PONE-D-19-24096

Shear stress-exposed pulmonary artery endothelial cells fail to upregulate HSP70 in chronic thromboembolic pulmonary hypertension

PLOS ONE

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

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The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Partly

Reviewer #2: Partly

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The authors showed a paradoxical reduction in the level of Hsp70 protein in shear-stress exposed pulmonary artery endothelial cells from chronic thromboembolic pulmonary hypertension. While their findings are interesting and intriguing, however, I have some concerns:

1) The authors claimed that the decreased hsp70 level was due to shear-stress induced proteostatic dysregulation. Though hsp70 is a major player in the protein quality control system, other proteins including CHIP, an E3 ubiquitin ligase play crucial roles. The authors cannot justify decreased hsp70 to increased protein degradation. If the authors want use the word proteostasis, they will have to back it up with data that decreased hsp70 expression is due to transcriptional or post transcriptional mechanism, both of which can be due to enhanced hsp70 degradation by ubiquitin-proteasomal signaling pathways.

2) The authors assessed the levels of Hsp70 in HPAECs and HPAEC from patients with chronic thromboembolic PH. However they failed to mention the specific isoform of hsp70. So far, there are about 12 -15 hsp70 isoforms, of which most of them are localized to the cytosol.

3) Importantly, what is the relevance of decreased hsp70 in the context of the pathogenesis of chronic thromboembolic PH?

4) The manuscript was written poorly with no flow to the story. It needs to be re-written and thoroughly edited for clarity.

Reviewer #2: this study from Saline-Filho et al evaluates the effects of shear stress on EC isolated from CTEPH patients compared to normal control PAEC. The role of proteostasis in pulmonary disease is in its infancy so studies which will increase our understanding of this are important for the field. However, this work has several major limitations that limit is potential impact on the field.

1. It is very descriptive and there is no mechanistic insight into why CTEPH PAEC cells down regulation molecular chaperones in response to flow. This needs to be addressed.

2. There are actually no studies presented that actually study proteostasis in the manuscript. It would seem that the authors need to evaluate at least a few targets of hsp70/hsp90 or other chaperones and see how these differ in EC isolated from CTEPH patients compared to normal control PAEC.

3. Can over or under expression of hsp70/hsp90 or other chaperones be used to restore proteostasis in EC from CTEPH patients or perturb it in normal control PAEC respectively? This is needed to demonstrate cause and effect relationships.

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Reviewer #1: No

Reviewer #2: No

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Shear stress-exposed pulmonary artery endothelial cells fail to upregulate HSP70 in chronic thromboembolic pulmonary hypertension

PONE-D-19-24096R1

Dear Dr. Salibe-Filho,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

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Michael Bader

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PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

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3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Nice work! The authors should endeavor to determine the underlying mechanisms leading to decreased Hsp70 and Hsp90 expression in chronic thromboembolic induced-PAH. Identifying these mechanisms can provide therapeutic strategies in the treatment of PAH.

Also, the manuscript needs more work. The authors need today particular attention to tenses

Reviewer #2: Thea authors ahem adequately addressed the concerns raised in the prior review. No remaining concerns need to be addressed.

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7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

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Reviewer #1: No

Reviewer #2: No