- Email: [email protected]
Immunomodulatory effects of enkephalin degrading enzymes inhibitors
EFFECT OF DTC AND CALF THYMUSEXTRACTON RESTORATION 'OF RESTRAINT STRESS SUPPRESED HUMORAL RESPONSE IN MICE B. Obmi~ska-Domoradzka and 3. O~bowy ~partment of PharmacoIogy and Toxicoiogy, Faculty of Veterinary Medicine, Agricultural University, Norwida 3i, 50-375 WROCLAW, POLAND.
IMMIR~OMODULATORY EFFECTS OF HNKFJPHALIlq D]~RADINO ENZYMP~ INI~BITORS ~ , S. Peli~, Z. Man~ev, l Radalovid and $. Z. Maiobabid
Department of Pharmacolo~, Faculty of MedicLrte, Univenlty of Nit, Brade Taskovid 81,18000 Nil, Yugodavia Be,statln/8 an inhibitor of anduopepttdue (EC 3. 4. 1L 2) azid (De~
The experiments were carried out on mice (Balb/c 6
weeks old) exposed to restraint stress. Mice were restrained for 12 h per day at nighttime and released at daytime for 2 consecutive days. Animals were immu~ nized i.p. immediately before the stress with 4xlO sheep red ~lood cell (SRBC).The control mice were only immunized with £RBC. Sodium diethyIdithiocarbamate (BTC, 20 mg/kg) was administered i.p.twice e.g. 4 a 2 days prior to SRBC in~ection and restraint stress.Cal~
l
Lhymus extract (TFX, 10 mg/kg) was in3ected i.p. four times at 24 h intervals prior to SRBC immunization and exoosure to stress. The ~ollowing indices were evaluated: ( i ) the number of sptenocytes producing anti-SRBC antibodies (plaque forming cells, PFC) on Says # and 7; (ii) the total (19S+7S) and 2 mercaptoethanol resistant (75) anti-SRBC serum haemagglutinins
on days 4, 7 and 10. It was found that restraint stress reduced humoraI £esponse to SRBC ~hiPh was reflected in the decreased
number of PFC and serum haemagglutinin t i t r e s (19S+7S and 7S). The supressing effect of restraint stress was observed £or lO days. Pretreatment with OTC or TFX resulted in partial restoration of the number o~ PFC and production of the anti-SReC haemagglutinins ~mpaired by restraint stress. The stronger effect restoring ~he humeral response to SRBC was observed ~or DTC.
Tyrl).Met-enkelYlmlln is a n tnln3~it~ of ~ r n ~ i m and ne.tral endopepl/dase 0Be 3. 4. 24. 11). eazyme,=involved in endoBenotm OldCdd peptide degradation. W e investigated effects of intraperRoneally (jp) admlni~terexl inl~'tmttOPtlOnhomoral an celllllar l e e , m e respt~se of ra1~ Bealalln ip ~dm~ni~ered at lower dose (0.2 ms/kg) exerted the ~mm~'alopoteatlatta8 effect on &amoral Jmmltne reRx>a=e mtr~m+l-ed by ~'e¢l: p]aqoe..tormlnS celt amlay(~mC) and direct ~ t t n a l ~ o n tedmique. l~maber of PFC/10~-~SD and annq~ody liter Oog~tSD) were s i s n i ~ a n t l y ]ldsher (p<0.001) then in control atdmala Drug ip aam{-i~ered at hisher dom~ (1 ms/kS) leads to lmmunosuEpr~siou (p<0.001) in both t ~ ]~.,m.atln~ aami-]qteredl at dose of 0.1 a ~ has no effeO~ on Atthus, reactioa and reaclloa ot delayed skin hype,rsen~tivity to bovine serum allmmi. ARea" ip admt-tstration at hieher dose of 2 ~ be~ leads to |mm~no~lppr~@~00 in both AIthllll reaction ( I ~ . 0 5 ) and reattioa of dat=yea ~ h y p e n e ~ U v i t y (p~0.0o:1). (De~-Tyr~).Met~eph,~- tp
ap]~ed at a dose ~0.2 mWkB ~ m e m t l y ine'eamm~nwmber of ~ (p< 0.00:t) and amtibody titer (p¢0.001). Number of ~ and antRxxly titer after (De~Tyrl)-Met-eakep]mllm at a dose of 1 ~ were also d~ifl. ,~aney ki~er tbea in co=ztrol ,.+mals. (Des-Tyr~)-Met-eakephal/n /p a ~ at a dose of 13,1 ms/kS lea& t~ imm~lnostimulalS, Cel (1~0,00I) in Arthus rea~ton and r e a o ~ t of delayed skin hyperse~tivity. ( D e ~ Tyrl)-Met-enkephann at a dose of 2 mll~ leads to imunosuppre~on (p~0.001) only in rea~¢m ¢~ delayed skin h y p e ~ ~_ttviry, Our experiments show that in~bitors o~ enkephalin de~radin8 enzymes exert dllal f m m ~ m o ~ t l l a t o r y effecUt o11 humoral and oellular immune respon~ of rats dependin__8of applied dose.
