The anti-CD2 monoclonal antibody BTI-322 generates unresponsiveness by activation-associated T cell depletion.

The antihuman CD2 MoAb BTI-322 (Lo-CD2a) effectively inhibits T cell responses in vitro to allogeneic cells, which is followed by unresponsiveness to the original stimulator in secondary stimulation. We studied the xenogeneic human antiporcine mixed lymphocyte reaction (MLR), and utilized anti-T cell receptor (TCR) Vbeta family antibody-induced cell proliferation to determine the specificity and mechanism. BTI-322 and its humanized version, MEDI-507, effectively inhibited the primary xenogeneic MLR. After suboptimal primary stimulation using lower numbers of xenogeneic stimulator cells, the unresponsiveness in secondary culture was apparent only for xenogeneic stimulator cells of the original SLA haplotype, and not for third-party stimulators or allogeneic cells. The inhibition of primary MLR was not observed for nylon-wool-purified T cells, but was seen after reconstitution of purified T cells with monocytes. Similarly, anti-Vbeta family-specific stimulation showed family-specific unresponsiveness in secondary culture. This required the presence of the whole BTI-322 molecule: a F(ab')2 fragment was not effective. T cells of a distinct Vbeta family were depleted after stimulation with an anti-Vbeta family-specific antibody and BTI-322. We conclude that the inhibition by BTI-322 of a primary xenogeneic MLR or the response to an anti-TCR Vbeta antibody is associated with unresponsiveness upon restimulation, due to activation-associated cell depletion. In this process, the interaction between monocytes and the Fc part of the antibody is involved. This unique characteristic of BTI-322 suggests the potential of the antibody for tolerance induction in vivo, besides the potential use as a T cell depleting agent.

Clonal lines of transgenic fibroblast cells derived from the same fetus result in different development when used for nuclear transfer in pigs.

Different factors are believed to influence the outcome of nuclear transfer (NT) experiments. Besides the cell cycle stage of both recipient cytoplast and donor karyoplast, the origin of the donor cells (embryonic, fetal, and adult) is of interest. We compared in vitro development of NT embryos derived from small serum-starved (G0) or small cycling (G1) porcine fetal fibroblast cells. Serum starvation did not have a positive effect on cleavage rate or the percentage of embryos that developed to the morula and blastocyst stages. Next, we investigated the development of porcine NT embryos derived from different transgenic clonal cell lines that had originated from the same fetus. When different clonal lines of fetal fibroblasts were fused to enucleated metaphase II oocytes, differences in fusion rates as well as in development to the morula and blastocyst stages were observed (P < 0.05). When oocytes derived from sow ovaries were used as recipient cytoplasts, significantly better cleavage (P = 0.03) and blastocyst formation (P < 0.014) was obtained when compared with oocytes derived from gilts. Our data indicate that not only different cell lines, but also different clones derived from one primary cell line, result in different development when used for NT. In addition, the use of sow oocytes as a cytoplast source also improves the efficiency of NT experiments.