Rheumatoid arthritis (RA) as a model of IL-1β-induced cachexia
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THIRD INTERNATIONAL WORKSHOP ON CYTOKINES / 515 391 394 RHEUMATOID ARTHRITIS @A) AS A MODEL OF IL-l&INDUCED CACHEXIA. R. Roubenoff. J. Can o . J. Kehayias. H. Zhuane. C. Dinarello. I. Rosenberg. Tufts Univeiik, Boston, MA 02111 Cachexia, or loss of lean body mass (LBM), is associated in disease with morbidity and mortality and wrth immune suppression, including reduced cytokine production. Starvation studies show that loss of 40% of LBM is fatal. We have shown that cachexia is common in RA (Am J Clin Nurr 1990; 52:1113). Paradoxically, RA is associated with increuwd ILlg production, and IL-l has been linked to anorexia and cachexia in animal studies. We investigated cytokine production in RA patients and age-, sex- race-, and weight-matched controls, examining their body composition using total body potassium (TBK) as a measure of LBM. The RA patients had 16.3% less LBM than matched controls (1.69 g TBK cm-l vs. 2.02 g cm-i, p < 0.005, n = 16). Isolated PBMC’s were cultured with or without LPS or S. epidermidis stimulation. Indomethacin was added to all cultures to minimize medication difference between RA and control subjects. Total IL-15 concentrations are shown (* p<O.O5 vs. control): RPM1 LPS LPS LPS 1 nelml S. Epi 10 “a/ml 100 nelml RA 2.56’ 12.1 14.5 24.1’ 20.6’ Cnrl 1 I 1 The 7-fold higher production of IL-l by unstimulated RA cells is consistent with autoimmune activation of PBMC’s. In contrast? the 2-to-3-fold reduction in IL-1 produced by RA cells with high-dose shmulation (100 ng LPS or S. eoi) is consistent with malnutrition. This studv orovides preliminary evidence that RA is a model of cytok&-a&ociated malnutrition, with excessive IL-15 production by PBMC’s at rest but exhaustion of production with stimulation. SEW4 IL6 A"0 ACUTE PnASE PaaT~lYPIa PATIEWT~ RECEWWG A ROWE-IVIRRW TRIWSPL.4YT~,,W. R.U.Seuertin . J.P.Domrllv , Y.Kortekaas . R.Melchers', L.A.Aarden and Th.de Witte‘. 1. Depwtrrnt of Medical Microbiology, 2. Department of Haemntology, University Hospital Wijmegen, Yijmegm, The Netherlands. 3. Department of Autoimvme Diseases. fentr. Lab. of the Dutch Bloodtransfusion Services, Amterdam, The "etherlands. lnterleukin 6 (IL61 is a plciotropic cytokine with pyrogmic proprties and a major iher of the acute phsoe response as reflected by an increase in ser~ll levels of c-reactive protein (CRPl and a,-entitrypsin (e-AI> end e reduction of pre-albmin (p-IN). 116 and CRP are often elevated in the 8em.m of patients with systemic inflermatory reactions of various origin with either acute or chronic character. Hwewr, the precise roLe of IL6 and acute phase proteins WP) in the dwelopnent and cwrse of these reactions has yet to be elucidated. Ye have studied prospectively the cwrse of IL.5, CRP, a-AT and p-ILg in 45 alloge- neic WI recipients. Serb YBS obtain.4 3x weekly frox edsission (a days pre-BMTl until either discharge or death. Thirty-nine patients developed an @sale of fever ttenperature 2 38.3 for more than 24 hours) 5-12 days fotlowiw BMT, while 17 patients shoved a second fever period betueen day 13-D. Mean ser~n IL6 levels were significantly higher in the first episode of fever (1.53 f 30 versus WI t 32 pglml) there being M difference in man tmperature betueen the two fever periods. The APP showed changes in the serw concentratiww which uere canpatible with the fluctuations in IL6, but their reaction profites were less pronounced. The first fever pried was associated with severe profwnd granulocytepenia (!&SC x.1 x 10 ,,qmxositTs, positive blwdcultures and L> while graft-versus host-reactions occurred mostly during the second period. High levels of IL6 ard CRP were associa- ted with high Imrtelity. From these data it can be concluded that the levels of IL6 may vary depending on the fever-inducing-stiwlus. 