Journal: PLOS Pathogens

Article Title: Salmonella infection and biofilms drive anti-C1q and anti-dsDNA autoantibodies in systemic autoimmunity

doi: 10.1371/journal.ppat.1014666

Figure Lengend Snippet: ( A-B ) Mouse serum was assayed using a multiplex Luminex platform and antibodies quantified by mean fluorescence intensity (MFI); only anti-C1q antibodies are shown here, see for other antigens. 6-8 week old ( A ) C57BL/6 mice and ( B ) NZBxW/F1 mice were injected intraperitoneally with curli purified from Salmonella Typhimurium (STm) biofilms, or sterile PBS for controls, weekly for ( A ) 12 weeks or ( B ) 5 weeks. ( C-F ) Anti-C1q antibodies in serum (diluted 1:50) were detected using ELISA, normalized to aliquots of serum from an untreated aged MRL/lpr mouse as a positive control (1:50, top dotted line) and a negative control without serum (bottom dotted line). ( C ) C57BL/6 mice were injected intraperitoneally with 100 µg curli purified from STm biofilms, 100 µg/kg LPS, 50 µg curli and 50 µg/kg LPS, or PBS for untreated controls, weekly for 3 weeks . ( D ) C57BL/6 mice were injected intraperitoneally with 100 µg curli purified from the biofilm of an E. coli isolate from the human microbiome (hEC), an E. coli isolate from the mouse microbiome (mEC), STm biofilms, or PBS for untreated controls, weekly for 4 weeks . ( E ) 129Sv mice were infected intragastrically with 10 7 CFU of wild-type (WT) STm strain IR715, the isogenic curli-deficient mutant csgBA , or LB broth for uninfected controls, in two replicate experiments, and Week 6 serum was assayed. ( F ) C57BL/6 mice were infected intragastrically with 10 7 CFU of STm spiB or invAspiB mutant, or LB broth for uninfected controls, and monitored for 10 weeks. Week -1 denotes baseline samples collected from each group one week before the first injection (Week 0). Statistics: ( A-B ) Unpaired t-test, ( C-E ) One-way ANOVA, or ( E ) Mixed-effects analysis, with Tukey’s multiple comparisons, comparisons within the legend indicating overall column factor (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001).

Article Snippet: Tissue was incubated in antibody cocktails including 1:30 goat anti-mouse IgG conjugated to AlexaFluor 405 (Invitrogen A31553), 1:25 rat anti-mouse C3 (11H9) conjugated to AlexaFluor 647 (Santa Cruz Biotechnology sc-58926), and 1:20 rat anti-mouse C1q (7H8) conjugated to FITC (Santa Cruz Biotechnology sc-58920) diluted in PBS-T with 1% bovine serum albumin for 2 hours at room temperature, covered with a small circle of parafilm and kept in the dark underneath a lid to prevent evaporation and bleaching.

Techniques: Multiplex Assay, Luminex, Fluorescence, Injection, Purification, Sterility, Enzyme-linked Immunosorbent Assay, Positive Control, Negative Control, Microbiome, Infection, Mutagenesis, Analysis

Journal: PLOS Pathogens

Article Title: Salmonella infection and biofilms drive anti-C1q and anti-dsDNA autoantibodies in systemic autoimmunity

doi: 10.1371/journal.ppat.1014666

Figure Lengend Snippet: Serum was collected from ( A, D ) pre-onset, ( B, E ) early-disease, and ( C,F ) late-disease NZBxW/F1 mice 24 hours after each weekly injection of curli purified from Salmonella Typhimurium biofilms or PBS for untreated controls. ( A-C ) ELISA assays detected anti-dsDNA antibodies in serum diluted ( A, B ) 1:250 or ( C ) 1:500 ( A-B, N = 8; C, N = 15). Week -1 denotes baseline samples collected from each group one week before the first injection (Week 0). ( D-F ) ELISA assays detected anti-C1q antibodies in serum diluted 1:50, normalized to the O. D. of positive control MRL/lpr serum, indicated by the top dotted line, and negative controls without serum are indicated by the bottom dotted line ( D-E, pre-bleed and Week 4 not assayed due to insufficient serum volume for the assay). Statistics: ( A-B, D-E ) Two-way ANOVA, or ( C, F ) Mixed-effects model, with Tukey’s multiple comparisons, comparisons within legends indicating overall column factor, (*p < 0.05, **p < 0.01).

