SARS-CoV-2 infection disrupts trophoblast function and placental syncytial barrier integrity via its accessory protein, ORF3a

Problem: Although it is rare for a SARS-CoV-2 infection to transmit vertically to the fetus during pregnancy, there is a significantly increased risk of adverse pregnancy outcomes due to maternalCOVID- 19. However, there is a poor understanding of such risks because mechanistic studies on how SARS-CoV-2 infection disrupts placental homeostasis are significantly lacking. The SARS-CoV-2 proteome includes multiple structural and non-structural proteins, including the non-structural accessory proteinORF3a. The roles of these proteins in mediating placental infection remain undefined. We and others have shown that autophagy activity in placental syncytium is essential for barrier function and integrity. Here, we have used clinical samples and cultured trophoblast cells to evaluate syncytial integrity of placenta exposed to SARS-CoV-2. The objective of our study was to investigate potential mechanisms through which SARS-CoV-2 impairs placental homeostasis and causes adverse pregnancy outcomes. We tested the central hypothesis that an essential SARS-CoV-2 non-structural and accessory protein, ORF3a, uniquely (amongst multiple viral proteins tested) with a novel three-dimensional structure andwith no homology to any other proteins is a key modulator of placental trophoblast cell dynamics via autophagy and intracellular trafficking of a tight junction protein (TJP), ZO-1. Method(s): We used clinical samples and cultured trophoblast cells to evaluate syncytial integrity of placentas exposed to SARS-CoV- 2. Autophagic flux was measured in placental villous biopsies from SARS-CoV-2-exposed and unexposed pregnant women by quantifying the expression of autophagy markers, LC3 and P62. Trophoblast cells (JEG-3, Forskolin-treated JEG-3, HTR8/SVneo, or primary human trophoblasts (PHTs)) were transfected with expression plasmids encoding SARS-CoV-2 proteins including ORF3a. Using western blotting, multi-label immunofluorescence, and confocal imaging, we analyzed the effect of ORF3a on the autophagy, differentiation, invasion, and intracellular trafficking of ZO-1 in trophoblasts. Using coimmunoprecipitation assays, we tested ORF3a interactions with host proteins. t-tests and one-way analyses of variance (ANOVAs) with post hoc tests were used to assess the data, with significance set at P < .05. Result(s): We discovered :1) increased activation of autophagy, but incomplete processing of autophagosome-lysosomal degradation;2) accumulation of protein aggregates in placentas exposed to SARS-CoV- 2. Mechanistically, we showed that the SARS-CoV-2 ORF3a protein, uniquely 3) blocks the autophagy-lysosomal degradation process;4) inhibits maturation of cytotrophoblasts into syncytiotrophoblasts (STBs);5) reduces production ofHCG-beta, a key pregnancy hormone that is also essential for STB maturation;and 6) inhibits trophoblast invasive capacity. Furthermore, ORF3a harbors an intrinsically disordered C-terminus withPDZ-bindingmotifs.We show for the first time that, 7) ORF3a binds to and co-localizes with the PDZ domain of ZO-1, a junctional protein that is essential for STB maturation and the integrity of the placental barrier. Conclusion(s): Our work outlines a new molecular and cellular mechanism involving the SARS-CoV-2 accessory protein ORF3a that may drive the virus's ability to infect the placenta and damage placental syncytial integrity. This implies that the mechanisms facilitating viral maturation, such as the interaction of ORF3a with host factors, can be investigated for additional functionality and even targeted for developing new intervention strategies for treatment or prevention of SARS-CoV-2 infection at the maternal-fetal interface.

Olmesartan alleviates SARS-CoV-2 envelope protein induced renal fibrosis by regulating HMGB1 release and autophagic degradation of TGF-ß1.

