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Cimetidine abrogates suppressor T cell function in vitro
Immunology Letters, 3 (1981) 33-37
© Elsevier/North-Holland Biomedical Press
C I M E T I D I N E A B R O G A T E S S U P P R E S S O R T C E L L F U N C T I O N IN V I T R O Ronald PALACIOS and Donato ALARCON-SEGOVIA Department o f Immunology and Rheumatology, lnstituto Nacional de la NutriciOn, Mexico 22 D. F., Mexico
(Received 20 December 1980) (Accepted 5 Januari 1981)
1. Summary
3. Materials and methods
Cimetidine increased the [3tt] thymidine incorporation of normal human mononuclear cells in culture both when unstimulated or when under the stimulus of phytohemagglutinin or pokeweed mitogen (PWM). It also increased their supernatant immunoglobulin production under PWM stimulus. These effects were higher when the cells were preincubated with cimetidine than when it was added simultaneously. To determine if this effect of cimetidine reflects an abrogation of suppression we studied concanavalin-A-induced suppressor function of normal mononuclear cells using both [3tl]thymidine incorporation and immunoglobulin synthesis as indicator systems and found that preincubation with cimetidine caused significant decrease in suppressor cell function in both systems.
3.1. Isolation o f cells
2. Introduction
Cimetidine is an antagonist of histamine-2 (H2) receptors that is being widely used in the treatment of acid peptic disease and related disorders [1,2]. H2 receptors, however, are not limited no gastric parietal cells but are also found in other cells [3,4], notably T-lymphocytes [5,6]. Effects of cimetidine on in vitro [7,8] and in vivo [9,10] immune functions have begun to be described, granting it further {herapeutic potential, as well as indicating the possibility of provoking undesirable side-effects of its use. tterein we describe the in vitro effects of cimetidine on the suppressor T-cell function and identify it as responsible for the previously described action of cimetidine on the blastogenic response to mitogens.
Peripheral blood mononuclear cells (MNC) were isolated from heparinized venous blood of 5 young adult volunteers of both sexes by density gradient centrifugation of Ficoll-Hypaque. Cells remaining at the interface were removed, washed 3 times in phosphate-buffered saline, and resuspended in minimum essential medium (MEM). 3.2. L y m p h o c y t e blastogenesis For the study of lymphocyte blastogenesis, we cultured MNC in flat-bottom microtiter plates (Costar Cambridge, Mass.) 1 × l0 s MNC/well in MEM supplemented with 10% fetal calf serum (Grand Island Biologicals, New York), 0.8 mg L-glutamine (Sigma Chemicals, S. Louis, Mo.) and 10/~g/ml of gentamicin (Schering, Kenilworth, N. J.). MNC were cultured with or without cimetidine (Smith Kline and French Laboratories, Mexico City, Mexico) at concentrations of 1.5/ag/ml and 3.0/ag/ml for 24 or 48 h at 37°C, in 5% CO2, 95% air, 100% humidity atmosphere for 4 days. Eighteen hours before ending the culture, we added 0.3/aCi of [3H] thymidine (New England Nuclear, Boston) to each well. After completing the culture period, we harvested the cells with a mash multichannel sample collector, placed them in scintillation liquid mixture and read them in a scintillation counter (Packard). All the experiments were done in triplicate. 3.3. Immunoglobin synthesis
The effect of cimetidine on immunoglobulin synthesis was studied by placing in 13 X 75 mm Falcon tubes 1 X l0 6 MNC in enriched MEM, where PWM 33
(final dilution 1:100) was added either at the same time as cimetidine or to cells that had been preincubated in cimetidine. In each instance cimetidine was added at final concentrations of 1.5/~g/ml or 3.0 #g/ml during 24 or 48 h. We also ran similarly preincubated controls but without cimetidine. The tubes were incubated at 37°C, in a 5% CO2 and 100% humidified atmosphere for 6 days. After this, we centrifuged the tubes and determined the amount of immunoglobulins in the supernatants by placing 0.1 ml'of each supernatant and 0.2 ml ofa 1:5 dilution of rabbit antiserum to human IgG, IgA or IgM (Behringwerke, Marburg Lahn, G.F.R.) in disposable-plastic cuvettes with low background and read them in a Helium-Neonlaser nephelometer (Behringwerke) [11 ].
