Category: Animals

Innate Immune Activation and Mitochondrial ROS Invoke Persistent Cardiac Conduction System Dysfunction after COVID-19

Abstract:

Background Cardiac risk rises during acute SARS-CoV-2 infection and in long COVID syndrome in humans, but the mechanisms behind COVID-19-linked arrhythmias are unknown. This study explores the acute and long term effects of SARS-CoV-2 on the cardiac conduction system (CCS) in a hamster model of COVID-19.

Methods Radiotelemetry in conscious animals was used to non-invasively record electrocardiograms and subpleural pressures after intranasal SARS-CoV-2 infection. Cardiac cytokines, interferon-stimulated gene expression, and macrophage infiltration of the CCS, were assessed at 4 days and 4 weeks post-infection. A double-stranded RNA mimetic, polyinosinic:polycytidylic acid (PIC), was used in vivo and in vitro to activate viral pattern recognition receptors in the absence of SARS-CoV-2 infection.

Results COVID-19 induced pronounced tachypnea and severe cardiac conduction system (CCS) dysfunction, spanning from bradycardia to persistent atrioventricular block, although no viral protein expression was detected in the heart. Arrhythmias developed rapidly, partially reversed, and then redeveloped after the pulmonary infection was resolved, indicating persistent CCS injury. Increased cardiac cytokines, interferon-stimulated gene expression, and macrophage remodeling in the CCS accompanied the electrophysiological abnormalities. Interestingly, the arrhythmia phenotype was reproduced by cardiac injection of PIC in the absence of virus, indicating that innate immune activation was sufficient to drive the response. PIC also strongly induced cytokine secretion and robust interferon signaling in hearts, human iPSC-derived cardiomyocytes (hiPSC-CMs), and engineered heart tissues, accompanied by alterations in electrical and Ca2+ handling properties. Importantly, the pulmonary and cardiac effects of COVID-19 were blunted by in vivo inhibition of JAK/STAT signaling or by a mitochondrially-targeted antioxidant.

Conclusions The findings indicate that long term dysfunction and immune cell remodeling of the CCS is induced by COVID-19, arising indirectly from oxidative stress and excessive activation of cardiac innate immune responses during infection, with implications for long COVID Syndrome.

Source: Deepthi Ashok, Ting Liu, Joseph Criscione, Meghana Prakash, Byunggik Kim, Julian Chow, Morgan Craney, Kyriakos N. Papanicolaou, Agnieszka Sidor, D. Brian Foster, Andrew Pekosz, Jason Villano, Deok-Ho Kim, Brian O’Rourke. Innate Immune Activation and Mitochondrial ROS Invoke Persistent Cardiac Conduction System Dysfunction after COVID-19. bioRxiv 2024.01.05.574280; doi: https://doi.org/10.1101/2024.01.05.574280 https://www.biorxiv.org/content/10.1101/2024.01.05.574280v1.full (Full text)

Central 5-HTergic hyperactivity induces myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS)-like pathophysiology

Abstract:

Objectives: Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a significant medical challenge, with no indisputable pathophysiological mechanism identified to date.

Methods: Based on clinical clues, we hypothesized that 5-hydroxytryptamine (5-HT) hyperactivation is implicated in the pathogenic causes of ME/CFS and the associated symptoms. We experimentally evaluated this hypothesis in a series of mouse models.

Results: High-dose selective serotonin reuptake inhibitor (SSRI) treatment induced intra- and extracellular serotonin spillover in the dorsal raphe nuclei of mice. This condition resulted in severe fatigue (rota-rod, fatigue rotating wheel and home-cage activity tests) and ME/CFS-associated symptoms (nest building, plantar and open field test), along with dysfunction in the hypothalamic-pituitary-adrenal (HPA) axis response to exercise challenge. These ME/CFS-like features induced by excess serotonin were additionally verified using both a 5-HT synthesis inhibitor and viral vector for Htr1a (5-HT1A receptor) gene knockdown.

