Atopic NC/Nga mice as a model for allergic asthma: severe allergic responses by single intranasal challenge with protein antigen

Since certain characters of allergic asthma are common with other allergic disorders like atopic dermatitis, the possible relationship in etiology is expected. Herein, we investigated whether NC/Nga mice, an inherent animal model for human atopic dermatitis, are inclined to allergic asthma. A single intranasal challenge of NC/Nga mice immunized with ovalbumin (OVA) resulted in an increase in plasma levels of OVA-specific IgE, and typical pathological aspects of allergic asthma characterized by infiltration of numerous eosinophils, mucus hyper production of bronchial epithelial cells. Moreover, airway hyperresponsiveness to inhaled acetylcholine and marked enhancement of airway resistance after the challenge were observed as compared to control BALB/c mice. Delayed expression of mRNA of eosinophil active chemokines, interleukin-5, eotaxin, macrophage inflammatory protein-1alpha in concert with eosinophilia was determined in the lung of NC/Nga mice. These results suggest that asthmatic responses developed in NC/Nga mice challenged with OVA are very similar to human allergic asthma, and that NC/Nga mice are a useful model to elucidate various aspects of allergic asthma.     

A lysosomal storage disorder in the BALB/c mouse: bone marrow transplantation

The morphological and biochemical consequences of transplanting affected bone marrow from donor BALB/c mice with a lysosomal storage disorder (BALB/c LSD) into normal recipient mice were studied. Bone marrow was removed from normal BALB/c and BALB/c LSD mice and transfused into normal BALB/c recipient mice four hours after the mice received 850 rads of irradiation. Tissues of the recipient mice were examined 240 days later. This study revealed that the defective cells that constituted the visceral lesions of BALB/c LSD could be transplanted to normal BALB/c mice by the use of bone marrow from affected BALB/c LSD homozygote; that the defective cells of BALB/c LSD proliferated and disseminated throughout the mononuclear phagocytic system of the recipient; that there were increases in cholesterol, sphingolipids, and cystine with decreases in sphingomyelinase and glucocerebrosidase activity in tissues of the recipients; and that the recipients survived substantially longer than BALB/c LSD homozygotes and their lifespan was compromised mainly by the secondary effects of irradiation. These lesions, although not as extensive as in homozygous BALB/c LSD, paralleled the lesions which develop in BALB/c LSD. Since the recipient mice were not compromised by the short life span (70 days) of the BALB/c LSD mice, they may be used to study the long-term chronic effects of these metabolic lesions.     

A role for balance of interferon-gamma and interleukin-4 production in protective immunity against Neospora caninum infection

A suitable balance in the production of Th1/Th2-type cytokines has a crucial role in the control of microbial infections. We investigated cytokine production patterns and effects during Neospora caninum infection, based on two mouse models and an in vitro system. In the acute infection of N. caninum, BALB/c-background IFN-gamma-deficient mice that were sensitive to the N. caninum infection showed high levels of IL-10 production, whereas significant levels of interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) production were observed in resistant wild type mice. BALB/c mice vaccinated with recombinant vaccinia virus expressing N. caninum surface protein NcSRS2 resisted parasite spread throughout the body, low levels of IFN-gamma production and high levels of IL-4 production were observed compared to unvaccinated animals. The treatment of N. caninum-infected cells with IFN-gamma or IL-10 decreased the host-cell viability in an in vitro system using mouse macrophage J774A.1 cells. On the other hand, IL-4, but not IL-10 administration, increased the viability of N. caninum-infected and IFN-gamma-treated cells. In the light of the balance of Th1/Th2-type cytokine production, an IFN-gamma/IL-4 balance may have a crucial role for the control of cellular responses against the parasite invasion.     

