Mechanism of macrophage activation by chitin derivatives

In order to analyze the detailed mechanisms responsible for macrophage activation by chitin derivatives, resident peritoneal macrophages were prepared and stimulated with chitin, chitosan and low-molecular weight chitosan. Our findings were as follows: (i) chitosan induced apoptosis of peritoneal macrophages, but this did not occur when chitin or water soluble low-molecular weight chitosan were used; (ii) chitosan treatment induced activation markers, such as the major histocompatibility complex (MHC) class I, class II, Fc receptors, transferrin receptor, mannose receptor, Fas, and macrophage inflammatory protein (MIP)-2, whereas chitin and low molecular weight soluble chitosan induced only the expression of MHC class I and II molecules; (iii) apoptosis induced by chitosan was mediated by the Fas signaling pathway, in response to phagocytosis via the mannose receptor. We conclude that since chitosan activates macrophages, this may be the mechanism by which it accelerates wound healing.     

Establishment of a chicken monoclonal antibody panel against mammalian prion protein

A panel of chicken monoclonal antibodies (mAbs) was developed against prion protein (PrP), the sequence of which is a highly conserved molecule among mammals. A portion of the splenocytes from chickens immunized with recombinant mouse PrP was fused with the chicken B cell line, MuH1. The remaining splenocytes were used to generate the recombinant mAbs by phage display. A total of 36 anti-PrP mAbs, 2 from cell fusion and 34 from phage display were established. The specificity of these mAbs was determined by Western blot and ELISA using various PrP antigens including recombinant PrPs, synthetic PrP peptides and PrPs from brains or scrapie-infected neuroblastoma cell line. These mAbs were classified into three main groups, protease K (PK)-sensitive (Group I), PK cleavage site proximal (Group II) and PK-resistant (Group III), based on their abilities to recognize PrP following PK-treatment. Some mAbs were found to selectively recognize different glycoforms of PrP as well as the metabolic fragments of PrP. Furthermore, we found that PrP recognition by chickens differed from that by PrP-knockout mouse. These results indicate that these newly generated PrP antibodies from chickens will help to research the PrP and to establish the diagnosis of prion disease.     

Relationship of disease progression and plasma histamine concentrations in 11 dogs with mast cell tumors

Plasma histamine concentrations (PHCs) were measured serially over 9 months or until death in 11 dogs with mast cell tumors (MCTs). Eight dogs had grossly visible disease and the other 3 dogs had microscopic disease. Initial PHCs in the dogs with gross disease were significantly higher than PHCs in healthy dogs (median, 0.73 ng/mL and 0.19 ng/mL respectively; P < .009), whereas initial PHCs in dogs with microscopic disease showed no difference from controls. Seven dogs subsequently had progressive increases in PHC, and developed hyperhistaminemia (median, 14.0 ng/mL; range, 5.11-30.1 ng/nL). These 7 dogs died from MCTs, and 1 had general weakness with rapid lysis of a large tumor burden after radiation therapy. PHCs of the other 4 dogs were less than 1 ng/mL during the study. These 4 dogs were still alive with adequate control of the tumor at the conclusion of the study. Four of the 11 dogs initially had gastrointestinal (G1) signs, which abated soon after administration of histamine-2 (H-2) blockers. No significant difference was found between PHCs in dogs with GI signs and those without GI signs (median, 0.86 ng/mL and 0.35 ng/mL. respectively). Thereafter, 7 dogs had serious GI complications for which H-2 blocker therapy was ineffective. PHCs in these 7 dogs were extremely high (median, 12.2 ng/mL; range, 3.42-30.1 ng/nL). Results of this study demonsrated that PHC was one factor related to disease progression, and indicated that marked hyperhistaminemia was associated with the GI signs refractory to H-2 blocker therapy in dogs with MCTs.     

Estrogen receptor alpha expression in the medial preoptic area and the medial basal hypothalamus under different physiological conditions in cattle

The distribution and morphology of immunoreactive estrogen receptor alpha (ERalpha)- containing cells were examined in the cattle brain. The brains were collected from two cows and a male calf. One of the cows was sacrificed one week after parturition. ERalpha expressions in the preoptic area of the rostral forebrain and the medial basal hypothalamic area were evaluated immunohistochemically using a mouse monoclonal antibody. In all animals, the signals that indicate ERalpha were detected in the medial preoptic area, the level of the organum vasculosum of the lamina terminalis and the bed nucleus of the stria terminalis. In the caudal hypothalamic region, they were detected in the arcuate nucleus, the periventricular and ventromedial hypothalamic nuclei. ERalpha-containing cells were characterized by a dense nuclear reaction product. In all the subjects examined, ERalpha signals were clearly detected in the cytoplasm as well. The density of ERalpha expression was different among the three cattle; the number of ERalpha containing cells in the postpartum cow was much lower than that in the other cow and the male calf. The present findings suggest that ERalpha expression in the forebrain is influenced by the reproductive status in the cattle.     

