We hypothesized that digit formation during foetal stage may be related to testicular development and testicular volume in adulthood. Therefore, this study was performed to investigate the relationship between digit ratio and adult testicular volume.

Testicular volume is known to reflect spermatogenesis potential and testicular function (Arai et al ., 1998 ). However, testicular volume was found to widely vary in large‐scale studies (right testicular volume: 15.9 ± 4.6 mL, left testicular volume: 15.3 ± 4.6 mL (Ku et al ., 2002 ); right testicular volume: 18.7 ± 5.1 mL, left testicular volume: 18.0 ± 5.0 mL (Sakamoto et al ., 2007 ); total testicular volume: 36.8 ± 9.7 mL (Sakamoto et al ., 2008a ); right testicular volume: 18.13 ± 3.85 mL, left testicular volume: 18.37 ± 3.62 mL (Bahk et al ., 2010 )). These findings indicate that adults do not always have similar testicular volumes. Up to now, few studies have been conducted to explain why one male's testicular volume is different from another's testicular volume in spite of normal puberty.

The relationship between male testosterone concentrations during the prenatal period and digit ratio is supported by the finding that Hox genes modulate the development of both digits and testes (Kondo et al ., 1997 ; Mortlock & Innis, 1997 ; Peichel et al ., 1997 ). After the differentiation of Leydig cells within the testes during the 8th week of gestation, testosterone production begins in the male foetus and secretion levels are maximized during the 14th gestational week (George et al ., 1981 ). Thus, many studies have attempted to identify the relationships between digit ratio, sex hormone production and fertility in conjunction with Hox gene expression during foetal development (Zákány et al ., 1997 ). Moreover, a few previous investigations demonstrated that the expression of Hox genes in spermatozoa after meiosis may affect sperm structure and/or activity (Erickson, 1990 ).

The second to fourth digit ratio (digit ratio) is known to be sexually dimorphic. This ratio is essentially established in utero and is fixed before the age of 2 years (Manning et al ., 1998 ). Lutchmaya et al . ( 2004 ) showed that the digit ratio of the right hand is negatively correlated with the foetal testosterone/foetal estradiol ratio. This may mean that a high concentration of testosterone leads to a low digit ratio and indicates high prenatal testicular activity.

The vertebrate Hox gene family is known to play an essential role in limb and genital development (Peichel et al ., 1997 ). In particular, Hoxa and Hoxd genes are indispensable not only for the growth and development of digits but also differentiation of the genital bud (Kondo et al ., 1997 ). In a human study, a mutation within the Hoxa gene was shown to cause hand–foot–genital syndrome, which is associated with anatomical defects in digits and genitalia (Mortlock & Innis, 1997 ). In mice, deregulation of Hoxd expression may affect the relative lengths of digits and influence growth of the genital bud (Kondo et al ., 1997 ; Peichel et al ., 1997 ). These findings demonstrate that a relationship may exist between patterns of digit growth and fertility (Manning et al ., 1998 ).

Relationships between study variables were analysed using Pearson's linear correlation. To identify the independent predictive factors influencing testicular volume, univariate and multivariate analyses were performed using linear regression models. Analyses were performed using SPSS 12.0 (SPSS, Chicago, IL, USA) and p ‐values less than 0.05 were considered statistically significant.

Using orchidometry, the testicular volume was measured by an experienced urologist who had no information about the patients’ digit ratios. Testicular volumes were obtained after stretching the scrotal skin over the testis in a warm room by comparing the testes with 12 solid ellipsoid models constituting the Prader orchidometry and ranging in volume from 1 to 25 mL (1–6, 8, 10, 12,15, 20 and 25 mL).

Before determining testicular volume, the second and fourth digits of the right hand were measured by a single investigator using a digital vernier calliper accurate to 0.01 mm. Digit length was measured on the ventral surface of the hand from the basal crease of the digit to the tip (Manning et al ., 1998 ). To reduce errors in measurement, the mean values of two digit ratios that were calculated from the duplicate measurements were used in the analysis. The intraclass correlation coefficient (ICC) of two repeated measures of digit ratio by a single investigator was 0.918, which was similar with that of the previous studies (ICC = 0.873–0.969) (Allaway et al ., 2009 ). This method is known to be associated with a high degree of repeatability (Scutt & Manning, 1996 ).

Patients who suffered from infertility and were receiving LHRH analogues or testosterone replacement therapy were excluded from the analysis. In addition, patients with a history of endocrine disease, testicular cancer, varicocoele, epididymoorchitis, testicular trauma, hydrocele/hernia, testicular torsion, cryptorchidism, or other scrotal surgical procedures that significantly affect testicular volume were also excluded from the analysis.

