Analysis of factors associated with undescended testis in patients with congenital diaphragmatic hernia

This study aimed to investigate the incidence and clinical factors associated with undescended testes (UDT) in patients with congenital diaphragmatic hernia (CDH). We retrospectively reviewed the incidence of UDT in male neonates admitted to our institution and underwent surgery for CDH between January 2006 and December 2022. Patients were divided into two groups based on the presence or absence of UDT, and risk factors for UDT were compared between the two groups. Among the 66 male neonates with CDH, 16 (24.2%) developed UDT. Patients with UDT had a significantly smaller gestational age (p = 0.026), lower birth weight (p = 0.042), and lower Apgar score at 1 min (p = 0.016) than those without UDT. They had a significantly higher incidence of large diaphragmatic defects (p = 0.005), received more patch closures (p = 0.020), had a longer mechanical ventilation period (p = 0.034), and longer hospital stay (p = 0.028). Multiple logistic regression analysis revealed that large diaphragmatic defect was an independent risk factor for UDT (adjusted odds ratio of 3.87). CDH and UDT are strongly correlated. In patients with CDH, the incidence of UDT was related not only to patients’ prematurity but also to the large diaphragmatic defect. Large diaphragmatic defect is an independent risk factor for UDT in patients with CDH.


Introduction
Congenital diaphragmatic hernia (CDH) is a relatively frequent congenital malformation, with an incidence of approximately one in 2000-3000 live births [1,2].It causes various degrees of respiratory distress, and the reported frequency of associated malformations ranges from 27 to 63% [3][4][5][6] The wide variation in the incidence of associated anomalies in CDH is due to inconsistent definitions across studies.A recent population-based cohort study on CDH revealed that 64% of the cases were isolated, 23% were multiples (additional unrelated major or unique minor malformations), and 13% were syndromic (genetic or chromosomal) [7].According to these reports, associated major and minor anomalies are frequent in patients with CDH.
UDT is a minor but common anomaly among those associated anomalies in male patients with CDH.Although the incidence of UDT in full-term infants is reported to be around 3% [8], some studies have reported a high incidence in patients with CDH [2,[8][9][10][11][12].However, the mechanisms underlying testicular maldescence in patients with CDH have not been elucidated, and clinical factors related to UDT in these patients are poorly understood.
Therefore, this study aimed to investigate the incidence of UDT and identify its risk factors in infants with CDH.

Methods
The data of male neonates admitted to our institution and who underwent surgery for CDH between January 2006 and December 2022 were included in this study.Patients who died before one year of age were excluded.We retrospectively reviewed the patients' medical records to obtain the following clinical information: gestational age, birth weight, Apgar score, prenatal diagnosis of CDH, the delivery method, diaphragmatic defect size, the diaphragmatic repair method, best oxygenation index (OI) within 24 h after birth, length of mechanical ventilation, length of hospital stay, and presence of concomitant malformations.The presence of liver-up, lung-to-thorax transverse area ratio, and observed/expected lung area-to-head circumference ratio were reviewed in patients with a prenatal CDH diagnosis.The diaphragmatic defect size was divided into two groups: small and large defects.Minor defects included defects A and B, and the large defects included defects C and in the standardised classification of diaphragmatic defect size [13].Subsequently, patient data were analysed to obtain information on spontaneous testicular descent during the observation period and the need for orchidopexy.We divided the patients into two groups based on the presence or absence of UDT and compared the clinical factors related to UDT between the two groups.
Values were expressed as median values and ranges.Statistical analyses were performed using the JMP software ver.13.0 (SAS Institute Inc., Cary, NC, USA).Analyses of categorical variables were performed using the chi-square test.Comparisons of means or medians were performed using the Mann-Whitney U test.Factors identified as significant (p<0.05) in the univariate analysis were input into multiple logistic regression analyses to determine independent risk factors and odds ratios (ORs).Statistical significance was defined as p < 0.05.

