Effects of Coenzyme Q10 Supplementation in Women with Metabolic Syndrome and Non-Alcoholic Fatty Liver Disease Evaluated by Magnetic Resonance Imaging—Coenzyme Q10 in Metabolic Syndrome and NAFLD

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

CoQ10 is decreased in metabolically unhealthy humans, and treatment by CoQ10 was previously shown to improve metabolic parameters.  This study investigates impact of oral CoQ10 delivery on key metabolic parameters after 12 weeks of treatment in a very small cohort of obese individuals. Novelty is lacking, but the findings add to the additional premise to the prior knowledge base on CoQ10 treatment in humans. Nonetheless, there are a number of major concerns about this article:

 

Selection criteria:

1) The selection criteria excludes a group of humans that have taken a list of treatments in the past.  However, it doesn’t explain the medication that was used by those who were included within the study.  For instance, if the placebo group include higher number of people who have undergone diabetes or hyperlipidemia treatment, their responsiveness might be blunted for additional treatment attempts.

 

2) BMI of placebo vs treatment groups are 4 points apart.  This is a large difference at the start point. The type of diet for each group should be specified (it is said that they continued as before).  If placebo group ate less healthy meals (their BMI is 4 point higher in average), did they continue to eat unhealthy meals for the next 12 weeks (and vice versa for the CoQ10 treated cohort)?

 

3) Table 1 values are repeated within Table 2, so Table 1 is redundant. There should be only one table after combining the two (after adding P values from Table 1 too).

 

4) Small cohort size: Although insignificant, there is a good trend toward increased TG in both groups. Also, there is a good trend in % liver fat fraction in treated group. Many of these observations could reach significance if the cohort numbers were larger.

 

Methods:

1) Oral bioavailability of CoQ10 should be discussed (which organ will it have its highest impact?). Since they used oral delivery method, do they expect the primary impact to be in the GI or is systemic exposure anticipated?  Please list prior pharmacokinetic reference if available.

 

2) The reliability of FG measurement is not certain. Authors should cite more references to establish the validity of this measurement and equation. Discussion refers to this as a pioneer in demonstration. If this method is being used for the first time, establishing such a new technique would require comparison to liver biopsy biochemistry. As is, it is not clear how accurate this measurement is.

 

3) HbA1c measurement (instead of glucose) would be more informative.

 

Minor:

1) Metabolic syndrome is a rarely used vague terminology, which combines other pathologies to come up with a non-descriptive nomenclature. The article would be more accurate by citing specific metabolic parameters that they intend to study and avoid the usage of vague terminology such as MetS.

 

2) A sentence elaborating on increased HDL (and tendency to increase TG) in the presence of decreased LDL and total cholesterol would be beneficial as HDL is predominantly made of cholesterol and LDL is predominantly made of TG. This is a conundrum otherwise.

Comments on the Quality of English Language

Attention to grammar, punctuation and overall sentence coherence is extremely weak. There are broken and incomplete sentences throughout the whole text. It is hard to read some of the incomprehensible sentences. Certain passages are out of context and do not make a point (i.e. Line 70 typo;

 

Lines 71, 132, 160, 166 and many more with no space between words; Lines 73 –74  mixed up sentences (copy pasted in wrong places); Line 131 after post intervention (after post?); Line 249 – remark about hydrogen peroxide is unclear.  Please explain (should patients be treated with hydrogen peroxide?); Line 252 – Please explain “markers that make up the lipid profile”). The language and the cohesiveness of the sentences should be very carefully edited.

 

Author Response

REVIEWER 1

 

Selection criteria:

 

Question 01 - The selection criteria excludes a group of humans that have taken a list of treatments in the past. However, it doesn't explain the medication that was used by those who were included within the study. For instance, if the placebo group includes higher number of people who have undergone diabetes or hyperlipidemia treatment, their responsiveness might be blunted for additional treatment attempts.

 

Reply. Thanks for the question. A questionnaire was previously applied when recruiting patients with medications that the literature describes as potential interferences in the absorption of CoQ10 (hypoglycemic drugs, hypocholesterolemic drugs, statins, etc.). The patients in the present study did not use any of the medications mentioned in the literature with the potential to alter CoQ10 levels.

 

Question 02 - BMI of placebo vs treatment groups are 4 points apart. This is a big difference at the start point. The type of diet for each group should be specified (it is said that they continued as before). If placebo group had less healthy meals (their BMI is 4 points higher on average), did they continue to eat unhealthy meals for the next 12 weeks (and vice versa for the CoQ10 treated cohort)?

