INTRODUCTION
Vitamin D is an important micronutrient which is necessary for the growth and maintenance of functional skeleton and helps to sustain health and improve longevity. 25(OH)D
3 is a form of vitamin D
3 that has been used in animal diets as a new source of vitamin D [
1,
2] because it is an intermediary metabolite that bypasses the liver metabolism and is readily available to animals. Moreover, it is five times more potent in raising vitamin D status of humans than an equivalent amount of vitamin D
3 [
3]. Increases in piglet body weight (BW) at birth and weaning were observed when dams were supplemented 2,000 IU/kg 25(OH)D
3 compared to piglets from dams supplemented 2,000 IU/kg vitamin D
3 [
4]. Thayer et al [
5] demonstrated that the feed efficiency of progeny was improved in the nursery from day 0 to 59 when the dams and nursery pigs were fed diets containing 1,500 IU/kg vitamin D
3+50 μg/kg 25(OH)D
3 compared with pigs from dams fed diets containing 500 IU/kg vitamin D
3+ 25 μg/kg 25(OH)D
3 but no differences in the growth performance of pigs in the finisher phases were observed. A previous study by Flohr et al [
6] noted that growing pigs from sows fed 50 μg/kg 25(OH)D
3 achieved higher average daily gain (ADG) than those pigs from sows fed 800 IU/kg vitamin D
3. Duffy et al [
7] demonstrated that pigs offered 25(OH)D
3 diets exhibited highest serum 25(OH)D
3 concentration and subsequently exhibited the highest
Longissimus thoracis total vitamin D activity. However, other studies reported that finishing growth was unaffected by the supplementation of vitamin D
3 alone or in combination with 25(OH)D
3 [
8–
10].
The benefits of vitamin D go beyond the function of the regulation of calcium and phosphorus homeostasis as it influences bone development, growth performance, immune status and production. Vitamin D metabolites control the expression of more than 200 genes through activation of the vitamin D receptor, which regulates or modulates gene expression within the target cell [
11]. This gives vitamin D a role in many functions in swine, including immunity, muscle function, and reproduction. The development of fetal muscle has far-reaching consequences for overall growth performance and health. In agricultural research, the importance of myostatin (
MSTN) and myogenic regulatory factors expression in early stages of development is well understood to impact meat quality and ultimate meat yield. Zhou et al [
10] demonstrated that supplementation with 25(OH)D
3 to dam’s diet can promote prenatal and postnatal skeletal muscle development of pig offspring by modulating the expressions of muscle transcription factors. A population-based mother-offspring cohort study in humans suggested that maternal vitamin D status during late pregnancy might influence muscle strength of offspring at 4 years of age [
12]. Thus, vitamin D supplementation has been demonstrated to exert a range of effects on the development of skeletal muscle of humans and animals [
10,
13,
14]. An alteration in fetal muscle characteristics was observed in fetuses from gilts fed the 25(OH)D
3 compared to fetuses from gilts fed vitamin D
3 [
15] when fed at concentrations above the basal requirement estimate.
Studies on how the performance and production characteristics of finishing pigs might be affected by supplementation of the dam’s diet with 25(OH)D3 followed by supplementation to their progeny diets throughout the wean to finish period are very limited but based on some of the human literature, it could have important scientific and commercial implications. Therefore, the objectives of the experiment herein were to: i) evaluate the performance and production characteristics of pigs fed dietary supplementation of 50 μg/kg 25(OH)D3 (equivalent to 2,000 IU/kg vitamin D3) during wean-to-finish period, ii) evaluate the influence of maternal 25(OH)D3 supplementation on the performance and production characteristics, blood metabolites, carcass characteristics and myostatin/myogenic regulatory factor gene expression of finishing pigs. We hypothesized that supplementation of the sow diets with 50 μg/kg 25(OH)D3 would improve reproduction and pre-weaning performance and that the effects would extend to post-weaning performance, muscle gene expression and meat quality. We also hypothesized that these maternal effects would be enhanced by 25(OH)D3 supplementation of the diets offered to the progeny after weaning.