Background and aims: Renal damage in severe coronavirus disease 2019 (COVID-19) is highly associated with mortality. Finding relevant therapeutic candidates that can alleviate it is crucial. Angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin-receptor blockers (ARBs) have been shown to be harmless to COVID-19 patients, but it remains elusive whether ACEIs/ARBs have protective benefits to them. We wished to determine if ACEIs/ARBs had a protective effect on the renal damage associated with COVID-19, and to investigate the mechanism. Methods: We used the envelope (E) protein of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) to induce COVID-19-like multiple organ damage and observed renal fibrosis. We induced the epithelial-mesenchymal transformation of HK-2 cells with E protein, and found that olmesartan could alleviate it significantly. The protective effects of olmesartan on E protein-induced renal fibrosis were evaluated by renal-function assessment, pathologic alterations, inflammation, and the TGF-ß1/Smad2/3 signaling pathway. The distribution of high-mobility group box (HMGB)1 was examined after stimulation with E protein and olmesartan administration. Results: E protein stimulated HMGB1 release, which triggered the immune response and promoted activation of TGF-ß1/Smad2/3 signaling: both could lead to renal fibrosis. Olmesartan regulated the distribution of HMGB1 under E protein stimulation. Olmesartan inhibited the release of HMGB1, and reduced the inflammatory response and activation of TGF-ß1/Smad2/3 signaling. Olmesartan increased the cytoplasmic level of HMGB1 to promote the autophagic degradation of TGF-ß1, thereby alleviating fibrosis further. Conclusion: Olmesartan alleviates E protein-induced renal fibrosis by regulating the release of HMGB1 and its mediated autophagic degradation of TGF-ß1.

Porcine epidemic diarrhoea virus (PEDV) infection activates AMPK and JNK through TAK1 to induce autophagy and enhance virus replication.

Autophagy plays an important role in defending against invading microbes. However, numerous viruses can subvert autophagy to benefit their replication. Porcine epidemic diarrhoea virus (PEDV) is an aetiological agent that causes severe porcine epidemic diarrhoea. How PEDV infection regulates autophagy and its role in PEDV replication are inadequately understood. Herein, we report that PEDV induced complete autophagy in Vero and IPEC-DQ cells, as evidenced by increased LC3 lipidation, p62 degradation, and the formation of autolysosomes. The lysosomal protease inhibitors chloroquine (CQ) or bafilomycin A and Beclin-1 or ATG5 knockdown blocked autophagic flux and inhibited PEDV replication. PEDV infection activated AMP-activated protein kinase (AMPK) and c-Jun terminal kinase (JNK) by activating TGF-beta-activated kinase 1 (TAK1). Compound C (CC), an AMPK inhibitor, and SP600125, a JNK inhibitor, inhibited PEDV-induced autophagy and virus replication. AMPK activation led to increased ULK1^S777 phosphorylation and activation. Inhibition of ULK1 activity by SBI-0206965 (SBI) and TAK1 activity by 5Z-7-Oxozeaenol (5Z) or by TAK1 siRNA led to the suppression of autophagy and virus replication. Our study provides mechanistic insights into PEDV-induced autophagy and how PEDV infection leads to JNK and AMPK activation.

Neuromuscular disease: 2022 update.

This review highlights ten important advances in the neuromuscular disease field that were reported in 2021. As with prior updates in this article series, the overarching topics include (i) advances in understanding of fundamental neuromuscular biology; (ii) new / emerging diseases; (iii) advances in understanding of disease etiology and pathogenesis; (iii) diagnostic advances; and (iv) therapeutic advances. Within this general framework, the individual disease entities that are discussed in more detail include neuromuscular complications of COVID-19 (another look at the topic first covered in the 2021 review), autosomal recessive myopathy caused by MLIP mutations, autosomal recessive neuromuscular disease caused by VWA1 mutations, Leber's hereditary optic neuropathy, myopathies with autophagic defects, tRNA synthetase-associated Charcot-Marie-Tooth disease, systemic sclerosis-associated myopathy, humoral immune endoneurial microvasculopathy, and late-onset Pompe disease. In addition, the review highlights a few other advances (including new insights into mechanisms of muscle and nerve regeneration and the use of gene expression profiling to better characterize different subtypes of immune-mediated myopathies) that will be of significant interest for clinicians and researchers who specialize in neuromuscular disease.

Cellular Metabolism: A Fundamental Component of Degeneration in the Nervous System.

