Chlorogenic Acid: A Systematic Review on the Biological Functions, Mechanistic Actions, and Therapeutic Potentials.

Chlorogenic acid (CGA) is a type of polyphenol compound found in rich concentrations in many plants such as green coffee beans. As an active natural substance, CGA exerts diverse therapeutic effects in response to a variety of pathological challenges, particularly conditions associated with chronic metabolic diseases and age-related disorders. It shows multidimensional functions, including neuroprotection for neurodegenerative disorders and diabetic peripheral neuropathy, anti-inflammation, anti-oxidation, anti-pathogens, mitigation of cardiovascular disorders, skin diseases, diabetes mellitus, liver and kidney injuries, and anti-tumor activities. Mechanistically, its integrative functions act through the modulation of anti-inflammation/oxidation and metabolic homeostasis. It can thwart inflammatory constituents at multiple levels such as curtailing NF-kB pathways to neutralize primitive inflammatory factors, hindering inflammatory propagation, and alleviating inflammation-related tissue injury. It concurrently raises pivotal antioxidants by activating the Nrf2 pathway, thus scavenging excessive cellular free radicals. It elevates AMPK pathways for the maintenance and restoration of metabolic homeostasis of glucose and lipids. Additionally, CGA shows functions of neuromodulation by targeting neuroreceptors and ion channels. In this review, we systematically recapitulate CGA's pharmacological activities, medicinal properties, and mechanistic actions as a potential therapeutic agent. Further studies for defining its specific targeting molecules, improving its bioavailability, and validating its clinical efficacy are required to corroborate the therapeutic effects of CGA.

Pharmacological Activities, Therapeutic Effects, and Mechanistic Actions of Trigonelline.

Trigonelline (TRG) is a natural polar hydrophilic alkaloid that is found in many plants such as green coffee beans and fenugreek seeds. TRG potentially acts on multiple molecular targets, including nuclear factor erythroid 2-related factor 2 (Nrf2), peroxisome proliferator-activated receptor γ, glycogen synthase kinase, tyrosinase, nerve growth factor, estrogen receptor, amyloid-ß peptide, and several neurotransmitter receptors. In this review, we systematically summarize the pharmacological activities, medicinal properties, and mechanistic actions of TRG as a potential therapeutic agent. Mechanistically, TRG can facilitate the maintenance and restoration of the metabolic homeostasis of glucose and lipids. It can counteract inflammatory constituents at multiple levels by hampering pro-inflammatory factor release, alleviating inflammatory propagation, and attenuating tissue injury. It concurrently modulates oxidative stress by the blockage of the detrimental Nrf2 pathway when autophagy is impaired. Therefore, it exerts diverse therapeutic effects on a variety of pathological conditions associated with chronic metabolic diseases and age-related disorders. It shows multidimensional effects, including neuroprotection from neurodegenerative disorders and diabetic peripheral neuropathy, neuromodulation, mitigation of cardiovascular disorders, skin diseases, diabetic mellitus, liver and kidney injuries, and anti-pathogen and anti-tumor activities. Further validations are required to define its specific targeting molecules, dissect the underlying mechanistic networks, and corroborate its efficacy in clinical trials.

Glutathione supplementation improves fat graft survival by inhibiting ferroptosis via the SLC7A11/GPX4 axis.

