Cell mediated immunity in patients with subacute sclerosing panencephalitis

Cell Mediated Immunity in Patients with Subacute Sclerosing Panencephalitis Sabiha Aysun, MD, bzden Sanal, MD, Yavuz Renda, MD, izzet Berkel, MD, Kalbiye Yalaz, MD, Fiigen Ersoy, MD and Emire Ozdirim, MD Cell mediated immunity was assessed on the basis of total lymphocytes (TL), total T lymphocytes (TTL) counts, delayed skin test responses and in vitro leucocyte migration inhibition test (LIF) production in 25 patients with SSPE, classified according to the clinical stages of -the disease. The patients in stage I of the disease did not show any defect in cellular immunity while the patients in stage II showed decreased TL and TTL counts, more negative skin test responses to PHA, SKSD and PPD, and unresponsiveness to SKSD of LIF production. When the patient group was evaluated as a whole, only the TL counts and the skin test responses against SKSD differed from those in the controls. These results suggest that the defects in cellular immunity demonstrated in the patients with SSPE may be due to SSPE or the measles virus itself rather than to a genetic factor predisposing patients to SSPE. Aysun S, SanalD, Renda Y, Berkel t, Yalaz K, Ersoy F, Dzdirim E. Cell mediated immunity in patients with subacute sclerosing panencephalitis. Brain Dev 1984; 6: 391-6

Subacute sclerosing panencephalitis (SSPE), a progressive encephalitis occurring in children and young adults [1-3], is regarded as a model of human slow virus infection associated with measles virus [4-6]. The features which are responsible for the persistence of this viral infection are not fully understood. Since the protection and recovery from measles infection largely depend on cellular immune response [7], the cell mediated immunity in patients with SSPE has been investigated by various

From the Departments of Neurology (SA, YR, KY, EO) and Immunology (OS, IB, FE), Institute of Child Health, Hacettepe University, Ankara. Received for publication: April 6, 1984. Accepted for publication: August 1, 1984.

Key words: Subacute sclerosing panencephalitis (SSPE), cell mediated immunity. Correspondence address: Dr. Sabiha Aysun, Department of Neurology, Institute of Child Health, Hacettepe University, Ankara, Turkey.

authors with controversial results [8-19] and the pathogenesis of SSPE is still unclear. In this study cell mediated immunity was investigated in SSPE with various parameters and the relationship between the clinical stage and immunological abnormalities was analyzed. Patients and Methods Between 1978 and 1980, 25 patients (19 males, 6 females) between 4-17 years of age with SSPE diagnosed on the basis of clinical and electroencephalographical findings, and increased measles antibodies in the cerebrospinal fluid and/or serum* were studied. Seven patients were in the first stage, 17 in the second and one in the third, according to Freeman's staging system [20] (Table 1). The control group included 21 age-matched healthy chil-

* Measles

antibody titers in CSF and serum were kindly determined by P. Lachmann.

dren. The delayed hypersensitivity (skin test) responses were evaluated in 19 patients and 19 controls by using a battery of antigens in appropriate doses. These antigens were: phytohemagglutinin (PHA) (pHAP-Difco Lab, 10 p.g/O.l ml), Candida (Cand) (Hollister-Stier Lab , 50 PNU/O.l ml), streptokinase-streptodornase (SK-SD) (Lederle Lab , SK, 50 U/O.l ml , SD, 12.5 U/O.l ml) and PPD (Refik Say dam Institute,S U/O.l ml). An induration measuring at least 5 mm in diameter after 48 hours was considered as a positive reaction.

The percentage of T lymphocytes was determined from the E rosettes as described by Jondal et al [21). Total lymphocytes (TL) were determined in 25 patients with SSPE and 21 controls, and total T lymphocyte counts (TTL) in 24 SSPE and 20 controls. A leucocyte migration inhibition test (UF) was performed in 19 patients using PH A, Cand and SK-SD antigens as described by Clausen [22]. The Mann-Whitney U test, the x 2 test with Yates co rrection where appropriate and Fisher 's test were used for stat istical analysis.

Table I Clinical features of patients with SSPE Sex

Duration a/disease (in mas)

12

F

6

Prednisolone - 20 days

10

M

3

None

12

M

12

Case

Age

1

2 3 4

9

M

5 6 7

4

M

Stage

Type of treatment and duration

Phenytoin -1 wk Clonazepam - 5 mos

20/30

Interval between treatm ent and immunologic testing 1 mo 5 mos Ongoing

None None

12

9

10 8 4 8

10 11

17 10

12

7

F

5

II

None

13

9

M

4

II

Steroid -1 wk Clonazepam

4 m os Ongoing

14

14

M

II

Steroid - 3 wks Carbamazepine - 3 wks

3 days

15

11 5 4 10 8

II

Clonazepam - 2 wks

3 wks

II

Clonazepam - 3 wks

Ongo ing

II

None

8

16 17 18 19

20

M

M M

F F M

M M M

1 1.5 9 1.5 2

6 1.5 15/30

F

None Clonazepam - 3 wks

10 days

II

Phenytoin - 3 wks

Ongoing

II

Phenytoin - 6 wks Clonazepam - 6 wks

6 wks

II

Clonazepam

Ongoing

II

Phenytoin Clonazepam

Ongoing

II

None Phenytoin - ? Phenobarbital- ?