Furthermore, when we analysed data of married patients with more than one child ( N = 156), the results were similar to those of whole study population ( N = 172) (Tables 5 and 6 ).

Multivariate analysis using linear regression models showed that digit ratio was the independent factor to predict all (right, left and total) testicular volumes (right testicular volume: β = −0.174, p = 0.023; left testicular volume: β = −0.181, p = 0.017; total testicular volume: β = −0.185, p = 0.014) (Table 4 ). However, weight was not independent factor to predict right testicular volume (Table 4 ). As shown in Fig. 1 , right and left testicular volumes were found to be negatively associated with digit ratio (Fig. 1 ).

Almost all patients (156/172, 90.7%) were married and had more than one child (2.1 ± 0.8 children) (Table 2 ). Comparing data between married ( N = 156) and unmarried ( N = 16) persons, there were significant differences only in age and number of offsprings. However, there were no significant differences between the two groups in height, weight, digit ratio, testosterone level and testicular volumes (Table 2 ).

The characteristics of all 172 participants are summarized in Table 1 . Mean age, height, weight and digit ratio of all subjects were 55.1 years, 169.3 cm, 68.9 kg and 0.941 respectively. Mean serum testosterone and free testosterone levels were 337.07 ng/dL and 8.77 pg/mL respectively. Mean right, left and total testicular volumes were 18.1, 17.8 and 35.9 mL respectively (Table 1 ).

Discussion

The testes have been known to play a main role in the production of spermatozoa and testosterone in male reproductive system. Testis size reflects spermatogenesis because seminiferous tubules account for about 80% of the testicular mass, and it is generally considered as an index of reproductive function (spermatogenesis or semen analysis) (Aribarg et al., 1986; Takihara et al., 1987; Wu et al., 1993; Arai et al., 1998). Accordingly, reduction in testicular volume reflects decrease in the number of germ cells and seminiferous tubules (Lipshultz & Corriere, 1977). Consequently, measuring of testicular volume may help evaluate patients with various disorders associated with testicular growth and function (Scutt & Manning, 1996; Arai et al., 1998).

In general, the normal range of testicular volume has been reported to be 14–30 mL (Takihara et al., 1987; Lenz et al., 1993; Arai et al., 1998; Sakamoto et al., 2008a,b). However, these data of human testicular volumes showed marked differences according to ethnicity (Dave & Tiwari, 1983; Diamond, 1986; Mittwoch, 1988). In fact, testicular volume of Caucasian has been reported larger than that of Oriental populations (Mittwoch, 1988). Compared with the results of Auger & Eustache (2011), our study population had smaller testicular volumes (total testicular volume: 35.9 ± 8.6 vs. 55.7 ± 18.7 mL). Thus, it is not appropriate to compare our Korean data with other Caucasian data directly. However, testicular volumes in this study were similar to those of large‐scale studies of healthy young Korean men (Ku et al., 2002; Bahk et al., 2010). These mean that the testicular volumes of our study population were normal since we excluded patients with conditions known to strongly influence testicular volume.

Several studies have been reported to prove the positive relationship between testicular volume and body weight (Aribarg et al., 1986; Ku et al., 2002; Bahk et al., 2010). In this study, both digit ratio and body weight had a significant correlation with testicular volumes in univariate analysis (Table 3). However, in multivariate analysis, body weight was not independent factor to predict right testicular volume. Only digit ratio was the independent factor to predict all (right, left and total) testicular volumes (Table 4). This is one of the novel findings in this study.

In addition, we found that the patients with a higher digit ratio may be more likely to have smaller testes compared to those with a lower digit ratio (Tables 3 and 4, Fig. 1). These results are similar to ones from previous studies of the relationship between digit ratio and fertility. Auger & Eustache (2011) showed that digit ratios are significantly and negatively associated with testicular volume and male reproductive health. Other investigations demonstrated that men with low digit ratios have high sperm counts and large family sizes (Manning et al., 1998). Moreover, Wood et al. (2003) asserted that testicular sperm retrieval in azoospermic men is more successful in men with low digit ratios than in ones with high digit ratios.

These findings are also supported by ones from studies assessing ethnic differences in digit ratio and testicular volume (testicular function) between Finnish and Danish males. In one study, Finnish males were shown to have relatively lower digit ratios than those of other European populations (Manning, 2002). Another study demonstrated that Danish men have a very high mean digit ratio (digit ratio = 1.02 ± 0.04) compared with other ethnic groups (Bang et al., 2005). Thus, these indicate that the digit ratio of Finnish males is lower than that of Danish males.

