Leupeptin maintains redox homeostasis via targeting ROS-autophagy-inflammatory axis in LPS-stimulated macrophages and cytokines dichotomy in Con-A challenged lymphocyte.

Information regarding cellular anti-inflammatory and immunomodulatory attributes of leupeptin with respect to modulation of perturbed macrophage function and lymphocytes has not yet been delineated, particularly in the context of ROS-cytokines-autophagy-inflammatory signalling cascades. Therefore, the present study identified the attributes and mechanisms of leupeptin, from actinomycetes, in relation to excessive oxidative stress mediated disrupted immune homeostasis and inflammatory mechanism in activated macrophages and lymphocytes. Results revealed that leupeptin treatment showed noticeable inhibition in the production of NO, ROS, mitochondrial membrane potential and phagocytosis activity in LPS-stimulated macrophages. These findings were accompanied by reduction in TNF-α, IL-1ß, IL-6, IFN-γ/IL-10 ratio, endopeptidases, oxidative effectors (Cox-2, IL-5, IL-15, IL-17, COX-2), iNOS with concomitant increase in Arg 1, Msr 1 and Mrc - 1exprssion in leupeptin treatment. Additionally, compared to LPS-challenged cells, marked alleviation in MDC, lysotracker staining, beclin-1, LC3B expression, and enhanced p62 levels in leupeptin exposed cells indicate the reversal of impaired autophagy flux. Subsequently, oxi-inflammatory signalling analysis demonstrated p-PTEN, p-NF-κB, p-PI3K, p-Akt, p-p38, and ERK1/2 upregulation decisively thwarted by leupeptin administration. In silico analysis further implied its target selectivity to these cascades. Furthermore, decreased proliferation index and Th1, Th2/IL-10 cytokines ratio in mitogen-challenged splenic lymphocytes confers its role in mitigating unwarranted inflammation mediated by disrupted regulation of adaptive immune cells. Together, these findings signify the attributes of leupeptin as an alternative anti-inflammatory strategy and affirm it as a promising natural entity to modulate immune-mediated response during inflammatory disorder.

Combating the Progression of Novel Coronavirus SARS-CoV-2 Infectious Disease: Current State and Future Prospects in Molecular Diagnostics and Drug Discovery.

A highly infectious and life-threatening virus was first reported in Wuhan, China, in late 2019, and it rapidly spread all over the world. This novel virus belongs to the coronavirus family and is associated with severe acute respiratory syndrome (SARS), causing respiratory disease known as COVID-19. In March 2020, WHO has declared the COVID-19 outbreak a global pandemic. Its morbidity and mortality rates are swiftly rising day by day, with the situation becoming more severe and fatal for the comorbid population. Many COVID-19 patients are asymptomatic, but they silently spread the infection. There is a need for proper screening of infected patients to prevent the epidemic transmission of disease and for early curative interventions to reduce the risk of developing severe complications from COVID-19. To date, the diagnostic assays are of two categories, molecular detection of viral genetic material by real-time RTpolymerase chain reaction and serological test, which relies on detecting antiviral antibodies. Unfortunately, there are no effective prophylactics and therapeutics available against COVID-19. However, a few drugs have shown promising antiviral activity against it, and these presently are being referred for clinical trials, albeit FDA has issued an Emergency Use Authorization (EUA) for the emergency use of a few drugs for SARSCoV- 2 infection. This review provides an insight into current progress, challenges and future prospects of laboratory detection methods of COVID-19, and highlights the clinical stage of the major evidence-based drugs/vaccines recommended against the novel SARS-CoV-2 pandemic virus.

Plant-derived immuno-adjuvants in vaccines formulation: a promising avenue for improving vaccines efficacy against SARS-CoV-2 virus.