3.4. Suppressor cell function assays Concanavalin-A- (Con-A) induced suppressor func tion was studied as previously described [ 12,13]. As indicator system we measured the 3 major imnmnoglobulins in the culture supernatants by laser nephelometry [11]. We also studied Con-A-induced suppression using [3H] thymidine incorporation as indicator system. For this, we treated the Con-A-stimulated MNC with mitomycin C. We added to them 1 × 10 6 fresh autologous MNC and PHA (final dilution 1:100) in each well of the microtiter plate and incubated the plates at 37°C, in a 5% CO2 and 100% humidity chamber for 4 days. Eighteen hours before ending the culture we added 0.2 #Ci of [3H] thymidine to each well. We
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I
i
1.5 3.O CJMC,'l Di ,', ~- °,;/ml
±
(b)
0 CIMETIDINE
I
1.5
l
3.0
ug / m l
Fig. 1. Effect o f cimetidine on the blastogenic transformation of MNC under the stimulus of (a) pokeweed mitogen (e), or (b) p h y t o h e m a g g l u t i n i n (e), as well as in the absence o f mitogen (o).
34
harvested the ceils with a mash collector, placed them in a scintillation mixture to count in a scintillation counter. All the experiments were done in triplicate. We calculated suppression with the following formula:
suppression = 1 -
Cimetidine enhanced the production of immunoglobulins G, A and M by MNC in culture upon PWM stimulation (Table 2). This effect of cimetidine was also higher when we incubated the cells with it 24 or
(IS of Con-A- and cimetidine-treated or non-treated cells) × 100 (IS of fresh mitogen-stimulated MNC)
where the indicator system (IS) represents the mean concentration ofimmunoglobulins in the supernatant or the CPM in the [3H] thymidine incorporation studies.
4. Results Cime tidine caused a significant increase (P < 0.0005 ) in the [3H] thymidine incorporation of cells under the mitogenic stimulus of either PHA or PWM. With either mitogen the effect on [3H] thymidine incorporation was higher at a final concentration of cimetidine of 1.5/ag/ml than at 3.0/2g/ml (Fig. 1). Addition of cimetidine to non-mitogen-stimulated cultures also showed a significant increase in [3H] thymidine incorporation (P < 0.005). This blastogenic effect was found to be higher on cells incubated with cimetidine 24 or 48 h before cultivating them with mitogen than when we added both cimetidine and mitogen simultaneously (Table 1). Preincubation with cimetidine also increased the [3HI thymidine incorporation of cells not stimulated with mitogens.
48 h prior to the addition of the mitogen than when we added both cimetidine and PWM simultaneously. Table 3 shows the inhibitory effect of cimetidine on Con-A-induced suppressor function with both indicator systems used. This inhibitory effect was also significantly higher at a 1.5/2g/ml concentration of cimetidine (P < 0.005) and occurred both when we incubated the cells with cimetidine prior to the ConA induction of suppression, as well as when we added cimetidine and Con-A simultaneously.
5. Discussion The effect of cimetidine on [3H] thymidine incorporation described by Gifford et al. [8] and confirmed here, and its effect on immunoglobulin synthesis shown here may both result from the abrogatory effect ofcimetidine on suppressor cell function that we found. This is suggested by the higher effect of preincubation with cimetidine than that obtained when cimetidine was added simultaneously to the cultures.
Table 1 Increase in [3H]thymidine incorporation of MNC after preincubation with cimetidine Mitogen
None PHA PWM
Cimetidine treatment Simultaneous addition a (cpm ± S.E.M.)
24 h preincubation b (cpm ± S.E.M.)
48 h preincubation b (cpm ± S.E.M.)
