Participants

This cross-sectional observational study used a convenience sample. Participants were recruited between March 30, 2023 and June 30, 2023. During this period, we enrolled 140 male middle school students affiliated with a soccer team, resulting in a cohort of 280 knees. Individuals with substantial lower limb deformities, history of lower limb surgery, or neurological symptoms affecting the lower extremities were excluded from the study cohort.

Experimental preparation

In a preliminary questionnaire, participants were asked to provide information regarding their height, weight, age, sex, dominant foot, medical history, and years of soccer experience. The dominant foot was defined as the foot used to kick the ball.

US evaluation

Patellar tendon thickening, hypoechoic areas, and neovascularization were observed using a high-frequency linear probe (L11–3; 3–11 MHz) attached to an ultrasound system (SONIMAGE MX1; Konica Minolta, Inc., Tokyo, Japan). The imaging depth was fixed at 3.5 cm for all participants, and gain settings were adjusted by the examiner at the beginning of each examination to ensure adequate visualization of the tendon. Patellar tendon thickening was measured by observing the patellar tendon in the longitudinal plane and measuring its thickness at a point 5 mm distal to the inferior pole of the patella [14]. The hypoechoic areas of the patellar tendon were assessed in the longitudinal direction by observing the area from the attachment site at the patella to the tibial tuberosity. Neovascularization was assessed using the color Doppler method. The measurements were conducted exclusively by a physical therapist with 5 years of experience in US.

Data processing

Muscle strength was evaluated twice, and the highest value was recorded and normalized to the body weight. Tendon thickness was defined as the distance between its superficial and deep layers [27], and it was measured using ImageJ (version 1.54) [14]. In accordance with a previous study, if a hypoechoic area measuring ≥ 2 mm in length and width was observed in the patellar tendon, it was considered hypoechoic-positive (+) [28]. Neovascularization was assessed by observing vascular structures longer than 1 mm in the sagittal plane of the tendon, classified as positive (+) for Doppler [14]. The image analysis process demonstrated high intra-rater reliability [14]. To minimize bias, the analyst was not informed of whether each knee was painful or not during the image evaluation.

Statistical analysis

The normality of data was assessed using the Shapiro–Wilk test. As the data were non-normally distributed, the Mann–Whitney U test was used to compare knees with and without patellar tendon-related pain. The proportions of hypoechoic- and Doppler-positive findings were compared using the chi-square test. Variables with a p-value < 0.05 in the initial analysis were included as independent variables, and the presence or absence of pain was set as the dependent variable in the logistic regression analysis using the forced entry method. The goodness of fit of the logistic regression model was assessed using the Hosmer–Lemeshow test. To address the potential non-independence of bilateral knees, a sensitivity analysis at the participant level was performed using one knee per participant; the painful knee was prioritized when present. However, for knee extension strength, the mean value of both knees was used to provide a single representative value for each participant. The same statistical procedures were applied as in the primary analysis. All statistical analyses were performed using SPSS (version 22.0; SPSS Inc., Chicago, IL, USA), with a significance level set at < 5%.

Ethics statements

This study was conducted in accordance with the Declaration of Helsinki. The Research Ethics Committee of Kio University (reference number: R4-48) approved this study, and all participants and their parents provided written informed consent before participation in this study. The authors did not have access to identifiable personal information during or after data collection.

Main findings

The findings of the present study support our hypothesis, as patellar tendon-related pain was significantly associated with hypoechoic findings. Furthermore, the multivariable logistic regression analysis demonstrated that hypoechoic findings in the patellar tendon were significantly associated with pain, whereas physical function measures showed weaker or no significant associations.

The participant-level sensitivity analysis yielded results that were largely consistent with those in the primary knee-level analysis, particularly regarding the significant association between hypoechoic findings and pain. Although Doppler findings reached statistical significance in the sensitivity analysis, this was not observed in the multivariable model, suggesting that its association with pain may be less robust and may have been influenced by the analytical approach or the limited sample size. Therefore, these findings should be interpreted cautiously, as the participant-level analysis was intended to support, not replace, the primary knee-level analysis.

Prevalence rate of patellar tendon-related pain

The prevalence rate of OSD is reported to be between 9.8% and 16% [6,10,29], whereas that of jumper’s knee ranges from 8.5% to 14.2% [30,31]. In the present study, the prevalence of patellar tendon-related pain disorders was 9.4%. Although no formal diagnosis was made, the prevalence observed in this study was comparable to those reported in previous studies.