The SARS-CoV-2 outbreak has posed a plethora of problems for the global healthcare system and socioeconomic burden. Despite valiant efforts to contain the COVID-19 outbreak, the situation has deteriorated to the point that there are no viable preventive therapies to treat this disease. The case count has skyrocketed globally due to the newly evolved variants. Despite vaccination drives, the re-occurrence of recent pandemic waves has reinforced the importance of innovation/utilization of immune-booster to achieve appropriate long-term vaccine protection. Plant-derived immuno-adjuvants, which have multifaceted functions, can impede infections by boosting the immune system. Many previous studies have shown that formulation of vaccines using plant-derived adjuvant results in long-lasting immunity may overcome the natural tendency of coronavirus immunity to wane quickly. Plant polysaccharides, glycosides, and glycoprotein extracts have reportedly been utilized as enticing adjuvants in experimental vaccines, such as Advax, Matrix-M, and Mistletoe lectin, which have been shown to be highly immunogenic and safe. When employed in vaccine formulation, Advax and Matrix-M generate long-lasting antibodies, a balanced robust Th1/Th2 cytokine profile, and the stimulation of cytotoxic T cells. Thus, the use of adjuvants derived from plants may increase the effectiveness of vaccines, resulting in the proper immunological response required to combat COVID-19. A few have been widely used in epidemic outbreaks, including SARS and H1N1 influenza, and their use could also improve the efficacy of COVID-19 vaccines. In this review, the immunological adjuvant properties of plant compounds as well as their potential application in anti-COVID-19 therapy are thoroughly discussed.

Berberis lycium fruit extract and its phytoconstituents berberine and rutin mitigate collagen-CFA-induced arthritis (CIA) via improving GSK3ß/STAT/Akt/MAPKs/NF-κB signaling axis mediated oxi-inflammation and joint articular damage in murine model.

Rheumatoid arthritis (RA), a chronic auto-immune disease, is often result of persistent and misdirectional inflammation and cannot be effectually resolved by single-target selective drugs. Present study attempted to uncover anti-arthritic efficacy and governing molecular mechanism of BLFE and its phytoconstituents berberine and rutin, with focus on dysregulated oxi-inflammation and structural integrity during articular damage using Collagen II-CFA-induced RA mice model. NMR-based phytometabolomic analysis revealed presence of phenolics and alkaloids such as berberine and rutin. BLFE, rutin and berberine remarkably mitigated Collagen II-CFA-induced disease severity index, articular damage, immune cells influx and pannus formation. An effective decrease in levels of TNF-α, IL-6, IL-1ß, IFN-γ, IL-13, IL-17, MMPs, RORγt, Ob-cadherin, Cox-2, iNOS and enhancement in IL-10, IL-4 and IL-5, BMP-6/7 was observed in BLFE, rutin and berberine treatments. Molecular mechanistic analysis demonstrated reduction in expression of p-STAT-1/3, p-PI3K, p-Akt, p-JNK, p-p38, p-IκB, p-NF-κB and ß-catenin via BLFE, rutin and berberine. Furthermore, reduced activation of p-ERK and p-GSK3ß and enhanced splenic Tregs was only noticed in BLFE and berberine. Thus, the signifying presence of these phytoconstituents could contribute to the above-mentioned findings. These findings imply that BLFE could be beneficial for assuaging deleterious aspects of RA mediated via perturbed inflammation.

Agnuside mitigates OVA-LPS induced perturbed lung homeostasis via modulating inflammatory, autophagy, apoptosis-fibrosis response and myeloid lineages in mice model of allergic asthma.

Attributes of agnuside, a nontoxic, iridoid glycoside have been advocated for inflammatory disorders. However, information on its efficacy in alleviating allergic asthma largely remain ambiguous and yet to be deciphered. Present study aimed to assess efficacy of agnuside in targeting vicious circle of oxi-inflammation, autophagy and fibrosis, together with investigating its underlying molecular mechanism during OVA-LPS induced allergic asthma. Results revealed that agnuside showed prophylactic effect in assuaging asthmatic lung architecture impairment (p ≤ 0.01) as indicated by suppression of inflammatory cell infiltration, congestion, fibrosis, airway remodeling and alveolar collapse in OVA-LPS sensitized group. Decreased expression level (p ≤ 0.05) of allergic inflammatory mediators such as IgE, Th1/Th2, IL-4/IFN-γ, IL-4/IL-10, chemokines, endopeptidases and TGF-ß, Smad2/4, Caspase9/3, connexin 43/50 observed in agnuside treatments. Analysis of redox molecular signaling cascade and autophagic proteins revealed concurrent upregulation in p-NF-κB, p-PI3K, p-Akt, p-p38, p-Stat3 activation, GATA3, LC3B expression and reduction in Bcl2/Bax, Beclin1 and p62 expression in sensitized mice (p ≤ 0.05) which were intensely counteracted by administration of agnuside. Suppression in myeloid cells activation and augmentation (p ≤ 0.001) of Tregs established modulatory attribute of agnuside for innate and adaptive immune response during allergic asthma. Collectively, these outcomes confer prophylactic attribute of agnuside and signify it as promising strategy to thwart allergic asthma.