BACKGROUND: Autologous fat grafting is hampered by unpredictable graft survival, which is potentially regulated by ferroptosis. Glutathione (GSH), a powerful antioxidant used in tissue preservation, has ferroptosis-regulating activity; however, its effects on fat grafts are unclear. This study investigated the effects and mechanisms of GSH in fat graft survival. METHODS: Human lipoaspirates were transplanted subcutaneously into the backs of normal saline-treated (control) or GSH-treated nude mice. Graft survival was evaluated by magnetic resonance imaging and histology. RNA sequencing was performed to identify differentially expressed genes and enriched pathways. GSH activity was evaluated in vitro using an oxygen and glucose deprivation (OGD) model of adipose-derived stem cells. RESULTS: Compared with control group, GSH induced better outcomes, including superior graft retention, appearance, and histological structures. RNA sequencing suggested enhanced negative regulation of ferroptosis in the GSH-treated grafts, which showed reduced lipid peroxides, better mitochondrial ultrastructure, and SLC7A11/GPX4 axis activation. In vitro, OGD-induced ferroptosis was ameliorated by GSH, which restored cell proliferation, reduced oxidative stress, and upregulated ferroptosis defense factors. CONCLUSIONS: Our study confirms that ferroptosis participates in regulating fat graft survival and that GSH exerts a protective effect by inhibiting ferroptosis. GSH-assisted lipotransfer is a promising therapeutic strategy for future clinical application.

Adipose-derived Mesenchymal Stem Cells are Ideal for the Cell-based Treatment of Refractory Wounds: Strong Potential for Angiogenesis.

Although Mesenchymal Stem Cells (MSCs)-based therapy has been proposed as a promising strategy for the treatment of chronic lower-extremity ulcers, their optimal sources, amounts, and delivery methods are urgently needed to be determined. In this study, we compared the heterogeneity of the human MSCs derived from bone marrow (BMSCs), umbilical cord (UCMSCs), and adipose tissue (ADSCs) in accelerating wound healing and promoting angiogenesis and explored the underlying mechanism. Briefly, a diabetic rat model with a full-thickness cutaneous wound on the dorsal foot was developed. The wound was topically administered with three types of MSCs. Additionally, we carried out in vitro and in vivo analysis of the angiogenic properties of the MSCs. Moreover, the molecular mechanism of the heterogeneity of the MSCs derived from the three tissues was explored by transcriptome sequencing. When compared with the BMSCs- and UCMSCs-treated groups, the ADSCs-treated group exhibited markedly accelerated healing efficiency, characterized by increased wound closure rates, enhanced angiogenesis, and collagen deposition at the wound site. The three types of MSCs formed three-dimensional capillary-like structures and promoted angiogenesis in vitro and in vivo, with ADSCs exhibiting the highest capacity for tube formation and pro-angiogenesis. Furthermore, transcriptome sequencing revealed that ADSCs had higher expression levels of angiogenesis-associated genes. Our findings indicate that MSCs-based therapy accelerates the healing of ischemia- and diabetes-induced lower-extremity ulcers and that adipose tissue-derived MSCs might be ideal for therapeutic angiogenesis and treatment of chronic ischemic wounds.

Single-cell RNA-seq and bulk-seq identify RAB17 as a potential regulator of angiogenesis by human dermal microvascular endothelial cells in diabetic foot ulcers.

Background: Angiogenesis is crucial in diabetic wound healing and is often impaired in diabetic foot ulcers (DFUs). Human dermal microvascular endothelial cells (HDMECs) are vital components in dermal angiogenesis; however, their functional and transcriptomic characteristics in DFU patients are not well understood. This study aimed to comprehensively analyse HDMECs from DFU patients and healthy controls and find the potential regulator of angiogenesis in DFUs. Methods: HDMECs were isolated from skin specimens of DFU patients and healthy controls via magnetic-activated cell sorting. The proliferation, migration and tube-formation abilities of the cells were then compared between the experimental groups. Both bulk RNA sequencing (bulk-seq) and single-cell RNA-seq (scRNA-seq) were used to identify RAB17 as a potential marker of angiogenesis, which was further confirmed via weighted gene co-expression network analysis (WGCNA) and least absolute shrink and selection operator (LASSO) regression. The role of RAB17 in angiogenesis was examined through in vitro and in vivo experiments. Results: The isolated HDMECs displayed typical markers of endothelial cells. HDMECs isolated from DFU patients showed considerably impaired tube formation, rather than proliferation or migration, compared to those from healthy controls. Gene set enrichment analysis (GSEA), fGSEA, and gene set variation analysis (GSVA) of bulk-seq and scRNA-seq indicated that angiogenesis was downregulated in DFU-HDMECs. LASSO regression identified two genes, RAB17 and CD200, as characteristic of DFU-HDMECs; additionally, the expression of RAB17 was found to be significantly reduced in DFU-HDMECs compared to that in the HDMECs of healthy controls. Overexpression of RAB17 was found to enhance angiogenesis, the expression of hypoxia inducible factor-1α and vascular endothelial growth factor A, and diabetic wound healing, partially through the mitogen-activated protein kinase/extracellular signal-regulated kinase signalling pathway. Conclusions: Our findings suggest that the impaired angiogenic capacity in DFUs may be related to the dysregulated expression of RAB17 in HDMECs. The identification of RAB17 as a potential molecular target provides a potential avenue for the treatment of impaired angiogenesis in DFUs.