 

Reply. Great question. Even with a difference of 4 points, it was not statistically significant when comparing the groups, considering similar samples. The same result was maintained after supplementation, showing no changes between and within groups. The recommendation given to all volunteers was to maintain the same diet and level of physical activity as usual.

 

Question 03 -Table 1 values ​​are repeated within Table 2, so Table 1 is redundant. There should be only one table after combining the two (after adding P values ​​from Table 1 too).

 

Reply. We thanked the suggestion, removed table 1 and described in the text that the groups were similar.

 

Question 04 -Small cohort size: Although insignificant, there is a good trend toward increased TG in both groups. Also, there is a good trend in % liver fat fraction in treated group. Many of these observations could reach significance if the cohort numbers were larger.

 

Reply. We agree with your statement. We work with a convenience sample. Prospective clinical trials that work with MRI exams in COQ10 supplementation usually work with small numbers of participants due to the complexity of the design and the high value involved in carrying out the exam. However, we are optimistic about continuing to investigate the matter.

 

Methods:

 

Question 01-Oral bioavailability of CoQ10 should be discussed (which organ will it have its highest impact?). Since they used oral delivery method, do they expect the primary impact to be in the GI or is systemic exposure anticipated? Please list prior pharmacokinetic reference if available.

 

Reply. CoQ10 is ingested in its oxidized form, and is subsequently transformed, at the level of erythrocytes, into its reduced form. Because it is hydrophobic and has a large molecular weight, the absorption of CoQ10 from the diet ends up being slow and limited, similar to lipids. After oral intake within 1-2 hours plasma CoQ10 levels begin to increase. Maximum concentration occurs in 6 to 8 hours and the half-life can reach 34 hours. Its absorption occurs mainly in the small intestine, and is then transported to the liver, forming the lipoprotein complex. To transport CoQ10, it is coupled to chylomicrons, captured by the liver and then incorporated into LDL, which transports 58% of CoQ10, 26% by HDL and 10% by VLDL. Subsequently, CoQ10 is distributed to all tissues, but in greater quantities in organs that demand high energy concentrations such as the brain, heart, kidneys and liver. The majority of CoQ10 supplemented orally is eliminated through feces and bile, and a small fraction of what is absorbed ends up being eliminated through urine.

 

MILES M. The uptake and distribution of Coenzyme Q10. Mithochondrion., v. 7, p. 72-77, 2007.

GREENBERG S.; FRISHMAN WH Co-enzyme Q10: a new drug for cardiovascular disease. J Clin Pharmacol., v. 30, p. 596-608, 1990.

 

BHAGAVAN H.; CHOPRA R. Plasma coenzyme Q10 response to oral ingestion of coenzyme Q10 formulations. Mitochondrion., v. 7, p. 78-88, 2007.

 

Question 02 -The reliability of FG measurement is not certain. Authors should cite more references to establish the validity of this measurement and equation. Discussion refers to this as a pioneer in demonstration. If this method is being used for the first time, establishing such a new technique would require comparison to liver biopsy biochemistry. As is, it is not clear how accurate this measurement is.

Reply. In the paper:

Fishbein M, Castro F, Cheruku S, Jain S, Webb B, Gleason T, Stevens WR. Hepatic MRI for fat quantitation: its relationship to fat morphology, diagnosis, and ultrasound. J Clin Gastroenterol. 2005 Aug;39(7):619-25. doi: 10.1097/00004836-200508000-00012. PMID: 16000931.

with autorship of one co-author of our paper, the conclusion was: Hepatic MRI and ultrasound are both useful in identifying heavy fat accumulation associated with nonalcoholic fatty liver disease. MRI is superior to ultrasound in detecting and quantifying minor degrees of fatty metamorphosis in the liver.

 

Question 03- HbA1c measurement (instead of glucose) would be more informative.

 

Reply. We appreciate the suggestion and in a future study it is a marker that we will measure.

 

Minor:

 

1 - Metabolic syndrome is a rarely used vague terminology, which combines other pathologies to come up with a non-descriptive nomenclature. The article would be more accurate by citing specific metabolic parameters that they intend to study and avoid the usage of vague terminology such as MetS.

 

Reply. We appreciate the suggestion. We have corrected the term SM for the variable presented throughout the text.

 

2 - A sentence elaborating on increased HDL (and tendency to increase TG) in the presence of decreased LDL and total cholesterol would be beneficial as HDL is predominantly made of cholesterol and LDL is predominantly made of TG. This is a conundrum otherwise.