Conclusions: Our findings support the involvement of 5-HTergic hyperactivity in the pathophysiology of ME/CFS. This ME/CFS-mimicking animal model would be useful for understanding ME/CFS biology and its therapeutic approaches.

Source: Lee JS, Kang JY, Park SY, Hwang SJ, Bae SJ, Son CG. Central 5-HTergic hyperactivity induces myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS)-like pathophysiology. J Transl Med. 2024 Jan 8;22(1):34. doi: 10.1186/s12967-023-04808-x. PMID: 38191373. https://translational-medicine.biomedcentral.com/articles/10.1186/s12967-023-04808-x (Full text)

Integrated ‘omics analysis for the gut microbiota response to moxibustion in a rat model of chronic fatigue syndrome

Abstract:

Objective: To observe the efficacy of moxibustion in the treatment of chronic fatigue syndrome (CFS) and explore the effects on gut microbiota and metabolic profiles.

Methods: Forty-eight male Sprague-Dawley rats were randomly assigned to control group (Con), CFS model group (Mod, established by multiple chronic stress for 35 d), MoxA group (CFS model with moxibustion Shenque (CV8) and Guanyuan (CV4), 10 min/d, 28 d) and MoxB group (CFS model with moxibustion Zusanli (ST36), 10 min/d, 28 d).

Open-field test (OFT) and Morris-water-maze test (MWMT) were determined for assessment the CFS model and the therapeutic effects of moxibustion.16S rRNA gene sequencing analysis based gut microbiota integrated untargeted liquid chromatograph-mass spectrometer (LC-MS) based fecal metabolomics were executed, as well as Spearman correlation analysis, was utilized to uncover the functional relevance between the potential metabolites and gut microbiota.

Results: The results of our behavioral tests showed that moxibustion improved the performance of CFS rats in the OFT and the MWMT. Microbiome profiling analysis revealed that the gut microbiomes of CFS rats were less diverse with altered composition, including increases in pro-inflammatory species (such as Proteobacteria) and decreases in anti-inflammatory species (such as Bacteroides, Lactobacillus, Ruminococcus, and Prevotella). Moxibustion partially normalized these changes in the gut microbiota.

Furthermore, CFS was associated with metabolic disorders, which were effectively ameliorated by moxibustion. This was demonstrated by the normalization of 33 microbiota-related metabolites, including mannose (P = 0.001), aspartic acid (P = 0.009), alanine (P = 0.007), serine (P = 0.000), threonine (P = 0.027), methionine (P = 0.023), 5-hydroxytryptamine (P = 0.008), alpha-linolenic acid (P = 0.003), eicosapentaenoic acid (P = 0.006), hypoxanthine (P = 0.000), vitamin B6 (P = 0.000), cholic acid (P = 0.013), and taurocholate (P = 0.002).

Correlation analysis showed a significant association between the perturbed fecal microbiota and metabolite levels, with a notable negative relationship between LCA and Bacteroides.

Conclusions: In this study, we demonstrated that moxibustion has an antifatigue-like effect. The results from the 16S rRNA gene sequencing and metabolomics analysis suggest that the therapeutic effects of moxibustion on CFS are related to the regulation of gut microorganisms and their metabolites. The increase in Bacteroides and decrease in LCA may be key targets for the moxibustion treatment of CFS.

Source: Chaoran LI, Yan Y, Chuwen F, Heng LI, Yuanyuan QU, Yulin W, Delong W, Qingyong W, Jing G, Tianyu S, Xiaowei S, Xue W, Yunlong H, Zhongren S, Tiansong Y. Integrated ‘omics analysis for the gut microbiota response to moxibustion in a rat model of chronic fatigue syndrome. J Tradit Chin Med. 2023 Oct;43(6):1176-1189. doi: 10.19852/j.cnki.jtcm.20231018.004. PMID: 37946480; PMCID: PMC10623263. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10623263/ (Full text)

SARS-CoV-2 viral persistence in lung alveolar macrophages is controlled by IFN-γ and NK cells

Abstract:

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA generally becomes undetectable in upper airways after a few days or weeks postinfection. Here we used a model of viral infection in macaques to address whether SARS-CoV-2 persists in the body and which mechanisms regulate its persistence.

