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Asian-Australas J Anim Sci > Volume 27(1); 2014 > Article
Park, Lee, Hong, Cho, Kim, and Park: Effect of Dietary Supplementation of Procyanidin on Growth Performance and Immune Response in Pigs

Abstract

This study was performed to determine the effect of dietary supplementation of procyanidin on growth performance, blood characteristics, and immune function in growing pigs. In experiment 1 (Exp. 1), thirty-two crossbred pigs with an initial BW of 19.2±0.3 kg were allocated into 4 treatments for an 8-wk experiment: i) CON (basal diet), ii) MOS 0.1 (basal diet+0.1% mannanoligosaccharide), iii) Pro-1 (basal diet+0.01% procyanidin), and iv) Pro-2 (basal diet+0.02% procyanidin). Pigs fed Pro-1 and Pro-2 diets had greater (p<0.05) gain:feed ratio compared with those fed CON or MOS 0.1 diets. Serum creatinine concentration was less (p<0.05) in Pro-2 treatment than those in CON, MOS 0.1 and Pro-1 treatments. In Exp. 2, twelve pigs (BW 13.4±1.3 kg) received basal diet with i) 0 (CON), ii) 0.02% (Pro-0.02%), and iii) 0.04% procyanidin (Pro-0.04%) for 4 wk. Concentration of platelets was lower (p<0.05) in the Pro-0.04% group compared to CON at 24 h after lipopolysaccharide (LPS) challenge. In addition, secretion of cytokines from cultured peripheral blood mononuclear cells (PBMC) in the presence or absence of procyanidin was examined. The levels of interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α were lower (p<0.05) in Pro (LPS-stimulated PBMCs+procyanidin) than those in CON (LPS-stimulated PBMCs+PBS) at 4 h after LPS challenge. These data suggest that dietary addition of procyanidin improves feed efficiency and anti-inflammatory cytokines of pigs.

INTRODUCTION

During the past decades, antibiotics have been used as feed additives to improve growth of animals and protect them from pathogenic microorganisms (Meng et al., 2010). However, in recent years, public concerns over the use of antibiotics in livestock have increased due to the emergence of antibiotic-resistant bacteria that may be a hazard to human health (Huang et al., 2010). Therefore, European Union (EU) started to ban the use of antibiotic as feed additives in 1999 and now are considering restricting its use outside EU (Windisch et al., 2008). In Korea, it has been required to develop antibiotic-free diets for domestic animals including pigs (Bae et al., 1999). Accordingly, considerable effort has been dedicated to identifying alternatives to antibiotics as growth promoters in the animal industry (Hong et al., 2004; Cho et al., 2006).
Procyanidins have been reported to exert antioxidant properties (Balu et al., 2005; Mohanasundari et al., 2005; Sangeetha et al., 2005; Yahara et al., 2005). They are present in the grape seed extracts and are composed of flavan units linked by C4-C6 or C4-C8 bonds forming dimmers, trimers, tetramers, and polymers up to 15 to 16 units (Prieur et al., 1994) with small amounts of catechin and epicatechin (Ricardo da Silva et al., 1990). Flavan compounds possessed many biological effects including scavenging free radicals, chelating transition metals, and modulating activities of antioxidant enzymes. Therefore, they have anti-aging effect, alleviate bone debilities, and protect renal tissues against toxicity. Procyanidins are also known to have therapeutic effects with anti-mutagenic (Faria et al., 2006), anti-cytotoxic (Cos et al., 2004), and anti-inflammatory (Selmi et al., 2006) activities. Therefore, this study was performed to determine the effect of oral administration of procyanidin on growth performance and immune response in growing pigs.

MATERIALS AND METHODS

Preparation of procyanidin

Procyanidin supplied by SINE-BIO (Seongnam, Korea) was collected and sterilized as described by Hwang et al. (2011). Grape seeds were powdered and soaked in a mixture of acetone/water/acetic acid (v/v/v 70:29.5:0.5). The mixture was filtered and dried under reduced pressure using a rotary evaporator at 40°C. It was further dried in a vacuum-freeze dryer and 70% acetone extracts were obtained. Extracts were then separated by n-hexane, and separated by an open column chromatography packed with silica gel. The stationary phase was sephadex LH-20 and the mobile phase consisted of a mixture of methanol and water (v/v 20:80). The flow rate was 1.2 mL/min and the sample was analyzed at 283 nm with a spectrometer. The fraction with a retention time of 18.7 min (89% purity) showed the highest activity and 18 mg of the procyanidin extract was collected from this fraction. Finally, it was dried using a freeze drier. Concentration of extracted procyanidin used in this experiment was mixed with feed at 0.265%.

Animal managements and experimental diets

The experimental protocols describing the management and care of animals were reviewed and approved by the Animal Care and Use at the National Institute of Animal Science (NIAS). All diets (Table 1) were formulated to meet the nutrient requirements recommended by the official Korean Feeding Standard for Swine (NIAS, 2007). Each pen was provided with a stainless steel feeder and one nipple drinker that allowed for ad libitum access to feed and water throughout the experiment.

Experiment 1: feeding trial

Experimental design and animals

A total of 32 barrows (Landrace×Yorkshire) with an average initial BW of 19.2±0.3 kg were selected for this 8 wk growth trial. Pigs were allocated to 1 of 4 dietary treatments according to their BW in a randomized complete block design with 8 pigs per treatment group. Dietary treatments included i) CON (basal diet), ii) MOS 0.1 (basal diet+0.1% mannanoligosaccharide), iii) Pro-1 (basal diet+0.01% procyanidin), and iv) Pro-2 (basal diet+0.02% procyanidin).

