PONE-D-23-00204Non-invasive assessment of steatohepatitis indicates increased risk of coronary artery diseasePLOS ONE

Dear Dr. Beer,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

 Please submit your revised manuscript by Apr 20 2023 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

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We look forward to receiving your revised manuscript.

Kind regards,

Shunsuke Mori, MD, PhD

Academic Editor

PLOS ONE

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Additional Editor Comments:

Our reviewers found some merits in this article but made a number of comments and suggestions. I ask the authors to clarify/correct each of the points that the reviewers indicated below. Additionally, in the text and tables, the authors should report p values with up to 2 significant digits and maximum 3 decimals (p = 0.21, p = 0.007, p = 0.001, etc.).

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. 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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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

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3. 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: No

Reviewer #2: Yes

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4. 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

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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: Reviewer : The authors have prospectively evaluated non-invasive fibrosis tests (FIB4, NFS, FAST and Fibroscan) in 120 patients with coronary arterial disease. MAFLD and advanced liver fibrosis (LSM ≥8 kPa) were observed in 41% and 6% (5% in the abstract?), respectively. A significant high score was present in patients needing coronary intervention.

Paragraph “Introduction”

• Page 3, line 73-74: The authors could add the more recent reference by Toh MR, (Global epidemiology and genetics of hepatocellular carcinoma. Gastroenterology 2023 Feb 2:S0016-5085(23)00105-1.) Tho et al. indicated that “Non-cirrhotic HCC is more common in NASH than the other etiologies; 39% of NASH-associated HCCs develop in absence of cirrhosis, compared with 22%, 6%, 9% for HBV, HCV, ALD respectively.”

• Page 3, line 78: Please, indicate the precise prevalence with its CI : 44.6% (95% CI, 36.0%–53.6%). Interestingly, the prevalence of CAD was even more important when invasive coronary angiogram was used : 60.5% (95% CI, 43.8%–75.1%)

Paragraph “Materials and Methods”

• Page 4, line 128: The authors need to specify conditions of blood samples. Surely, in a fasting condition

• Page 5: The authors could add the reference by Castera L, et al (Gastroenterology 2019) for the threshold (8 kPa) used to define significant fibrosis

• Page 5, line 147 : “Patients with SLCD exceeding 25 mm were examined with the XL-probe”. This recommendation is similar to that also proposed by EASL using a BMI cut-off of 30 kg/m2 to select the M or XL probe (EASL-ALEH clinical practice guidelines. J Hepatol 2015;63:237–264). The authors could add this BMI cut-off for clinicians since BMI cut-off is more often used than SLCD.

• Page 5, line152: The authors used AAP value ≥302 dB/m to diagnose any grade of steatosis. However, different cut-off values have been proposed elsewhere and, for EASL, a CAP (> 275 dB/m) is currently suggested (J Hepatol. 2021; 75:659-689). This point could be discussed.

Paragraph “Results”

• In this section, baseline biological and demographic characteristics of the 120 patients should be clearly reported in a dedicated table. All the components of the MAFLD definition need to be reported (waist circumference, hs-CRP, etc.) together with liver enzymes, PT, albumin, platelets, etc. and % of patients with MAFLD.

• Instead of describing characteristics of patients with and without coronary intervention in the total cohort, the authors should complete this table 1 by describing the non-MAFLD population

• In this section, the authors could specify the indications of routine (n= …) or emergency angiography (n=…).

• Page 9 : the authors write “LSM was valid and reliable in 120 (94% of eligible) patients”. The 128 “eligible” patients could be shown in the flow chart (Figure 1) for a more comprehensive reading.

• Table 2 : the authors need to define the ULN of AST and ALT in their centre. All percentages have been rounded in this table. I suggest to indicate the right numbers (for instance 4/70 = 5.7% instead of 6%). The authors may round figures in the discussion

• Page 11, line 241 : the authors write “None of the high-risk FIB4 patients and only one of the high-risk NFS patients had increased LSM values”. I suggest to create a table in supplementary appendix to characterize the 113 patients with a low risk (< 8kPa) and the 7 patients with a high risk (≥ 8 kPa), even though the number of high-risk cases is low. We need to better characterize the small group of CAD patients with LSM ≥ 8 kPa

• Table 3 : the title of this table does not indicate clearly what the authors are really looking for. I suggest to writ “Variables associated with coronary intervention using multivariate analysis”.

