names. Within the field of proteomics, researchers seem to use the name metalloproteinase inhibitor 1. Please see the link below and click on “Names & Taxonomy” in the left side of the screen:

https://www.uniprot.org/uniprot/P01033#names_and_taxonomy

 

Methods

Please define how replicates were used. Were quantitative values averaged?

96-97: Which kind of kit was used? Was it an ELISA-based kit?

Line 98: “were ran” is not a correct verb form. Please replace with “was run”.

Lines 104-107: Please be more details. Did the authors use student’s t-test, Mann-Whitney U etc.?

 

Discussion

Line 146: Inter-ethnical differences is a broad term. Differences in etiology or pathogenesis may be better terms to use, but they refer to what the authors are studying.

Lines 153-157: The sentence is too long and seems incorrect.

The conclusion is excellent.

Author Response

Reviewer comment #1:

Sørensen and co-workers report on MMP-9 and TIMP-1 levels in a Caucasian population with polypoidal choroidal vasculopathy (PCV). Plasma levels of MMP-9 and TIMP-1 were quantified in PCV patients and age-matched controls. A sample size of n = 26 PCV patients is a decent sample size in a Caucasian population.

In a Chinese population serum MMP-9 has been reported to be twice as high in PCV compared to healthy age-matched controls. The authors tested the hypothesis that plasma MMP-9 and inhibitor TIMP-1 levels were higher in PCV in a Caucasian population compared to age-matched Caucasian controls. No difference in MMP-9 levels and TIMP-1 between the groups was identified indicating a different etiology in Caucasian PCV.

Overall, the study is well conducted and the authors provide a good explanation of the rationale behind the study. The study is a valuable contribution to the journal as it brings insights into differences in PCV etiology between Caucasian and Asian populations.

The study is well-suited for publication in Vision after minor corrections. English language must be fine tuned. Parts of the manuscript has rather informal English language that is best avoided in written English. In some sections of the manuscript the authors use general terms that should be replaced with more specific terms – please see specific comments. Also, the authors must revise their description of findings in PCV.

 

Authors’ comment:

Thank you for your time and comments.

 

 

Reviewer comment #2:

Abstract

Please revise the abstract. A better explanation of the background is needed. The reader should be able to understand why it is relevant to quantify MMP-9 and TIMP-1

 

Authors’ comment:

We have now revised the abstract as suggested.

 

 

Reviewer comment #3:

Introduction

The term “inter-ethnical differences” is used several times in the manuscript. The authors may be more specific. For example, the authors may refer to differences in inter-ethnical etiology or pathogenesis.

 

Authors’ comment:

We have now provided more specific phrasing as suggested.

 

 

Reviewer comment #4:

Line 34: ”choroidal originated vessels” should be replaced with “vessels originating from the choroid”

 

Authors’ comment:

Changed as suggested.

 

 

Reviewer comment #5:

Lines 34-37: Please state differences in treatment between neovascular AMD and PCV. It will be useful for the reader.

 

Authors’ comment:

We have now specified as suggested.

 

 

Reviewer comment #6:

Lines 38-43: Introducing pulsation on ICGA as a principle finding in PCV may not be correct. Pulsation on ICGA may not be the first lesion to look for. More frequent findings on ICGA may include branching network of inner choroidal vessels, nodular polypoidal aneurysms, dilations at the edge of these abnormal vessel networks, which correspond to orange sub­retinal nodules.

 

Authors’ comment:

We agree that the phrasing could be misinterpreted and have revised this accordingly.

 

 

Reviewer comment #7:

Lines 60-63: Please rephrase. Instead of writing “we wondered” and finishing with a question mark, it would be more elegant to write “we hypothesized” or “we tested the hypothesis that…”.

 

Authors’ comment:

Changed as suggested.

 

 

Reviewer comment #8:

Lines 60-62: The term ‘interracial differences’ is not sufficiently specific. Differences in etiology may be more correct.

 

Authors’ comment:

Changed as suggested.

 

 

Reviewer comment #9:

Line 64: please consider replacing “investigate” with “quantifiy” because what is done in the study is do quantify the proteins. Investigate may refer to many ways of characterizing a protein including folding, posttranslational modifications and interactions.

 

Authors’ comment:

Changed as suggested.