Pharmaco-immunomodulatory interventions for averting cytokine storm-linked disease severity in SARS-CoV-2 infection.

The year 2020 is characterised by the COVID-19 pandemic that has quelled more than half a million lives in recent months. We are still coping with the negative repercussions of COVID-19 pandemic in 2021, in which the 2nd wave in India resulted in a high fatality rate. Regardless of emergency vaccine approvals and subsequent meteoric global vaccination drives in some countries, hospitalisations for COVID-19 will continue to occur due to the propensity of mutation in SARS-CoV-2 virus. The immune response plays a vital role in the control and resolution of infectious diseases. However, an impaired immune response is responsible for the severity of the respiratory distress in many diseases. The severe COVID-19 infection persuaded cytokine storm that has been linked with acute respiratory distress syndrome (ARDS), culminates into vital organ failures and eventual death. Thus, safe and effective therapeutics to treat hospitalised patients remains a significant unmet clinical need. In that state, any clue of possible treatments, which save patients life, can be treasured for this time point. Many cohorts and clinical trial studies demonstrated that timely administration of immunomodulatory drugs on severe COVID-19 patients may mitigate the disease severity, hospital stay and mortality. This article addresses the severity and risk factors of hypercytokinemia in COVID-19 patients, with special emphasis on prospective immunomodulatory therapies.

Vitexin restores lung homeostasis by targeting vicious loop between inflammatory aggravation and autophagy mediated via multiple redox cascade and myeloid cells alteration in experimental allergic asthma.

BACKGROUND: Allergic asthma is one of the leading respiratory diseases with complex pathology. Attributes of vitexin, a trihydroxyflavone, has been studied to alleviate Th2 cytokines response in allergic asthma. However, its efficacy and underlying mechanism in mitigating allergic asthma particularly mediated by oxi-inflammatory stress, autophagy and apoptosis, yet to be delineated. PURPOSE: Present study aimed to decipher efficacy and governing molecular mechanism of vitexin in mitigating allergic asthma particularly mediated by vicious loop of oxi-inflammatory stress, autophagy and apoptosis. METHODS: To ascertain this, OVA-LPS induced mice model was used and protective attributes of vitexin for different mediators, pathological facets and sensing pathways of allergic asthma were evaluated. RESULTS: Vitexin treatment remarkably inhibited OVA-LPS induced inflammatory cell infiltration, mast cell activation, alveolar collapse, congestion, fibrosis in lung architecture. These results were accompanied by suppression of immune cells hyperactivation, mucus secretion, goblet cell proliferation, persistent inflammation which were affirmed by alleviation in levels of IgE, Th1/Th2/Th17, IL-4/IFN-γ, chemokines, endopeptidases (MMP-1, MMP-13), oxidative effectors with concomitant increase in IL-15, IL-10, MMP-9 and MMP-3. Additionally, noticeable decline in p-connexin 43, p-c-Fos, TGF-ß, Smad2/3/4, Caspase9/3, LC3A/B expression and upregulation in beclin-1, p62 co-localization and Bcl2/Bax indicate reversal of lung vascular permeability, mast cell degranulation, fibrosis, apoptosis, autophagosome impairment. Subsequent allergic inflammatory cascades analysis revealed p-NF-κB, p-PI3K, p-Akt, p-p38, p-Stat3, GATA3 upregulation and p-PTEN downregulation in sensitized mice, which were decisively counteracted by vitexin. In silico studies signified target specificity of vitexin with these proteins. Suppression in myeloid cells activation and enhancements of Tregs demonstrated immunomodulatory potential of vitexin in allergic airways. CONCLUSION: Collectively, to our knowledge, this is the first report that confers vitexin meditated multi-faceted protective attribute in mitigation of allergic asthma that could be linked to its suppressive effects on vicious cycle of pathological process particularly regulated via oxi-inflammation, autophagy and apoptosis. Thus, signify vitexin as safe therapeutic strategy.

