**4.2 DENV infection and the host innate immune responses**

Although plasma leakage in severe dengue occurs at the end of the acute illness, there is substantial evidence that the pathophysiologic processes start at the earliest stages of DENV infection [95, 96]. Introduction of DENV particles along with mosquito saliva triggers a variety of host innate immune responses leading to the production of antiviral and pro-inflammatory cytokines mostly from the immune cells exposed to DENV [57, 62, 138]. At this stage, innate immune cells are the first to respond to infection through stimulation of patterns recognition receptors (PRRs) recognizing pathogen-associated molecular patterns (PAMPs) as well as endogenous molecules released from damaged cells, termed damage-associated molecular patterns (DAMPs) [161, 162]. PRR recognition triggers the production of cytokines and chemokines, which induces a local antiviral state [54, 55]. This local innate response could potentially play an important role in modulating local viremia and virus dissemination by recruiting susceptible target cells for DENV infection at the inoculation site [57, 62, 144].

PRRs include transmembrane proteins such as the Toll-like receptors (TLRs) and C-type lectin receptors (CLRs) as well as cytoplasmic proteins such as the retinoic acid-inducible gene (RIG)-I-like receptors (RLRs) and NOD-like receptors (NLRs) [161]. These are an essential part of the innate immune response against the virus, sensing viral replication in the cytoplasm [161, 163]. The PRRs that are associated with DENV recognition after infecting target cells are the cytoplasmic retinoic acid-inducible gene I (RIG-I) and the melanoma differentiation-associated protein 5 (MDA5) and the endosomal Toll-like receptor 3 (TLR3) and TLR7 [164–166]. Recognition of DENV RNA by TLR-3 results in the production of type I IFN and chemokines such as IL-8 via sensing of phosphate-containing RNA and long double-stranded RNA (dsRNA) in the cytoplasm or inside endosomal compartments [167, 168]. DENV infection in nonhuman primates demonstrated that the administration of TLR-3 and TLR7 agonists resulted in significantly decreased viral replication and increased production of pro-inflammatory chemokines as well as increased production of antibodies targeting DENV [169], indicating a protective role for TLRs during DENV infection.

Additional pathways such as the cyclic GMP-AM synthase (cGAS), a DNA-sensor pathway which triggers the simulator of IFN genes (STING) pathway are also activated during DENV infection leading to the production of type I IFN and activation of TLRs (TLR9), an endosomal PRR that recognizes cytoplasmic DNA originated from mitochondrial damage [170–172]. In addition to type I IFN production, small RNAs such as micro RNAs (miRNA) and the complement system are important components of the innate immune response against viral infections [173, 174]. miR-NAs are processed by and interact with the proteins in the RNA interference (RNAi) pathway, such as Dicer, Drosha, Argo1, and Argo2 [175]. RNA interference (RNAi) is an important antiviral defense response in plants and invertebrates [176]. In DENV infection, knockdown of these components resulted in increased DENV replication in mammalian cells, suggesting that the RNAi pathways may play important roles in the cellular anti-DENV responses [177, 178]. Additional evidence showed that DENV can interfere with RNAi pathways in human hepatocytes cells via NS4B and subgenomic flavivirus RNA (sfRNA) interactions with Dicer's ability to process small

*Dengue Fever in a One Health Perspective*

combat DENV infection [54, 141].

In animal models such as the alpha/beta (IFN-α/β)-deficient mice (*Ifnar*−/−) and nonhuman primates, DENV has been recovered from the spleen, liver, peripheral lymph nodes, and the central nervous system [123–127]. However, the absence of an appropriate animal disease model has largely hampered with the understanding of the role played by DENV tropism *in vivo*. Sustained viral replication and severe manifestations have been observed in *Ifnar*−/− mice after infection with DENV, which gives a clear advantage to study DENV pathogenesis *in vivo*, but the absence of intact IFN signaling