Growth performance measurements, blood sampling and analysis

Individual pig BW and feed consumption were recorded on d 0 and at the end of the experimental period on d 56 and ADG, ADFI, and gain:feed (G:F) were calculated. Blood samples (10 mL) were collected via anterior vena cava puncture after the pigs had been starved for 12 h at the beginning, 3 wk, and 8 wk of experimental period. Whole blood sample was centrifuged at 2,000×g for 10 min at 4°C, and the serum was collected. Serum biochemical profile was determined using a chemistry analyzer (COBAS MIRA plus, ROCHE diagnostics. Block Scientific, Inc. Bohemia, NY, USA).

Superoxide dismutase (SOD) activity

In our study, SOD was measured to investigate the effect of in vivo antioxidant activity of hesperidin. The SOD was determined by the method of Marklund and Marklund (1974). In a cap tube containing 100 μL of serum sample (9 wk serum), 1.5 mL of Tris EDTA HCl buffer (50 mM Tris, 1 mM EDTA, 6 N HCl) with 50 μL of pyrogallol (5 mM) was incubated at 25°C for 30 min. Thereafter, 50 μL of 1 N HCl was added and vortexed for several seconds. The absorbance of the solution was measured at 420 nm. One unit was the amount of enzyme required to cause 50% inhibition of pyrogallol oxidation, and SOD activity was expressed as unit/mg protein.

Experiment 2: Lipopolysaccharide (LPS) challenge trial

Experimental design and animals

A total of 12 barrows (Landrace×Yorkshire) with an average initial BW of 13.4±1.3 kg maintained in individual pens were assigned to 3 dietary treatments in a randomized complete block design based on BW for a 4-wk period. Three treatments included: i) CON (basal diet), ii) Pro-0.02% (basal diet+0.02% procyanidin), and iii) Pro-0.04% (basal diet+0.04% procyanidin). The level of procyanidin was increased to 0.04% since pigs needed to have more procyanidin under LPS challenge to give rise to the effect of procyanidin on pro-inflammatory cytokines. At the end of 4 wk, all pigs in each dietary treatment were intraperitoneally injected with Escherichia coli (serotype 0111:B4) LPS (Sigma Chemical Co., St, Louis, MO, USA) at a level of 50 μg/kg of BW. The LPS dosage was referenced on the results of previous studies (Matteri et al., 1998; Wright et al., 2000; Kim et al., 2010).

Blood sampling and measurements

Blood samples (15 mL) were collected via anterior vena cava puncture at 0 and 24 h after LPS injection after the pigs had been starved for 12 h. Blood samples were collected into both non-heparinized tubes (10 mL) and vacuum tubes (5 mL) containing K2 EDTA (Becton, Dickinson and Co., Franklin Lakes, NJ, USA) to obtain serum and whole blood, respectively. Blood samples were centrifuged (2,000×g) for 10 min at 4°C. The white blood cells, red blood cells and platelet concentration in the whole blood were determined using an automatic blood analyzer (Hemavet 950, CDC. Drew Scientific Inc. Dallas, TX, USA). Serum immunoglobulin A (IgA), G (IgG) and M (IgM) concentrations were evaluated using commercially available ELISA Starter Accessory kits (Bethyl Laboratories, Inc. Montgomery, TX, USA). The absorbance was read at 450 nm using a microplate reader (Versamax, Molecular Devices, Inc. Sunnyvale, CA, USA).

Peripheral blood mononuclear cells (PBMCs) culture and cytokine secretion

Peripheral blood samples were collected in heparin-containing tubes (Becton, Dickinson and Co., Franklin Lakes, NJ, USA) from 2 healthy piglets that were not used in the experiments. Blood samples were diluted with PBS and layered over a Ficoll-Hypaque (GE Healthcare Bio-Sciences AB. Rapsgatan 7, Uppsala, Sweden) gradient and centrifuged at 2,200×g for 17 min at 10°C. PBMCs were harvested from the interface layer, washed twice with PBS and then counted. Cells were resuspended in RPMI (Roswell Park Memorial Institute) 1640 GlutaMax (Invitrogen, GIBCO. Carlsbad, CA, USA). Concentration of PBMC was adjusted to 1.25×106 viable cells/mL after estimation of viability by trypan blue exclusion assay. For the cytokine secretion assay, 200 μL of a 1.25×106 cell suspension was cultured with an equal volume of flavonoid 25 μg/mL (procyanidin 25 μg/mL) or PBS. We set the concentration of flavonoids used in this study based on prior dose response analysis conducted by our laboratory (Sanbongi et al., 1997; Mao et al., 1999).
Isolated PBMCs were stimulated with 100 ng/mL LPS and then cultured in addition of PBS (CON) or procyanidin (Pro) for 2 h or 4 h at 38°C with 5% CO2. PBMC interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α concentrations were evaluated using commercially available ELISA kits (Quantikine, R&D systems, Minneapolis, MN, USA) and the absorbance was measured at 450 nm.

Statistical analysis

In this experiment, data were analyzed by using GLM procedure of SAS (SAS Institute, 2008). Differences among treatments were analyzed by Duncan’s multiple range test (Duncan, 1955). Probability values less than 0.05 were considered significant. Each pen was considered as an experimental unit. In Exp. 2, Cytokine secretion data were analyzed using t-test of SAS (SAS Institute, 2008). Each animal was considered as an experimental unit. Probability values less than 0.05 were considered significant.

RESULTS

Experiment 1: feeding trial

Growth performance

During the overall period, no difference was observed in BW, ADG, and ADFI among treatments (Table 2). However, dietary Pro-1 (0.425) and Pro-2 (0.458) treatments increased (p<0.05) G:F compared to CON (0.381) or MOS 0.1 (0.380) group.

Serum biochemical profile

The effects of procyanidin supplementation on serum profile in growing pigs are shown in Table 3. No effects were observed on cholesterol, total protein, albumin and blood urea nitrogen among treatments. However, serum creatinine concentration was significantly lower (p<0.05) in Pro-2 treatment than those in CON, MOS 0.1 and Pro-1 treatments. The glutamic oxalate transaminase concentration was greater (p<0.05) in MOS 0.1 treatment than that in CON treatment on 8 wk. Concentration of lactate dehydrogenase was lower (p<0.05) in CON treatment than all other groups at 3 wk.

