L-tryptophan (TRP) is reported to be a limiting AA in growing lambs [
16] and cattle [
11] during the process of non-protein N utilization. Thus, supplementation of TRP in a rumen-protected form has positive effects on growth performance. L-tryptophan metabolites can affect growth, development, and health of beef cattle [
11]. It is also known as a precursor of a stress-relieving neurotransmitter called serotonin [
11,
20]. Therefore, in the present study, we can postulate that animals in the RPT group were more tolerant of the cold stress situation, and thus showing higher ADG and lower FCR.
We observed higher FI in the RPT group compared to the control group (
Table 4). Ma et al. [
13] supplied two dosages of rumen-protected TRP to cashmere goats and observed an increase in the final BW and ADG in the supplemented group. Higher total FI intake could also postulate the fact that the steers in the RPT group tended to consume more feed in order to receive higher amounts of RPT, enabling them to better cope with cold stress conditions during the experiment period. On the contrary, higher amounts of RPT (over 0.5% of TMR, dry matter (DM)) have been documented to cause decreased FI due to lower palatability [
5,
10,
21]. However, no changes in FI was observed by supplementing 0.1% RPT in Korean native steers in a previous study in our laboratory [
11] under normal environmental conditions. In contrast, in the present study, higher FI in the RPT group (
Table 4) implies that the amount of RPT up to 0.1% did not decrease the palatability of the feed. As stated before, tryptophan is known as a precursor of a stress-relieving neurotransmitter called serotonin [
11,
20]. Given this phenomenon, tryptophan supplementation may indirectly and positively help the animals to deal with cold stress in this experiment and thus indirectly induce FI in the tryptophan supplemented group.
In this study, higher blood glucose of cattle in the RPT group, compared with the control group, was shown on day 27 of the experiment period, and the glucose level returned to the normal values on day 48 of the experiment (
Table 5). Abeni et al. [
22] stated that decreasing glucose content in blood in heat-stressed lactating Friesian cows could be the result of reduced energy intake. They also suggested that heat stress negatively affects gluconeogenesis as an endocrine acclimation to the stress conditions [
22]. L-tryptophan has been suggested not only to upregulate gluconeogenesis in the liver but its metabolites, including serotonin, modulate glucose uptake into muscle and thus suppress the rise in glucose. However, the reason why blood glucose was higher in the RPT group, to the authors, is unknown. In this study, we postulated that higher glucose levels in blood provides sufficient energy for thermoregulatory purposes in order to better dealing with cold stress.
In a previous study in our laboratory [
11], we observed that relative mRNA expression levels, including MYF6, MyoG, FABP4, and LPL genes, were higher in the RPT supplemented group than those in the control group in Korean native steers. While the MYF6 and MyoG are representative of muscle differentiation [
23], FABP4 is documented to be involved in intracellular transport and fatty acid metabolism [
24,
25]. Thus, 0.1% RPT alone may not be effective in altering the expression of muscle metabolism genes, including MYF6, MyoD, and Desmin. However, with respect to the fat metabolism and its related genes, 0.1% RPT could decrease the expression of PPARγ, C/EBPα, and FABP4 genes. The putative role of C/EBPα on nitrogen, glucose, and lipid metabolism is well documented [
26,
27]. C/EBPα is one of the main transcriptional mediators of the early stage of adipogenesis [
28,
29]. A study on leptin-deficient mice suggested that hepatic C/EBPα positively regulates lipogenesis in the mice [
27]. They suggested that C/EBPα may regulate nuclear factor Y (YF-Y) or Sp1, whereas the NF-Y consensus sequence CCAAT is included in most genes involved in lipogenesis, including farnesyl diphosphate synthetase (FPP) and 7-dehydrocholesterol reductase (7DCR). Loss of expression of C/EBPα resulted in attenuated induction of typical lipogenic genes and FPP and 7DCR, in the leptin-deficient mice [
27]. On the other hand, acetyl-CoA and NADPH required for fatty acid synthesis and β-oxidation were dysregulated in the liver of Δ proline-histidine rich domain (Δ PHR), but not in adipose tissue, which resulted in increased hepatic triglyceride production [
26]. They found that distinct C/EBPα motifs regulate lipogenic and gluconeogenic gene expression in mice. It is thought that FABPs roles include fatty acid uptake, transport, and metabolism. The decrease in gene expression of PPARγ, C/EBPα, and FABP4 in the present study may be attributed to the muscle tissues that utilized free fatty acid for myocyte differentiation. This phenomenon could be caused by direct involvement of TRP as a blocking block or by indirect involvement of metabolic components related to TRP or both. Given the above review, it can be suggested that dietary supplementation of RPT may alter intracellular transportation of fatty acids by inhibiting the catabolism of fat in muscle. The intramuscular fat and intramuscular fatty acid concentration are important in meat quality improvement [
30,
31]. It has been stated that peroxisome proliferator-activated receptor γ (PPARγ) is the pivotally important gene in relation to lipid metabolism in muscle tissue [
32]. Recently, Yang et al. [
33] investigated the effects of diets with different energy levels on fat deposition and the fatty acid profile of the
longissimus dorsi muscle in yak. They concluded that the high energy diets promoted the deposition and partial fatty acid content of
longissimus dorsi muscle mainly by up-regulation of mRNA expression of ACACA, SCD, FASN, SREBP-1c, PPARγ, and FABP4. However, in the present study, since the energy and amino acid are adversely related, up-regulation of mRNA expression of PPARγ, C/EBPα, and FABP4 could be seen in the non-supplemented RPT group (control), which is in line with the aforementioned study. Different energy or protein levels, herein the supplementation of TRP, may alter intramuscular fat deposition into muscle by regulating PPARγ. PPARγ is in charge of some promotion, including adipocyte proteins or enzymes such as fatty acid binding protein (FABP4), fatty acid synthase (FASN), and lipoprotein lipase (LPL) [
34]. Since very little information is available regarding the effect of RPT on fatty acid gene expressions in
longissimus dorsi muscle, further investigations are necessary in order to confirm these results and to bring more insights to the available knowledge.