Immunobiotic lactobacilli reduce viral-associated pulmonary damage through the modulation of inflammation–coagulation interactions

Abstract

The exacerbated disease due to immune- and coagulative-mediated pulmonary injury during acute respiratory viruses infection results in severe morbidity and mortality. Identifying novel approaches to modulate virus-induced inflammation–coagulation interactions could be important alternatives for treating acute respiratory viruses infections. In this study we investigated the effect of the probiotic strain Lactobacillus rhamnosus CRL1505 on lung TLR3-mediated inflammation, and its ability to modulate inflammation–coagulation interaction during respiratory viral infection. Our findings reveal for the first time that a probiotic bacterium is able to influence lung immune-coagulative reaction triggered by TLR3 activation, by modulating the production of proinflammatory and anti-inflammatory cytokines as well as expression of tissue factor and thrombomodulin in the lung. We also demonstrated that the preventive treatment with the probiotic bacteria beneficially modulates the fine tune balance between clearing respiratory viruses (respiratory syncytial virus and influenza virus) and controlling immune-coagulative responses in the lung, allowing normal lung function to be maintained in the face of a viral attack. Our data also pinpoint a crucial role for IL-10 in the immune protection induced by L. rhamnosus CRL1505 during respiratory viral infections. These observations might be helpful to propose new preventive or therapeutic approaches to better control virus-inflammatory lung damage using probiotic functional foods.

Introduction

Influenza virus (IFV) and respiratory syncytial virus (RSV) are common causes of upper respiratory tract infection and pneumonia. Although several studies have examined the host inflammatory/immune responses to these viruses, some investigations have demonstrated an important role of the hemostatic system in the outcome of viral respiratory infections. Besides inflammatory pathways, respiratory viruses can trigger the coagulation system. They increase the expression of tissue factor (TF), the main initiator of coagulation, in endothelial cells and monocytes inducing a prothrombotic state by concurrent stimulation of coagulation and inhibition of fibrinolysis [1], [2], [3], [4]. Although enhanced coagulation may be considered host protective in containing the infection [5], excessive procoagulant activity may result in alveolar fibrin formation and enhancement of inflammation and lung injury. Moreover, much information has accrued demonstrating the close interaction of inflammation, atherosclerosis and thrombosis. Several case–control studies have repeatedly confirmed the common clinical observation that viral respiratory tract infections often shortly precede or accompany acute ischemic strokes or acute myocardial infarctions [6], [7].

Inflammatory and hemostatic alterations in respiratory viral infections have been associated to double-stranded RNA (dsRNA) intermediates produced during the replication of respiratory viruses such as IFV and RSV, which are recognized by a variety of pattern-recognition receptors (PRRs) in respiratory epithelial, endothelial and immune cells, including Toll-like receptor (TLR)-3 and retinoic acid-inducible gene I (RIG-I). In vivo studies using mice have demonstrated that the viral-associated molecular pattern polyinosinic:polycytidylic acid (poly(I:C)), treatment results in TLR3- and CXCR2-dependent neutrophilic pulmonary inflammation, interstitial edema, bronchiolar epithelial hypertrophy, and altered lung function [8], [9]. These changes were accompanied by elevated levels of proinflammatory cytokines and type I interferons in broncho-alveolar lavages (BAL) [8] and, increased airway epithelial cell TLR3 protein expression [9]. In addition, studies have reported that poly(I:C) can upregulate TF and downregulate thrombomodulin (TM) expression on endothelial cells. Moreover, in vivo application of poly(I:C) induces similar changes in the aortic endothelium of mice and increases D-dimer levels indicating enhanced coagulation and fibrinolysis [10].

Certain probiotic lactic acid bacteria (LAB) strains can exert their beneficial effect on the host through their immunomodulatory activity. These strains, termed immunobiotics [11], have been used for the development of functional foods with the ability to stimulate mucosal immunity. Moreover, studies have demonstrated that some immunobiotic LAB can stimulate the common mucosal immune system to provide protection in other mucosal sites distant from the gut [12]. In this regard, several lines of evidence demonstrated that oral administration of immunobiotics is able to increase resistance against respiratory viral infections. It has been described that several aspects of respiratory antiviral immunity can be beneficially modulated by immunobiotics, including the production of type I interferons, the activity of NK cells, the generation of Th1 responses as well as the production of specific antibodies and the regulation of inflammatory lung injury [11]. We recently initiated a series of studies seeking to establish the capacity of Lactobacillus rhamnosus CRL1505 to improve respiratory antiviral immunity. Our research work has demonstrated that mucosal (oral and nasal) administration of the CRL1505 strain is able to beneficially modulate the immune response triggered by TLR3 activation in the respiratory tract and to increase the resistance to RSV challenge [13], [14], [15]. Moreover, L. rhamnosus CRL1505 administration efficiently reduces inflammatory lung tissue damage produced by poly(I:C) or RSV through its capacity to beneficially modulate proinflammatory/IL-10 and Th1/Th2 balances in the respiratory tract [13], [14], [15]. On the other hand, we demonstrated that some immunobiotic strains such as L. casei CRL431 or L. rhamnosus CRL1505 are able to beneficially modulate the inflammation–coagulation interaction during respiratory infections, indicating that LAB is able to modulate the immune-coagulative response [16], [17], [18], [19]. Much research of our group has been done on coagulation activation during severe bacterial infections, and no data on the modulation of coagulation/inflammation interaction by immunobiotics in viral infections are available.

The exacerbated disease due to immune- and coagulative-mediated pulmonary injury during acute respiratory viruses infection results in severe morbidity and mortality. Then, identifying novel approaches to modulate virus-induced inflammation–coagulation interactions could be important alternatives for treating acute respiratory viruses infections. In this sense, studying the effect of orally administered immunobiotics on the immune-coagulative response triggered by respiratory activation of TLR3 would contribute to the knowledge of the mechanism of probiotics' protective effect against respiratory viral infections. Therefore, the aim of the present study was to deepen the understanding of the mechanisms of L. rhamnosus CRL1505 immunoregulatory activity by evaluating a) its effects on lung TLR3-mediated inflammation; b) its ability to modulate inflammation–coagulation interaction; c) and its influence on the outcome of respiratory viruses challenges.