【病毒外文文献】2004 Severe Acute Respiratory Syndrome, a Pathological Immune Response to the New Coronavirus_Implications for Understan

535 VIRAL IMMUNOLOGY Volume 17 Number 4 2004 Mary Ann Liebert Inc Pp 535 544 Severe Acute Respiratory Syndrome a Pathological Immune Response to the New Coronavirus Implications for Understanding of Pathogenesis Therapy Design of Vaccines and Epidemiology JESUS F BERMEJO and M ANGELES MU OZ FERNANDEZ ABSTRACT Findings coming from autopsies and serum of SARS patients suggest an important immune in flammatory implication in the evolution to respiratory distress Conditions such as HIV infection or treatment with immunosuppressors in cancer or autoimmune diseases are not among the bad prognosis factors for development of distress To date there have been no reported case fatalities in children probably due to their more immature immune system Our conclusions follow 1 The milder form of SARS in children and the apparent protective factor that immunosupression rep resent rules out a significant viral cytopathic effect they would be the most affected 2 The evi dence for immune implication in distress strongly supports immunomodulators for therapy phos phodiesterase inhibitors due to their down modulating activity on proinflammatory cytokines inhaled corticoids aimed at producing a local immunomodulation teophylline or nedocromil sodium which prevents inflammatory cell recruitment into the airway wall 3 An early im munomodulatory therapy based on the levels of proinflammatory cytokines and clinical parame ters to evaluate the respiratory function such as arterial oxygen saturation could prevent the oc currence of distress 4 Vaccine design should consider the immune origin of distress 5 Physicians should be aware of mildly symptomatic patients children immuno compromised hosts to avoid transmission to immunocompetent adults Laboratorio de Inmunobiologia Molecular Hospital General Universitario Gregorio Mara on Madrid Spain INTRODUCTION S EVERE ACUTE RESPIRATORY SYNDROME SARS pro duced 774 deaths worldwide during the first and so far the main outburst in 2003 SARS has returned in 2004 in the form of limited focuses that have been con trolled satisfactorily The causal agent was identified as a new Coronavirus that fulfilled Koch s postulates 13 Once the causal agent was identified the following steps were taken to understand the viral cycle transmission and disease spread better study of viral tropism identi fication of tissues and fluids where the virus was present characterization of possible reservoirs and achievement of an animal model 30 36 However we feel there are still unsolved questions regarding SARS pathogenesis What is the underlying cause of respiratory distress Is it a direct viral cytopathic damage or an immune medi ated damage in response to the viral infection What is the relevance and weight of both components in the pathogenesis of SARS In adult SARS patients respiratory distress is the prin cipal cause of mortality 14 When the scientific com 5354 08 p535 544 12 6 04 2 30 PM Page 535 munity deals with a viral infectious disease the first ra tional approach is to attribute the damage to a direct vi ral action SARS has not been an exception This explains why the most important international initiatives have in volved achieving an effective vaccine to test drugs that could show an antiviral effect and to design new ones for this purpose However there is strong evidence that the immune system is implicated in SARS pathogenesis Anatomopathological findings The first evidence came from autopsies 42 which showed flooding of alveolar lumina with edema fluid mixed with inflamma tory cells a pronounced increase in macrophages in the alveoli and the interstitium of the lung and hemophago cytosis Hemophagocytosis has been previously attrib uted to dysregulation of T lymphocytes and proinflam matory cytokines 24 As suggested by Nicholls et al proinflammatory cytokines released by stimulated macrophages in the alveoli could have a prominent role in SARS pathogenesis 42 Cytokine profile In concordance with these findings various authors have observed elevation of several pro inflammatory cytokines and chemokines in the serum samples of SARS patients Table 1 This fact may be associated with lung infiltration and proliferation 2 12 53 56 In contrast Zhang et al have recently re ported finding no elevation of proinflammatory cytokines in SARS patients except interleukin 6 IL 6 57 The possible reason for these differences will be discussed later Wong et al found increases in interferon IFN gamma in addition to a number of other cytokines and chemokines during the 2 weeks after onset 53 IFN gamma plays an important role in regulating the balance BERMEJO AND MUN OZ FERNANDEZ between Th1 and Th2 cells Firstly it increases the syn thesis of IL 12 in antigen presenting cells IL 12 is the primary effector that drives developing CD4 H11001 T cells to become Th1 cells Secondly IFN gamma prevents the development of Th2 cells by