Journal: Disease Models & Mechanisms

Article Title: RhoB blockade selectively inhibits autoantibody production in autoimmune models of rheumatoid arthritis and lupus

doi: 10.1242/dmm.029835

Figure Lengend Snippet: Anti-RhoB Ig inhibits autoantibody production and attenuates arthritis. K/BxN mice were treated with 500 µg anti-RhoB Ig at (A-D,F) 21 days or (E) 28 days of age. (A) The number of anti-GPI ASCs was measured by ELISpot at the termination of the experiment (6 weeks of age). Data show means±s.e.m. for n =19 control-Ig- and n =16 anti-RhoB-Ig-treated mice. (B) Titers of anti-GPI Ig in the serum were also measured at 6 weeks of age by ELISA. Data show means±s.e.m. for n =8 mice of each treatment group. (C,E) Joint inflammation was measured by the change in ankle thickness. Data show means±s.e.m. for (C) n =12 control Ig (▲) and n =14 anti-RhoB Ig (◊), and (E) n =9 control Ig (▲) and n =10 anti-RhoB Ig (◊)-treated mice. (D) Metatarsal joint harvested at 6 weeks of age stained with H&E. Representative sections from a total of n =5 mice for each treatment group are shown. Scale bar: 100 µm. (F) K/BxN mice were treated with 500 µg anti-RhoB Ig at 21 days of age. Three weeks later, the LNs draining the arthritic joints were harvested, and the isolated cells were stimulated with PMA+ionomycin overnight. Inflammatory cytokines were measured in culture supernatants using the cytometric bead array and flow cytometry. Each symbol depicts an individual mouse (control Ig, n =18; anti-RhoB Ig, n =15) with the mean indicated by a solid bar. *** P <0.001. ** P <0.01.

Article Snippet: Recombinant proteins RhoB (OPPA00123), RhoA (OPPA00043), Rac1 (OPPA01858) and Cdc42 (OPPA00141) were purchased from Aviva Systems Biology (San Diego, CA, USA).

Techniques: Enzyme-linked Immunospot, Control, Enzyme-linked Immunosorbent Assay, Staining, Isolation, Flow Cytometry

Journal: Disease Models & Mechanisms

Article Title: RhoB blockade selectively inhibits autoantibody production in autoimmune models of rheumatoid arthritis and lupus

doi: 10.1242/dmm.029835

Figure Lengend Snippet: Arthritis, but not autoantibody level, is attenuated in RhoB ko mice. The RhoB ko allele was crossed into the KRN model on the C57BL/6 background. (A) Joint inflammation was determined by measuring ankle thickness. Data show means±s.e.m. for n =25 KRN.g7 and n =14 RhoB ko KRN.g7 mice. (B) Metatarsal joint harvested at 6 weeks of age stained with H&E. Representative sections from a total of n =5 mice for each treatment group are shown. Scale bar: 100 µm. At 6 weeks of age, (C) the number of anti-GPI ASCs was measured by ELISpot; data show means±s.e.m. for n =15 KRN.g7 and n =17 RhoB ko KRN.g7 mice; n.s., not significant; and (D) titers of anti-GPI Ig in the serum were measured by ELISA. Data show means±s.e.m. for n =23 KRN.g7 and n =11 RhoB ko KRN.g7 mice. (E) At 6 weeks of age, LNs draining the arthritic joints were harvested and the isolated cells were stimulated with PMA+ionomycin overnight. Inflammatory cytokines were measured in culture supernatants using the cytometric bead array and flow cytometry. Each symbol depicts an individual mouse with the mean indicated by a solid bar (KRN B6.g7, n =25; RhoB ko KRN B6.g7, n =12).

Article Snippet: Recombinant proteins RhoB (OPPA00123), RhoA (OPPA00043), Rac1 (OPPA01858) and Cdc42 (OPPA00141) were purchased from Aviva Systems Biology (San Diego, CA, USA).