3364-+ 197 30,010 ± 1059 c 21,690± 262 c
9920± 1768 39,847 ± 1556 36,046± 569
9147± 550 46,090 ± 374 39,901 ± 1130
aCimetidine added at the same time as mitogen bMitogen added after 24 or 48 h preincubation with cimetidine. CDifference from cimetidine-preincubated MNC significant at the P < 0.005 level. 35
Table 2 Effect of cimetidine treatment on immunoglobulin synthesis by peripheral blood MNC Preincubation time a (h)
lg synthesis Cimetidine treatment b
0 24 48
without (ng/106 cells)
1.5 ~zg/ml (ng/106 cells)
3/~g/ml (ng/l 06 cells)
11,999 ± 1713 12,436 t 982 12,875 -~ 1324
15,667 + 1425 21,833+ 602 25,384 + 1248
15,000 ± 1324 19,000+ 1287 21,867 + 1048
apWM added simultaneously, or after 24 and 48 h of preincubation with cimetidine. bAll differences between cimetidine-treated and non-treated MNC were statistically significant at P < 0.005 level. Differences between MNC treated with 1.5 ug or 3/~g/ml of cimetidine are not statistically significant. A l t h o u g h there are suppressor T-cells that have been shown to have H2 receptors we cannot tell w h e t h e r the effect o f cimetidine shown here was due to their action on such cells or their receptors. In the absence o f histamine it c a n n o t be ascribed to decreased release o f histamine-induced suppressor factor by H2 receptor-bearing cells [ 14,16]. The c o n c e n t r a t i o n s o f cimetidine used here are akin to those f o u n d in the serum o f patients w h o receive this m e d i c a t i o n for t r e a t m e n t of peptic ulcer [3,8]. Higher c o n c e n t r a t i o n reduced rather than enhanced the effect, indicating that it is not due to
toxicity. If these effects o f cimetidine on suppressor cell f u n c t i o n in vitro can be shown to take place upon its administration in vivo new indications and contraindications o f its use may arise. A m o n g the indications there m a y be diseases where a u g m e n t e d suppressor f u n c t i o n leads to i m m u n e deficiencies: e. g. c o m m o n variable h y p o g a m m a g l o b u l i n e m i a in adults [ 16], where as a m o n g the contradictions there may be diseases where defective suppressor f u n c t i o n has been docum e n t e d : e.g. systemic lupus e r y t h e m a t o s u s [13,17] and m i x e d connective tissue disease [18]. I n d u c t i o n o f a u t o i m m u n e h e m o l y t i c anemia by cimetidine has
Table 3 Effect of cimetidine on concanavalin-A-induced suppressor function '?; suppression Without cimetidine
With cimetidine
1.5 ~g/ml [3Hlthymidine incorporation (A) b 53.4+2.3 a (B) b 52.3±3.4
3 ug/ml
IgS synthesis
[aH]thymidine incorporation
IgS synthesis
[ 3H] thymidine incorporation
lgS synthesis
63.6+4.5 a 61.8+3.1
15.0±4.3 14.6±4.5
33.5+3.5 25.6+4.5
25.9±1.2 25.4±8.8
33.7~4.4 29.0t4.4
P < 0.0005 N.S.
P < 0.0005 N.S.
P < 0.0005 N.S.
P < 0.0005 N.S.
Significance cimetidine treated vs non-treated A vs B
aMean -+ S.E.M. of five experiments. b(A) Con-A added at the same time as cimetidine. (B) Con-A added to MNC preincubated in cimetidine 24 h. 36
been recorded [19], perhaps by causing inhibition of suppression as methyl-DOPA has been proposed to cause [20]. Abrogation of suppressor function by cimetidine may explain the increased delayed hypersensitivity response shown in normal subjects and in patients with chronic mucocutaneous candidiasis [2l] as well as the increased response to PHA of peripheral blood lymphocytes from peptic ulcer patients receiving cimetidine [10]. It may also explain the rejection of renal allografts following cimetidine treatment [22,23].
Acknowledgements We thank Dr. Marco Aurelio Botey, Smith, Kline and French de M~xico for a gift of cimetidine used in these studies and Ms. Eugenia Fishbein for the quantitation of immunoglobulins in culture supernatants. This work was supported by Grant 1485 from the Programa Nacional de Salud, Consejo Nacional de Ciencia y Tecnolog/a, M~xico.
