Capsaicin, one of the pungent principles of hot pepper, has been reported to cause a cessation of increases in body weight and fat gain induced by high-fat feeding. Especially, in body weight and feeding control, cholecystokinin (CCK) has been well known as a satiety signal and neuropeptide Y (NPY) has been described as one of the most potent orexigenic signals. This study was carried out to investigate the effect of capsaicin on CCK- and NPY- immunoreactivities (IR) in the brain of high-fat fed rats. The animals were divided into normal-fat diet (NF), high-fat diet (HF) and high-fat diet containing capsaicin (HF-CAP) groups. Mean body weight gain (MBWG) of HF group was higher than that of NF group. However, in HF-CAP group, MBWG was lower than that of HF group. CCK-IR in suprachiasmatic nucleus (SCN), paraventricular nucleus (PVN), median eminence (ME), arcuate nucleus (ARC) and amygdala was not prominent in all the groups. In cerebral cortex, CCK-IR was more reduced in HF-CAP group than in the other groups. In the HF-CAP group, NPY-IR in the hypothalamic nuclei, amygdala and cerebral cortex was more poorly found than in the NF and HF groups. It is concluded that (1) NPY-IR may react more sensitively on capsaicin than CCK-IR, (2) no rapid increase of body weight in capsaicin treated rats may result from the diminished food intake through the low expression of NPY in hypothalamus in HF-CAP group.
Renal fibrotic change, extreme accumulation of extracellular matrix (ECM) components in glomeruli and tubulointerstitum, is one of the characteristic features of ICR-derived glomerulonephritis (ICGN) mice. Decreased degradation of ECMs by matrixmetalloproteinases was demonstrated in kidneys of ICGN mice. To determine the balance between production and degradation of ECMs in kidneys of ICGN mice, we examined expression of mRNAs of ECMs in those. To demonstrate the localization of type I, III and IV collagen mRNAs in kidney sections of ICGN and control ICR mice, in situ hybridization using digoxigenin-labeled oligonucleotide antisense probes for procollagen-α1 (I), -α1 (III) and -α1 (IV) mRNAs, respectively, was performed. Negative or trace expressions of type I and III collagen mRNAs were observed in the kidneys of control mice, but stronger expressions of those were seen in glomeruli and injured renal tubules of the kidneys of ICGN mice. Moderate expression of type IV collagen mRNA was demonstrated in a part of glomeruli and renal tubules of both control and ICGN mice, and no remarkable difference was seen between them. Severe renal fibrosis, extreme accumulation of interstitial type I and III collagens is caused by increased production and decreased degradation in the kidneys of ICGN mice. Thus, the profiles of metabolism between interstitial and membranous collagens may be different in the kidneys of ICGN mice, and excessive production of interstitial collagens may be the dominant cause of renal disease in them.
An 11-month-old female ostrich (Struthio camelus) had become gradually emaciated over a 2-week period and subsequently died. Necropsy revealed white to green mold growth on the walls of caseous thickened air sac membranes and multiple white necrotic foci in the lungs and liver. Histologically, the multiple exudative, necrotic and granulomatous lesions were compatible with mycotic infection in the air sacs and lungs, and hyphae positively reacted with a monoclonal antibody (Mab-WF-AF-1) to Aspergillus fumigatus wall fractions. Multifocal hepatic necrosis was also found, and several spores were observed in the blood vessels. Fungal culture of these lesions yielded pure growth of A. fumigatus. This is an established case of fatal A. fumigatus infection in an ostrich reared in Japan.
Catechol O-methyltransferase (COMT) inactivates catecholamines and catechol-containing drugs such as L-DOPA. The common genetic polymorphism Val158Met in the human COMT gene is suspected to be associated with "persistence" or risk for schizophrenia. In this study, we attempted to identify the canine COMT gene fragment and to find a similar polymorphism and to reveal its genetic distribution among five representative canine breeds. We found that the amplified gene consisted of 663 bp nucleotides and was 84% homologous with the human COMT gene. The single nucleotide polymorphisms, guanine adenine substitution, were observed at the 39th, 216th and 482nd nucleotides. From the genotyping of the 216th polymorphism among 266 dogs by the polymerase chain reaction-restriction fragment length polymorphism method with restriction enzyme EagI, and that of the 482nd polymorphism with restriction enzyme SfcI, we found inter-breed variations of genotypes as well as of allelic frequencies for both of these polymorphic regions. These results suggest that the identified polymorphisms will be useful tools in elucidating the genetic background of canine behavioral traits.