The SARS-CoV-2 spike protein induces long-term transcriptional perturbations of mitochondrial metabolic genes, causes cardiac fibrosis, and reduces myocardial contractile in obese mice.

BACKGROUND: As the pandemic evolves, post-acute sequelae of CoV-2 (PASC) including cardiovascular manifestations have emerged as a new health threat. This study aims to study whether the Spike protein plus obesity can exacerbate PASC-related cardiomyopathy. METHODS: A Spike protein-pseudotyped (Spp) virus with the proper surface tropism of SARS-CoV-2 was developed for viral entry assay in vitro and administration into high fat diet (HFD)-fed mice. The systemic viral loads and cardiac transcriptomes were analyzed at 2 and 24 h, 3, 6, and 24 weeks post introducing (wpi) Spp using RNA-seq or real time RT-PCR. Echocardiography was used to monitor cardiac functions. RESULTS: Low-density lipoprotein cholesterol enhanced viral uptake in endothelial cells, macrophages, and cardiomyocyte-like H9C2 cells. Selective cardiac and adipose viral depositions were observed in HFD mice but not in normal-chow-fed mice. The cardiac transcriptional signatures in HFD mice at 3, 6, and 24 wpi showed systemic suppression of mitochondria respiratory chain genes including ATP synthases and nicotinamide adenine dinucleotide:ubiquinone oxidoreductase gene members, upregulation of stress pathway-related crucial factors such as nuclear factor-erythroid 2-related factor 1 and signal transducer and activator of transcription 5A, and increases in expression of glucose metabolism-associated genes. As compared with the age-matched HFD control mice, cardiac ejection fraction and fractional shortening were significantly decreased, while left ventricular end-systolic diameter and volume were significantly elevated, and cardiac fibrosis was increased in HFD mice at 24 wpi. CONCLUSION: Our data demonstrated that the Spike protein could induce long-term transcriptional suppression of mitochondria metabolic genes and cause cardiac fibrosis and myocardial contractile impairment in obese mice, providing mechanistic insights to PASC-related cardiomyopathy.

Corrigendum: The m6A-methylated mRNA pattern and the activation of the Wnt signaling pathway under the hyper-m6A-modifying condition in the keloid.

[This corrects the article DOI: 10.3389/fcell.2022.947337.].

The SARS-CoV-2 Spike protein induces long-term transcriptional perturbations of mitochondrial metabolic genes, causes cardiac fibrosis, and reduces myocardial contractile in obese mice.