Superoxide dismutase (SOD) activity

The serum SOD activity in the Pro-2 group was higher (p<0.05) than that in CON, MOS 0.1 and Pro-1 groups (Table 4).

Experiment 2: LPS challenge trial

Blood characteristics

White blood cell, red blood cell, hemoglobin, hematocrit, immunoglobulin A, G, and M were not influenced by dietary treatments (Table 5). Pigs administrated with Pro-0.04% had lower (p<0.05) platelet concentrations at 24 h after LPS injection compared to control group.

Cytokine secretion following LPS activation

Secretion of IL-1β and TNF-α were lower (p<0.05) in Pro (LPS-stimulated PBMCs+procyanidin) treatment than those in CON (LPS-stimulated PBMCs+PBS) treatment at 2 h and 4 h after LPS challenge, and IL-6 was lower (p<0.05) in Pro treatment than in CON treatment at 4 h after LPS challenge (Table 6).

DISCUSSION

Experiment 1: feeding trial

Growth performance

Endotoxin activity of procyanidin was tested by using Lumulus amebocyte lysate (LAL) and no detectable endotoxin was found (Holderness et al., 2007). Procyanidin’s endotoxin free property was again tested in various in vivo and in vitro studies including many animals and cell types (Bentivegna, 2002; Yamakoshi et al., 2002; Daughenbaugh et al., 2011; Lluis et al., 2011). Concomitant with these results, pigs treated with very low level of procyanidin, which was compared to those used in toxicity studies, had no sign of harmful effects on health or pro-inflammatory cytokines.
Discrepancies have been found in the effect of procyanidin on feed intake or growth performance that are fully or partly related to the differences in level of procyanidin in the experimental diets or in animal species used. Supplementation of antioxidant improves growth performance and feed efficiency (Lohakare et al., 2005). Previous studies reported that procyanidin helps reduce stress-related problems and improve growth rate in humans (Wooden et al., 1984; Slayback and Ronald, 2006). Procyanidin also showed these beneficial effects in rabbits (Garcia et al., 2002) and broiler chicks (Brenes et al., 2010). Concomitant with these data, results from the present study showed that administration of procyanidin increased the G:F ratio (Table 2) as well as the antioxidant activity in growing pigs (Table 4). Its beneficial effects on inflammation could reduce energy needs for immune responses thereby maintain animal health and improve growth performance. Therefore, it is possible that procyanidin be used as a feed additive for improving animal health and productivity.

Serum biochemical profile

Blood urea nitrogen (BUN) and serum creatinine (Scr) are well-known indicators of the renal function and health (Chen et al., 2003). However, Scr is a more specific indicator than BUN, since level of BUN can be affected by protein intake and liver function (Mathieson, 2003). Shi et al. (2003) reported that the antioxidant effect of procyanidin is approximately 50 times greater than that of vitamin C or vitamin E. Procyanidin alleviates the gentamicin induced-kidney injury (Jeong et al., 2005; Han et al., 2008; Hwang et al., 2008; Safa et al., 2010). Animal experiments have also demonstrated that procyanidin decreases the Scr concentrations in mice and rats (Kalantari et al., 2007; Yanarates et al., 2008). Concentration of Scr was also decreased by procyanidin treatment in our study (Table 3), indicating that procyanidin may have beneficial effects on renal health. However, it will be important to identify the relationship between procyanidin and serum creatinine level and how it helps improve renal function.

Superoxide dismutase (SOD) activity

Superoxide is formed in the red blood cells by auto-oxidation of hemoglobin into methemoglobin (Nordberg and Arnér, 2001). Khan (1999) reported that SOD and glutathione peroxidase (GPX) have roles in an antioxidant defense system. These enzymes are involved in the clearance of superoxide and H2O2 to maintain the structure and function of biological membranes (McCord, 2000). Oguntibeju et al. (2012) reported that Wistar rats administered with procyanidin (0.87 mg/kg BW) showed increased SOD level compared to controls. Dietary supplementation of procyanidin also improved (p<0.05) the SOD-like activity in growing pigs from the present study (Table 4). This result is consistent with our hypothesis that supplementation of procyanidin may result in an increase in plasma flavanol level and an increase in the antioxidant capacity of plasma.

Experiment 2: LPS challenge trial

Blood characteristics

As a constituent of gram-negative bacteria cell walls, LPS induces the expression of many genes necessary for immune defense function (Tracey et al., 1994). Therefore, it is plausible to detect the inhibitory effect of procyanidin on the expression of inflammatory cytokines with a LPS challenge. Consequently, the present results confirm its ability to inhibit the production of multiple cytokines and thereby improve anti-inflammatory effect in pigs. Procyanidin has positive effects on the platelet concentration after LPS challenge (Zhang et al., 2006; Martinez-Micaelo et al., 2012). Zhang et al. (2009) reported that LPS promotes platelet activity by inducing the secretion of both α and dense granules, thus amplifying secretion-dependent platelet aggregation. Previous studies showed that platelets are involved in the pathogenesis of severe sepsis (Taylor et al., 1997; Pu et al., 2001; Zhao et al., 2002), and that LPS stimulates thrombosis, the formation of platelet microaggregates (Rumbaut et al., 2006).
Procyanidins are polymeric compounds derived from catechin and epicatechin (Pasinetti et al., 2010). They modulate platelet function, thus reduced the risk of clot formation (Karen et al., 2003). Pignatelli et al. (2000) suggested that procyanidin decreases platelet production induced by hydrogen peroxide. Procyanidin interferes oxidation process, decreases platelet activation and increases eicosanoid synthesis (Karen et al., 2003). These responses result in lowering the platelet activation, thereby decreases the concentration of platelets. Procyanidin also increases concentration of plasma prostacyclin. Prostacyclin decreases platelet aggregation in vivo and ex vivo by elevating platelet cyclic AMP concentration and inhibiting agonist-induced increase in glycoprotein IIb-IIa expression and phosphorylation. Administration of procyanidin inhibits platelet aggregation in rats (Chang and Hsu, 1989; Ruf et al., 1995), dogs (Demrow et al., 1995) and humans (Rein et al., 2000; Freedman et al., 2001). Concomitant with these results, pigs administrated with Pro-0.04% treatment also showed lower (p<0.05) platelet concentration compared to control group in this study (Table 5), suggesting that administration of procyanidin derived from catechin and epicatechin inhibits platelet aggregation in LPS-challenged pigs.