Article Snippet: Tissue was incubated in antibody cocktails including 1:30 goat anti-mouse IgG conjugated to AlexaFluor 405 (Invitrogen A31553), 1:25 rat anti-mouse C3 (11H9) conjugated to AlexaFluor 647 (Santa Cruz Biotechnology sc-58926), and 1:20 rat anti-mouse C1q (7H8) conjugated to FITC (Santa Cruz Biotechnology sc-58920) diluted in PBS-T with 1% bovine serum albumin for 2 hours at room temperature, covered with a small circle of parafilm and kept in the dark underneath a lid to prevent evaporation and bleaching.

Techniques: Injection, Purification, Enzyme-linked Immunosorbent Assay, Positive Control

Journal: PLOS Pathogens

Article Title: Salmonella infection and biofilms drive anti-C1q and anti-dsDNA autoantibodies in systemic autoimmunity

doi: 10.1371/journal.ppat.1014666

Figure Lengend Snippet: ( A ) Experimental scheme: MRL/lpr mice at age 6 weeks underwent weekly intraperitoneal injections of curli, purified from Salmonella Typhimurium biofilms or PBS for untreated controls, and periodic serum samples, taken 24 hours after each injection, for 6 weeks. ( B-C ) ELISA assays detected anti-dsDNA antibodies in serum diluted ( B ) 1:250 or ( C ) 1:500. ( D ) ELISA assay detected anti-C1q antibodies in serum diluted 1:50, normalized to the O.D. of the positive control untreated MRL/lpr serum. Dotted lines indicate relative O.D. of MRL/lpr positive control serum (top line) or negative control with no serum added (bottom line). Week -1 denotes baseline samples collected from each group one week before the first injection (Week 0). Statistics: Two-way ANOVA with Sidak’s multiple comparisons, comparisons within legends indicating overall column factor, (*p < 0.05, **p < 0.01).

Article Snippet: Tissue was incubated in antibody cocktails including 1:30 goat anti-mouse IgG conjugated to AlexaFluor 405 (Invitrogen A31553), 1:25 rat anti-mouse C3 (11H9) conjugated to AlexaFluor 647 (Santa Cruz Biotechnology sc-58926), and 1:20 rat anti-mouse C1q (7H8) conjugated to FITC (Santa Cruz Biotechnology sc-58920) diluted in PBS-T with 1% bovine serum albumin for 2 hours at room temperature, covered with a small circle of parafilm and kept in the dark underneath a lid to prevent evaporation and bleaching.

Techniques: Purification, Blood/Plasma/Serum Samples, Injection, Enzyme-linked Immunosorbent Assay, Positive Control, Negative Control

Journal: PLOS Pathogens

Article Title: Salmonella infection and biofilms drive anti-C1q and anti-dsDNA autoantibodies in systemic autoimmunity

doi: 10.1371/journal.ppat.1014666

Figure Lengend Snippet: MRL/lpr mice were intraperitoneally injected weekly with ( A ) PBS or ( B ) curli purified from Salmonella Typhimurium for 6 weeks and euthanized at 13 weeks old. Frozen tissue sections were stained with goat anti-mouse IgG-AlexaFluor405 (blue), rat anti-mouse C3-AlexaFluor647 (red), and rat anti-mouse C1q-FITC (green). ( A-B ) Representative images of glomeruli by confocal microscopy (63X objective). Sections shown are from two individual mice per group, scale bars represent 20 µm. ( C-F ) Images were quantified in ImageJ for mean fluorescence intensity (MFI) inside glomeruli of ( C ) IgG, ( D ) C3, ( E ) C1q, and ( F ) area was measured, within at least 15 glomeruli per mouse and averaged. Statistics: ( C-F ) Unpaired t-test (**p < 0.01).

