Arch. Oto-Rhino-Laryng. 220, 163-165 (1978)
Archivesof Oto-Rhino-Laryngology 9 Springer-Verlag 1978
T Lymphocytes in the Effusions of Secretory Otitis Media* P. Sipil/i ~, P. Ryh/inen 2, and P. Karma 3 Departments of OtolaryngologyI and Anaesthesiology2, University of Oulu, Finland, Institute of Clinical Sciences3, University of Tampere, Teiskontie 35, SF-33520 Tampere 52, Finland
Summary. Smears were made from 39 middle ear effusions in 29 children with secretory otitis media (SOM) and stained with acid oz-naphthyl acetate esterase (ANAE) to demonstrate T lymphocytes. The effusions showed on the average 21% of the lymphocytes to be A N A E positive, while simultaneously drawn peripheral blood revealed the proportion of A N A E positive cells to be 53% (P < 0.001, the Student's t-test). Our finding suggests that T cell dependent defence mechanisms may be locally disturbed in SOM.
Key words: Secretory otitis media - T cells - Acid a-naphthyl acetate esterase.
It is proposed (Tomasi, 1976) that local cell mediated immune reactions, which have been demonstrated to take place in the respiratory tract (Waldman et al., 1972), may play an important role in the protection of the middle ear against infection. However, although there have been numerous immunological studies - which have failed to show any systemic or antibody mediated local immune defect as a cause of secretory otitis media (SOM) - the possible role of local cell mediated immune mechanisms in SOM have not, so far as we know, been studied. We present here the preliminary results of our studies concerning the occurrence and role of T lymphocytes, as marked with acid ce-naphthyl acetate esterase (ANAE) according to the method presented by Mueller et al. (1975), in the middle ear effusions of children with SOM.
Material and Methods Effusion was aspirated from 39 middle ears in 29 children (mean age 5.1 years, range 0.4-11.3 years) who came for tympanostomybecause of SOM. A sample of peripheral bloodwas drawn simultaneously. Immediately after taking the specimens, smears from blood and the secretion were made and the * Supported by the grant from the Emil Aaltonen Foundation, Finland Offprint requests to: Dr. P. Karma (address see above)
0302--9530/78/0220/0163/5 01.00
164
P. Sipil/i et al.
slides were allowed to dry for 30 min. The fixation and staining of the smears for the demonstration of ANAE activity in the lymphocytes were done according to Mueller et al. (1975), with the exception of the smears being counterstalned with 1% aqueous toluidine blue solution for 60 min (Ranki et al., 1976). After the procedure, ANAE positive lymphocytes could be distinguished by a single or a few reddish-brown distinct spots in their cytoplasms. The percentage of ANAE positive lymphocytes of total lymphocytes in each specimen was counted.
Results
A s s e e n f r o m T a b l e 1, t h e b l o o d s a m p l e s u s u a l l y s h o w e d a d i s t i n c t l y g r e a t e r p r o p o r tion o f l y m p h o c y t e s to be A N A E positive than the respective ear samples. Only t w i c e t h e p r o p o r t i o n w a s slightly g r e a t e r in effusion. T h e m e a n p e r c e n t a g e for T cells in all t h e e f f u s i o n s w a s 2 1 % . T h i s figure, a well as t h o s e f o r t h e r i g h t ( 2 0 % ) a n d t h e left ( 2 1 % ) ear, w a s h i g h l y s i g n i f i c a n t l y ( P < 0.001, t h e S t u d e n t ' s t-test) s m a l l e r t h a n t h e c o r r e s p o n d i n g p e r c e n t a g e for b l o o d s a m p l e s (53%).
Table 1. Percentages of ANAE positive cells of lymphocytes in middle ear effusions and blood in SOM Patient No. 1
2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 Mean S.D.