COMPLEX
MODIFICATION OF LYMPHOCYTES OF PEYER'S PATCHES MESENTERIC LYMPH NODES - SPLEEN BY EGG-WHITE
G. Migliorati, R. Moraca, G. Nocentini, A. Bartoli, S. Ronchetti, G. Testa and C. Riccardi. DMCPP, Section of Pharmacology, Univ. of Perugia, 06100 Perugia, Italy.
LYSOZYME. Pacer S, Sava G, Bergamo A*, Giacomello E, Gagliardi R*, BoccO E^, Zacchigna M^. Department of Biomedical Sciences, (^) Department of Pharmaceutical Sciences, University of Trieste, (*) Foundation Callerio, Institutes of Biological Research, Trieste, Italy.
Glucocorticoid hormones (GCH) are anti-inflammatory and immunosuppressive agents able to inhibit T-cell growth and activation. Since the T-cell receptor (TCR)/CD3 complex mediates T-lymphocyte activation, we studied the effect of in vitro treatment with dexamethasone (DEX), a synthetic GCH, On TCR/CD3 complex expression. DEX-treatment of a murine hybridoma T-cell line (3DO), induced a decrease of the TCR/CD3 membrane expression as evidenced after 4 days of treatment. Following 4 weeks of DEX contact, TCR/CD3 was undetectable in cell membrane. However, the amount of mRNAs coding TCR/CD3 chains, including TCRcz, TCRI3, CD3.y, CD35 and CD38, as well as the amount of CD3s protein, did not change after drug treatment. On the contrary, a decrease of the mRNAs deriving from the T C R ( gene locus (TCRr~, TCRq and TCRe alternative splicings), which are responsible for the membrane expresssion of the TCR/CD3 complex, was induced. These data suggest that the down-modulation of TCR expression might be due to the diminution of mRNAs coded by the T C R ( gene in DEX-treated cells.
The mechanism by which lysozyme, after oral administration, potentiates host's immune responses, either in animal models or in humans, has not been fully elucidated yet. The aim of the present investigation was that of examinig the effects of lysozyme on lymphocytes of the intestine (intraepithelial and of Peyer's patches), of the mesenteric lymph nodes and spleens of mice bearing MCa mammary carcinoma. Lysozyme or PEGlysozyme (lysozyme coupled with PEG - 7Mol PEG per Mol lysozyme) was given per os (admixed with the powdered food) at the dose of 100 mg/kg/day. 24 hr after last day of treatment, mice were sacrificed and lymphocytes were isolated and studied for in vitro response to polyclonal activators and for the expression of CD3, CD4, CD8, CD25 and C D l l b antigens. Lysozyme enzymatic activity is preserved in the gut also at jejunum level. Both lysozymes activate immune cells as shown by the increased response to growth factors and mitogens of in vitro cultures of mixed macrophage-lymphocyte cells and by the significant modifications of the antigenic profiles of lymphocytes harvested from the above tissues. On metastasis formation, PEG-lysozyme was more effective than lysozyme. These data indicate that lysozyme activates systemic immunity by stimulation of gut associated lymphocytes. Acknowledgements: work done with contributions from MURST 40%, CNR Contract 94.01002.CT04 and Cassa di RisparmioFondazione Trieste. A Bergamo is recipient of a fellowship grant from Fondazione Callerio.
