FIGURE 3.

B7-H3 induced on synovial monocytes from RA patients is preferentially the 4Ig isoform. A, genomic structure and alternatively spliced isoforms of the human B7-H3 gene. PCR primers (arrowheads) common to both 2Ig and 4Ig sequences were designed. The amino acid-coding exons are presented as gray boxes. These primers allow for discrimination between the two isoforms using RT-PCR based on the sized of the amplified products. B, RT-PCR analysis of the two distinct B7-H3 gene isoforms in PBMO and SFMO from two patients using the primer sets described in A. 4Ig gives a 1.6-kb PCR product, whereas 2Ig gives a 0.95-kb PCR product. C, goat anti-human B7-H3 polyclonal Ab used in immunoblotting can detect recombinant 4Ig and 2Ig proteins of human B7-H3. D, expression pattern of the two isoforms of B7-H3 in whole cell lysates prepared from freshly purified PBMO of healthy donors and PBMO and SFMO of RA patients. The samples loaded in all lanes were derived from each individual RA patient or healthy control except RA SFMO sample 1, which was pooled from two different RA patients. β-Actin was used as the protein loading control. HC, healthy control.