Rutin prevents inflammation-associated colon damage via inhibiting the p38/MAPKAPK2 and PI3K/Akt/GSK3ß/NF-κB signalling axes and enhancing splenic Tregs in DSS-induced murine chronic colitis.

A large body of emerging evidence has revealed the role of p38/MK2 and PI3K/Akt/GSK3ß cascades in the orchestrating process of colitis. Rutin, a bioflavonoid present in many fruits and vegetables, has been recognized to offer therapeutic attributes in acute colitis. However, its role in chronic colitic condition has not yet been delineated in reference to p38/MK2 and PI3K/Akt/GSK3ß signalling. The present investigation assessed the efficacy and underlying molecular mechanism of rutin in alleviating DSS-induced chronic colitis. The analysis of signalling pathways demonstrated the robust activation of PI3K/Akt/GSK3ß/MAPKs/NF-κB and p38/MK2 in DSS-induced colitis in animals, which was efficiently alleviated following the rutin treatment. In silico studies indicated its target specificity with these pathways. Rutin administration markedly improved the disease activity score, colon length, goblet cell loss and compromised colon epithelial integrity in colitic mice. Decreased expression of oxi-inflammatory markers such as IgM, IgE, iNOS, ICAM-1, HO-1 and Th1/IL-10 cytokines ratios after treatment suggests its efficacy in regulating effector, regulatory and B cell homeostasis. Additionally, rutin demonstrated its role in restoring epithelial integrity by modulating the transcript levels of tight junction proteins, mucus-secreting proteins, epithelial cell proliferation and apoptosis. Treg expansion revealed that rutin supplementation also exhibits an immune regulatory potential and suppresses inflammatory aggravation mediated by adaptive immune responses. Overall, results indicate that the modulation of p38/MK2 and PI3K/Akt/GSK3ß/NF-κB pathways by rutin represents a novel therapeutic approach in chronic colitis that help to curb dysregulated intestinal integrity, cytokine ratio and splenic Tregs.

Vitex negundo Linn. extract alleviates inflammatory aggravation and lung injury by modulating AMPK/PI3K/Akt/p38-NF-κB and TGF-ß/Smad/Bcl2/caspase/LC3 cascade and macrophages activation in murine model of OVA-LPS induced allergic asthma.

ETHNOPHARMACOLOGICAL RELEVANCE: There is growing inclination towards developing bioactive molecule-based strategies for the management of allergic airway inflammation associated respiratory diseases. Vitex negundo Linn., also known as Nirgundi, is one such medicinal plant enriched with phytochemicals and used for inflammatory and respiratory disorders including asthma in traditional system of medicine. Preliminary studies have claimed anti-tussive and bronchodilator potential of V. negundo Linn. However, its attributes as well as molecular mechanism (s) in modulation of asthma mediated by allergic inflammation are yet to be delineated scientifically. AIM OF THE STUDY: Present study attempted to assess the effectiveness of Vitex negundo leaf extract (VNLE) in mitigation of allergen induced inflammation associated asthmatic lung damage with emphasis to delineate its molecular mechanism (s). MATERIALS AND METHODS: Allergic lung inflammation was established in Balb/c mice using Ovalbumin-lipopolysaccharide (OVA-LPS). Several allergic inflammatory parameters, histopathological changes, alveolar macrophage activation and signalling pathways were assessed to examine protective effects of VNLE. UHPLC-DAD-QTOF-ESI-IMS was used to characterize VLNE. RESULTS: VNLE administration effectively attenuated LPS-induced oxi-inflammatory stress in macrophages suggesting its anti-inflammatory potential. Further, VNLE showed protective effect in mitigating asthmatic lung damage as evident by reversal of pathological changes including inflammatory cell influx, congestion, fibrosis, bronchial thickness and alveolar collapse observed in allergen group. VNLE suppressed expressions of inflammatory Th1/Th2 cytokines, chemokines, endopeptidases (MMPs), oxidative effector enzyme (iNOS), adhesion molecules, IL-4/IFN-γ release with simultaneous enhancement in levels of IL-10, IFN-γ, MUC3 and tight junction proteins. Subsequent mechanistic investigation revealed that OVA-LPS concomitantly enhanced phosphorylation of NF-κB, PI3K, Akt and p38MAPKs and downregulated AMPK which was categorically counteracted by VNLE treatment. VNLE also suppressed OVA-LPS induced fibrosis, apoptosis, autophagy and gap junction proteins which were affirmed by reduction in TGF-ß, Smad2/3/4, Caspase9/3, Bax, LC3A/B, connexin 50, connexin 43 and enhancement in Bcl2 expression. Additionally, suppression of alveolar macrophage activation, inflammatory cells in blood and elevation of splenic CD8+T cells was demonstrated. UHPLC-DAD-QTOF-ESI-IMS revealed presence of iridoids glycoside and phenolics which might contribute these findings. CONCLUSION: These findings confer protective effect of VNLE in attenuation of allergic lung inflammation and suggest that it could be considered as valuable medicinal source for developing safe natural therapeutics for mitigation of allergic inflammation during asthma.

