The Antimalaria Drug Artesunate Inhibits Porcine Reproductive and Respiratory Syndrome Virus Replication via Activating AMPK and Nrf2/HO-1 Signaling Pathways

2021 ◽  
Author(s):  
Feixiang Long ◽  
Mingxin Zhang ◽  
Xia Yang ◽  
Xiaohuan Liang ◽  
Lizhan Su ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Alveolar Macrophages ◽  
Energy Homeostasis ◽  
Ex Vivo ◽  
Antiviral Agents ◽  
Antimalarial Drugs ◽  
Effective Drug ◽  
Respiratory Syndrome Virus ◽  
Pork Industry ◽  
Novel Therapeutics

Porcine Reproductive and Respiratory Syndrome virus (PRRSV) causes significant economic losses to the pork industry worldwide. Currently, vaccine strategies provide limited protection against PRRSV transmission, and no effective drug is commercially available. Therefore, there is an urgent need to develop novel antiviral strategies to prevent PRRSV pandemics. This study showed that artesunate (AS), one of the antimalarial drugs, potently suppressed PRRSV replication in Marc-145 cells and ex vivo primary porcine alveolar macrophages (PAMs) at micromolar concentrations. Furthermore, we demonstrated that this suppression was closely associated with AS-activated AMPK (energy homeostasis) and Nrf2/HO-1 (inflammation) signaling pathways. AS treatment promoted p-AMPK, Nrf2 and HO-1 expression, and thus inhibited PRRSV replication in Marc-145 and PAM cells in a time- and dose-dependent manner. These effects of AS were reversed when AMPK or HO-1 gene was silenced by siRNA. In addition, we demonstrated that AMPK works upstream of Nrf2/HO-1 as its activation by AS is AMPK-dependent. Adenosine phosphate analysis showed that AS activates AMPK via improving AMP/ADP:ATP ratio rather than direct interaction with AMPK. Altogether, our findings indicate that AS could be a promising novel therapeutics for controlling PRRSV and that its anti-PRRSV mechanism, which involves the functional link between energy homeostasis and inflammation suppression pathways, may provide opportunities for developing novel antiviral agents. Importance Porcine reproductive and respiratory syndrome virus (PRRSV) infections have been continuously threatened the pork industry worldwide. Vaccination strategies provide very limited protection against PRRSV infection, and no effective drug is commercially available. We show that artesunate (AS), one of the antimalarial drugs, is a potent inhibitor against PRRSV replication in Marc-145 cells and ex vivo primary porcine alveolar macrophages (PAMs). Furthermore, we demonstrate that AS inhibits PRRSV replication via activation of AMPK-dependent Nrf2/HO-1 signaling pathways, revealing a novel link between energy homeostasis (AMPK) and inflammation suppression (Nrf2/HO-1) during viral infection. Therefore, we believe that AS may be a promising novel therapeutics for controlling PRRSV, and its anti-PRRSV mechanism may provide a potential strategy to develop novel antiviral agents.

Hyperactivation of RAP1 and JAK/STAT Signaling Pathways Contributes to Fibrosis during the Formation of Nasal Capsular Contraction

2021 ◽  
pp. 1-12
Author(s):  
Meng Wu ◽  
Ming Li ◽  
Hong-Ju Xie ◽  
Hong-Wei Liu
Keyword(s):  
Signaling Pathways ◽  
Type I Collagen ◽  
Ex Vivo ◽  
Capsular Contracture ◽  
Silicone Implant ◽  
Type I ◽  
Loss Of Function ◽  
Sequencing Data ◽  
Functional Changes ◽  
Stat Signaling

