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CHIKV induced TNF facilitates TCR driven T cell activation in vitro. Both CHIKV infected and mock cell culture supernatants were harvested and used to culture for T cells activation assay in vitro. (A) Dot plot analysis showing the expression of CD69 in different conditions. (B) Graphical representation depicting the percent positive cells for CD69 in T cells. Data represent mean ± SEM of three independent experiments. p < 0.05 was considered as statistically significant difference between the groups. (**p ≤ 0.01; ***p ≤ 0.001).
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Chikungunya virus (CHIKV), a mosquito-borne Alphavirus, is endemic in different parts of the globe. The host macrophages are identified as the major cellular reservoirs of CHIKV during infection and this virus triggers robust TNF production in the host macrophages, which might be a key mediator of virus induced inflammation. However, the molecular...
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Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes persistent arthritis in a subset of human patients. We report the isolation and functional characterization of monoclonal antibodies (mAbs) from two patients infected with CHIKV in the Dominican Republic. Single B cell sorting yielded a panel of 46 human mAbs of diverse germ...
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... Several clinical and experimental studies have revealed that the Chikungunya virus (CHIKV) infection leads to the profound production of pro-inflammatory cytokines and chemokines such as tumor necrosis factor (TNF), interleukin (IL)-6, 4, 1b and 12 in human as well as in mouse macrophages via p38 and Jun Nterminal protein kinase (JNK)-mitogen-activated protein kinase (MAPK) mediated pathway, which may aggravate host immune system towards CHIKV infection mediated fever (CHIKF) and polyarthralgia (19)(20)(21)(22). However, the initial pathways behind CHIKV-driven pro-inflammatory responses are still unexplored. ...
... CHIKV-induced host cell activation and a rise in associated pro-inflammatory responses are already reported by us and others (19,22,33). Earlier studies have revealed that the pro-inflammatory cytokines along with MAPKs are induced during CHIKV infection in the host macrophages (19,22). ...
... CHIKV-induced host cell activation and a rise in associated pro-inflammatory responses are already reported by us and others (19,22,33). Earlier studies have revealed that the pro-inflammatory cytokines along with MAPKs are induced during CHIKV infection in the host macrophages (19,22). Since TLR4 activation could be connected with TNF response and MAPK activation (34,35), the possible interaction of TLR4 with CHIKV infection along with subsequent regulation of host immune responses, if any, needs to be explored. ...
Toll like receptor 4 (TLR4), a pathogen-associated molecular pattern (PAMP) receptor, is known to exert inflammation in various cases of microbial infection, cancer and autoimmune disorders. However, any such involvement of TLR4 in Chikungunya virus (CHIKV) infection is yet to be explored. Accordingly, the role of TLR4 was investigated towards CHIKV infection and modulation of host immune responses in the current study using mice macrophage cell line RAW264.7, primary macrophage cells of different origins and in vivo mice model. The findings suggest that TLR4 inhibition using TAK-242 (a specific pharmacological inhibitor) reduces viral copy number as well as reduces the CHIKV-E2 protein level significantly using p38 and JNK-MAPK pathways. Moreover, this led to reduced expression of macrophage activation markers like CD14, CD86, MHC-II and pro-inflammatory cytokines (TNF, IL-6, MCP-1) significantly in both the mouse primary macrophages and RAW264.7 cell line, in vitro. Additionally, TAK-242-directed TLR4 inhibition demonstrated a significant reduction of percent E2-positive cells, viral titre and TNF expression in hPBMC-derived macrophages, in vitro. These observations were further validated in TLR4-knockout (KO) RAW cells. Furthermore, the interaction between CHIKV-E2 and TLR4 was demonstrated by immuno-precipitation studies, in vitro and supported by molecular docking analysis, in silico. TLR4-dependent viral entry was further validated by an anti-TLR4 antibody-mediated blocking experiment. It was noticed that TLR4 is necessary for the early events of viral infection, especially during the attachment and entry stages. Interestingly, it was also observed that TLR4 is not involved in the post-entry stages of CHIKV infection in host macrophages. The administration of TAK-242 decreased CHIKV infection significantly by reducing disease manifestations, improving survivability (around 75%) and reducing inflammation in mice model. Collectively, for the first time, this study reports TLR4 as one of the novel receptors to facilitate the attachment and entry of CHIKV in host macrophages, the TLR4-CHIKV-E2 interactions are essential for efficient viral entry and modulation of infection-induced pro-inflammatory responses in host macrophages, which might have translational implication for designing future therapeutics to regulate the CHIKV infection.