Replication-competent virus was detected in bronchioalveolar lavage (BAL) macrophages beyond 6 months postinfection. Viral propagation in BAL macrophages occurred from cell to cell and was inhibited by interferon-γ (IFN-γ). IFN-γ production was strongest in BAL NKG2r+CD8+ T cells and NKG2Alo natural killer (NK) cells and was further increased in NKG2Alo NK cells after spike protein stimulation.

However, IFN-γ production was impaired in NK cells from macaques with persisting virus. Moreover, IFN-γ also enhanced the expression of major histocompatibility complex (MHC)-E on BAL macrophages, possibly inhibiting NK cell-mediated killing. Macaques with less persisting virus mounted adaptive NK cells that escaped the MHC-E-dependent inhibition.

Our findings reveal an interplay between NK cells and macrophages that regulated SARS-CoV-2 persistence in macrophages and was mediated by IFN-γ.

Source: Huot N, Planchais C, Rosenbaum P, Contreras V, Jacquelin B, Petitdemange C, Lazzerini M, Beaumont E, Orta-Resendiz A, Rey FA, Reeves RK, Le Grand R, Mouquet H, Müller-Trutwin M. SARS-CoV-2 viral persistence in lung alveolar macrophages is controlled by IFN-γ and NK cells. Nat Immunol. 2023 Nov 2. doi: 10.1038/s41590-023-01661-4. Epub ahead of print. PMID: 37919524. https://www.nature.com/articles/s41590-023-01661-4 (Full text)

Comparative single-cell analysis reveals IFN-γ as a driver of respiratory sequelae post COVID-19

Abstract:

Post-acute sequelae of SARS-CoV-2 infection (PASC) represents an urgent public health challenge, with its impact resonating in over 60 million individuals globally. While a growing body of evidence suggests that dysregulated immune reactions may be linked with PASC symptoms, most investigations have primarily centered around blood studies, with few focusing on samples derived from post-COVID affected tissues. Further, clinical studies alone often provide correlative insights rather than causal relationships. Thus, it is essential to compare clinical samples with relevant animal models and conduct functional experiments to truly understand the etiology of PASC.

In this study, we have made comprehensive comparisons between bronchoalveolar lavage fluid (BAL) single-cell RNA sequencing (scRNAseq) data derived from clinical PASC samples and relevant PASC mouse models. This revealed a strong pro-fibrotic monocyte-derived macrophage response in respiratory PASC (R-PASC) in both humans and mice, and abnormal interactions between pulmonary macrophages and respiratory resident T cells.

IFN-g emerged as a key node mediating the immune anomalies in R-PASC. Strikingly, neutralizing IFN-g post the resolution of acute infection reduced lung inflammation, tissue fibrosis, and improved pulmonary gas-exchange function in two mouse models of R-PASC. Our study underscores the importance of performing comparative analysis to understand the root cause of PASC for developing effective therapies.

Source: Jie Sun, Chaofan Li, Wei Qian, Xiaoqin Wei. Comparative single-cell analysis reveals IFN-γ as a driver of respiratory sequelae post COVID-19. bioRxiv 2023.10.03.560739; doi: https://doi.org/10.1101/2023.10.03.560739 https://www.biorxiv.org/content/10.1101/2023.10.03.560739v1

Recuperative herbal formula Jing Si maintains vasculature permeability balance, regulates inflammation and assuages concomitants of “Long-Covid”

Abstract:

Coronavirus disease 2019 (COVID-19) is a worldwide health threat that has long-term effects on the patients and there is currently no efficient cure prescribed for the treatment and the prolonging effects. Traditional Chinese medicines (TCMs) have been reported to exert therapeutic effect against COVID-19.