DEXAMETHASONE EXPRESSION
INHIBITS
TCR/CD3
--
363 --
IMMIR~OMODULATORY EFFECTS OF HNKFJPHALIlq D]~RADINO ENZYMP~ INI~BITORS ~ , S. Peli~, Z. Man~ev, l Radalovid and $. Z. Maiobabid
Department of Pharmacolo~, Faculty of MedicLrte, Univenlty of Nit, Brade Taskovid 81,18000 Nil, Yugodavia Be,statln/8 an inhibitor of anduopepttdue (EC 3. 4. 1L 2) azid (De~
The experiments were carried out on mice (Balb/c 6
weeks old) exposed to restraint stress. Mice were restrained for 12 h per day at nighttime and released at daytime for 2 consecutive days. Animals were immu~ nized i.p. immediately before the stress with 4xlO sheep red ~lood cell (SRBC).The control mice were only immunized with £RBC. Sodium diethyIdithiocarbamate (BTC, 20 mg/kg) was administered i.p.twice e.g. 4 a 2 days prior to SRBC in~ection and restraint stress.Cal~
l
Lhymus extract (TFX, 10 mg/kg) was in3ected i.p. four times at 24 h intervals prior to SRBC immunization and exoosure to stress. The ~ollowing indices were evaluated: ( i ) the number of sptenocytes producing anti-SRBC antibodies (plaque forming cells, PFC) on Says # and 7; (ii) the total (19S+7S) and 2 mercaptoethanol resistant (75) anti-SRBC serum haemagglutinins
on days 4, 7 and 10. It was found that restraint stress reduced humoraI £esponse to SRBC ~hiPh was reflected in the decreased
number of PFC and serum haemagglutinin t i t r e s (19S+7S and 7S). The supressing effect of restraint stress was observed £or lO days. Pretreatment with OTC or TFX resulted in partial restoration of the number o~ PFC and production of the anti-SReC haemagglutinins ~mpaired by restraint stress. The stronger effect restoring ~he humeral response to SRBC was observed ~or DTC.
Tyrl).Met-enkelYlmlln is a n tnln3~it~ of ~ r n ~ i m and ne.tral endopepl/dase 0Be 3. 4. 24. 11). eazyme,=involved in endoBenotm OldCdd peptide degradation. W e investigated effects of intraperRoneally (jp) admlni~terexl inl~'tmttOPtlOnhomoral an celllllar l e e , m e respt~se of ra1~ Bealalln ip ~dm~ni~ered at lower dose (0.2 ms/kg) exerted the ~mm~'alopoteatlatta8 effect on &amoral Jmmltne reRx>a=e mtr~m+l-ed by ~'e¢l: p]aqoe..tormlnS celt amlay(~mC) and direct ~ t t n a l ~ o n tedmique. l~maber of PFC/10~-~SD and annq~ody liter Oog~tSD) were s i s n i ~ a n t l y ]ldsher (p<0.001) then in control atdmala Drug ip aam{-i~ered at hisher dom~ (1 ms/kS) leads to lmmunosuEpr~siou (p<0.001) in both t ~ ]~.,m.atln~ aami-]qteredl at dose of 0.1 a ~ has no effeO~ on Atthus, reactioa and reaclloa ot delayed skin hype,rsen~tivity to bovine serum allmmi. ARea" ip admt-tstration at hieher dose of 2 ~ be~ leads to |mm~no~lppr~@~00 in both AIthllll reaction ( I ~ . 0 5 ) and reattioa of dat=yea ~ h y p e n e ~ U v i t y (p~0.0o:1). (De~-Tyr~).Met~eph,~- tp
ap]~ed at a dose ~0.2 mWkB ~ m e m t l y ine'eamm~nwmber of ~ (p< 0.00:t) and amtibody titer (p¢0.001). Number of ~ and antRxxly titer after (De~Tyrl)-Met-eakep]mllm at a dose of 1 ~ were also d~ifl. ,~aney ki~er tbea in co=ztrol ,.+mals. (Des-Tyr~)-Met-eakephal/n /p a ~ at a dose of 13,1 ms/kS lea& t~ imm~lnostimulalS, Cel (1~0,00I) in Arthus rea~ton and r e a o ~ t of delayed skin hyperse~tivity. ( D e ~ Tyrl)-Met-enkephann at a dose of 2 mll~ leads to imunosuppre~on (p~0.001) only in rea~¢m ¢~ delayed skin h y p e ~ ~_ttviry, Our experiments show that in~bitors o~ enkephalin de~radin8 enzymes exert dllal f m m ~ m o ~ t l l a t o r y effecUt o11 humoral and oellular immune respon~ of rats dependin__8of applied dose.
