Toshiharu Hayashi

Less sensitivity for late airway inflammation in males than females in BALB/c mice

Several studies have investigated allergic airway inflammation, a T helper 2 (Th2)‐type immune response, using a mouse model of asthma. At present, however, no reports have described sex differences in the sensitivity of late airway inflammation (LAI). The LAI induced by ovalbumin in adult BALB/c mice was compared in males and females or sham‐operated males and castrated males. The males showed less severe bronchial‐bronchiolar inflammation with infiltration of eosinophils and lymphocytes and lower content of such cells in bronchoalveolar fluid than the females. Moreover, interleukin‐4 (IL‐4) mRNA expression levels in splenic cells were lower in the males than in the females. Castrated males performed like the females. Moreover, when compared with the sham‐operated males, the castrated males showed lower testosterone levels in the blood. The present results suggest that less sensitivity for LAI in the males may be because of the decreased Th2 cell responses compared with the females. Moreover the testosterone, at least in part, may be responsible for the decreased Th2 cell responses in males in vivo.

Ultrastructure of goblet-cell metaplasia from Clara cell in the allergic asthmatic airway inflammation in a mouse model of asthma in vivo

Mucus overproduction from goblet cells, a characteristic feature of the allergic asthmatic inflammation induced by ovalbumin (OVA) in mice, was examined morphologically. In OVA-untreated (normal) mice, there were no goblet cells in intrapulmonary bronchus and bronchiole. However, goblet cells with or without hyperplasia in the mucosa of inflamed bronchus–bronchiole were recognized in the allergic asthmatic mice. The non-ciliated epithelium containing electron lucent granules (mucus) showed many similarities to Clara cells, which have characteristic secretory granules and many mitochondria, except for the less-developed smooth endoplasmic reticulum seen in normal mice. Ciliated Clara cells with or without mucus were rarely recognized. In addition, mucus was found in neither ciliated nor basal epithelium. The present study suggests that goblet-cell metaplasia in the bronchus and bronchiole of inflamed mucosa may be derived, at least in part, from Clara cells.

Elimination of CD4+CD25+ T cell accelerates the development of glomerulonephritis during the preactive phase in autoimmune-prone female NZB × NZW F1 mice

The role of CD4+CD25+ T cell in glomerulonephritis (GN) development during the preactive phase was investigated in autoimmune‐prone female NZB × NZW F1 (B/WF1) mice. The administration of anti‐mouse CD25+ T‐cell monoclonal antibody (PC61.5) 3 days after birth induced the development of GN with an increase in IgG2a antinuclear antibody, productions of IL‐6 and IFN‐γ, whereas TGF‐β1 production decreased, compared to untreated control mice. The present study results suggest that CD4+CD25+ T cells may, at least in part, downregulate the development of GN during the preactive phase in B/WF1 mice.

Dysfunction of Lacrimal and Salivary Glands in Sjögren's Syndrome: Nonimmunologic Injury in Preinflammatory Phase and Mouse Model

Sjögrens syndrome (SjS) is a chronic autoimmune disorder characterized by dry eyes and dry mouth due to dacryoadenitis and sialoadenitis with SS-A/Ro and/or SS-B/La autoantibodies in genetically predisposed individuals. Destruction of lacrimal and salivary glands by autoimmune reactions may lead to clinical manifestation. However, the mechanisms behind the decreased volume of secretions in tears and saliva are complex and are not fully understood. Exocrine gland dysfunction may precede autoimmunity (acquired immunity) or represent a process independent from inflammation in the pathogenesis of SjS. The preceded functional and morphologic changes of those tissues by nonimmunologic injury before the development of inflammation at the sites of target organs have been implicated. This paper focuses on the several factors and components relating to glandular dysfunction and morphologic changes by nonimmunologic injury during the preinflammatory phase in mouse model, including the factors which link between innate immunity and adaptive immunity.

Interferon-γ plays a role in pancreatic islet-cell destruction of reovirus type 2 -induced diabetes–like syndrome in DBA/1 suckling mice

Reovirus type 2 (Reo‐2) infection in DBA/1 suckling mice causes insulitis, which leads to pancreatic islet‐cell destruction,resulting in a diabetes–like syndrome. T‐helper (Th)1 cytokines are thought to play a key role in islet inflammation in insulin‐dependent diabetes mellitus. We examined this hypothesis in the Reo‐2‐induced diabetes–like syndrome. We used reverse transcriptase polymerase chain reaction (PCR) and quantitative PCR techniques to examine mRNA expression of interferon (IFN)‐γ (Th1 type cytokine), and interleukin (IL)‐4 (Th2 type cytokine) in splenic cells. We observed that in Reo‐2 infected mice the level of IFN‐γ expression increases with the development of insulitis, whereas expression of message for IL‐4 is minimal to detectable with the immuno‐inflammatory process 10 days after infection. The treatment of monoclonal antibody (mAb) against mouse IFN‐γ during the expansion phase of insulitis (5–9 days after infection) inhibited the development of insulitis and the elevation of blood glucose concentrations in a dose dependent manner. Furthermore altered CD4+/CD8+ cell ratio compared with uninfected mice in the splenic cells by the infection was recovered to the ratio of uninfected mice by the treatment of mAb against mouse IFN‐γ, suggesting normalization of T cell balance in immune system. These results suggest that Reo‐2‐triggered autoimmune insulitis may be mediated by Th1 lymphocytes and IFN‐γ may play a role in islet inflammation leading to islet cell destruction.