392 395 CYTOKINE-MEDIATED ENDOTHELIAL CELL ACTIVATION IN HODGKIN'S DISEASE. L.P.Ruco, O.Pomponi and M.Pittiglio. Univ. "La Sapienza", 00161 Roma. Neoplastic cells of Hodgkin's disease (HD) are powerful cytokine producers. We have investigated cytokine production (IL-l/TNF) in 21 HD lymph nodes using immunocytochemistry and/or in situ hybridization. Furthermore, we have correlated cytokine production with expression of activation antigens (HLA-DR, ICAM-1, ELAM-1, VCAM-11 in vascular endothelium. Our results indicate that neoplastic Hodgkin's cells (HCI and tumour associated reactive macrophages were positive for IL-l/TNF in 18 cases. Moreover, we have noted that two distinct patterns of endothelial cell activation were present. In the histological subtype "mixed cellularity" blood vessels were mostly HLA-DR+/ICAM-lt; in "nodular sclerosis" they were ICAM-lt/ELAM-l+/VCAM-lt. In the latter histological variant, high expression of ELAM-1 on endothelium was associated with a significantly increased number of perivascular neutrophils. Our data suggest that cytokine-mediated endothelial cell activation may contribute to determine the histological aspect of HD lesions. 393 INTSRLBUKIN 1 (IL-11 8TrMnLATES SBCRETIGN OF INTERLBUKIN 6 (II‘-6) ?%ND cHwIoT*mIc *CTMTY BY CYSTIC B1BR0818 cm1 AIRNAY BPITHELIAL CELLS. Uf. s. Schl&lel. D.M. JeffersQn. P.D. L&w. s. S”ta Univ. Geneva, 1211 Geneva, Switzerland and Tufts Univ.. Boston, t.m 02111,USA Local accumulation of neutrophils is pronounced in the inflama- tory response (IR) to infection in the always of CF-patients. we studied the hypothesis that CF epithelial cells contribute to the IR bv secretion of mediators with effects on T- end B-cells IIL-61 or local effects on the accumulation of neutrophils (chemotactic activity). The CF cell line JME/CP 15 (homozygous for deletion Phe 508; Am J Physiol 259: L 496-505, 1990)was cultured as described. Monolayers were stimulated with recombinant IL-1 or Lipopoly- saccharide (LPS). Conditioned medium (CM) was tested for IL-5 bio- activitv (B9 as&w) and chemotactic activitv for human neutroohils (modifi;dBoyden &amber). Unstimulated JMEiCF 15 cells secrete IL- 6 in (mean 8580 U/ml). Stimulation with "IL-1 results in a dose dependent increase of IL-6 secretion (34033 U/ml 18 hours after IL-l). A similar response is observed with LPS. CM from un- stimulated cells contains chemotactic activity, which is detectable even after 200.fold dilution (122% of controll.After z-IL-1 the chemotactic activity of CM increased moderately Ci29% of control at 200.fold dilution). Our data demonstrate that CF airway epithelial cells contribute to the local IR by secretion of IL-6 and chemo- tactic activity. Inflammatory mediators such as IL-1 and LPS, which are likely to be present in the airway environment during infection, upregulate the secretion of IL-6 markedly, whereas the effect of IL-1 on the level of chemotactic activity is modest. Studies of the regulation of cytoklne secretion by airway epithelial cells of CF patients and controls as well as the characterization of the factor(s) responsible for the observed chemotactic activity are needed to improve our understanding of the immune pathogenesis of CF aiway disease. DEVELOPMENT AND FUNCTION OF T CELLS IN MICE RENDERED IL-Z DEFICIENT BY TARGETED DISRUPTION OF THE IL-2 LOCUS. A.SchimD1. Institute of Virology and Immunobiology, Univer- sity of Wiirzburg, Wiirzburg, D-8700 Germany. 11-2 minus mice with a null mutation in the 11-2 locus were established by targeted recombination in pluripotent mouse embryonic stem cells. In mitogen stimulated T cells of the mutants neither 11-2 mRNA nor 11-2 protein can be detected. Analysis at four days and four weeks of age showed a normal distribution of CD4+CD8+ CD4-CD8+ and CD4+CD8- cells in the thymus. The distribution of peripheral T cells was also normal. However, proliferation of mitogen stimulated thymus, spleen and lymph node cells was usually greatly reduced. It could be fully restored by addition of 11-2. The residual responses suggest some use of alternate growth factor pathways. The CD8 compartment seems to be more severely compromised as suggested by the fact that after long term culture <l% CD8+ cells could be detected. Immunoglobulin isotype distribution in sera of nonimmunized 11-2 minus mutants was drastically changed. They showed lo-100 fold increases in IgGl, IgG2b and IgE, with IgG3 being lower than in the sera of controls, suggesting a disbalance between THl and TH2 cells. 