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482 Unique suppression of human blood mononuclear leukocyte (ML) production of IgG by prolonged exposure to vasoactive intestinal peptide (VIP)
260
481
J. ALLERGY
Abstracts
VASOACTIVE INTESTINAL PEPTIDE INHIBITS PHORBOL ESTER-ENHANCEDI@l SYNTHESIS IN A HUHAN B CELL LINE. 1 h n Alain Robichon. and Edward J. Coetzl. San Francisco. California. Vasoactive intestinal peptide (VIP) is a 28-amino acid neuromediator, which also exerts potent effects on smooth muscle, blood vessels, epithelial cells, glands and leukocytes. VIP influences immunity by altering the migration, proliferation, and synthetic functions of mixed populations of lymphocytes. To increase our understanding of the regulation of individual components of immunity b VIP, we examined the capacity of lo-“M to 10‘ zH VIP to alter IgM secretion by the SKW 6.4 cell line, which contains the message for VIP receptors by Northern blot analysis and expresses highaffinity membrane receptors for [12?] VIP. The production of IgM by cultured SKW 6 4 cells, quantified by an ELISA assay, was enhanced by phorbol myristate acetate (PMA), which activates protein kinase C. The exposure SKW 6.4 cells to VIP for 7 of PMA-stimulated days inhibited the secretion of IgM in a concentration-dependent manner with maximum mean (&SD) inhibition of 4022% at 10-l”M VIP, whereas unstimulated IgM secretion was unaffected. Thus VIP is a potent inhibitor of immunoglobulin production with apparent specificity for stimulated B cells.
k&w&& San Franc&o, CA. Distinct subsets of mammalian blood and tissue B and T lymphocytes express high-affinity receptors for VIP. Mixed lymphocytes in vitro respond to acute exposures to VIP with reduced secretion of IgA and IgM. but rot IgG. To develop a model for studying the effects of chronic exposure of VIP on lymphocytes, replicate aliquots of 1x10” normal ML, depleted of monocytes by adherence, were cultured in complete RPM1 with suboptimally stimutating concentrations (O.~%W/V) of PWM andlo-8~ to 10-‘2# VIP which was added twice daily for 18 days. Production of IgG and IgM in culture supsrnatants was determined by standard ELlSA at days 59.14 and 18. A dose dependent inhibition of IgG (range-41 -77%) and IgM (range=32-70%) synthesis was observed. Maximum inhibition of Ig production was observed at lvgM, when compared to control cultures lacking VIP. No inhibition of Ig synthesis was seen at the highest (10m8M) and bwest (lo-‘*M) concentrations. The effect of VIP on both IgG and IgM production was maximal on day 9, persisted through day 14, and was no foager evident on day 18. VIP concentiatiins were quantiffed by HPLC and RJA on days 5 and 9 and were found to be 4% and 30% of expected respectively, suggesting that the in vitro effects were not due to accumulation of the intact peptide. Therefore chronic exposure of lymphocytes to VIP suppresses IgG and IgM secretion, in contrast to the lack of effect of brief treatment with VIP on IgG secretion. The time-course and the concentratior’l of VIP thus are both critical determinants of the immunoregulatory effects.