The fact that DENV can infect many mammalian and insect cell types *in vitro* and *in vivo* suggest there are different molecules or cellular routes that might be controlling virus attachment and internalization, resulting in productive infection [63]. Numerous studies have shown that C-type lectins including DC-SIGN (CD209) and C-type lectin domain family 5, member A (CLEC5A) expressed on dendritic cells and macrophages act as cellular receptors for DENV [129–131]. Other extensively studied DC receptors are the mannose receptor (MR), Langerins, Fc-receptors, TIM3, TIM4, and AXL [63, 65, 132, 133]. Contrary to the DC-SIGN that may primarily function as a viral attachment factor, DENV binding to CLEC5A (C-type lectin domain family 5, member A), highly expressed by monocytes, macrophages, neutrophils, and dendritic cells, has been shown to induce the production of antiviral and pro-inflammatory cytokines suggesting that this C-type lectin may act as a cognate receptor for dengue virion [131, 134]. These cytokines include type I IFNs and chemotactic factors such as migration inhibition factor (MIF), monocyte chemotactic factor (MCP), and IL-8 [102, 134]. DENV infection of DCs also induces the production of matrix metalloproteinases (MPPs), MMP-2 and MMP-9, which induces migration of DCs to lymph nodes where virus further replicates before it disseminates into the blood circulation [135]. In the skin, DENV also infects mast cells that can be activated leading to degranulation and increased secretion of various inflammatory cytokines (IL-1, IL-6, TNF-α, and IFN-α), chemokines (CCL5, CXCL12, and CX3CL1), and chymase, the latter being a protease found circulating at high levels in the blood of dengue patients, suggesting a potential role in the development of severe dengue that contributes to vascular leakage [115, 136–140]. All these innate immune processes together lead to an antiviral state in nearby cells, generating an inflammatory response and recruitment of natural killer (NK) cells to

Along with DCs, monocytes and macrophages are also the primary targets of DENV infection [142, 143]. In lymphoid and nonlymphoid tissues, macrophages are considered the primary reservoirs of DENV after its dissemination from the skin [144]. Macrophages susceptible to DENV have been found in different organs in the mouse models or human autopsies, namely, Kupfer cells in the liver, alveolar macrophages in the lungs, dermal macrophages, microglial cells (brain and spinal cord), and monocytes in the peripheral blood [118, 120, 122, 145–147]. Comparable to DENV infection of DCs, DENV can use an array of cell surface receptors to infect monocytes and macrophages, including mannose receptor (CD205), CD14 associated protein, heat shock proteins (HSP70/HSP90), DC-SIGN (CD209), CD300a, AXL, TIM4, PD1, and the Fc receptors, particularly FcγRI (CD64) and FcγRII (CD32, 63). These two Fc-Rs play major roles in enhancing DENV infection of monocytes and macrophages, particularly during secondary infections [148–151]. Other populations of immune cells including NK cells can also be activated during DENV infection, particularly in patients with DHF compared to those with DF [141, 152, 153]. Additionally, B cells and T cells have been studied to test permissiveness to DENV, but these studies have resulted in contradictory results [154–156]. *In vitro* studies using B cell and T cell lines (e.g., *Raji* cells, Daudi, and Jurkat) and primary B cells derived from healthy human peripheral blood mononuclear cells

is a limitation that must be considered when interpreting data [128].

**68**

RNA *in vitro* [179]. sfRANs are abundant noncoding RNA sequences derived from the stalling of the host 5′-3′ exoribonuclease XRN1/Pacman in the 3′-untranslated regions (3′UTRs) of the viral genomic RNA [180]. sfRNAs have been shown to block exonuclease XRN1, increasing the overall messenger RNA stability within the host cell which may also benefit the viral RNA [181–183]. However, evidence for RNAi contribution to mammalian antiviral defense are few and still controversial [179]. miRNAs have been shown to regulate TLRs and IL-1 signaling pathways in response to viral infection, which provides control of host innate immune responses [184]. So far, there have been reported several cellular miRNAs (miRNAome) that are modulated during DENV infection of mammalian cells, mainly related to the regulation of IFN-β signaling pathways [185–188]. Some of these miRNAs have been proposed to be used as biomarkers in dengue-infected patients [189, 190]. Interestingly, modulation of microRNA expressions have been also described upon DENV infection of insect cells (e.g., C6/36 cells) as well as adult vector mosquitoes such as *Aedes albopictus* [191–193], suggesting that DENV might also regulate the activation of these antiviral RNAi pathways in vector mosquitoes, potentially avoiding viral clearance that promotes viral replication and transmission [194]. Despite RNAi constituting an evolutionarily conserved phenomenon of the mosquito-innate immune response to virus infections [195], viruses have found ways to subvert the RNAi-mediated antiviral responses in vector mosquitoes to manipulate miRNA profiles to their own benefit [196].