DISCUSSION
The massive change in growth potential and management of commercial pigs has initiated a new concept of optimum vitamin nutrition so as to supply appropriate levels of vitamins during specific physiological phases of animals leading to positive results that go beyond the initial objective of preventing deficiency. To meet the body needs during different phases of growth, the vitamin D
3 dose recommended by DSM Nutritional Products Limited [
25] (above National Research Council [
16] recommendation levels) was fortified in the premix of the basal diets in the present study and considered as the control diet.
There is also increased interest in understanding how maternal nutrient supplement can impact progeny growth and health. A study by Hines et al [
15] concluded that the increase in fetal muscle development was due to the increase in maternal 25(OH)D
3. The main purpose of the current study was to evaluate the supplementation of 50 μg/kg 25(OH)D
3 to the basal diets of sows and their progeny on the growth performance, blood profile, muscle gene expression and production of wean to finish pigs.
In the current study, 25(OH)D
3 supplementation of the diets fed during gestation and lactation improved pre-weaning growth rate and growth and feed efficiency in the period to day 42 post weaning. Weber et al [
4] noted that the weaning weight of pigs born to sows fed 50 μg/kg 25(OH)D
3 was higher than those fed 2,000 IU/kg vitamin D
3, indicating the source of vitamin D was influential in improving the growth of pigs. In addition, Zhou et al [
26] reported that feeding a combined supplement of the two vitamin D
3 sources (50 μg/kg each 25(OH)D
3 and vitamin D
3) during gestation and lactation increased piglet growth during the first 2 weeks of lactation. In the study reported herein, the sow diet 25(OH)D
3 supplementation had no effect on growth performance after 42 days post weaning but supplementation of 50 μg/kg 25(OH)D
3 to the basal diets offered post weaning led to heavier BW at days 42, 98, and 140 and a higher ADG from weaning to day 42, between days 98 and 140 and over the entire wean to finish period. Tousignant et al [
27], reported that the oral administration of vitamin D
3 to suckling pigs resulted in higher BW at weaning and 7 days post weaning but weights did not differ at 26 days post weaning compared with control. However, several other reports indicated that nursery and finishing growth was unaffected by the supplementation of vitamin D
3 alone or in combination with 25(OH)D
3 [
8–
10]. The difference in outcomes between our study and these others may be due to the doses of vitamin D tested. For example, Thayer et al [
5], investigated total vitamin D (D
3 alone or combined with 25(OH)D
3) levels per kg diet ranging from 1,500 to 3,500 IU, 1,000 to 2,000 IU, and 800 to 1,600 IU for sow, nursery pigs, and growing and finishing pigs respectively and reported no effects of treatments on reproduction or pig growth performance. In the current study the total vitamin D/kg diet (basal D
3 alone or basal D
3 plus 2,000 IU 25(OH)D
3) ranged from 2,000 to 4,000 IU, 2,500 to 4,500 IU, and 1,750 to 3,750 IU for sows, nursery and growing and finishing pigs, respectively. It’s probable that that contemporary pigs may need higher dietary vitamin D levels to maximize their lifetime performance than previously thought [
28]. The interplay between vitamin D and the growth hormone (GH)/insulin-like growth factor (IGF)-1 system is not fully understood. Insulin-like growth factor-1 is produced by the liver in response to GH stimulation. Both GH and IGFs form part of the somatropic axis, which promotes whole body growth and development via action on key metabolic organs including the liver, skeletal muscle and bone. Growth hormone directly regulates renal 1α-hydroxylase (catalyzed the conversion of 25(OH)D
3 to 1α,25 (OH)
2D
3 and therefore modulates vitamin D metabolism mediated by IGF-1 [
29]. It has been suggested that the supplementation of vitamin D to the diet of humans with vitamin D deficiency served as a link between the proliferating cartilage cells of the growth plate and GH/IGF-1 secretion and the increase in IGF-1 and 25(OH)D
3 levels. The improvement in growth of pigs receiving 25(OH)D
3 either from the sows or through their nursery-finish diets might partly be due to the activation of the GH/IGF-1 axis.
