This correlation analysis suggests that cocoa consumption during early life might be correlated to both TC incidence among young men aged 20–34 years and hypospadias, a reproductive congenital defects supposed to underline the same pathogenetic mechanism of TC.
In particular, the increasing incidence of TC over time in developed countries is consistent with increases in the consumption of cocoa. The intake of some of the main sweet ingredients, in fact, such as cocoa and sweeteners increased significantly during the last 45 years in developed countries [8
]. Over the years 1961–2004 cocoa consumption overall in developed countries grew at an average rate of 1 % per year [9
]. For example, in Denmark cocoa consumption more than doubled in the period 1961–2004. Similar trends have been observed in other European countries [8
]. The magnitude of these increases is similar to those noted for incidence rates of testicular cancer and other MRD. The reduced risk for testicular cancer for the cohorts of Danish, Norwegian, Swedish and Japanese men born during World War II is also consistent with a greatly reduced supply of sweet foods and cocoa during the war.
Data from food balance sheets (FAOSTAT) indicate that the consumption of cocoa in Denmark, where TC and hypospadias rates are elevated, is among the highest in the world and is more than three times that in Finland, where MRD rates are rather low [9
Is there any evidence that this association may be causal?
Cocoa powder is a complex substance containing several biologically active compounds, including theobromine, caffeine, serotonin, phenylethylamine and cannabinoid-like fatty acids [11
Various studies reported that theobromine, the main stimulant of cocoa, exerts toxic effects on the testis, inducing testicular atrophy accompanied by aspermatogenesis or oligospermatogenesis and that even low doses of cocoa impair sperm quality [10
Friedman et al.
reported that feeding theobromine to male Osborne-Mendel rats at a dietary level of 0.5% for 64 weeks resulted in severe testicular atrophy in 94% of animals, with aspermatogenesis in 82% [20
]. The results were confirmed in another strain of rats; following 19 weeks of feeding theobromine, all rats showed atrophy, and 79% had aspermatogenesis [10
Tarka et al.
found that feeding theobromine at levels of 0.2–1.0 % in the diet (90–140 to 500–600 mg/kg bw per day) for a period of 28 days to rats produced severe testicular atrophy at the 0.8% level and seminiferous tubular-cell degenerationat the 0.6% level [13
]. These authors also studied the potential reversibility of this phenomenon by feeding proven breeder male Sprague-Dawley Rats 0.2, 0.6 or 0.8% theobromine (88, 244 or 334 mg/kg bw per day, respectively) for 49 days, performing unilateral orchiectomy at that time and allowing rats to recover on a theobromine-free diet for an additional 49 days. Histologically, the effects at the two highest dose levels were largely irreversible [13
Funabashi et al.
administered orally theobromine to male Sprague-Dawley rats at dose levels of 250 and 500 mg/kg for 2 weeks starting at the age of 6 or 8 weeks, and for 4 weeks from the age of 6 weeks [21
]. Histopathological examination of reproductive organs revealed toxic findings in the testis such as degeneration/necrosis and desquamation of spermatids and spermatocytes, vacuolization of seminiferous tubules, and multinucleated giant cell formation at 500 mg/kg after 2 weeks of dosing at both ages, and at 250 and 500 mg/kg after 4 weeks of dosing [21
There was also genetic effects of theobromine in animals. In vivo
, theobromine induced sister chromatid exchange and micronuclei in the bone marrow of Chinese hamsters. In human cells in vitro
, theobromine induced sister chromatid exchange and chromosomal breaks. In cultured mammalian cells, it induced gene mutations and sister chromatid exchange [10
]. In particular, theobromine was found to produce a higher and more linear rate of sister chromatid exchange damage than caffeine [10
]. In addition, progeny from the mice fed chocolate presented considerable morphometric abnormalities in the kidney structure, with the lower number of glomeruli per mm2
and their increased diameter [15
These findings from animal studies can be compared with the only human study on exposure to theobromine during pregnancy and early childhood [22
]. This study examined the level of theobromine in mother’s milk after ingestion of 113 g (4 ounces) milk chocolate (containing a total of 240 mg of theobromine). Peak theobromine concentrations of 3.7 to 8.2 mg/L were found in all fluids including breast milk at 2 to 3 hour after ingestion of the chocolate [22
]. According to the European Food Safety Authority (EFSA), if this amount of chocolate were to be ingested four times a day, it would potentially lead to an exposure of the breast fed infant of about 10 mg theobromine/day (corresponding to 1–2 mg/kg b.w.) [23
]. Such an exposure might result in pharmacologically active theobromine levels since newborn babies and infants have very low CYP1A2 activity and would metabolise theobromine much more slowly than adults [23
An alternative explanation of the relationship between cocoa consumption and occurrence of MRD might be that consumption of cocoa and chocolate-containing confectionery could be related to the hormonal equilibrium of the mother during pregnancy and of the infant in the development of MRD in later life. Consumption of energy-dense foods during pregnancy or childhood could negatively effect the balance between free and bound estrogens. The foetus could be exposed to increased estrogen levels through the availability of the high maternal estradiol levels occurring during pregnancy. Bioavailability of estradiol during pregnancy is mainly regulated by the levels of sex hormone-binding globulin (SHBG), whose most important endogenous suppressor is insulin [24
]. It is well-established that sweet and cocoa consumption is associated with increased levels of insulin. In fact, though cocoa and chocolate-containing confectionery elicits only a low glycemic response due to its poor sugar content, insulin responses to chocolate is disproportionately high, being almost 50–60 % higher than its glucose score [25
Fung et al.
found recently that a Western pattern, which represents a higher intakes of sweets and desserts, was associated with a higher level of estradiol and lower concentration of SHBG in women [26
In particular, it was observed that pregnancy intake of sweets was significantly related to the reduction of serum sex hormone-binding globulin (SHBG) at the 16th gestational week [27
]. The exposure to free estrogens during pregnancy was suggested to increase the risk of MRD in the offspring later in life [4
Finally, cocoa beans are some of the principal dietary sources of ochratoxin A, a mycotoxin that causes adducts in testicular DNA of rats and is an established cause of liver and kidney tumors in rodents and is a testicular toxicant [28
]. In addition, there is some evidence that ochratoxin might be a cause of TC [29
]. Surprisingly, there were no previous studies on the cocoa consumption in the development of MRD in humans, whereas toxic effects on male reproduction in animals are established.
We chose specifically hypospadias and TC trying to avoid the risk of possible underestimation reported for other congenital defects of the male reproductive tract, such as chryptorchidism, and because the available data on these conditions are rather accurate. The ecological approach used in this analysis has well known limitations. For example, it is not to be excluded that cocoa consumption may be simply a surrogate marker of affluence or other environmental factor. The data on the prevalence of hypospadias could be incomplete or non uniform, while the food consumption data may not apply to the cases under study. In fact, ecological studies correlating prevalence and/or incidence rates of a disease with the dietary practices in various geographical areas can only generate hypotheses for further studies.
Food consumption data provided by the FAO present other types of limitations, including differences in classification and definition of food items, completeness, and spatial and temporal variability and could not be adjusted for age, gender and other possible confounding factors, such as alcohol intake.
In addition, FAO consumption data may be particularly incomplete in developing countries such as India and China. However, we separated the analysis excluding these two countries and found the same results.
These findings, therefore, suggest that the associations found may not be spurious, and might deserve further studies of hypospadias and TC at the individual level to investigate the role of dietary intake of cocoa.