Regarding the complement system, this multifaceted pathway has been shown to limit DENV replication; however, excessively activated complement components have been also associated with disease severity [197]. The complement cascade constitutes an integral component of the immune system, composed of many plasma proteins that once activated can initiate a proteolytic cascade, resulting in the release of chemokines, facilitation of particle phagocytosis via opsonization, and deposition of the cell-killing membrane attack complex (MAC) designed to target and destroy foreign pathogens such as viruses [174]. Activation of the complement system occurs via three convergent pathways: the classical, the lectin, and the alternative pathways [174]. *In vitro* experiments showed that DENV replication enhances complement activation [197–199]. Additionally, clinical and *in vivo* studies have shown that excessive consumption of some complement components (e.g., C3, C4, and factor B) contributed to severe manifestations by increasing the levels of complement-activated products which enhance vascular permeability to cause severe dengue disease [9, 108, 109, 200]. In fact, increased circulation of anaphylatoxins (C3a, C4a, and C5a) in the blood of severe patients correlated with symptoms of vascular leakage [109, 200]. In autopsy studies from children who died of acute severe dengue manifestations (DHF/DSS), augmented deposition of complement components from both classical and alternative pathways were found on hepatocytes which results in severe liver damage and death [120]. Altogether, these data support the hypothesis that exacerbated complement activation influences dengue disease immunopathogenesis leading to disease severity [197].

#### **4.3 DENV subversion of antiviral responses**

The first barrier to overcome for successful viral infection is the rapid innate immune responses of the host, including type I IFNs, inflammatory cytokines, complement responses, NK cells, apoptosis, and autophagy [201, 202]. These innate immune responses are meant to defeat viral infections by engaging specific viral components (e.g., RNA and DNA) leading to activation of immediate protective defense mechanisms such as the rapid recognition of PAMP in nonimmune and innate immune cells [161]. IFN production is a key goal of PRR activation for viral pathogens, and DENV is highly susceptible to effective induction of both type

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I (IFN α/β) and type II (IFN γ) interferons [124, 203, 204]. Accordingly, *in vivo* DENV infection of wild-type mice causes little disease; in contrast, in mice lacking

Secreted type I IFNs trigger autocrine and paracrine induction of cellular antiviral responses and warning signals to noninfected adjacent cells, such as the expression of the interferon stimulated genes (ISGs) [205, 206]. ISGs have been shown to exert numerous antiviral effector functions, many of which are still not fully described [207]. Upon DENV infection, RLRs are activated to trigger antiviral responses based on the induction of type I IFN and pro-inflammatory cytokines [208]. The binding of type I IFN with its receptor activates multi-subsets of ISGs through JAK-STAT signaling which amplifies and sustains the initial antiviral responses [207, 209, 210]. However, ISGs can also be activated in IFN-independent pathways during DENV infection [211]. DENV infection has been shown to trigger the transcriptional activation of ISGs *in vivo* and *in vitro* [208, 212–215]. For instance, a tripartite motif (TRIM) protein encoding gene, *TRIM69*, is induced during DENV infection as an ISG. TRIM69 restricts DENV replication by direct interaction with DENV NS3, which mediates its polyubiquitination and degradation in a process called ISGylation [216]. In addition to ISGs, activation of the transcription factors IRF-3, IRF-7, and NF-κB through either the TLR or RIG-I/MDA5 pathways results in the production of type I IFN which contributes to anti-DENV immunity [217, 218]. IRF-3 and IRF-7 are part of the interferon regulatory factors (IRFs) considered the master regulators of the type I IFN production that contribute to the suppression of viruses [219]. Due to the central importance in viral defense, many pathogenic viruses, including DENV, have evolved mechanisms to suppress IRF signaling. In the case of DENV, the nonstructural proteins restrict IRF3 and IFN response which facilitate DENV replication and virulence [220].

In recent years, considerable advances have been made toward understanding of the specific IFN antagonistic mechanisms evolved by DENV to subvert these intracellular antiviral mechanisms and directly inhibiting these cellular signaling cascades, which results in enhanced virus infection, pathogenesis, and disease [167, 221]. This is supported by the increased susceptibility of mice deficient in IFN-α/β and IFN-γ receptors (AG129) to DENV infection as compared to wild-type mice [124, 126, 127]. Although IFN response is antagonized in mouse, human cells still induce high levels of IFN production in response to DENV, so this pathway is not entirely abrogated in humans during infection [203, 222]. Accordingly, humans infected with DENV have high levels of circulating of type I and type II IFNs [223–225]. Strong IFN-α responses have shown to correlate with milder dengue clinical conditions [226]. Similarly, the levels of the dengue-related gene expression of ISGs have been reported to be lower in patients with more severe disease [227–229] suggesting that DENV may abrogate IFN

responses to facilitate viral infection which results in severe manifestations.

