Infection of a human respiratory epithelial cell line with rhinovirus. Induction of cytokine release and modulation of susceptibility to infection by cytokine exposure.

Rhinovirus infections cause over one third of all colds and are a contributing factor to exacerbations of asthma. To gain insights into the early biochemical events that occur in infected epithelial cells, we develop, for the first time, a model in which a pure respiratory epithelial cell population can be routinely infected by rhinovirus. Viral infection was confirmed by demonstrating that viral titers of supernatants and lysates from infected cell increased with time and by PCR. Infection by rhinovirus 14 was inhibited by homotypic antiserum and by antibodies to intercellular adhesion molecule-1 (ICAM-1), the receptor for this virus. Susceptibility of epithelial cells to infection by rhinovirus 14 (but not rhinovirus 2, an ICAM-1 independent strain) can be increased by preexposure of cells to TNF alpha, whereas IFN gamma reduces susceptibility to infection by both rhinovirus strains. Rhinovirus infection per se does not markedly alter ICAM-1 expression on epithelial cells. Finally, we demonstrate that rhinovirus infection induced increased production of IL-8, IL-6, and GM-CSF from epithelial cells. Production of IL-8 correlated with viral replication during the first 24 h after infection. This model should provide useful insights into the pathogenesis of rhinovirus infections.     

Asthmatic bronchial epithelial cells have a deficient innate immune response to infection with rhinovirus

Rhinoviruses are the major trigger of acute asthma exacerbations and asthmatic subjects are more susceptible to these infections. To investigate the underlying mechanisms of this increased susceptibility, we examined virus replication and innate responses to rhinovirus (RV)-16 infection of primary bronchial epithelial cells from asthmatic and healthy control subjects.Viral RNA expression and late virus release into supernatant was increased 50- and 7-fold, respectively in asthmatic cells compared with healthy controls. Virus infection induced late cell lysis in asthmatic cells but not in normal cells. Examination of the early cellular response to infection revealed impairment of virus induced caspase 3/7 activity and of apoptotic responses in the asthmatic cultures. Inhibition of apoptosis in normal cultures resulted in enhanced viral yield, comparable to that seen in infected asthmatic cultures. Examination of early innate immune responses revealed profound impairment of virus-induced interferon-beta mRNA expression in asthmatic cultures and they produced >2.5 times less interferon-beta protein. In infected asthmatic cells, exogenous interferon-beta induced apoptosis and reduced virus replication, demonstrating a causal link between deficient interferon-beta, impaired apoptosis and increased virus replication. These data suggest a novel use for type I interferons in the treatment or prevention of virus-induced asthma exacerbations.     

Levofloxacin inhibits rhinovirus infection in primary cultures of human tracheal epithelial cells

Respiratory virus infections, including infections with rhinoviruses (RVs), are related to exacerbations of chronic obstructive pulmonary disease (COPD). A new quinolone antibiotic, levofloxacin (LVFX), has been used to treat bacterial infections that cause COPD exacerbations as well as bacterial infections that are secondary to viral infection in COPD patients. However, the inhibitory effects of LVFX on RV infection and RV infection-induced airway inflammation have not been studied. We examined the effects of LVFX on type 14 rhinovirus (RV14) (a major human RV) infection of human tracheal epithelial cells pretreated with LVFX. LVFX pretreatment reduced the RV14 titer, the level of cytokines in the supernatant, the amount of RV14 RNA in the cells after RV14 infection, and the cells' susceptibility to RV14 infection. LVFX pretreatment decreased the mRNA level of intercellular adhesion molecule 1 (ICAM-1), a receptor for RV14, in the cells and the concentration of the soluble form of ICAM-1 in the supernatant before RV14 infection. LVFX pretreatment also decreased the number and the fluorescence intensity of the acidic endosomes from which RV14 RNA enters the cytoplasm. LVFX pretreatment inhibited the activation of nuclear factor κB proteins, including p50 and p65, in nuclear extracts. LVFX pretreatment did not reduce the titers of RV2 (a minor human RV) but reduced the titers of RV15 (a major human RV). These results suggest that LVFX inhibits major-group rhinovirus infections in part by reducing ICAM-1 expression levels and the number of acidic endosomes. LVFX may also modulate airway inflammation in rhinoviral infections.     

