Journal: Journal of the American Society of Nephrology

Article Title: Negative Modulation of B Cell Activation by Melanocortin 1 Receptor Signaling Protects against Membranous Nephropathy

doi: 10.1681/asn.2022050605

Figure Lengend Snippet: Figure 8. Loss of MC1R potentiates B-cell activation and en- hances autologous humoral immune responses in rats with PHN. (A) Animals were treated as elaborated in Figure 1F. Splenocytes derived from WT or KO rats with PHN were pro- cessed for flow cytometry analysis of CD1381 cells. (B) Arbitrary levels of sheep IgG-specific rat IgG in sera collected from WT or KO rats with PHN, as estimated by ELISA of diluted sera; aP,0.05 versus WT rats with PHN (n56). (C) Schematic diagram depicting the working model for negative regulation of B-cell activation by MC1R-mediated melanocortinergic signaling. As a typical G-protein coupled receptor, MC1R is expressed by B lympho- cytes and is able to trigger the cAMP signaling pathway on ac- tivation by diverse melanocortins, such as pan-MCR agonists like ACTH and NDP-MSH, or the selective MC1R agonist MS05. Subsequently, cAMP signaling downstream of MC1R activates MITF, one of the major signaling transducers of MC1R pathway and a critical regulator of a plethora of key cellular processes, including melanogenesis in melanocytes and cellular differenti- ation in nonmelanocytic cells. In B lymphocytes, MITF acts as a repressor of IRF4, a characteristic lymphoid transcription factor essential for B-cell development and maturation. MC1R-medi- ated melanocortinergic signaling suppresses B-cell activation and plasmacytic differentiation and reduces IgG production via the cAMP/MITF/IRF4 pathway, which could be intercepted by SQ22536, an adenylyl cyclase inhibitor. This MC1R-mediated B- cell regulatory signaling is defective in B cells with MC1R KO and could be partially restored by the cAMP mimetic dBcAMP.

Article Snippet: IgG levels in cell culture supernatants were detected by using a rat IgG ELISA kit (Aviva Systems Biology, San Diego, CA) according to the manufacturer’s instruction.

Techniques: Activation Assay, Derivative Assay, Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Clinical and Experimental Immunology

Article Title: The effects of maternal anti-alpha-enolase antibody expression on the brain development in offspring

doi: 10.1093/cei/uxac086

Figure Lengend Snippet: the levels of ENO1Ab in the H-ENO1Ab dams and their pups. Serum and amniotic fluid samples were collected as depicted in . Anti-ENO1 total IgG and its subtypes were measured by enzyme-linked immunosorbent assay (ELISA). The results are shown above for the dams (A, n = 6-14/group, ** P < 0.01, *** P < 0.001 vs. the control group) and the pups (B, n = 14-19 for total IgG, n = 5-8 for IgG1, n = 5-9 for IgG2a, IgG2b and IgG3, ** P < 0.01, *** P < 0.001, not significantly (NS) vs. the pups of the control group).

Article Snippet: After 2 h incubation at RT followed by repeated washing, the plates were further incubated with diluted horseradish peroxidase (HRP)-labeled goat anti-mouse total IgG (1:200 diluted for TgAb;1:10 000 for ENO1Ab), IgG1(1:1000 for ENO1Ab), IgG2a (1:1000 for ENO1Ab), IgG2b (1:1000 for ENO1Ab), or IgG3 antibodies (1:1000 for ENO1Ab), which were all from Bethyl (USA).

Techniques: Enzyme-linked Immunosorbent Assay, Control

Journal: Clinical and Experimental Immunology

Article Title: The effects of maternal anti-alpha-enolase antibody expression on the brain development in offspring

doi: 10.1093/cei/uxac086

Figure Lengend Snippet: IgG deposition and the distribution of C3, MAC, and CD16 in the brain tissue of the pups from H-ENO1Ab dams. All the pups were obtained as depicted in . (A) Representative images of IgG deposition in the brain tissues from the pups of H-ENO1Ab dams and those of the control group on P10 and P40 ( n = 4/group) after immunofluorescence (IF) staining with FITC-conjugated goat anti-mouse IgG (green, ×200). (B) Representative images for the colocalization of IgG and ENO1, NeuN, and CD34 in the brain tissues from the pups of H-ENO1Ab dams and those of the control group on P10 ( n = 4/group) after double IF staining with FITC-conjugated goat anti-mouse IgG (green) and rabbit anti-ENO1 and NeuN, rat anti-CD34 (red, ×200). (C and E) Representative images of the colocalization of IgG and C3 and CD16 in the brain tissues from the pups of H-ENO1Ab dams and those of the control group on P10 and P40 ( n = 4/group) after double IF staining with FITC-conjugated goat anti-mouse IgG (green) and rabbit anti-C3 and CD16 (red, ×200). (D) Representative images of MAC expression in the brain tissues from the pups of H-ENO1Ab dams and the those of the control group on P10 and P40 ( n = 4/group) after IF staining with mouse anti-C5b-9 (green, ×200). Nuclei were counterstained with DAPI (blue).

Article Snippet: After 2 h incubation at RT followed by repeated washing, the plates were further incubated with diluted horseradish peroxidase (HRP)-labeled goat anti-mouse total IgG (1:200 diluted for TgAb;1:10 000 for ENO1Ab), IgG1(1:1000 for ENO1Ab), IgG2a (1:1000 for ENO1Ab), IgG2b (1:1000 for ENO1Ab), or IgG3 antibodies (1:1000 for ENO1Ab), which were all from Bethyl (USA).

Techniques: Control, Immunofluorescence, Staining, Expressing