| Backgroud and ObjectiveAsthma is defined as a chronic inflammation of the bronchial airways and characterized by reversible airway obstruction, which increased mucus production and infiltration of the airway with eosinophils, neutrophils, mast cells and T-lymphocytes. Asthma is one of the most common chronic lung diseases in children and adults, but it is a disease that we are currently unable to prevent or cure. Worldwide epidemiological survey found that prevalence rate of children asthma is3.3%～29%and prevalence rate of adult asthma is1.2%-25.5%, it was about3hundred million people suffer from asthma in the worldwide. Global asthma death rate was about1/100000, there were about250000people die of asthma each year worldwide, which is a large proportion of young people. America asthma mortality rate increased by2.5times from1979to1998years. Our children asthma prevalence survey found that children asthma prevalence rate increased from0.91%to1.54%, up64.84%between ten years. Adult epidemiological survey showed that asthma prevalence have regional difference. According to WHO estimates that asthma disability adjusted life years (DALYs) reached15000000each year, about1%in the global burden of disease. In the world, the economy of asthma related cost far more than the total number of pulmonary tuberculosis and AIDS, it is very heavy economic burden for governments, families and patients, at the same time, it is a great impact for the patient’s daily activities and patient’s the psychological and social function.Interleukins35(IL-35) is composed of P35subunit and Ebi3subunit, it is a kind of anti-inflammatory and immune suppression of cytokine belong to the IL-12famimlies and secreted by Treg cells.Its main performance for promoting Inhibitory T cell proliferation, promoting inhibitory cytokine TNF and IL-10-β secretion and inhibiting the expression of proinflammatory cytokines IL-17. Currently, IL-35related diseases research mainly focused on infectious diseases, allergic and autoimmune diseases. Nevertheless, IL-35cytokine can promote syndrome of asthma model in mice by inhibiting numbers of Th17cells in vitro, we hypothesized that overexpression of IL-35may be adjusting the immune cells numbers of Treg cells and Th17cells in IL-35transgenic mice, which affect the inflammation process of asthma. It is likely to provide important clues for allergy treatment by studying the regulation effect of IL-35in allergic reaction.However, there is no research reports about of allergic regulating effect in IL-35transgenic mice, In this study,we used IL-35transgenic mice and asthma model challenged by OVA to study the inflammation role of IL-35in the suppression of allergic respect and regulating mechanism on the immunization.MethodsUsing molecular cloning techniques and microinjection technology to build overexpression IL-35transgenic mice, using western blotting technique to detect the expression of EBI3subunit and p35subunit of IL-35, the expression of IL-12and IL-27in IL-35transgenic mice and negative littermates (NTG) mice respectively. Analysis the distribution of EBI3subunit and p35subunit of IL-35in blood, spleen and muscle cells of IL-35transgenic mice and negative littermates mice by using western reblot. Quantitatively analyzed the percentage of Treg cells and Thl7cells blood, spleen and pulmonary washing lotion in IL-35transgenic mice and negative littermates mice by flow cytometry. In order to establish mouse model of asthma in IL-35transgenic mice and negative littermates mice, we continuously intraperitoneally inoculated with1%OVA and aluminium adjuvant once a week for3weeks, and the fourth week in a row with four days challeng by5%OVA atomization. Detected airway hyperresponsiveness of mouse model of asthma by the pulmonary function testing, the comparativly analysize asthma syndrome of IL-35transgenic mice and nontransgenic mice. Pathology observed the inflammatory cell exudation and bronchial mucus infiltration of lungs in OVA-challenged IL-35transgenic mice and OVA-challenged negative littermates mice. Comparativly analysize the percentage of Treg cells and Thl7cells of pulmonary washing lotion in mouse model of IL-35transgenic mice and negative littermates mice by flow cytometry.ResultsThe results of the experiments in vivo supported that:(1) Established transgenic over-expression of IL-35, IL-35expression quantity higher than that of IL-12and IL-27was proved by western blotting, the percentage of Treg cells increased in peripheral blood and spleen in overexpression IL-35transgenic mice.