From the viral perspective, DENV uses its nonstructural (NS) proteins to block and inhibit the antiviral sensing pathways in infected cells. NS2a, NS3, NS4a, NS4b, and NS5 prevent the virus from being sensed by RIG-I, inhibiting IFNβ induction [230–233]. NS2a, NS4a, and NS4b complex inhibits STAT1 signaling after IFNAR activation *in vitro* [233, 234]. NS5 induces proteasomal degradation of STAT2 which inhibits IFN-mediated response [230]. NS2b induces degradation of cGAS, which prevents DNA sensing resulting from mitochondrial damage [170, 171, 235]. The NS2b/3 protease complex cleaves STING which inhibits IFN production [236]. This phenomenon has been shown to occur in human but not nonhuman primates, suggesting that DENV may have evolved to increase viral titers in human populations, while maintaining decreased titers and pathogenicity in rare animals would serve as a sustainable reservoir in nature [237].

In addition to NS proteins, flavivirus sfRNAs have been described to regulate the innate immune responses via binding and inactivating RNA-binding proteins which are crucial for innate immunity [180, 238]. DENV 3′UTRs possess RNA structures

of type I IFN receptors (IFANR), DENV infection causes mortality [126].

*DOI: http://dx.doi.org/10.5772/intechopen.93140*

*Dengue Immunopathogenesis: A Crosstalk between Host and Viral Factors Leading to Disease… DOI: http://dx.doi.org/10.5772/intechopen.93140*

I (IFN α/β) and type II (IFN γ) interferons [124, 203, 204]. Accordingly, *in vivo* DENV infection of wild-type mice causes little disease; in contrast, in mice lacking of type I IFN receptors (IFANR), DENV infection causes mortality [126].

Secreted type I IFNs trigger autocrine and paracrine induction of cellular antiviral responses and warning signals to noninfected adjacent cells, such as the expression of the interferon stimulated genes (ISGs) [205, 206]. ISGs have been shown to exert numerous antiviral effector functions, many of which are still not fully described [207]. Upon DENV infection, RLRs are activated to trigger antiviral responses based on the induction of type I IFN and pro-inflammatory cytokines [208]. The binding of type I IFN with its receptor activates multi-subsets of ISGs through JAK-STAT signaling which amplifies and sustains the initial antiviral responses [207, 209, 210]. However, ISGs can also be activated in IFN-independent pathways during DENV infection [211]. DENV infection has been shown to trigger the transcriptional activation of ISGs *in vivo* and *in vitro* [208, 212–215]. For instance, a tripartite motif (TRIM) protein encoding gene, *TRIM69*, is induced during DENV infection as an ISG. TRIM69 restricts DENV replication by direct interaction with DENV NS3, which mediates its polyubiquitination and degradation in a process called ISGylation [216]. In addition to ISGs, activation of the transcription factors IRF-3, IRF-7, and NF-κB through either the TLR or RIG-I/MDA5 pathways results in the production of type I IFN which contributes to anti-DENV immunity [217, 218]. IRF-3 and IRF-7 are part of the interferon regulatory factors (IRFs) considered the master regulators of the type I IFN production that contribute to the suppression of viruses [219]. Due to the central importance in viral defense, many pathogenic viruses, including DENV, have evolved mechanisms to suppress IRF signaling. In the case of DENV, the nonstructural proteins restrict IRF3 and IFN response which facilitate DENV replication and virulence [220].

In recent years, considerable advances have been made toward understanding of the specific IFN antagonistic mechanisms evolved by DENV to subvert these intracellular antiviral mechanisms and directly inhibiting these cellular signaling cascades, which results in enhanced virus infection, pathogenesis, and disease [167, 221]. This is supported by the increased susceptibility of mice deficient in IFN-α/β and IFN-γ receptors (AG129) to DENV infection as compared to wild-type mice [124, 126, 127]. Although IFN response is antagonized in mouse, human cells still induce high levels of IFN production in response to DENV, so this pathway is not entirely abrogated in humans during infection [203, 222]. Accordingly, humans infected with DENV have high levels of circulating of type I and type II IFNs [223–225]. Strong IFN-α responses have shown to correlate with milder dengue clinical conditions [226]. Similarly, the levels of the dengue-related gene expression of ISGs have been reported to be lower in patients with more severe disease [227–229] suggesting that DENV may abrogate IFN responses to facilitate viral infection which results in severe manifestations.