In this study, the therapeutic effects of Jing Si herbal tea (JSHT) against COVID-19 infection and associated long-term effects were evaluated in different in vitro and in vivo models. The anti-inflammatory effects of JSHT were studied in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells and in Omicron pseudotyped virus-induced acute lung injury model. The effect of JSHT on cellular stress was determined in HK-2 proximal tubular cells and H9c2 cardiomyoblasts.

The therapeutic benefits of JSHT on anhedonia and depression symptoms associated with long COVID were evaluated in mice models for unpredictable chronic mild stress (UCMS). JSHT inhibited the NF-ƙB activities, and significantly reduced LPS-induced expression of TNFα, COX-2, NLRP3 inflammasome, and HMGB1. JSHT was also found to significantly suppress the production of NO by reducing iNOS expression in LPS-stimulated RAW 264.7 cells.

Further, the protective effects of JSHT on lung tissue were confirmed based on mitigation of lung injury, repression in TMRRSS2 and HMGB-1 expression and reduction of cytokine storm in the Omicron pseudotyped virus-induced acute lung injury model. JSHT treatment in UCMS models also relieved chronic stress and combated depression symptoms. The results therefore show that JSHT attenuates the cytokine storm by repressing NF-κB cascades and provides the protective functions against symptoms associated with long COVID-19 infection.

Source: Chiang CY, Lin YJ, Weng WT, Lin HD, Lu CY, Chen WJ, Shih CY, Lin PY, Lin SZ, Ho TJ, Shibu MA, Huang CY. Recuperative herbal formula Jing Si maintains vasculature permeability balance, regulates inflammation and assuages concomitants of “Long-Covid”. Biomed Pharmacother. 2023 Apr 26;163:114752. doi: 10.1016/j.biopha.2023.114752. Epub ahead of print. PMID: 37116351; PMCID: PMC10130602. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10130602/ (Full text)

Effect of transcutaneous electrical acupoint stimulation on learning and memory ability of chronic fatigue syndrome rats and its mechanisms

Abstract:

in English, Chinese

Objective: To observe the effect of transcutaneous electrical acupoint stimulation (TEAS) on the histomorphological manifestations of hippocampal CA1 region and the expression of extracellular regulatory protein kinase (ERK), cyclic adenosine response element binding protein (CREB) and brain-derived neurotrophic factor (BDNF) in chronic fatigue syndrome (CFS) rats, so as to explore the mechanisms of TEAS in improving the learning and memory abilities of CFS rats.

Methods: Forty male Wistar rats were randomly divided into normal group (10 rats) and modeling group (30 rats); then after modeling, they were selected and randomly divided into model group (10 rats) and TEAS group (10 rats). CFS rats model was prepared by sleep deprivation combined with weight-bearing swimming. Rats in the TEAS group were stimulated with Han’s acupoint nerve stimulator at bilateral “Zusanli” (ST36) and “Shenshu” (BL23) (2 Hz/15 Hz, 1-2 mA), 20 min each time, once a day for 4 weeks with 1 d rest every 6 d. The score of general conditions of rats was evaluated. The learning and memory ability was tested with Morris water maze. The morphology and ultrastructure of hippocampal CA1 region were observed by HE staining and transmission electron microscopy. The expression levels of ERK, CREB and BDNF mRNAs and proteins in hippocampus were detected by real time quantitative PCR and Western blot, respectively.

Results: Compared with the normal group, the score of general condition was increased (P<0.01); the escape latency was prolonged (P<0.05, P<0.01) and the times of crossing the original platform was decreased (P<0.05); the expression levels of ERK, CREB and BDNF mRNAs and proteins in hippocampus were decreased (P<0.05, P<0.01) in the model group. Compared with the model group, the scores of general condition on the 42nd and 49th day were decreased (P<0.05, P<0.01); the escape latency was shortened (P<0.01, P<0.05)and the times of crossing the original platform were increased (P<0.05); the expression levels of ERK, CREB and BDNF mRNAs and proteins in hippocampus were increased (P<0.01, P<0.05) in the TEAS group. The morphology of neurons in hippocampal CA1 region was normal in the normal group. In the model group, the number of neurons in hippocampal CA1 region decreased, the arrangement of nerve cells was scattered, the number of apoptotic cells increased, some nuclear structures disappeared, nuclear heterochromatin increased, the cell membrane wrinkled, the chromatin appeared empty bright area, and the crista was incomplete. Compared with the model group, the nerve cells morphology in hippocampal CA1 region was more regular, the number of apoptotic cells decreased, the chromatin and the cytoplasm were uniformly distributed, and the crista was relatively intact in the TEAS group.