ABCs of Asthma

Asthma results from chronic airway inflammation involving a diversity of activated cells including mast cells, eosinophils, T-lymphocytes, neutrophils, macrophages, and epithelial cells. These cells release proinflammatory cytokine mediators that augment and regulate airway inflammation, leading to airway hyperresponsiveness responsible for the chronic asthma symptoms of dyspnea, wheezing, and chest tightness. It is hypothesized, but unproven, that inflammatory effects can lead to irreversible structural and functional airway changes. Early intervention with anti-inflammatory agents mitigates inflammatory changes, reverses airway obstruction, and may possibly prevent progression of airway remodeling. Current asthma guidelines recommend that initial management should be based on pretreatment assessments of asthma severity as determined by measures of clinical and spirometric impairment in individual patients; subsequent adjustments of pharmacotherapy and avoidance recommendations should be performed at regular follow-up visits and guided by frequent assessments of asthma control. Physicians and providers should continually educate asthmatic patients about proper use of asthma controller medications, avoidance of asthma triggers, and self-management of asthma exacerbations.     

A common cold virus, rhinovirus 16, potentiates airway inflammation after segmental antigen bronchoprovocation in allergic subjects.

Many patients with asthma have increased wheezing with colds. We hypothesized that rhinovirus colds might increase asthma by augmenting airway allergic responses (histamine release and eosinophil influx) after antigen challenge. Seven allergic rhinitis patients and five normal volunteers were infected with rhinovirus type 16 (RV16) and evaluated by segmental bronchoprovocation and bronchoalveolar lavage. Segmental challenge with saline and antigen was performed 1 mo before infection, during the acute infection, and 1 mo after infection. Lavage was performed immediately and 48 h after antigen challenge. Data were analyzed by two-way analysis of variance, and a P value of < or = 0.05 was considered to be significant. All volunteers inoculated with RV16 developed an acute respiratory infection. BAL fluid obtained from allergic rhinitis subjects during the acute viral infection, and 1 mo after infection, showed the following significant RV16-associated changes after antigen challenge: (a) an enhanced release of histamine immediately after local antigen challenge; (b) persistent histamine leak 48 h afterwards; and (c) a greater recruitment of eosinophils to the airway 48 h after challenge. These changes were not seen in non-allergic volunteers infected with RV16 and challenged with antigen, nor in allergic volunteers repetitively challenged with antigen but not infected with RV16, nor in RV16 infected allergic volunteers sham challenged with saline. We conclude that rhinovirus upper respiratory infection significantly augments immediate and late allergic responses in the airways of allergic individuals after local antigen challenge. These data suggest that one mechanism of increased asthma during a cold is an accentuation of allergic responses in the airway which may then contribute to bronchial inflammation.     

2019－2021年长春市儿童急性呼吸道感染病例中鼻病毒感染状况及其分子分型研究

  目的   为了解2019年6月至2021年5月吉林省长春市急性呼吸道感染住院儿童病例中鼻病毒感染状况并对鉴定出的鼻病毒进行分子分型研究。  方法   采用实时荧光聚合酶链式反应技术对采集的1 251份咽拭子标本进行常见8种呼吸道病毒的检测。 针对鉴定结果为阳性的鼻病毒标本,使用巢式反转录聚合酶链式反应扩增VP1区并进行序列测定和分析。   结果   61.87%（774/1 251）的标本至少检出有1种呼吸道病毒,其中呼吸道合胞病毒和鼻病毒检出率较高,分别为19.18%（240/1 251）和18.63%（233/1 251）。 男女性间鼻病毒检出率比较差异无统计学意义,检出年龄主要集中在1～3岁组,占所有鼻病毒感染病例的67.81%;呈现为以夏秋季为感染高峰的季节性流行,9—10月为鼻病毒的检出高峰。 本研究从233份阳性鼻病毒标本中共获得161条鼻病毒VP1序列,通过分子分型鉴定属于55个型别,其中A种鼻病毒37个,B种鼻病毒4个,C种鼻病毒14个。 RV-A12和RV-A49两个型别检出数目最多,均为12条。   结论  2019年6月至2021年5月长春市急性呼吸道感染病例咽拭子标本中鼻病毒检出阳性率位居第二,仅次于呼吸道合胞病毒。 长春市儿童急性呼吸道感染相关的鼻病毒流行以A种鼻病毒为主,C种次之,B种最少;RV-A12和RV-A49是长春市2019—2021年流行的优势型别。     

