During T cell-B cell collaboration, plasma cell (PC) differentiation and Ig production are known to require T cell-derived soluble factors. However, the exact nature of the cytokines produced by activated T cells that costimulate PC differentiation is not clear. Previously, we reported that costimulation of purified human B cells with IL-21 and anti-CD40 resulted in efficient PC differentiation. In this study, we addressed whether de novo production of IL-21 was involved in direct T cell-induced B cell activation, proliferation, and PC differentiation. We found that activated human peripheral blood CD4+ T cells expressed mRNA for a number of cytokines, including IL-21, which was confirmed at the protein level. Using a panel of reagents that specifically neutralize cytokine activity, we addressed which cytokines are essential for B cell activation and PC differentiation induced by anti-CD3-activated T cells. Strikingly, neutralization of IL-21 with an IL-21R fusion protein (IL-21R-Fc) significantly inhibited T cell-induced B cell activation, proliferation, PC differentiation, and Ig production. Inhibition of PC differentiation was observed even when the addition of IL-21R-Fc was delayed until after initial B cell activation and expansion had occurred. Importantly, IL-21 was found to be involved in PC differentiation from both naive and memory B cells. Finally, IL-21R-Fc did not inhibit anti-CD3-induced CD4+ T cell activation, but rather directly blocked T cell-induced B cell activation and PC differentiation. These data are the first to document that B cell activation, expansion, and PC differentiation induced by direct interaction of B cells with activated T cells requires IL-21.

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Cutaneous insulin allergy remains a clinical problem despite the use of highly purified human insulins. We used in vitro lymphocyte-transformation studies to examine the reactivity of various insulin formulations in diabetic patients with (n = 4) and without (n = 8) cutaneous allergies. Nonspecific response to concanavalin A demonstrated a greater than 40-fold response in both groups. Control patients did not respond to the addition of commercial insulin preparations (stimulation index [SI] less than 4), whereas allergic patients had an 11-fold response to beef (P less than 0.01), a 10-fold response to pork (P less than 0.01), and a 6-fold response to human (P less than 0.01) insulins. This response was limited to a single insulin manufacturer's preparations and was uniform in all three species tested. Efforts to identify the offending agent revealed no lymphoblast transformation when crystalline insulin was used or when commercial preparations were purified to a single peak by high-performance liquid chromatography (HPLC). Pure crystalline insulin dimers of beef, pork, and human species were tested; control subjects responded with mean SIs of 1.9, 1.9, and 1.8, respectively, whereas allergic patients showed greater reactivity to beef (SI 7.3) and pork (SI 14.8). The lymphoblast-transformation response to crystalline human dimer was dose dependent with mean SIs of 0.9 at low concentration (2.8 ng/ml) and 19.2 at a higher concentration (20.4 ng/ml). The commercial insulin preparations were run on size-exclusion HPLC to determine high-molecular-weight aggregate content. Independent of species, a single manufacturer had products demonstrating aggregate levels 3- to 6-fold higher than those found in other manufacturers' preparations.(ABSTRACT TRUNCATED AT 250 WORDS)