CLIN. iMMUi%Oi.. JANUARY 1991
463
VIRAL PEPTIRE MIMICS VIP-INDUCED SUPPRESSION 0)i___SPECIFIC LYMPHOPROLIFERATSON.Soledad Rarkor,BSc and Oscar L. Frick,MD. San Francisco, C!. A peptide has been synthesized that .is idcn:i<~al in part of the parainfluenza-3 hemagg!nt ininii’TXiA) and closely resembles one-third of human VIP pcj!ypeptide. We showed last year that b<>th ViP and PI3HA inhibited PHA-induced lymphoprsl iferstioti. This has been extended to include spccif ic Fmmuirr response to tetanus toxoid. Human voir;otcers wrre given tetanus toxoid bcosters and bivod vas l.aXrn at subsequent interval.s.I,ymphoprolif~‘r~l .ion asfiays were done with 3H-thymidine in 7 day r~ult.ures with tetanus toxoid 1 :ltl to !:30. ?drall.*l -~.!it.ur~~s contained either human VIP 10-8 tr.t :U-i’?l andi:,rPI3HA-like peptide at IO-I, IO-3,c.r !?‘ i CJ:?. .?1? cultures had tetanus toxoid stim ind:::i !:;l)of i-17. In presence of VIP,8/11 expts showcf! il!hibitii;r, up to 50% of lymphoprolifcraiio~, &if.:i!’ h:i: slight (
484
iN VIVO EFFECT OF PREDNISONE ON SPECIFIC .ANTIBODY RESPONSE (SAR) AND SERI!M Igr, SUBCLASSES. John Paul Cella. 54.Qlz_nd Sudhir r;uDtn. M.D., Ph.D. Irvine ?.A Treatment sit,h cnrticosteroids can be associa‘ed Iwith increased froquencv of bactoriai infecTions. promptjnp lhe investigation ?f in vivo vPfec1. r:t’ prednisone on S.4R and serum 1gG in patients with bronchial asthma. S.&R naainst tetanus toxoid (TT) and diphtheria iD) were measured pre- snd post-vaccination by ELISA in 6 normal controls (NC) 2nd 6 bronchial asthmatics rtn prednisone (P). Serum IgG subclasses were measurea pre-vsccina.rion b\. radial l,mmunodiffusion. P mean increase of 653tT;)“b was not :;ignificnnrly differeric r‘r3m NC mean 12crease of 157%278%. :\ subgroup of P on long term steroid >I0 years (n=2) showed :a mean Increase of TT antibody (Ab) 1648%+148% which was significantly different (~<0.001) Prom the remaining J patients with short duration of steroid ([C? years1 mean increase l&5%+44%3 and from NC. There was no significant difference between NC and short duration steroid patients. There was no significant difference in mean D Ab increase between NC and P (665+125894 vs 70?%943%) and no subgroup of hiah responders was identified. Total serum JgG was low in 3 patients: nil hnd &Cl subclass deficiency xnd rune had low IgC4. i)ne aUditiona patient had selective igG3 deficiencv with normal t3tciI IgG. !gG did not correlate rvrth the durat.ion of steroid treatment. The above data show that corticosteroid treatment in vivo is associated with a decrease in IgG levels predominantly of’ the lgGl subclass. However. no inhibitory effect was observed on SAR to protein antigens.
481
J. ALLERGY
Abstracts
VASOACTIVE INTESTINAL PEPTIDE INHIBITS PHORBOL ESTER-ENHANCEDI@l SYNTHESIS IN A HUHAN B CELL LINE. 1 h n Alain Robichon. and Edward J. Coetzl. San Francisco. California. Vasoactive intestinal peptide (VIP) is a 28-amino acid neuromediator, which also exerts potent effects on smooth muscle, blood vessels, epithelial cells, glands and leukocytes. VIP influences immunity by altering the migration, proliferation, and synthetic functions of mixed populations of lymphocytes. To increase our understanding of the regulation of individual components of immunity b VIP, we examined the capacity of lo-“M to 10‘ zH VIP to alter IgM secretion by the SKW 6.4 cell line, which contains the message for VIP receptors by Northern blot analysis and expresses highaffinity membrane receptors for [12?] VIP. The production of IgM by cultured SKW 6 4 cells, quantified by an ELISA assay, was enhanced by phorbol myristate acetate (PMA), which activates protein kinase C. The exposure SKW 6.4 cells to VIP for 7 of PMA-stimulated days inhibited the secretion of IgM in a concentration-dependent manner with maximum mean (&SD) inhibition of 4022% at 10-l”M VIP, whereas unstimulated IgM secretion was unaffected. Thus VIP is a potent inhibitor of immunoglobulin production with apparent specificity for stimulated B cells.