Background: As the pandemic evolves, post-acute sequelae of CoV-2 (PACS) including cardiovascular manifestations have emerged as a new health threat. This study aims to study whether the Spike protein plus obesity can exacerbate PACS-related cardiomyopathy. Methods: A Spike protein-pseudotyped (Spp) virus with the proper surface tropism of SARS-CoV-2 was developed for viral entry assay in vitro and administration into high fat diet (HFD)-fed mice. The systemic viral loads and cardiac transcriptomes were analyzed at 2 and 24 hrs, 3, 6, and 24 weeks post introducing (wpi) Spp using RNA-seq or real time RT-PCR. Echocardiography was used to monitor cardiac functions. Results: Low-density lipoprotein cholesterol enhanced viral uptake in endothelial cells, macrophages, and cardiomyocyte-like H9C2 cells. Selective cardiac and adipose viral depositions were observed in HFD mice but not in normal-chow-fed mice. The cardiac transcriptional signatures in HFD mice at 3, 6, and 24 wpi showed systemic suppression of mitochondria respiratory chain genes including ATP synthases and nicotinamide adenine dinucleotide:ubiquinone oxidoreductase gene members, upregulation of stress pathway-related crucial factors such as nuclear factor-erythroid 2-related factor 1 and signal transducer and activator of transcription 5A, and increases in expression of glucose metabolism-associated genes. As compared with the age-matched HFD control mice, cardiac ejection fraction and fractional shortening were significantly decreased, while left ventricular end-systolic diameter and volume were significantly elevated, and cardiac fibrosis was increased in HFD mice at 24 wpi. Conclusion: Our data demonstrated that the Spike protein could induce long-term transcriptional suppression of mitochondria metabolic genes and cause cardiac fibrosis and myocardial contractile impairment, providing mechanistic insights to PACS-related cardiomyopathy.

The Interplay Between Autophagy and Regulated Necrosis.

Significance: Autophagy is critical to cellular homeostasis. Emergence of the concept of regulated necrosis, such as necroptosis, ferroptosis, pyroptosis, and mitochondrial membrane-permeability transition (MPT)-derived necrosis, has revolutionized the research into necrosis. Both altered autophagy and regulated necrosis contribute to major human diseases. Recent studies reveal an intricate interplay between autophagy and regulated necrosis. Understanding the interplay at the molecular level will provide new insights into the pathophysiology of related diseases. Recent Advances: Among the three forms of autophagy, macroautophagy is better studied for its crosstalk with regulated necrosis. Macroautophagy seemingly can either antagonize or promote regulated necrosis, depending upon the form of regulated necrosis, the type of cells or stimuli, and other cellular contexts. This review will critically analyze recent advances in the molecular mechanisms governing the intricate dialogues between macroautophagy and main forms of regulated necrosis. Critical Issues: The dual roles of autophagy, either pro-survival or pro-death characteristics, intricate the mechanistic relationship between autophagy and regulated necrosis at molecular level in various pathological conditions. Meanwhile, key components of regulated necrosis are also involved in the regulation of autophagy, which further complicates the interrelationship. Future Directions: Resolving the controversies over causation between altered autophagy and a specific form of regulated necrosis requires approaches that are more definitive, where rigorous evaluation of autophagic flux and the development of more reliable and specific methods to quantify each form of necrosis will be essential. The relationship between chaperone-mediated autophagy or microautophagy and regulated necrosis remains largely unstudied. Antioxid. Redox Signal. 38, 550-580.

Nrf2-Mediated Dichotomy in the Vascular System: Mechanistic and Therapeutic Perspective.

Nuclear factor-erythroid 2-related factor 2 (Nrf2), a transcription factor, controls the expression of more than 1000 genes that can be clustered into different categories with distinct functions ranging from redox balance and metabolism to protein quality control in the cell. The biological consequence of Nrf2 activation can be either protective or detrimental in a context-dependent manner. In the cardiovascular system, most studies have focused on the protective properties of Nrf2, mainly as a key transcription factor of antioxidant defense. However, emerging evidence revealed an unexpected role of Nrf2 in mediating cardiovascular maladaptive remodeling and dysfunction in certain disease settings. Herein we review the role of Nrf2 in cardiovascular diseases with a focus on vascular disease. We discuss the negative effect of Nrf2 on the vasculature as well as the potential underlying mechanisms. We also discuss the clinical relevance of targeting Nrf2 pathways for the treatment of cardiovascular and other diseases.

The m6A-methylated mRNA pattern and the activation of the Wnt signaling pathway under the hyper-m6A-modifying condition in the keloid.