Article Snippet: Tissue was incubated in antibody cocktails including 1:30 goat anti-mouse IgG conjugated to AlexaFluor 405 (Invitrogen A31553), 1:25 rat anti-mouse C3 (11H9) conjugated to AlexaFluor 647 (Santa Cruz Biotechnology sc-58926), and 1:20 rat anti-mouse C1q (7H8) conjugated to FITC (Santa Cruz Biotechnology sc-58920) diluted in PBS-T with 1% bovine serum albumin for 2 hours at room temperature, covered with a small circle of parafilm and kept in the dark underneath a lid to prevent evaporation and bleaching.

Techniques: Injection, Purification, Tissue, Staining, Confocal, Microscopy, Fluorescence

Journal: Frontiers in Neuroscience

Article Title: Microglial regulation of synaptic plasticity in transsynaptic degeneration of glaucoma

doi: 10.3389/fnins.2026.1828085

Figure Lengend Snippet: Microglia-mediated synaptic plasticity (schematic diagram, created with BioRender). Under physiological conditions in the CNS, microglia precisely regulate synaptic plasticity through multiple canonical signaling pathways. (1) “Find-me” signals: CX3CL1 exists in membrane-bound (mCX3CL1) and soluble forms (sCX3CL1), mediating adhesion and chemotaxis, respectively, both of which bind to microglial CX3CR1 to promote targeted recruitment to synaptic sites. (2) “Eat-me” signals: PS exposure on less active synapses is recognized by microglial receptors such as GPR56 and MERTK, promoting synaptic engulfment. Complement signaling also plays a central role, with C1q and C3 deposition facilitating C3b generation and binding to CR3 on microglia, thereby driving phagocytosis. (3) “Do not-eat-me” signals: neuronal CD47 and CD200 interact with microglial SIRP α and CD200R, respectively, suppressing excessive synaptic pruning. In glaucoma, microglia show increased numbers and morphological activation, colocalizing with degenerating RGC synaptic elements, alongside abnormal complement activation (notably upregulated C1q), suggesting their involvement in synaptic remodeling, although the precise mechanisms remain to be elucidated.

Article Snippet: Regarding complement-targeted inhibition strategies, the C1q inhibitor ANX007 (Annexon Biosciences) has completed a phase I clinical trial.

Techniques: Protein-Protein interactions, Membrane, Chemotaxis Assay, Binding Assay, Activation Assay

Journal: Journal of Neuroinflammation

Article Title: Targeting microglial C1q alleviates blood–brain barrier disruption in the thalamus after cortical infarction

doi: 10.1186/s12974-026-03822-2

Figure Lengend Snippet: Single-cell RNA sequencing identifies microglia as the primary source of C1q in ipsilateral VPN of thalamus after dMCAO. A Scheme of scRNA-seq. B Umap of 11 major cell populations identified in the ipsilateral VPN in the Sham and dMCAO groups. Left: Integration of data from the Sham and dMCAO groups. Top middle: The Sham group. Bottom middle: The dMCAO group. Right: Stacked bar graphs of cell type proportions in the Sham and dMCAO groups ( n = 4 in each group). C Cell type annotation was performed on the basis of the expression of well-established marker genes. D Feature plots showing the expression of known cell markers. E Feature plots showing the expression of Tmem119, C1qa, C1qb and C1qc. F Co-stained with C1q (red) and Iba-1 (green), GFAP (green), NeuN (green) within ipsilateral VPN in Sham and dMCAO groups. Scale bar: 25 µm. G Umap of eight cell populations identified by microglial reclustering. H Proportions of all microglial subclusters in Sham and dMCAO groups. I Feature plots showing the expression of eight microglial subclusters

Article Snippet: Endothelial cells were treated with 1% DMSO, LPS, LPS-treated BV2 cell culture medium, C1q recombinant protein (Medchemexpress, Cat# HY-NP0194C), or C1q neutralizing antibody (GeneTex, Cat# GTX54404), separately.