Histopathological studies of visceralized Leishmania (Leishmania) amazonensis in mice experimentally infected

BALB/c, C57BL/6, and DBA/2 mice were subcutaneously infected in the left footpad by injecting 10(4) Leishmania (Leishmania) amazonensis amastigotes. Mice were sacrificed 20, 30, 40, 60 and 90 days post-infection. Fragments of liver, kidney, spleen, skin, and draining lymph node were collected for histological examination. Light microscopy showed that at 20 days after infection BALB/c mice presented discrete inflammatory infiltrates in the skin made up of eosinophils, lymphocytes, and rare parasitized macrophages. Ninety days post-infection, the dermis showed necrotic tissue, large numbers of mononuclear cells and vacuolated macrophages filled with amastigotes. Forty days post-infection, the draining lymph nodes showed hyperplastic germinal centers, increase of high endothelial venules and apoptosis in germinal center cells. After the first 3 months post-infection, the involvement of spleen, kidney and liver was discreet, being characterized by multifocal inflammatory infiltrates. Eight months after infection, the animals presented metastatic lesions in the contralateral footpad and nose. In deep dermis, there was remarkable proliferation of fibroblasts associated with collagen fibers. The liver showed multifocal granulomas and mononuclear infiltrate around the blood vessels, but no parasites were observed, except in one animal. In some mice there were immature cells of the hematopoietic lineage. Both BALB/c and C57BL/6 mice presented osteonecrosis, which is characterized by pycnotic osteocytes and empty lacunae at the point of inoculation and subsequently, replacement of this tissue by fibrous connective tissue and colonization of the bone marrow. A diffuse inflammatory process composed of mononuclear cells and rare parasites were seen in the kidneys. In one mouse, bone marrow cells were observed in the renal medulla along with where free amastigotes. DBA/2 mice developed a mild infection and they did not visceralize. In conclusion, our data demonstrates that in susceptible mice L. (L.) amazonensis, a causative agent of tegumentary leishmaniasis, causes pathological changes similar to those produced by Leishmania (L.) infantum in both humans and canids.     

Epsilon-polylysine microparticle adjuvant drives cytokine production to Th1 profile

Epsilon-polylysine micro particles (SGEPL) and polyethyleneimine micro particles (SGPEI) were developed by the addition of a hydrophobic group and the immunological characterization of these micro particles and aluminum hydroxide (ALUM) was investigated. BALB/c mice were injected intraperitoneally with ovalbumin (OVA) as an antigen and SGEPL, SGPEI or ALUM as an adjuvant. The results showed that the mice injected with SGEPL produced a significant portion of anti-OVA antibody subclass IgG2a in the sera and suppressed interleukin (IL)-4 and IL-5, but enhanced IL-12 and interferon-gamma (IFN-gamma) from the spleen cells. Similar results relating to cytokines were also obtained, even without OVA. Direct stimulation with SGEPL to na?ve BALB/c mouse spleen cells induced IL-12 and IFN-gamma. Both spleen and purified B cells produced IgG1 and IgE after stimulation with IL-4 and the anti-CD40 monoclonal antibody. With the addition of SGEPL, the IgE production from the cells was suppressed as a result of enhanced IFN-gamma production. Furthermore, IgE production was also suppressed in the purified B cells without the influence of IFN-gamma or IL-12. Thus, we suggest SGEPL drives cytokine production to Th1 profile. It will be a novel promising adjuvant based on this viewpoint.     

The radioprotective effects of the hexane and ethyl acetate extracts of Callophyllis japonica in mice that undergo whole body irradiation