396 AUTOCRINE REGULATION OF ICAM- EXPRESSION ON HUMAN MELA- NOCYTES By CYTOKIEES (IL-$, IL-7, TNFl AND "V LIGBT ~.;,r,",";~=~:~~~,,p",~~~~~~: ,&E;;:;$tt, A.KBck+, @Urbanski *LBI-DVS, Lab.Cellbiol., Dept.DermII, Univ:Vienna and Hospital Lains, Austria; Dept.Derm., U"lv.Miinster, FRG The expression of intercellular adhesion molecule (ICAE) 1 in melanoma was found to correlate with increased risk of metastasis. Therefore the regulation of ICAM- expression on human melanocytes and melanoma cells was investigated. Foreskin derived melanoc tes G361) were incubated wit x and melanoma cell lines (A375, different cytokines and ICAM- expression was evaluated by FACS analysis. IFN-,, IL-6, IL-7, TNFa and TNFR significantly upregulated ICAM- expression in a dose dependent manner. Most interestingly, IL-6 which does not influence adhesion molecule expression on other cells upregulated melanocyte and melanoma cell ICAM- expression. This effect was dose dependent and could be blocked by an IL-6 antibody. Irradiation with UVB light did not influence constitutive ICAB- expression on melanoma cells and melano- cytee but suppressed cytokine induced ICAM- expression when cells were harvested 16 hrs after irradiation. These findings were further confirmed by Northern blot analysis, showing a marked accumulation of ICAM- mRNA after cytokine treatment which was reduced by irradiation with UVB-light. However, when WB-exposed melanoma cells were cultured for at least 48 hrs induction of ICAB- expression was observed. This effect could be partially inhibited by a TNFol antibody suggesting a" autocrine regulatory pathway. These data indicate that similar to other cells ICAM- expression on melanoma cells and melanocytes is regulated by cytokines and that UVB light affects ICAM- expression on melanocytic cells in s biphasic manner. I" addition, the present study demonstrates for the first time that IL-6 can influence adhesion molecule expression.
Transcript of Rheumatoid arthritis (RA) as a model of IL-1β-induced cachexia
THIRD INTERNATIONAL WORKSHOP ON CYTOKINES / 515
391 394
RHEUMATOID ARTHRITIS @A) AS A MODEL OF IL-l&INDUCED CACHEXIA. R. Roubenoff. J. Can o . J. Kehayias. H. Zhuane. C. Dinarello. I. Rosenberg. Tufts Univeiik, Boston, MA 02111
Cachexia, or loss of lean body mass (LBM), is associated in disease with morbidity and mortality and wrth immune suppression, including reduced cytokine production. Starvation studies show that loss of 40% of LBM is fatal. We have shown that cachexia is common in RA (Am J Clin Nurr 1990; 52:1113). Paradoxically, RA is associated with increuwd ILlg production, and IL-l has been linked to anorexia and cachexia in animal studies. We investigated cytokine production in RA patients and age-, sex- race-, and weight-matched controls, examining their body composition using total body potassium (TBK) as a measure of LBM. The RA patients had 16.3% less LBM than matched controls (1.69 g TBK cm-l vs. 2.02 g cm-i, p < 0.005, n = 16). Isolated PBMC’s were cultured with or without LPS or S. epidermidis stimulation. Indomethacin was added to all cultures to minimize medication difference between RA and control subjects. Total IL-15 concentrations are shown (* p<O.O5 vs. control):
RPM1 LPS LPS LPS 1 nelml
S. Epi 10 “a/ml 100 nelml
RA 2.56’ 12.1 14.5 24.1’ 20.6’ Cnrl 1 I 1 The 7-fold higher production of IL-l by unstimulated RA cells is consistent with autoimmune activation of PBMC’s. In contrast? the 2-to-3-fold reduction in IL-1 produced by RA cells with high-dose shmulation (100 ng LPS or S. eoi) is consistent with malnutrition. This studv orovides preliminary evidence that RA is a model of cytok&-a&ociated malnutrition, with excessive IL-15 production by PBMC’s at rest but exhaustion of production with stimulation.