Purpose: The present study was carried out to investigate the global m6A-modified RNA pattern and possible mechanisms underlying the pathogenesis of keloid. Method: In total, 14 normal skin and 14 keloid tissue samples were first collected on clinics. Then, three samples from each group were randomly selected to be verified with the Western blotting to determine the level of methyltransferase and demethylase. The total RNA of all samples in each group was isolated and subjected to the analysis of MeRIP sequencing and RNA sequencing. Using software of MeTDiff and htseq-count, the m6A peaks and differentially expressed genes (DEGs) were determined within the fold change >2 and p-value < 0.05. The top 10 pathways of m6A-modified genes in each group and the differentially expressed genes were enriched by the Kyoto Encyclopedia of Genes and Genomes signaling pathways. Finally, the closely associated pathway was determined using the Western blotting and immunofluorescence staining. Results: There was a higher protein level of WTAP and Mettl3 in the keloid than in the normal tissue. In the keloid samples, 21,020 unique m6A peaks with 6,573 unique m6A-associated genetic transcripts appeared. In the normal tissue, 4,028 unique m6A peaks with 779 m6A-associated modified genes appeared. In the RNA sequencing, there were 847 genes significantly changed between these groups, transcriptionally. The genes with m6A-methylated modification and the upregulated differentially expressed genes between two tissues were both mainly related to the Wnt signaling pathway. Moreover, the hyper-m6A-modified Wnt/ß-catenin pathway in keloid was verified with Western blotting. From the immunofluorescence staining results, we found that the accumulated fibroblasts were under a hyper-m6A condition in the keloid, and the Wnt/ß-Catenin signaling pathway was mainly activated in the fibroblasts. Conclusion: The fibroblasts in the keloid were under a cellular hyper-m6A-methylated condition, and the hyper-m6A-modified highly expressed Wnt/ß-catenin pathway in the dermal fibroblasts might promote the pathogenesis of keloid.

Graphene-incorporated hyaluronic acid-based hydrogel as a controlled Senexin A delivery system.

Perivascular delivery of therapeutic agents against established aetiologies for occlusive vascular remodelling has great therapeutic potential for vein graft failure. However, none of the perivascular drug delivery systems tested experimentally have been translated into clinical practice. In this study, we established a novel strategy to locally and sustainably deliver the cyclin-dependent kinase 8/19 inhibitor Senexin A (SenA), an emerging drug candidate to treat occlusive vascular disease, using graphene oxide-hybridised hyaluronic acid-based hydrogels. We demonstrated an approach to accommodate SenA in hyaluronic acid-based hydrogels through utilising graphene oxide nanosheets allowing for non-covalent interaction with SenA. The resulting hydrogels produced sustained delivery of SenA over 21 days with tunable release kinetics. In vitro assays also demonstrated that the hydrogels were biocompatible. This novel graphene oxide-incorporated hyaluronic acid hydrogel offers an optimistic outlook as a perivascular drug delivery system for treating occlusive vascular diseases, such as vein graft failure.

The Spike Protein of SARS-CoV-2 Impairs Lipid Metabolism and Increases Susceptibility to Lipotoxicity: Implication for a Role of Nrf2.

Coronavirus disease 2019 (COVID-19) patients show lipid metabolic alterations, but the mechanism remains unknown. In this study, we aimed to investigate whether the Spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) impairs lipid metabolism in host cells. We generated a Spike cell line in HEK293 using the pcDNA vector carrying the Spike gene expression cassette. A control cell line was generated using the empty pcDNA vector. Gene expression profiles related to lipid metabolic, autophagic, and ferroptotic pathways were investigated. Palmitic acid (PA)-overload was used to assess lipotoxicity-induced necrosis. As compared with controls, the Spike cells showed a significant increase in lipid depositions in cell membranes as well as dysregulation of expression of a panel of molecules involving lipid metabolism, autophagy, and ferroptosis. The Spike cells showed an upregulation of nuclear factor erythroid 2-related factor 2 (Nrf2), a multifunctional transcriptional factor, in response to PA. Furthermore, the Spike cells exhibited increased necrosis in response to PA-induced lipotoxicity compared to control cells in a time- and dose-dependent manner via ferroptosis, which could be attenuated by the Nrf2 inhibitor trigonelline. We conclude that the Spike protein impairs lipid metabolic and autophagic pathways in host cells, leading to increased susceptibility to lipotoxicity via ferroptosis which can be suppressed by a Nrf2 inhibitor. This data also suggests a central role of Nrf2 in Spike-induced lipid metabolic impairments.