 

Reply.The effect of triglyceride concentration on CoQ is somewhat surprising because VLDL contains only a minor portion of CoQ in plasma. The possibility that ubiquinol increases in triglyceride-rich lipoproteins, VLDL, with the presence of metabolic syndrome may be important given the evidence that small amounts of ubiquinol in VLDL provide highly efficient antioxidant protection. The relatively strong correlations between triglycerides and ubiquinol, ubiquinone, and total CoQ suggest that triglyceride content may be more important in CoQ transport than previously thought. And in relation to LDL, the antioxidant function of CoQ10 proved to be effective, as it was the first antioxidant to be depleted when these lipoproteins were exposed to oxidative stress (CHEN, 2019; STOCKER, 1991).CoQ10 also promotes fatty acid oxidation by activating AMPK and PPARα. As a result, this mechanism increases lipoprotein lipase and apolipoprotein AV (APO-AV) gene expression, potentially reducing TG and VLDL levels. PPARα inhibits the maturation of sterol regulatory element binding protein (SREBP), thereby reducing the production of fatty acids and triglycerides and increasing the size of LDL-C particles.

 

Attention to grammar, punctuation and overall sentence coherence is extremely weak. There are broken and incomplete sentences throughout the entire text. It is difficult to read some of the incomprehensible sentences. Certain passages are out of context and do not make a point (ie Line 70 typo;

 

Lines 71, 132, 160, 166 and many more with no space between words; Lines 73 –74 mixed up sentences (copy pasted in wrong places); Line 131 after post intervention (after post?); Line 249 – remark about hydrogen peroxide is unclear. Please explain (should patients be treated with hydrogen peroxide?); Line 252 – Please explain “markers that make up the lipid profile”). The language and the cohesiveness of the sentences should be very carefully edited.

 

Reply. Our sincere apologies for the translation. After suggestion, we re-edited the entire text.

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript entitled “Effects of coenzyme Q10 supplementation in patients with metabolic syndrome and Non-alcoholic fatty liver disease evaluated by magnetic resonance imaging: Coenzyme Q10 in metabolic syndrome and NAFLD” by Casagrande et al., tried to evaluate the effects of CoQ10 supplementation in 22 female patients with metabolic syndrome and NAFLD. In this study, NAFLD, abdominal and visceral fat volume were analyzed by Magnetic Resonance Imaging. In addition, carrying out and anthropometric analysis, blood, blood pressure and some markers of metabolic syndrome. The topic of the manuscript is quite interesting, but the following questions should be considered during review.

 

Line 73-74: The text has some errors in the writing, I suppose the correct thing is “in patients with hyperglycemia was its significant ability to reduce levels of plasma glucose”. [15] “Farsi et al. (2016), administered 100 mg/day of CoQ10 fasting for 12 weeks.”

 

Line 159: Please change “mm2” into mm².

 

Line 197: Delete the period between the word pressure and arterial.

 

Line 249: Please change “H2O2 into H₂O₂

 

In the Table 1. Add a title to table 1.

 

In the Table 2. Add to Table 2 the levels of LDL-cholesterol for each group before and after the experiment since in their results and discussion the mention this parameter with a significance decreases.

 

In their study they mention that for the diagnosis of NAFLD, this disease can be confirmed with the elevation of liver enzymes. I suggest putting the before and after of the liver enzymes on one the tables and talking about the results you observed.

Comments for author File: Comments.pdf

Author Response

Response to Reviewer´s Comments

Dear reviewer´s, the considerations improved this paper.

 

The manuscript entitled “Effects of coenzyme Q10 supplementation in patients with metabolic syndrome and Non-alcoholic fatty liver disease evaluated by magnetic resonance imaging: Coenzyme Q10 in metabolic syndrome and NAFLD” by Casagrande et al., tried to evaluate the effects of CoQ10 supplementation in 22 female patients with metabolic syndrome and NAFLD. In this study, NAFLD, abdominal and visceral fat volume were analyzed by Magnetic Resonance Imaging. In addition, carrying out and anthropometric analysis, blood, blood pressure and some markers of metabolic syndrome. The topic of the manuscript is quite interesting, but the following questions should be considered during review.

 

Line 73-74: The text has some errors in the writing, I suppose the correct thing is “in patients with hyperglycemia was its significant ability to reduce levels of plasma glucose”. [15] “Farsi et al. (2016), administered 100 mg/day of CoQ10 fasting for 12 weeks.”