An etiological investigation of domestic cats with conjunctivitis and upper respiratory tract disease in Japan

Chlamydophila felis (C. felis), feline herpesvirus-1 (FHV-1) and feline calicivirus (FCV) were detected in 39 (59.1%), 11 (16.7%) and 14 (21.2%) cats respectively, from 66 domestic cats presented with conjunctivitis and upper respiratory tract disease (URTD) in 9 prefectures of Japan. Dual and multiple infections were found in 7 (10.6%) cats with both C. felis and FHV-1, 10 (15.2%) cats with both C. felis and FCV, and 1 (1.5%) cat with all three agents. C. felis was isolated from 11 (28.2%) of 39 PCR positive cats. Antigenic difference was found in a 96 kDa protein of our isolates and Fe/145 strain isolated in USA. In conclusion, C. felis is the most common agent of feline conjunctivitis and URTD, and the coinfection with C. felis, FHV-1 and FCV are also common in cats in Japan.     

Incidence of Salmonella infection in healthy dogs in Gifu Prefecture,Japan

A total of 1,013 feces samples and 8 mesenteric lymphonodus samples obtained from apparently healthy dogs were examined for the incidence of salmonella infection. One strain of S. typhimurium (ST) was isolated from feces of one dog, and S. enteritidis (SE) was isolated from the mesenteric lymphonodus of one dog. Sera obtained from 330 apparently healthy dogs were examined for Salmonella antibodies using an ELISA with heated whole cells of SE and ST. Fifty-one of the 330 serum samples were considered to be positive for salmonella antibodies, including 12 which were SE-positive and 39 which were ST-positive. These results indicate that dogs cause possible environmental problems as Salmonella carriers.     

In vitro bulblet formation from leaf segments of lilies,especially Lilium rubellum Baker

Bulblets were induced from 5-mm lily leaf segments, cultured on Murashige and Skoog's mineral salts medium (1962) together with α-naphthalene acetic acid (NAA) 1 mg/l, 6-benzylaminopurine (BA) 0.1 mg/l and several organic elements.Bulblet formation was affected by several factors, especially the time of excision and the origin of the explant; bulblets developed well in leaf explants excised before flowering, and mainly in those excised within 15 mm of the leaf base. However, bulblets were seldom induced in explants excised after flowering, regardless of their origin.NAA was almost indispensable for bulblet formation, but BA had a slight stimulatory effect only when NAA was also present. Sucrose concentration and light also affected bulblet regeneration.     

Behavior, physiology, performance and physical condition of layers in conventional and large furnished cages in a hot environment

The aim of the present study was to compare old layers' level of welfare and performance between conventional and large furnished cages under hot environmental conditions. At the age of 80 weeks, 104 Boris Brown layers were divided into two groups: 12 conventional cages (two hens/cage) and four large furnished cages (20 hens/cage, 240 cm wide × 62 cm deep). The room temperature was set to fluctuate between 25 and 33°C in a day. The hens' behavior, immune response, performance and physical condition were measured. Aggression and moving were more frequent in furnished than in conventional cages (P < 0.05 for both items). Egg production (P < 0.05), egg mass (P < 0.05) and feed conversion ratio (P < 0.01) were better in conventional cages than in furnished cages. No significant differences were found in the birds' immune responses or physical condition between the cage designs. In conclusion, under a hot environment, the performance of old layers in large furnished cages was lower compared with that in small conventional cages, which might be due to their greater aggressive interactions. However, it was not evident that welfare level was lower in large furnished cages compared with conventional cages. Hereafter, additional studies about large furnished cages using young layers over a long term should be performed.     

Frequencies of bovine PrP gene polymorphisms in Holstein and Japanese Black bulls in Japan

We screened for Japanese Black and Holstein bull sire samples to detect single nucleotide polymorphisms (SNPs) involving animo‐acid substitutions in the bovine prion gene in the entire coding region of the PRNP gene. Although three silent SNPs were found, we could not detect any SNP with animo‐acid substitution. We also examined the polymorphism of the octapeptide repeat number in these samples. There was no homozygous bull with repeat number 5. The frequency of heterozygous (6/5) bulls was 8% in the Japanese Black bull and 4% in the Holstein bull, respectively. The bull samples used in this study contain popular elite sires, so it appears that the polymorphisms of prion protein (PrP) are rather difficult to find in these two breeds in Japan, except for polymorphism of the octapeptide repeat number.