The novel peptides based on a putative active site of defensin, an anti-bacterial peptide from the beetle Allomyrina dichotoma, were synthesized. These synthetic oligopeptides exhibited strong anti-bacterial activity in vitro, even against antibiotic-resistant pathogenic bacteria. Then, anti-bacterial activity of two newly synthesized peptides, RLYLRIGRR-NH2 (peptide A) and RLRLRIGRR-NH2 (peptide B) was also examined by macroscopic and histopathologic assessment during the course of infection in mice inoculated with antibiotic-resistant pathogenic Escherichia coli (E. coli) in vivo. Peptide B decreased the mortality of mice inoculated with antibiotic-resistant pathogenic E. coli. The results of macroscopic and histopathologic examinations revealed that peptide B could protect the mice from infection. In contrast, peptide A failed to protect mice from infection with antibiotic-resistant pathogenic E. coli. Also, modified peptides A and B produced no toxicity or side effects in mice. These results suggest that peptide B is useful for developing novel antibiotics against antibiotic-resistant pathogenic bacteria.
The existence of CD3+CD4-CD8- T cells in thymus and spleen has already been known. However, because of the presence of large amounts of thrombocytes in peripheral blood (PB), the proportion of CD3+CD4-CD8- T cells in PB has yet to be investigated. Therefore, the proportion of peripheral T cell-subsets was investigated in 6-week-old chickens. The percentage of CD3+ cells, CD4+ cells, CD8α+ cells, CD8β+, and CD3+CD4-CD8- cells was 76%, 41%, 14%, 5%, and 15%, respectively. The proportion of CD3+CD4-CD8- cells in PB increased during egg-laying periods and in chickens treated with an analog of estrogen, while it decreased with age and in response to restraint stress. All of the CD3+CD4-CD8- cells expressed TCR1, and did not have NK activity. CD3+CD4-CD8- cells represent about 60% of peripheral TCR1+ cells. These findings indicate that the proportion of CD3+CD4-CD8- cells is regulated by the endocrine and nerve systems.
To test whether glycosphingolipids (GSLs) on the intestinal mucosa of rainbow trout (Oncorhynchus mykiss) serve as a binding receptor for Vibrio anguillarum, we analyzed neutral GSLs from rainbow trout intestinal mucosa and investigated the binding of bacteria to neutral GSLs. Two kinds of neutral GSLs, designated N-1 and N-2, were identified on high-performance thin-layer chromatography (TLC) plates. In TLC immunostaining tests, V. anguillarum bound only to galactosylceramide (GalCer), lactosylceramide and N-1 having the same TLC mobility as GalCer, but neither to glucosylceramide nor to N-2. These results suggest that N-1 is GalCer (Galβ1-1Cer) and also that N-1 (GalCer) on rainbow trout intestinal mucosa act as a receptor for V. anguillarum.
To develop an experimental animal model for immune-mediated glomerulonephritis and nephrotic syndrome, nine healthy dogs were sensitized by intravenous injection with 1 μg of endotoxin and 5 mg of native bovine serum albumin. After 1 week, 120 mg of cationized bovine serum albumin was injected intravenously 5 times a week. Among nine dogs, five dogs were confirmed as having developed glomerulonephritis and nephrotic syndrome by increase of urine protein-to-creatinine ratio (>1.0), hypoalbuminemia (<1.5 g/dl), hypercholesterolemia (>240 mg/dl), and edema. This model should be useful for studying immune-mediated glomerulonephritis and nephrotic syndrome.