Right ear 32 -10 34 25 14 0 5 I0
Left ear
Blood
-48 18 20 --42 25 4 6 -24 28 -20 12
12 8 4 ----12 -14 42 -19 -14 50 56 30 10 30 12 22 10 40 6 28 6
45 55 20 40 77 60 54 44 32 52 60 57 38 64 50 44 53 55 68 46 60 56 66 66 55 65 42 70 37
20 13
21 16
53 13
--
T Cells in Secretory Otitis Media
165
Discussion Sloyer et al. (1976) found rosette forming lymphocytes and the phytohaemagglutinin stimulation index in the middle ear effusion to be greatly decreased when compared with peripheral blood in children with acute otitis media. However, these functional methods specific for T cells cannot be used in studying often rather viscous and gluelike effusions of SOM. ANAE marks specifically human T cells directly from cell smears (Ranki et al., 1976). Also monocytes stain with this method but very strongly and diffusely (Ranki et al., 1976), which, associated with their different morphology, makes them easy to distinguish from lymphocytes. In our earlier studies (Palva et al., 1976) we found lymphocytes and neutrophils to be the dominant cell types in the effusions of SOM, thus supporting the idea of inflammatory origin of the disease. The present study shows that the distribution of T and non-T (principally B) cells in SOM effusions significantly differs from that of peripheral blood, and that there is paucity of T cells in the effusion. When we further remember that the normal middle ear mucosa is almost totally devoid of inflammatory cells, our finding suggests relative local shortage of T cells in SOM. This may disturb local T and B cell interactions (Sloyer et al., 1976) or weaken the possible cell mediated immune defence systems in the middle ear and thus it may be a link in the chain of events leading to SOM. However, further studies concerning the nature of different types of lymphocytes in the effusions and mucosa of the middle ear in various otitic processes are needed before the significance of our finding can be assessed. We have such studies in progress.
References Mueller, J., Brun del Re, G., Buerki,H., Keller,H.-U., Hess, M. W., Cottier,H.: Europ. J. Immunol. 5, 270--274 (1975) Palva, T., Karma, P., Holopainen, E.: Ann. Otol. (St. Louis) Suppl. 25, 103-109 (1976) Ranki, A., T&terman, T. H., H~iyry,P.: Scand. J. Imrnunol. 5, 1129-1138 (1976) Sloyer, J. L., Jr., Ploussard, J. H., Howie, V. M.: Ann. Otol. (St. Louis) Suppl. 25, 130-134 (1976) Tomasi, T. B.: Ann. Otol. (St. Louis) Suppl. 25, 87-89 (1976) Waldman, R. H., Spencer, C. S., Johnson, J. E., III.: CeUul. Immunol. 3, 294-300 (1972) Received December 6, 1977
Archivesof Oto-Rhino-Laryngology 9 Springer-Verlag 1978
T Lymphocytes in the Effusions of Secretory Otitis Media* P. Sipil/i ~, P. Ryh/inen 2, and P. Karma 3 Departments of OtolaryngologyI and Anaesthesiology2, University of Oulu, Finland, Institute of Clinical Sciences3, University of Tampere, Teiskontie 35, SF-33520 Tampere 52, Finland
Summary. Smears were made from 39 middle ear effusions in 29 children with secretory otitis media (SOM) and stained with acid oz-naphthyl acetate esterase (ANAE) to demonstrate T lymphocytes. The effusions showed on the average 21% of the lymphocytes to be A N A E positive, while simultaneously drawn peripheral blood revealed the proportion of A N A E positive cells to be 53% (P < 0.001, the Student's t-test). Our finding suggests that T cell dependent defence mechanisms may be locally disturbed in SOM.
Key words: Secretory otitis media - T cells - Acid a-naphthyl acetate esterase.
It is proposed (Tomasi, 1976) that local cell mediated immune reactions, which have been demonstrated to take place in the respiratory tract (Waldman et al., 1972), may play an important role in the protection of the middle ear against infection. However, although there have been numerous immunological studies - which have failed to show any systemic or antibody mediated local immune defect as a cause of secretory otitis media (SOM) - the possible role of local cell mediated immune mechanisms in SOM have not, so far as we know, been studied. We present here the preliminary results of our studies concerning the occurrence and role of T lymphocytes, as marked with acid ce-naphthyl acetate esterase (ANAE) according to the method presented by Mueller et al. (1975), in the middle ear effusions of children with SOM.
