Disordered metabolism in mice lacking irisin
Authors: Charles C Xu
Hongxia Deng
Xiaoyong Qiao
Liangzhi Xu
Yunyao Luo
Yaxian Ma
Journal: Scientific Reports (2020)
Discipline: Multidisciplinary
Keywords: Irisin; Fndc5; adipokine secretion; osteoblasts; osteoclasts
Affiliations: Reproductive Endocrinology and Regulation Laboratory West China Second University Hospital, Sichuan University, #20 Section 3, Ren Min Nan Road, Chengdu, 610041, Sichuan, People's Republic of China., The Joint Laboratory for Reproductive Medicine of Sichuan University, The Chinese University of Hong Kong, Hong Kong, People's Republic of China., Key Laboratory of Birth Defects and Related Diseases of Women and Children (Sichuan University), Ministry of Education, Chengdu, People's Republic of China., Department of Obstetrics and Gynecology, West China Second University Hospital, Sichuan University, Chengdu, People's Republic of China., College of Engineering, The Ohio State University, Columbus, OH, USA., Reproductive Endocrinology and Regulation Laboratory West China Second University Hospital, Sichuan University, #20 Section 3, Ren Min Nan Road, Chengdu, 610041, Sichuan, People's Republic of China. ., The Joint Laboratory for Reproductive Medicine of Sichuan University, The Chinese University of Hong Kong, Hong Kong, People's Republic of China. ., Key Laboratory of Birth Defects and Related Diseases of Women and Children (Sichuan University), Ministry of Education, Chengdu, People's Republic of China. ., Department of Obstetrics and Gynecology, West China Second University Hospital, Sichuan University, Chengdu, People's Republic of China. .

Abstract:  Irisin is a product of fibronectin type III domain-containing protein (Fndc5) and is involved in the regulation of adipokine secretion and the differentiation of osteoblasts and osteoclasts. In this study, we aimed to determine whether irisin lacking affects glucose/lipid and bone metabolism. We knocked out the Fndc5 gene to generate irisin-lacking mice. Remarkable, irisin lacking was related to poor 'browning response', with a bigger size of the intraperitoneal white adipose cell and decreased a number of brown adipose cells in brown adipose of interscapular tissue. The irisin lacking mice had hyperlipidemia and insulin resistance, reduced HDL-cholesterol level, increased LDL-cholesterol level, and decreased insulin sensitivity. The lacking of irisin was associated with reduced bone strength and bone mass in mice. The increased number of osteoclasts and higher expression of RANKL indicated increased bone resorption in irisin lacking mice. The level of IL-6 and TNF-Œ± also increased in irisin lacking mice. The results showed that irisin lacking was related to decreased 'browning response', glucose/lipid metabolic derangement, and reduced bone mass with increased bone resorption. Further studies are needed to confirm these initial observations and explore the mechanisms underlying the effects of irisin on glucose/lipid and bone metabolism. 