... ii) Inhibition of macrophages polarization. It has been shown that small molecule inhibitors of c-Jun amino-terminal kinase (JNK) play protective roles in kidney injury in different forms of kidney disease by inhibiting renal inflammation, apoptosis, and fibrosis (101). JNK inhibitors (CC-401) can suppress M1 polarization and maintain effectiveness in renal IRI induced fibrotic models (102). ...
Acute kidney injury (AKI) is a renal disease with a high incidence and mortality. Currently, there are no targeted therapeutics for preventing and treating AKI. Macrophages, important players in mammalian immune response, are involved in the multiple pathological processes of AKI. They are dynamically activated and exhibit a diverse spectrum of functional phenotypes in the kidney after AKI. Targeting the mechanisms of macrophage activation significantly improves the outcomes of AKI in preclinical studies. In this review, we summarize the role of macrophages and the underlying mechanisms of macrophage activation during kidney injury, repair, regeneration, and fibrosis and provide strategies for macrophage-targeted therapies.
... Both enzymes are promising targets for the development of small molecule inhibitors which can be used in anti-inflammatory therapy [28]. Moreover, it was reported earlier by our group that P38MAPK is crucial for CHIKV infection [29]. Besides, our group has formerly reported an Indian outbreak strain IS, to exhibit a faster replication rate than the CHIKV prototype strain, PS in vitro [30]. ...
Chikungunya virus (CHIKV) epidemics around the world have created public health concern with the unavailability of effective drugs and vaccines. This emphasizes the need for molecular understanding of host-virus interactions for developing effective targeted antivirals. Microarray analysis was carried out using CHIKV strain (Prototype and Indian) infected Vero cells and two host isozymes, MAPK activated protein kinase 2 (MK2) and MAPK activated protein kinase 3 (MK3) were selected for further analysis. The substrate spectrum of both enzymes is indistinguishable and covers proteins involved in cytokines production, endocytosis, reorganization of the cytoskeleton, cell migration, cell cycle control, chromatin remodeling and transcriptional regulation. Gene silencing and drug treatment were performed in vitro and in vivo to unravel the role of MK2/MK3 in CHIKV infection. Gene silencing of MK2 and MK3 abrogated around 58% CHIKV progeny release from the host cell and a MK2 activation inhibitor (CMPD1) treatment demonstrated 68% inhibition of viral infection suggesting a major role of MAPKAPKs during late CHIKV infection in vitro . Further, it was observed that the inhibition in viral infection is primarily due to the abrogation of lamellipodium formation through modulation of factors involved in the actin cytoskeleton remodeling pathway. Moreover, CHIKV-infected C57BL/6 mice demonstrated reduction in the viral copy number, lessened disease score and better survivability after CMPD1 treatment. In addition, reduction in expression of key pro-inflammatory mediators such as CXCL13, RAGE, FGF, MMP9 and increase in HGF (a CHIKV infection recovery marker) was observed indicating the effectiveness of the drug against CHIKV. Taken together it can be proposed that MK2 and MK3 are crucial host factors for CHIKV infection and can be considered as important target for developing effective anti-CHIKV strategies.
... Additional viral proteins modulating p38 signaling have been described. Chikungunya virus protein nsP2 interacted with phos-p38 and phos-JNK protein [56]. Herpes simplex virus type-1 (HSV-1) virion transactivator protein VP16 stimulated p38/JNK activation, however the mechanism of action is not known [57]. ...
Viruses have evolved mechanisms to subvert critical cellular signaling pathways that regulate a wide range of cellular functions, including cell differentiation, proliferation and chemotaxis, and innate immune responses. Here, we describe a novel ORFV protein, ORFV113, that interacts with the G protein-coupled receptor Lysophosphatidic acid receptor 1 (LPA 1 ). Consistent with its interaction with LPA 1 , ORFV113 enhances p38 kinase phosphorylation in ORFV infected cells in vitro and in vivo , and in cells transiently expressing ORFV113 or treated with soluble ORFV113. Infection of cells with virus lacking ORFV113 (OV-IA82Δ113) significantly decreased p38 phosphorylation and viral plaque size. Infection of cells with ORFV in the presence of a p38 kinase inhibitor markedly diminished ORFV replication, highlighting importance of p38 signaling during ORFV infection. ORFV113 enhancement of p38 activation was prevented in cells in which LPA 1 expression was knocked down and in cells treated with LPA 1 inhibitor. Infection of sheep with OV-IA82Δ113 led to a strikingly attenuated disease phenotype, indicating that ORFV113 is a major virulence determinant in the natural host. Notably, ORFV113 represents the first viral protein that modulates p38 signaling via interaction with LPA 1 receptor.
