Maternal physiological adaptations are impaired by cancer development and can lead to impaired foetal growth and development. Here, we clearly show that in the presence of Walker-256 tumour growth, foetal development was jeopardised, especially with respect to muscle protein metabolism linked to increased proteolysis and inhibited protein synthesis. These effects were likely associated with the lack of an anti-oxidative response, which reduced foetal weight. More importantly, we observed that nutritional supplementation with leucine led to a marked modulatory effect that minimised foetal proteolysis, maintained the balance of similar levels of protein synthesis and degradation, and restored the anti-oxidant responses.
Previous studies found that the presence of a tumour affected foetal weight [4, 5]. This decrease in foetal weight was related to a reduction in placental weight, which may be related to changes in placental tissue, and decreased the foetal/placental weight ratio (Figure 1), leading to impaired foetal development. Foetal growth in rats is more intense during the third phase of pregnancy (19th - 21st day), which coincides with the greatest tumour weight [3, 5]. The high activity of tumour cells that promotes exponential growth occurs during the same period of foetal growth. Thus, we verified that, at the time points when the tumour weighed approximately 10 to 12% of the maternal body weight, the W foetuses also displayed reductions in body weight, total serum protein, albumin and glucose that was coupled with a decline in muscle protein synthesis and increased proteolysis. All of these findings are also the main metabolic changes that occur during tumourigenesis , as was found in the group W foetuses. Proteolysis is the main process during cancer-cachexia, and this can be assessed by a decrease in albumin content, followed by the catabolism of skeletal muscle cells, promoting atrophy and apoptosis [43, 44]. Although the total muscle protein decreased in both of the tumour-bearing groups (W and LW), a leucine-rich diet maintained protein synthesis and reduced protein degradation in the LW group. This observation suggests that the LW group had a less intense catabolism of lean body mass, as was verified in the W group, which was likely related to the degradation of foetal muscle protein that was enhanced by oxidative stress [38, 45]. According to our previous studies, lean body mass was reduced during tumour growth, and dietary supplementation with leucine minimised this loss [3, 5, 8, 24]. Intense protein catabolism, primarily in the skeletal muscle, may have provided substrates, predominantly amino acids and gluconeogenic substrates, to be used by neoplastic cells [25, 46, 47]. Consequently, it is thought that tumour development induces a deviation of nutrients from the placenta and foetus (which is the reason for the decrease in foetal weight and reduction in the foetus/placenta ratio that is associated with increased protein degradation and oxidative stress). In parallel, the foetuses from mothers that were subjected to graduated food restriction (Cp and Lp) showed reduced weight, but their serum parameters and total muscle protein were similar to those of the control group animals. Nutritional supplements are particularly important for foetal viability and foetus/placenta unity, but foetuses have the capacity to adapt metabolically to acute and chronic changes in maternal and foetal serum nutrients content [48, 49]. In contrast, we show that foetuses from tumour-bearing mothers, primarily those in group W, were not able to efficiently adapt their metabolism under cancer conditions compared to those under food restriction, as found in pair-fed foetuses (Cp). This fact may be related to the action of substances that are produced by cancer cells, as we have previously reported [4, 7, 50, 51]. Carbo and colleagues  found deficits in the amino acid mobilisation of foetuses in pregnant rats that received tumour necrosis factor during pregnancy, and this was associated with decreased placental blood flow due to the action of this cytokine. We previously verified that pregnant rats that were injected with ascitic liquid (obtained from tumour-bearing animals without tumour cells) exhibited an increase in foetal reabsorption and a reduction in foetal and placental weight; similar deleterious effects were found in tumour-bearing mothers [3, 4, 50]. In these experiments, we proposed that these changes were induced by substances produced by cancer cells and did not result from only the effect of reducing nutrients, as was observed in this study when comparing the W and Cp foetuses.
Pregnancy changes, especially the increase in muscle protein breakdown, are related to the effects of the tumour on the body. In the cancer host, the catabolism that is associated with the reduction and/or maintenance of the protein synthesis process is increased [24, 53] and is related to increased oxidative stress [38, 45, 50]. In this case, the foetuses from the rats in group W displayed reduced protein synthesis, which was also associated with an increase in proteolysis caused by the activation of proteolytic systems and a high oxidative stress process. The ubiquitin-proteasome pathway can be modulated by a leucine-rich diet [6, 8]. As has been reported in many other studies, the activity of this process was higher and more widely observed in foetal cells. The 20S subunit, which comprises the catalytic core of the ubiquitin-proteasome proteolytic pathway, is most likely involved in protein turnover, leading to differentiation and development of embryonic and foetal tissues. However, we showed that although the 20S subunit was expressed at lower levels in the W and LW foetuses, the greater chymotrypsin-like activity and oxidative stress that were found in group W were sufficient to promote protein wasting in those foetuses. More importantly, under leucine nutritional supplementation, the LW foetuses showed less increased protein degradation because leucine diminished the increase in chymotrypsin activity and maintained the cathepsin enzyme while increasing the antioxidant response; these results suggest a beneficial effect of leucine in minimising proteolysis. Moreover, tumour growth produces alterations that affect placental metabolism and interact with the placenta-foetus, as evidenced by intrauterine growth restriction with increased foetal-muscle catabolism [3, 4, 6, 24, 52].
Nutritional supplementation with leucine provides several benefits by activating cellular synthesis processes [6, 24, 25]. These data suggest that the foetuses that received nutritional leucine supplementation, even those that were suffering from their mother’s tumours, showed diminished deleterious effects due to tumour growth compared to those that did not receive leucine; the foetuses in the LW group had similar muscle protein synthesis to the control foetuses, and more importantly, the process of protein catabolism was minimised and was equal to that of the control group [4, 5, 54]. Further studies are underway in our laboratory to determine whether leucine supplementation can counteract the damage to foetal growth during cancer development and the mechanism underlying this effect. These data indicate the importance of co-adjuvant therapy in improving the state of host responses to clinical treatment and particularly the importance of leucine supplementation, which has a positive effect on the host mother and on foetal development.