References [ 1] Henn, R. M., Isenberg, J. I., Maxwell,V. and Sturdevant, R. A. L. (1975) New Engl. J. Med. 293,371-375. [2] Longstreth, G. F., Go, V. L. W. and Malagelade, J.-R. (1976) New Engl. J. Med. 294,801-804. [ 3 ] Grennan, D. M., Rooney, P. J., Gilberston, E. and Carson, Dick, W. (1974) Nature (London) 294,368-370. [4] Feldman, M. and Richardson, C. T. (1978) Adv. Intern. Med. 23, 1 24.
[5] Rocklin, R. E., Greineder, G. and Littraan, B. H. (1978) Cell Immunol. 37,162-173. [6] Gupta, S. and Good, R. A. (1978) Sere. Hematol. 17, 1-29. [7] Daman, L. A. and Rosenberg, E. W. (1977) Lancet 2, 1087. [8] Gifford, R. R. M., Hatfield, S. M. and Schmidtke, J. R. (1980) Transplantation 29,143 148. [9] Avella, J., Binder, H. J., Madsen, J. E. and Askenase, P.W.(1978) Lancet l, 624 626. [10] Robertson, A. J., Peden, N. R., Saunders, J. H. B., Gibbs, J. H., Potts, R. C., Brown, R. A., Wormsley, K. G., Beck, J. S. (1979) Lancet 2,420-421. [11] Alarcdn-Segovia,D. and Fishbein, E. (1980) Rev. Invest. Clin. 32, 295-297. [12] Alarc6n-Segovia,D., Ruiz-Arguelles, A. and Llorente, L. (1979) J. Immunol. 122, 1855-1863. [ 13 ] Ruiz-Arguelles,A., Ala~c6n-Segovia,D., Llorente, L. and del Guidice-Knipping, J. A. (1980) Arthr. Rheum. 23, 1004-1009. [14] Rocklin, R. E., Greineder, D. K., Littman, B. H. and Melman, K. L. (1979) Cell Immunol. 44,405-415. [15] Rocklin, R. E. (1977) J. Immunol. 118, 1734-1738. [16] Waldmann, T. A., Durm, M., Brodes, S., Blackman, M., Blaese, R. M. and Strober, W. (1974) Lancet 2,609-613. [ 17 ] Abdou, N. I., Sadawa, A., Pascual, E., Hebert, J. and Sadeghee, S. (i 976)Clin. Immunol. Immunopathol. 6, 192-199. [ 18] Alarc6n-Segovia,D. and Ru~z-Arguelles,A. (1980) Arthritis Rheum. 23,314-318. [19] Rotoli, B., Formisano, S. and Alfinito, F. (1979) Lancet 2,583. [20] Kirtland, H. H. lII, Mohler, D. N. and Horowitz, D. A. (1980) New Engl. J. Med. 302,825-832. [21 ] Jorizzo, J. L., Sams, W. M., Jegasothy, B. V., Olansky, A. J. (1980) Ann. Intern. Med~ 92, 192-195. [22] Primack, W. A. (1978) Lancet 1,824-825. [23] Zammit, M. and Toledo-Pereyra, L. H. (1979) Transplantation 27, 358-359.
37
© Elsevier/North-Holland Biomedical Press
C I M E T I D I N E A B R O G A T E S S U P P R E S S O R T C E L L F U N C T I O N IN V I T R O Ronald PALACIOS and Donato ALARCON-SEGOVIA Department o f Immunology and Rheumatology, lnstituto Nacional de la NutriciOn, Mexico 22 D. F., Mexico
(Received 20 December 1980) (Accepted 5 Januari 1981)
1. Summary
3. Materials and methods
Cimetidine increased the [3tt] thymidine incorporation of normal human mononuclear cells in culture both when unstimulated or when under the stimulus of phytohemagglutinin or pokeweed mitogen (PWM). It also increased their supernatant immunoglobulin production under PWM stimulus. These effects were higher when the cells were preincubated with cimetidine than when it was added simultaneously. To determine if this effect of cimetidine reflects an abrogation of suppression we studied concanavalin-A-induced suppressor function of normal mononuclear cells using both [3tl]thymidine incorporation and immunoglobulin synthesis as indicator systems and found that preincubation with cimetidine caused significant decrease in suppressor cell function in both systems.