• Page 11, line 249 : The authors write “The results are very similar in the MAFLD subgroup”. The authors provide in Suppl. Appendix regarding this multivariate analysis regarding the subgroup of 49 MAFLD patients. Since only 22 coronary interventions were performed in this subgroup, only two variables can be entered in the model.

Paragraph “Discussion”

• Page 12, line 273-74: It is surprising that metabolic factors are not associated with advanced liver fibrosis in this study. Indeed, liver fibrosis development in NAFLD patients may result from a long-term exposure to cardiometabolic risk factors such as diabetes, arterial hypertension, dyslipidaemia and overweight (Baratta F, et al. Gastroenterol Hepatol 2020;18:2324-31). Genetic factors may also impact the relationship between NAFLD and CAD (Miao Z, et al. HGG Adv 2021 Aug 24;3:100056) ; The pathophysiology of the relationship between NAFLD and CAD could be explained briefly with the main causal factors.

• Page 12, line 285: “As a secondary finding, we observed associations of higher liver stiffness within the limits of normal in patients with relevant CAD compared to those without need of revascularization therapy” : these data are not shown.

• Please, check the references 7, 8, 9, etc.

Reviewer #2: The study presents the results of an interesting and original research. Experiments, statistics, and other analyses are performed to a high technical standard and are described in a detailed manner.

Conclusions are presented in an appropriate fashion and are supported by the data, but the authors must describe if are any group of patients with coronary interventions that must do the non-invasive assessment of liver fibrosis, probably patients with metabolic syndrome or T2DM should be examined with transient elastography for liver fibrosis. Also, the authors must explain if are any correlations between LSM values and severity of CAD according to the grade of stenosis. Also if are any correlations between them in which patients are most commen met?

In addition, they must put a space before the references in thes text.

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​

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Reviewer #1: Yes: Thevenot

Reviewer #2: No

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PONE-D-23-00204R1Non-invasive assessment of steatohepatitis indicates increased risk of coronary artery diseasePLOS ONE

Dear Dr. Karlas,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

This second version of the manuscript has been revised well. Reviewer #1 requested the authors to clarify just a few points. Please reply to the reviewer’s comments and make appropriate revision if necessary.

Please submit your revised manuscript by Jun 18 2023 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

• A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Shunsuke Mori, MD, PhD

Academic Editor

PLOS ONE

Journal Requirements:

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

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

**********

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: Minor corrections to provide :

Page 4, line 115-117: the references should be put at the end of the sentence : “….severe congestive heart failure …………… and pulmonary hypertension (WHO III or IV) (28, 29).”.

Revised Table 1 : Quantitative data should be expressed in median with IQR and qualitative data in absolute numbers with %

For instance :

“Male gender n (%): 80 (67%)” and only one column is sufficient

The line “AST % of ULN” is write twice ?

AST should be expressed in µkat/L, like GGT and AP

Please correct “gGT” by “GGT”

Instead of “exp9/l”, write “109/L” or “mm-3”

Is is really useful to write “HbA1c NSGP/DCCT”? It should be simpler to write only “HbA1c”

Page 9, line 238: write “≥ S2”

Page 11 : In the following sentence, the authors could specify that “high-risk”patients were defined according to the specific cut-off of NFS : “Only 5 patients (10%) of the interventional group and 4 patients (6%) of the non- intervention group (p=0.36) were identified as high-risk”

Page 11, line 266 : it would be better to write in full “MI” for a better understanding of this sentence

Reviewer #2: The work of Beer et al. is an exceptional original paper, which includes new data regarding liver fibrosis and CAD. Also, this paper open a new path in the links between liver fibrosis and CAD.

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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.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: Yes: Thevenot

Reviewer #2: No

**********

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

Non-invasive assessment of steatohepatitis indicates increased risk of coronary artery disease

PONE-D-23-00204R2

Dear Dr. Karlas,

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.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Shunsuke Mori, MD, PhD

Academic Editor

PLOS ONE