Cytokine Secretion Following LPS Activation

Inflammation and acute phase responses after LPS challenge have been well characterized in a pig model (Johnson, 1997; Wright et al., 2000). Several pro-inflammatory cytokines, such as TNF-α, IL-6, and IL-1, are encoded by target genes in the NFkB activation pathway (Yang et al., 2008). NFkB acts as a crucial transcriptional activator of pro-inflammatory cytokines and its expression is triggered by pro-inflammatory stimuli and genotoxic stress, including bacterial cell-wall components (Karin and Greten, 2005).
IL-1β, a pro-inflammatory cytokine, is a mediator of LPS toxicity in vivo and in vitro (Lee, 2007). Mao et al. (2005) reported that IL-1β level was increased by LPS, but pigs fed Astragalus membranaceus β-glucan (AMG) had a lower level of IL-1β than pigs from control group when they were challenged with LPS. TNF-α is produced at early onset of inflammation followed by waves of IL-1 and IL-6 (Tizard, 2000). However, TNF-α causes a detrimental effect on animal performance by compromising immunity and nutrient metabolism (Spurlock, 1997). These results may highlight the important role of these cytokines in regulation of immune and inflammatory responses. Procyanidin decreases the expression of IL-6 and TNF-α (Mackenzie et al., 2004; Erlejman et al., 2008; Terra et al., 2009). The potent anti-inflammatory property of procyanidin has been shown in rats and mice (Li et al., 2001; Sakaguchi et al., 2006). In this study, we assessed serum concentration of cytokines in association with dietary administration of procyanidin with experimentally imposed stimulation of the immune system by LPS. As expected, procyanidin decreased LPS-induced IL-1β, IL-6 and TNF-α production in PBMCs. Our results indicate that procyanidin may have the potential to suppress the inflammatory response and be effective for ameliorating the bacteria challenged anti-inflammatory response in pigs.

CONCLUSION

Taken together, dietary supplementation of procyanidin may help prevent over-stimulation of the immune system in growing pigs after an immunological challenge. In addition, procyanidin improves feed efficiency and decreases the serum creatinine concentrations in pigs. Further research, however, is necessary to identify the specific mechanisms by which procyanidin improves immune function and growth performance in growing pigs.

ACKNOWLEDGEMENTS

This work was carried out with the support of “cooperative research program for Agriculture Science & Technology Development (Project No. PJ009340)”, Rural Development Administration, Republic of Korea.

Table 1
Composition of diets used in Exp. 1 and Exp. 2 (as-fed basis)
Item %
Ingredients
 Corn 56.20
 Soybean meal 28.00
 Wheat bran 10.00
 Soy oil 3.00
 L-lysine-HCl (78%) 0.10
 Limestone 0.90
 Calcium phosphate, Dibasic 1.00
 Salt 0.30
 Vitamin-mineral mixture1 0.50
 Total 100.00
Chemical composition
 ME (Mcal/kg) 3.40
 CP (%) 19.63
 Lysine (%) 1.26
 Crude fat (%) 5.03
 Crude fiber (%) 3.68
 Crude ash (%) 4.56
 Ca (%) 0.79
 P (%) 0.58

1 The vitamin-mineral premix provided the following quantities of vitamins and minerals per kilogram of diets: vitamin A, 10,000 IU; vitamin D3, 2,000 IU; vitamin E, 250 IU; vitamin K3, 0.5 mg; vitamin B1, 0.49 mg as mononitrate; thiamin, 0.49 mg as thiamin mononitrate; riboflavin, 1.50 mg; pyridoxine, 1 mg as pyridoxine hydrochloride; vitamin B12, 0.01 mg; niacin, 10 mg as nicotinic acid; pantothenic acid, 5 mg as calcium pantothenate; folic acid, 1 mg; biotin as d-biotin, 0.1 mg; choline, 125 mg as choline chloride; Mn, 60 mg as manganese sulfate; Zn, 75 mg as zinc sulfate; Fe, 20 mg as ferrous sulfate; Cu, 3 mg as cupric sulfate; I, 1.25 mg as calcium iodate; Co, 0.5 mg as cobaltous carbonate; Mg, 10 mg as magnesium oxide.

Table 2
Effect of dietary supplementation of procyanidin on growth performance in growing pigs (Exp. 1)1
Item Treatment2 SEM

CON MOS 0.1 Pro-1 Pro-2
Initial BW (kg) 19.53 19.08
Final BW (kg) 68.9 72.18 72.4 70.58 1.96
ADG (kg) 0.78 0.84 0.85 0.81 0.02
ADFI (kg) 1.95 2.12 1.89 1.78 0.11
G:F 0.381b 0.380b 0.425a 0.458a 0.012

1 Each least square mean represents 8 observations.

2 CON = Basal diet, MOS 0.1 = Basal diet+mannanoligosaccharide 0.1%, Pro-1 = Basal diet+procyanidin 0.01%, Pro-2 = Basal diet+ procyanidin 0.02%.

ab Means within the same row differ (p<0.05).