Spike protein of SARS-CoV-2 activates macrophages and contributes to induction of acute lung inflammation in male mice.

The spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) plays a crucial role in mediating viral entry into host cells. However, whether it contributes to pulmonary hyperinflammation in patients with coronavirus disease 2019 is not well known. In this study, we developed a spike protein-pseudotyped (Spp) lentivirus with the proper tropism of the SARS-CoV-2 spike protein on the surface and determined the distribution of the Spp lentivirus in wild-type C57BL/6J male mice that received an intravenous injection of the virus. Lentiviruses with vesicular stomatitis virus glycoprotein (VSV-G) or with a deletion of the receptor-binding domain (RBD) in the spike protein [Spp (∆RBD)] were used as controls. Two hours postinfection (hpi), there were 27-75 times more viral burden from Spp lentivirus in the lungs than in other organs; there were also about 3-5 times more viral burden from Spp lentivirus than from VSV-G lentivirus in the lungs, liver, kidney, and spleen. Deletion of RBD diminished viral loads in the lungs but not in the heart. Acute pneumonia was observed in animals 24 hpi. Spp lentivirus was mainly found in SPC+ and LDLR+ pneumocytes and macrophages in the lungs. IL6, IL10, CD80, and PPAR-γ were quickly upregulated in response to infection in the lungs as well as in macrophage-like RAW264.7 cells. Furthermore, forced expression of the spike protein in RAW264.7 cells significantly increased the mRNA levels of the same panel of inflammatory factors. Our results demonstrated that the spike protein of SARS-CoV-2 confers the main point of viral entry into the lungs and can induce cellular pathology. Our data also indicate that an alternative ACE2-independent viral entry pathway may be recruited in the heart and aorta.

Cullin Deneddylation Suppresses the Necroptotic Pathway in Cardiomyocytes.

Cardiomyocyte death in the form of apoptosis and necrosis represents a major cellular mechanism underlying cardiac pathogenesis. Recent advances in cell death research reveal that not all necrosis is accidental, but rather there are multiple forms of necrosis that are regulated. Necroptosis, the earliest identified regulated necrosis, is perhaps the most studied thus far, and potential links between necroptosis and Cullin-RING ligases (CRLs), the largest family of ubiquitin E3 ligases, have been postulated. Cullin neddylation activates the catalytic dynamic of CRLs; the reverse process, Cullin deneddylation, is performed by the COP9 signalosome holocomplex (CSN) that is formed by eight unique protein subunits, COPS1/CNS1 through COPS8/CNS8. As revealed by cardiomyocyte-restricted knockout of Cops8 (Cops8-cko) in mice, perturbation of Cullin deneddylation in cardiomyocytes impairs not only the functioning of the ubiquitin-proteasome system (UPS) but also the autophagic-lysosomal pathway (ALP). Similar cardiac abnormalities are also observed in Cops6-cko mice; and importantly, loss of the desmosome targeting of COPS6 is recently implicated as a pathogenic factor in arrhythmogenic right ventricular dysplasia/cardiomyopathy (ARVD/C). Cops8-cko causes massive cardiomyocyte death in the form of necrosis rather than apoptosis and rapidly leads to a progressive dilated cardiomyopathy phenotype as well as drastically shortened lifespan in mice. Even a moderate downregulation of Cullin deneddylation as seen in mice with Cops8 hypomorphism exacerbates cardiac proteotoxicity induced by overexpression of misfolded proteins. More recently, it was further demonstrated that cardiomyocyte necrosis caused by Cops8-cko belongs to necroptosis and is mediated by the RIPK1-RIPK3 pathway. This article reviews these recent advances and discusses the potential links between Cullin deneddylation and the necroptotic pathways in hopes of identifying potentially new therapeutic targets for the prevention of cardiomyocyte death.