3.1. Isolation o f cells
2. Introduction
Cimetidine is an antagonist of histamine-2 (H2) receptors that is being widely used in the treatment of acid peptic disease and related disorders [1,2]. H2 receptors, however, are not limited no gastric parietal cells but are also found in other cells [3,4], notably T-lymphocytes [5,6]. Effects of cimetidine on in vitro [7,8] and in vivo [9,10] immune functions have begun to be described, granting it further {herapeutic potential, as well as indicating the possibility of provoking undesirable side-effects of its use. tterein we describe the in vitro effects of cimetidine on the suppressor T-cell function and identify it as responsible for the previously described action of cimetidine on the blastogenic response to mitogens.
Peripheral blood mononuclear cells (MNC) were isolated from heparinized venous blood of 5 young adult volunteers of both sexes by density gradient centrifugation of Ficoll-Hypaque. Cells remaining at the interface were removed, washed 3 times in phosphate-buffered saline, and resuspended in minimum essential medium (MEM). 3.2. L y m p h o c y t e blastogenesis For the study of lymphocyte blastogenesis, we cultured MNC in flat-bottom microtiter plates (Costar Cambridge, Mass.) 1 × l0 s MNC/well in MEM supplemented with 10% fetal calf serum (Grand Island Biologicals, New York), 0.8 mg L-glutamine (Sigma Chemicals, S. Louis, Mo.) and 10/~g/ml of gentamicin (Schering, Kenilworth, N. J.). MNC were cultured with or without cimetidine (Smith Kline and French Laboratories, Mexico City, Mexico) at concentrations of 1.5/ag/ml and 3.0/ag/ml for 24 or 48 h at 37°C, in 5% CO2, 95% air, 100% humidity atmosphere for 4 days. Eighteen hours before ending the culture, we added 0.3/aCi of [3H] thymidine (New England Nuclear, Boston) to each well. After completing the culture period, we harvested the cells with a mash multichannel sample collector, placed them in scintillation liquid mixture and read them in a scintillation counter (Packard). All the experiments were done in triplicate. 3.3. Immunoglobin synthesis
The effect of cimetidine on immunoglobulin synthesis was studied by placing in 13 X 75 mm Falcon tubes 1 X l0 6 MNC in enriched MEM, where PWM 33
(final dilution 1:100) was added either at the same time as cimetidine or to cells that had been preincubated in cimetidine. In each instance cimetidine was added at final concentrations of 1.5/~g/ml or 3.0 #g/ml during 24 or 48 h. We also ran similarly preincubated controls but without cimetidine. The tubes were incubated at 37°C, in a 5% CO2 and 100% humidified atmosphere for 6 days. After this, we centrifuged the tubes and determined the amount of immunoglobulins in the supernatants by placing 0.1 ml'of each supernatant and 0.2 ml ofa 1:5 dilution of rabbit antiserum to human IgG, IgA or IgM (Behringwerke, Marburg Lahn, G.F.R.) in disposable-plastic cuvettes with low background and read them in a Helium-Neonlaser nephelometer (Behringwerke) [11 ].
3.4. Suppressor cell function assays Concanavalin-A- (Con-A) induced suppressor func tion was studied as previously described [ 12,13]. As indicator system we measured the 3 major imnmnoglobulins in the culture supernatants by laser nephelometry [11]. We also studied Con-A-induced suppression using [3H] thymidine incorporation as indicator system. For this, we treated the Con-A-stimulated MNC with mitomycin C. We added to them 1 × 10 6 fresh autologous MNC and PHA (final dilution 1:100) in each well of the microtiter plate and incubated the plates at 37°C, in a 5% CO2 and 100% humidity chamber for 4 days. Eighteen hours before ending the culture we added 0.2 #Ci of [3H] thymidine to each well. We
32000-I
T
I
2i 000 I80OO 15OOO
_/~.,~
28000-, 24000]
74J
200OO-i
i2000.