Conclusion: TEAS can improve the learning and memory ability of CFS rats, the mechanisms may be related to improving the neural structure of hippocampal CA1 region and up-regulating the expression levels of ERK/CREB/BDNF.

Source: Zhong XL, Tong BY, Yang YH, Zeng HL, Lin C, Jing Y, He LL, You SJ. [Effect of transcutaneous electrical acupoint stimulation on learning and memory ability of chronic fatigue syndrome rats and its mechanisms]. Zhen Ci Yan Jiu. 2023 Apr 25;48(4):317-24. Chinese. doi: 10.13702/j.1000-0607.20221032. PMID: 37186194. https://pubmed.ncbi.nlm.nih.gov/37186194/

Astragalus polysaccharide ameliorated complex factor-induced chronic fatigue syndrome by modulating the gut microbiota and metabolites in mice

Abstract:

Chronic fatigue syndrome (CFS) is a debilitating disease with no symptomatic treatment. Astragalus polysaccharide (APS), a component derived from the traditional Chinese medicine A. membranaceus, has significant anti-fatigue activity. However, the mechanisms underlying the potential beneficial effects of APS on CFS remain poorly understood.

A CFS model of 6-week-old C57BL/6 male mice was established using the multiple-factor method. These mice underwent examinations for behavior, oxidative stress and inflammatory indicators in brain and intestinal tissues, and ileum histomorphology. 16 S rDNA sequencing analysis indicated that APS regulated the abundance of gut microbiota and increased production of short chain fatty acids (SCFAs) and anti-inflammatory bacteria.

In addition, APS reversed the abnormal expression of Nrf2, NF-κB, and their downstream factors in the brain-gut axis and alleviated the reduction in SCFAs in the cecal content caused by CFS. Further, APS modulated the changes in serum metabolic pathways induced by CFS.

Finally, it was verified that butyrate exerted antioxidant and anti-inflammatory effects in neuronal cells. In conclusion, APS could increase the SCFAs content by regulating the gut microbiota, and SCFAs (especially butyrate) can further regulate the oxidative stress and inflammation in the brain, thus alleviating CFS.

This study explored the efficacy and mechanism of APS for CFS from the perspective of gut-brain axis and provides a reference to further explore the efficacy of APS and the role of SCFAs in the central nervous system.

Source: Wei X, Xin J, Chen W, Wang J, Lv Y, Wei Y, Li Z, Ding Q, Shen Y, Xu X, Zhang X, Zhang W, Zu X. Astragalus polysaccharide ameliorated complex factor-induced chronic fatigue syndrome by modulating the gut microbiota and metabolites in mice. Biomed Pharmacother. 2023 May 9;163:114862. doi: 10.1016/j.biopha.2023.114862. Epub ahead of print. PMID: 37167729. https://www.sciencedirect.com/science/article/pii/S0753332223006522?via%3Dihub (Full study)

Mouse Adapted SARS-CoV-2 Model Induces “Long-COVID” Neuropathology in BALB/c Mice

Abstract:

The novel coronavirus SARS-CoV-2 has caused significant global morbidity and mortality and continues to burden patients with persisting neurological dysfunction. COVID-19 survivors develop debilitating symptoms to include neuro-psychological dysfunction, termed “Long COVID”, which can cause significant reduction of quality of life. Despite vigorous model development, the possible cause of these symptoms and the underlying pathophysiology of this devastating disease remains elusive.