Supplementation of D
3 or 25(OH)D
3 results in increased concentrations of Ca
2+ in the blood and muscle tissue [
30,
31]. Higher level of calcium in muscles activates calpain proteinase system (CPS), which consists of μ-calpain and m-calpain activated by Ca
2+ ions and endogenous calpain inhibitor-calpastatin. In addition to playing a role in post-mortem meat tenderness, CPS also regulates skeletal muscle growth [
32]. In our study, the activity of CPS was not measured, but it was highly likely that CPS was activated when feeding D
3 plus 25(OH)D
3 from wean-finish and was involved in regulating muscle growth, resulting in increased growth performance as evidenced in the current study.
Several researchers have suggested the role of vitamin D
3 and its active metabolite 1α, 25(OH)
2D
3 in modulating immune response [
33,
34]. In a human study, it was suggested that there is a direct link between 25(OH)D
3 and IgG [
35]. Vitamin D also has an effect on the inflammatory profile of monocytes by down-regulating the expression and production of several pro-inflammatory cytokines including TNF-α, IL-1β, IL-6, and IL-8 [
36,
37]. In the present study, only IL-6 and IL-1 were affected by 25(OH)D
3 supplementation of the diets offered post weaning. The former was reduced at 42 days and the latter increased at 140 days suggesting positive effects of 25(OH)D
3 on some immune related markers. The longer-term consequences on animal health remain to be established. Several studies reported that supplementation of 25(OH)D
3 resulted in increased serum 25(OH)D
3 response in sows, neonatal pigs, nursery pig, grower, and finisher pigs [
4,
5,
7,
38–
40]. In the present study, an increase in serum 25(OH)D
3 and Ca concentration was detected in growing pigs that received diets supplemented with 50 μg/kg 25(OH)D
3 post weaning. Conversely no significant sow diet effects were observed for these parameters in pigs. The lack of sow diet effect on serum 25(OH)D
3 concentration of their progeny during the nursery and grower-finisher period is unclear but this may have differed if it was measured at weaning.
Backfat thickness is one of the traits that have an important influence on the profitability of swine industry. It is used as a tool to figure out the dietary requirements for the optimization of growth as well as to determine the price [
41]. In the present study, the backfat thickness of growing-finish pigs on days 70 and 140 tended to be higher for pigs raised from sows fed the 25(OH)D
3 supplemented diets. The increase in backfat thickness could be in part due to the higher growth rate of the 25(OH)D
3 group in the nursery phase. Otherwise the small increase in fat thickness is difficult to explain as the sow treatments had minor effects on pig performance after day 42 post weaning.
Vitamin D may play an influential role in enhancing pork quality [
7,
42]. In a recent study, Duffy et al [
7] demonstrated that pigs offered 25(OH)D
3 diets exhibited highest serum 25(OH)D
3 concentration and subsequently exhibited the highest
Longissimus thoracis total vitamin D activity while Wilborn et al [
43] demonstrated that supplementation of 2,000 IU/kg vitamin D
3 to finishing pigs did not show detectable effects on a* or b* values but resulted in lower L* values compared to the meat evaluated from pigs fed control diets. While Wiegand et al [
44] demonstrated that the supplementation of 500,000 IU of vitamin D
3 for 3 days lowered L* values, increased a* values and did not affect b* values at 7 and 14 days post-mortem compared to control animals. In the present study, the sensory evaluation of meat, meat color, cooking loss, and LM area were unaffected by supplementation of 50 μg/kg 25(OH)D
3 during the wean-to-finish period or through maternal transfer. The drip loss was reduced after 5 and 7 days of storage by supplementation of the sow and post weaning diets with 25(OH)D
3 with the latter having the greater effect. Other aspects of meat quality affected by treatment included WHC and muscle pH. The former was enhanced in the meat from pigs offered the supplemented diets after weaning and the latter reduced by the same diets but increased in the muscle of pigs born to sows fed the supplemented diets.