 

 

Reviewer comment #10:

Lines 64-66. This reviewer misses the rationale behind quantification of TIMP-1. Did the study by Zeng et al., which is referred to, also quantify TIMP-1? A couple of lines about how TIMP-1 inhibits MMP-9 would be useful for the reader. In the introduction please state that TIMP-1 is also known by other names. Within the field of proteomics, researchers seem to use the name metalloproteinase inhibitor 1. Please see the link below and click on “Names & Taxonomy” in the left side of the screen:

https://www.uniprot.org/uniprot/P01033#names_and_taxonomy

 

Authors’ comment:

Yes, Zeng et al. also quantified TIMP-1, which is an inhibitor of MMP-9. Measuring one without the other would give insufficient information on the topic. We have now elaborated on this as suggested.

 

 

Reviewer comment #11: 

Methods

Please define how replicates were used. Were quantitative values averaged?

 

Authors’ comment:

Yes, quantitative values were averaged. This is now specified in the manuscript.

 

 

Reviewer comment #12:

96-97: Which kind of kit was used? Was it an ELISA-based kit?

 

Authors’ comment:

U-PLEX is an electrochemiluminescent-based multiplex immunoassay. This is now specified in the manuscript.

 

 

Reviewer comment #13:

Line 98: “were ran” is not a correct verb form. Please replace with “was run”.

 

Authors’ comment:

Changed as suggested.

 

 

Reviewer comment #14:

Lines 104-107: Please be more details. Did the authors use student’s t-test, Mann-Whitney U etc.?

 

Authors’ comment:

We used the Mann-Whitney U test for comparisons. This is now specified in the manuscript.

 

 

Reviewer comment #15:

Discussion

Line 146: Inter-ethnical differences is a broad term. Differences in etiology or pathogenesis may be better terms to use, but they refer to what the authors are studying.

 

Authors’ comment:

This is now revised to more specific phrasing as suggested.

 

 

Reviewer comment #16:

Lines 153-157: The sentence is too long and seems incorrect.

 

Authors’ comment:

This sentence is now revised into shorter sentences as suggested.

 

 

Reviewer comment #17:

The conclusion is excellent.

 

Authors’ comment:

Thank you.

Reviewer 3 Report

This paper by Dr. Sorensen and colleagues investigates a possible association between polypoidal choroidal vasculopathy and the increase of plasma levels of MMP-9 and TIMP-1 in Caucasian patients, similar to an association of the same kind that has been reported in Asian patients. The results do not indicate differences between Caucasian PCV patients and controls, thus confirming the inter-ethnical differences characterizing this pathology.

The study is well-conducted and the conclusions are quite clear, therefore I do not have particular comments. A couple of suggestions:

- The first part of the introduction explains that there are some difficulties at the diagnostic level in differentiating PCV from neovascular AMD: do the Authors imply that a possible association between PVC and MMP-9 and TMP-1 plasma levels may help this differentiation? Otherwise, what is the logic of a comparison between PCV and neovascular AMD in the introduction?

- In the discussion, the Authors could briefly discuss how the same type of pathology can show this apparent lack of similarities in the factors associated to it (including genetic features, retinal location of the vascular lesions, and, as demonstrated here, plasmatic markers) in Asian and Caucasian patients.

Author Response

Review comment:

This paper by Dr. Sorensen and colleagues investigates a possible association between polypoidal choroidal vasculopathy and the increase of plasma levels of MMP-9 and TIMP-1 in Caucasian patients, similar to an association of the same kind that has been reported in Asian patients. The results do not indicate differences between Caucasian PCV patients and controls, thus confirming the inter-ethnical differences characterizing this pathology.

The study is well-conducted and the conclusions are quite clear, therefore I do not have particular comments. A couple of suggestions:

 

Authors’ comment:

Thank you for your time and comments.

 

 

Reviewer comment #1:

- The first part of the introduction explains that there are some difficulties at the diagnostic level in differentiating PCV from neovascular AMD: do the Authors imply that a possible association between PVC and MMP-9 and TMP-1 plasma levels may help this differentiation? Otherwise, what is the logic of a comparison between PCV and neovascular AMD in the introduction?

 

Authors’ comment:

As the reviewer highlights, our introduction comments on the differences between PCV and neovascular AMD. Classically, PCV is seen in patients suspected of exudative AMD and who undergo detailed retinal diagnosis. It is therefore seen as an important differential diagnosis of AMD. Focusing the introduction on PCV could be argued, as the reviewer comments. However, the other reviewer recommends more details into the differences in treatment between PCV and AMD. We propose to include these descriptions of differences between PCV and AMD.

 

 

Reviewer comment #2:

- In the discussion, the Authors could briefly discuss how the same type of pathology can show this apparent lack of similarities in the factors associated to it (including genetic features, retinal location of the vascular lesions, and, as demonstrated here, plasmatic markers) in Asian and Caucasian patients.

 

Authors’ comment:

Thank you for this comment. We have now elaborated on this aspect in the discussion.