... However, these changes could also decrease T cell functionality by impacting other relevant functions. For instance, p38 is involved in the induction of senescence and IFNg and TNFa secretion (188), and its inhibition have diminished these cytokines in different inflammation or virus infection models (189,190). Moreover, blockage of DDR and p53 involves a risk of DNA damage on T cells that might induce malignancy (191). ...
Adoptive cellular immunotherapy using chimeric antigen receptor (CAR)-modified T cells and Natural Killer (NK) cells are common immune cell sources administered to treat cancer patients. In detail, whereas CAR-T cells induce outstanding responses in a subset of hematological malignancies, responses are much more deficient in solid tumors. Moreover, NK cells have not shown remarkable results up to date. In general, immune cells present high plasticity to change their activity and phenotype depending on the stimuli they receive from molecules secreted in the tumor microenvironment (TME). Consequently, immune cells will also secrete molecules that will shape the activities of other neighboring immune and tumor cells. Specifically, NK cells can polarize to activities as diverse as angiogenic ones instead of their killer activity. In addition, tumor cell phagocytosis by macrophages, which is required to remove dying tumor cells after the attack of NK cells or CAR-T cells, can be avoided in the TME. In addition, chemotherapy or radiotherapy treatments can induce senescence in tumor cells modifying their secretome to a known as “senescence-associated secretory phenotype” (SASP) that will also impact the immune response. Whereas the SASP initially attracts immune cells to eliminate senescent tumor cells, at high numbers of senescent cells, the SASP becomes detrimental, impacting negatively in the immune response. Last, CAR-T cells are an attractive option to overcome these events. Here, we review how molecules secreted in the TME by either tumor cells or even by immune cells impact the anti-tumor activity of surrounding immune cells.
... Also, it has been shown that alphaviruses, such as Sindbis virus, can induce an early phosphorylation of eIF2α, leading to global translational shut-off [26]. The role of the PI3K-AKT-mTOR and the p38 MAPK pathways have been studied in CHIKV infection [27,28]. However, much information is lacking on the cross-talk between the PI3K-Akt-mTOR, RAS-RAF-MEK-ERK and p38 MAPK pathways, which are key players in regulating cellular cap-dependent translation, and autophagy during CHIKV infection. ...
The 5′ capped, message-sense RNA genome of Chikungunya virus (CHIKV) utilizes the host cell machinery for translation. Translation is regulated by eIF2 alpha at the initiation phase and by eIF4F at cap recognition. Translational suppression by eIF2 alpha phosphorylation occurs as an early event in many alphavirus infections. We observe that in CHIKV-infected HEK293 cells, this occurs as a late event, by which time the viral replication has reached an exponential phase, implying its minimal role in virus restriction. The regulation by eIF4F is mediated through the PI3K-Akt-mTOR, p38 MAPK and RAS-RAF-MEK-ERK pathways. A kinetic analysis revealed that CHIKV infection did not modulate AKT phosphorylation, but caused a significant reduction in p38 MAPK phosphorylation. It caused degradation of phospho-ERK 1/2 by increased autophagy, leaving the PI3K-Akt-mTOR and p38 MAPK pathways for pharmacological targeting. mTOR inhibition resulted in moderate reduction in viral titre, but had no effect on CHIKV E2 protein expression, indicating a minimal role of the mTOR complex in virus replication. Inhibition of p38 MAPK using SB202190 caused a significant reduction in viral titre and CHIKV E2 and nsP3 protein expression. Furthermore, inhibiting the two pathways together did not offer any synergism, indicating that inhibiting the p38 MAPK pathway alone is sufficient to cause restriction of CHIKV replication. Meanwhile, in uninfected cells the fully functional RAS-RAF-MEK-ERK pathway can circumvent the effect of p38 MAPK inhibition on cap-dependent translation. Thus, our results show that host-directed antiviral strategies targeting cellular p38 MAPK are worth exploring against Chikungunya as they could be selective against CHIKV-infected cells with minimal effects on uninfected host cells.