The study by Wilborn et al [
43] also reported a trend in reduction in drip loss after 8 days of storage in the meat obtained from pigs supplemented with high levels (40,000 or 80,000 IU) of vitamin D
3. The variation in findings with regards to meat quality among different studies may be due to the difference in the sources of vitamin D used, treatment duration and dose of vitamin D. Interestingly, supplementation of the sow diets and progeny diets with 25(OH)D
3 had opposite effects on muscle pH. The former increased and the latter reduced it. Though pork tenderness was not assessed in this study, calcium ions can improve tenderness when they are introduced into meat by injection [
45] and infusion [
46]. Vitamin D and 25(OH)D
3 naturally increase serum and muscle Ca levels [
30] and activates both μ-calpain and m-calpain and their inhibitor calpastatin, improving pork meat tenderness [
32]. Dietary supplementation of pigs with 25(OH)D
3 may be an effective strategy to increase pork meat tenderness without generating concerns of high vitamin D3 residues in meat. Unpublished field data by our group (T. K. Chung, personal communication, June 8, 2020) showed that feeding pigs with 25(OH)D
3 dosed at 50 μg per kg diet in the presence of dietary vitamin D
3 from wean to finish produced fresh pork with significantly lower Warner Bratzler Forces and cooked pork with significantly better tenderness scores assessed by trained pork meat panelists. Tenderness is a major driver of consumer perceptions of pork eating quality and the role of vitamin D in improving it warrants further research.
In the last few decades, a growing number of studies concerning the muscular effects of vitamin D supplementation and research on the vitamin D receptor in muscle cells have contributed to understanding the role and actions of vitamin D in muscle tissue and on physical performance. Vitamin D and its receptor are important for normal skeletal muscle development and in optimizing muscle strength and performance [
47]. A study by Olsson et al [
48] indicated the expression of vitamin D receptor at the cellular level and noted there is a direct effect of vitamin D on human skeletal muscle precursor cells. In another study, mice lacking vitamin D receptor showed the skeletal muscle phenotype having smaller and variable muscle fibers and immature muscle gene expression that persisted even during the adult age suggested the role of vitamin D in muscle development [
49,
50]. In a study by Garcia et al [
51], it was demonstrated that inclusion of 25(OH)D
3 to C
2C
l2 skeletal muscle cells induced an expression of several myogenic markers such as
MYOD, myogenin at different stages of differentiation, and reduced the expression of
MSTN, which is the negative regulator of muscle mass. In the present study, we also evaluated the expression of several genes that regulate muscle growth and differentiation. The inclusion of 50 μg/kg 25(OH)D
3 in the sow and progeny diets reduced the expression of
MSTN gene in finishing pig. It has been reported that the
MSTN gene is the negative regulator of muscle mass [
52,
53].
Previous studies suggested that there is a direct effect of 25(OH)D
3 on increasing the expression of follistatin during muscle cell differentiation which antagonizes
MSTN by a direct protein interaction, preventing the inhibitory effects of
MSTN [
51,
54,
55]. Increased expression of the pro-myogenic skeletal markers
MYOD and
MYF5 was observed for finishing pigs from sows fed 25(OH)D
3 supplemented diets in gestation and lactation. Expression of
MYF5 but not
MYOD was also enhanced in finishing pigs receiving 25(OH)D
3 supplemented diets after weaning. However, no effect on
MYOG gene expression was seen which agrees with the results of Braga et al [
56] to some extent who indicated the downregulation of
MSTN gene and upregulation of
MYOD,
MYOG in satellite cells from 8-week old C57/BL6 mice incubated with 1α,25(OH)
2D
3. The results show that, 25(OH)D
3 supplementation in sow diets exerted a long-term effect on muscle gene expression with the changes indicative of enhanced muscle development. The results need to be confirmed and the implications on animal performance and carcass traits remain to be elucidated.