Virucidal hand treatments for prevention of rhinovirus infection

Rhinovirus infections are associated with substantial morbidity and economic cost. The available common cold remedies are of limited utility and specific antiviral approaches have been unsuccessful. Viral contamination of the hands appears to play an important role in the transmission of rhinovirus from person-to-person. Interruption of this step in transmission presents a potential target for intervention. Initial studies demonstrated that the common cold could be prevented by treatment of hands with iodine. Inactivation of the rhinoviruses by acid is well known and a survey of organic acids considered safe for consumer use revealed that salicylic acid and pyroglutamic acid have potent virucidal activity for the rhinoviruses that persists for several hours after application to the hands. A subsequent evaluation in human volunteers confirmed the prevention of rhinovirus infections by these acids and suggested that these agents have promise as cosmetically acceptable virucidal agents for interruption of the transmission of these infections.     

The seasonality of rhinovirus infections and its implications for clinical recognition

BACKGROUND: Rhinoviruses are the most common cause of acute respiratory infections. Isolation of rhinoviruses occurs in a distinct and consistent seasonal pattern that can be used to help determine whether an acute respiratory illness is caused by a rhinovirus. OBJECTIVE: This article reviews information on the seasonality of rhinovirus infection derived from early and recent studies of rhinovirus occurrence and treatment. METHODS: PubMed was searched from 1965 to the present to identify all potentially relevant papers. The search terms used were rhinovirus and seasonality. A total of 1998 papers were screened. RESULTS: Rhinoviruses comprise more than three quarters of viruses circulating in early autumn. In some years and perhaps some geographic areas, spring is an even more important time for rhinovirus transmission. Although overall rates of respiratory illness are lower in summer, rhinoviruses are the most frequently isolated virus at this time of year. Other viral agents, including influenza viruses and respiratory syncytial virus (particularly with parainfluenza virus), predominate in the winter. Thus, for most of the year, rhinoviruses are the cause of the majority of acute viral respiratory infections. CONCLUSION: Understanding the seasonal incidence of rhinovirus infection may help determine how best to employ currently available antirhinoviral agents in patients presenting with symptoms of an acute viral respiratory infection.     

Inflammatory and bronchospastic factors in asthma exacerbations caused by upper respiratory tract infections

It is still uncertain how viral respiratory infections cause acute exacerbations of bronchial asthma, although several mechanisms have been proposed. We studied the relationship between the airway narrowing and the inflammatory and bronchospastic factors in peripheral venous blood and urine, in 30 patients with asthma at the exacerbations caused by upper respiratory tract infections (URTIs). Acute exacerbations caused decreases in peak expiratory flow rate (PEFR) in all 30 patients with asthma. Asthma exacerbations caused the rises in serum levels of interleukin-6, soluble intercellular adhesion molecule-1 and eosinophil cationic protein, concentrations of urinary leukotriene E4 and plasma histamine, compared with those in patients with asthma at a stable condition and those in 30 control subjects (p < 0.05). The values of PEFR at the exacerbations correlated with the levels of these factors. Treatment with oral glucocorticoids reversed the decreases in PEFR and the increases in these factors. At the onset of URTIs, rhinovirus and influenza type A virus were identified in 13 and 7 patients, respectively. Each of parainfluenza virus, adenovirus, and enterovirus was identified in one patient. These findings suggest that respiratory viral infections may cause acute asthma exacerbations via the production of mediators that induce inflammation and bronchospasm.     