Autophagy Controls Nrf2-Mediated Dichotomy in Pressure Overloaded Hearts.

Burgeoning evidence has indicated that normal autophagy is required for nuclear factor erythroid 2-related factor (Nrf2)-mediated cardiac protection whereas autophagy inhibition turns on Nrf2-mediated myocardial damage and dysfunction in a setting of pressure overload (PO). However, such a concept remains to be fully established by a careful genetic interrogation in vivo. This study was designed to validate the hypothesis using a mouse model of PO-induced cardiomyopathy and heart failure, in which cardiac autophagy and/or Nrf2 activity are genetically inhibited. Myocardial autophagy inhibition was induced by cardiomyocyte-restricted (CR) knockout (KO) of autophagy related (Atg) 5 (CR-Atg5KO) in adult mice. CR-Atg5KO impaired cardiac adaptations while exacerbating cardiac maladaptive responses in the setting of PO. Notably, it also turned off Nrf2-mediated defense while switching on Nrf2-operated tissue damage in PO hearts. In addition, cardiac autophagy inhibition selectively inactivated extracellular signal regulated kinase (ERK), which coincided with increased nuclear accumulation of Nrf2 and decreased nuclear translocation of activated ERK in cardiomyocytes in PO hearts. Mechanistic investigation revealed that autophagy is required for the activation of ERK, which suppresses Nrf2-driven expression of angiotensinogen in cardiomyocytes. Taken together, these results provide direct evidence consolidating the notion that normal autophagy enables Nrf2-operated adaptation while switching off Nrf2-mediated maladaptive responses in PO hearts partly through suppressing Nrf2-driven angiotensinogen expression in cardiomyocytes.

Spike Protein of SARS-CoV-2 Activates Macrophages and Contributes to Induction of Acute Lung Inflammations in Mice.

Background: Coronavirus disease 2019 (COVID-19) patients exhibit multiple organ malfunctions with a primary manifestation of acute and diffuse lung injuries. The Spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is crucial to mediate viral entry into host cells; however, whether it can be cellularly pathogenic and contribute to pulmonary hyper-inflammations in COVID-19 is not well known. Methods and Findings: In this study, we developed a Spike protein-pseudotyped (Spp) lentivirus with the proper tropism of SARS-CoV-2 Spike protein on the surface and tracked down the fate of Spp in wild type C57BL/6J mice receiving intravenous injection of the virus. A lentivirus with vesicular stomatitis virus glycoprotein (VSV-G) was used as the control. Two hours post-infection (hpi), Spp showed more than 27-75 times more viral burden in the lungs than other organs; it also exhibited about 3-5 times more viral burden than VSV-G lentivirus in the lungs, liver, kidney and spleen. Acute pneumonia was evident in animals 24 hpi. Spp lentivirus was mainly found in LDLR+ macrophages and pneumocytes in the lungs, but not in MARC1+ macrophages. IL6, IL10, CD80 and PPAR-γ were quickly upregulated in response to infection of Spp lentivirus in the lungs in vivo as well as in macrophage-like RAW264.7 cells in vitro. We further confirmed that forced expression of the Spike protein in RAW264.7 cells could significantly increase the mRNA levels of the same panel of inflammatory factors. Conclusions: Our results demonstrate that the Spike protein of SARS-CoV-2 alone can induce cellular pathology, e.g. activating macrophages and contributing to induction of acute inflammatory responses.

Autophagy Inhibition Enables Nrf2 to Exaggerate the Progression of Diabetic Cardiomyopathy in Mice.