Table 3
Effects of dietary supplementation of procyanidin on serum biochemical profile in growing pigs (Exp. 1)1
Treatments2 SEM

CON MOS 0.1 Pro-1 Pro-2
Creatinine (mg/dL)
 0 week 1.6 1.6 1.6 1.6 0.66
 3 week 2.2a 2.1a 2.1a 1.5b 0.13
 8 week 2.4a 2.0ab 2.0ab 1.9b 0.08
Glutamic oxalate transaminase (U/L)
 0 week 65.9 92.3 79.3 84.4 10.77
 3 week 42.6 41.1 48.9 56.1 8.51
 8 week 28.0b 62.5a 38.3ab 38.8ab 11.22
Lactate dehydrogenase (mg/dL)
 0 week 1,323.8 1,461.5 1,264.0 1,343.0 88.34
 3 week 624.3b 913.6a 1,085.6a 1,014.6a 79.40
 8 week 563.5b 685.4a 607.5ab 585.5ab 58.84
Cholesterol (mg/dL)
 0 week 76.0 76.1 71.4 74.6 4.61
 3 week 70.6 79.6 65.7 66.5 4.52
 8 week 78.6 89.1 73.8 73.4 7.17
Total protein (g/dL)
 0 week 5.2 5.2 5.4 5.2 0.16
 3 week 6.5 5.9 6.2 5.8 0.16
 8 week 6.1 6.4 6.4 6.3 0.12
Albumin (g/dL)
 0 week 3.0 3.0 2.8 3.0 1.47
 3 week 3.6 3.4 3.6 3.3 0.13
 8 week 3.7 3.9 3.8 3.8 0.08
Blood urea nitrogen (mg/dL)
 0 week 23.8 24.3 19.8 23.4 1.26
 3 week 18.9 20.6 19.7 20.9 1.80
 8 week 26.8 32.4 28.0 27.4 2.83

1 Each least squares mean represents 8 observations.

2 CON = Basal diet, MOS 0.1 = Basal diet+mannanoligosaccharide 0.1%, Pro-1 = Basal diet+procyanidin 0.01%, Pro-2 = Basal diet+procyanidin 0.02%.

ab Means in the same row with different superscripts differ (p<0.05).

Table 4
Superoxide dismutase (SOD) activity in serum of pigs (Exp. 1)1
Treatment2 SEM

CON MOS 0.1 Pro-1 Pro-2
SOD (unit/mg protein) 51.23 b 51.38 b 51.42 b 60.45a 2.68

1 Each least squares mean represents 8 observations.

2 CON = Basal diet, MOS 0.1 = Basal diet+mannanoligosaccharide 0.1%, Pro-1 = Basal diet+procyanidin 0.01%, Pro-2 = Basal diet+ procyanidin 0.02%.

ab Means in the same row with different superscripts differ (p<0.05).

Table 5
Concentrations of white blood cells, red blood cells, platelets and immunoglobulins before and after an immunological challenge with lipopolysaccharide (LPS) following a dietary treatment in growing pigs (Exp. 2)1
Item Treatment2 SEM

CON Pro-0.02% Pro-0.04%
White blood cell (103/μL)
 Before LPS 17.85 16.96 13.19 2.74
 After 24 h 16.57 16.27 16.42 1.94
Red blood cell (106/μL)
 Before LPS 6.62 5.78 6.37 0.30
 After 24 h 6.63 6.82 6.38 0.28
Hemoglobin (g/dL)
 Before LPS 9.77 8.17 9.13 0.47
 After 24 h 9.03 9.22 8.63 0.58
Hematocrit (%)
 Before LPS 29.8 25.54 28.00 1.50
 After 24 h 30.50 32.33 28.05 1.99
Platelet (103/μL)
 Before LPS 406.3 412.2 295.8 76.1
 After 24 h 487.0a 385.6ab 359.0b 64.3
Immunogloblin A (mg/mL)
 Before LPS 1.31 1.18 1.12 0.11
 After 24 h 1.32 1.04 1.07 0.12
Immunogloblin G (mg/mL)
 Before LPS 7.4 7.58 6.21 0.93
 After 24 h 9.81 7.33 6.97 1.25
Immunogloblin M (mg/mL)
 Before LPS 1.28 1.66 1.49 0.18
 After 24 h 1.35 1.64 1.51 0.16

1 Each least squares mean represents 4 observations.

2 CON = Basal diet, Pro-0.02% =Basal diet+prcyanidin 0.02%, Pro-0.04% = Basal diet+prcyanidin 0.04%.

ab Means in the same row with different superscripts differ (p<0.05).

Table 6
Effects of procyanidin on secretion of pro-inflammatory cytokine by cultured peripheral blood mononuclear cells from piglets following an immunological challenge with lipopolysaccharide in vitro (Exp. 2)1
Items (pg/mL) Treatment2 SEM

CON Pro
IL-1β
 2 h 0.64b 0.20a 0.05
 4 h 1.07b 0.20a 0.05
IL-6
 2 h 60.45 61.32 10.25
 4 h 115.32b 55.25a 1.05
TNF-α
 2 h 29.83b 14.57a 0.65
 4 h 103.34b 14.59a 1.53

1 Each least squares mean represents 3 observations.

2 CON = LPS-stimulated PBMCs+PBS (Phosphate Buffer Saline), Pro = LPS-stimulated PBMCs+Procyanidin.

ab Means in the same row with different superscripts differ (p<0.01).

REFERENCES

Bae KH, Ko TG, Kim JH, Cho WT, Han YK, Han IK. 1999. Use of metabolically active substances to substitute for antibiotics in finishing pigs. Korean J Anim Sci 41:23–30.