Reply. Thanks for the comment, correction made.

 

In the Table 2. Add to Table 2 the levels of LDL-cholesterol for each group before and after the experiment since in their results and discussion the mention of this parameter with a significance decreases.

Reply. It was not added because we did not measure this analyte in serum, the value found was through thecalculation of LDL-cholesterol using the Friedewald formula (LDL= Total Cholesterol – HDL – Triglycerides/5).

In their study they mention that for the diagnosis of NAFLD, this disease can be confirmed with the elevation of liver enzymes. I suggest putting the before and after of the liver enzymes on one of the tables and talking about the results you observed.

Reply. We appreciate the suggestion, but in this study we did not perform pre and post measurement of liver enzymes. In the recruitment phase, patients brought recent liver enzyme tests (from the last three months) to confirm NAFLD and subsequent sample stratification.

Reviewer 3 Report

Comments and Suggestions for Authors

The article is devoted to describing the results of a study assessing the effect of taking coenzyme Q10 in women with metabolic syndrome and NAFLD.

My comments:

1. In the Title of the article, the word Patients should be replaced with Women.

2. You need to add analyzed literary sources for 2023 to the list of references.

3. The English language of the article must be checked throughout the entire text. There are many stylistic errors (lines 72-73, 79-83, and so on throughout the text).

4. Why does the metabolic syndrome criteria indicate that the HDL-C level is ≥ 50 mg/dL?

5. It is indicated that 30 people were included in the study (lines 104-105), why are there only 22 women in the results of the article?

6. The tables show the results as mean ± SD. Due to the small number of groups, the distribution of features is clearly not parametric. You need to present the data as a median (25%; 75%).

7. The Discussion of the results is very extensive, long, and difficult to understand. This section needs to be rewritten more clearly, more concisely and more specifically.

8. The last paragraph (lines 365-376) of the text from the Discussion section should be made a separate Conclusion section.

9. Why is Figure 1 in the Discussion section?

Overall, the article is not very strong, it needs to be significantly reworked.

Comments on the Quality of English Language

The English language of the article must be checked throughout the entire text. There are many stylistic errors (lines 72-73, 79-83, and so on throughout the text).

Author Response

Response to Reviewer´s Comments,

Dear reviewer´s, the considerations improved this paper.

 

1. In the Title of the article, the word Patients should be replaced with Women.

Reply. Thanks for the comment, correction made.

 

2. You need to add analyzed literary sources for 2023 to the list of references.

Reply.Thank you for your comment, we have added recent studies to the list of references.

 

3. The English language of the article must be checked throughout the entire text. There are many stylistic errors (lines 72-73, 79-83, and so on throughout the text).

Reply. Our sincere apologies for the translation. After suggestion, we re-edited the entire text.

 

4. Why does the metabolic syndrome criteria indicate that the HDL-C level is ≥ 50 mg/dL?

Reply.Thanks for the comment, correction made.

 

5. It is indicated that 30 people were included in the study (lines 104-105), why are there only 22 women in the results of the article?

Reply.The sample consisted of 30 women, who attended the first consultation and received study instructions. During the 12-week intervention period, some volunteers interrupted treatment and were excluded from the research. The reasons for exclusion were: withdrawal by patients during the protocol (4 volunteers from the CoQ10 group and 2 from the placebo group), forgetfulness (1 volunteer from the placebo group) and pregnancy (1 volunteer from the placebo group). 22 patients participated in the study. We have corrected the information in the text.

 

6. The tables show the results as mean ± SD. Due to the small number of groups, the distribution of features is clearly not parametric. You need to present the data as a median (25%; 75%).

Reply. We appreciate the comment.

 

7. The discussion of the results is very extensive, long, and difficult to understand. This section needs to be rewritten more clearly, more concisely and more specifically.

Reply. Our sincere apologies for the translation. After suggestions, we rewrote the text and submitted it to an English proofreader

 

8. The last paragraph (lines 365-376) of the text from the Discussion section should be made a separate Conclusion section.

Reply. We appreciate the suggestion and follow the recommendations.

 

9. Why is Figure 1 in the Discussion section?

Reply.We appreciate the suggestion and have removed table 1.

 

 

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

Thank you for reviewing and responding to my suggestions.

Reviewer 3 Report

Comments and Suggestions for Authors

Dear editor, The authors made major corrections to the article according to my comments. I have no further comments. The article has been improved and can be published.