Letter to the editor concerning the article “The role of exercise selection in regional Muscle Hypertrophy: A randomized controlled trial” by Zabaleta-Korta et al. (2021)

We read with great interest the article by Zabaleta-Korta et al. (2021) titled “The role of exercise selection in regional Muscle Hypertrophy: A randomized controlled trial” due to its coherent proposal that aimed to verify, with a randomized controlled trial, whether muscle regional hypertrophy can be influenced by the selection of exercise modality. We praise the authors, as understanding the mechanisms regulating the heterogeneity of muscle hypertrophy is needed in training science literature. It follows that the methods used to quantify muscle hypertrophy become crucial in such investigations. That said, we believe that the morphological measures presented in Zabaleta-Korta et al. study, claimed as muscle cross-section area (CSA), are not representative of true muscle CSA and this may lead to erroneous interpretation and potentially erroneous replication studies. The intent of our letter is not to discredit the authors for the study design, but solely to draw attention to the methodology employed, which we believe has led to inconsistent results. Our point of view is explained below. First, we would like to address the methods used by the authors to quantify the rectus femoris (RF), vastus lateralis (VL) and vastus medialis CSA. Canonically, muscle CSA is defined as the area of a muscle delimited by the surrounding of its aponeurosis, which can be obtained via magnetic resonance imaging (MRI) or computed tomography (CT) scans, and even by ultrasound (US) (Noorkoiv et al., 2010). Therefore, to measure CSA from large muscles (such as the VL) using B-mode US, it is expected to either use the extended-field-of-view (EFOV) technique for panoramic image acquisition (Franchi et al., 2020a, 2018a; Ritsche et al., 2021) or to collect several images and make a collage using contour matching, the socalled “stitching images technique” (Reeves et al., 2004; Stokes et al., 2021), so the aponeurosis surrounding the whole muscle can be imaged and thus CSA assessed in its entirety. The use of either EFOV or stitching techniques is warranted when using US as one of the main limitations of US imaging is represented by the small field of view of the transducers, commonly ranging from 40 to 60 mm (Franchi et al., 2020b, 2018a; Sarto et al., 2021a, 2021b). In this regard, although presenting various methodological challenges, we believe that panoramic image acquisitions are more practical than collecting and overlapping single scans manually. In fact, EFOV technique employs a similar fitting method but with a larger number of images taken automatically as the transducer is moved along the skin (e Franchi et al., 2018a; Lima et al., 2012), being also a reliable alternative to MRI and CT – gold standard methods (Ahtiainen et al., 2010; Franchi et al., 2020a; Kositsky et al., 2020; Noorkoiv et al., 2010). However, in the study of Zabaleta-Korta et al., the US images were acquired at a single muscle site using a transducer with a very narrow field of view (not reported), as the authors were unable to even see the surrounding fascia of the RF, the smallest head of the quadriceps with regards to peak CSA (Figure 1 of Zabaleta-Korta et al., 2021). Thereby the authors only quantified the muscle area visible below the transducer (see Figure 1 a and b), claiming that to be the respective muscles CSA. This is, in our opinion, not correct, since that does not represent the true CSA of a muscle, with the scanned region being only a small part of it. In this regard, as the direction of hypertrophy is still unknown (i.e., where new muscle mass is added) and the method employed does not quantify muscle size changes at its lateral and medial extremities, the results shown cannot hold truth to the actual true CSA variations. In order to support the protocol employed, the authors cited Hacker et al. (2016): in the latter study, the RF CSA was measured, but the chosen muscle site was the most proximal thigh position so that the entire RF could be visualized in all participants even with the linear transducer used. Further, ZabaletaKorta et al. referenced Ruas et al. (2017), as very similar CSA assessment was performed in that investigation. Ruas et al. (2017) aimed to test the reliability of measuring the muscle visible area inside the transducer field of view collected on a specific muscle site, claiming that the EFOV US imaging could be more complicated, less time efficient and requires specific technique. Although we could agree in principle that EFOV US is not simple, the area measured by Ruas et al. was still not representative of the true muscle CSA, thus the two techniques cannot be deemed as interchangeable. It would be necessary to carry out a validation study of such proposed measurements in order to verify if this method is suitable to quantify muscle size changes, such as the ones carried out for the EFOV technique (Ahtiainen et al., 2010; Franchi et al., 2020a; Monti et al., 2020).