Silicone implant-based augmentation rhinoplasty or mammoplasty induces capsular contracture, which has been acknowledged as a process that develops an abnormal fibrotic capsule associated with the immune response to allogeneic materials. However, the signaling pathways leading to the nasal fibrosis remain poorly investigated. We aimed to explore the molecular mechanism underlying the pathogenesis of nasal capsular contracture, with a specific research interest in the signaling pathways involved in fibrotic development at the advanced stage of contracture. By examining our recently obtained RNA sequencing data and global gene expression profiling between grade II and grade IV nasal capsular tissues, we found that both the RAP1 and JAK/STAT signaling pathways were hyperactive in the contracted capsules. This was verified on quantitative real-time PCR which demonstrated upregulation of most of the representative component signatures in these pathways. Loss-of-function assays through siRNA-mediated Rap1 silencing and/or small molecule-directed inhibition of JAK/STAT pathway in ex vivo primary nasal fibroblasts caused a series of dramatic behavioral and functional changes, including decreased cell viability, increased apoptosis, reduced secretion of proinflammatory cytokines, and synthesis of type I collagen, compared to control cells, and indicating the essential role of the RAP1 and JAK/STAT signaling pathways in nasal capsular fibrosis. Our results sheds light on targeting downstream signaling pathways for the prevention and therapy of silicone implant-induced nasal capsular contracture.

scholarly journals Lung macrophages drive mucus production and steroid-resistant inflammation in chronic bronchitis

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Kristina Andelid ◽  
Karolina Öst ◽  
Anders Andersson ◽  
Esha Mohamed ◽  
Zala Jevnikar ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Chronic Bronchitis ◽  
Cross Talk ◽  
Alveolar Macrophages ◽  
Ex Vivo ◽  
Bronchial Epithelial Cells ◽  
Steroid Resistance ◽  
Bronchial Epithelial ◽  
Steroid Resistant ◽  
Copd Patients

Abstract Background Patients with chronic obstructive pulmonary disease (COPD) frequently suffer from chronic bronchitis (CB) and display steroid-resistant inflammation with increased sputum neutrophils and macrophages. Recently, a causal link between mucus hyper-concentration and disease progression of CB has been suggested. Methods In this study, we have evaluated the steroid sensitivity of purified, patient-derived sputum and alveolar macrophages and used a novel mechanistic cross-talk assay to examine how macrophages and bronchial epithelial cells cross-talk to regulate MUC5B production. Results We demonstrate that sputum plug macrophages isolated from COPD patients with chronic bronchitis (COPD/CB) are chronically activated and only partially respond to ex vivo corticosteroid treatment compared to alveolar macrophages isolated from lung resections. Further, we show that pseudo-stratified bronchial epithelial cells grown in air–liquid-interface are inert to direct bacterial lipopolysaccharide stimulation and that macrophages are able to relay this signal and activate the CREB/AP-1 transcription factor complex and subsequent MUC5B expression in epithelial cells through a soluble mediator. Using recombinant protein and neutralizing antibodies, we identified a key role for TNFα in this cross-talk. Conclusions For the first time, we describe ex vivo pharmacology in purified human sputum macrophages isolated from chronic bronchitis COPD patients and identify a possible basis for the steroid resistance frequently seen in this population. Our data pinpoint a critical role for chronically activated sputum macrophages in perpetuating TNFα-dependent signals driving mucus hyper-production. Targeting the chronically activated mucus plug macrophage phenotype and interfering with aberrant macrophage-epithelial cross-talk may provide a novel strategy to resolve chronic inflammatory lung disease.

Aging does not alter the mechanosensitivity of the p38, p70S6k, and JNK2 signaling pathways in skeletal muscle

2005 ◽  
Vol 98 (4) ◽  
pp. 1562-1566 ◽  
Author(s):  
Troy A. Hornberger ◽  
R. D. Mateja ◽  
E. R. Chin ◽  
J. L. Andrews ◽  
K. A. Esser
Keyword(s):  
Skeletal Muscle ◽  
Signaling Pathways ◽  
Ex Vivo ◽  
Mechanical Stimuli ◽  
S6 Kinase ◽  
Diminished Capacity ◽  
Ribosomal S6 Kinase ◽  
Passive Stretch ◽  
Old Mice ◽  
Dependent Mechanism