k&w&& San Franc&o, CA. Distinct subsets of mammalian blood and tissue B and T lymphocytes express high-affinity receptors for VIP. Mixed lymphocytes in vitro respond to acute exposures to VIP with reduced secretion of IgA and IgM. but rot IgG. To develop a model for studying the effects of chronic exposure of VIP on lymphocytes, replicate aliquots of 1x10” normal ML, depleted of monocytes by adherence, were cultured in complete RPM1 with suboptimally stimutating concentrations (O.~%W/V) of PWM andlo-8~ to 10-‘2# VIP which was added twice daily for 18 days. Production of IgG and IgM in culture supsrnatants was determined by standard ELlSA at days 59.14 and 18. A dose dependent inhibition of IgG (range-41 -77%) and IgM (range=32-70%) synthesis was observed. Maximum inhibition of Ig production was observed at lvgM, when compared to control cultures lacking VIP. No inhibition of Ig synthesis was seen at the highest (10m8M) and bwest (lo-‘*M) concentrations. The effect of VIP on both IgG and IgM production was maximal on day 9, persisted through day 14, and was no foager evident on day 18. VIP concentiatiins were quantiffed by HPLC and RJA on days 5 and 9 and were found to be 4% and 30% of expected respectively, suggesting that the in vitro effects were not due to accumulation of the intact peptide. Therefore chronic exposure of lymphocytes to VIP suppresses IgG and IgM secretion, in contrast to the lack of effect of brief treatment with VIP on IgG secretion. The time-course and the concentratior’l of VIP thus are both critical determinants of the immunoregulatory effects.
CLIN. iMMUi%Oi.. JANUARY 1991
463
VIRAL PEPTIRE MIMICS VIP-INDUCED SUPPRESSION 0)i___SPECIFIC LYMPHOPROLIFERATSON.Soledad Rarkor,BSc and Oscar L. Frick,MD. San Francisco, C!. A peptide has been synthesized that .is idcn:i<~al in part of the parainfluenza-3 hemagg!nt ininii’TXiA) and closely resembles one-third of human VIP pcj!ypeptide. We showed last year that b<>th ViP and PI3HA inhibited PHA-induced lymphoprsl iferstioti. This has been extended to include spccif ic Fmmuirr response to tetanus toxoid. Human voir;otcers wrre given tetanus toxoid bcosters and bivod vas l.aXrn at subsequent interval.s.I,ymphoprolif~‘r~l .ion asfiays were done with 3H-thymidine in 7 day r~ult.ures with tetanus toxoid 1 :ltl to !:30. ?drall.*l -~.!it.ur~~s contained either human VIP 10-8 tr.t :U-i’?l andi:,rPI3HA-like peptide at IO-I, IO-3,c.r !?‘ i CJ:?. .?1? cultures had tetanus toxoid stim ind:::i !:;l)of i-17. In presence of VIP,8/11 expts showcf! il!hibitii;r, up to 50% of lymphoprolifcraiio~, &if.:i!’ h:i: slight (
484
iN VIVO EFFECT OF PREDNISONE ON SPECIFIC .ANTIBODY RESPONSE (SAR) AND SERI!M Igr, SUBCLASSES. John Paul Cella. 54.Qlz_nd Sudhir r;uDtn. M.D., Ph.D. Irvine ?.A Treatment sit,h cnrticosteroids can be associa‘ed Iwith increased froquencv of bactoriai infecTions. promptjnp lhe investigation ?f in vivo vPfec1. r:t’ prednisone on S.4R and serum 1gG in patients with bronchial asthma. S.&R naainst tetanus toxoid (TT) and diphtheria iD) were measured pre- snd post-vaccination by ELISA in 6 normal controls (NC) 2nd 6 bronchial asthmatics rtn prednisone (P). Serum IgG subclasses were measurea pre-vsccina.rion b\. radial l,mmunodiffusion. P mean increase of 653tT;)“b was not :;ignificnnrly differeric r‘r3m NC mean 12crease of 157%278%. :\ subgroup of P on long term steroid >I0 years (n=2) showed :a mean Increase of TT antibody (Ab) 1648%+148% which was significantly different (~<0.001) Prom the remaining J patients with short duration of steroid ([C? years1 mean increase l&5%+44%3 and from NC. There was no significant difference between NC and short duration steroid patients. There was no significant difference in mean D Ab increase between NC and P (665+125894 vs 70?%943%) and no subgroup of hiah responders was identified. Total serum JgG was low in 3 patients: nil hnd &Cl subclass deficiency xnd rune had low IgC4. i)ne aUditiona patient had selective igG3 deficiencv with normal t3tciI IgG. !gG did not correlate rvrth the durat.ion of steroid treatment. The above data show that corticosteroid treatment in vivo is associated with a decrease in IgG levels predominantly of’ the lgGl subclass. However. no inhibitory effect was observed on SAR to protein antigens.