Similar to digit ratio, testicular volume also varied between these two ethnic groups. Main et al. (2006) demonstrated that Danish boys have smaller testicular size in comparison with Finnish boys. Furthermore, there is also considerable evidence that the degree of reproductive health among Danish men is lower than that of Finnish males. Finnish males were found to have better semen quality than Danish males (Jensen et al., 2000; Jørgensen et al., 2002). In addition, there is a higher prevalence of cryptorchidism and hypospadias among newborn males in Denmark compared with the ones in Finland (Boisen et al., 2004).

Like these, several studies about the racial difference of digit ratio and testicular volume (testicular function) may support that testicular volume and function in adulthood might be associated with in utero milieu (prenatal androgen exposures). Dysfunctional germ cell proliferation and differentiation during embryogenesis might result in male infertility (Lue et al., 2007). Primordial germ cells differentiate into gonocytes that subsequently transform into foetal spermatogonia between the 10th and 22nd week after conception (Ehmcke et al., 2006). Digit ratio is associated with prenatal testosterone levels (Berenbaum et al., 2009) and the balance of androgen/oestrogen concentrations during foetal development (Lutchmaya et al., 2004). Therefore, digit ratio could reflect the hormonal environment of the foetal testis. In addition, finger development and genital bud differentiation are also regulated by Hox genes during embryogenesis. In this study, adult testicular volume was associated with digit ratio that is reflective of the in utero environment, particularly prenatal androgen levels. This means that testicular volume and function in adulthood might be determined during the foetal period.

Although digit ratio is known to be an indicator of prenatal testosterone that impacts the development of male genital and testicular growth, digit ratio in this study was not associated with serum testosterone or free testosterone level (Table 3). These findings are similar to those of previous studies about the association between digit ratio and adult sex hormone levels (Bang et al., 2005; Hönekopp et al., 2007).

When we consider the range of testosterone level, the value of standard deviation in this study was similar to that of other Korean study. In a large‐scale study of 2172 Korean men aged 21–79 years, the mean total testosterone level was 487.61 ± 152.74 ng/dL (Hong et al., 2013). This value of standard deviation is similar to that of the present study (testosterone level: 337.07 ± 121.62 ng/dL). This means that the range of testosterone level in this study is not very large. However, testosterone level in our study was found to be lower than that of the large‐scale Korean study. We think that the difference of testosterone level between the two studies may come from the difference of patients’ age between the two studies (55.1 ± 12.0 vs. 49.4 ± 8.8 years).

We excluded the patients who suffered from infertility. Thus, almost all patients (156/172, 90.7%) in our study were married and had more than one child, which means that the fertility of our study population was indirectly proven. Unlike our study, Auger & Eustache (2011) enrolled 122 men who were partners of pregnant women.

Furthermore, our study first reported digit ratio as the only independent factor to predict all (right, left and total) testicular volumes in multivariate analyses using linear regression models. Although Auger & Eustache (2011) showed the association between digit ratio and fertility, they did not consider confounding factor such as body weight to clarify the independent factor to predict testicular volumes.

Although there were the differences in testicular volumes according to racial differences, this study showed that digit ratio in Oriental (Korean) population was also correlated with testicular volumes. Furthermore, this study also supports their suggestions that digit ratio could represent a useful indicator of fertility.

The major limitation of the study is that it is not based on a general population. Participants in our investigation were recruited from among patients who were hospitalized for urological surgery at a single tertiary academic centre. Thus, our study population was relatively older (age: 55.1 ± 12.0 years) than that of other studies and may therefore not accurately represent the general population.

Another major limitation of the study is that semen analyses were not performed. However, we excluded the patients who suffered from infertility. Thus, in this study, almost all patients (156/172, 90.7%) were married and had more than one child. We think that this might show enough reproductive information. Furthermore, testicular volumes in this study were similar to those of large‐scale studies of healthy young Korean men (Ku et al., 2002; Bahk et al., 2010). Therefore, we believe that our study has produced sufficient evidence of a relationship between digit ratio and adult testicular volumes.

In summary, this study showed that digit ratio was negatively associated with adult testicular volume. Furthermore, digit ratio was found to be the only independent factor to predict all (right, left and total) testicular volumes in multivariate analyses. Thus, it could be suggested that digit ratio may be indicative of testicular size and function in adulthood.