Treatment and prevention of COPD exacerbation

Airway inflammation, mucosal edema, epithelial hyperpermeability, mucus secretion and airway smooth muscle contraction induced by airway bacterial, virus infection and exposure to air pollution may be associated with COPD exacerbation. Severity of COPD exacerbation is estimated by blood gas analysis, serum CRP values and the chest radiograph. Patients with COPD exacerbations are recommended to be treated with additional inhalations of beta-2 agonists and anti -cholinergic agents, systemic administered glucocorticosteroids, oxygen inhalation, and, in cases with purulent sputum, antibiotics. Glucocorticosteroids, beta-2 agonists and anti-cholinergic agents reduce the frequency of COPD exacerbation. We reported the inhibitory effects of glucocorticosteroids on rhinovirus infection, the major cause of common colds, and the inhibitory effects of L-carbocisteine and erythromycin on COPD exacerbations and rhinovirus infection.     

Understanding the mechanisms of viral induced asthma: new therapeutic directions

Asthma is a common and debilitating disease that has substantially increased in prevalence in Western Societies in the last 2 decades. Respiratory tract infections by respiratory syncytial virus (RSV) and rhinovirus (RV) are widely implicated as common causes of the induction and exacerbation of asthma. These infections in early life are associated with the induction of wheeze that may progress to the development of asthma. Infections may also promote airway inflammation and enhance T helper type 2 lymphocyte (Th2 cell) responses that result in exacerbations of established asthma. The mechanisms of how RSV and RV induce and exacerbate asthma are currently being elucidated by clinical studies, in vitro work with human cells and animal models of disease. This research has led to many potential therapeutic strategies and, although none are yet part of clinical practise, they show much promise for the prevention and treatment of viral disease and subsequent asthma.     

Th2 cytokine inhibition and cough in asthmatic and bronchitic patients

BACKGROUND: Activated T helper lymphocytes are present in the airway and their production of cytokines is important in the pathogenesis of asthma, however, the relationship between T helper lymphocyte-derived cytokines and airway cough reflex sensitivity remains unknown. METHODS: The effect of the orally active Th2 cytokine inhibitor suplatast tosilate on cough response to inhaled capsaicin was examined in eleven patients with stable atopic asthma and compared with patients having non-atopic asthma and chronic bronchitis (the latter of which is not related to Th2 cytokines). Capsaicin cough threshold, defined as the lowest concentration of capsaicin eliciting five or more coughs, was measured as an index of airway cough reflex sensitivity. Concentration of serum total IgE level was also measured after treatment with suplatast tosilate. RESULTS: The cough threshold after two weeks treatment with suplatast tosilate was significantly greater than the value with placebo accompanied by decrease of serum IgE level in atopic asthmatics. This significance was not observed in patients with non-atopic asthma or chronic bronchitis. CONCLUSIONS: Th2 cytokines may be possible modulators augmenting airway cough reflex sensitivity in atopic asthmatic airways but not in non-atopic asthmatic or bronchitic airways.     

Th2 cytokine inhibition and cough in asthmatic and bronchitic patients

BACKGROUND: Activated T helper lymphocytes are present in the airway and their production of cytokines is important in the pathogenesis of asthma, however, the relationship between T helper lymphocyte‐derived cytokines and airway cough reflex sensitivity remains unknown. METHODS: The effect of the orally active Th2 cytokine inhibitor suplatast tosilate on cough response to inhaled capsaicin was examined in eleven patients with stable atopic asthma and compared with patients having non‐atopic asthma and chronic bronchitis (the latter of which is not related to Th2 cytokines). Capsaicin cough threshold, defined as the lowest concentration of capsaicin eliciting five or more coughs, was measured as an index of airway cough reflex sensitivity. Concentration of serum total IgE level was also measured after treatment with suplatast tosilate. RESULTS: The cough threshold after two weeks treatment with suplatast tosilate was significantly greater than the value with placebo accompanied by decrease of serum IgE level in atopic asthmatics. This significance was not observed in patients with non‐atopic asthma or chronic bronchitis. CONCLUSIONS: Th2 cytokines may be possible modulators augmenting airway cough reflex sensitivity in atopic asthmatic airways but not in non‐atopic asthmatic or bronchitic airways.     

