Molecular cloning, tissue expression, and subcellular localization of porcine peptidoglycan recognition proteins 3 and 4

Peptidoglycan recognition proteins (PGRPs) are innate immune molecules that are present in most invertebrates and vertebrates. Mammals have four PGRPs, PGLYRP1-4. In the present study, we cloned the cDNAs encoding porcine PGLYRP3 and 4 from the esophagus of adult swine. The length of the complete open reading frames of porcine PGLYRP3 and 4 are identical and contain 1125bp encoding 374 amino acid residues. The amino acid sequences of these two proteins were more similar to their human orthologs (78.9% [PGLYRP3] and 73.9% [PGLYRP4]) than to their mouse orthologs (71.3% [PGLYRP3] and 67.9% [PGLYRP4]). Expression analysis revealed that both PGLYRP3 and 4 were more strongly expressed in digestive tract, especially the esophagus, than in immune organs such as spleen or mesenteric lymph nodes in both newborn and adult swine. To analyze the subcellular distribution of porcine PGLYRP1-4, we constructed transfectant cell lines. Western blot and flow cytometric analyses revealed that porcine PGLYRP3 and 4 are not only secreted, but also expressed on the cell surface, unlike PGLYRP1 and 2. These results should help contribute to the understanding of PGLYRP3- and 4-mediated immune responses via their recognition of intestinal microorganisms in newborn and adult swine.     

Central insulin suppresses feeding behavior via melanocortins in chicks

Growing evidence suggests that insulin interacts with both orexigenic and anorexigenic peptides in the brain for the control of feeding behavior in mammals. However, the action of central insulin in chicks has not yet been identified. In the present study, we investigated the effects of central injection of insulin on feeding behavior in chicks. Intracerebroventricular (ICV) administration of insulin, at doses that do not influence peripheral glucose levels, significantly inhibited food intake in chicks. Central injection of insulin in chicks significantly increased expression of pro-opiomelanocortin (POMC) mRNA, and decreased that of neuropeptide Y (NPY) mRNA. Finally, co-injection of the melanocortin antagonist (SHU9119 or HS014) prevented the reduction in food intake caused by ICV administration of insulin. These data suggest that insulin functions in chicks as an appetite-suppressive peptide in the central nervous system, and that the central melanocortin system mediates this anorexic effect of insulin, as in mammals.     

Inosine‐5'‐monophosphate is a candidate agent to resolve rigor mortis of skeletal muscle

The object of the present study was to reveal the action of inosine‐5'‐monophosphate (IMP) toward myofibrils in postmortem muscles. IMP solubilized isolated actomyosin within a narrow range of KCl concentration, 0.19‐0.20 mol/L, because of the dissociation of actomyosin into actin and myosin, but it did not solubilize the proteins in myofibrils with 0.2 mol/L KCl. However, IMP could solubilize both proteins in myofibrils with 0.2 mol/L KCl in the presence of 1 m mol/L pyrophosphate or 1.0–3.3 m mol/L adenosine‐5'‐diphosphate (ADP). Thus, we presumed that pyrophosphate and ADP released thin filaments composed of actin, and thick filaments composed of myosin from restraints of myofibrils, and then both filaments were solubilized through the IMP‐induced dissociation of actomyosin. Thus, we concluded that IMP is a candidate agent to resolve rigor mortis because of its ability to break the association between thick and thin filaments.     

Influence of Polyunsaturated Fatty Acid Intake on Kidney Functions of Rats with Chronic Renal Failure

Arachidonic acid (ARA), an omega-6 (ω-6) polyunsaturated fatty acid (PUFA), is involved in the development and maintenance of renal functions, whereas docosahexaenoic acid (DHA) is an omega-3 (ω-3) PUFA that has anti-inflammatory effects and attenuates nephropathy. However, their effects on the progression of chronic kidney disease (CKD) remain unknown. The aim of this study was to assess the effects of feeding ARA, DHA, and ARA and DHA-containing diets on rats with 5/6 nephrectomized kidneys. Urine and feces were collected every 4 weeks, and the kidneys were collected at 16 weeks after surgery. Urinary albumin (U-ALB) excretion increased gradually with nephrectomy, but the U-ALB excretion was attenuated by feeding the rats with an ARA + DHA-containing diet. Reactive oxygen species (ROS) levels in the kidneys were lower in the ARA + DHA group than in the other groups. At 4 weeks after surgery, the lipid peroxide (LPO) levels in the plasma of the ARA + DHA groups decreased significantly after surgery compared to the control CKD group, but this did not happen at 16 weeks post-surgery. There was a significant negative correlation between LPO levels in the plasma at 4 weeks and creatinine clearance, and a positive correlation with urinary albumin levels. These results suggest that the combination of ARA and DHA inhibit the progress of early stage CKD.     