From the viral perspective, DENV uses its nonstructural (NS) proteins to block and inhibit the antiviral sensing pathways in infected cells. NS2a, NS3, NS4a, NS4b, and NS5 prevent the virus from being sensed by RIG-I, inhibiting IFNβ induction [230–233]. NS2a, NS4a, and NS4b complex inhibits STAT1 signaling after IFNAR activation *in vitro* [233, 234]. NS5 induces proteasomal degradation of STAT2 which inhibits IFN-mediated response [230]. NS2b induces degradation of cGAS, which prevents DNA sensing resulting from mitochondrial damage [170, 171, 235]. The NS2b/3 protease complex cleaves STING which inhibits IFN production [236]. This phenomenon has been shown to occur in human but not nonhuman primates, suggesting that DENV may have evolved to increase viral titers in human populations, while maintaining decreased titers and pathogenicity in rare animals would serve as a sustainable reservoir in nature [237].

In addition to NS proteins, flavivirus sfRNAs have been described to regulate the innate immune responses via binding and inactivating RNA-binding proteins which are crucial for innate immunity [180, 238]. DENV 3′UTRs possess RNA structures

*Dengue Fever in a One Health Perspective*

RNA *in vitro* [179]. sfRANs are abundant noncoding RNA sequences derived from the stalling of the host 5′-3′ exoribonuclease XRN1/Pacman in the 3′-untranslated regions (3′UTRs) of the viral genomic RNA [180]. sfRNAs have been shown to block exonuclease XRN1, increasing the overall messenger RNA stability within the host cell which may also benefit the viral RNA [181–183]. However, evidence for RNAi contribution to mammalian antiviral defense are few and still controversial [179]. miRNAs have been shown to regulate TLRs and IL-1 signaling pathways in response to viral infection, which provides control of host innate immune responses [184]. So far, there have been reported several cellular miRNAs (miRNAome) that are modulated during DENV infection of mammalian cells, mainly related to the regulation of IFN-β signaling pathways [185–188]. Some of these miRNAs have been proposed to be used as biomarkers in dengue-infected patients [189, 190]. Interestingly, modulation of microRNA expressions have been also described upon DENV infection of insect cells (e.g., C6/36 cells) as well as adult vector mosquitoes such as *Aedes albopictus* [191–193], suggesting that DENV might also regulate the activation of these antiviral RNAi pathways in vector mosquitoes, potentially avoiding viral clearance that promotes viral replication and transmission [194]. Despite RNAi constituting an evolutionarily conserved phenomenon of the mosquito-innate immune response to virus infections [195], viruses have found ways to subvert the RNAi-mediated antiviral responses in

vector mosquitoes to manipulate miRNA profiles to their own benefit [196].

**4.3 DENV subversion of antiviral responses**

Regarding the complement system, this multifaceted pathway has been shown to limit DENV replication; however, excessively activated complement components have been also associated with disease severity [197]. The complement cascade constitutes an integral component of the immune system, composed of many plasma proteins that once activated can initiate a proteolytic cascade, resulting in the release of chemokines, facilitation of particle phagocytosis via opsonization, and deposition of the cell-killing membrane attack complex (MAC) designed to target and destroy foreign pathogens such as viruses [174]. Activation of the complement system occurs via three convergent pathways: the classical, the lectin, and the alternative pathways [174]. *In vitro* experiments showed that DENV replication enhances complement activation [197–199]. Additionally, clinical and *in vivo* studies have shown that excessive consumption of some complement components (e.g., C3, C4, and factor B) contributed to severe manifestations by increasing the levels of complement-activated products which enhance vascular permeability to cause severe dengue disease [9, 108, 109, 200]. In fact, increased circulation of anaphylatoxins (C3a, C4a, and C5a) in the blood of severe patients correlated with symptoms of vascular leakage [109, 200]. In autopsy studies from children who died of acute severe dengue manifestations (DHF/DSS), augmented deposition of complement components from both classical and alternative pathways were found on hepatocytes which results in severe liver damage and death [120]. Altogether, these data support the hypothesis that exacerbated complement activation influences dengue disease immunopathogenesis leading to disease severity [197].