16000t
9000-
12000-
6000.
8 000-
3ooo_
4ooo-
13_
i
(O)
I
i
1.5 3.O CJMC,'l Di ,', ~- °,;/ml
±
(b)
0 CIMETIDINE
I
1.5
l
3.0
ug / m l
Fig. 1. Effect o f cimetidine on the blastogenic transformation of MNC under the stimulus of (a) pokeweed mitogen (e), or (b) p h y t o h e m a g g l u t i n i n (e), as well as in the absence o f mitogen (o).
34
harvested the ceils with a mash collector, placed them in a scintillation mixture to count in a scintillation counter. All the experiments were done in triplicate. We calculated suppression with the following formula:
suppression = 1 -
Cimetidine enhanced the production of immunoglobulins G, A and M by MNC in culture upon PWM stimulation (Table 2). This effect of cimetidine was also higher when we incubated the cells with it 24 or
(IS of Con-A- and cimetidine-treated or non-treated cells) × 100 (IS of fresh mitogen-stimulated MNC)
where the indicator system (IS) represents the mean concentration ofimmunoglobulins in the supernatant or the CPM in the [3H] thymidine incorporation studies.
4. Results Cime tidine caused a significant increase (P < 0.0005 ) in the [3H] thymidine incorporation of cells under the mitogenic stimulus of either PHA or PWM. With either mitogen the effect on [3H] thymidine incorporation was higher at a final concentration of cimetidine of 1.5/ag/ml than at 3.0/2g/ml (Fig. 1). Addition of cimetidine to non-mitogen-stimulated cultures also showed a significant increase in [3H] thymidine incorporation (P < 0.005). This blastogenic effect was found to be higher on cells incubated with cimetidine 24 or 48 h before cultivating them with mitogen than when we added both cimetidine and mitogen simultaneously (Table 1). Preincubation with cimetidine also increased the [3HI thymidine incorporation of cells not stimulated with mitogens.
48 h prior to the addition of the mitogen than when we added both cimetidine and PWM simultaneously. Table 3 shows the inhibitory effect of cimetidine on Con-A-induced suppressor function with both indicator systems used. This inhibitory effect was also significantly higher at a 1.5/2g/ml concentration of cimetidine (P < 0.005) and occurred both when we incubated the cells with cimetidine prior to the ConA induction of suppression, as well as when we added cimetidine and Con-A simultaneously.
5. Discussion The effect of cimetidine on [3H] thymidine incorporation described by Gifford et al. [8] and confirmed here, and its effect on immunoglobulin synthesis shown here may both result from the abrogatory effect ofcimetidine on suppressor cell function that we found. This is suggested by the higher effect of preincubation with cimetidine than that obtained when cimetidine was added simultaneously to the cultures.
Table 1 Increase in [3H]thymidine incorporation of MNC after preincubation with cimetidine Mitogen
None PHA PWM
Cimetidine treatment Simultaneous addition a (cpm ± S.E.M.)
24 h preincubation b (cpm ± S.E.M.)
48 h preincubation b (cpm ± S.E.M.)