(2) Established mouse model of asthma. It confirmed that transgenic overexpression of IL-35can inhibit airway hyperresponsiveness in mice by using this model. Reduced the inflammatory cells infiltration and mucus secretion in the surrounding of bronchus, this explain that IL-35can inhibit mouse asthma symptoms in vivo. IL-35can activate the Treg cells in vivo, inhibit the secrete of IL-17by inhibiting the numbers of Th17cells, the signaling pathway IL-35inhibit mouse asthma is one of the important pathways by inhibiting proinflammatory IL-17cytokine.ConclusionsTaken togerther, our results indicated that:(1) IL-35has obvious inhibitory effect in mouse asthma.It affect by promoting Treg cells and inhibiting proliferation and activation of Th17cells.(2) IL-35could inhibit inflammation and mucus secretion in mouse astnma by promoting Treg cell proliferation and inhibiting the secretion of IL17cytokine. In conclusion, overexpression of IL-35can significantly improve the mouse asthma symptoms, it play the role in mainly inhibiting production of proinflammatory cytokine. Necrotizing myositis caused restrictive hypoventilation in human enterovirus71infected-mouse modelBackgroud and ObjectiveEV71infection is associated with a high prevalence of Hand, foot and mouth disease (HFMD) in children and occasionally causes lethal complications. Most of the infections were self-limited, while the complications including aseptic meningitis, encephalitis or poliomyelitis-like acute flaccid paralysis, and neurological originated pulmonary edema or hemorrhage were the main causes of lethal symptoms, the pathogenesis of which were remained to be clarified. Hence we presumed that caused hypoventilation caused by Necrotizing inflammation is one of the causes of death in mice.MethodsEV71virus inoculated two weeks old ICR mice by intraperitoneal (i.p.) injection, then we observe situation of the mice body generally, the weight of the mice bady and survival situation of mice, detecting infected viral load of tissues and organs by fluorescence quantitative PCR (qRT-PCR), pathological and immunohistochemical analysis inflammation and viral load of skeletal muscle in infected mice, we can analyze inspiratory time, respiratory rate, tidal volume and respiratory total volume of virus infection group of mice and control group of mice by pulmonary function detector.Results In present study, two-week-old ICR mouse were infected with a mouse-adapted EV71strain, and then the infected-mice demonstrated progressive paralysis and died within12days post infection. EV71mainly replicates in skeletal muscle tissues and caused severe necrotizing myositis, whereas the lesion in CNS and other tissues was not observed.ConclusionsThe necrotizing myositis of respiratory-related muscles caused severe restrictive hypoventilation and subsequently hypoxic, which was supposed as one reason of death for the EV71-infected mouse, suggesting that necrotic myositis caused may be a reason of paralysis and death beside of CNS injury in EV71-infected mice. Human SCARB2enhanced the infection of human enterovirus71on transgenic miceBackgroud and ObjectiveHuman enterovirus71(EV71) is a RNA virus of the micro RNA virus enterovirus, it can cause hand, foot and mouth disease in children under6years old.But some of EV71infections will spontaneous recovery, and EV71infection accompany serious complications occasionally, including the brain stem, aseptic meningitis, encephalitis, pulmonary edema and pulmonary hemorrhage, heart failure and acute flaccid paralysis and even death.the virus receptors play a pole role in the early days ofthe virus infection, mainly include host range and tropism of tissue.Human SCARB2was identified as one of the functional receptors of EV71in vitro, so establish more higher infection ability transgenic mice by establishment of hunman SCARB2transgenic mice in present study.MethodsThe transgenic vector was constructed by inserting the human SCARB2gene under the CMV promoter and then were subjected to establish transgenic mice by microinjection. the genotype of transgenic line was identified by PCR and the expression level of target protein was detected by Western blot. Viral load in the tissues of transgenic mice was detected by immunohistochemical staining and quantitative real-time PCR.Results One line of transgenic mice in C57BL/6J background with high levels of SCARB2expression in skeletal muscle and brain was identified. Upon infection with EV71, the virus burden of4to5times in muscle and brain of transgenic mice were significantly higher than that of wild type mice.ConclusionhSCARB2is a functional receptor of EV71in vivo, as expression of it could promote the infection of EV71on transgenic mice. |
PDF Full Text Request