COMPLEX
MODIFICATION OF LYMPHOCYTES OF PEYER'S PATCHES MESENTERIC LYMPH NODES - SPLEEN BY EGG-WHITE
G. Migliorati, R. Moraca, G. Nocentini, A. Bartoli, S. Ronchetti, G. Testa and C. Riccardi. DMCPP, Section of Pharmacology, Univ. of Perugia, 06100 Perugia, Italy.
LYSOZYME. Pacer S, Sava G, Bergamo A*, Giacomello E, Gagliardi R*, BoccO E^, Zacchigna M^. Department of Biomedical Sciences, (^) Department of Pharmaceutical Sciences, University of Trieste, (*) Foundation Callerio, Institutes of Biological Research, Trieste, Italy.
Glucocorticoid hormones (GCH) are anti-inflammatory and immunosuppressive agents able to inhibit T-cell growth and activation. Since the T-cell receptor (TCR)/CD3 complex mediates T-lymphocyte activation, we studied the effect of in vitro treatment with dexamethasone (DEX), a synthetic GCH, On TCR/CD3 complex expression. DEX-treatment of a murine hybridoma T-cell line (3DO), induced a decrease of the TCR/CD3 membrane expression as evidenced after 4 days of treatment. Following 4 weeks of DEX contact, TCR/CD3 was undetectable in cell membrane. However, the amount of mRNAs coding TCR/CD3 chains, including TCRcz, TCRI3, CD3.y, CD35 and CD38, as well as the amount of CD3s protein, did not change after drug treatment. On the contrary, a decrease of the mRNAs deriving from the T C R ( gene locus (TCRr~, TCRq and TCRe alternative splicings), which are responsible for the membrane expresssion of the TCR/CD3 complex, was induced. These data suggest that the down-modulation of TCR expression might be due to the diminution of mRNAs coded by the T C R ( gene in DEX-treated cells.
The mechanism by which lysozyme, after oral administration, potentiates host's immune responses, either in animal models or in humans, has not been fully elucidated yet. The aim of the present investigation was that of examinig the effects of lysozyme on lymphocytes of the intestine (intraepithelial and of Peyer's patches), of the mesenteric lymph nodes and spleens of mice bearing MCa mammary carcinoma. Lysozyme or PEGlysozyme (lysozyme coupled with PEG - 7Mol PEG per Mol lysozyme) was given per os (admixed with the powdered food) at the dose of 100 mg/kg/day. 24 hr after last day of treatment, mice were sacrificed and lymphocytes were isolated and studied for in vitro response to polyclonal activators and for the expression of CD3, CD4, CD8, CD25 and C D l l b antigens. Lysozyme enzymatic activity is preserved in the gut also at jejunum level. Both lysozymes activate immune cells as shown by the increased response to growth factors and mitogens of in vitro cultures of mixed macrophage-lymphocyte cells and by the significant modifications of the antigenic profiles of lymphocytes harvested from the above tissues. On metastasis formation, PEG-lysozyme was more effective than lysozyme. These data indicate that lysozyme activates systemic immunity by stimulation of gut associated lymphocytes. Acknowledgements: work done with contributions from MURST 40%, CNR Contract 94.01002.CT04 and Cassa di RisparmioFondazione Trieste. A Bergamo is recipient of a fellowship grant from Fondazione Callerio.
DEXAMETHASONE EXPRESSION
INHIBITS
TCR/CD3
--
363 --