Journal: The Journal of Clinical Investigation

Article Title: Combined FXIII-C3 autoantibodies elicit bleeding and complement dysfunction in autoimmune FXIII deficiency

doi: 10.1172/JCI192619

Figure Lengend Snippet: ( A and B ) Quantification of platelets’ activation in 8 different groups of mice via CD62p surface expression analyzed by flow cytometry. A representative dot plot ( A ) and statistical chart of activated platelets ( B ) are shown. ( C ) Platelet activation upon in vitro co-culture of platelets with C . albicans . CD62p expression (left) and GPIIbIIIa expression (right) on platelet surfaces were analyzed by flow cytometry upon in vitro co-culture of platelets (1 × 10 7 /sample) with different doses of C . albicans (1 × 10 5 , 1 × 10 6 and 1 × 10 7 /sample) in the presence of Gly-Pro-Arg-Pro (Pefa 6003) to prevent platelet aggregation. ( D ) Soluble CD40L levels in plasma of 8 groups of mice on day 7 after infection. ( E ) Soluble CD40L levels in plasma of patients with FXIII deficiency and in healthy control donors. ( F ) CD40L release in the platelet- C . albicans co-culture supernatant. Data are shown as mean ± SD; 1-way ANOVA was used for statistical analysis. ** P < 0.01, *** P < 0.001. FSC-A, forward scatter area; PLA, proximity ligation assay; APC-A, allophycocyanin area.

Article Snippet: Purified CD40L (Elabscience, PKSH032214) was used as a positive control.

Techniques: Activation Assay, Expressing, Flow Cytometry, In Vitro, Co-Culture Assay, Clinical Proteomics, Infection, Control, Proximity Ligation Assay

Journal: The Journal of Clinical Investigation

Article Title: Combined FXIII-C3 autoantibodies elicit bleeding and complement dysfunction in autoimmune FXIII deficiency

doi: 10.1172/JCI192619

Figure Lengend Snippet: ( A ) Binding of CD40L to B cells analyzed by flow cytometry. ( B ) Co-localization CD40L and CD40 on the surface of B cells. Memory B cells isolated from P1 (1 × 10 6 /sample) were incubated with the platelet– C . albicans culture supernatant. Co-localization of CD40L and CD40 was examined by confocal microscopy. Representative images (out of 3) are shown. Scale bar: 10 μm. ( C ) Combined autoantibody release detected by ELISA. ( D ) Specificity of released autoantibody against C3 analyzed by ELISA. ( E ) Specificity of released autoantibody against FXIII analyzed by Far Western blot. ( F ) Neutralization of CD40L blocked autoantibody release by memory B cells. ( C – F ) CD40L-containing supernatant was added to the isolated memory B cells in the presence or absence of anti-CD40L. Purified CD40L was used as a positive control. Data are shown as mean ± SD; 1-way ANOVA was used for statistical analysis. *** P < 0.001. NC, negative control; PC, positive control; HB, memory B cells isolated from healthy donors; PB, memory B cells isolated from patient 1.

Article Snippet: Purified CD40L (Elabscience, PKSH032214) was used as a positive control.

Techniques: Binding Assay, Flow Cytometry, Isolation, Incubation, Confocal Microscopy, Enzyme-linked Immunosorbent Assay, Far Western Blot, Neutralization, Purification, Positive Control, Negative Control

Journal: The Journal of Clinical Investigation

Article Title: Combined FXIII-C3 autoantibodies elicit bleeding and complement dysfunction in autoimmune FXIII deficiency

doi: 10.1172/JCI192619

Figure Lengend Snippet: A combined FXIII-C3 autoantibody was identified in patients with autoimmune FXIII deficiency. The combined autoantibody, by forming a complex with C3, blocks C3 cleavage by C3 convertase, thereby inhibiting complement activation and complement mediated T cell activation, which increases host susceptibility to pathogenic C . albicans infections. Uncontrolled exogenous infections, by triggering platelets’ activation and platelet-related CD40L release, further promote autoantibody generation, resulting in progressive consumption of FXIII and C3. This process likely forms a self-enhanced amplification loop for autoantibody generation and progressive consumption of FXIII and C3 antigens.

Article Snippet: Purified CD40L (Elabscience, PKSH032214) was used as a positive control.

Techniques: Activation Assay, Amplification