Journal: Journal of the American Society of Nephrology

Article Title: Complement C3a and C3a Receptor Activation Mediates Podocyte Injuries in the Mechanism of Primary Membranous Nephropathy

doi: 10.1681/asn.2021101384

Figure Lengend Snippet: Figure 4. The treatments of C3aR antagonist alleviated plasma level of C3a and glomerular expression of C3aR on Heymann nephritis rats. Sprague-Dawley rats were injected with anti-Fx1A antibody serum via the tail vein. The plasma C3a was significantly elevated in disease controls (A, C). After C3aR antagonist treatments, it was decreased remarkably, both in the early-treatment groups and in the late-treatment groups: SB290157 (B), JR14a (D). In the kidneys, the staining of C3aR was strong in the rats with Heymann nephritis (E), and was attenuated in both treatment groups (F, G). *P,0.05, **P,0.01, ***P,0.001.

Article Snippet: Conditionally immortalized human podocytes, kindly provided by Prof. Jochen Reiser (Rush University Medical Center, Chicago, IL), were cultivated and allowed to differentiate as described previously.16 The differentiated podocytes on sixwell plates were cultured in RPMI 1640 medium for 6 hours to achieve synchronization, then stimulated in RPMI 1640 medium containing 10% plasma from patients with primary MN, 10% MN plasma 1 C3aR antagonist SB290157 (HY101502A; MedChemExpress, Monmouth Junction, NJ), 10% MN plasma 1 C3aR antagonist JR14a (HY-138161; MedChemExpress), 10% MN plasma 1 recombinant human C3a (2 mg/ml, 3677-C3; R&D Systems, Minneapolis, MN), 10% plasma from healthy controls, C3a, or 5% FBS (10100147; Gibco, CA).

Techniques: Clinical Proteomics, Expressing, Injection, Staining

Journal: Journal of the American Society of Nephrology

Article Title: Complement C3a and C3a Receptor Activation Mediates Podocyte Injuries in the Mechanism of Primary Membranous Nephropathy

doi: 10.1681/asn.2021101384

Figure Lengend Snippet: Figure 5. The treatments of C3aR antagonist inhibited specific IgG and complement activation on Heymann nephritis rats. Sprague-Dawley rats were injected with anti-Fx1A antibody serum via the tail vein. In the early-treatment groups, C3aR antagonists (SB290157 or JR14a) was administrated daily from day 0. In the late-treatment groups, both C3aR antagonists were administrated from week 3 after proteinuria was detected. Compared with the disease controls treated with normal saline, the plasma level of spe- cific anti-sheep IgG was decreased in treatment groups of both C3aR antagonists, SB290157 (A) and JR14a (B), whereas the level of total IgG was comparable among the groups. In the glomeruli, the deposit of sheep IgG (C) was comparable among the disease controls and treatment groups. However, the deposit of rat IgG (D) was alleviated in both treatment groups. Along with that, the staining of C1q (E), factor B (F), and C5b-9 (G) was significantly decreased in the treatment groups. *P,0.05, **P,0.01, ***P,0.001.

Article Snippet: Conditionally immortalized human podocytes, kindly provided by Prof. Jochen Reiser (Rush University Medical Center, Chicago, IL), were cultivated and allowed to differentiate as described previously.16 The differentiated podocytes on sixwell plates were cultured in RPMI 1640 medium for 6 hours to achieve synchronization, then stimulated in RPMI 1640 medium containing 10% plasma from patients with primary MN, 10% MN plasma 1 C3aR antagonist SB290157 (HY101502A; MedChemExpress, Monmouth Junction, NJ), 10% MN plasma 1 C3aR antagonist JR14a (HY-138161; MedChemExpress), 10% MN plasma 1 recombinant human C3a (2 mg/ml, 3677-C3; R&D Systems, Minneapolis, MN), 10% plasma from healthy controls, C3a, or 5% FBS (10100147; Gibco, CA).

Techniques: Activation Assay, Injection, Saline, Clinical Proteomics, Staining

Journal: iScience

Article Title: Feeder-free generation and characterization of endocardial and cardiac valve cells from human pluripotent stem cells

doi: 10.1016/j.isci.2023.108599

Figure Lengend Snippet:

Article Snippet: Isolated VICs and VECs were plated on 12-chamber microscopy slides (Ibidi) coated with human fibronectin plasma (1:100, Sigma).

Techniques: Recombinant, SYBR Green Assay, Software, Plasmid Preparation, Cell Counting

Journal: The Lancet. Rheumatology

Article Title: Autoantibodies against interleukin-1 receptor antagonist in multisystem inflammatory syndrome in children: a multicentre, retrospective, cohort study

doi: 10.1016/S2665-9913(22)00064-9

Figure Lengend Snippet: Neutralising and functional effect of anti-IL-1Ra antibodies in MIS-C (A) Free IL-1Ra plasma concentrations as measured by ELISA in patients with MIS-C (n=21), Kawasaki disease (n=6) and systemic juvenile idiopathic arthritis (n=10). Horizontal lines represent the mean and SD. Data were analysed by Brown–Forsythe and Welch ANOVA with Dunnett's T3 multiple comparisons. (B) IL-1β-signalling reporter assay on selected MIS-C plasma compared with an adult critical COVID-19 plasma sample (both 1:20 dilution) as well as commercially available anti-IL-1Ra antibody or control (anti-SLP2) antibody. The absorbance of secreted embryonic alkaline phosphatase, as a marker for IL-1β pathway activation in HEK IL-1β reporter cells, was detected at 650 nm. Error bars show mean (SD). MIS-C=multisystem inflammatory syndrome in children. IL-1Ra=interleukin-1 receptor antagonist. KD=Kawasaki disease. sJIA=systemic juvenile idiopathic arthritis. TNF=tumour necrosis factor. IL-1β=interleukin-1β.

Article Snippet: Recombinant IL-1Ra at 40 ng/mL (Biozol, Eching, Germany) alone or with either rabbit antihuman IL-1Ra antibody at 5 μg/mL (antibodies-online, Aachen, Germany), recombinant anti-SLP2 antibody at 5 μg/mL (Abcam, Cambridge, UK), diluted plasma (1:20) from a patient with acute MIS-C and high-titred (1:800) anti-IL-1Ra antibodies, or diluted plasma (1:20) from the same patient 7 months after onset of MIS-C but without detectable anti-IL-1Ra antibodies, were preincubated for 2 h at room temperature.

Techniques: Functional Assay, Enzyme-linked Immunosorbent Assay, Reporter Assay, Marker, Activation Assay