Many observations have demonstrated great heterogeneity in the magnitude of the response to the physical exercise stimulus. Only a few studies have investigated the effects of high-intensity interval training (HIIT) on the prevalence of non-responders (NRs) in adolescents while considering health-related fitness measurements and other co-variables. Therefore, the aim of the current work was twofold: (1) to assess the effects of ten weeks of school-based HIIT on the prevalence of responders (Rs) and NRs for body composition, resting blood pressure (BP), cardiorespiratory and musculoskeletal fitness, and (2) to assess the relationship of this prevalence with sex, body mass category, and biological maturation status, while estimating the likelihood of a response. Adolescents aged 16 years (n = 73) were included in the study. Waist-to-hip ratio, body fat percentage (BFP), skeletal muscle mass (SMM), BP, cardiorespiratory fitness (CRF), and musculoskeletal fitness were the primary outcomes. Co-variables included sex, body mass index (BMI), and maturity offset. The percentages of the Rs and NRs were analyzed based on changes (Δ) between post-intervention and pre-intervention values. The typical error (TE) method was used to identify Rs and NRs. Results showed a variation in the relative size of changes (% of changes) depending on the variable and sex. The greatest changes were observed in females’ abdomen muscle strength (Δ% = 23.89%), and the difference from that of males (Δ% = 5.98%) was statistically significant (p < 0.001) with very large effect size of (Cohen’s d = 0.941). Similar significant differences in relative changes were observed in body composition, but in the reverse direction. Males gained more from HIIT in the decrease in the body fat percentage (Δ% = −8.24%) and increase in skeletal muscle mass (Δ% = 3.38%) comparing to females (Δ% = 0.46, p = 0.040 and Δ% = 1.06%, p = 0.007, respectively). Effect size for body fat was small (Cohen’s d = 0.469), and for skeletal muscle mass it was moderate (Cohen’s d = −0.659). The results also showed positive significant differences in the prevalence of Rs compared to NRs in BFP, SMM, systolic BP (SBP), diastolic BP (DBP), CRF, and abdominal muscular strength (ABS) (p = 0.047, p = 0.047, p < 0.001, p = 0.003, p = 0.014, and p = 0.014, respectively). The effect was greatest and largest for SBP (Cohen’s ω = 0.67). Sex was related to the prevalence of Rs for ABS and close to significance for SMM. More males than females benefited from HIIT in SMM (p = 0.058), but more females than males had ABS benefits (p = 0.050). Males were more likely (2.5-fold) to be Rs than females in SMM. BMI was not related to the prevalence of Rs and NRs. Biological maturation was related to the prevalence of Rs and NRs in SMM (p = 0.036) and SBP (p = 0.016). In SBP, 100% of the early matured derived benefit from HIIT, but the effect size was small (Cramer’s V = 0.27). Those who maturated later were more likely (almost 70%) to be Rs. Thus, the HIIT program introduced to a typical physical education (PE) lesson can be considered partially effective. Therefore, there is a need to continue the search for an optimal and effective program for all health-related parameters. The close relationship between analyzed values and sex and biological maturation, but not BMI, indicates that the intervention programs should be tailored specifically for males and females.
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