The radioprotective activity of extracts from the red seaweed Callophyllis (C.) japonica was investigated in mice that underwent whole-body exposure to gamma radiation. A methanol extract of C. japonica and its fractions [hexane, ethyl acetate (EtOAc), butanol and the remaining H₂O] were used. Each fraction (100 mg/kg body weight) was administered intraperitoneally (i.p.) 2 times into the BALB/c mice, once at 1 and once at 24 h before exposure to 9 Gray (Gy) of gamma radiation. Pre-irradiation administration of the hexane and EtOAc fractions saved the mice, with their survival rates being greater than 80% at 30 days post-irradiation; the mice that were pretreated with the other fractions showed survival rates lower than 20% over the same time period. To examine the effect of each C. japonica fraction on the survival of intestinal and bone marrow stem cells, the number of intestinal crypts and bone marrow cells in the gamma-irradiated mice were examined. Pre-treatment of mice (i.p., 100 mg/kg body weight at 1 and 24 h before irradiation) with the hexane or EtOAc fraction prior to 6-Gy irradiation significantly protected the number of jejunal crypts and bone marrow cells at 9 days after irradiation. These findings suggest that certain extracts from C. japonica, when they are administered prior to irradiation, play an important role in the survival of irradiated mice, and this is possibly due to the extracts protecting the hematopoietic cells and intestinal stem cells against gamma irradiation.     

Chronic eosinophilic leukemia in a cat: cytochemical and immunophenotypical features

A 3-year-old, male, domestic shorthaired cat was presented with a 3-day history of anorexia and depression. The cat was moderately dehydrated, had pale, slightly icteric, mucous membranes, oral ulcerations, and mild hepatosplenomegaly. A feline leukemia virus (FeLV) antigen test was positive. CBC results obtained at initial presentation included severe normocytic, normochromic, nonregenerative anemia, severe thrombocytopenia, and marked leukocytosis (>100,000/microL) with 77% eosinophils. After 15 days of treatment with prednisone and doxycycline, the cat had persistent severe nonregenerative anemia (HCT 3.4%), thrombocytopenia (28,000/microL), and extreme eosinophilia (total eosinophils, 123.1 x 10(3)/microL; segmented 103.0 x 10(3)/microL; immature 20.1 X 10(3)/microL). Cytologic examination of aspirates from bone marrow, liver, lymph nodes, and spleen revealed a predominance of mature and immature eosinophils, many with dysplastic changes. The M:E ratio was 96.4. On histopathologic examination, multiple organs were infiltrated by eosinophilic granulocytes. Neoplastic cells in blood and bone marrow stained positive for alkaline phosphatase and were negative for myeloperoxidase, chloroacetate esterase, and alpha-naphthyl acetate esterase. On flow cytometric analysis of peripheral blood, the neoplastic cells were positive for CD11b and CD14. These findings were consistent with chronic eosinophilic leukemia. To our knowledge, this is the first report of chronic eosinophilic leukemia in a cat associated with naturally acquired FeLV infection, in which flow cytometry was used to characterize the neoplastic cells.     

表达猪附红细胞体ENO基因的质粒DNA的构建及免疫效果研究

试验旨在构建表达猪附红细胞体ENO基因的质粒DNA,并测定其免疫效果。将猪附红细胞体ENO基因克隆到PVAX1真核表达载体上,然后转染到Vero细胞中进行表达并测定其免疫效果。将18只BALB/c小鼠（雌雄各半）随机分为3组（PVAX1-ENO质粒DNA免疫组、PVAX1空载体对照组及PBS对照组）,每组6只,每组小鼠对应免疫接种PVAX1-ENO质粒DNA、PVAX1空质粒和PBS。分离血清并通过ELISA方法测定血清中ENO蛋白抗体效价、IgG₁和IgG_2a抗体水平及IFN-γ和IL-4细胞因子水平。三免2周后每组选取3只小鼠检测CD4⁺和CD8⁺含量。结果显示,本试验成功构建了PVAX1-ENO质粒DNA,经PCR和酶切鉴定正确,并能在Vero细胞中成功表达。PVAX1-ENO质粒DNA免疫组ENO蛋白抗体水平、IgG₁和IgG_2a抗体水平、IFN-γ和IL-4细胞因子水平及CD4⁺和CD8⁺含量均显著高于PVAX1空载体对照组及PBS对照组（P<0.05）。结果表明,PVAX1-ENO质粒DNA可显著提高BALB/c小鼠的体液免疫水平,并在一定程度上刺激BALB/c小鼠细胞免疫。     

