BREAKING! Indiana University Finds That Transcriptome Of Human Pluripotent Stem Cell-Derived Cardiomyocytes Compromised By SARS-CoV-2 Nsp6, Nsp8 And M Genes!

SARS-CoV-2-Heart: Researchers from Indiana University School of Medicine have in a new study discovered that SARS-CoV-2 Nsp6, Nsp8 and M genes compromise the transcriptome of human pluripotent stem cell-derived cardiomyocytes.


 
Cardiac manifestations are commonly observed in COVID-19 patients and prominently contributed to overall mortality.
 
To date, it is already known that human myocardium could be infected by SARS-CoV-2, and human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) are susceptible to SARS-CoV-2 infection. However, molecular mechanisms of SARS-CoV-2 gene-induced injury and dysfunction of human CMs remain elusive.
 
The study team found that overexpression of three SARS-CoV-2 coding genes, Nsp6, Nsp8 and M, could compromise transcriptome of hPSC-CMs.
 
Integrated transcriptomic analyses of hPSC-CMs infected by SARS-CoV-2 with hPSC-CMs of Nsp6, Nsp8 or M overexpression identified concordantly activated genes enriched into apoptosis and immune/inflammation responses, whereas reduced genes related to heart contraction and functions.
 
It was further found that Nsp6, Nsp8 or M overexpression induce prominent apoptosis and electrical dysfunctions of hPSC-CMs.
 
Detailed interactome analysis found that Nsp6, Nsp8 and M all interact with ATPase subunits, leading to significantly reduced cellular ATP level of hPSC-CMs.
 
The study team interestingly fund that two existing U.S. FDA-approved drugs, ivermectin and meclizine, could enhance the ATP level, and ameliorate cell death and dysfunctions of hPSC-CMs overexpressing Nsp6, Nsp8 or M.
 
The study findings on the whole uncovered the detrimental impacts of SARS-CoV-2 genes Nsp6, Nsp8 and M on the whole transcriptome and interactome of hPSC-CMs, defined the crucial role of ATP level reduced by SARS-CoV-2 genes in CM death and functional abnormalities, and explored the potentially pharmaceutical approaches to ameliorate SARS-CoV-2 genes-induced CM injury and abnormalities.
 
The study findings were published on a preprint server are currently being peer reviewed. https://www.biorxiv.org/content/10.1101/2022.01.20.477147v1
 
The study team used whole messenger RNA (mRNA)-seq to investigate the overall effects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral genes Nsp6, Nsp8, and M on the transcriptome of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs).
 
It is already known that low adenosine triphosphate (ATP) levels can affect intracellular Ca2+ signaling and cardiomyocytes (CM) contractility.
 
The SARS-CoV-2-Heart study team also explored pharmacological strategies to increase the cellular ATP levels of hPSC-CMs overexpressing Nsp6, Nsp8, or M.
 
Various heart or cardiac symptoms are prevalent in COVID-19 infected patients, and they play a significant role in total mortality. SARS-CoV-2 can infect the human myocardium, and hPSC-CMs are susceptible to the virus infect ion.
 
The study team overexpressed three SARS-CoV-2 genes - Nsp6, Nsp8, and M - in hPSC-derived CMs. The team cultured human embryonic stem cell (hESC) line H9 and human-induced PSC (hiPSC) line S3 in mTesR1 102 medium and differentiated cardiomyocytes. Whole mRNA-sequencing was used to evaluate total RNA for quality and quantity.
 
The study team sequenced pooled cDNA libraries and generated approximately 30-40M 118 reads per library. Co-immunoprecipitation mass spectrometry (Co-IP-MS) was performed on the total cell proteins extracted, and Western blotting was used to analyze Co-IP protein samples.
 
Detailed data comparison between the gene overexpression and control groups was conducted with the help of an unpaired two-tailed t-test.
 
The research findings showed that ATP hemostasis impairment may play a key role in SARS-CoV-2 gene-induced CM damage in the heart and other organs/tissues that are SARS-CoV-2 targets.
 
Alarmingly, while both Nsp6OE, Nsp8OE, and MOE infection and SARS-CoV-2 infection resulted in concordant transcriptomic changes in hPSC-CMs, SARS-CoV-2 infection was responsible for 70% of differential expression genes (DEGs), implying that the other SARS-CoV-2 genes may also influence the transcriptome of human CMs through different targets or mechanisms.
 
As the SARS-CoV-2 coronavirus-induced pathways were linked to apoptosis, gene transcription, and several metabolic processes, the study findings suggest that at least some other SARS-CoV-2 genes may contribute to CM damage and impact the metabolism of human CMs.
 
The specific vulnerability of hPSC-CMs to individual SARS-CoV-2 genes was determined in this investigation. Nsp6 and Nsp8 are the SARS-CoV-2 non-structural proteins. M, the amplest structural protein in the viral particle, is the structural protein of SARS-CoV-2.
 
The study team discovered that forcing Nsp6, Nsp8, or M expression was enough to cause apoptosis and dysfunction in hPSC-CMs, which phenocopied SARS-CoV-2 infected hPSC-CMs from prior studies.
 
The whole mRNA-seq demonstrated global transcriptional alterations of Nsp6OE, Nsp8OE, and MOE human embryonic stem cell (hESC)-CMs compared to control hESC-CMs, especially with differentially expressed genes enriched in activated cellular damage and immunological responses, as well as reduced calcium/gap junction signaling.
 
The study findings suggest that exogenous SARS-CoV-2 viral genes could significantly affect the gene expression patterns of human CMs, potentially leading to CM abnormalities in patients with COVID-19.
 
The study team discovered that Nsp6, Nsp8, and M interacted with ATPase subunits and affected the cellular ATP level in hPSC-derived CMs by examining their interactome in hESC-CMs.
 
The research findings point to ATP homeostasis playing a key role in SARS-CoV-2-induced tissue damage in CMs and other SARS-CoV-2-sensitive tissue cells in the lung and kidney, while the mechanisms by which Nsp6, Nsp8, or M could hijack ATPase are still unknown.
Typically, heart muscle cells consume a lot of energy due to their constant contractions, making them one of the most sensitive cell types to a lack of ATP supply.
 
The study team examined pharmaceutical techniques to increase cellular ATP levels and revealed that two U.S. Food and Drug Administration (FDA)-approved medications, ivermectin, and meclizine, greatly decreased SARS-CoV-2 gene-induced electrical dysfunctions and cell death in human CMs.
 
Despite the fact that ivermectin is used to treat parasitic infections, it has been discovered to be a mitochondrial ATP protector in CMs and boost mitochondrial ATP generation in human CMs, which was confirmed in this investigation. https://pubmed.ncbi.nlm.nih.gov/28942281/
 
Meclizine an antihistamine may also raise ATP levels in hPSC-CMs, preventing cell death caused by the SARS-CoV-2 gene. It has been reported that meclizine had cardio-protection effect through promoting glycolysis of CMs, which increased ATP synthesis. https://pubmed.ncbi.nlm.nih.gov/20160716/
 
Overall, the study team identified the negative effects of SARS-CoV-2 genes Nsp6, Nsp8, and M on the entire transcriptome and interactome of hPSC-CMs, defined the critical role of ATP level reduction caused by SARS-CoV-2 genes in CM death and functional abnormalities, and investigated potential pharmaceutical approaches to alleviate SARS-CoV-2 genes-induced CM injury and abnormalities.
 
The study findings are critical in terms of understanding the possible causes of cardiac related issues in both COVID-19 patients and also in long COVID and ways to prevent them.
 
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