Nuclear factor-erythroid factor 2-related factor 2 (Nrf2) may either ameliorate or worsen diabetic cardiomyopathy. However, the underlying mechanisms are poorly understood. Herein we report a novel mechanism of Nrf2-mediated myocardial damage in type 1 diabetes (T1D). Global Nrf2 knockout (Nrf2KO) hardly affected the onset of cardiac dysfunction induced by T1D but slowed down its progression in mice independent of sex. In addition, Nrf2KO inhibited cardiac pathological remodeling, apoptosis, and oxidative stress associated with both onset and advancement of cardiac dysfunction in T1D. Such Nrf2-mediated progression of diabetic cardiomyopathy was confirmed by a cardiomyocyte-restricted (CR) Nrf2 transgenic approach in mice. Moreover, cardiac autophagy inhibition via CR knockout of autophagy-related 5 gene (CR-Atg5KO) led to early onset and accelerated development of cardiomyopathy in T1D, and CR-Atg5KO-induced adverse phenotypes were rescued by additional Nrf2KO. Mechanistically, chronic T1D leads to glucolipotoxicity inhibiting autolysosome efflux, which in turn intensifies Nrf2-driven transcription to fuel lipid peroxidation while inactivating Nrf2-mediated antioxidant defense and impairing Nrf2-coordinated iron metabolism, thereby leading to ferroptosis in cardiomyocytes. These results demonstrate that diabetes over time causes autophagy deficiency, which turns off Nrf2-mediated defense while switching on an Nrf2-operated pathological program toward ferroptosis in cardiomyocytes, thereby worsening the progression of diabetic cardiomyopathy.

An inhibitor role of Nrf2 in the regulation of myocardial senescence and dysfunction after myocardial infarction.

Cellular senescence, a process whereby cells enter a state of permanent growth arrest, appears to regulate cardiac pathological remodeling and dysfunction in response to various stresses including myocardial infarction (MI). However, the precise role as well as the underlying regulatory mechanism of cardiac cellular senescence in the ischemic heart disease remain to be further determined. Herein we report an inhibitory role of Nrf2, a key transcription factor of cellular defense, in regulating cardiac senescence in infarcted hearts as well as a therapeutic potential of targeting Nrf2-mediated suppression of cardiac senescence in the treatment of MI-induced cardiac dysfunction. MI was induced by left coronary artery ligation for 28 days in mice. Heart tissues from the infarct border zone were used for the analyses. The MI-induced cardiac dysfunction was associated with increased myocardial cell senescence, oxidative stress and apoptosis in adult wild type (WT) mice. In addition, a downregulated Nrf2 activity was associated with upregulated Keap1 levels and increased phosphorylation of JAK and FYN in the infarcted border zone heart tissues. Nrf2 Knockout (Nrf2-/-) enhanced the MI-induced myocardial, cardiac dysfunction and senescence. Qiliqiangxin (QLQX), a herbal medicine which could reverse the MI-induced suppression of Nrf2 activity, significantly inhibited the MI-induced cardiac senescence, apoptosis, and cardiac dysfunction in WT mice but not in Nrf2-/- mice. These results indicate that MI downregulates Nrf2 activity thus promoting oxidative stress to accelerate cellular senescence in the infarcted heart towards cardiac dysfunction and Nrf2 may be a drug target for suppressing the cellular senescence-associated pathologies in infarcted hearts.

The Dark Side of Nrf2 in the Heart.

Nuclear factor-erythroid factor 2-related factor 2 (Nrf2) is a critical transcription factor that regulates the expression of over 1000 genes in the cell under normal and stressed conditions. These transcripts can be categorized into different groups with distinct functions, including antioxidative defense, detoxification, inflammatory responses, transcription factors, proteasomal and autophagic degradation, and metabolism. Nevertheless, Nrf2 has been historically considered as a crucial regulator of antioxidant defense to protect against various insult-induced organ damage and has evolved as a promising drug target for the treatment of human diseases, such as heart failure. However, burgeoning evidence has revealed a detrimental role of Nrf2 in cardiac pathological remodeling and dysfunction toward heart failure. In this mini-review, we outline recent advances in structural features of Nrf2 and regulation of Nrf2 activity and discuss the emerging dark side of Nrf2 in the heart as well as the potential mechanisms of Nrf2-mediated myocardial damage and dysfunction.