迷迭香酸对哮喘小鼠氧化性肺损伤的保护作用

为评价迷迭香酸对哮喘小鼠模型氧化性肺损伤的保护作用,本研究用卵清蛋白（OVA）致敏、激发雌性BALB/c小鼠建立哮喘模型,并用OVA和H₂O₂联合激发小鼠作为氧化肺损伤阳性对照模型。在最后一次滴鼻激发24 h后,取支气管肺泡灌洗液（BALF）进行细胞计数并测定活性氧（ROS）、超氧化物歧化酶（SOD）和谷胱甘肽过氧化物酶（GSH-Px）水平,取左侧肺脏固定做HE染色。结果显示,迷迭香酸可明显减少BALF中细胞总数和嗜酸性粒细胞数目,显著抑制肺组织和BALF中ROS的产生,升高SOD和GSH-Px水平,改善肺组织病理变化。本试验结果表明,迷迭香酸对氧化肺损伤起到明显的保护作用。     

Protective Effects of Rosmarinic Acid on Oxidative Lung Damage in Asthmatic Mice

In order to evaluate the protective effect of rosmarinic acid on oxidative lung damage in asthmatic mice, in this study, the asthma model was established by sensitizing and challenging female BALB/c mice with ovalbumin (OVA), and some mice were challenged by OVA and H₂O₂ as a positive control of oxidative lung damage. The BALF, lung tissues of mice were harvested 24 h after the final OVA challenge and the number of total cells and eosinophils were measured, ROS,SOD and GSH-Px levels were detected, and pathological changes of lung tissue were observed using HE staining. The results showed that rosmarinic acid significantly reduced the number of total cells and eosinophils in BALF, significantly inhibited the production of ROS in lung tissue and BALF, increased the levels of SOD and GSH-Px, and ameliorated pathological changes of lung tissue. The results suggested that rosmarinic acid could protect oxidative lung damage in asthmatic mice.     

Hypereosinophilia associated with transitional cell carcinoma in a cat.

A 14-year-old spayed female domestic shorthair cat was evaluated because of an abdominal mass and eosinophilia. Widely disseminated, transitional cell carcinoma of the urinary bladder was diagnosed histologically. To further characterize the eosinophilia, eosinophils were separated from other leukocytes and cultured in vitro. Harvested cells were evaluated for density and for in vitro survivability. Results of these tests, hyperplasia of bone marrow eosinophil precursors, and lack of tumor tissue eosinophilic infiltrates suggested that an eosinophilopoietic stimulus of undetermined origin was likely the cause of this cat's hypereosinophilia.     

Idiopathic hypereosinophilic syndrome in 3 Rottweilers

Three Rottweilers with marked peripheral eosinophilia and infiltration of the liver, spleen, lungs, and bone marrow with eosinophils were diagnosed with idiopathic hypereosinophilic syndrome (IHES). Mean serum immunoglobulin E concentrations were markedly high. On cytogenetic analysis, no evidence of karyotypic abnormalities was found in bone marrow aspirates. Despite an extensive search, no underlying cause for the eosinophilia could be identified. In this study, cytogenetic analysis and measurement of serum IgE concentrations were used to differentiate IHES and eosinophilic leukemia.     

Eosinophilic leukaemoid reaction in a dog

A haematological disorder in a dog characterised by a massive leucocytosis, mainly composed of eosinophils and their precursors is reported here. The normal composition of cells in the bone marrow was displaced in favour of the eosinophils and their precursors. No apparent cause for the pronounced eosinophilia could be determined by clinical, haematological, clinical-chemical, radiological or pathological examinations. A diagnosis of eosinophilic leukaemoid reaction was suggested as the criteria for the diagnosis of eosinophilic leukaemia in the dog but this was not firmly established.     