The first barrier to overcome for successful viral infection is the rapid innate immune responses of the host, including type I IFNs, inflammatory cytokines, complement responses, NK cells, apoptosis, and autophagy [201, 202]. These innate immune responses are meant to defeat viral infections by engaging specific viral components (e.g., RNA and DNA) leading to activation of immediate protective defense mechanisms such as the rapid recognition of PAMP in nonimmune and innate immune cells [161]. IFN production is a key goal of PRR activation for viral pathogens, and DENV is highly susceptible to effective induction of both type

**70**

necessary for viral genome cyclization, viral RNA synthesis, translation, and replication [239]. sfRNAs regulate the pathogenicity in both mammalian and mosquito cells after interacting with proteins such as TRIM25 to inhibit RIG-I signaling and translation of ISGs [73, 240, 241]. Interestingly, reduced IFN responses have been found during DENV outbreaks where the infecting DENV serotype produced greater levels of sfRNA than the less pathogenic strains [100, 240]. Thus, high levels of sfRNAs may cause an epidemiological fitness of DENV, which results in lower stimulation of RIG-IMDA5 RNA sensors and reduced production of IFN, causing higher viremia levels that could be translated in more infections and severe diseases.

On the other hand, DENV utilizes the endoplasmic reticulum (ER) of host cells for replication and assembly. In this process, the ER undergoes extensive rearrangements and expansion that requires *de novo* synthesis of viral proteins [71]. Accumulation of unfolded proteins in the ER lumen leads to an unfolded protein response (UPR), a pro-survival cellular reaction induced in response to DENV-mediated ER stress [242, 243]. DENV has evolved to manipulate the UPR to cope with ER stress which hijacks the host cell machinery to evade the host immunity, facilitating viral replication [112]. Distinct *in vitro* and *in vivo* studies have shown that DENV induced ER stress and manipulates the host metabolism and protein production by increasing the autophagy (lipophagy) activity, viral replication, and pathogenesis through UPR signaling pathways [244–247]. Autophagy is the lysosomal degradation of cytoplasmic contents, which results in the recycling of cellular macromolecules as well as the activation of cellular host responses to starvation or stress [248]. Autophagy has been implicated as an innate immune response that would engulf and destroy pathogens by degrading cytosolic contents [249, 250]. In DENV infection, functional autophagy components have been shown to either promote or restrict viral RNA replication and virus production [251–253]. However, DENV has found ways of preventing autophagic processing and degradation of viral components [254, 255]. Several studies have linked DENV induction of autophagy to the regulation of lipid metabolism, leading to increased degradation of lipid droplets that produces more fatty acid material important for viral replication [247]. In this process, the NS3, NS1, and C proteins of DENV have been found to increase fatty acid biosynthesis and recruitment of lipid droplets to the DENV replication complex, facilitating viral particle assembly [256, 257].

Furthermore, several studies have shown that lipids and lipoproteins play a role in modifying DENV infectivity in both mammal and insect cells *in vitro* [258, 259]. Modulation of cholesterol levels in the host cells facilitates viral entry, replication, virus assembly, and control type I IFN response [260, 261]. This modulation involves the regulation of cholesterol levels, expression of cholesterol receptors as well as changes in cholesterol synthesis related to important modifications in the cellular metabolism [114, 262, 263]. Interestingly, clinical studies have found that levels of total serum cholesterol and LDL-C levels are modulated over the course of dengue illness, with generally lower levels associated with increased dengue severity [264–266]. In general, low cholesterol levels have been associated with critical illness related to sepsis and vascular disorders [267]. Thus, the association of cholesterol with severe dengue outcome may be an important indicator of the pathophysiology of DHF/DSS.

