Journal: Research Square

Article Title: Immune responses in COVID-19 respiratory tract and blood reveal mechanisms of disease severity

doi: 10.21203/rs.3.rs-802084/v1

Figure Lengend Snippet: Higher anti-RBD IgM seroconversion rate in respiratory samples compared to paired blood samples of COVID-19 patients. a ELISA titration curves against the SARS-CoV-2 receptor-binding domain (RBD) for IgM, IgG, and IgA in COVID-19 respiratory and paired blood samples and non-COVID-19 respiratory samples. Dotted lines within each graph indicates the cut-off used to determine end-point titres. b Endpoint titres of SARS-CoV-2 RBD antibodies between (i) respiratory samples of COVID-19 and non-COVID-19 patients, and (ii) plasma and respiratory samples of COVID-19 patients. (i) Bars indicate median with interquartile range. Dotted line indicates the detection level. (ii) Dotted lines connect the most closely matched plasma and respiratory samples from each patient. Statistical significance was determined with Mann-Whitney test. c ELISA titration curves against the SARS-CoV-2 RBD for 3 COVID-19 patients with serial respiratory samples. d Heatmap of percentage (%) inhibition tested by surrogate virus neutralization test (sVNT) and anti-RBD ELISA titres. e Correlation between anti-RBD antibody titres and (%) sVNT inhibition. Correlation was determined with Spearman’s correlation. f Number of (i) samples and (ii) patients with seroconverted anti-RBD IgM, IgG, IgA and positive % sVNT inhibition. Pink curved lines surrounding the donut graphs indicate the samples/patients with seroconverted IgM. Earliest samples were used for each patient when determining seroconversion which was defined as average titre +2xSD of non-COVID-19 samples. Positive % sVNT inhibition was defined as % sVNT inhibition ≥ 20%.

Article Snippet: DuoSet ELISA ancillary reagent kit (R&D Systems) was used for respiratory fluids and in-house reagents with the same composition were used for plasma samples.

Techniques: Enzyme-linked Immunosorbent Assay, Titration, Binding Assay, Clinical Proteomics, MANN-WHITNEY, Inhibition, Virus, Neutralization

Journal: bioRxiv

Article Title: TRPC6-Mediated Ca 2+ Influx Activates MAPK and NFκB Signaling and Elicits Pro-Inflammatory and Catabolic Responses in Human Intervertebral Disc Cells

doi: 10.64898/2026.02.07.704609

Figure Lengend Snippet: TRPC6 activation induces and sustains inflammatory gene expression and protein secretion in human IVD cells. (A) Transcriptional profiling of inflammatory markers after 18 h of Hyp9 (1 μM) treatment. COX-2, IL-6, and IL-8 mRNA levels are presented as fold change relative (2 −ΔΔCt ) to vehicle control (dashed line, n= 17). (B-C) Time-dependent secretion of IL-6 (B) and IL-8 (C) protein into the culture media, quantified by ELISA and normalized to total DNA content presented as fold change relative to vehicle control (dashed line, n=5). (D-F) Longitudinal gene expression analysis of COX-2 (D), IL-6 (E), and IL-8 (F) over 18, 24, and 48 h, demonstrating sustained transcriptional activation (n=5, symbols matched with protein secretion). Data are presented as mean ± SEM, with individual donor points; n denotes biological replicates. Statistical significance (**p<0.01, #p<0.0001 vs. vehicle control) was determined by the Mann-Whitney test compared to vehicle control.

Article Snippet: Media were centrifuged to remove debris and analyzed using human DuoSet ELISA kits (R&D Systems, Minneapolis, MN, USA; DY008B) following the manufacturer’s instructions.

Techniques: Activation Assay, Gene Expression, Control, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

Journal: bioRxiv

Article Title: TRPC6-Mediated Ca 2+ Influx Activates MAPK and NFκB Signaling and Elicits Pro-Inflammatory and Catabolic Responses in Human Intervertebral Disc Cells

doi: 10.64898/2026.02.07.704609

Figure Lengend Snippet: TRPC6 activation elicits selective collagenolytic and aggrecan-associated transcriptional remodeling in human IVD cells. (A) Transcriptional profiling of Collagenolytic MMP markers after 18 h of Hyp9 (1 μM) treatment. MMP-1, MMP-2, MMP-3, and MMP-13 mRNA levels are presented as fold change (2 −ΔΔCt ) relative to vehicle control (dashed line, n= 17). (B-C) Time-dependent secretion of MMP-1 (B) and MMP-3 (C) protein into the culture media, quantified by ELISA and normalized to total DNA content, presented as fold change relative to vehicle control (dashed line, n=5). (D-F) Longitudinal gene expression analysis of MMP-1 (D), MMP-2 (E) MMP-3 (F), and MMP-13 (G) over 18, 24, and 48 h, demonstrating sustained transcriptional activation. (Symbols represent matched biological donors across time points; n = 5). (H) Aggrecan (ACAN) and aggrecanase gene expression following 18 h of Hyp9 treatment under serum-free conditions, including ACAN, ADAMTS4, and ADAMTS5, presented as fold change relative to vehicle control (n = 17). (I–K) Temporal transcriptional regulation of ACAN (I), ADAMTS4 (J), and ADAMTS5 (K) under serum-free conditions following Hyp9 treatment at 18, 24, and 48 h (n = 5). Data are presented as mean ± SEM with individual donor points; n denotes biological replicates. Statistical significance (**p<0.01, ***p<0.001, #p<0.0001 vs. vehicle control) was determined by the Mann-Whitney test compared to vehicle control.

Article Snippet: Media were centrifuged to remove debris and analyzed using human DuoSet ELISA kits (R&D Systems, Minneapolis, MN, USA; DY008B) following the manufacturer’s instructions.

Techniques: Activation Assay, Control, Enzyme-linked Immunosorbent Assay, Gene Expression, MANN-WHITNEY

Journal: bioRxiv

Article Title: TRPC6-Mediated Ca 2+ Influx Activates MAPK and NFκB Signaling and Elicits Pro-Inflammatory and Catabolic Responses in Human Intervertebral Disc Cells

doi: 10.64898/2026.02.07.704609

Figure Lengend Snippet: TRPC6 activation modestly modulates neurogenic and angiogenic mediators in human IVD cells. (A) Transcriptional profiling of neuro-angiogenic markers after 18 h of Hyp9 (1 μM) treatment. NGF, BDNF, and VEGF mRNA levels are presented as fold change (2 −ΔΔCt ) relative to vehicle control (dashed line, n= 17). (B-C) Time-dependent secretion of NGF (B) and VEGF (C) protein into the culture media following Hyp9 treatment, quantified by ELISA under serum-replete (10%) conditions and normalized to total DNA content, presented as fold change relative to vehicle control (dashed line, n=5). (D-F) Longitudinal gene expression analysis of NGF (D), BDNF (E), and VEGF (F) over 18, 24, and 48 h, illustrating distinct temporal regulation patterns among neurotrophic and angiogenic mediators (n=5, symbols matched with protein secretion). Data are presented as mean ± SEM with individual donor points; n denotes biological replicates. Statistical significance (**p<0.01, #p<0.0001 vs. vehicle control) was determined by the Mann-Whitney test compared to vehicle control.

Article Snippet: Media were centrifuged to remove debris and analyzed using human DuoSet ELISA kits (R&D Systems, Minneapolis, MN, USA; DY008B) following the manufacturer’s instructions.

Techniques: Activation Assay, Control, Enzyme-linked Immunosorbent Assay, Gene Expression, MANN-WHITNEY