Results
The study included 66 male neonates who underwent surgery for CDH at our institution and survived for > 1 year during the study period; 16 (24.2%)developed UDT.
Table 1 illustrates the clinical features of patients with CDH with and without UDT.Those with UDT had a significantly smaller gestational age (p = 0.026), lower birth weight (p = 0.042), and lower Apgar score at 1 min (p = 0.016) than those without UDT.Additionally, patients who developed UDT had a significantly higher large diaphragmatic defects incidence (p = 0.005), received more patch closures (p = 0.020), had a longer mechanical ventilation period (p = 0.034), and extended hospital stays (p = 0.028).Despite a trend towards a higher incidence of liver-up, lower incidence of isolated CDH, and higher best OI within 24 h after birth in patients with UDT than in those without UDT, these differences were not statistically significant (p = 0.118, p = 0.090, p = 0.062, respectively).Other clinical features showed no significant differences between patients with CDH associated with UDT and those without.Univariate analyses revealed that, among the seven factors significantly related to the development of UDT, gestational age and length of mechanical ventilation were not significant.Multiple logistic regression analysis revealed that large diaphragmatic defect was an independent risk factor for UDT (adjusted OR of 3.87) (Table 2).
There were four cases of non-isolated CDH in each group.These included congenital tracheal stenosis, single atrium single ventricle, Matthew-Wood Syndrome, and One patient had right-sided CDH among the 16 patients with CDH associated with UDT.The affected side of the UDT was bilateral in six patients, left-sided in eight, rightsided in two, and 22 testes were undescended.Most of the locations of the UDT were in the inguinal canal, with an incidence of 81.8%; three patients had testes in the upper scrotum, and one had an intra-abdominal testis.All 16 patients who developed UDT required subsequent orchidopexy.The median age at the time of orchidopexy was 13 months (Table 3).