3364-+ 197 30,010 ± 1059 c 21,690± 262 c
9920± 1768 39,847 ± 1556 36,046± 569
9147± 550 46,090 ± 374 39,901 ± 1130
aCimetidine added at the same time as mitogen bMitogen added after 24 or 48 h preincubation with cimetidine. CDifference from cimetidine-preincubated MNC significant at the P < 0.005 level. 35
Table 2 Effect of cimetidine treatment on immunoglobulin synthesis by peripheral blood MNC Preincubation time a (h)
lg synthesis Cimetidine treatment b
0 24 48
without (ng/106 cells)
1.5 ~zg/ml (ng/106 cells)
3/~g/ml (ng/l 06 cells)
11,999 ± 1713 12,436 t 982 12,875 -~ 1324
15,667 + 1425 21,833+ 602 25,384 + 1248
15,000 ± 1324 19,000+ 1287 21,867 + 1048
apWM added simultaneously, or after 24 and 48 h of preincubation with cimetidine. bAll differences between cimetidine-treated and non-treated MNC were statistically significant at P < 0.005 level. Differences between MNC treated with 1.5 ug or 3/~g/ml of cimetidine are not statistically significant. A l t h o u g h there are suppressor T-cells that have been shown to have H2 receptors we cannot tell w h e t h e r the effect o f cimetidine shown here was due to their action on such cells or their receptors. In the absence o f histamine it c a n n o t be ascribed to decreased release o f histamine-induced suppressor factor by H2 receptor-bearing cells [ 14,16]. The c o n c e n t r a t i o n s o f cimetidine used here are akin to those f o u n d in the serum o f patients w h o receive this m e d i c a t i o n for t r e a t m e n t of peptic ulcer [3,8]. Higher c o n c e n t r a t i o n reduced rather than enhanced the effect, indicating that it is not due to
toxicity. If these effects o f cimetidine on suppressor cell f u n c t i o n in vitro can be shown to take place upon its administration in vivo new indications and contraindications o f its use may arise. A m o n g the indications there m a y be diseases where a u g m e n t e d suppressor f u n c t i o n leads to i m m u n e deficiencies: e. g. c o m m o n variable h y p o g a m m a g l o b u l i n e m i a in adults [ 16], where as a m o n g the contradictions there may be diseases where defective suppressor f u n c t i o n has been docum e n t e d : e.g. systemic lupus e r y t h e m a t o s u s [13,17] and m i x e d connective tissue disease [18]. I n d u c t i o n o f a u t o i m m u n e h e m o l y t i c anemia by cimetidine has
Table 3 Effect of cimetidine on concanavalin-A-induced suppressor function '?; suppression Without cimetidine
With cimetidine
1.5 ~g/ml [3Hlthymidine incorporation (A) b 53.4+2.3 a (B) b 52.3±3.4
3 ug/ml
IgS synthesis
[aH]thymidine incorporation
IgS synthesis
[ 3H] thymidine incorporation
lgS synthesis
63.6+4.5 a 61.8+3.1
15.0±4.3 14.6±4.5
33.5+3.5 25.6+4.5
25.9±1.2 25.4±8.8
33.7~4.4 29.0t4.4
P < 0.0005 N.S.
P < 0.0005 N.S.
P < 0.0005 N.S.
P < 0.0005 N.S.
Significance cimetidine treated vs non-treated A vs B
aMean -+ S.E.M. of five experiments. b(A) Con-A added at the same time as cimetidine. (B) Con-A added to MNC preincubated in cimetidine 24 h. 36
been recorded [19], perhaps by causing inhibition of suppression as methyl-DOPA has been proposed to cause [20]. Abrogation of suppressor function by cimetidine may explain the increased delayed hypersensitivity response shown in normal subjects and in patients with chronic mucocutaneous candidiasis [2l] as well as the increased response to PHA of peripheral blood lymphocytes from peptic ulcer patients receiving cimetidine [10]. It may also explain the rejection of renal allografts following cimetidine treatment [22,23].
Acknowledgements We thank Dr. Marco Aurelio Botey, Smith, Kline and French de M~xico for a gift of cimetidine used in these studies and Ms. Eugenia Fishbein for the quantitation of immunoglobulins in culture supernatants. This work was supported by Grant 1485 from the Programa Nacional de Salud, Consejo Nacional de Ciencia y Tecnolog/a, M~xico.
References [ 1] Henn, R. M., Isenberg, J. I., Maxwell,V. and Sturdevant, R. A. L. (1975) New Engl. J. Med. 293,371-375. [2] Longstreth, G. F., Go, V. L. W. and Malagelade, J.-R. (1976) New Engl. J. Med. 294,801-804. [ 3 ] Grennan, D. M., Rooney, P. J., Gilberston, E. and Carson, Dick, W. (1974) Nature (London) 294,368-370. [4] Feldman, M. and Richardson, C. T. (1978) Adv. Intern. Med. 23, 1 24.
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