F

3

II

M

8 11

Clonazepam-4 mos

Ongoing

M

6 4

II

21

II

Clonazepam - 2 mos

1 mo

22

6

M

6

II

Clonazepam - 1 mo Mysoline - 1 mo

2 wks

23

13 7

M

3

II

None

24

F

1

II

Phenytoin -1 wk Phenobarbital-l wk

Ongoing

25

8

M

3

III

Phenytoin - 2 mos

Ongoing

392 Brain & Development, Vol 6, No 4, 1984

Results ' The TL and TTL in the patients and the controls are shown in Figs 1 and 2. Both TL and TTL numbers of the patients in stage II of the disease were significantly lower than those in the controls but no significant difference was

I

5500

P< 0.05

l

found between patients in stage I and the controls (Figs 1 and 2). When the patient group was evaluated as a whole, the TL number was found to be significantly lower while TTL did not show any significant difference in comparison with controls.

I.

5500

•

P> 0.05

r- P <0.05,

5000

5000 o

o

4500

4500 o

4000

-I

o

3500

f-

o

•

1:

5 3000

u

.....III>. o

~a. 2500 E ~

~

.8 2000 ~

1500

o

ME

o

E

~ -I

o

3500

!::.

•

III

o

4000-

o

M~ ~

...c

o o

•

• • • •

5 3000-

u

II .....

~ o

-a o

• I• ••

o

8 8

• • • ••

E

x )(

>.

-I

~

500

o

o o

x

2500

2000

•

1500

x

~

x

I

o

•

x

• •

III

•

•• 1000

,

x

x

x x

x x

§

0

o o

8

8 o

x x

1000-

•

x

•

500-

Z .. 1. 84

•

o

L

P·0.0336

• PATIENTS

o CONTROLS

• STAGE I

LZ=194 ~ P=00262 Z=1.46 P=0.072

I

xSTAGE'[

oCONTROLS

Fig 1 The distribution of TL in patients with SSPE and controls.

Aysun et al: Cell mediated immunity in SSPE 393

III the skin test responses, there was no difference between the patients in the first stage and the controls but there was a significant difference in the responses to PHA, SKSD and PPD between the patients in the second stage and the controls. When the patient group was evaluated as a whole only the skin test response

4000

to SKSD showed a significant difference in comparison with controls (Table 2). The rate of positive responses to PHA and Candida with LIF production in the patients in stage I and stage II of the disease was not different from that in the controls, however , the positive response to SKSD was significantly

4000 o

o

3500

3500

3000

3000

0 0

Me

E

...... ..J

~

2500

-i=! ......

••

0

I-

...c

8

0

u III

(j'

•

0

8

:J 0

U

2000

OJ

0

t

E

>-

1500

"E .....

I•

• •

500

{j'

2000-

>-'

to!....c

1500

••

~

• • • • • • •

•

8

1000

0

8

§0

~X

0 0

0

x x

•

E

0

0

0

.c a.


I 1000

0

0 0

§0

• •

,!:!

....

0

0

.&: Q.

..J ~

2500

."

III

1: :J +'

8

ME E

• •

~

x

x

x

x x

8 0

•

0 Z .. 0·91 P-0·1814

• PATIENTS

o

CONTROLS

L

0

• STAGE I

Fig 2 The distribution of TTL in patients with SSPE and controls.

394 Brain & Development, Vol 6, No 4, 1984

L

Z = 2.789 P=00026

---.J

Z=1352 P .. 0.0885 x STAGE ][

0

0

x

x

500

0

oCONTROLS

0

Table 3 The results of the LIF test in patients with SSPE and controls

Table 2 Delayed hypersensitivity (skin test) responses in patients with SSPE and controls Patients No* %** PHA Stage I Stage II Total*** Candida Stage I Stage II Total*** SKSD Stage I Stage II Total*** PPD Stage I Stage II Total***

2/7 8/12 10/20

29 67 50

4/7 12/12 17/ 20

Controls No* %**

Patients No* %**

16

p > 0.05 p <0.05 p> 0.05

57 100 85

16/ 19 84

p> 0.05 p > 0.05 p > 0.05

4/7 12/12 17/20

57 100 85

9/19 48

p >0.05 p < 0.005 p < 0.05

3/7 9/ 12 13/20

43 75 65

3/19

7/19

37

PHA Stage I Stage II Total*** Candida Stage I Stage II Total*** SKSD Stage I Stage II Total***

Controls No* %**

4/7 6/12 11 / 20

57 60 55

7/18 39

p > 0.05 p > 0.05 p > 0.05

5/7 9/ 12 15/ 20

71 75 75

10/19 53

p > 0.05 p > 0.05 p > 0.05

5/ 7 11/12 17/20

71 92 85

8/19 48

p > 0.05 p <0.05 p > 0.05

*: number with negative results/number tested, **: percent negative, ***: includes the patients in stage III of the disease.

