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Keywords:

  • Lymphocyte;
  • granulosa;
  • luteinizing hormone;
  • porcine

ABSTRACT: Lymphocyte conditioned medium (CM) was prepared by incubating 0 (cell-free control) or 4 × 106 lymphocytes/ml in serum-supplemented RPMI containing 0, 10−9, 10−7, and 10−5 M luteinizing hormone releasing hormone (LHRH), 10−5 M LHRH antagonist (LHRHA), or 10−7 M LHRH + 10−5 M LHRHA. Treatments were applied with and without 10 μg/ml concanavalin A (con A), and media were analyzed for LH. Aliquots of the CM from cultures incubated for 48 h were later applied to porcine granulosa cell cultures (suspended to 1.25 × 105 cells in 450 μl). Thereafter, 50 μl of CM were added to granulosa cell cultures. Media were collected after 12, 24, and 48 h and progesterone determined. Immunoreactive LH increased with time of incubation in lymphocyte CM but not cell-free CM. LH content of lymphocyte CM increased as LHRH concentration increased. LHRHA significantly reduced the amount of LH measured. The presence of con A in the medium resulted in maximal concentrations of LH, irrespective of dose of LHRH or LHRHA. Cell-free CM containing LHRH, LHRHA, and/or con A did not affect progesterone production by granulosa cells at any of the time periods. Lymphocyte CM containing LHRH caused a dose-dependent increase in progesterone production at 48 h. This stimulation was blocked by lymphocyte CM containing LHRHA. Lymphocyte CM containing con A also stimulated progesterone production at all of the LHRH concentrations studied. This response was not inhibited by lymphocyte CM containing the LHRHA. Therefore, LHRH and con A stimulate porcine lymphocytes to produce a factor that resembles LH and that stimulates progesterone production by granulosa cells in vitro. The stimulation due to con A appears to be nonspecific while that of LHRH is a specific and dose-dependent response.