SEW4 IL6 A"0 ACUTE PnASE PaaT~lYP Ia PATIEWT~ RECEWWG A ROWE-IVIRRW TRIWSPL.4YT~,,W. R.U.Seuertin . J.P.Domrllv , Y.Kortekaas . R.Melchers', L.A.Aarden and Th.de Witte‘. 1. Depwtrrnt of Medical Microbiology, 2. Department of Haemntology, University Hospital Wijmegen, Yijmegm, The Netherlands. 3. Department of Autoimvme Diseases. fentr. Lab. of the Dutch Bloodtransfusion Services, Amterdam, The "etherlands.
lnterleukin 6 (IL61 is a plciotropic cytokine with pyrogmic proprties and a major iher of the acute phsoe response as reflected by an increase in ser~ll levels of c-reactive protein (CRPl and a,-entitrypsin (e-AI> end e reduction of pre-albmin (p-IN). 116 and CRP are often elevated in the 8em.m of patients with systemic inflermatory reactions of various origin with either acute or chronic character. Hwewr, the precise roLe of IL6 and acute phase proteins WP) in the dwelopnent and cwrse of these reactions has yet to be elucidated.
Ye have studied prospectively the cwrse of IL.5, CRP, a-AT and p-ILg in 45 alloge- neic WI recipients. Serb YBS obtain.4 3x weekly frox edsission (a days pre-BMTl until either discharge or death.
Thirty-nine patients developed an @sale of fever ttenperature 2 38.3 for more than 24 hours) 5-12 days fotlowiw BMT, while 17 patients shoved a second fever period betueen day 13-D. Mean ser~n IL6 levels were significantly higher in the first episode of fever (1.53 f 30 versus WI t 32 pglml) there being M difference in man tmperature betueen the two fever periods.
The APP showed changes in the serw concentratiww which uere canpatible with the fluctuations in IL6, but their reaction profites were less pronounced. The first fever pried was associated with severe profwnd granulocytepenia (!&SC x.1 x 10
,,qmxositTs, positive blwdcultures and L> while graft-versus host-reactions
occurred mostly during the second period. High levels of IL6 ard CRP were associa- ted with high Imrtelity.
From these data it can be concluded that the levels of IL6 may vary depending on the fever-inducing-stiwlus.
392 395
CYTOKINE-MEDIATED ENDOTHELIAL CELL ACTIVATION IN HODGKIN'S DISEASE. L.P.Ruco, O.Pomponi and M.Pittiglio. Univ. "La Sapienza", 00161 Roma.
Neoplastic cells of Hodgkin's disease (HD) are powerful cytokine producers. We have investigated cytokine production (IL-l/TNF) in 21 HD lymph nodes using immunocytochemistry and/or in situ hybridization. Furthermore, we have correlated cytokine production with expression of activation antigens (HLA-DR, ICAM-1, ELAM-1, VCAM-11 in vascular endothelium. Our results indicate that neoplastic Hodgkin's cells (HCI and tumour associated reactive macrophages were positive for IL-l/TNF in 18 cases. Moreover, we have noted that two distinct patterns of endothelial cell activation were present. In the histological subtype "mixed cellularity" blood vessels were mostly HLA-DR+/ICAM-lt; in "nodular sclerosis" they were ICAM-lt/ELAM-l+/VCAM-lt. In the latter histological variant, high expression of ELAM-1 on endothelium was associated with a significantly increased number of perivascular neutrophils. Our data suggest that cytokine-mediated endothelial cell activation may contribute to determine the histological aspect of HD lesions.
393 INTSRLBUKIN 1 (IL-11 8TrMnLATES SBCRETIGN OF INTERLBUKIN 6 (II‘-6) ?%ND cHwIoT*mIc *CTMTY BY CYSTIC B1BR0818 cm1 AIRNAY BPITHELIAL CELLS. Uf. s. Schl&lel. D.M. JeffersQn. P.D. L&w. s. S”ta
Univ. Geneva, 1211 Geneva, Switzerland and Tufts Univ.. Boston, t.m 02111,USA
Local accumulation of neutrophils is pronounced in the inflama- tory response (IR) to infection in the always of CF-patients. we studied the hypothesis that CF epithelial cells contribute to the IR bv secretion of mediators with effects on T- end B-cells IIL-61 or local effects on the accumulation of neutrophils (chemotactic activity). The CF cell line JME/CP 15 (homozygous for deletion Phe 508; Am J Physiol 259: L 496-505, 1990)was cultured as described. Monolayers were stimulated with recombinant IL-1 or Lipopoly- saccharide (LPS). Conditioned medium (CM) was tested for IL-5 bio- activitv (B9 as&w) and chemotactic activitv for human neutroohils (modifi;dBoyden &amber). Unstimulated JMEiCF 15 cells secrete IL- 6 in (mean 8580 U/ml). Stimulation with "IL-1 results in a dose dependent increase of IL-6 secretion (34033 U/ml 18 hours after IL-l). A similar response is observed with LPS. CM from un- stimulated cells contains chemotactic activity, which is detectable even after 200.fold dilution (122% of controll.After z-IL-1 the chemotactic activity of CM increased moderately Ci29% of control at 200.fold dilution). Our data demonstrate that CF airway epithelial cells contribute to the local IR by secretion of IL-6 and chemo- tactic activity. Inflammatory mediators such as IL-1 and LPS, which are likely to be present in the airway environment during infection, upregulate the secretion of IL-6 markedly, whereas the effect of IL-1 on the level of chemotactic activity is modest. Studies of the regulation of cytoklne secretion by airway epithelial cells of CF patients and controls as well as the characterization of the factor(s) responsible for the observed chemotactic activity are needed to improve our understanding of the immune pathogenesis of CF aiway disease.