inhibiting the production of IL 4 which is required for Th2 cell formation IFN gamma also plays an important role in macrophage acti vation 18 A group of Canadian researchers of the Uni versity Health Network Toronto and the Canadian Network for Vaccines and Immunotherapeutics CAN VAC have studied the role of chemokines in SARS CXCL10 IFN gamma inducible protein 10 IP 10 is a chemokine associated with inflammation and prolifera tion In a study on SARS patients they observed that plasma levels of IFN gamma were significantly elevated at onset of symptoms Levels of CXCL10 were also sig nificantly increased in SARS patients at onset but re mained specifically elevated in critical patients and were highest in dying SARS patients Furthermore CXCL10 and its receptor CXCR3 were elevated locally in lung specimens from deceased patients 25 They hypothe size that the maintenance of high CXCL10 expression in the lung in SARS patients may generate a positive feed back loop resulting in the continual recruitment of acti vated T cells This would lead to severe lung inflamma tion Thus IFN gamma could be promoting the secretion of proinflammatory chemokines in SARS in conse quence IFN gamma down modulation could translate into clinical benefit On the other hand interferon alpha and beta are known to induce cells into the antiviral state Preliminary studies with interferon alpha and beta re veal a promising antiviral activity of these drugs 536 TABLE 1 CYTOKINE PROFILE IN SARS Beijing Group of National Duan et al Research Project for SARS Wong et al Xie et al IL 1b XX IL 2 X IL 4 X IL 6 X IL 8 XX X IL 10 X IL 12 X IL 13 X IL 16 X TNF alpha XX X TGF beta1 X IFN gamma X MCP 1 X IP 10 X Proinflammatory cytokines and chemokines are bold faced IL interleukine TNF tumour necrosis factor TGF transforming growth factor IFN interferon MCP monocyte chemoattractant protein IP 10 IFN gamma inducible protein 10 5354 08 p535 544 12 6 04 2 30 PM Page 536 10 20 48 The authors of these studies suggest their clin ical testing for SARS treatment To conclude the virus is believed to come from an animal host with further transmission to humans Based on this it does not seem to be hazardous to suppose that an incorrect or exacer bated immune response against this uncommon virus could lead to an intense inflammatory response In the opinion of Duan et al this inflammatory response is sys temic 12 focusing mainly on the lungs Table 1 Viral load evolution An important event that supports the immune implication in the development of respira tory distress is the evolution of viral load The timing of the IgG seroconversion in SARS patients which starts on day 10 seems to correlate with falls in viral load which occur from day 10 to 15 Severe clinical worsen ing also occurs at this time which cannot be explained by an uncontrolled viral replication The findings suggest that the lung damage at this phase is related to an im munopathological damage resulting from an overly exu berant host response rather than an uncontrolled viral replication 45 Clinical course in children The first reports about SARS in pediatric patients yielded a surprising conclu sion SARS in children seems to appear in a more benign form than in adults There is currently abundant evidence that the disease in children under 12 is less severe than in adults while adolescents clinical features are similar to those found in adults To date there have been no re ported fatalities in children with this disease 5 8 11 22 31 50 54 Three possible explanations have been suggested a children were kept relatively isolated from the outside world during the epidemic therefore avoiding the infection b their immature immune sys tem does not cause acute lung injury and c children have anti SARS antibodies GuanFu and ZhongYuan studied the possibility of the existence of protective anti SARS antibodies in children as a result of multiple vac cinations in the childhood 19 Their studies with vac cinated mice showed that there was essentially no cross immunoreactivity between the SARS coronavirus and the 13 common vaccinal immunogens tested This study along with the communications reporting that anti SARS activity can be detected in very few if any non SARS children s serum samples 33 34 leads to the con clusion that the presence of anti SARS antibodies in children can be apparently ruled out as the cause of the low morbidity In regard to the children s isolation the ory in our opinion it is difficult to maintain that in the affected Asiatic countries with an elevated population density and strong familial communities children could be kept away from infected adults Thus the immaturity of children s immune system appears to be the most plau sible explanation for the low incidence of respiratory dis tress in children under 12 years old and in general for SARS A PATHOLOGICAL IMMUNE RESPONSE TO THE NEW CORONAVIRUS the milder course of the disease in children These con clusions again support a pathogenic role of immune re sponse against the virus