The capacity for skeletal muscle to recover its mass following periods of unloading (regrowth) has been reported to decline with age. Although the mechanisms responsible for the impaired regrowth are not known, it has been suggested that aged muscles have a diminished capacity to sense and subsequently respond to a given amount of mechanical stimuli (mechanosensitivity). To test this hypothesis, extensor digitorum longus muscles from young (2–3 mo) and old (26–27 mo) mice were subjected to intermittent 15% passive stretch (ex vivo) as a source of mechanical stimulation and analyzed for alterations in the phosphorylation of stress-activated protein kinase (p38), ribosomal S6 kinase (p70S6k), and the p54 jun N-terminal kinase (JNK2). The results indicated that the average magnitude of specific tension (mechanical stimuli) induced by 15% stretch was similar in muscles from young and old mice. Young and old muscles also revealed similar increases in the magnitude of mechanically induced p38, p70S6k (threonine/serine 421/424 and threonine 389), and JNK2 phosphorylation. In addition, coincubation experiments demonstrated that the release of locally acting growth factors was not sufficient for the induction of JNK2 phosphorylation, suggesting that JNK2 was activated by a mechanical rather than a mechanical/growth factor-dependent mechanism. Taken together, the results of this study demonstrate that aging does not alter the mechanosensitivity of the p38, p70S6k, and JNK2 signaling pathways in skeletal muscle.

Porcine reproductive and respiratory syndrome virus infection upregulates negative immune regulators and T-cell exhaustion markers

2021 ◽  
Author(s):  
Jayeshbhai Chaudhari ◽  
Chia-Sin Liew ◽  
Jean-Jack M. Riethoven ◽  
Sarah Sillman ◽  
Hiep L. X. Vu
Keyword(s):  
T Cell ◽  
Fluorescent Protein ◽  
Ex Vivo ◽  
Acute Stage ◽  
Respiratory Syndrome Virus ◽  
Porcine Alveolar Macrophage ◽  
Host Immune System ◽  
T Cell Exhaustion ◽  
Swine Industry ◽  
Cell Exhaustion

Porcine alveolar macrophage (PAM) is one of the primary cellular targets for PRRSV, but less than 2% of PAMs are infected with the virus during the acute stage of infection. To comparatively analyze the host transcriptional response between PRRSV-infected PAMs and bystanders PAMs that remained uninfected but were exposed to the inflammatory milieu of an infected lung, pigs were infected with a PRRSV strain expressing green fluorescent protein (PRRSV-GFP) and GFP + (PRRSV infected) and GFP – (bystander) cells were sorted for RNA-sequencing (RNA-seq). Approximately 4.2% of RNA reads from GFP + and 0.06% reads from GFP – PAMs mapped to the PRRSV genome, indicating that PRRSV-infected PAMs were effectively separated from bystander PAMs. Further analysis revealed that inflammatory cytokines, interferon-stimulated genes, and antiviral genes were highly upregulated in GFP + as compared to GFP – PAMs. Importantly, negative immune regulators including NF-κB inhibitors (NFKBIA, NFKBID, NFKBIZ, and TNFAIP3), and T-cell exhaustion markers (PD-L1, PD-L2, IL10, IDO1, and TGFB2) were highly upregulated in GFP + cells as compared to GFP – cells. By using in situ hybridization assay, RNA transcripts of TNF and NF-κB inhibitors were detected in PRRSV-infected PAMs cultured ex vivo and lung sections of PRRSV-infected pigs during the acute stage of infection. Collectively, the results suggest that PRRSV infection upregulates expression of negative immune regulators and T-cell exhaustion markers in PAMs to modulate the host immune response. Our findings provide further insight into PRRSV immunopathogenesis. Importance PRRSV is widespread in many swine producing countries, causing substantial economic loses to the swine industry. PAM is considered the primary target for PRRSV replication in pigs. However, less than 2% of PAM from an acutely infected pigs are infected with the virus. In the present study, we utilized a PRRSV-GFP strain to infect pigs and sorted infected- and bystander- PAMs from the pigs during the acute stage of infection for transcriptome analysis. PRRSV infected PAMs showed a distinctive gene expression profile and contained many uniquely activated pathways compared to bystander PAMs. Interestingly, upregulated expression of and NF-κB signaling inhibitors and T-cell exhaustion molecules were observed in PRRSV-infected PAMs. Our findings provide additional knowledge on the mechanisms that PRRSV employs to modulate the host immune system.