It is estimated that, at minimum, 500 million individuals suffer from cellular metabolic dysfunction, such as diabetes mellitus (DM), throughout the world. Even more concerning is the knowledge that metabolic disease is intimately tied to neurodegenerative disorders, affecting both the central and peripheral nervous systems as well as leading to dementia, the seventh leading cause of death. New and innovative therapeutic strategies that address cellular metabolism, apoptosis, autophagy, and pyroptosis, the mechanistic target of rapamycin (mTOR), AMP activated protein kinase (AMPK), growth factor signaling with erythropoietin (EPO), and risk factors such as the apolipoprotein E (APOE-ε4) gene and coronavirus disease 2019 (COVID-19) can offer valuable insights for the clinical care and treatment of neurodegenerative disorders impacted by cellular metabolic disease. Critical insight into and modulation of these complex pathways are required since mTOR signaling pathways, such as AMPK activation, can improve memory retention in Alzheimer's disease (AD) and DM, promote healthy aging, facilitate clearance of ß-amyloid (Aß) and tau in the brain, and control inflammation, but also may lead to cognitive loss and long-COVID syndrome through mechanisms that can include oxidative stress, mitochondrial dysfunction, cytokine release, and APOE-ε4 if pathways such as autophagy and other mechanisms of programmed cell death are left unchecked.

Strategies for the Management of Spike Protein-Related Pathology.

In the wake of the COVID-19 crisis, a need has arisen to prevent and treat two related conditions, COVID-19 vaccine injury and long COVID-19, both of which can trace at least part of their aetiology to the spike protein, which can cause harm through several mechanisms. One significant mechanism of harm is vascular, and it is mediated by the spike protein, a common element of the COVID-19 illness, and it is related to receiving a COVID-19 vaccine. Given the significant number of people experiencing these two related conditions, it is imperative to develop treatment protocols, as well as to consider the diversity of people experiencing long COVID-19 and vaccine injury. This review summarizes the known treatment options for long COVID-19 and vaccine injury, their mechanisms, and their evidentiary basis.

The metabolic effects of APOL1 in humans.

Harboring apolipoprotein L1 (APOL1) variants coded by the G1 or G2 alleles of the APOL1 gene increases the risk for collapsing glomerulopathy, focal segmental glomerulosclerosis, albuminuria, chronic kidney disease, and accelerated kidney function decline towards end-stage kidney disease. However, most subjects carrying APOL1 variants do not develop the kidney phenotype unless a second clinical condition adds to the genotype, indicating that modifying factors modulate the genotype-phenotype correlation. Subjects with an APOL1 high-risk genotype are more likely to develop essential hypertension or obesity, suggesting that carriers of APOL1 risk variants experience more pronounced insulin resistance compared to noncarriers. Likewise, arterionephrosclerosis (the pathological correlate of hypertension-associated nephropathy) and glomerulomegaly take place among carriers of APOL1 risk variants, and these pathological changes are also present in conditions associated with insulin resistance, such as essential hypertension, aging, and diabetes. Insulin resistance may contribute to the clinical features associated with the APOL1 high-risk genotype. Unlike carriers of wild-type APOL1, bearers of APOL1 variants show impaired formation of lipid droplets, which may contribute to inducing insulin resistance. Nascent lipid droplets normally detach from the endoplasmic reticulum into the cytoplasm, although the proteins that enable this process remain to be fully defined. Wild-type APOL1 is located in the lipid droplet, whereas mutated APOL1 remains sited at the endoplasmic reticulum, suggesting that normal APOL1 may participate in lipid droplet biogenesis. The defective formation of lipid droplets is associated with insulin resistance, which in turn may modulate the clinical phenotype present in carriers of APOL1 risk variants.

Endomembrane remodeling in SARS-CoV-2 infection.

During severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, the viral proteins intimately interact with host factors to remodel the endomembrane system at various steps of the viral lifecycle. The entry of SARS-CoV-2 can be mediated by endocytosis-mediated internalization. Virus-containing endosomes then fuse with lysosomes, in which the viral S protein is cleaved to trigger membrane fusion. Double-membrane vesicles generated from the ER serve as platforms for viral replication and transcription. Virions are assembled at the ER-Golgi intermediate compartment and released through the secretory pathway and/or lysosome-mediated exocytosis. In this review, we will focus on how SARS-CoV-2 viral proteins collaborate with host factors to remodel the endomembrane system for viral entry, replication, assembly and egress. We will also describe how viral proteins hijack the host cell surveillance system-the autophagic degradation pathway-to evade destruction and benefit virus production. Finally, potential antiviral therapies targeting the host cell endomembrane system will be discussed.

Establishment and validation of the autophagy-related ceRNA network in irreversible pulpitis.

BACKGROUND: The molecular mechanisms underlying the onset and progression of irreversible pulpitis have been studied for decades. Many studies have indicated a potential correlation between autophagy and this disease. Against the background of the competing endogenous RNA (ceRNA) theory, protein-coding RNA functions are linked with long noncoding RNAs (lncRNAs) and microRNAs (miRNAs). This mechanism has been widely studied in various fields but has rarely been reported in the context of irreversible pulpitis. The hub genes selected under this theory may represent the key to the interaction between autophagy and irreversible pulpitis. RESULTS: Filtering and differential expression analyses of the GSE92681 dataset, which contains data from 7 inflamed and 5 healthy pulp tissue samples, were conducted. The results were intersected with autophagy-related genes (ARGs), and 36 differentially expressed ARGs (DE-ARGs) were identified. Functional enrichment analysis and construction of the protein‒protein interaction (PPI) network of DE-ARGs were performed. Coexpression analysis was conducted between differentially expressed lncRNAs (DElncRNAs) and DE-ARGs, and 151 downregulated and 59 upregulated autophagy-related DElncRNAs (AR-DElncRNAs) were identified. StarBase and multiMiR were then used to predict related microRNAs of AR-DElncRNAs and DE-ARGs, respectively. We established ceRNA networks including 9 hub lncRNAs (HCP5 and AC112496.1 ↑; FENDRR, AC099850.1, ZSWIM8-AS1, DLX6-AS1, LAMTOR5-AS1, TMEM161B-AS1 and AC145207.5 ↓), which were validated by a qRT‒PCR analysis of pulp tissue from patients with irreversible pulpitis. CONCLUSION: We constructed two networks consisting of 9 hub lncRNAs based on the comprehensive identification of autophagy-related ceRNAs. This study may provide novel insights into the interactive relationship between autophagy and irreversible pulpitis and identifies several lncRNAs that may serve as potential biological markers.

Respiratory Syncytial Virus (RSV) infections in children: recent concepts in immunopathogenesis and diagnostic dilemma