that could explain the pathogenic events in immunocompetent adults 4 Immunosuppression as a probable protective fac tor In the context of a viral disease immunosuppressed hosts are thought to be the most affected Fowler et al 14 reported a cohort of critically ill patients with SARS Patients who died were more often older people with co morbidities such as diabetes However immunosuppres sive conditions such as HIV infection or treatment with immunosuppressors employed in cancer patients or au toimmune disorders for example were not listed among the negative prognosis factors The Chinese Ministry of Health has estimated that more than 1 million people were infected by HIV in China by the end of 2002 In contrast to date no reports indicating that HIV infection is a risk factor for development of SARS have been published in China neither from other affected countries such as Tai wan Canada or Singapore There has been only one doc umented case of an HIV patient infected with the SARS associated coronavirus 52 He was on highly active antiretroviral therapy HAART including lopinavir ri tonavir Kaletra efavirenz abacavir and tenofovir The patient s disease ran a relatively mild course and in 39 days he was declared SARS free The authors offer two possible explanations the protective effect of HAART or the immunosuppression due to HIV infection HIV pa tients in contact with SARS patients have been shown not to develop respiratory distress 7 Most of these HIV patients were in treatment with HAART so a protective effect of HAART in SARS has been hypothesized Some of the drugs used for HIV treatment could display an tiviral activity against the SARS associated Coronavirus Chu et al observed an apparently favourable clinical re sponse in their patients treated with a combination of anti HIV protease inhibitors PI lopinavir ritonavir Kaletra with ribavirin 9 Consequently the use of anti HIV drugs has been suggested for patients with SARS and in the prophylaxis of patients close contacts 7 However if the case of the only HIV patient reported with SARS infection is considered the patient was in treatment with PI before SARS infection As the authors comment it could be argued that if Kaletra was active against Coro navirus it should have prevented the infection in the first place 52 In our opinion the positive role of HIV PI in SARS could be attributed not only to an antiviral ac tivity but also to an immunomodulatory activity Nuclear factor kappa B NF kB is an ubiquitous transcriptional factor and a pleiotropic regulator of many genes involved in inflammatory responses and immuno regulatory ac tivities In most cell types NF kB is associated with the inhibitor IkB in the cytoplasm The NF kB activation fol lows this sequence of events in response to external sig 537 5354 08 p535 544 12 6 04 2 30 PM Page 537 nals IkB binds to NF kB factor Activation requires IkB phosphorylation at serine residues followed by phos phorylation dependent multi ubiquitination at lysine residues degradation of IkB by an ubiquitin dependent proteasome and finally the release of free NF kB tran scription factor Free NF kB moves to the nucleus and induces expression of certain genes kB degradation is an efficient process that can be inhibited by serine protease inhibitors suggesting that it is an obligatory step in NF kB activation 17 Monini et al have reported that IP are capable of inhibiting inflammatory cytokine produc tion and modulating antigen presentation and T cell re sponses 37 In particular ritonavir has recently been shown to inhibit the expression of adhesion molecules and the production or release of inflammatory cytokines or chemokines including tumour necrosis factor TNF alpha IL 6 or IL 8 by endothelial cells 44 HIV PIs used at therapeutic concentrations affect pathways in volved in cell invasion and matrix metallo proteases MMP activity particularly MMP 2 proteolytic activa tion 37 MMPs are known to be involved in several im mune and immunomodulatory functions Specifically MMPs are required for leucocyte transmigration and tis sue infiltration by inflammatory cells In this context ri tonavir has been shown to abolish cytotoxic T lympho cyte CTL dependent inflammatory responses in a murine model of lymphocytic choriomeningitis virus in fection 1 Andr et al found that ritonavir appeared to inhibit the chymotrypsin like activity of isolated 20S pro teasomes in vitro being able to reduce proteasome cellu lar functions such as the degradation of ubiquitin conju gates and IkB 1 In addition Tovo has reported a study on two HIV uninfected subjects in prophylaxis with HAART including indinavir At the end of prophylaxis in both subjects who did not acquire HIV the percentage of peripheral blood mononuclear cells expressing TNF alpha IL 2 and IFN gamma was reduced when compared with simultaneously tested normal controls 51 All these studies attempt to examine the role that the immunomod ulatory properties of IP could play in SARS Corticoids in the treatment In general the