Rumen ciliate species and composition were surveyed on the reindeer kept in Inner Mongolia, China. As a result of survey, all the reindeer had the same 18 species of 8 genera of the ciliates in their rumen. Appearance of Entodinium parvum is the first record in this host species. Of the species detected, 15 species including Entodinium anteronucleatum, E. bicornutum, Enoploplastron confluens and Epidinium gigas which have been detected only from reindeer were common with those in the reindeer kept in the other areas, such as Alaska, Russia and Finland, indicating that every rumen ciliate faunae in reindeer has high similarity to each other, and suggesting that these hosts have been isolated from the other ruminants for long time and established characteristic ciliate fauna. The ciliate density was estimated as 1.1-2.5 × 10 6/ml, which value was fairly higher than those in domestic ruminants. Surface structures of characteristic species were observed by SEM.
Temperature dependence, heat stability and metal ions-dependent activity were examined on the Family I inorganic pyrophosphatase (PPase) recently identified from Ascaris suum. Recombinant A. suum PPase (rAsPPase) showed an optimal activity at 55°C. The rAsPPase was heat stable at 40°C in the absence of added Mg2+ and at 50°C in its presence. The enzyme required divalent metal ions for its activity. The preferences for the metal ions (5 mM concentration) were in the order: Mg2+> Co2+> Cu2+> Fe2+> Zn2+> Mn2+. On the contrary, enzyme activity was inhibited by Ca2+. These findings suggest that catalytic features of AsPPase are consistent with the Family I PPases reported from a wide range of organisms.
The relationship between the canine distemper virus (CDV) infection and apoptosis in the canine lymphoid tissues was investigated using immunostaining for single stranded DNA (ssDNA), TdT-mediated dUTP-biotin nick end-labeling (TUNEL) method, and electron microscopy. Twenty-six lymphoid tissues from 8 spontaneously CDV-infected dogs and 1 non-infected dog were used, and lesions were classified into 4 groups according to frequency of the CDV-antigen. Histologically, the degree of lymphoid depletion tended to depend on amount of CDV antigen. The numbers of ssDNA- and TUNEL-labeling cells were significantly high in the lymphoid tissues with abundant viral antigen. However, ssDNA- and TUNEL-positive lymphocytes were also frequently found even in the lymphoid tissues where there was only a small amount of CDV-antigen in sinus histiocytes. The incidence and distribution of apoptotic cells in the CDV-antigens-negative lymphoid tissues from infected dogs were equal to those from a non-infected dog. Double labeling immunostaining using a ssDNA and a CDV nucleocapsid protein (CDV-NP) antibody revealed that there were ssDNA positive but CDV-NP negative cells besides those stained doubly positive. Ultrastructurally, lymphocytes in the CDV-infected lymphoid tissues frequently had characteristic morphological features of apoptosis such as apoptotic bodies. All these results suggest that CDV leads to lymphocytic apoptosis directly or indirectly, resulting in severe lymphoid depletion and immunosuppression in acute or subacute phase of CDV infection.
A 10-year-old male mixed breed cat died after six months history of intermittent vomiting and weight loss. At necropsy, large white-colored foci were found in both kidneys, and whitish thickening of the gastric wall was present at the pyloric part of the stomach. Histopathological examination revealed that both lesions consisted of proliferation of large-sized neoplastic lymphocytes intermingled with multinucleated giant cells. Immunohistochemically, the neoplastic cells were positive for both B-cell antigen receptor complex (CD 79 α cy) and MHC class II, although multinucleated giant cells were negative. The present case was diagnosed as B-cell immunoblastic lymphoma with multinucleated giant cells.
Lactoferrin (LF) is a ubiquitous protein which exists in milk, plasma, synovial fluids, cerebrospinal fluid and other biological fluids. LF is also well known as a natural immunomodulator. Recently, we found that bovine milk-derived LF (BLF) produced μ-opioid receptor-mediated analgesia. In this study, we examined whether oral administration of BLF causes anti-nociceptive and anti-inflammatory effects, and also whether it modulates LPS-induced TNF-α and IL-10 production in rat model of rheumatoid arthritis (RA), rat adjuvant arthritis. BLF was administrated once daily, starting 3 hr before (preventive experiment) or 19 days after (therapeutic experiment) adjuvant injection. In both experiments, BLF suppressed the development of arthritis and the hyperalgesia in the adjuvant-injected paw. The single-administered BLF produced a dose-dependent analgesia, which was reversed by naloxone, in the adjuvant arthritis rats. Both repeated and single administration of BLF suppressed TNF-α production and increased IL-10 production in the LPS-stimulated adjuvant arthritis rats. These results suggest that orally administered BLF has both preventive and therapeutic effects on the development of adjuvant-induced inflammation and pain. Moreover, the immunomodulatory properties of BLF, such as down-regulation of TNF-α and up-regulation of IL-10, could be beneficial in the treatment of RA. Thus, we concluded that LF can be safely used as a natural drug for RA patients suffering from joint pain.