Mouse adapted (MA10) SARS-CoV-2 is a novel mouse-based model of COVID-19 which simulates the clinical symptoms of respiratory distress associated with SARS-CoV-2 infection in mice. In this study, we evaluated the long-term effects of MA10 infection on brain pathology and neuroinflammation. 10-week and 1-year old female BALB/cAnNHsd mice were infected intranasally with 10 4 plaque-forming units (PFU) and 10 3 PFU of SARS-CoV-2 MA10, respectively, and the brain was examined 60 days post-infection (dpi).

Immunohistochemical analysis showed a decrease in the neuronal nuclear protein NeuN and an increase in Iba-1 positive amoeboid microglia in the hippocampus after MA10 infection, indicating long-term neurological changes in a brain area which is critical for long-term memory consolidation and processing. Importantly, these changes were seen in 40-50% of infected mice, which correlates to prevalence of LC seen clinically.

Our data shows for the first time that MA10 infection induces neuropathological outcomes several weeks after infection at similar rates of observed clinical prevalence of “Long COVID”. These observations strengthen the MA10 model as a viable model for study of the long-term effects of SARS-CoV-2 in humans. Establishing the viability of this model is a key step towards the rapid development of novel therapeutic strategies to ameliorate neuroinflammation and restore brain function in those suffering from the persistent cognitive dysfunction of “Long-COVID”.

Source: Gressett TE, Leist SR, Ismael S, Talkington G, Dinnon KH, Baric RS, Bix G. Mouse Adapted SARS-CoV-2 Model Induces “Long-COVID” Neuropathology in BALB/c Mice. bioRxiv [Preprint]. 2023 Mar 20:2023.03.18.533204. doi: 10.1101/2023.03.18.533204. PMID: 36993423; PMCID: PMC10055301. https://www.biorxiv.org/content/10.1101/2023.03.18.533204v1.full (Full text)

SARS-CoV-2 infection induces DNA damage, through CHK1 degradation and impaired 53BP1 recruitment, and cellular senescence

Abstract:

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the RNA virus responsible for the coronavirus disease 2019 (COVID-19) pandemic. Although SARS-CoV-2 was reported to alter several cellular pathways, its impact on DNA integrity and the mechanisms involved remain unknown. Here we show that SARS-CoV-2 causes DNA damage and elicits an altered DNA damage response.

Mechanistically, SARS-CoV-2 proteins ORF6 and NSP13 cause degradation of the DNA damage response kinase CHK1 through proteasome and autophagy, respectively. CHK1 loss leads to deoxynucleoside triphosphate (dNTP) shortage, causing impaired S-phase progression, DNA damage, pro-inflammatory pathways activation and cellular senescence. Supplementation of deoxynucleosides reduces that. Furthermore, SARS-CoV-2 N-protein impairs 53BP1 focal recruitment by interfering with damage-induced long non-coding RNAs, thus reducing DNA repair.

Key observations are recapitulated in SARS-CoV-2-infected mice and patients with COVID-19. We propose that SARS-CoV-2, by boosting ribonucleoside triphosphate levels to promote its replication at the expense of dNTPs and by hijacking damage-induced long non-coding RNAs’ biology, threatens genome integrity and causes altered DNA damage response activation, induction of inflammation and cellular senescence.

Source: Gioia U, Tavella S, Martínez-Orellana P, Cicio G, Colliva A, Ceccon M, Cabrini M, Henriques AC, Fumagalli V, Paldino A, Presot E, Rajasekharan S, Iacomino N, Pisati F, Matti V, Sepe S, Conte MI, Barozzi S, Lavagnino Z, Carletti T, Volpe MC, Cavalcante P, Iannacone M, Rampazzo C, Bussani R, Tripodo C, Zacchigna S, Marcello A, d’Adda di Fagagna F. SARS-CoV-2 infection induces DNA damage, through CHK1 degradation and impaired 53BP1 recruitment, and cellular senescence. Nat Cell Biol. 2023 Mar 9. doi: 10.1038/s41556-023-01096-x. Epub ahead of print. PMID: 36894671. https://www.nature.com/articles/s41556-023-01096-x (Full text)