Berberis lycium fruit extract attenuates oxi-inflammatory stress and promotes mucosal healing by mitigating NF-κB/c-Jun/MAPKs signalling and augmenting splenic Treg proliferation in a murine model of dextran sulphate sodium-induced ulcerative colitis.

PURPOSE: There is a growing interest in developing phytomolecule-based therapies for the management of inflammatory disorders owing to rising cost of treatments and unwarranted effects. The present work attempted to assess the efficacy and mechanisms of Berberis lycium Royle fruit extract (BLFE) in mitigating chemical-induced colitis in mice. METHODS: Colitis was induced in Balb/C mice using dextran sulphate sodium (DSS) and protective effects of BLFE were examined. Several oxi-inflammatory parameters, histopathological changes, epithelial barrier integrity and activation of NF-κB/c-Jun/MAPKs in colon tissue were determined. Splenic T cell subpopulations were also gauged to evaluate the systemic effects of BLFE in the modulation of immune responses. RESULTS: BLFE treatment effectively improved animal survival rate, DAI score, colon length and structural damage in DSS-exposed mice. Expression of oxi-inflammatory markers such as MPO, IgE, iNOS, ICAM-1, MCP-1 and RANTES as well as Th1/Th2/Th17 cytokines were decreased in BLFE treated animals. On the other hand, an increased mRNA expression of anti-inflammatory cytokines (IL-4/IL-10), tight junction proteins and IgA levels were also observed during BLFE treatment. BLFE appeared to modulate intestinal epithelial cell proliferation (PCNA) and apoptosis (Bcl2/Bax), thereby suggesting its role in the maintenance of intestinal integrity. Analysis of inflammatory signalling pathways indicated robust activation and expression of NF-κB/c-Jun/MAPKs (JNK and p38) in DSS treated animal which was strongly abrogated by BLFE treatment. BLFE supplementation also enhanced the proliferation of CD3+CD4+CD25+ Treg cells indicating suppression of inflammatory activation. CONCLUSION: These observations provide compelling evidence that BLFE could be considered as a viable natural strategy in the prevention and management of ulcerative colitis.

MAPKAPK2 plays a crucial role in the progression of head and neck squamous cell carcinoma by regulating transcript stability.

BACKGROUND: Head and neck squamous-cell carcinoma (HNSCC) ranks sixth among cancers worldwide. Though several molecular mechanisms of tumor initiation and progression of HNSCC are known, others remain unclear. Significance of p38/MAPKAPK2 (Mitogen-activated protein kinase-activated protein kinase-2) pathway in cell stress and inflammation is well established and its role in tumor development is being widely studied. METHODS: We have elucidated the role of MAPKAPK2 (MK2) in HNSCC pathogenesis using clinical tissue samples, MK2-knockdown (MK2KD) cells and heterotropic xenograft mice model. RESULTS: In patient-derived tissue samples, we observed that MK2 is reproducibly overexpressed. Increased stability of cyclin-dependent kinase inhibitor 1B (p27), mitogen-activated protein kinase phosphatase-1 (MKP-1) transcripts and decreased half-life of tumor necrosis factor-alpha (TNF-α) and vascular endothelial growth factor (VEGF) transcripts in MK2KD cells suggests that MK2 regulates their transcript stability. In vivo xenograft experiments established that knockdown of MK2 attenuates course of tumor progression in immunocompromised mice. CONCLUSION: Altogether, MK2 is responsible for regulating the transcript stability and is functionally important to modulate HNSCC pathogenesis.