... Importantly, the observed effects of MAPK inhibitors on MAYV replication did not appear to result from virucidal activity by these compounds. Previous reports have shown that p38 inhibition affects the replication of viruses from different families, among them, enterovirus 71, hepatitis C virus, influenza virus, Chikungunya virus and SARS-CoV-2 [27,35,[51][52][53]. ...
Mayaro virus (MAYV) hijacks the host´s cell machinery to effectively replicate. The mitogen-activated protein kinases (MAPKs) p38, JNK and ERK1/2 have emerged as crucial cellular factors implicated in different stages of the viral cycle. However, whether MAYV uses these MAPKs to competently replicate has not yet been determined. The aim of this study was to evaluate the impact of MAPKs inhibition on MAYV replication using primary human dermal fibroblasts (HDFs) and HeLa cells. Viral yields in supernatants from MAYV-infected cells treated or untreated with inhibitors SB203580, SP600125, U0126 or Losmapimod were quantified using plaque assay. Also, viral protein expression was analyzed using immunoblot and immunofluorescence. Knockdown of p38⍺/p38β isoforms was performed in HDFs using the PROTACs molecule NR-7h. Our data demonstrated that HDFs are highly susceptible to MAYV infection. SB203580, a p38 inhibitor, reduced MAYV replication in a dose-dependent manner in both HDFs and HeLa cells. Additionally, SB203580 significantly decreased viral E1 protein expression. Similarly, knockdown or inhibition of p38⍺/p38β isoforms with NR-7h or Losmapimod, respectively, affected MAYV replication in a dose-dependent manner. Collectively, these findings suggest that p38 could play an important role in MAYV replication and could serve as a therapeutic target to control MAYV infection.
... Methods used to assess rheumatoid arthritis are often applied to other diseases, such as CHIKV arthritis [107]. CHIKV frequently involves the small joints of the [72,[76][77][78] Macrophage and B cell activation, Th1 differentiation, increased MHC 1 and MHC II TNF [72,79,80] Pro-inflammatory signaling, neutrophil activation IL-1β [72,81,82] Pro-inflammatory signaling, Th17 differentiation IL-6 [72,77,81,[83][84][85][86] Th17 differentiation, T reg inhibition, synthesis of acute phase proteins, stimulation of antibody production IL-8 [72,77,86,87] Neutrophil chemotaxis IL-10 [72,81,88,89] Suppresses expression of pro-inflammatory cytokines, inhibition of neutrophil recruitment IL-12 [72,90] Th1 differentiation, IFNγ synthesis in NK and T cells CCL2 [56,72,86] Recruits T cells, monocytes, and dendritic cells to sites of inflammation hands and feet and as such some instruments may underestimate that the severity of the arthritis may be due to lack of inclusion of these joints. In contrast to RA, swollen joints in CHIKV arthritis are poor predictors of outcomes while joint pain and stiffness may be independently associated with disability and quality of life assessments. ...
Purpose of Review
Persistent joint pain is a common manifestation of arthropod-borne viral infections and can cause long-term disability. We review the epidemiology, pathophysiology, diagnosis, and management of arthritogenic alphavirus infection.
Recent findings
The global re-emergence of alphaviral outbreaks has led to an increase in virus-induced arthralgia and arthritis. Alphaviruses, including Chikungunya, O’nyong’nyong, Sindbis, Barmah Forest, Ross River, and Mayaro viruses, are associated with acute and/or chronic rheumatic symptoms. Identification of Mxra8 as a viral entry receptor in the alphaviral replication pathway creates opportunities for treatment and prevention. Recent evidence suggesting virus does not persist in synovial fluid during chronic chikungunya infection indicates that immunomodulators may be given safely.
Summary
The etiology of persistent joint pain after alphavirus infection is still poorly understood. New diagnostic tools along and evidence-based treatment could significantly improve morbidity and long-term disability.
... In macrophages, downregulation of JNK phosphorylation using natural compounds like Esculentoside B (Esb), inhibited inflammatory response by reducing the secretion of proinflammatory cytokines, iNOS, COX-2, TNF-α, IL-1β, and IL-6 [17]. Another study used chikungunya virus (CHIKV), a mosquito-borne Alphavirus to induce the production of TNF-α in macrophages that were mediated by JNK and p38 MAPK pathways [18]. Others examined fluctuations in glucose concentration on the activation of TLR4-JNK pathway in mediating diabetes-related inflammation in macrophages using THP-1 cells (human monocytes) [19]. ...