Techniques: Single Cell, RNA Sequencing, Expressing, Marker, Staining

Journal: Journal of Neuroinflammation

Article Title: Targeting microglial C1q alleviates blood–brain barrier disruption in the thalamus after cortical infarction

doi: 10.1186/s12974-026-03822-2

Figure Lengend Snippet: Knockdown of microglial C1qa alleviates BBB disruption in the VPN of ipsilateral thalamus at 2 w after dMCAO. A The AAV vector maps and study design/timeline of left VPN of thalamus with AAV vectors carrying the CX3CR1 promoter. B Representative photomicrographs show the co-localization of GFP (green) and Iba-1 (red) in ipsilateral VPN at 2 w after dMCAO from animals with AAV- shC1qa injection. Scale bar: 25 μm. C Representative images show the co-localization of GFP (green) and C1q (red) in ipsilateral VPN at 2 w after dMCAO from animals with AAV injection. Scale bar: 25 μm. D Representative western blot showing the expression of C1qa in different AAV- shC1qa treatment sequence in Sham groups. Statistical analysis was performed using one-way ANOVA followed by Tamhane’s T2 test. Each bar represents the mean ± SD. * p < 0.05 vs AAV-shCon group at 2 w after dMCAO ( n = 4 in each group). E On day 14 post operation, the expressions of C1qa, ZO-1, OCLN, CLDN5, and VE-cad in ipsilateral VPN from Sham and dMCAO animals with AAV- shC1qa injection were showed through representative western blot. F - J On day 14 post operation, quantitative analyses of C1qa, ZO-1, OCLN, CLDN5, and VE-cad level relative to GAPDH in the ipsilateral VPN from Sham and dMCAO animals with AAV- shC1qa injection were conducted ( n = 4 in each group). Statistical analysis was performed using one-way ANOVA followed by Tamhane’s T2 test. (K) On day 14 post operation, representative images of extravascular albumin, IgG or fibrinogen deposits and CD8a positive cells in ipsilateral VPN from Sham and dMCAO animals with AAV- shC1qa injection were displayed. Scale bar: 25 μm. (L-O) On day 14 post operation, quantitative analyses of the area of albumin, IgG and fibrinogen leakage and the number of CD8a positive cells in the ipsilateral VPN from Sham and dMCAO animals with AAV- shC1qa injection were conducted ( n = 6 in each group). Statistical analysis was performed using Kruskal–Wallis H test. Each bar represents the mean ± SD. * p < 0.05 vs Sham + AAV-shCon group, # p < 0.05 vs dMCAO + AAV-shCon group, & p < 0.05 vs Sham + AAV- shC1qa group

Article Snippet: Endothelial cells were treated with 1% DMSO, LPS, LPS-treated BV2 cell culture medium, C1q recombinant protein (Medchemexpress, Cat# HY-NP0194C), or C1q neutralizing antibody (GeneTex, Cat# GTX54404), separately.

Techniques: Knockdown, Disruption, Plasmid Preparation, Injection, Western Blot, Expressing, Sequencing

Journal: Journal of Neuroinflammation

Article Title: Targeting microglial C1q alleviates blood–brain barrier disruption in the thalamus after cortical infarction

