Substance-P Inhibits Cardiac Microvascular Endothelial Dysfunction Caused by High Glucose-Induced Oxidative Stress

Round 1

Reviewer 1 Report

The paper is interesting, while several concerns existed to be reconsidered.

1. Were the words ‘high glucose (HG) and hyperglycemia’ differently used throughout the text?
2. Can the results obtained by using CMECs be applied to systemic diabetic vascular pathology? If possible, the authors discuss it.
3. Substance P inhibition may be caused by ACE inhibitor agents commonly used in diabetes and cardiac diseases. Can the use of these agents support the authors’ experiment data?
4. How large is the effect of substance P on vasculature compared to that of NO in theory?
5. In line 71-77 and line 83-85 of Introduction, the sentences should have some references.
6. The line 138 should have the period, after …Switzerland) at the end of the sentence.
7. In line 143-144, the space should be set between lines.
8. In the Cytokine assay (2.8) of Methods, the authors should describe all kinds of cytokines to be measured.
9. In line 356 of Discussion, the sentence should have some references.
10. In subheadings, large and small capitals were complexed, for instance as ‘Assay’ and ‘assay’. These should be uniformed.
11. References had doble labelled number (e.g., 1. 1…). These should be corrected.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Reviewer 2 Report

The authors demonstrated that high glucose (HG) treatment reduced cell viability of cardiac microendothelial cells (CMECs) and the Akt/glycogen synthase kinase 3 beta (Gsk 3β) signaling, along with increased ROS production. These effect were rescued by the co-treatment with Substance-p (SP). SP was also able to improve angiogenesis, NO production and cytokine profile in HG-treated CMECs.

The work is interesting and experiments are well performed. SP may be considered an important therapeutic agent to reduce cardiovascular abnormalities induced by diabetes.

I have only few comments:

1) In addition to NO production, phospho-eNOS should be evaluated in SP+HG treated cells

2) I encourage to assess apoptosis (TUNEL assay, cleaved caspase 3 levels).

3) Is SP expressed by endothelial cells?

4) Please specify at which passage the cells were used for the experiments

5) I suggest to add “ROS level” instead to “fluorescence intensity” in the legend of graphs in Figure 1E and 2C

6) Figure 4B, please add statistics.

Author Response

Response to reviewer's comment was attached as a separate file.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

The paper has been improved.

Reviewer 2 Report

The authors addressed all my comments. I have no further comments.