REGULATION OF IMMUNE RESPONSES IN HUMANS AND NON-HUMAN PRIMATES

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I) To further elucidate B cell isotype differentiation in humans, we investigated the regulation of germline and mature C-alpha mRNA transcript expression in human peripheral blood B cells. Focusing first on regulation of germline (I-alpha-C-alpha) transcripts (GLT), we showed that C-alpha-1 GLT are induced by TGF-beta alone, but such induction is enhanced by S. aureus, Cowan 1 (SAC). In contrast, C-alpha-2 GLT are optimally induced by TGF-beta1 in the absence of a B cell stimulus. Turning next to transcripts, and such induction is IL-10-dependent because it is enhanced by exogenous IL-10 and abrogated by the presence of anti- IL-10. In contrast, C-alpha-2 transcripts are induced by T cell stimuli such as anti-CD4O (presented by CDw32-transfected L cells) in the presence of TGF-beta1/IL-10 or by PWM-activated T cells. In summary, these studies show that C-alpha-1 and C-alpha-2 germline and mature transcripts induction is differentially regulated. In addition, this shows that IL-1O appears to be uniquely supportive of the induction of both C-alpha-1 and C-alpha-2 mature transcripts. II) IL-10 has been shown to play an important role in inflammation. To further investigate this property, we determined if polymorphonuclear cells (PMNs) are similar to other phagocytic cell types in their capacity to produce IL-10. Accordingly, human PMNs were isolated from buffy coat preparations and then cultured in the presence of absence of LPS; resulting IL-10 production was assayed at the mRNA level using RT-PCR and at the protein secretion level using IL-1O-specific ELISA. We found that after 24 h of incubation, LPS-stimulated PMNs produced a substantial amount of IL-1O mRNA and secreted IL-10 protein (600-3000 pg/ml). Such production, however, was not seen in highly purified PMNs unless mononuclear cells were added back; thus, it is likely that another cell population secretes a factor which induces stimulated PMNs to produce IL- 10. Finally, we showed that the above production of IL-10 by PMNs has potential self-regulatory effects: LPS-induced expression of the adhesion molecule Mac-1 is suppressed by IL-10 and enhanced by anti-Il-10. In addition, antibody to IL-10 caused a 3-10-fold enhancement of TNF-alpha, IL-1alpha and IL-10 mRNA and/or protein production. These studies support the view that PMNs are a major source of IL-10 at inflammatory sites and IL-10 is a self-regulator of PMNs.