About the complement pathway, DENV has evolved strategies to limit recognition and activation of the complement cascade [108, 165]. NS1 is the only flavivirus protein that is secreted by infected cells and has been shown to modulate the complement pathway [14, 268]. NS1 promotes efficient degradation of C4 to C4b to protect DENV from complement-dependent neutralization [13, 269]. The NS1 protein of DENV and other flaviviruses such as WNV NS1 interacts with some components of the alternative complement pathway such as the C3bBb convertase, which limits the formation of C5b-9 membrane attack complex (MAC) [268, 270]. Additional studies have found that NS1 proteins from DENV, WNV, and YFV all attenuate classical and

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*Dengue Immunopathogenesis: A Crosstalk between Host and Viral Factors Leading to Disease…*

lectin pathway activation by directly interacting with C4, which reduces C4b deposition and C3 convertase (C4b2a) activity [13, 271]. Also, anti-NS1 antibodies have been shown to induce complement consumption and C5b-9 generation [272]. Overall, through protein-to-protein interactions between the viral and host factors involved in antiviral responses and careful manipulation of cellular processes, such as ER expansion, autophagy and lipid metabolism, and complement pathways, DENV hijacks many host antiviral responses to facilitate virus replication leading to pathogenesis.

Dengue is the most prevalent arboviral disease transmitted by mosquitoes, which poses an enormous burden to the public health systems worldwide as more than 40% of the world population is at risk of infection. The infection with any of the four DENV serotypes (DENV1–4) can lead to a wide spectrum of clinical manifestations that range from the asymptomatic or inapparent to moderate flu-like symptoms, known as dengue fever (DF), and life-threatening manifestations identified by the WHO, known as the dengue hemorrhagic fever and dengue shock syndrome (DHF/ DSS), also known as severe dengue, with or without warning signs. In endemic areas where multiple DENV serotypes can seasonally circulate, distinct epidemiological studies have demonstrated that an individual human being can be exposed to sequential infections with distinct DENV serotypes, which poses a risk of developing severe manifestations such as DHF/DSS. This phenomenon has been attributed to the potential enhancement activity that the preexisting antibody response elicited from a previous infection with one serotype (e.g., DENV-1) may have on the infection with a different serotype (e.g., DENV-2). This process leads to an increased viral burden that triggers a series of immunological and cellular events (e.g., ADE, cytokine storm, skewed T cell responses, and complement pathways), which despite being intended to prevent the invasion and infection of the infecting viral pathogens, can induce host tissue damage leading to pathology and disease. The cellular and molecular mechanisms involved in this phenomenon will be explained in more detail in the Part II of this chapter entitled "Adaptive immune response and NS1 pathogenesis." As an arthropod-transmitted virus (arbovirus), DENV is initially transmitted by an infected vector mosquito in which the virus has already been amplified after replication in its distinct tissues, starting at the midgut to finalize in the salivary glands, where a new transmission cycle begins after blood feeding from a new host (**Figure 2**). Following inoculation from the bite of an infected mosquito, viruses undergo replication in the local tissues such as the skin. In the skin, infectious virus particles along with mosquito saliva components including proteases and immunomodulatory proteins among others are sown in the epidermis and dermis, leading to an activation of a cascade of events including the recruitment of skin resident cells (e.g., Langerhans cells, mast cells, and keratinocytes) and new cells (e.g., T cells and neutrophils) into the site of the infection that later serve as viral targets for viral replication. After infection of target cells, sensing of viral products (e.g., PAMPs and DAMPs) results in the activation of innate immune responses (e.g., type I IFN chemokines), the first line of defense, which establishes inflammatory and antiviral states intended to prevent the virus to colonize and to replicate in the skin; however, DENV has elaborated several pathogenic mechanisms to hijack these responses and escape from the normal immune system processing, which results in its dissemination and seeds into the lymph nodes. There, DENV further replicates in monocyte lineage cells, resulting in a primary viremia after its systemic dissemination through the circulatory bloodstream, which results in the subsequent infection of peripheral tissues such as the liver, spleen, and kidney. Overall, the skin represents not only the first line of

*DOI: http://dx.doi.org/10.5772/intechopen.93140*

**5. Concluding remarks (Part I)**

*Dengue Immunopathogenesis: A Crosstalk between Host and Viral Factors Leading to Disease… DOI: http://dx.doi.org/10.5772/intechopen.93140*

lectin pathway activation by directly interacting with C4, which reduces C4b deposition and C3 convertase (C4b2a) activity [13, 271]. Also, anti-NS1 antibodies have been shown to induce complement consumption and C5b-9 generation [272]. Overall, through protein-to-protein interactions between the viral and host factors involved in antiviral responses and careful manipulation of cellular processes, such as ER expansion, autophagy and lipid metabolism, and complement pathways, DENV hijacks many host antiviral responses to facilitate virus replication leading to pathogenesis.