doi: 10.1186/s12974-026-03822-2

Figure Lengend Snippet: The interaction between microglia and endothelial cells and the activation of endothelial cells in the ipsilateral VPN of thalamus after dMCAO. A List of the molecular ligand-receptor pairing between microglia and endothelial cells from Sham and dMCAO groups. B Feature plots showing the expression of vWF and CD93. C Double-staining of Laminin (green) with CD93 (red) in the ipsilateral VPN in Sham and dMCAO groups. Scale bar: 25 μm. D Quantitative analyses of the expression of CD93 in Sham and dMCAO groups ( n = 6 in each group). Statistical analysis was performed using two-tailed t -test. E Representative western blot showing the expression of CD93 in VPN at 1–4 w after dMCAO. Statistical analysis was performed using one-way ANOVA followed by Tamhane’s T2 test. F Immunoprecipitation assay shows the levels of C1qa and CD93 in ipsilateral VPN in Sham and dMCAO groups ( n = 4 in each group). Statistical analysis was performed using two-tailed t -test. G Double-staining of C1q (red) with CD93 (green) in the ipsilateral VPN in Sham and dMCAO groups. Scale bar: 25 μm. H Representative western blot showing the expressions of VCAM-1 and ICAM-1 in VPN at 1–4 w after dMCAO. Quantitative analyses of VCAM-1 and ICAM-1 level relative to GAPDH ( n = 4 in each group). Statistical analysis was performed using one-way ANOVA followed by Tamhane’s T2 test. Each bar represents the mean ± SD. * p < 0.05 vs Sham group, # p < 0.05 vs contra. I Double-staining of VCAM-1 (green) with RECA-1 (red) in the ipsilateral VPN in Sham and dMCAO groups. Scale bar: 25 μm. J Colocalization analyses of VCAM-1 (green) and RECA-1 (red) in Sham and dMCAO groups

Article Snippet: Endothelial cells were treated with 1% DMSO, LPS, LPS-treated BV2 cell culture medium, C1q recombinant protein (Medchemexpress, Cat# HY-NP0194C), or C1q neutralizing antibody (GeneTex, Cat# GTX54404), separately.

Techniques: Activation Assay, Expressing, Double Staining, Two Tailed Test, Western Blot, Immunoprecipitation

Journal: Journal of Neuroinflammation

Article Title: Targeting microglial C1q alleviates blood–brain barrier disruption in the thalamus after cortical infarction

doi: 10.1186/s12974-026-03822-2

Figure Lengend Snippet: Microglial C1q activates endothelial cells via CD93 in the ipsilateral VPN of thalamus after dMCAO. A Representative images show the co-localization of RECA-1 (red) and VCAM-1 (grey) in ipsilateral VPN from Sham and dMCAO animals with AAV- shC1qa injection. Scale bar: 25 μm. B Immunoprecipitation assay shows the levels of C1qa and CD93 in ipsilateral VPN in Sham and dMCAO animals with AAV- shC1qa injection ( n = 4 in each group). Statistical analysis was performed using one-way ANOVA followed by Bonferroni test. C Representative western blot showing the expressions of VCAM-1 in ipsilateral VPN in Sham and dMCAO animals with AAV- shC1qa injection ( n = 4 in each group). Quantitative analysis of VCAM-1 level relative to GAPDH. Statistical analysis was performed using one-way ANOVA followed by Tamhane’s T2 test. * p < 0.05 vs Sham + AAV-shCon group, # p < 0.05 vs dMCAO + AAV-shCon group. D Representative western blot showing the expression of C1qa in BV2 cells with LPS treatment ( n = 4 in each group). Quantitative analysis of C1qa level relative to GAPDH. E Representative western blot showing the expression of VCAM-1 in brain endothelial cells with LPS treatment ( n = 4 in each group). Quantitative analysis of VCAM-1 level relative to GAPDH ( n = 4 in each group). Statistical analysis was performed using one-way ANOVA followed by Tamhane’s T2 test. Each bar represents the mean ± SD. * p < 0.05 vs Veh group. F Representative western blot showing the expression of VCAM-1 in each group ( n = 4 in each group). Quantitative analysis of VCAM-1 level relative to GAPDH. Each bar represents the mean ± SD. * p < 0.05 vs Veh group, # p < 0.05 vs BV2 or r-C1q groups. G Flow chart illustrates drug administration and experimental design. Rats received intracerebroventricular injection of DCBY02 or Veh at 3 days prior to dMCAO and every 2 days thereafter until the end of experiment. H – L Representative western blot showing the expressions of VCAM-1, ZO-1, OCLN, CLDN5, and VE-cad in ipsilateral VPN, respectively. Quantitative analyses of VCAM-1, ZO-1, OCLN, CLDN5, and VE-cad level relative to GAPDH from Sham and dMCAO animals with DCBY02 or Veh injection ( n = 4 in each group). Statistical analysis was performed using one-way ANOVA followed by Bonferroni test. Each bar represents the mean ± SD. * p < 0.05 vs Sham + Veh group, # p < 0.05 vs dMCAO + Veh group