Balu M, Sageetja P, Haripriya D, Panneerselvam C. 2005. Rejuvenation of antioxidant system in central nervous system of aged rats by grape seed extract. Neurosci Lett 383:295–300.
crossref pmid
Bentivegna SS, Whitney KM. 2002. Subchronic 3-month oral toxicity study of grape seed and grape skin extracts. Food Chem Toxicol 40:1731–1743.
crossref pmid
Brenes A, Viveros A, Goni I, Centeno C, Saura-Calixto F, Arija I. 2010. Effect of grape seed extract on growth performance, protein and polyphenol digestibilities, and antioxidant activity in chickens. Span J Agric Res 8:326–333.
crossref
Chang WC, Hsu FL. 1989. Inhibition of platelet aggregation and arachidonate metabolism in platelets by procyanidins. Prostaglandins Leukot. Essent Fatty Acids 38:181–1888.
crossref pmid
Chen ST, Peng SJ, Chen JR. 2003. Effects of dietary protein on renal function and lipid metabolism in five-sixths nephrectomized rats. Br J Nutr 89:491–497.
crossref pmid
Cho JH, Chen YJ, Min BJ, Kim HJ, Kwon OS, Shon KS, Kim IH, Kim SJ, Asamer A. 2006. Effects of essential oils supplementation on growth performance, IgG concentration and fecal noxious gas concentration of weaned pigs. Asian-Aust J Anim Sci 19:80–85.
crossref
Cos P, de Bruyne T, Hermans N, Apers S, Berghe DV, Vlietinck AJ. 2004. Proanthocyanidins in health care: Current and new trend. Curr Med Chem 11:1345–1359.
crossref pmid
Daughenbaugh KF, Holderness J, Graff JC, Hedges JF, Freedman B, Graff JW, Jutila MA. 2011. Contribution of transcript stability to a conserved procyanidin-induced cytokine response in γδT cells. Genes Immun 12:378–389.
crossref pmid pmc
Demrow HS, Slane PR, Folts JD. 1995. Administration of wine and grape juice inhibits in vivo platelet activity and thrombosis in stenosed canine coronary arteries. Circulation 91:1182–1188.
crossref pmid
Duncan DB. 1955. Multiple range and multiple F tests. Biometrics 11:1–42.
crossref
Erlejman AG, Jaggers G, Fraga CG, Oteiza PI. 2008. TNFα-induced NF-kB activation and cell oxidant production are modulated by hezameric procyanidins in Caco-2 cells. Arch Biochem Biophys 476:186–195.
crossref pmid
Faria A, Calhau C, de Freitas V, Mateus N. 2006. Procyanidins as antioxidants and tumor cell growth modulators. J Agric Food Chem 54:2392–2397.
crossref pmid
Freedman JE, Parker C, Li L, Perlman JA, Frei B, Ivanov V, Deak LR, Iafrati MD, Folts JD. 2001. Select flavonoids and whole juice from purple grapes inhibit platlet function and enhance nitric oxide release. Circulation 103:2792–2798.
crossref pmid
Garcia J, Nicodemus N, Carabano R, de Blass JC. 2002. Effect of inclusion of defated grape seed meal in the diet on digestion and performance of growing rabbits. J Anim Sci 80:162–170.
crossref pmid
Han JY, Sung JH, Kim DJ, Jeong HS, Lee JS. 2008. Inhibitory effect of methanol extract and its fractions from grape seeds on mushroom tyrosinase. J Korean Soc Food Sci Nutr 37:1679–1683.
crossref
Holderness J, Jackiw L, Kimmel E, Kerns H, Radke M, Hedges JF, Petrie C, McCurley P, Glee PM, Palecanda A, Jutila MA. 2007. Select plant tannins induce IL-2R up-regulation and augment cell division in γδT cells. J Immunol 179:6468–6478.
crossref pmid
Hong JW, Kim IH, Kwon OS, Min BJ, Lee WB, Shon KS. 2004. Influences of plant extract supplementation on performance and blood characteristics in weaned pigs. Asian-Aust J Anim Sci 17:374–378.
crossref
Huang Y, Yoo JS, Kim HJ, Wang Y, Chen YJ, Cho JH, Kim IH. 2010. Effects of dietary supplementation with blended essential oils on growth performance, nutrient digestibility, blood profiles and fecal characteristics in weanling pigs. Asian-Aust J Anim Sci 23:607–613.
crossref
Hwang DS, Shin SY, Lee YG, Hyun JY, Yong YJ, Park JC, Lee YH, Lim YH. 2011. A compound isolated from Schisandra chinensis induces apoptosis. Bioorg Med Chem Lett 21:6054–6057.
crossref pmid
Hwang IW, Lee HR, Kim SK, Zheng HZ, Choi JU, Lee SH, Lee SH, Chung SK. 2008. Proanthocyanidin content and antioxidant characteristics of grape seeds. Korean J Food Preserv 15:859–863.

Jeong SM, Kim SY, Ha JU, Lee SC. 2005. Effect of far-infrared irradiation on the antioxidant activity of extracts from grape seed. J Korean Soc Food Sci Nutr 34:1619–1624.
crossref
Johnson RW. 1997. Inhibition of growth by pro-inflammatory cytokines: An integrated view. J Anim Sci 75:1244–1255.
crossref pmid
Kalantari H, Rashidi I, Bazgir S, Dibaei A. 2007. Protective effects of hydroalcoholic extract of red grape seed (VITIS VENIFERA) in nephrotoxicity induced by amikacin in mice. Jund. J. Nat. Parm Products 2:87–93.

Karen JK, Andriana KC, Indu S, Maureen AF, Helen M, Marilyn JP, Alan HT, Neil JM, Andrew JS. 2003. Dietary flavanols and procyanidin oligomers from cocoa (Theobroma cacao) inhibit platelet function. Am J Clin Nutr 77:1466–1473.
crossref pmid
Karin M, Greten FR. 2005. NFkB: Linking inflammation and immunity to cancer development and progression. Nat Rev Immunol 5:749–759.
crossref pmid
Khan MA. 1999. Chemical composition and medicinal properties of Nigella sativa Linn. Inflammopharmacology 7:15–35.
crossref pmid
Kim BG, Lindemann MD, Cromwell GL. 2010. The effects of dietary chromium (III) picolinate on growth performance, vital signs, and blood measurements of pigs during immune stress. Biol Trace Elem Res 135:200–210.
crossref pmid
Lee E. 2007. Anti-inflammatory effect of Scutellariae Radix. Korean J Pant Res 20:548–552.

Li WG, Zhang XY, Wu YJ, Tian X. 2001. Anti-inflammatory effect and mechanism of proanthocyanidins from grape seeds. Acta Pharmacol Sin 22:1117–1120.
pmid
Lluis L, Munoz M, Nogues MR, Martos VS, Romeu M, Giralt M, Valls J, Sola R. 2011. Toxicology evaluation of a procyanidin-rich extract from grape skins and seeds. Food Chem Toxicol 49:1450–1454.
crossref pmid
Lohakare JD, Ryu MH, Hahn TW, Lee JK, Chae BJ. 2005. Effects of supplemental ascorbic acid on the performance and immunity of commercial broilers. J Appl Poult Res 14:10–19.
crossref
Mackenzie GG, Carrasquedo F, Delfino JM, Keen CL, Fraga CG, Oteiza PI. 2004. Epicatechin, catechin, and dimeric procyanidins inhibit PMA-induced NF-kB activation at multiple steps in Jurkat T cells. FASEB J 18:167–169.
crossref pmid
Mao TK, Powell JJ, van de Water J, Keen CL, Schmitz HH, Gershwin ME. 1999. The influence of cococa procyanidins on the transcription of interleukin-2 in peripheral blood mononuclear cells. Int J Immunother 15:23–29.

Mao XF, Piao XS, Lai CH, Li DF, Xing JJ, Shi BL. 2005. Effect of β-glucan obtained from the chinese herb Astragalus membranaceus and lipopolysaccharide challenge on performance, immunological, adrenal, and somatotropic responses of weanling pigs. J Anim Sci 83:2775–2782.
crossref pmid
Marklund S, Marklund G. 1974. Involvement of the superoxide anion radical in the autoxidation of pyrogallol and a convenient assay for superoxide dismutase. Eur J Biochem 47:469–474.
crossref pmid
Martinez-Micaelo N, González-Abuín N, Terra X, Richart C, Ardèvol A, Pinent M, Blay M. 2012. Omega-3 docosahexaenoic acid and procyanidins inhibit cyclo-oxygenase activity and attenuate NF-kB activation through a p105/p50 regulatory mechanism in macrophage inflammation. Biochem J 441:653–663.
crossref pmid
Mathieson PW. 2003. Immune dysregulation in minimal change nephropathy. Nephrol Dial Transplant 6:26–29.
crossref
Matteri RL, Klir JJ, Fink BN, Johnson RW. 1998. Neuroendocrine-immune interactions in the neonate. Domest Anim Endocrinol 15:397–407.
crossref pmid
McCord JM. 2000. The evolution of free radicals and oxidative stress. Am J Med 108:652–659.
crossref pmid
Meng QW, Yan L, Ao X, Jang HD, Cho JH, Kim IH. 2010. Effects of chito-oligosaccharide supplementation on egg production, nutrient digestibility, egg quality and blood profiles in laying hens. Asian-Aust J Anim Sci 23:1476–1481.
crossref pdf
Mohanasundari M, Sabesan M, Sethupathy S. 2005. Renoprotective effect of grape seeds extract in ethylene glycol induced nephrotoxic mice. Indian J Exp Biol 43:356–359.
pmid
National Institute of Animal Science (NIAS). 2007. Korean feeding standard for Swine. Korea:

Nordberg J, Arnér ES. 2001. Reactive oxygen species, antioxidants, and the mammalian thioredoxin system. Free Radic Biol Med 31:1287–1312.
crossref pmid
Oguntibeju OO, Fashola AN, Cole-Showers CL. 2012. Effects of procyanidin on antioxidant enzyme status in kidney and heart homogenates of Wistar rats. J Food Agric Environ 10:34–37.

Pasinetti GM, Ksiezak-Reding H, Santa-Maria I, Wang J, Ho L. 2010. Development of grape seed polyphenolic extract with anti-oligomeric activity as a novel treatment in progressive supranuclear palsy and other tauopathies. J Neurochem 114:1557–1568.
crossref pmid pmc
Pignatelli P, Pulcinelli FM, Celestini A, Lenti L, Ghiselli A, Gazzaniga PP, Violi F. 2000. The flavonoids quercetin and catechin synergistically inhibit platelet function by antagonizing the intracellular production of hydrogen peroxidase. Am J Clin Nutr 72:1150–1155.
crossref pmid
Prieur C, Rigaud J, Cheyner V, Moutounet M. 1994. Oligomeric and polymeric procyanidin from grape seed. Phytochemistry 36:781–784.
crossref
Pu Q, Wiel E, Corseaux D, Bordet R, Azrin MA, Ezekowitz MD, Lund N, Jude B, Vallet B. 2001. Beneficial effect of glycoprotein IIb/IIIa inhibitor (AZ-1) on endothelium in Escherichia coli endotoxin-induced shock. Crit Care Med 29:1181–1188.
crossref pmid
Rein D, Paglieroni TG, Pearson DA, Wun T, Schmitz HH, Gosselin R, Keen CL. 2000. Cocoa and wine polyphenols modulate platelet activation and function. J Nutr 130:2120S–2126S.
crossref
Ruf JC, Berger JL, Renaud S. 1995. Platelet rebound effect of alcohol withdrawal and wine drinking in rats. Relation to tannins and lipid peroxidation. Arterioscler Thromb Vasc Biol 15:140–144.
crossref pmid
Rumbaut RE, Bellera RV, Randhawa JK, Shrimpton CN, Dasgupta SK, Dong JF, Burns AR. 2006. Endotoxin enhaces microvascular thrombosis in mouse cremaster venules via a TKR4-dependent, neutrophil-independent mechanism. Am J Physiol Heart Circ Physiol 290:H1671–1679.
crossref pmid
Ricardo da Silva JM, Rosec JP, Bourzeix M, Heredia N. 1990. Separation and quantitative determination of grape and wine procyanidins by high performance reversed phase liquid chromatography. J Sci Food Agric 53:85–92.
crossref
Sanbongi C, Sizuki N, Sakane T. 1997. Polyphenols in chocolate, which have antioxidant activity, modulate immune function in humans in vitro. Cell Immunol 177:129–136.
crossref pmid
SAS Institute. 2008. SAS user’s guide. Statistical Analysis System Inst. Inc; Cary NC:

Safa J, Argani H, Bastani B, Nezami N, Rahimi Ardebili B, Ghorbanihaghjo A, Kalagheichi H, Amirfirouzi A, Mesgari M, Soleimany Rad J. 2010. Protective effect of grape seed extract on gentamicin-induced acute kidney injury. Iran J Kidney Dis 4:285–291.
pmid
Sakaguchi Y, Shirahase H, Kunishiro K, Ichikawa A, Kanda M, Uehara Y. 2006. Effect of combination of nitric oxide synthase and cyclooxygenase inhibitors on carrageenan-induced pleurisy in rats. Life Sci 79:442–447.
crossref pmid
Sangeetha P, Balu M, Haripriya D, Panneerselvam C. 2005. Age associated change in erythrocyte membrane surface charge: Modulatory role of grape seed proanthocyanidin. Exp Gerontol 40:820–828.
crossref pmid
Selmi C, Mao TK, Keen CL, Schmitz HH, Eric Gershwin M. 2006. The anti-inflammatory properties of cocoa flavanols. J Cardiovasc Pharmacol 47:S163–S171.
crossref pmid
Shi J, Yu J, Pohorly JE, Kakuda Y. 2003. Polyphenolics in grape seeds-biochemistry and functionality. J. Med Food 6:291–299.
crossref pmid
Slayback LD, Ronald RW. 2006. Bioflavonoids and cardiovascular health: tea, red wine, cocoa and pycnogenol®. J Am Nutrac Assoc 9:16–21.

Spurlock ME. 1997. Regulation of metabolism and growth during immune challenge: an overview of cytokine function. J Anim Sci 75:1773–1783.
crossref pmid
Taylor FB, Coller BS, Chang AC, Peer G, Jordan R, Engellener W, Esmon CT. 1997. 7E3 F(ab′)2, a monoclonal antibody to the platelet GPIIb/IIIa receptor, protects against microangiopathic hemolytic anemia and microvascular thrombotic renal failure in baboons treated with C4b binding protein and a sublethal infusion of Escherichia coli. Blood 89:4078–4084.
pmid
Terra X, Montagut G, Bustos M, Llopiz N, Ardèvol A, Bladé C, Fernández-Larrea J, Pujadas G, Salvadó J, Arola L, Blay M. 2009. Grape-seed procyanidins prevent low-grade inflammation by modulating cytokine expression in rats fed a high-fat diet. J Nutr Biochem 20:210–218.
crossref pmid
Tizard IR. 2000. Veterinary immunology: An introduction. W.B. Sauders Company; Philadelphia, PA:

Tracey KJ, Cerami A. 1994. Tumor Necrosis Factor: A pleiotropic cytokine and therapeutic target. Annu Rev Med 45:491–503.
crossref pmid
Windisch WM, Schedle K, Plitzner C, Kroismayr A. 2008. Use of phytogenic products as feed additives for swine and poultry. J Anim Sci 86:E140–148.
crossref pmid
Wooden GR, Crane CS, Beisel CG. 1984. An investigation of the effect of hesperidin complex and lemon bioflavonoid complex on growth and development of thoroughbred horses. J Anim Sci 59:1529–1535.
crossref pmid
Wright KJ, Balaji R, Hill CM, Dritz SS, Knoppel EL, Minton JE. 2000. Integrated adrenal, somatotropic, and immune responses of growing pigs to treatment with lipopolysaccharide. J Anim Sci 78:1892–1899.
crossref pmid
Yahara N, Tofani I, Maki K, Kojima K, Kijima Y, Kimura M. 2005. Mechanical assessment of effects of grape seed proanthocyanidins extract on tibial bone diaphysis in rats. J Musculoskelet Neuronal Interact 5:162–169.
crossref pmid
Yamakoshi J, Saito M, Kataoka S, Kikuchi M. 2002. Safety evaluation of proanthocyanidin-rich extract from grape seeds. Food Chem Toxicol 40:599–607.
crossref pmid
Yanarates O, Guven A, Sizlan A, Uysal B, Akgul O, Atim A, Ozcan A, Korkmaz A, Kurt E. 2008. Ameliorative effects of proanthocyanidin on renal ischemia/reperfusion injury. Ren Fail 30:931–938.
crossref pmid
Yang X, Guo Y, He X, Yuan J, Yang Y, Wang Z. 2008. Growth performance and immune responses in chickens after challenge with lipopolysaccharide and modulation by dietary different oils. Animal 2:216–223.
crossref pmid
Zhang G, Han J, Welch EJ, Ye RD, Voyno-Yasenetskaya TA, Malik AB, Du X, Li Z. 2009. Lipopolysaccharide stimulates platelet secretion and potentiates platelet aggregation via TLR4/MyD88 and the cGMP-dependent protein kinase pathway. J Immunol 182:7997–8004.
crossref pmid pmc
Zhang WY, Liu HQ, Xie KQ, Yin LL, Li Y, Kwik-Uribe CL, Zhu XZ. 2006. Procyanidin dimer B2 [epicatechin-(4β-8)-epicatechin] suppresses the expression of cyclooxygenase-2 in endotoxin-treated monocytic cells. Biochem Biophys Res Commun 345:508–515.
crossref pmid
Zhao L, Ohtaki Y, Yamaguchi K, Matsushita M, Fujita T, Yokochi T, Takada H, Endo Y. 2002. LPS-induced platelet response and rapid shock in mice: contribution of O-antigen region of LPS and involvement of the lectin pathway of the complement system. Blood 100:3233–3239.
crossref pmid


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