use of steroids for SARS seems to be beneficial The differences encountered in the clinical results with corticoids are prob ably explained by the fact that the optimal timing dosage and duration of treatment have not been determined yet 15 However as discussed in the second part of this arti cle the adverse effects of systemic corticoids are them selves a cause of mortality and morbidity Fig 1 ANALYSIS The evidences recovered here could contribute to a bet ter understanding of the disease On the basis of these BERMEJO AND MUN OZ FERNANDEZ evidences the responsibility of an abnormal immune re sponse in the pathogenesis of respiratory distress appears to be clear This conclusion could lead to the develop ment of better treatment approaches It also affects vac cine design epidemiological control of the disease and prevention of spreading Relevance for therapy strategies An important ques tion to be answer for SARS treatment is the degree of vi ral pathogenicity If a direct viral cytopathic effect were the principal cause of the lung damage in SARS then young children would be the most affected patients due to their immune system s immaturity 4 The same ratio nal could be applied to immuno suppressed hosts How ever the reality is that children under 12 years old do not develop respiratory distress with no need of oxi genotherapy Additionally immunosuppressed hosts are not reported to show an increased mortality Even more it seems to represent a protective factor Pharmacological research screening of drugs with antiviral activity against the new Coronavirus and design of new ones is undoubt edly necessary in SARS However perhaps we currently have drugs in our pharmacies that could prevent respira tory distress The role that immunomodulators could play in SARS therapy acquires more relevance we have pre viously proposed phosphodiesterase PDE inhibitors as drugs to be considered because of their activity in de creasing proinflammatory cytokines They present other interesting pharmacological properties for example their ability to decrease the in vitro replication of several kinds of viruses 3 In addition we propose here other alterna tive immunomodulators inhaled corticoids teophylline cromolyn sodium ketotifen and nedocromil sodium PDE inhibitors a Non specific PDE inhibitors Pentoxifylline PTX PTX is able to decrease the secretion of TNF IL 1 and IL 6 as well as IP 10 CXCL10 modulating neutrophil 538 FIG 1 Evidence of the immune implication in SARS 5354 08 p535 544 12 6 04 2 30 PM Page 538 and macrophage activation 6 27 29 40 46 By blocking the inflammatory action of IL 1 and TNF on neutrophils PTX may reduce the tissue damage caused by neutrophils in conditions such as septic shock adult respiratory dis tress syndrome cardiopulmonary bypass lung damage and myocardial reperfusion injury 47 Other proof or its efficacy modulating inflammation is that PTX can in hibit cytokine release from alveolar macrophages in pul monary sarcoidosis 49 a NIH clinical trial to test PTX in this disease is currently ongoing Additionally PTX can inhibit ICAM 1 expression and chemokine produc tion IL 8 and MCP 1 induced by proinflammatory cy tokines in human pulmonary epithelial cells 28 In a previous study our group examined the in vitro activity of PTX on cell proliferation cytokine production viral replication and CD4 H11001 depletion in acutely HIV 1 in fected human T cells PTX was able to inhibit with sim ilar potency IFN gamma TNF alpha and cell prolifera tion 38 PTX also has an additional advantage Evidence on its down modulating effect on proinflammatory cy tokines does not come only from in vitro assays It has also been tested in patients patients with other diseases with an important inflammatory component In this con text PTX administration provides clinical benefit Ex amples of this are HTLV I associated myelopathy tropi cal spastic paraparesis asthma acute respiratory distress syndrome in oncological patients or mucosal leishmani asis 3 It has also been tested successfully in animal models for P carinii lung infection and experimental bleomycin induced fibrosing alveolitis two other in flammatory conditions 3 In relation to the mechanism of action of PTX we showed in another work that PTX and Rolipram a specific PDE IV inhibitor discussed later decreased transcription of IL 2 and TNF alpha pro moters in transiently transfected normal T cells More over they inhibited the activation of NF kB and nuclear factor of activated T cells NFAT and stimulated acti vator protein 1 AP 1 and cAMP response element binding proteins CREBs These data indicate that block ade of PDE IV regulates transcription of inflammatory cytokines through inhibition of NF kB and NFAT and stimulation of AP 1 and CREB 23 But the complete mechanisms of action of PTX remain to be elucidated Haddad et al concluded in their elegant work with pri mary cultures of alveolar epithelia that taken together the potent anti inflammatory potential of PTX points to a multifaceted mechanism of action