Discussion
The study incidence of UDT in male patients with CDH was 24.2%, which was significantly higher than the incidence of UDT in the normal population [14].In CDH patients with UDT, gestational age, birth weight, and Apgar score at 1 min were significantly lower than in those without UDT.In addition, CDH patients with UDT had significantly higher incidence of large diaphragmatic defects and patch closure.They required longer mechanical ventilation periods and extended hospital stays.
Several studies have reported an association between CDH and UDT and cited incidence rates ranging from 3.6 to 30% [9][10][11][12].Janssen et al. reported that 36% of male patients with CDH were diagnosed with UDT, of whom 29% underwent orchidopexy.They demonstrated no significant differences between the UDT and non-UDT groups regarding demographics or CDH-specific characteristics, including birth weight, gestational age, method of diaphragmatic repair, surgical approach, or use of Extracorporeal Membrane Oxygenation (ECMO) [2].In contrast, Azarow et al. demonstrated that 18% of male CDH patients required orchidopexy.There was no difference between patients with and without UDT, except for the need for ECMO, patch closure, and oxygen dependence at 28 days of life [9].Although a high incidence of UDT in patients with CDH has been recognised, the clinical factors related to UDT in these patients were poorly understood over a long time.Our study is the UDT is strongly associated with both premature birth and small foetal size [15], as 15-30% of premature males have UDT in the first few months after birth [16].Similar to previous reports, our data demonstrated significant differences in measures of immaturity, such as birth weight and gestational age, between patients with and without UDT.In addition, multiple logistic regression analyses revealed that large diaphragmatic defect was an independent risk factor for UDT.These results suggest that the size of the diaphragmatic defect may be more crucial than prematurity in the development of UDT in patients with CDH.
Recent studies have reported that the incidence of UDT in patients with abdominal wall defects is as high as 40% [17,18].Various theories, including mechanical, hormonal, and genetic mechanisms, have been proposed to describe testicular descent [15,19].The effect of intra-abdominal pressure on testicular descent has been reported in several studies.For example, an experimental rat model has demonstrated that intra-abdominal pressure contributes to testicular descent [20].In addition, we reported that the incidence of UDT in gastroschisis was high and was related to the defect size of the abdominal wall and the timing of abdominal wall closure, indicating an insufficient increase in intra-abdominal pressure during the foetal period may affect the development of UDT [21].The present study revealed a significantly higher incidence of large diaphragmatic defects and patch closure, and CDH patients with UDT required longer mechanical ventilation and hospital stay.Furthermore, it was revealed that large diaphragmatic defect was an independent risk factor for developing UDT.These results indicate that size of the diaphragmatic defect might be related to the development of UDT in patients with CDH, which implies that an insufficient increase in intra-abdominal pressure during the foetal period might contribute to the development of UDT in patients with CDH, as well as to the associated abdominal wall defect.
UDT incidence among full-term infants declines from 3% at birth to approximately 1% at 3 months and remains approximately 1% at 1 year [16,22].Therefore, spontaneous testicular descent during early infancy is observed in more than half of the UDT cases in the general population.In the present study, all patients who developed UDT eventually required orchidopexy.Our study may have missed a patient who had UDT at birth, followed by spontaneous testicular descent and did not require orchiopexy.In addition, some cases had gradually elevated testes later, as in cases 4, 6, 7, and 8.Although the speculation that corrective surgery for CDH causes a secondary testicular ascent owing to operative handling and scar tissue formation pulling at the retroperitoneum cannot be denied [2], the true mechanisms of secondary testicular ascent in CDH patients are still unclear.Hence, careful long-term observation of the testicular location is necessary when male patients with CDH are followed-up.
Regarding the laterality of CDH and UDT, although leftsided CDH is much higher than right-sided CDH [23], the prevalence of left-sided UDT is similar to that of right-sided UDT in the normal population [24].Janssen et al. reported that only 3 out of 27 patients developed unilateral UDT contralateral to the side of the CDH, and 8 out of 27 patients developed bilateral UDT [2], while Azarow et al. reported that no patient developed unilateral UDT contralateral to the side of the CDH and 2 out of 12 CDH patients developed bilateral UDT [10].Thus, Azalow et al. suggested that the absence of diaphragm tissue may disrupt the regression of the craniosuspensory ligament, resulting in testicular maldescent in CDH [9].Only 3 of the 16 patients had a unilateral UDT contralateral to the side of the diaphragmatic defect.Of the 15 patients with UDT and left-sided CDH, 13 had unilateral or bilateral UDT.From our results, as advocated by Azarow et al., the absence of diaphragmatic tissue might impact UDT in patients with CDH.
The present study had several limitations.First, this retrospective cohort study included only a small number of patients with CDH.Second, the parameters of disease severity in this study, such as defect size, method of diaphragmatic repair, presence of liver-up, and lung-to-thorax transverse area ratio, do not directly reflect intra-abdominal pressure during the process of foetal testicular descent.Third, as CDH is a relatively rare and life-threatening condition, some patients with CDH with UDT may have been missed, suggesting the true incidence of UDT in patients with CDH is difficult to assess.Therefore, further prospective studies with more patients must validate our results and clarify the key factors associated with male testicular descent in patients with CDH.
In conclusion, we observed a strong association between CDH levels and testicular descent.In patients with CDH, UDT incidence is related to the prematurity of patients and CDH severity.A large diaphragmatic defect was an independent risk factor.These results indicate that the diaphragmatic defect size may be crucial in developing UDT in patients with CDH.

Table 1
Comparisons between CDH patients with and without UDT *Statistically significantUDT undescended testis, CDH congenital diaphragmatic hernia, L/T ratio lung-to-thorax transverse area ratio, o/e LHR observed/expected lung area-to-head circumference ratio, OI oxygenation index

Table 2
Univariate and multivariate analysis of risk factors for the development of UDT in CDH