Respiratory viral infections are important cause of morbidity and mortality in early life. The relative influence of host and viral factors possibly contribute to the disease pathogenesis. Predisposing conditions like prematurity, Low birth weight and congenital heart diseases etc. have been incriminated in the disease progression. The development of cough, wheezing, and tachypnea, usually peaking on days 4 to 5, go parallel with host cytokine responses and viral load. Various host cytokines, chemokines and molecules involved in the immune response against RSV infection might be responsible for the outcome of the disease process. Nasopharyngeal aspirates (NPAs) from children (n = 349) between 2013-2017 were subjected for IL-17A, IFN-gamma, TNF-alpha, IL-10, IL-6 levels by CBA and MMP-9 and TIMP-1 levels by ELISA. The viral load in RSV positive samples and cytokine levels were correlated with the WHO criteria for acute lower respiratory tract illness (ALRTI). RSV viral load, Pro-inflammatory cytokine (TNF-alpha) levels in severe ALRTI patients were significantly higher than the ALRTI patients [p<0.001]. Whereas Th17 cytokine (IL-17) was found to be significantly higher (p<0.05) in ALRTI patients than severe patients. MMP-9 is secreted in higher levels in severe ALRTI patients (n = 77) in comparison to Acute LRTI patients (n = 35) with an increase of thirty seven fold (p<0.001). Thus, the study highlights the role of TNF -alpha, IL-17 and Th2 cytokine biasness in the pathogenesis of RSV disease with the possible contribution of higher MMP-9/TIMP-1 ratio as a bad prognostic marker towards disease severity. To study the gene expression of autophagy and mTOR signalling pathways in RSV infected children with ALRTI. Nasopharyngeal aspirate (NPA) samples (n = 145) from children suffering from ALRTI were subjected for detection of RSV (Oct 2019 to March 2020). Semi-quantitative gene expression analysis for 5 representative genes each of mTOR signalling and autophagy pathway were performed in respiratory tract epithelial cells using 25 RSV positive cases and 10 healthy controls subjects. Autophagy gene expression analysis revealed significant upregulation in NPC1 and ATG3 autophagy genes. mTOR, AKT1 and TSC1 genes of mTOR pathway were significantly down-regulated in RSV positive patients except RICTOR gene which was significantly upregulated. Thus, survival of RSV within autophagosome might have been facilitated by upregulation of autophagy and downregulation of mTOR signalling genes. To assess the impact of SARS-CoV2 pandemic on RSV, samples were collected from children with ALRTIs admitted to emergency, PICU and indoor admissions during pre-pandemic period (October 2019 to February 2020;n = 166) and during COVID-19 Pandemic (July 2021 to July 2022;n = 189, SARS-CoV2 negative). These NP swabs were analyzed for pdm InfA H1N1, InfA H3N2, Inf B, RSV, hMPV, hBoV, hRV, PIV-2 and PIV-3 by PCR. Higher proportion of children with ALRTIs have had virus/es isolated during pre-pandemic period than during pandemic period (p<0.001). During pre-pandemic period, significantly higher proportion of children had RSV positivity (p<0.001);and significantly lower positivity for hRV (p<0.05), hMPV (p<0.05), and hBoV (p <= 0.005). The occurrence of COVID-19 pandemic has significantly impacted the frequency and pattern of detection of RSV among hospitalized children with LRTIs. RSV Fusion protein plays a critical role in the entry of the virus into the host cell by initiating the fusion of host and viral membranes. It happens to be a target of neutralizing antibodies paving the way as a vaccine candidate. Hence effort was made to introduce point mutation in hRSV fusion protein which can confer stability in its prefusion form. In-silico a stable structure of RSV fusion protein was generated making it a potential vaccine candidate. The timely diagnosis of RSV infection in this population is important for initiating therapy and instituting appropriate infection prevention measures. Serological testing is not widely used for the diagnosis of RSV. C ll Cultures including shell vial culture were used for RSV diagnosis. However, culture approaches lack sensitivity, often quite significantly, compared to nucleic acid amplification assays for the diagnosis of RSV infections. Molecular multiplex assays now offer increased sensitivity for a more accurate diagnosis. However issues with the use of these types of commercial panel assays include the requirement for substantial training, quality systems, and infrastructure to maintain and run these assays and many a times identification of viruses where the true pathogenic potential of those multiple viruses are debatable. Studies are available with laboratory- developed nucleic acid amplification test systems for the detection of RSVA and RSVB in clinical specimens either by PCRbased technologies or RT-LAMP. Gene targets of laboratory-developed molecular assays point towards M gene and the N gene in RSVA and -B with the benefits of flexibility to modify assays when targets are under evolutionary pressure to change, as well as a perceived initial low cost to carry out testing.