A dramatic rise in the number of resistant Campylobacter to quinolones has been documented in human patients and domestic animals. In this study, the mechanism of acquisition of quinolone resistance was studied by detecting point mutations in the gyrA gene of Campylobacter strains obtained from broilers and strains with in vitro-induced resistance. The minimal inhibitory concentrations (MICs) of norfloxacin (NFLX) and ofloxacin (OFLX) for the strains that had no point mutation were slightly increased from the source strain (Campylobacter jejuni ATCC 33560). The MICs of nalidixic acid (NA), NFLX, and OFLX for the strains that had the point mutation at Thr-86 were 100 or 200 μg/ml, 50 μg/ml, and 25 μg/ml, respectively. The MIC of NA for the strain that had a point mutation at Asp-90 higher than those for the strains that had the point mutation at Thr-86, but the MICs of NFLX and OFLX were relatively lower than those for the strains that had point mutation at Thr-86. These findings suggest that the degree of antimicrobial resistance against NA, NFLX, and OFLX in the in vitro-induced C. jejuni strains was associated with the location of the point mutation in gyrA. On the other hand, a point mutation in all seven resistant strains isolated from broilers was located only at Thr-86, while the MICs of the three quinolones varied in each wild strain. This suggests that another mechanism might also be involved in the acquisition of quinolone resistance in C. jejuni wild strains.
To study whether or not passive immunity of neonatal calves against Shiga toxin-producing Escherichia coli (STEC) O26, O111, and O157 was obtained by colostrum administration, serum antibodies in calves after the feeding were determined by enzyme-linked immunosorbent assay (ELISA) in comparison with antibodies in colostrum and sera from donor dams. The highest antibody titers to STEC in colostrum from dams were detected soon after parturition. The antibody titers were found to be elevated in sera of neonatal calves (4-9 hr after birth) orally administered with colostrum with high antibody titers, suggesting that passive immunity of neonatal calves to STEC infection may be obtained by feeding colostrum. These results suggest that colostrum administration to neonatal calves may play an important role in elevating serum antibodies against STEC in neonatal calves.
The prevalence of porcine endogenous retrovirus (PERV) proviral DNA among various pig breeds raised in Japan was investigated by polymerase chain reaction (PCR). Moreover, potential infection of PERV was investigated by PCR and reverse transcriptase-polymerase chain reaction (RT-PCR) in experimentally induced diabetic dogs (n=5) implanted with the diffusion chamber type bio-artificial endocrine pancreas (Bio-AEP) containing porcine pancreatic endocrine (PE) cells. No immunosupplessant was used after the transplantation. PERV gag, pol, env-A and env-B genes were detected in any pigs examined. In two of three Landrace breeds, env-C gene was absent. PERV proviral DNAs and viral RNAs were also detected from the cultured porcine PE-cells. In the peripheral blood mononuclear cells and the spleen obtained at 6, 30, 32, 36, 79 weeks of xenotransplantation in dogs, however, no evidence of microchimerism, infection and viremia were confirmed. These results suggested that the risk of PERV infection through xenotransplantation of Bio-AEP containing porcine islet cells without immunosuppressants may be quite low.
The effect of Korean red ginseng (KRG) on morphologic change and function of the liver was investigated after 70% hepatectomy in rats. The liver weight and hepatocyte proliferation of the KRG treated groups significantly increased compared to those of the control group. KRG inhibited the increase of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels, and the number and area of lipid droplets. On the basis of these results, it could be concluded that KRG accelerated the liver regeneration and ameliorated liver injury after hepatectomy in rats.
