Since the emergence of the COVID-19 pandemic, the effects of SARS-CoV-2 have been extensively researched. While much is already known about the acute phase of the infection, increasing attention has turned to the prolonged symptoms experienced by a subset of individuals, commonly referred to as long COVID-19 patients. This study aims to delve deeper into the immune landscape of patients with prolonged symptoms by implementing single-cell mRNA analysis. A 71-year-old COVID-19 patient presenting with persistent viral pneumonia was recruited, and peripheral blood samples were taken at 3 and 2 years post-acute infection onset. Patients and control peripheral blood mononuclear cells (PBMCs) were isolated and single-cell sequenced. Immune cell population identification was carried out using the ScType script. Three months post-COVID-19 patients’ PBMCs contained a significantly larger immature neutrophil population compared to 2-year and control samples. However, the neutrophil balance shifted towards a more mature profile after 18 months. In addition, a notable increase in the CD8+ NKT-like cells could be observed in the 3-month patient sample as compared to the later one and control. The subsequent change in these cell populations over time may be an indicator of an ongoing failure to clear the SARS-CoV-2 infection and, thus, lead to chronic COVID-19 complications.

Bacteriophage-derived dsRNA, known as Larifan, is a nationally well-known broad-spectrum antiviral medication. This study aimed to ascertain the antiviral activity of Larifan against the novel SARS-CoV-2 virus. Larifan’s effect against SARS-CoV-2 in vitro was measured in human lung adenocarcinoma (Calu3) and primary human small airway epithelial cells (HSAEC), and in vivo in the SARS-CoV-2 infection model in golden Syrian hamsters. Larifan inhibited SARS-CoV-2 replication both in vitro and in vivo. Viral RNA copy numbers and titer of infectious virus in the supernatant of Calu3 cells dropped significantly: p = 0.0296 and p = 0.0286, respectively. A reduction in viral RNA copy number was also observed in HSAEC, especially when Larifan was added before infection (p = 0.0218). Larifan markedly reduced virus numbers in infected hamsters’ lungs post-infection, with a more pronounced effect after intranasal administration (p = 0.0032). The administration of Larifan also reduced the amount of infections virus titer in the lungs (p = 0.0039). Improvements in the infection-induced pathological lesion severity in the lungs of animals treated with Larifan were also demonstrated. The inhibition of SARS-CoV-2 replication in vitro and the reduction in the viral load in the lungs of infected hamsters treated with Larifan alongside the improved lung histopathology suggests a potential use of Larifan in also controlling the COVID-19 disease in humans.

Resistance to Vemurafenib (VEM), a targeted BRAFV600E inhibitor, was examined in the metastatic paratetraploid (XX, abnormal Y chromosome) melanoma cell line, SkMel28. During the first week of treatment, pERK suppression coincided with transcriptomic and phenotypic changes related to senescence, autophagy/mitophagy, neuro-melanogenesis, and cell co-alignment. By the second week, MAPK-ERK signalling was restored, coinciding with surmounting the G1/S checkpoint, G2M checkpoint delay, mitotic slippage (MS), and downregulation of senescence and melanogenesis. The dynamics of melanogenesis and MS were highly correlated. By days 12-15, ∼8% of cells with melanin remnants exhibited hyperploidy and multinucleation, some arranged as rosettes, encased by a Zona pellucida-positive structure reminiscent of oocytes, zygotes, or blastulae, occasionally yielding cellularised sub-cells or stalling in diapause. These parasexual processes eventually ceased; cells resumed proliferative clonogenic growth and their initial mito-meiotic, mesenchymal profile. Transcriptomic analysis confirmed the reversal of their cell fate direction: from senescence-induced neuro-melanogenesis to its suppression and activation of female meiosis-like and mitosis states. The transition point of this cell-fate reversal coincided with S-phase resumption, highlighted by replication delay and activation of the FOS-TEAD/Hippo axis of the "female pregnancy" (stress-response, embryonal placentation, vascularisation, stemness, anti-apoptosis) gene ontology module. We conclude that resistance to VEM in SkMel28 cells encompasses the transition between three possible cell fates: (1) senescence/differentiation, (2) reprogramming/blastulation, and (3) recovery of the proliferative mito-meiotic profile. The coexpression of senescence, reprogramming and gametogenetic genes in a dataset of late-stage melanoma patient samples supports these results.

Background: IgA nephropathy (IgAN) is characterised by the production of galactose-deficient IgA1 (Gd-IgA1) antibodies. As the source of pathogenic antibodies, B cells are central to IgAN pathogenesis, but the B cell activation pathways as well as the potential B cell source of dysregulated IgA-secretion remain unknown. Methods: We carried out flow cytometry analysis of peripheral blood B cells in patients with IgA nephropathy and control subjects with a focus on IgA-expressing B cells to uncover the pathways of B cell activation in IgAN and how these could give rise to pathogenic GdIgA1 antibodies. Results: In addition to global changes in the B cell landscape – expansion of naive and reduction in memory B cells – IgAN patients present with an increased frequency of IgA-expressing B cells that lack the classical memory marker CD27, but are CD21 ^pos . IgAN patients further have an expanded population of IgA ^pos antibody-secreting cells, which correlate with serum IgA levels. Both IgA ^pos plasmabalsts and CD27 ^neg B cells co-express GdIgA1. Implicating dysregulation at mucosal surfaces as the driver of such B cell differentiation, we found a correlation between lipopolysaccharide (LPS) in the serum and IgA ^pos CD27 ^neg B cell frequency. Conclusion: We propose that dysregulated immunity in the mucosa may drive de novo B cell activation within germinal centres, giving rise to IgA ^pos CD27 ^neg B cells and subsequently IgA-producing plasmablasts. These data integrate B cells into the paradigm of IgAN pathogenesis and allow to further investigate this pathway to uncover biomarkers and develop therapeutic interventions. Key learning points: What was known: Patients with IgA nephropathy (IgAN) have aberrant production of galactose-deficient IgA1 (Gd-IgA1) and antibodies against it, which together form immune complexes that are deposited in the renal mesangium and lead to kidney damage; this is known as the multi-hit model of IgAN pathogenesis. The multi-hit model centrally implicates B cells as they produce both Gd-IgA1 and antibodies against it, yet B cell activation pathways that lead to aberrant antibody production are absent from the model. Only isolated reports exist describing specific features of B cells that are altered in patients with IgAN, including a reduction in regulatory B cells, increase in toll-like receptor 7 expression in total peripheral blood B cells and elevated frequency of circulating CCR9 ⁺ IgA ⁺ B cells. This study adds: In addition to changes in the overall circulating B cell landscape, differentiation of IgA ⁺ plasmablasts is enhanced in patients with IgAN and their levels correlate with serum IgA. IgA-expressing plasmablast frequency correlates with that of IgA ⁺ CD21 ⁺ B cells, that lack the classical memory B cell marker CD27. Both IgA ⁺ plasmablasts and IgA-expressing CD27 ^- B cells co-express GdIgA1 receptors. IgA ⁺ CD27 ^- CD21 ⁺ B cell frequency correlates with serum lipopolysaccharide (LPS) levels, implicating mucosa in their activation. Potential impact: We uncover the previously unknown B cell activation pathway that appears to be associated with pathogenic IgA secretion in IgAN and integrate this into the multi-hit model of IgAN pathogenesis. This pathway holds potential for further investigation to identify biomarkers and therapeutic targets in IgAN.