AUTOPHAGY AND Acyl-CoA-BINDING PROTEIN INFLUENCE COVID-19 SEVERITY

Background: Autophagy, a cytosolic-structure degradation pathway, allows production of IL21 by CD4 T-cells and efficient cytolytic responses by CD8 T-cells. Autophagy is in part regulated by acyl-CoA-binding protein (ACBP) which has two functions. Intracellular ACBP favors autophagy, whereas secreted extracellular ACBP inhibits autophagy. Herein, we assessed whether autophagy and the ACBP pathway were associated with COVID-19 severity. Method(s): Through the BQC-19 Quebec biobank, somalogic proteomic analysis was performed on 5200 proteins in plasma samples collected between March 2020 and December 2021. Plasma from 903 patients (all data available) during the acute phase of COVID-19 were assessed. COVID-19 severity was stratified using WHO criteria. In vitro, ACBP intracellular levels, autophagy levels (LC3II) and IL21 production were assessed by flow in PBMCs after a 24h stimulation with IL6, phorbol myristate acetate (PMA)+ionomycin or lipopolysaccharide (LPS). Plasma levels of anti-SARS-CoV-2 (full spike protein or RBD) IgG were assessed by ELISA. Result(s): Median age of the cohort was 62 yo, 48% were female, 55% had comorbidities (see table). Increasing plasma levels of ACBP were found with severity (mild, moderate, severe and fatal groups having 5.3, 7.3, 9.5 and 10.6 RFU/50muL of plasma, respectively, p< 0.001 for all comparisons). Patients with comorbidities had higher plasma ACBP levels (7.4 vs 6.4 RFU/50muL, p< 0.001). Plasma ACBP levels were higher during the delta and omicron-variant periods (8.4 vs 6.8 RFU/50muL;p< 0.001). Plasma ACBP levels correlated with LC3II levels (r=0.51, P< 0.001) and IL6 (r=0.41, p< 0.001), but neither with markers IL1beta nor IL8. ACBP levels negatively correlated with IL21 levels (r=-0.27, p< 0.001), independently of age, sex, and severity. ACBP levels were not associated with levels of anti-SARS-CoV-2 IgG levels. In vitro, IL6 stimulation of healthy control PBMC induced extracellular ACBP release. Moreover, adding recombinant ACBP: 1) reduced autophagy in lymphocytes and monocytes upon polyclonal stimulation with PMA/ionomycin or LPS;2) reduced intracellular production of IL21 in T-cells after PMA/ ionomycin stimulation. Conclusion(s): Plasma ACBP levels were inversely linked with IL21 levels, suggesting that autophagy and IL21 allow control of SARS-CoV-2 infection, independently of the level of SARS-CoV-2 antibody secretion. ACBP is a targetable autophagy checkpoint and its extracellular inhibition may improve SARS-CoV-2 immune control. (Table Presented).

Lysosomal nanotoxicity: Impact of nanomedicines on lysosomal function.

Although several nanomedicines got clinical approval over the past two decades, the clinical translation rate is relatively small so far. There are many post-surveillance withdrawals of nanomedicines caused by various safety issues. For successful clinical advancement of nanotechnology, it is of unmet need to realize cellular and molecular foundation of nanotoxicity. Current data suggest that lysosomal dysfunction caused by nanoparticles is emerging as the most common intracellular trigger of nanotoxicity. This review analyzes prospect mechanisms of lysosomal dysfunction-mediated toxicity induced by nanoparticles. We summarized and critically assessed adverse drug reactions of current clinically approved nanomedicines. Importantly, we show that physicochemical properties have great impact on nanoparticles interaction with cells, excretion route and kinetics, and subsequently on toxicity. We analyzed literature on adverse reactions of current nanomedicines and hypothesized that adverse reactions might be linked with lysosomal dysfunction caused by nanomedicines. Finally, from our analysis it becomes clear that it is unjustifiable to generalize safety and toxicity of nanoparticles, since different particles possess distinct toxicological properties. We propose that the biological mechanism of the disease progression and treatment should be central in the optimization of nanoparticle design.

Autophagy Receptor p62 Regulates SARS-CoV-2-Induced Inflammation in COVID-19.