Ex Vivo Plasmodium malariae Culture Method for Antimalarial Drugs Screen in the Field

2021 ◽  
Author(s):  
Laurent Dembele ◽  
Nouhoum Diallo ◽  
Fanta Sogore ◽  
Bintou Diarra ◽  
Fatoumata I. Ballo ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Culture Method ◽  
Plasmodium Malariae ◽  
Antimalarial Drugs

Modulation by dexamethasone of phospholipase A2 activities in endotoxemic guinea pigs

1995 ◽  
Vol 79 (4) ◽  
pp. 1271-1277 ◽  
Author(s):  
C. M. De Castro ◽  
M. F. Bureau ◽  
M. A. Nahori ◽  
C. H. Dumarey ◽  
B. B. Vargaftig ◽  
...  
Keyword(s):  
Bronchoalveolar Lavage ◽  
Phospholipase A2 ◽  
Alveolar Macrophages ◽  
Guinea Pigs ◽  
Ex Vivo ◽  
Neutrophil Infiltration ◽  
Tnf Alpha ◽  
Necrosis Factor Alpha ◽  
Intracellular Enzyme ◽  
Lps Challenge

One hour after lipopolysaccharide (LPS) administration (intravenous) in guinea pigs, alveolar macrophages are primed for an ex vivo increased secretion of arachidonic acid metabolites from the cyclooxygenase and the lipoxygenase pathways, with challenge by a second stimulus. At the same time, maximal levels of tumor necrosis factor-alpha (TNF-alpha) are observed in the circulation and in the bronchoalveolar lavage fluid. An extracellular form of phospholipase A2, corresponding probably to the low-molecular-mass type II enzyme, known to accumulate in inflammatory exudates, appears later in the serum of guinea pigs, to reach maximal levels 6 h after the LPS. Unlike the intracellular enzyme, extracellular phospholipase A2 is not increased by LPS in alveolar macrophages or in bronchoalveolar lavage fluids. After 24 h, at the time when neither TNF-alpha nor extracellular phospholipase A2 is present and priming of macrophages is over, maximal neutrophil infiltration is observed in the alveolar space of LPS-treated guinea pigs. Dexamethasone administered repeatedly during 3 days (subcutaneous) before the LPS challenge prevented both early events such as the macrophage priming and the TNF-alpha appearance and later events such as extracellular phospholipase A2 release and neutrophil recruitment.

scholarly journals Porcine Arterivirus Infection of Alveolar Macrophages Is Mediated by Sialic Acid on the Virus

2004 ◽  
Vol 78 (15) ◽  
pp. 8094-8101 ◽  
Author(s):  
Peter L. Delputte ◽  
Hans J. Nauwynck
Keyword(s):  
Sialic Acid ◽  
Alveolar Macrophages ◽  
Sialic Acids ◽  
Respiratory Syndrome Virus ◽  
Specific Monoclonal Antibody ◽  
Neuraminidase Treatment ◽  
Acid Binding Protein ◽  
Sialic Acid Binding ◽  
High Mannose ◽  
Binding Lectin

ABSTRACT Recently, we showed that porcine sialoadhesin (pSn) mediates internalization of the arterivirus porcine reproductive and respiratory syndrome virus (PRRSV) in alveolar macrophages (Vanderheijden et al., J. Virol. 77:8207-8215, 2003). In rodents and humans, sialoadhesin, or Siglec-1, has been described as a macrophage-restricted molecule and to specifically bind sialic acid moieties. In the current study, we investigated whether pSn is a sialic acid binding protein and, whether so, whether this property is important for its function as a PRRSV receptor. Using untreated and neuraminidase-treated sheep erythrocytes, we showed that pSn binds sialic acid. Furthermore, pSn-specific monoclonal antibody 41D3, which blocks PRRSV infection, inhibited this interaction. PRRSV attachment to and infection of porcine alveolar macrophages (PAM) were both shown to be dependent on the presence of sialic acid on the virus: neuraminidase treatment of virus but not of PAM blocked infection and reduced attachment. Enzymatic removal of all N-linked glycans on the virus with N-glycosidase F reduced PRRSV infection, while exclusive removal of nonsialylated N-linked glycans of the high-mannose type with endoglycosidase H had no significant effect. Free sialyllactose and sialic acid containing (neo)glycoproteins reduced infection, while lactose and (neo)glycoproteins devoid of sialic acids had no significant effect. Studies with linkage-specific neuraminidases and lectins indicated that α2-3- and α2-6-linked sialic acids on the virion are important for PRRSV infection of PAM. From these results, we conclude that pSn is a sialic acid binding lectin and that interactions between sialic acid on the PRRS virion and pSn are essential for PRRSV infection of PAM.

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