Retracted: No

PubPeer comments:
 We have discussed the article in our group and identified several points needing clarification: 1.tThe (female) Fndc5-KO mice are significantly heavier than the WT mice. This is in contrast to all other reports using Fndc5-KO mice (Lui et al. 2016 u2013 males; Kim et al. 2018 u2013 males and females; Xiong et al. 2019 u2013 males and females). Is there any explanation? 2.tIt is stated: u201eThe liver, u2026 weights were comparable between the two groups (Table 1).u201c In Table 1 the liver ratio (%) is 5-fold higher in KO compared to WT: 0.058 u00b1 0.01 vs. 0.011 u00b1 0.0002. Is this difference not significant? 3.tThe first heading reads: u201eReduced browning response and hyperlipidemia in irisin lac[k]ing miceu201c. The lipid data (Figure 3) show no differences in total serum cholesterol and triglycerides, just increased LDL and decreased HDL cholesterol. Is it correct to consider this as hyperlipidemia? 4.tTibia strength was characterized by a maximum bending load (N) of 25.71 u00b1 18.78 in WT vs. 15.60 u00b1 6.31** in KO. How can this difference be significant given the huge SEM given in Table 2? 5.tPro-inflammatory cytokine concentrations are given in Figure 7. There is a significant increase in IL-6 and a significant decrease in TNF-alpha in KO-mice. Eight of the cytokines did not differ. Why do you discuss u201cincreased inflammatory factors in irisin lacking miceu201d as a major outcome of this study. 6.tYour results on Fndc5-KO are opposite to those of Kim et al. (2018). You speculate that this discrepancy comes from different animal models. Your statement: u201cThey performed germline deletion of Fndc5(irisin residues 30u201369 deletion, 70u2013112 remaining). In this study, we clipped the 18th, and 19th nucleotides in exon three led to a frameshift mutation, resulting in irisin residues 43u2013112 deletion.u201d This is not true because Kim et al. (2018) created u201cu2026 germline deletion of Fndc5 (Exon 2 and 3).u201d thus targeting the same domain like you did. , 
 It appears that images in Figure 6 overlap, yet are described differently. ***   , 
 Images of mice seem to have been reused between two papers. https: pubpeer.com publications 271CEA66F9669CD6B6705D23B64023 https: pubpeer.com publications BCCD2BBBBE08E529E83A8DCD1F7C8E#2  ***  , 
 Dear Professor: Thank you for your attention and for the valuable comments concerning our manuscript entitled u201cDisordered metabolism in mice lacking irisinu201d. We have studied comments and analysed the original data carefully. The responds to some important comments are as follows: #1 1.Our results suggested that KO mice had lower weight when compared to WT mice (21.11u00b1 3.37 g VS 28.78 u00b14.31 g). The result was consistent with Xiong et al.2019, which similarly reported a decreased trend in weight of KO mice. In Kim et al.2018, they performed another method to generate KO mice and the weight of mice was not mentioned in their study. 2.Here, the original data showed significantly differences of liver,u2026 weight between KO and WT groups. We will add * to mark the significant differences in the Corrigendum letter. 3.To date, there is no standard for the definition of hyperlipidemia in animal models. Thus, our study referred to the diagnostic criteria of human hyperlipidemia, with the association of at least 1 of the 4 by following criteria: TCu22656.2mmol L; LDLu22654.1mmol Luff1bTGu22652.3mmol Luff1bHDL 1.0mmol L. Our results indicated a hyperlipidemia in KO mice, which had increased LDL level and decreased HDL level in serum. 4.We performed independent-samples T test to analyzed the data, and the results indeed showed a statistical difference between the two groups. 5.Our results showed KO mice had elevated TNF-u03b1 level than WT miceuff081.83 u00b1 0.04 pg ml VS 1.18 u00b1 0.03 pg mluff09. However, the data of KO and WT were inverted when drawing with GraphPad Prism software. We are sorry about that and we will make corrections in the corrigendum. 6. Kim et al.(2018) deleted second and third exons of Fndc5 in germline cell via Cre-loxp method, while we used TALEN technology in fertilized egg, which targeted the third exon contained a 2-nt deletion in the exon3, and this frameshift mutation generated 41 mutated amino acids ( 73MLRFIQEVNTTTRSCALWDLEEDTE YIVHVQAIS I Q G Q SPA113 to73IPVHSG GEHHHPVLRSLG PGGG HRIYR PCAGHLHPGTEPSQ 113) and a stop codon (TGA),leading to a truncated Fndc5 protein with only 113 aa, which completely destroyed the functions of Fndc5. In addition, our results suggested positive effects of irisin on bone metabolism, which is consistent with most other studies. In a recent study, Zhu et al. performed conditional Fndc5 knockout in osteocyte. Their results consistently showed the promoting effects of irisin on bone metabolism (DOI: 10.1002 jcp.29894). #2 1.In image acquisition, we usually capture several visual fields on one slice. We were likely to make a mistake in the process of image upload which led to the overlap of Figure 6. We had sufficient original pictures and we will make the corrections in the Corrigendum letter.    2.The generation of Fndc5 KO mice was the critical method in our study. Based on this, we observed the effects of irisin deficiency on metabolism and reproduction of mice. The timing of the submissions of the two papers were close; thus, it was necessary to describe the appearance of the mice in both papers. We would like to put the generation method and appearance of KO mice into the supplementary material in the second paper. Tips: You can download parts of the original data from Baidu Cloud . Besides, we have contacted the editor to modified the corrections. Thank you for your comments. , 
 #1 Dear professor: We have submitted the Correction letter to the Editorial on 5 June. Besides, we want to know if the responses answers your questions and if you have any other questions. , 
 #2 Dear professor: We have submitted the Correction letter to the Editorial on 5 June. Besides, we want to know if the responses answers your questions and if you have any other questions. , 
 Published: 04 November 2021 The original version of this Article contained errors in Figure 6 where the arrows were incorrectly placed, and in Figure 7 where the data of TNF-u03b1 in WT and KO mice was incorrect. The original Figures 6 and 7 and accompanying legends appear below. The original Article has been corrected.   ,