As autophagy can promote or inhibit inflammation, we examined autophagy-inflammation interplay in COVID-19. Autophagy markers in the blood of 19 control subjects and 26 COVID-19 patients at hospital admission and one week later were measured by ELISA, while cytokine levels were examined by flow cytometric bead immunoassay. The antiviral IFN-α and proinflammatory TNF, IL-6, IL-8, IL-17, IL-33, and IFN-γ were elevated in COVID-19 patients at both time points, while IL-10 and IL-1ß were increased at admission and one week later, respectively. Autophagy markers LC3 and ATG5 were unaltered in COVID-19. In contrast, the concentration of autophagic cargo receptor p62 was significantly lower and positively correlated with TNF, IL-10, IL-17, and IL-33 at hospital admission, returning to normal levels after one week. The expression of SARS-CoV-2 proteins NSP5 or ORF3a in THP-1 monocytes caused an autophagy-independent decrease or autophagy-inhibition-dependent increase, respectively, of intracellular/secreted p62, as confirmed by immunoblot/ELISA. This was associated with an NSP5-mediated decrease in TNF/IL-10 mRNA and an ORF3a-mediated increase in TNF/IL-1ß/IL-6/IL-10/IL-33 mRNA levels. A genetic knockdown of p62 mimicked the immunosuppressive effect of NSP5, and a p62 increase in autophagy-deficient cells mirrored the immunostimulatory action of ORF3a. In conclusion, the proinflammatory autophagy receptor p62 is reduced inacute COVID-19, and the balance between autophagy-independent decrease and autophagy blockade-dependent increase of p62 levels could affect SARS-CoV-induced inflammation.

Autophagy and SARS-CoV-2-Old Players in New Games.

At present it is well-defined that autophagy is a fundamental process essential for cell life but its pro-viral and anti-viral role has been stated out with the COVID pandemic. However, viruses in turn have evolved diverse adaptive strategies to cope with autophagy driven host defense, either by blocking or hijacking the autophagy machinery for their own benefit. The mechanisms underlying autophagy modulation are presented in the current review which summarizes the accumulated knowledge on the crosstalk between autophagy and viral infections, with a particular emphasizes on SARS-CoV-2. The different types of autophagy related to infections and their molecular mechanisms are focused in the context of inflammation. In particular, SARS-CoV-2 entry, replication and disease pathogenesis are discussed. Models to study autophagy and to formulate novel treatment approaches and pharmacological modulation to fight COVID-19 are debated. The SARS-CoV-2-autophagy interplay is presented, revealing the complex dynamics and the molecular machinery of autophagy. The new molecular targets and strategies to treat COVID-19 effectively are envisaged. In conclusion, our finding underline the importance of development new treatment strategies and pharmacological modulation of autophagy to fight COVID-19.

SARS-CoV-2 Nsp8 induces mitophagy by damaging mitochondria.

Autophagy plays an important role in the interaction between viruses and host cells. SARS-CoV-2 infection can disrupt the autophagy process in target cells. However, the precise molecular mechanism is still unknown. In this study, we discovered that the Nsp8 of SARS-CoV-2 could cause an increasing accumulation of autophagosomes by preventing the fusion of autophagosomes and lysosomes. From further investigation, we found that Nsp8 was present on mitochondria and can damage mitochondria to initiate mitophagy. The results of experiments with immunofluorescence revealed that Nsp8 induced incomplete mitophagy. Moreover, both domains of Nsp8 orchestrated their function during Nsp8-induced mitophagy, in which the N-terminal domain colocalized with mitochondria and the C-terminal domain induced auto/mitophagy. This novel finding expands our understanding of the function of Nsp8 in promoting mitochondrial damage and inducing incomplete mitophagy, which helps us to understand the etiology of COVID-19 as well as open up new pathways for creating SARS-CoV-2 treatment methods.

A new autophagy-driven protective pathway of Haematococcus astaxanthin against chemical damage in wistar rats with gentamicin-induced acute kidney injury