Material and Methods Effusion was aspirated from 39 middle ears in 29 children (mean age 5.1 years, range 0.4-11.3 years) who came for tympanostomybecause of SOM. A sample of peripheral bloodwas drawn simultaneously. Immediately after taking the specimens, smears from blood and the secretion were made and the * Supported by the grant from the Emil Aaltonen Foundation, Finland Offprint requests to: Dr. P. Karma (address see above)
0302--9530/78/0220/0163/5 01.00
164
P. Sipil/i et al.
slides were allowed to dry for 30 min. The fixation and staining of the smears for the demonstration of ANAE activity in the lymphocytes were done according to Mueller et al. (1975), with the exception of the smears being counterstalned with 1% aqueous toluidine blue solution for 60 min (Ranki et al., 1976). After the procedure, ANAE positive lymphocytes could be distinguished by a single or a few reddish-brown distinct spots in their cytoplasms. The percentage of ANAE positive lymphocytes of total lymphocytes in each specimen was counted.
Results
A s s e e n f r o m T a b l e 1, t h e b l o o d s a m p l e s u s u a l l y s h o w e d a d i s t i n c t l y g r e a t e r p r o p o r tion o f l y m p h o c y t e s to be A N A E positive than the respective ear samples. Only t w i c e t h e p r o p o r t i o n w a s slightly g r e a t e r in effusion. T h e m e a n p e r c e n t a g e for T cells in all t h e e f f u s i o n s w a s 2 1 % . T h i s figure, a well as t h o s e f o r t h e r i g h t ( 2 0 % ) a n d t h e left ( 2 1 % ) ear, w a s h i g h l y s i g n i f i c a n t l y ( P < 0.001, t h e S t u d e n t ' s t-test) s m a l l e r t h a n t h e c o r r e s p o n d i n g p e r c e n t a g e for b l o o d s a m p l e s (53%).
Table 1. Percentages of ANAE positive cells of lymphocytes in middle ear effusions and blood in SOM Patient No. 1
2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 Mean S.D.
Right ear 32 -10 34 25 14 0 5 I0
Left ear
Blood
-48 18 20 --42 25 4 6 -24 28 -20 12
12 8 4 ----12 -14 42 -19 -14 50 56 30 10 30 12 22 10 40 6 28 6
45 55 20 40 77 60 54 44 32 52 60 57 38 64 50 44 53 55 68 46 60 56 66 66 55 65 42 70 37
20 13
21 16
53 13
--
T Cells in Secretory Otitis Media
165
Discussion Sloyer et al. (1976) found rosette forming lymphocytes and the phytohaemagglutinin stimulation index in the middle ear effusion to be greatly decreased when compared with peripheral blood in children with acute otitis media. However, these functional methods specific for T cells cannot be used in studying often rather viscous and gluelike effusions of SOM. ANAE marks specifically human T cells directly from cell smears (Ranki et al., 1976). Also monocytes stain with this method but very strongly and diffusely (Ranki et al., 1976), which, associated with their different morphology, makes them easy to distinguish from lymphocytes. In our earlier studies (Palva et al., 1976) we found lymphocytes and neutrophils to be the dominant cell types in the effusions of SOM, thus supporting the idea of inflammatory origin of the disease. The present study shows that the distribution of T and non-T (principally B) cells in SOM effusions significantly differs from that of peripheral blood, and that there is paucity of T cells in the effusion. When we further remember that the normal middle ear mucosa is almost totally devoid of inflammatory cells, our finding suggests relative local shortage of T cells in SOM. This may disturb local T and B cell interactions (Sloyer et al., 1976) or weaken the possible cell mediated immune defence systems in the middle ear and thus it may be a link in the chain of events leading to SOM. However, further studies concerning the nature of different types of lymphocytes in the effusions and mucosa of the middle ear in various otitic processes are needed before the significance of our finding can be assessed. We have such studies in progress.
References Mueller, J., Brun del Re, G., Buerki,H., Keller,H.-U., Hess, M. W., Cottier,H.: Europ. J. Immunol. 5, 270--274 (1975) Palva, T., Karma, P., Holopainen, E.: Ann. Otol. (St. Louis) Suppl. 25, 103-109 (1976) Ranki, A., T&terman, T. H., H~iyry,P.: Scand. J. Imrnunol. 5, 1129-1138 (1976) Sloyer, J. L., Jr., Ploussard, J. H., Howie, V. M.: Ann. Otol. (St. Louis) Suppl. 25, 130-134 (1976) Tomasi, T. B.: Ann. Otol. (St. Louis) Suppl. 25, 87-89 (1976) Waldman, R. H., Spencer, C. S., Johnson, J. E., III.: CeUul. Immunol. 3, 294-300 (1972) Received December 6, 1977