Type 4 phosphodiesterase inhibitors attenuate respiratory syncytial virus-induced airway hyper-responsiveness and lung eosinophilia

Viral respiratory infections are considered one of the triggers of exacerbations of asthma. In a model of virus-induced airway hyper-responsiveness (AHR), mice infected with human respiratory syncytial virus (RSV) were shown to develop AHR accompanied by lung eosinophilia. Inhibitors of cyclic nucleotide phosphodiesterase (PDE) have been shown to affect airway responsiveness and pulmonary allergic inflammation. In this study, we assessed the effects of type 4 PDE (PDE4) inhibitors on AHR following RSV infection and compared them with a PDE3 inhibitor. In mice infected by intranasal inoculation of RSV, treatment with the PDE4 inhibitor rolipram or Ro-20-1724 reduced both AHR and the eosinophil infiltration of the airways. In contrast, the PDE3 inhibitor, milrinone, did not influence airway responsiveness or eosinophilic inflammation. These results demonstrate that PDE4 inhibitors can modulate RSV-induced AHR and lung eosinophilia and indicate that they have a potential role in treating exacerbations of asthma triggered by viral infection.     

Autocrine interaction between IL-5 and IL-1β mediates altered responsiveness of atopic asthmatic sensitized airway smooth muscle

T-helper type 2 (Th2) cytokines have been implicated in the pathogenesis of the pulmonary inflammatory response and altered bronchial responsiveness in allergic asthma. To elucidate the mechanism of Th2-dependent mediation of altered airway responsiveness in the atopic asthmatic state, the expression and actions of specific cytokines were examined in isolated rabbit and human airway smooth muscle (ASM) tissues and cultured cells passively sensitized with sera from atopic asthmatic patients or nonatopic/nonasthmatic (control) subjects. Relative to control tissues, the atopic asthmatic sensitized ASM exhibited significantly enhanced maximal isometric contractility to acetylcholine and attenuated relaxation responses to isoproterenol. These proasthmatic changes in agonist responsiveness were ablated by pretreating the atopic sensitized tissues with either an IL-5 receptor blocking antibody (IL-5ra) or the human recombinant IL-1 receptor antagonist (IL-1ra), whereas an IL-4 neutralizing antibody had no effect. Moreover, relative to controls, atopic asthmatic sensitized ASM cells demonstrated an initial, early (after 3 hours of incubation) increased mRNA expression and protein release of IL-5. This was followed (after 6 hours of incubation) by an enhanced mRNA expression and release of IL-1beta protein, an effect that was inhibited in sensitized cells pretreated with IL-5ra. Extended studies demonstrated that naive ASM exposed to exogenously administered IL-5 exhibited an induced upregulated mRNA expression and protein release of IL-1beta associated with proasthmatic-like changes in ASM constrictor and relaxant responsiveness, and that these effects were ablated in tissues pretreated with IL-1ra. Taken together, these observations provide new evidence that (a) the Th2 cytokine IL-5 and the pleiotropic proinflammatory cytokine IL-1beta are endogenously released by atopic asthmatic sensitized ASM and mechanistically interact to mediate the proasthmatic perturbations in ASM responsiveness; and (b) the nature of this interaction is given by an initial endogenous release of IL-5, which then acts to induce the autologous release of IL-1beta by the sensitized ASM itself, resulting in its autocrine manifestation of the proasthmatic phenotype.     