Identification and kinetic study of tyrosinase inhibitors found in sake lees

The present study found that the n-hexane extract of freeze-dried sake lees inhibits tyrosinase activity and showed that the constituents isolated from the n-hexane extract are the mixture of triacylglycerols. The inhibitory effects of triolein and trilinolein found as the triacylglycerols were examined using tyrosinases from mushroom and Streptomyces castaneoglobisporus. The IC50 values of the triacylglycerol mixture for the oxidase activity on mushroom and Streptomyces tyrosinases were 20 and 0.14 microg/mL, respectively. The IC50 values of trilinolein for the oxidase activity on mushroom and Streptomyces tyrosinases were 8.4 and 0.1 microM, respectively. However, the inhibitory effect of triolein (IC50=30 microM) was lower than that of trilinolein, even when the Streptomyces tyrosinase was used for the assay. Kinetic analyses indicate that both trilinolein and triolein inhibit the tyrosinase activity noncompetitively. When transformed with a plasmid carrying the Streptomyces tyrosinase gene, the melanin-synthesizing ability of the transformed Escherichia coli host was dose-dependently interfered with by trilinolein.     

Intrafollicular Concentrations of Steroid Hormones and PGF2α in Relation to Follicular Development in the Mares during the Breeding Season

The concentrations of androstenedione, estradiol-17β, progesterone and PGF_2α contained in the follicular fluid produced by the follicles in collected ovaries of mares that have had estrous phase during the breeding season were measured and analyzed the relation between the growth stage of follicles and the hormone levels in the follicular fluid. An ultrasonographic diagnostic instrument was used to measure the diameter of the follicles in order to categorize the follicles into three groups the following: 8 small follicles (from 1.0 to less than1.5 cm), 8 medium follicles (from 1.5 to less than 3.0 cm), and 8 large follicles (from 3.0 to 5.0 cm), respectively. The analysis of the follicular fluid in ovaries of estrous mares showed that the concentrations of androstenedione were significantly higher in the medium or large follicles than in the small follicles and the concentrations of estradiol-17β were significantly higher in larger follicles than in the small or medium follicles (P<0.05). The concentrations of progesterone and PGF_2α, on the other hand, did not significantly vary regardless of follicluar size. In the follicles within the mare ovaries that have had estrous stage, the concentrations of the hormones related the ovulation, namely androstenedione and estradiol-17β, were higher with larger follicles.     

Advanced molecular immunoassay system for immunobiotic lactic acid bacteria using a transfectant of Toll-like receptor 2

Toll‐like receptor 2 (TLR2) is a receptor for a variety of microbial components, and it also mediates activation signals in the cell relating to the innate immune system. In order to evaluate the precise molecular immunoregulation by various strains of lactic acid bacteria (LAB) via TLR2, the swine TLR2 (sTLR2)‐expressing transfectant was constructed using human embryonic kidney (HEK) 293 cells. It is demonstrated that intact immunobiotic LAB can induce immune responses through TLR2, and that different nuclear factor‐κB (NF‐κB) activities of various strains can be accurately detected by sTLR2‐expressing HEK293 cells. Furthermore, cellular activation of NF‐κB via TLR2 is reflected in enhanced binding and uptake of LAB. The sTLR2‐expressing HEK293 cells were also useful for characterizing the expression pattern of type I helper T (Th1) and type II helper T (Th2) cytokines by the stimulation of immunobiotic LAB. These results suggest that sTLR2‐expressing HEK293 cells may be useful in certain molecular immunoassay systems for producing new physiologically functional foods with intestinal immunomodulatory abilities, such as the maintenance of Th1/Th2 polarization.     

Interval mapping of growth quantitative trait loci in Japanese Black beef cattle using microsatellite DNA markers and half-sib regression analysis

A confirmatory scan for the regions of bovine chromosome 1 segregating the quantitative trait loci (QTL) influencing birthweight, weaning weight, yearling weight, and preweaning and postweaning average daily gains was performed by genotyping half‐sib progeny of four Japanese Black sires using microsatellite DNA markers. Data were analyzed by generating an F‐statistic every 1 cM on a linkage map by the regression of phenotype on the probabilities of inheriting an allele from the sire after adjusting for the fixed effects of sire, sex, parity and season of birth as well as age as a covariate. Permutation tests at chromosome‐wide significance thresholds were carried out over 10 000 iterations. A significant QTL for birthweight at 114 cM was detected in the sire 2 family. This identification of a birthweight QTL in Japanese Black cattle may be useful for the implementation of marker‐assisted selection.     

Effects of cannabidiol without delta-9-tetrahydrocannabinol on canine atopic dermatitis: A retrospective assessment of 8 cases

Objective We aimed to examine the effects of cannabidiol (CBD)-containing hemp oil without delta-9-tetrahydrocannabinol (THC) as a supplemental treatment for canine atopic dermatitis (CAD), as well as its adverse effects, and effects on concurrent drug use in dogs.Animal In this retrospective case series, 8 dogs with CAD were diagnosed by veterinary dermatologists certified by the Japanese Society of Veterinary Dermatology.Procedure The medical records of dogs supplemented with CBD-containing hemp oil were evaluated with respect to signalment, physical examination, plasma C-reactive protein concentrations, pharmacologic management, the CAD Extent and Severity Index (4th iteration), and the Pruritus Visual Analog Scale.Results Overall, CBD, used as a supplement in combination with other drugs, was well-tolerated over a wide dose range and decreased the occurrence of pruritus in dogs with CAD when ingested twice a day.Conclusion This study provides the first report of supplementation with CBD without THC that was effective in controlling pruritic behavior in dogs with CAD.Clinical relevance Further controlled studies are required to investigate the dose range, efficacy, and safety.