Immune responses of immunocompetent and immunocompromised mice experimentally infected with Bartonella henselae

The aim of this study is to understand host immune responses in immunocompetent and immunocompromised mice against Bartonella henselae infection. BALB/c and nude (BALB/c nu/nu) mice were inoculated intraperitoneally with 10(8) colony forming units of B. henselae (Houston-1 strain). Blood, brain, liver, spleen, kidney and bone marrow samples were collected 0, 3, 7, 14, 21 and 28 days after infection and submitted to bacteriological, serological and genetical examinations. B. henselae was isolated only from the liver 3 days after infection. DNA of the inoculums was detected by polymerase chain reaction from blood, liver, and spleen samples collected from BALB/c and blood from nude mice 3 and 7 days after infection. No bacterial DNA was detected from both BALB/c and nude mice thereafter during 4 weeks observation periods. These results indicate that the T-cell may not participate in the effective elimination of the organisms from mice. In addition, western blot analysis revealed that the antigens of 27.3- and 31.5-kDa reacted with IgM antibodies from the blood of BALB/c and nude mice after 3 days of infection, suggesting that these antigens were recognized by thymus-independent mechanism. Furthermore the antigens were detected from the culture-supernatants of B. henselae, indicating that these antigens were secreted from the organisms.     

Immunization with recombinant surface antigens p26 with Freund's adjuvants against Babesia rodhaini infection

The surface proteins of Babesia rodhaini have previously been shown to induce a high degree of protective immunity. In the present study, one of those proteins, B. rodhaini antigen p26 was expressed in Escherichia coli and in insect cells infected with a recombinant baculovirus. These proteins were recognized by immune serum from a drug-cured BALB/c mouse. While BALB/c mice immunized with both recombinant antigens and Freund's adjuvants showed 40-100% survival rate against challenge infection with B. rodhaini, saponin failed to induce protection, although significant levels of B. rodhaini-specific antibodies were produced in both immunized mice (1:1,000-2,000 by indirect immunofluorescent antibody test). The immunization of IFN-gamma-deficient mice with the recombinant proteins was not protective against B. rodhaini infection, indicating that IFN-gamma is one of the important factors for the survival against lethal B. rodhaini infection.     

Immune response and protective efficacy against homologous challenge in BALB/c mice vaccinated with DNA vaccine encoding Toxoplasma gondii actin depolymerizing factor gene

A DNA vaccine (pVAX1-TgADF) encoding Toxoplasma gondii actin depolymerizing factor (ADF) gene was constructed and the immune response and protective efficacy of this vaccine against homologous challenge in BALB/c mice were evaluated. High titers of specific antibody and increases in the percentage of CD4(+) and CD8(+) T lymphocyte cells were observed from BALB/c mice vaccinated with pVAX1-TgADF (P<0.05), when PBS group was used as control. The survival time of BALB/c mice in pVAX1-TgADF group was longer than those in control groups. The numbers of brain cysts in the experimental BALB/c mice immunized with pVAX1-TgADF reduced significantly compared with those in PBS group (P<0.05), and the rate of reduction could reach to around 42.8%. These results suggested that the DNA vaccine pVAX1-TgADF could generate specific humoral and cellular immune responses, prolong survival times, and reduce brain cysts load against T. gondii infection in BALB/c mice.     

Hypereosinophilic syndrome in cats: a report of three cases.

The clinical, clinicopathological and pathological findings in three cats with hypereosinophilic syndrome are described. The cats chosen for the study had marked eosinophilia and evidence of tissue infiltration by eosinophils. Necropsies were performed on two cats, biopsy and blood samples were provided for the third cat. At necropsy, there was diffuse reddening of femoral bone marrow with ulceration and thickening of the duodenum. The livers had an enhanced lobular pattern with multiple, white, 1-3 mm nodules throughout the parenchyma. One cat had splenomegaly and the other had several enlarged, white, firm lymph nodes. Histopathologically, there was eosinophil infiltration of intestine, lymph nodes, liver, spleen, adrenal medulla and beneath the endocardium. Ultrastructurally, the eosinophils from lymph node and bone marrow of cat II were morphologically normal. The rigid criteria for eosinophilic leukemia were not fulfilled by these cases and the etiology of the eosinophilia in each case is not known. Possible pathogenic mechanisms are discussed.     