p > 0.05 p < 0.05 p > 0.05

*: number with negative results/number tested, **: percent negative, ***: includes the patients in stage III of the disease.

lower in stage II. When the patient group was evaluated as a whole, this difference was not significant (Table 3). Discussion The cell mediated immunity was studied in SSPE patients by various groups of investigators and controversial results were reported . In most of the studies the number of patients were limited and did not allow analysis of the relationship between the stages of the disease and the severity of the immune defect. Ahmed et al [23] suggested that the assay systems and antigens applied, the stage of the disease , secondary infections and therapeutic agents administered to the patients may influence the results of related studies. Jabbour et al [12] reported that neither the stage and duration of the disease nor the serum measles titers had any relationship with the delayed hypersensitivity (skin test) responses in these patients. However, in the study of Valdimarsson et al [17] , the anergy to common skin test antigens was more pronounced when the disease was more advanced.

In our study we also observed that anergy became more striking in the advanced stage of the disease. Yamanaka et al [24] found markedly decreased IL numbers but normal numbers of TTL in patients with SSPE. Derakhshan et al [25] noted that the number of active T cells was normal whereas the TTL was lower than in the controls. In neither of these studies were immune parameters evaluated according to the stage of the disease. We found a significant decrease in TL numbers when the patient group was evaluated as a whole, while the TTL was within normal limits. When the patients were evaluated according to their clinical stages, neither TL nor TTL numbers in stage I showed any significant difference in comparison with the controls, but TL and TTL were significantly lower in the patients in stage II of the disease. The decreased numbers of TL and TTL, the occurrence of more patients with skin test anergy and the greater number of patients showing unresponsiveness to SKSD of LIF production in stage II suggest that there is a generalized suppression in cellular immunity in patients with the advanced disease . This suppression may be caused by the SSPE virus or the measles virus itself, but not by a genetic defect. In another analytical study we were

Aysun et al: Cell mediated immunity in SSPE 395

not able to demonstrate any increase in HLA antigen frequency in SSPE patients [26]. If a genetic defect is responsible for this suppression, then generalized suppression in cellular immunity should also be seen in the first stage. The conflicting results in earlier studies may be due to a stage dominancy . In studies with more patients in stage I of the disease, a prominent defect in cellular immunity would not be found. Whereas in studies with more patients in the second stage, this defect would be more prevalent. In conclusion we consider that the reduction in cellular immunity in patients with SSPE may be the result of the immunosuppressive effect of the virus, medication used or malnutrition rather than a predisposing immune response defect. Acknowledgments This study was supported by a grant (TAG 397) from the Technical and Research Council of Turkey. References 1. Canal N, Torck P. An epidemiological study of subacute sclerosing panencephalitis in Belgium. J Neural Sci 1964;1:380-9. 2. Lundon JR, Barnett HJ. Subacute inclusion body encephalitis in an Asian in the third decade. Can Med Assoc J 1966;95 :1151-3. 3. Dayan AD, Cuming IN. An infantile case of su bacute sclerosing panencephalitis with an abnormal ganglioside pattern in the brain. Arch Dis Child 1969;40:187-96. 4 . Barry SJ, Old stone MBA. Immunologic injury in measles virus infection. II . Suppression of immune injury through antigenic modulation. J Exp Med 1975;142 :864-76 . 5. Ter Meulen V, Enders-Ruckle G, Miiller D. Immunohistological, microscopical and neurochemical studies on encephalitis. III. Subacute progressive panencephalitis. Virological and immunohistological studies. Acta Neuropathol (Berl) 1969; 12 :244-59. 6. Dayan AD, Gastling JVT, Greaves JL, et al. Evidence of a pseudomyxovirus in the brain in subacute aclerosing panencephalitis. Lancet 1967; 1 :980-1. 7. Burnet FM. Measles as an index of immunological function. Lancet 1968;2 :611-3 . 8. Klajman A, Sternbach M, Ranon L, et al. Impaired delayed hypersensitivity in subacute sclerosing panencephalitis. Acta Paediatr Scand 1973 ;62:523-6. 9. Moulias RL, Reinert PH, Goust JM. Immunologic abnormalities in subacute sclerosing panencephalitis. N Engl J Med 1971;285: 1090.

396 Brain & Development, Vol 6, No 4, 1984

10. Gerson Kl , Haslem RHA. Subtle immunologic abnormalities in four boys with subacute sclerosing panencephalitis. N Engl J Med 1971 ; 285: 72-82 . 11. Sharma MK, Grover WD, Huff DS, et al. Studies of immunologic competence in subacute sclerosing panencephalitis (SSPE). Clin R es 1971; 19:730. 12. Jabbour JT, Roane JA, Sever J1. Studies of delayed dermal hypersensitivity in patients with subacute sclerosing panencephalitis. Neurology 1969;19:929-31. 13. Kreth HW, Kaeckell YM, ter Meulen V. Cellular immunity in SSPE. Med Microbiol Immllnol 1974; 160:191-9. 14. