Effects of GHRP-2 and Cysteamine Administration on Growth Performance,Somatotropic Axis Hormone and Muscle Protein Deposition in Yaks (Bos grunniens) with Growth Retardation

The objective of this study was to investigate the effects of growth hormone-releasing peptide-2 (GHRP-2) and cysteamine (CS) administration on growth performance in yaks with growth retardation and try to elucidate its regulatory mechanisms. Trial 1, thirty-six 1-year-old Qinghai high plateau yaks (body weight 38–83.2 kg) were randomly chosen for body weight and jugular blood samples collection. The relationship between body weight and serum GHRH (P < 0.05, R = 0.45), GH (P < 0.05, R = 0.47), IGF-1 (P < 0.05, R = 0.62) was significantly correlated in yaks colonies with lighter body weights. Trial 2, fifteen 1-year-old Qinghai high plateau yaks with growth retardation (average body weight 54.8 ± 8.24 kg) were randomly selected and assigned to negative control group (NG), GHRP-2 injection group (GG) and cysteamine feeding group (CG), with 5 yaks per group. Another five 1-year-old Qinghai high plateau yaks with normal growth performance (average body weight 75.3 ± 2.43 kg) were selected as positive control group (PG). The average daily gain (ADG) of the GG and CG were significantly higher than those in the PG and NG (P < 0.05). Both GHRP-2 and CS administration significantly enhanced the myofiber diameter and area of skeletal muscle (P<0.05). GHRP-2 significantly enhanced the serum GH and IGF-1 levels (P < 0.05), and up-regulated GHR, IGF-1 and IGF-1R mRNA expression in the liver and skeletal muscle (P < 0.05), enhanced the mRNA expression of PI₃K, AKt and mTOR in the skeletal muscle (P<0.05). CS significantly reduced the serum SS levels and the hypothalamus SS mRNA expression (P < 0.05), and enhanced GHR and IGF-1 mRNA expression in the liver (P < 0.05), decreased the mRNA expression of muscle atrophy F-box (Atrogin-1) and muscle ring finger 1 (MuRF1) mRNA (P < 0.05). Conclusions: Growth retardation in yaks was primarily due to somatotropic axis hormones secretion deficiency. Both GHRP-2 and CS administration can accelerate growth performance and GH, IGF-1 secretion in yaks with growth retardation. GHRP-2 enhanced muscle protein deposition mainly by up-regulated the protein synthesis pathways, whereas CS worked mainly by down-regulated the ubiquitin-proteasome pathway.     

Human Muscle Protein Synthetic Responses during Weight-Bearing and Non-Weight-Bearing Exercise: A Comparative Study of Exercise Modes and Recovery Nutrition

Effects of conventional endurance (CE) exercise and essential amino acid (EAA) supplementation on protein turnover are well described. Protein turnover responses to weighted endurance exercise (i.e., load carriage, LC) and EAA may differ from CE, because the mechanical forces and contractile properties of LC and CE likely differ. This study examined muscle protein synthesis (MPS) and whole-body protein turnover in response to LC and CE, with and without EAA supplementation, using stable isotope amino acid tracer infusions. Forty adults (mean ± SD, 22 ± 4 y, 80 ± 10 kg, VO_2peak 4.0 ± 0.5 L∙min^-1) were randomly assigned to perform 90 min, absolute intensity-matched (2.2 ± 0.1 VO₂ L∙m^-1) LC (performed on a treadmill wearing a vest equal to 30% of individual body mass, mean ± SD load carried 24 ± 3 kg) or CE (cycle ergometry performed at the same absolute VO₂ as LC) exercise, during which EAA (10 g EAA, 3.6 g leucine) or control (CON, non-nutritive) drinks were consumed. Mixed-muscle and myofibrillar MPS were higher during exercise for LC than CE (mode main effect, P < 0.05), independent of dietary treatment. EAA enhanced mixed-muscle and sarcoplasmic MPS during exercise, regardless of mode (drink main effect, P < 0.05). Mixed-muscle and sarcoplasmic MPS were higher in recovery for LC than CE (mode main effect, P < 0.05). No other differences or interactions (mode x drink) were observed. However, EAA attenuated whole-body protein breakdown, increased amino acid oxidation, and enhanced net protein balance in recovery compared to CON, regardless of exercise mode (P < 0.05). These data show that, although whole-body protein turnover responses to absolute VO₂-matched LC and CE are the same, LC elicited a greater muscle protein synthetic response than CE.     

Effects of Choline on Meat Quality and Intramuscular Fat in Intrauterine Growth Retardation Pigs

The aim of this study was to investigate the effects of choline supplementation on intramuscular fat (IMF) and lipid oxidation in IUGR pigs. Twelve normal body weight (NBW) and twelve intrauterine growth retardation (IUGR) newborn piglets were collected and distributed into 4 treatments (Normal: N, Normal+Choline: N+C, IUGR: I, and IUGR+Choline: I+C) with 6 piglets in each treatment. At 23 d of age, NBW and IUGR pigs were fed basal or choline supplemented diets. The results showed that the IUGR pigs had significantly lower (P<0.05) BW as compared with the NBW pigs at 23 d, 73 d, and 120 d of age, however, there was a slight decreased (P>0.05) in BW of IUGR pigs than the NBW pigs at 200 d. Compared with the NBW pigs, pH of meat longissimus dorsi muscle was significantly lower (P<0.05), and the meat color was improved in IUGR pigs. The malondialdehyde (MDA) levels were significantly decreased (P<0.05), while triglyceride (TG) and IMF contents were significantly higher (P<0.05) in the IUGR pigs than the NBW pigs. IUGR up-regulated the mRNA gene expression of fatty acid synthetase (FAS) and acetyl-CoA carboxylase (ACC). Dietary choline significantly increased (P<0.05) the BW at 120d of age, however, significantly decreased (P<0.05) the TG and IMF contents in both IUGR and NBW pigs. FAS and sterol regulatory element-binding proteins 1 (SREBP1) mRNA gene expressions were increased (P<0.05) while the muscle-carnitine palmityl transferase (M-CPT) and peroxisome proliferators-activated receptorγ (PPARγ) mRNA (P<0.05) gene expressions were decreased in the muscles of the IUGR pigs by choline supplementation. Furthermore, choline supplementation significantly increased (P<0.05) the MDA content as well as the O₂•¯ scavenging activity in meat of IUGR pigs. The results suggested that IUGR pigs showed a permanent stunting effect on the growth performance, increased fat deposition and oxidative stress in muscles. However, dietary supplementation of choline improved the fat deposition via enhancing the lipogenesis and reducing the lipolysis.     

Signaling Pathways Related to Protein Synthesis and Amino Acid Concentration in Pig Skeletal Muscles Depend on the Dietary Protein Level,Genotype and Developmental Stages

Muscle growth is regulated by the homeostatic balance of the biosynthesis and degradation of muscle proteins. To elucidate the molecular interactions among diet, pig genotype, and physiological stage, we examined the effect of dietary protein concentration, pig genotype, and physiological stages on amino acid (AA) pools, protein deposition, and related signaling pathways in different types of skeletal muscles. The study used 48 Landrace pigs and 48 pure-bred Bama mini-pigs assigned to each of 2 dietary treatments: lower/GB (Chinese conventional diet)- or higher/NRC (National Research Council)-protein diet. Diets were fed from 5 weeks of age to respective market weights of each genotype. Samples of biceps femoris muscle (BFM, type I) and longissimus dorsi muscle (LDM, type II) were collected at nursery, growing, and finishing phases according to the physiological stage of each genotype, to determine the AA concentrations, mRNA levels for growth-related genes in muscles, and protein abundances of mechanistic target of rapamycin (mTOR) signaling pathway. Our data showed that the concentrations of most AAs in LDM and BFM of pigs increased (P<0.05) gradually with increasing age. Bama mini-pigs had generally higher (P<0.05) muscle concentrations of flavor-related AA, including Met, Phe, Tyr, Pro, and Ser, compared with Landrace pigs. The mRNA levels for myogenic determining factor, myogenin, myocyte-specific enhancer binding factor 2 A, and myostatin of Bama mini-pigs were higher (P<0.05) than those of Landrace pigs, while total and phosphorylated protein levels for protein kinase B, mTOR, and p70 ribosomal protein S6 kinases (p70S6K), and ratios of p-mTOR/mTOR, p-AKT/AKT, and p-p70S6K/p70S6K were lower (P<0.05). There was a significant pig genotype-dependent effect of dietary protein on the levels for mTOR and p70S6K. When compared with the higher protein-NRC diet, the lower protein-GB diet increased (P<0.05) the levels for mTOR and p70S6K in Bama mini-pigs, but repressed (P<0.05) the level for p70S6K in Landrace pigs. The higher protein-NRC diet increased ratio of p-mTOR/mTOR in Landrace pigs. These findings indicated that the dynamic consequences of AA profile and protein deposition in muscle tissues are the concerted effort of distinctive genotype, nutrient status, age, and muscle type. Our results provide valuable information for animal feeding strategy.     

Energy Dense,Protein Restricted Diet Increases Adiposity and Perturbs Metabolism in Young,Genetically Lean Pigs

Animal models of obesity and metabolic dysregulation during growth (or childhood) are lacking. Our objective was to increase adiposity and induce metabolic syndrome in young, genetically lean pigs. Pre-pubertal female pigs, age 35 d, were fed a high-energy diet (HED; n = 12), containing 15% tallow, 35% refined sugars and 9.1–12.9% crude protein, or a control corn-based diet (n = 11) with 12.2–19.2% crude protein for 16 wk. Initially, HED pigs self-regulated energy intake similar to controls, but by wk 5, consumed more (P<0.001) energy per kg body weight. At wk 15, pigs were subjected to an oral glucose tolerance test (OGTT); blood glucose increased (P<0.05) in control pigs and returned to baseline levels within 60 min. HED pigs were hyperglycemic at time 0, and blood glucose did not return to baseline (P = 0.01), even 4 h post-challenge. During OGTT, glucose area under the curve (AUC) was higher and insulin AUC was lower in HED pigs compared to controls (P = 0.001). Chronic HED intake increased (P<0.05) subcutaneous, intramuscular, and perirenal fat deposition, and induced hyperglycemia, hypoinsulinemia, and low-density lipoprotein hypercholesterolemia. A subset of HED pigs (n = 7) was transitioned back to a control diet for an additional six weeks. These pigs were subjected to an additional OGTT at 22 wk. Glucose AUC and insulin AUC did not improve, supporting that dietary intervention was not sufficient to recover glucose tolerance or insulin production. These data suggest a HED may be used to increase adiposity and disrupt glucose homeostasis in young, growing pigs.     

Dietary Lactobacillus rhamnosus GG Supplementation Improves the Mucosal Barrier Function in the Intestine of Weaned Piglets Challenged by Porcine Rotavirus

Lactobacillus rhamnosus GG (LGG) has been regarded as a safe probiotic strain. The aim of this study was to investigate whether dietary LGG supplementation could alleviate diarrhea via improving jejunal mucosal barrier function in the weaned piglets challenged by RV, and further analyze the potential roles for apoptosis of jejunal mucosal cells and intestinal microbiota. A total of 24 crossbred barrows weaned at 21 d of age were assigned randomly to 1 of 2 diets: the basal diet and LGG supplementing diet. On day 11, all pigs were orally infused RV or the sterile essential medium. RV infusion increased the diarrhea rate, increased the RV-Ab, NSP4 and IL-2 concentrations and the Bax mRNA levels of jejunal mucosa (P<0.05), decreased the villus height, villus height: crypt depth, the sIgA, IL-4 and mucin 1 concentrations and the ZO-1, occludin and Bcl-2 mRNA levels of jejunal mucosa (P<0.05), and affected the microbiota of ileum and cecum (P<0.05) in the weaned pigs. Dietary LGG supplementation increased the villus height and villus height: crypt depth, the sIgA, IL-4, mucin 1 and mucin 2 concentrations, and the ZO-1, occludin and Bcl-2 mRNA levels of the jejunal mucosa (P<0.05) reduced the Bax mRNA levels of the jejunal mucosa (P<0.05) in weaned pigs. Furthermore, dietary LGG supplementation alleviated the increase of diarrhea rate in the weaned pigs challenged by RV (P<0.05), and relieve the effect of RV infection on the villus height, crypt depth and the villus height: crypt depth of the jejunal mucosa (P<0.05), the NSP4, sIgA, IL-2, IL-4, mucin 1 and mucin 2 concentrations of jejunal mucosa (P<0.05), the ZO-1, occludin, Bax and Bcl-2 mRNA levels of the jejunal mucosa (P<0.05), and the microbiota of ileum and cecum (P<0.05) in the weaned pigs challenged by RV. These results suggest that supplementing LGG in diets alleviated the diarrhea of weaned piglets challenged by RV via inhibiting the virus multiplication and improving the jejunal mucosal barrier function, which was possibly due to the decreasing apoptosis of jejunal mucosal cells and the improvement of intestinal microbiota.     

Dietary Leucine Supplementation Improves the Mucin Production in the Jejunal Mucosa of the Weaned Pigs Challenged by Porcine Rotavirus

The present study was mainly conducted to determine whether dietary leucine supplementation could attenuate the decrease of the mucin production in the jejunal mucosa of weaned pigs infected by porcine rotavirus (PRV). A total of 24 crossbred barrows weaned at 21 d of age were assigned randomly to 1 of 2 diets supplemented with 1.00% L-leucine or 0.68% L-alanine (isonitrogenous control) for 17 d. On day 11, all pigs were orally infused PRV or the sterile essential medium. During the first 10 d of trial, dietary leucine supplementation could improve the feed efficiency (P = 0.09). The ADG and feed efficiency were impaired by PRV infusion (P<0.05). PRV infusion also increased mean cumulative score of diarrhea, serum rotavirus antibody concentration and crypt depth of the jejunal mucosa (P<0.05), and decreased villus height: crypt depth (P = 0.07), goblet cell numbers (P<0.05), mucin 1 and 2 concentrations (P<0.05) and phosphorylated mTOR level (P<0.05) of the jejunal mucosa in weaned pigs. Dietary leucine supplementation could attenuate the effects of PRV infusion on feed efficiency (P = 0.09) and mean cumulative score of diarrhea (P = 0.09), and improve the effects of PRV infusion on villus height: crypt depth (P = 0.06), goblet cell numbers (P<0.05), mucin 1 (P = 0.08) and 2 (P = 0.07) concentrations and phosphorylated mTOR level (P = 0.08) of the jejunal mucosa in weaned pigs. These results suggest that dietary 1% leucine supplementation alleviated the decrease of mucin production and goblet cell numbers in the jejunal mucosa of weaned pigs challenged by PRV possibly via activation of the mTOR signaling.     

l-arginine supplementation of milk liquid or dry diets fed to pigs after weaning has a positive effect on production in the first three weeks after weaning at 21 days of age

A total of 48, 21-day-old weaned pigs, was used in a 2 × 2 factorial arrangement of treatments with the factors being diet type (milk liquid replacer vs. dry feed) and l-arginine (ARG) supplementation (0 vs. 6 g ARG/kg) to test the hypothesis that dietary supplementation with ARG would increase performance of pigs after weaning. Pigs were fed the experimental diets for 10 d (days) after weaning and then transitioned over a 3-d period to a dry Phase II diet fed in meal form devoid of supplemental ARG. The study ended at d 21. There were five replicates (pens) per treatment (a total of 12 pigs per treatment). Blood samples were collected from two pigs per replicate on d 7 and 16 of the experiment, and free amino acids (AA) and plasma urea nitrogen (PUN) levels analysed. Milk-fed pigs outperformed (P<0.001) dry-fed pigs for the first 10 d of the experiment as well as for the total 21-d period. At d 7, milk-fed pigs had higher (P<0.05) levels of most free indispensable and dispensable amino acids in their plasma. In both the milk-fed and dry-fed pigs supplemented with ARG, average daily feed intake (ADFI, P<0.05) and average daily gain (ADG, P<0.05) were increased during the dietary transition period (d 11–14), when pigs were being changed to the Phase II diet. The difference in production in the transition period caused a tendency for ARG-supplemented pigs to eat more feed (P<0.1) and grow faster (P<0.5) over the 21-d experimental period. Pigs supplemented with ARG had higher plasma ARG levels (P<0.05) at d 7 after weaning and lower plasma urea levels (P<0.05) at both d 7 and 16 after weaning. These data show the benefits of feeding a milk liquid diet as well as of ARG supplementation after weaning on production indices.     

Serum levels of appetite-regulating hormones and pro-inflammatory cytokines are ameliorated by a CLA diet and endurance exercise in rats fed a high-fat diet

[Purpose]

This study examined whether conjugated linoleic acid (CLA) supplementation and endurance exercise affect appetite-regulating hormones and pro-inflammatory cytokines in rats.

[Methods]

Seven-week-old male Sprague-Dawley rats were divided randomly into the high-fat diet sedentary group (HS, n=8), the 1.0% CLA supplemented high-fat diet sedentary group (CS, n=8), and the 1.0% CLA supplemented high-fat diet exercise group (CE, n=8). Rats in the CE group swam 60 min/day, 5 days/week for 4 weeks.

[Results]

Leptin and insulin levels in the CS and CE groups were significantly lower than those in the HS group (p<0.001), whereas leptin (p<0.01) and insulin (p<0.05) levels decreased significantly in the CE compared to those in the CS group. Interleukin (IL)-1β (p<0.001) and IL-6 (p<0.01) levels in the CS and CE groups decreased significantly compared to those in the HS group. Leptin (IL-1β: r=0.835, p<0.001), IL-6 (r=0.607, p<0.05), insulin (IL-1β: r=0.797, p<0.01), and IL-6 (r=0.827, p<0.01) levels were positively related with pro-inflammatory cytokine levels.

[Conclusion]

Endurance exercise may play an important role during CLA supplementation of rats on a high-fat diet.     

Multi-Functional Roles of Chitosan as a Potential Protective Agent against Obesity

Chitosan, a natural polysaccharide comprising copolymers of glucosamine and N-acetylglucosamine, has been shown to have anti-obesity properties. Two experiments (Exp. 1 and Exp. 2) were performed to determine the role of chitosan on dietary intake, body weight gain, and fat deposition in a pig model, as well as identifying potential mechanisms underlying the anti-obesity effect of chitosan. In Exp. 1, the nutrient digestibility experiment, 16 pigs (n = 4/treatment) were randomly allocated to one of four dietary treatments as follows: 1) basal diet; 2) basal diet plus 300 ppm chitosan; 3) basal diet plus 600 ppm chitosan; 4) basal diet plus 1200 ppm chitosan. The main observation was that crude fat digestibility was lower in the 1200 ppm chitosan group when compared with the control group (P<0.05). In Exp. 2, a total of 80 pigs (n = 20/treatment) were offered identical dietary treatments to that offered to animals in Exp. 1. Blood samples were collected on day 0, day 35 and at the end of the experiment (day 57). Animals offered diets containing 1200 ppm chitosan had a lower daily dietary intake (P<0.001) and body weight gain (P<0.001) from day 35 to 57 when compared with all the other treatment groups. Animals offered diets containing 1200 ppm chitosan had a significantly lower final body weight (P<0.01) when compared with all the other treatment groups. The decreased dietary intake observed in the 1200 ppm chitosan group was associated with increased serum leptin concentrations (P<0.001) and a decrease in serum C-reactive protein (CRP) concentrations (P<0.05). In conclusion, the results of this study highlight novel endocrine mechanisms involving the modulation of serum leptin and CRP concentrations by which chitosan exhibits anti-obesity properties in vivo.     

Maternal dietary omega-3 fatty acid intake increases resolvin and protectin levels in the rat placenta

Placental inflammation is associated with several pregnancy disorders. Inflammation is limited by anti-inflammatory and proresolving mechanisms, the latter partly mediated by resolvins and protectins derived from omega-3 polyunsaturated fatty acids (n-3PUFA). We examined effects of dietary n-3PUFAs on levels of resolvins, protectins, and lipoxygenase (ALOX) enzymes in the rat placenta. Rats consumed standard (Std) or high n-3PUFA (Hn3) diets from day 1 of pregnancy; tissues were collected on day 17 or 22 (term = day 23). Maternal Hn3 diet increased resolvin and protectin precursors, 18R/S-HEPE (P < 0.001), and 17R/S-HDHA (P < 0.01) at both days. Resolvins (17R-RvD1 and RvD1) increased at day 22 (P < 0.001) after Hn3 consumption, coincident with higher Alox15b and Alox5 mRNA expression, while RvD2 increased at both days (P < 0.05). Protectins, PD1, and 10S,17S-DiHDHA increased over late gestation (P < 0.001), coincident with higher Alox15 mRNA expression (P < 0.001) and further increased with Hn3 diet (P < 0.05). Maternal systemic and placental proinflammatory mediators were not suppressed by Hn3 diet; systemic IL1β, placental Il1β, and Il6 mRNA expression increased marginally with Hn3 at day 22 (P < 0.001), while Ptgs1 (Cox1) expression increased both days (P < 0.05). Our data indicate that maternal n-3PUFA supplementation enhances expression of enzymes in the n-3PUFA metabolic pathway and increases placental levels of resolvins and protectins.     

Mitochondrial Regulation of NADPH Oxidase in Hindlimb Unweighting Rat Cerebral Arteries

Exposure to microgravity results in post-flight cardiovascular deconditioning and orthostatic intolerance in astronauts. Vascular oxidative stress injury and mitochondrial dysfunction have been indicated in this process. To elucidate the mechanism for this condition, we investigated whether mitochondria regulated NADPH oxidase in hindlimb unweighting (HU) rat cerebral and mesenteric arteries. Four-week HU was used to simulate microgravity in rats. Vascular superoxide generation, protein and mRNA levels of Nox2/Nox4, and the activity of NADPH oxidase were examined in the present study. Compared with control rats, the levels of superoxide increased in cerebral (P<0.001) but not in mesenteric vascular smooth muscle cells. The protein and mRNA levels of Nox2 and Nox4 were upregulated significantly (P<0.001 and P<0.001 for Nox2, respectively; P<0.001 and P<0.001 for Nox4, respectively) in HU rat cerebral arteries but not in mesenteric arteries. NADPH oxidases were activated significantly by HU (P<0.001) in cerebral arteries but not in mesenteric arteries. Chronic treatment with mitochondria-targeted antioxidant mitoTEMPO attenuated superoxide levels (P<0.001), decreased the protein and mRNA expression levels of Nox2/Nox4 (P<0.01 and P<0.05 for Nox2, respectively; P<0.001 and P<0.001 for Nox4, respectively) and the activity of NADPH oxidase (P<0.001) in HU rat cerebral arteries, but exerted no effects on HU rat mesenteric arteries. Therefore, mitochondria regulated the expression and activity of NADPH oxidases during simulated microgravity. Both mitochondria and NADPH oxidase participated in vascular redox status regulation.     

The Association of Uremic Toxins and Inflammation in Hemodialysis Patients

Background

Cardiovascular disease is the leading cause of mortality in hemodialysis patients and is associated with chronic inflammation. Elevation of uremic toxins, particular protein-bound uremic toxins, is a possible cause of hyper-inflammation in hemodialysis patients. But the association between uremic toxins and inflammatory markers in hemodialysis is still unclear.

Methods

We conducted a cross-sectional study to evaluate the association of the serum uremic toxins and inflammatory markers in hemodialysis patients.

Results

The uremic toxins were not associated with inflammatory markers- including high sensitivity C-reactive protein, IL(Interleukin) -1β, IL-6, tumor necrosis factor-α. In multiple linear regression, serum levels of total p-cresol sulfate (PCS) were independently significantly associated with serum total indoxyl sulfate (IS) (standardized coefficient: 0.274, p<0.001), and co-morbidity of diabetes mellitus (DM) (standardized coefficient: 0.342, p<0.001) and coronary artery disease (CAD) (standardized coefficient: 0.128, p = 0.043). The serum total PCS levels in hemodialysis with co-morbidity of DM and CAD were significantly higher than those without co-morbidity of DM and CAD (34.10±23.44 vs. 16.36±13.06 mg/L, p<0.001). Serum levels of total IS was independently significantly associated with serum creatinine (standardized coefficient: 0.285, p<0.001), total PCS (standardized coefficient: 0.239, p = 0.001), and synthetic membrane dialysis (standardized coefficient: 0.139, p = 0.046).

Conclusion

The study showed that serum levels of total PCS and IS were not associated with pro-inflammatory markers in hemodialysis patients. Besides, serum levels of total PCS were independently positively significantly associated with co-morbidity of CAD and DM.     

Hydroxyurea affects in vitro porcine oocyte maturation through increased apoptosis and oxidative stress

Hydroxyurea (HU) is an FDA-approved drug used to treat a variety of diseases, especially malignancies, but is harmful to fertility. We used porcine oocytes as an experimental model to study the effect of HU during oocyte maturation. Exposure of cumulus–oocyte complexes (COCs) to 20 µM (P<0.01) and 50 µM (P<0.001) HU reduced oocyte maturation. Exposure to 20 µM HU induced approximately 1.5- and 2-fold increases in Caspase-3 (P<0.001) and P53 (P<0.01) gene expression levels in cumulus cells, respectively, increased Caspase-3 (P<0.01) and P53 (P<0.001) protein expression levels in metaphase II (MII) oocytes and increased the percentage of apoptotic cumulus cells (P<0.001). In addition, HU decreased the mitochondrial membrane potential (Δφm) (P<0.01 and P<0.001) and glutathione (GSH) levels (P<0.01 and P<0.001) of both cumulus cells and MII oocytes, while increasing their reactive oxygen species (ROS) levels (P<0.001). Following parthenogenetic activation of embryos derived from MII oocytes, exposure to 20 µM HU significantly reduced total blastocyst cell numbers (P<0.001) and increased apoptosis of blastocyst cells (P<0.001). Moreover, HU exposure reduced the rate of development of two-celled, four- to eight-celled, blastocyst, and hatching stages after parthenogenetic activation (P<0.05). Our findings indicate that exposure to 20 µM HU caused significant oxidative stress and apoptosis of MII oocytes during maturation, which affected their developmental ability. These results provide valuable information for safety assessments of HU.     

Effects of Dietary Pantothenic Acid on Growth,Intestinal Function,Anti-Oxidative Status and Fatty Acids Synthesis of Juvenile Blunt Snout Bream Megalobrama amblycephala

Four groups of juvenile Megalobrama amblycephala were fed three times daily with six semi-purified diets containing 3.39 (PA unsupplied diet), 10.54, 19.28, 31.04, 48.38 and 59.72 mg kg^-1 calcium D-pantothenate. The results showed that survival rate, final weight, specific growth rate, protein efficiency ratio and nitrogen retention efficiency all increased significantly (P<0.01) as dietary PA levels increased from 3.39 to 19.28 mg kg^-1, whereas the opposite was true for feed conversion ratio. Whole-body crude protein increased as dietary PA levels increased, while the opposite pattern was found for the crude lipid content. Intestinal α-amylase, lipase, protease, Na⁺-K⁺-ATPase, alkaline phosphatase and gamma-glutamyl transferase activities were all elevated in fish fed PA-supplemented diets. Hepatic catalase activities improved with increases in dietary PA, while the opposite was true for malondialdehyde contents. The liver PA concentration and coenzyme A content rose significantly (P<0.01), up to 31.04 mg kg^-1, with increasing dietary PA levels and then plateaued. The percentage of hepatic saturated fatty acids increased significantly (P<0.01) as dietary PA levels increased, while the percentages of monounsaturated fatty acids and polyunsaturated fatty acid (PUFA) decreased as dietary PA increased. Fish fed diets containing 19.28 and 31.04 mg kg^-1 PA exhibited higher (P<0.01) docosahexaenoic acid and PUFA percentages in muscle than those fed with other diets. The expression of the gene encoding pantothenate kinase was significantly up-regulated (P<0.01) in fish fed PA-supplemented diets. Hepatic Acetyl-CoA carboxylase α, fatty acid synthetase, stearoyl regulatory element-binding protein 1 and X receptor α genes all increased significantly (P<0.01) as dietary PA levels increased from 3.39 to 31.04 mg kg^-1. Based on broken-line regression analyses of weight gain, liver CoA concentrations and PA contents against dietary PA levels, the optimal dietary PA requirements of juvenile blunt snout bream were estimated to be 24.08 mg kg^-1.     

Prognostic Importance and Therapeutic Implications of PAK1, a Drugable Protein Kinase,in Gastroesophageal Junction Adenocarcinoma

Gastroesophageal junction (GEJ) adenocarcinoma is a lethal cancer with rising incidence, yet the molecular biomarkers that have strong prognostic impact and also hold great therapeutic promise remain elusive. We used a data mining approach and identified the p21 protein-activated kinase 1 (PAK1), an oncogene and drugable protein kinase, to be among the most promising targets for GEJ adenocarcinoma. Immunoblot analysis and data mining demonstrated that PAK1 protein and mRNA were upregulated in cancer tissues compared to the noncancerous tissues. Immunohistochemistry revealed PAK1 overexpression in 72.6% of primary GEJ adenocarcinomas (n = 113). A step-wise increase in PAK1 levels was noted from paired normal epithelium, to atypical hyperplasia and adenocarcinoma. PAK1 overexpression in tumor was associated with lymph node (LN) metastasis (P<0.001), advanced tumor stage (P<0.001), large tumor size (P = 0.006), residual surgical margin (P = 0.033), and unfavorable overall survival (P<0.001). Multivariate analysis showed PAK1 overexpression is an independent high-risk prognostic predictor (P<0.001). Collectively, PAK1 is overexpressed during tumorigenic progression and its upregulation correlates with malignant properties mainly relevant to invasion and metastasis. PAK1 expression could serve as a prognostic predictor that holds therapeutic promise for GEJ adenocarcinoma.     

High Expression of Heat Shock Protein 90 Is Associated with Tumor Aggressiveness and Poor Prognosis in Patients with Advanced Gastric Cancer

The heat shock protein 90 (HSP90) is overexpressed and highly associated with poor prognosis in many malignancies. However, the role of HSP90 in gastric cancer has not been thoroughly elucidated. The aim of this study is to investigate the relationship of HSP90 expression with clinicopathological parameters and prognosis in advanced gastric cancer, and estimate the alteration of HSP90 expression after neoadjuvant chemotherapy. HSP90 and matrix metallopeptidase 9 (MMP-9) antigen expression was evaluated by immunohistochemistry in 322 advanced gastric carcinoma samples. The relationships between HSP90 and clinicopathological parameters and prognosis were analyzed. The response of HSP90 level was assessed in chemotherapeutic effect in 54 patients received 1–2 cycles of neoadjuvant chemotherapy. The positive expression of HSP90 was found to be 69.6% in 322 advanced gastric carcinoma samples. HSP90 protein expression was significantly associated with depth invasion (P<0.001), lymph node metastasis (P<0.001) and stage of disease (P<0.001). The positive rates of HSP90 expression were higher in both prominent serosal invasion group (P<0.001) and lymph node metastasis group (P<0.001). Moreover, HSP90 was significantly correlated with MMP-9 among 322 gastric cancer tissues (P<0.001). In univariate and multivariate analyses, HSP90 was an independent prognostic factor for both recurrence-free survival (RFS) and overall survival (OS). These results suggested that HSP90 may play an important role on tumor invasion, metastasis and prognosis, and might act as a promising target for prognostic prediction.     

Intestinal Microbiota and Microbial Metabolites Are Changed in a Pig Model Fed a High-Fat/Low-Fiber or a Low-Fat/High-Fiber Diet

The intestinal microbiota and its metabolites appear to be an important factor for gastrointestinal function and health. However, research is still needed to further elaborate potential relationships between nutrition, gut microbiota and host’s health by means of a suitable animal model. The present study examined the effect of two different diets on microbial composition and activity by using the pig as a model for humans. Eight pigs were equally allotted to two treatments, either fed a low-fat/high-fiber (LF), or a high-fat/low-fiber (HF) diet for 7 weeks. Feces were sampled at day 7 of every experimental week. Diet effects on fecal microbiota were assessed using quantitative real-time PCR, DNA fingerprinting and metaproteomics. Furthermore, fecal short-chain fatty acid (SCFA) profiles and ammonia concentrations were determined. Gene copy numbers of lactobacilli, bifidobacteria (P<0.001) and Faecalibacterium prausnitzii (P<0.05) were higher in the LF pigs, while Enterobacteriaceae were more abundant in the HF pigs (P<0.001). Higher numbers of proteins affiliated to Enterobacteriaceae were also present in the HF samples. Proteins for polysaccharide breakdown did almost exclusively originate from Prevotellaceae. Total and individual fecal SCFA concentrations were higher for pigs of the LF treatment (P<0.05), whereas fecal ammonia concentrations did not differ between treatments (P>0.05). Results provide evidence that beginning from the start of the experiment, the LF diet stimulated beneficial bacteria and SCFA production, especially butyrate (P<0.05), while the HF diet fostered those bacterial groups which have been associated with a negative impact on health conditions. These findings correspond to results in humans and might strengthen the hypothesis that the response of the porcine gut microbiota to a specific dietary modulation is in support of using the pig as suitable animal model for humans to assess diet-gut-microbiota interactions.Data are available via ProteomeXchange with identifier PXD003447.