Astaxanthin was established to conserve kidney function and subcellular structure through anti-oxidation and/or the free radical scavenging system, yet little research linked a new protective effect to autophagy or lysosomes. We pre-fed Wistar rats with natural astaxanthin, β-carotene, or placebo and induced acute kidney injury using gentamicin, before examining renal tissues and measuring physiological indices. Qualitative evidence from histopathological and subcellular images, along with quantitative evidence showing treatment effects on blood urea nitrogen and serum creatinine (p < 0.01), indicated that esterified Haematococcus astaxanthin surpassed β-carotene at effectively counteracting chemical damage and protecting the kidneys from injury. Proliferation of enlarged lysosomes and mediation analysis results revealing enhanced lysosomal acid phosphatase activity were consistent with the hypothesized autophagy-lysosomal pathway being up-regulated by astaxanthin intake (p < 0.05). In conclusion, the protective effect of astaxanthin against acute kidney injury exerted through the autophagy-lysosomal detoxification pathway, which totally different from the anti-oxidation and/or conventional SOD-dependent free radical scavenging system, was demonstrated with strong evidence. In light of the pandemic outbreak of novel coronavirus pneumonia associated with a virus preferentially targeting the renal tubular cells, dietary astaxanthin may help bring down incidence rate of coronavirus disease, cases of acute kidney injury secondary to the disease, and mortality rate from acute kidney injury, especially when a standard of care treatment for the infectious disease is pending. © 2023 Elsevier B.V.

Trend of research on the medical use of molecular hydrogen: a bibliometric analysis.

The medical use of molecular hydrogen, including hydrogen-rich water and hydrogen gas, has been extensively explored since 2007. This article aimed to demonstrate the trend in medical research on molecular hydrogen. A total of 1126 publications on hydrogen therapy were retrieved from the PubMed database until July 30, 2021. From 2007 to 2020, the number of publications in this field had been on an upward trend. Medical Gas Research, Scientific Report and Shock have contributed the largest number of publications on this topic. Researchers by the name of Xue-Jun Sun, Ke-Liang Xie and Yong-Hao Yu published the most studies in the field. Analysis of the co-occurrence of key words indicated that the key words "molecular hydrogen," "hydrogen-rich water," "oxidative stress," "hydrogen gas," and "inflammation" occurred most frequently in these articles. "Gut microbiota," "pyroptosis," and "COVID-19" occurred the most recently among the keywords. In summary, the therapeutic application of molecular hydrogen had attracted much attention in these years. The advance in this field could be caught up by subscribing to relevant journals or following experienced scholars. Oxidative stress and inflammation were the most important research directions currently, and gut microbiota, pyroptosis, and coronavirus disease 2019 might become hotspots in the future.

Severe acute respiratory syndrome coronavirus-2 accessory proteins ORF3a and ORF7a modulate autophagic flux and Ca2+ homeostasis in yeast.

Virus infection involves the manipulation of key host cell functions by specialized virulence proteins. The Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) small accessory proteins ORF3a and ORF7a have been implicated in favoring virus replication and spreading by inhibiting the autophagic flux within the host cell. Here, we apply yeast models to gain insights into the physiological functions of both SARS-CoV-2 small open reading frames (ORFs). ORF3a and ORF7a can be stably overexpressed in yeast cells, producing a decrease in cellular fitness. Both proteins show a distinguishable intracellular localization. ORF3a localizes to the vacuolar membrane, whereas ORF7a targets the endoplasmic reticulum. Overexpression of ORF3a and ORF7a leads to the accumulation of Atg8 specific autophagosomes. However, the underlying mechanism is different for each viral protein as assessed by the quantification of the autophagic degradation of Atg8-GFP fusion proteins, which is inhibited by ORF3a and stimulated by ORF7a. Overexpression of both SARS-CoV-2 ORFs decreases cellular fitness upon starvation conditions, where autophagic processes become essential. These data confirm previous findings on SARS-CoV-2 ORF3a and ORF7a manipulating autophagic flux in mammalian cell models and are in agreement with a model where both small ORFs have synergistic functions in stimulating intracellular autophagosome accumulation, ORF3a by inhibiting autophagosome processing at the vacuole and ORF7a by promoting autophagosome formation at the ER. ORF3a has an additional function in Ca2+ homeostasis. The overexpression of ORF3a confers calcineurin-dependent Ca2+ tolerance and activates a Ca2+ sensitive FKS2-luciferase reporter, suggesting a possible ORF3a-mediated Ca2+ efflux from the vacuole. Taken together, we show that viral accessory proteins can be functionally investigated in yeast cells and that SARS-CoV-2 ORF3a and ORF7a proteins interfere with autophagosome formation and processing as well as with Ca2+ homeostasis from distinct cellular targets.