Article Snippet: Endothelial cells were treated with 1% DMSO, LPS, LPS-treated BV2 cell culture medium, C1q recombinant protein (Medchemexpress, Cat# HY-NP0194C), or C1q neutralizing antibody (GeneTex, Cat# GTX54404), separately.

Techniques: Injection, Immunoprecipitation, Western Blot, Expressing

Journal: Journal of Neuroinflammation

Article Title: Targeting microglial C1q alleviates blood–brain barrier disruption in the thalamus after cortical infarction

doi: 10.1186/s12974-026-03822-2

Figure Lengend Snippet: Minocycline alleviates BBB disruption by downregulating microglial C1q-mediated VCAM-1 upregulation in ipsilateral VPN after dMCAO. A Binding conformation of minocycline to C1q. Left: overall view of minocycline in the binding position of C1q. Middle: three-dimensional (3D) close-up view of the binding conformation of minocycline on the C1q structure. Right: the enlarged image of the binding conformation of minocycline on the C1q structure. B Free energy distribution profile of the C1q-minocycline complex. C Experimental design/timeline of minocycline injection. D Representative western blot showing the expression of C1qa in different dose of minocycline treatment (+ : 10 mg/kg; + + : 20 mg/kg; + + + : 40 mg/kg, n = 3 in each group). Statistical analysis was performed using one-way ANOVA followed by Tamhane’s T2 test. Each bar represents the mean ± SD. * p < 0.05 vs Sham + Veh group, # p < 0.05 vs dMCAO + Veh group. E Representative western blot showing the expressions of C1qa, VCAM-1, ZO-1, OCLN, CLDN5, and VE-cad in ipsilateral VPN, respectively. F - K Quantitative analyses of C1qa, VCAM-1, ZO-1, OCLN, CLDN5, and VE-cad level relative to GAPDH from Sham and dMCAO animals with MC or Veh administration ( n = 4 in each group). Statistical analysis was performed using one-way ANOVA followed by Tamhane’s T2 test. L Representative images show the co-localization of Iba-1 (green), C1q (red) and DAPI (blue) in ipsilateral VPN from dMCAO animals with MC or Veh administration. Scale bar: 25 μm. M Quantitative analysis of Iba-1 + -C1q + cells from Sham and dMCAO animals with MC or Veh administration ( n = 6 in each group). Statistical analysis was performed using one-way ANOVA followed by Tamhane’s T2 test. N Representative images show the co-localization of VCAM-1 (green) and RECA-1 (red) in ipsilateral VPN from dMCAO animals with MC administration. Scale bar: 25 μm. O Representative microphotographs of immunostaining of NeuN, GFAP, and Iba-1 in ipsilateral VPN. Scale bar: 75 µm. (P-R) Quantitative analyses of NeuN positive cells ( P ), and GFAP ( Q ) and Iba-1 ( R ) positive densities were showed from ( O ) in ipsilateral VPN from Sham and dMCAO animals with MC or Veh administration ( n = 6 in each group). Statistical analysis was performed using one-way ANOVA followed by Bonferroni test. Each bar represents the mean ± SD. * p < 0.05 vs Sham + Veh group, # p < 0.05 vs dMCAO + Veh group, & p < 0.05 vs Sham + MC group

Article Snippet: Endothelial cells were treated with 1% DMSO, LPS, LPS-treated BV2 cell culture medium, C1q recombinant protein (Medchemexpress, Cat# HY-NP0194C), or C1q neutralizing antibody (GeneTex, Cat# GTX54404), separately.

Techniques: Disruption, Binding Assay, Injection, Western Blot, Expressing, Immunostaining