Rhinovirus respiratory infections and asthma

Viral infections, particularly respiratory illnesses caused by rhinovirus, are the most common cause of asthma exacerbations in children and contribute in large part to asthma morbidity in adults. Epidemiologic studies and increasingly sophisticated viral detection methodologies have helped to define the role of rhinovirus as a potential causative agent in asthma exacerbations. Rhinovirus-induced lung disease is multifaceted and can be characterized in terms of a variety of physiologic, immunologic, and viral processes. The precise direct and indirect mechanisms of viral contribution to exacerbations must still be elucidated. Understanding them will have an impact on the design of future treatment modalities.     

鼻病毒感染的实验室诊断

鼻病毒（HRV）是引起呼吸道感染的重要病原体,与上呼吸道感染以及慢性肺病、哮喘加重均密切相关.针对鼻病毒的实验室诊断方法有细胞培养方法、免疫检测法和核酸检测法等,掌握这些方法的原理和优缺点对鼻病毒的诊断具有重要意义.     

Orosomucoid-like protein 3, rhinovirus and asthma

The genetic variants of orosomucoid-like protein 3 (ORMDL3) gene are associated with highly significant increases in the number of human rhinovirus (HRV)-induced wheezing episodes in children. Recent investigations have been focused on the mechanisms of ORMDL3 in rhinovirus infection for asthma and asthma exacerbations. ORMDL3 not only regulates major human rhinovirus receptor intercellular adhesion molecule 1 expression, but also plays pivotal roles in viral infection through metabolisms of ceramide and sphingosine-1-phosphate, endoplasmic reticulum (ER) stress, ER-Golgi interface and glycolysis. Research on the roles of ORMDL3 in HRV infection will lead us to identify new biomarkers and novel therapeutic targets in childhood asthma and viral induced asthma exacerbations.     

Fibroblastic reticular cells and their role in viral hemorrhagic fevers

Molecular methods of viral screening have demonstrated that human rhinoviruses (HRVs) are associated with lower respiratory tract infections (LRTIs, including bronchiolitis and pneumonia), exacerbations of chronic pulmonary disease and the development of asthma. Patients with severe chronic diseases are at greater risk of developing major clinical problems when infected by HRVs, particularly if they are immunocompromised or have a chronic lung disease. Analysing the characteristics of HRVs does not provide any certainty concerning the risk of a poor prognosis and, although viremia seems to be associated with an increased risk of severe HRV infection, the available data are too scanty to be considered conclusive. However, a chest x-ray showing alveolar involvement suggests the potentially negative evolution of a bacterial superinfection. There is therefore an urgent need for more effective diagnostic, preventive and therapeutic measures in order to prevent HRV infection, and identify and treat the patients at highest risk.     

Pathogenesis of persistent lymphatic vessel hyperplasia in chronic airway inflammation

Edema occurs in asthma and other inflammatory diseases when the rate of plasma leakage from blood vessels exceeds the drainage through lymphatic vessels and other routes. It is unclear to what extent lymphatic vessels grow to compensate for increased leakage during inflammation and what drives the lymphangiogenesis that does occur. We addressed these issues in mouse models of (a) chronic respiratory tract infection with Mycoplasma pulmonis and (b) adenoviral transduction of airway epithelium with VEGF family growth factors. Blood vessel remodeling and lymphangiogenesis were both robust in infected airways. Inhibition of VEGFR-3 signaling completely prevented the growth of lymphatic vessels but not blood vessels. Lack of lymphatic growth exaggerated mucosal edema and reduced the hypertrophy of draining lymph nodes. Airway dendritic cells, macrophages, neutrophils, and epithelial cells expressed the VEGFR-3 ligands VEGF-C or VEGF-D. Adenoviral delivery of either VEGF-C or VEGF-D evoked lymphangiogenesis without angiogenesis, whereas adenoviral VEGF had the opposite effect. After antibiotic treatment of the infection, inflammation and remodeling of blood vessels quickly subsided, but lymphatic vessels persisted. Together, these findings suggest that when lymphangiogenesis is impaired, airway inflammation may lead to bronchial lymphedema and exaggerated airflow obstruction. Correction of defective lymphangiogenesis may benefit the treatment of asthma and other inflammatory airway diseases.