Therapeutic strategies for SLE involving cytokines: mechanism-oriented therapies especially IFN-gamma targeting gene therapy.

Systemic lupus erythematosus (SLE: lupus) is a chronic complicated autoimmune disease and pathogenesis is still unclear. However, key cytokines have been recognized. Interferon (IFN)-γ and also IFNα/β are of particular importance. Depending on the concept that lupus is a helper T(Th)1 disease and that dendritic cells (DCs) determine the direction of lupus, balance shift of Th1/Th2 and immunogenic/tolerogenic DCs is reviewed for therapy. (IFN)-γ- and IFN-α/β-targeted (gene) therapies are introduced. These consist of Th1/Th2 balance shift and elimination of IFN-γ and IFN-γ-related cytokines such as (interleukin)IL-12 and IL-18. Other approaches include suppression of immunocompetent cells, normalization of abnormal T-cell function, costimulation blockade, B lymphocyte stimulator (Blys) blockade, and suppression of nephritic kidney inflammation. Moreover, balance shift of IFN-α/β and tumor necrosis factor (TNF)-α together with regulatory T(Treg) cells are briefely introduced. Clinical application will be discussed.

A serological survey of chickens, Japanese quail, pigeons, ducks and crows for antibodies to chicken anaemia virus (CAV) in Japan

The chicken has been considered as the natural host for chicken anaemia virus (CAV). To examine the prevalence of CAV in domestic and wild birds in Japan, we analyzed serum samples collected from 211 chickens, 168 Japanese quail, 105 pigeons, 113 ducks and 116 crows for the presence of antibodies to CAV by a micro-scale virus neutralization (VN) test. Nine of the 13 chicken flocks (69.2%) were found to be positive for VN antibodies to CAV. Of the 211 individual chicken serum samples, 127 (60.2%) were positive. VN antibodies were detected in 103 of the 168 (61.3%) quail samples with titres ranging from 1:20 to >/= 1:2560. Serum samples collected from quail in 1992 were found to possess a lower rate of antibody-positive sera (7.7%) and lower antibody titres (1:20 to 1:80) than those collected in 1995 (90.2% and 1:20 to >/= 1:2560, respectively). None of the pigeon, duck and crow samples neutralized CAV at a 1:20 dilution. These results indicate that quail might be one of the hosts of CAV or CAV-like virus in Japan. This is the first report of VN antibodies to CAV in a species other than chicken.

Possible Involvement of IL‐12 in Reovirus Type‐2‐Induced Diabetes in Newborn DBA/1 Mice

This study extends our previous observations that the reovirus type‐2(Reo‐2) can induce autoimmune insulitis, which may be mediated by T‐helper(Th)1‐dependent mechanisms, resulting in diabetes in newborn DBA/1 mice. In this study mRNA expression for Th1‐related cytokines including Th1 and Th2 cytokines in splenic cells was examined by reverse transcriptase polymerase chain reaction (RT–PCR) in relation to the development of insulitis. Furthermore, the effect of monoclonal antibody (MoAb) against interleukin (IL)‐12(p40) on the development of insulitis and the mRNA expression in the splenic cells was examined. The mRNA expression for IL‐12(p40), IL‐18, and interferon (IFN)‐γ, but not IL‐5, increased in the spleen in parallel with the development of insulitis. The treatment with MoAb to IL‐12(p40) reduced the insulitis with diabetes which was associated with a decrease in the mRNA expression for IL‐12(p40), IL‐18 and IFN‐γ, and an increase of IL‐4 mRNA expression in the spleen. The present study suggested that Th1‐dominant systemic immune responses, being responsible for the development of autoimmune insulitis, might be induced by IL‐12‐induced and IL‐18‐activated mechanisms.

Oncolytic Reovirus in Canine Mast Cell Tumor

The usage of reovirus has reached phase II and III clinical trials in human cancers. However, this is the first study to report the oncolytic effects of reovirus in veterinary oncology, focusing on canine mast cell tumor (MCT), the most common cutaneous tumor in dogs. As human and canine cancers share many similarities, we hypothesized that the oncolytic effects of reovirus can be exploited in canine cancers. The objective of this study was to determine the oncolytic effects of reovirus in canine MCT in vitro, in vivo and ex vivo. We demonstrated that MCT cell lines were highly susceptible to reovirus as indicated by marked cell death, high production of progeny virus and virus replication. Reovirus induced apoptosis in the canine MCT cell lines with no correlation to their Ras activation status. In vivo studies were conducted using unilateral and bilateral subcutaneous MCT xenograft models with a single intratumoral reovirus treatment and apparent reduction of tumor mass was exhibited. Furthermore, cell death was induced by reovirus in primary canine MCT samples in vitro. However, canine and murine bone marrow-derived mast cells (BMCMC) were also susceptible to reovirus. The combination of these results supports the potential value of reovirus as a therapy in canine MCT but warrants further investigation on the determinants of reovirus susceptibility.

Superficial necrolytic dermatitis associated with extrapancreatic glucagonoma in a dog.

An 11-year-old Shih Tzu presented with crusting and erythema, mainly on the abdomen and the root of the tail. Based on histopathological findings, blood examinations and necropsy findings, the condition was diagnosed as superficial necrolytic dermatitis associated with a glucagon-secreting extrapancreatic neuroendocrine tumour. Gross necropsy revealed tumour invasion into the spleen, liver, adrenal glands and mesenteric lymph nodes. Immunohistochemical analysis of the neoplastic cells revealed that the tumour was a glucagonoma, consistent with earlier findings of persistent glucagonaemia and hypoaminoacidaemia.