A large subcutaneous mass at the left cervical site in a 9-year-old male Siberian husky was removed surgically. Histopathologically, the mass was mainly consisted of a proliferation of spindle-shaped neoplastic cells arranging in solid sheath and partially vascular channels containing few blood cells. The tumor cells exhibited highly invasive activity to the surrounding tissues. In addition, the tumor cells were immunopositive for Factor VIII-related antigen. On the basis of these findings, the tumor was diagnosed as lymphangiosarcoma. Recurrent mass was noticed 3 weeks after surgery but completely disappeared after the doxorubicin treatment. Neither recurrence nor metastasis was observed for 9 months after the remission.
Massive lobar emphysema in the middle lobe of the right lung was observed in a dog brought to our clinic with sudden onset of tension pneumothorax, and lobectomy was performed to excise it. Pathological examination resulted in a diagnosis of congenital bronchiectasis associated with bronchial cartilage hypoplasia. Two cases of diagnosis and successful treatment of congenital lobar emphysema have been reported in dogs.
Interferon-tau (IFN-τ), produced by the embryonic trophectoderm, is a member of type I IFNs required for the establishment of pregnancy in the ruminant ungulates. Although this IFN possesses antiviral activity similar to other type I IFNs, the effectiveness of IFN-τ as an antiviral agent has not been well characterized. To investigate possible antiviral effects of ovine IFN-τ (oIFN-τ), oIFN-τ-GST fusion protein was expressed in E. coli BL21, from which the purified protein isolated possessed anti-viral activity. An apathogenic human foamy virus (hFV) was then used to establish a potential recombinant live vector consisting of oIFN-τ cDNA sense (+) or antisense (-) sequence, oIFN-τ(+)/hFV or oIFN-τ(-)/hFV, respectively. Human hematopoietic and other mammalian cell lines that had been transduced with hFV vector consisting of no oIFN-τ, oIFN-τ(+)/hFV or oIFN-τ(-)/hFV construct were cultured initially for 12 days, and three of cell lines were then maintained for up to 90 days. These cells with oIFN-τ expression directed by hFV exhibited the in vitro cytopathic effect minimally. Transduced cell lines that had been cultured for 90 days were subjected to studies on human immunodeficiency virus type-1 (HIV-1) infection, which was measured with infectivity of viral particles resulted from the GFP inserted T-cell tropic HIV SF2 or macrophage tropic HIV SF162: the number of HIV-1 positive cells was reduced by the hFV driven-intra-cellular oIFN-τ expression. Since oIFN-τ/hFV transduced cells exhibited the resistance to HIV-1 infection and/or replication, oIFN-τ could be considered as one of effective antiviral agents against HIV-1. These results suggest that the hFV genome could be an effective recombinant live vector for the expression of a targeted gene in various cell types.
Pestiviruses can be distinguished as two biotypes, cytopathogenic (cp) and noncytopathogenic (noncp), by the morphological changes that they induce during growth in cultured cells. In this study, the cp phenotype of several classical swine fever viruses (CSFV) was evaluated by the detections of the nonstructural proteins NS2-3 and NS3 using immunoprecipitation and Western blotting in different porcine cell lines. Most CSFVs that showed the exaltation of Newcastle disease virus (END) phenomenon (END+ viruses) did not induce cytopathic effect (CPE) in any cell line, and detections of NS2-3 and NS3 showed a strong signal for NS2-3 in the END+ virus-infected cells. However, clear CPE was observed in serum-free cultured cells (FS-L3 and CPK-NS) infected with viruses that induce intrinsic interference but did not show the END phenomenon (END- viruses), and signal of NS3 was strongly detected than that of NS2-3 in these cells at 72 hr after infection. As the results of the analysis of FS-L3 cells infected with ALD (END+ virus) and ALD-END- virus (END- virus) at several incubations, the signal of NS3 detected was strengthened with CPE that become evident progressively. These results suggest that CPE is associated with the accumulation of NS3, which is promoted in serum-free cell lines infected with END- viruses. Thus, indicating there is a close relationship between CPE and the quantity of NS3 produced in END- CSFV infection.