In vivo and in vitro production of IFN-β and IFN-γ during graft vs host disease

We have shown previously that high levels of IFN-.beta. were generated in vitro from spleen cells obtained from mice experiencing graft vs host disease (GVHD). However, very little or no IFN-.gamma. was found in these cultures even when IL-2 or Con A was added as a stimulant of IFN-.gamma. production. This study was undertaken to determine if the IFNs were similarly produced in vivo during the GVH reaction and to further explore the inability of GVHD spleen cells to produce IFN-.gamma. in vitro. GVHD was induced across minor histocompatibilities by the i.v. injection of B10.D2 spleen cells into sub-lethally irradiated BALB/c mice. Using cytoplasmic immunofluorescence to detect IFN-.beta. and -.gamma., both IFNs were readily detectable in vivo in spleens of mice undergoing GVHD. IFN-.gamma. demonstrated a distinct distribution pattern, localizing in the peri-arteriolar lymphoid regions of the spleen, whereas IFN-.beta. immunofluorescence appeared diffusely in all areas. Expression of both IFN-.beta. and -.gamma. was shown to be dependent on the GVH reaction, inasmuch as syngenetic controls and mice given T cell-depleted donor cells had little immunofluorescence. These results contradict in vitro data in that IFN-.gamma. cannot be found in GVHD spleen cell cultures even in the presence of Con A. This in vitro unresponsiveness appeared to be due to the mixing of different cell populations as a result of preparing splenic single-cell suspensions. Percoll gradient fractionation of GVH spleen cells yielded a cell population which, when stimulated with Con A, produced IFN-.gamma. and underwent cell proliferation. This study represents the first description of the in vivo splenic distributions of IFN-.beta. and -.gamma. during GVHD, and presents data that suggest that in vitro results may not truly reflect in vivo immune responsiveness. Thus, the IFNs may play a critical role in the complex events leading to the VHD syndrome.