Pretreatment of macrophages with the combination of IFN-gamma and IL-12 induces resistance to Leishmania major at the early phase of infection

Interferon (IFN)-gamma is essential but not sufficient to control leishmaniasis. It is known that IFN-gamma is one of the major macrophage-activating cytokines, and the activated macrophages are a principal source of interleukin (IL)-12, which induces autocrine macrophage activation. In this study, the combined effect of IFN-gamma and IL-12 on the susceptibility of macrophages to Leishmania major infection was evaluated. Macrophages pretreated with IFN-gamma and/or IL-12 were infected with the parasites. Four hr post-infection (p.i.), the levels of infection and parasite load in the macrophages treated with the combination of IFN-gamma and IL-12 (IFN-gamma/IL-12) were significantly lower than those in the nontreated cells. However, the macrophages treated with either IFN-gamma or IL-12 did not show resistance to L. major infection. In addition, 72 hr p.i., the IFN-gamma/IL-12-treated and IFN-gamma-treated macrophages showed significantly lower levels of infection and parasite load than the nontreated cells, and higher levels of resistance was observed in the IFN-gamma/IL-12-treated macrophages than in the IFN-gamma-treated macrophages. Although IFN-gamma/IL-12 treatment of macrophages prior to the infection led to the induction of resistance, as described above, this resistance was not induced when these cytokines and the parasites were added simultaneously to the macrophage culture. These results suggest that IFN-gamma/IL-12 treatment prior to the infection restricts the early phase of the infection.     

C5a/C5aR信号在微小隐孢子虫感染中的免疫调节作用研究

旨在探究C5a/C5aR信号在微小隐孢子虫感染过程中对宿主CD4⁺ T细胞免疫反应的调控作用。本研究以微小隐孢子虫（Cryptosporidium parvum）为研究对象,以BALB/c乳鼠和C5aR抑制BALB/c乳鼠为感染模型,应用实时荧光定量PCR和免疫组织化学技术检测了C.parvum感染前后乳鼠回肠组织中C5aR的表达变化,利用实时荧光定量PCR检测了隐孢子虫HSP70基因和CD4⁺ T细胞亚群（Th1、Th2、Th17细胞和Treg细胞）主效应细胞因子（IFN-γ、IL-4、IL-17和TGF-β）的转录变化,并通过病理组织切片观察乳鼠回肠黏膜的损伤情况。结果显示：与对照组乳鼠相比,C.parvum感染可以引起乳鼠回肠组织中C5aR的mRNA和蛋白表达水平显著上调（P<0.05）,以及IFN-γ表达水平显著上调（P<0.05）;与C.parvum感染组乳鼠相比,C5aR抑制剂处理可引起C.parvum感染乳鼠回肠组织中Th1细胞、Th2细胞和Treg细胞的主效应细胞因子IFN-γ、IL-4和TGF-β显著下调表达（P<0.05）,以及Th17细胞主效应细胞因子IL-17显著上调表达（P<0.05）。病理学观察发现,抑制C5aR能显著改善C.parvum感染引起的乳鼠回肠组织的绒毛直径和黏膜厚度变化（P<0.05）,但不能改善绒毛长度、绒毛长度与隐窝深度比值。隐孢子虫HSP70基因的mRNA水平检测发现,抑制C5aR能显著影响C.parvum在回肠组织中的增殖（P<0.05）。C5a/C5aR信号可能通过动态调节CD4⁺ T细胞亚群主效应细胞因子的表达来参与宿主与隐孢子虫相互作用的过程,为深入理解隐孢子虫与宿主的互作机制提供了参考。