INTRODUCTION: Ultrasonography is a non-invasive and safe method for assessing muscle morphology. Among its parameters, echo intensity (EI), derived from grayscale image analysis, has emerged as a promising indicator of muscle quality and intramuscular fat infiltration. This study aims to validate EI as a marker for evaluating muscle quality in a population of Czech children, through integration with gold-standard assessments of muscle strength and body composition. The primary aim of this study is to assess the reliability and construct validity of quadriceps muscle EI using ultrasound as a proxy measure of morphological muscle quality in children aged 10-14 years. METHODS AND ANALYSIS: Children aged 10-14 years will undergo ultrasound assessment of the quadriceps femoris (QF). EI will be derived from longitudinal scans of each QF head and the cross-sectional area (CSAQF) from panoramic mid-thigh images. Muscle function will be assessed as maximal voluntary contraction (MVC) of isometric knee extension with muscle quality expressed as MVC/CSAQF. A 30 s sit-to-stand test (30STS) will be used as an additional functional measure. EI reliability (intra-rater, inter-rater and test-retest) will be evaluated with intraclass correlation coefficients (ICC), Bland-Altman plots and complementary indices. Exploratory known-groups validity will be tested by comparing EI between weight-status groups. Control variables include dual-energy X-ray absorptiometry (DXA)-derived body composition, skeletal age (as determined by DXA hand scans) and physical activity (assessed using 7-day accelerometry).This study will include 200 children (100 girls and 100 boys) aged 10-14 years using an a priori power analysis based on the primary objective of assessing construct validity through multiple linear regression, assuming an alpha level of 0.05 and 80% power. Participants will be recruited from paediatric outpatients of the Paediatric Obesity Clinic and individuals reached through a recruitment campaign. Inclusion criteria require general good health, while exclusion criteria include a history or symptoms of cardiovascular, pulmonary, metabolic or neurological disease, as well as the use of over-the-counter or prescribed medications. Informed consent and assent will be obtained from all participants.Reliability of ultrasound-derived EI will be assessed for intra-rater, inter-rater and test-retest agreement using ICC coefficients, Bland-Altman plots and complementary indices such as SE of measurement, coefficient of variation and minimal detectable change at 95% CI, following Consensus-based Standards for the selection of health Measurement Instruments guidelines. Construct validity will be examined by modelling associations between EI and functional muscle quality (MVC/CSAQF), with 30STS as an additional functional measure. Known-groups validity will be tested by comparing EI across weight groups, using generalised linear regression models adjusted for skeletal age, body composition and physical activity. All validity analyses will be conducted separately for girls and boys. Ultrasound-derived EI of the QF is expected to show high reliability (ICC≥0.80) and acceptable test-retest reproducibility. Construct validity should be supported by moderate associations with functional muscle quality (MVC/CSAQF), while known-groups validity is expected to reveal higher EI values in children with obesity and/or insufficient physical activity. ETHICS AND DISSEMINATION: The study will be conducted in accordance with the Declaration of Helsinki and was approved by the Ethics Committee of the Faculty of Physical Education and Sport, Charles University (EK 101/2024). Written parental consent and verbal assent from children will be obtained, with all data handled confidentially and anonymised. Results will be disseminated transparently to participants and their families in line with ethical principles of respect, beneficence and justice. TRIAL REGISTRATION NUMBER: NCT06792279.

Charles University Faculty of Physical Education and Sport Prague Czech Republic

University of Padua School of Medicine and Surgery Padua Veneto Italy

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NCT06792279