The western diet and overuse of anti-inflammatory medication have caused a great deal of stress on the liver. Obesity and the associated inflammatory state in insulin-responsive tissues result in the release of pro-inflammatory cytokine that activates the stress-responsive MAPKs, p38 MAPK, and JNK. These MAPKs have figured prominently as critical effectors in physiological and pathophysiological hepatic inflammation. In contrast, evidence for a role for ERK1/2 in hepatic inflammation has been less well developed. In this review article, we describe recent insights into the physiology and pathophysiology of the role of stress-responsive MAPKs in hepatic inflammation during obesity and liver injury with a focus on macrophages, hepatocytes and hepatic stellate cells. In response to metabolic stress and liver injury, JNK activation in macrophages and hepatocytes promotes the secretion of inflammatory cytokines and macrophage and neutrophil infiltration. p38 MAPK plays an important role in contributing to the progression of hepatic inflammation in response to various hepatic cellular stresses, although the precise substrates mediating these effects in hepatocytes and hepatic stellate cells remain to be identified. Both JNK and p38 MAPK promotes profibrotic behavior in hepatic stellate cells.
... CHIKV infects a wide range of both immune and non-immune cells. Amongst them, monocytes/macrophages play important roles in modulating adaptive immune responses [35,[37][38][39]. ...
... CHIKV-infection-associated immune responses in the host macrophages have been reported recently [34,35,38,39,54,55]. However, the potential involvement of TRPV1 during CHIKV infection in host macrophages has not been investigated. ...
... The RAW 264.7 (ATCC® TIB-71™) mouse monocyte/macrophage cell line was maintained in Roswell Park Memorial Institute 1640 medium (RPMI-1640; PAN Biotech, Aidenbach Germany) supplemented with antibiotics, including penicillin (100 U/ml) and streptomycin (0.1 mg/ml), 2.0 mM L-glutamine (Himedia Laboratories Pvt. Ltd., Mumbai, India), and 10% heat-inactivated fetal bovine serum (FBS; PAN Biotech, Aidenbach Germany) at 37 °C in a humidified incubator with 5% CO 2 . Vero cells (an African green monkey kidney epithelial cell line), were maintained in Dulbecco's modified Eagle's medium (DMEM; Himedia) with the same supplements and conditions listed above [35,39,56]. ...
Chikungunya virus (CHIKV), a virus that induces pathogenic inflammatory host immune responses, is re-emerging worldwide, and there are currently no established antiviral control measures. Transient receptor potential vanilloid 1 (TRPV1), a non-selective Ca²⁺-permeable ion channel, has been found to regulate various host inflammatory responses including several viral infections. Immune responses to CHIKV infection in host macrophages have been reported recently. However, the possible involvement of TRPV1 during CHIKV infection in host macrophages has not been studied. Here, we investigated the possible role of TRPV1 in CHIKV infection of the macrophage cell line RAW 264.7. It was found that CHIKV infection upregulates TRPV1 expression in macrophages. To confirm this observation, the TRPV1-specific modulators 5ˊ-iodoresiniferatoxin (5ˊ-IRTX, a TRPV1 antagonist) and resiniferatoxin (RTX, a TRPV1 agonist) were used. Our results indicated that TRPV1 inhibition leads to a reduction in CHIKV infection, whereas TRPV1 activation significantly enhances CHIKV infection. Using a plaque assay and a time-of-addition assay, it was observed that functional modulation of TRPV1 affects the early stages of the viral lifecycle in RAW 264.7 cells. Moreover, CHIKV infection was found to induce of pNF-κB (p65) expression and nuclear localization. However, both activation and inhibition of TRPV1 were found to enhance the expression and nuclear localization of pNF-κB (p65) and production of pro-inflammatory TNF and IL-6 during CHIKV infection. In addition, it was demonstrated by Ca²⁺ imaging that TRPV1 regulates Ca²⁺ influx during CHIKV infection. Hence, the current findings highlight a potentially important regulatory role of TRPV1 during CHIKV infection in macrophages. This study might also have broad implications in the context of other viral infections as well.