DEVELOPMENT AND FUNCTION OF T CELLS IN MICE RENDERED IL-Z DEFICIENT BY TARGETED DISRUPTION OF THE IL-2 LOCUS. A.SchimD1.
Institute of Virology and Immunobiology, Univer- sity of Wiirzburg, Wiirzburg, D-8700 Germany.
11-2 minus mice with a null mutation in the 11-2 locus were established by targeted recombination in pluripotent mouse embryonic stem cells. In mitogen stimulated T cells of the mutants neither 11-2 mRNA nor 11-2 protein can be detected. Analysis at four days and four weeks of age showed a normal distribution of CD4+CD8+ CD4-CD8+ and CD4+CD8- cells in the thymus. The distribution of peripheral T cells was also normal. However, proliferation of mitogen stimulated thymus, spleen and lymph node cells was usually greatly reduced. It could be fully restored by addition of 11-2. The residual responses suggest some use of alternate growth factor pathways. The CD8 compartment seems to be more severely compromised as suggested by the fact that after long term culture <l% CD8+ cells could be detected. Immunoglobulin isotype distribution in sera of nonimmunized 11-2 minus mutants was drastically changed. They showed lo-100 fold increases in IgGl, IgG2b and IgE, with IgG3 being lower than in the sera of controls, suggesting a disbalance between THl and TH2 cells.
396 AUTOCRINE REGULATION OF ICAM- EXPRESSION ON HUMAN MELA- NOCYTES By CYTOKIEES (IL-$, IL-7, TNFl AND "V LIGBT ~.;,r,",";~=~:~~~,,p",~~~~~~: ,&E;;:;$tt, A.KBck+, @Urbanski *LBI-DVS, Lab.Cellbiol., Dept.DermII, Univ:Vienna and
Hospital Lains, Austria; Dept.Derm., U"lv.Miinster, FRG
The expression of intercellular adhesion molecule (ICAE) 1 in melanoma was found to correlate with increased risk of metastasis. Therefore the regulation of ICAM- expression on human melanocytes and melanoma cells was investigated. Foreskin derived melanoc tes G361) were incubated wit x
and melanoma cell lines (A375, different cytokines and ICAM-
expression was evaluated by FACS analysis. IFN-,, IL-6, IL-7, TNFa and TNFR significantly upregulated ICAM- expression in a dose dependent manner. Most interestingly, IL-6 which does not influence adhesion molecule expression on other cells upregulated melanocyte and melanoma cell ICAM- expression. This effect was dose dependent and could be blocked by an IL-6 antibody. Irradiation with UVB light did not influence constitutive ICAB- expression on melanoma cells and melano- cytee but suppressed cytokine induced ICAM- expression when cells were harvested 16 hrs after irradiation. These findings were further confirmed by Northern blot analysis, showing a marked accumulation of ICAM- mRNA after cytokine treatment which was reduced by irradiation with UVB-light. However, when WB-exposed melanoma cells were cultured for at least 48 hrs induction of ICAB- expression was observed. This effect could be partially inhibited by a TNFol antibody suggesting a" autocrine regulatory pathway. These data indicate that similar to other cells ICAM- expression on melanoma cells and melanocytes is regulated by cytokines and that UVB light affects ICAM- expression on melanocytic cells in s biphasic manner. I" addition, the present study demonstrates for the first time that IL-6 can influence adhesion molecule expression.