Techniques: Staining, Enzyme-linked Immunospot, Enzyme-linked Immunosorbent Assay, Isolation, Flow Cytometry

Journal: Disease Models & Mechanisms

Article Title: RhoB blockade selectively inhibits autoantibody production in autoimmune models of rheumatoid arthritis and lupus

doi: 10.1242/dmm.029835

Figure Lengend Snippet: Anti-RhoB Ig has no effect in RhoB ko arthritic mice. RhoB ko KRN.g7 mice were treated with 500 µg anti-RhoB Ig at 21 days of age. Data show means±s.e.m. for n =5 control-Ig- and n =5 anti-RhoB-Ig-treated mice. n.s., not significant. (A) Joint inflammation was measured by the change in ankle thickness. At 6 weeks of age, (B) titers of anti-GPI Ig in the serum were measured by ELISA and (C) the number of anti-GPI ASCs was measured by ELISpot.

Article Snippet: Recombinant proteins RhoB (OPPA00123), RhoA (OPPA00043), Rac1 (OPPA01858) and Cdc42 (OPPA00141) were purchased from Aviva Systems Biology (San Diego, CA, USA).

Techniques: Control, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot

Journal: Disease Models & Mechanisms

Article Title: RhoB blockade selectively inhibits autoantibody production in autoimmune models of rheumatoid arthritis and lupus

doi: 10.1242/dmm.029835

Figure Lengend Snippet: Serum from RhoB ko KRN.g7 mice does not induce arthritis in a serum-transfer model. (A) Pooled serum from K/BxN mice was injected into wt or RhoB ko C57BL/6 mice. Joint inflammation was measured by the change in ankle thickness. Data show means±s.e.m. from a representative experiment of two, with n =5 mice for each group. (B) Pooled serum from 6-week-old KRN.g7 or RhoB ko KRN.g7 mice was equalized for anti-GPI titer and injected into C57BL/6 recipient mice. Data show mean ankle thickness±s.e.m. from n =8 mice for each group. * P <0.05.

Article Snippet: Recombinant proteins RhoB (OPPA00123), RhoA (OPPA00043), Rac1 (OPPA01858) and Cdc42 (OPPA00141) were purchased from Aviva Systems Biology (San Diego, CA, USA).

Techniques: Injection

Journal: Disease Models & Mechanisms

Article Title: RhoB blockade selectively inhibits autoantibody production in autoimmune models of rheumatoid arthritis and lupus

doi: 10.1242/dmm.029835

Figure Lengend Snippet: Targeting RhoB does not affect immune-cell repertoire or function in either RhoB ko or anti-RhoB-Ig-treated C57BL/6 mice. (A) The frequency of individual lymphoid populations in wt or RhoB ko C57BL/6 mice was measured by flow cytometry. Data show means±s.e.m., n =6 mice of each genotype. Gating strategies are shown in Fig. S6 B . (B) The amount of serum Ig in wt or RhoB ko C57BL/6 mice was measured by ELISA. n =6 mice of each genotype. (C) wt and RhoB ko C57BL/6 mice or (D) control-Ig- and anti-RhoB-Ig-treated C57BL/6 mice were immunized with 100 µg NP-KLH. Serum was harvested 10 days later and serum anti-NP titers were measured by ELISA. Data show means±s.e.m. for n =5 mice of each group. All analyses were performed three times.

Article Snippet: Recombinant proteins RhoB (OPPA00123), RhoA (OPPA00043), Rac1 (OPPA01858) and Cdc42 (OPPA00141) were purchased from Aviva Systems Biology (San Diego, CA, USA).

Techniques: Flow Cytometry, Enzyme-linked Immunosorbent Assay, Control

Journal: Disease Models & Mechanisms

Article Title: RhoB blockade selectively inhibits autoantibody production in autoimmune models of rheumatoid arthritis and lupus

doi: 10.1242/dmm.029835

Figure Lengend Snippet: Anti-RhoB Ig lowers autoantibody levels in the MRL/ lpr mouse model of SLE. Weekly doses (500 µg) of anti-RhoB IgG or control mouse IgG were administered to MRL/ lpr mice starting at 4 weeks of age. At 16 weeks of age, serum dsDNA-autoantibody titers were determined by ELISA. Data show means±s.e.m. from two separate experiments combined for n =14 mice per group. Comparisons were made between control and anti-RhoB IgG groups at the specified weeks of age. * P <0.05.

Article Snippet: Recombinant proteins RhoB (OPPA00123), RhoA (OPPA00043), Rac1 (OPPA01858) and Cdc42 (OPPA00141) were purchased from Aviva Systems Biology (San Diego, CA, USA).

Techniques: Control, Enzyme-linked Immunosorbent Assay