Relationship between patellar tendon-related pain and morphological abnormalities of the patellar tendon

Hypoechoic findings commonly reflect inflammation-related changes, such as edema or fluid accumulation and are often associated with pain [17,32]. Reportedly, soccer players impose a high mechanical load on the patellar tendon due to the sport’s substantial demands on leg extensor velocity and power [33]. In the adolescent soccer players included in this study, prolonged exposure to mechanical stress contributed to microdamage in the patellar tendon, leading to increased proteoglycan production, collagen separation, and angiogenesis [34,35]. The proliferation of proteoglycans increases the water content within the tendon [33], which is visualized as a hypoechoic finding on US. Structural damage to the tendon has been demonstrated to be correlated with pain severity, with a higher likelihood of increased pain intensity [17]. These findings suggest that the association between a hypoechoic finding and patellar tendon-related pain in this study may be attributed to these pathological changes.

Doppler findings are considered to reflect neovascularization and have been suggested to be associated with pain in tendon disorders [36,37]. In contrast, other studies have reported that Doppler findings do not necessarily indicate tendon-related pain [38]. Similarly, in this study, we did not identify a significant association between Doppler-detected neovascularization and patellar tendon pain. Previous research has indicated that Doppler signals may persist even after pain resolution and can be observed for as long as 3–4 years following the onset of patellar tendinopathy [36]. Doppler findings suggest that neovascularization alone may not be sufficient to elicit pain and that other factors, such as neural ingrowth [39] and biochemical mediators [40], may contribute to symptom manifestation. Therefore, Doppler US findings should be interpreted with caution and in conjunction with clinical symptoms when assessing tendon pain. Recent evidence further supports the multifactorial nature of patellar tendon-related pain. Tayfur et al. [41,42] reported that, in addition to structural changes, psychosocial and clinical factors are significantly associated with patellar tendinopathy in jumping athletes, suggesting that ultrasonographic abnormalities represent only one aspect of the pain experience. Moreover, a recent systematic review and meta-analysis demonstrated that altered landing mechanics and impaired neuromuscular control are also associated with patellar tendinopathy. Together, these findings indicate that tendon-related pain arises from the interaction of structural, psychosocial, biomechanical, and neuromuscular factors, and they cannot be fully explained by imaging findings alone. Previous studies have reported that repetitive overload induces a non-inflammatory proliferative response, resulting in uniform thickening of the tendon and an increase in its cross-sectional area, which helps alleviate the stress (force per unit area) applied to the tendon [34,35]. The thickness of the patellar tendon itself is not necessarily linked to pain; rather, it may be a physiological response to maintaining tendon strength [43]. Additionally, in adult athletes, it has been reported that a patellar tendon thickness ≥ 4 mm at the proximal insertion site can be interpreted as an early sign of degeneration [40]. In the present study, the mean patellar tendon thickness in the pain and no-pain groups was < 4 mm. These findings suggest that, although excessive thickening of the patellar tendon may be associated with pain beyond a certain threshold, hypertrophy itself is a physiological response to maintaining tendon strength, which may explain why no association with pain was observed in this study. However, data on the relationship between patellar tendon hypertrophy and pain in middle school students remain limited; this subject therefore warrants further investigation.

Limitations

This study has certain limitations. The sample size was small, which may limit the generalizability of the findings. Additionally, data from 69 knees were excluded owing to missing measurements or lack of consent, as illustrated in the study flowchart. Although missing data were primarily related to logistical or consent-related issues, the possibility that the missingness was not completely random cannot be excluded, and this may have introduced selection bias. This study was also limited to middle school students in soccer teams. Due to differences in the growth process, it remains unclear whether the findings are applicable to other age groups, such as elementary or high school students. Although an association between hypoechoic findings and pain was observed, the cross-sectional design precluded determination of causality. Hypoechoic areas were defined as ≥ 2 mm based on a prior study [28]; however, evidence supporting this cutoff value is limited, and the optimal threshold remains unclear. Hypoechoic areas may reflect various underlying conditions, including ones that are reversible early and have specific pathological changes associated with pain. Further, all ultrasonographic measurements and image analyses were performed by a single examiner. Intra-rater reliability was assessed and found to be acceptable; however, inter-rater reliability was not evaluated, which may have introduced potential measurement bias. The number of painful knees was limited, resulting in a low events-per-variable ratio in the multivariable logistic regression analysis. Although clinically relevant variables were prespecified to reduce the risk of overfitting, the limited number of events may have affected the stability of the regression estimates. Finally, all participants were soccer players, which may have introduced selection bias and limited the generalizability of the results to athletes of other sports. Additionally, other factors that may influence the presence of hypoechoic changes and pain, such as training volume, level of sports participation, and type of sport, were not thoroughly investigated.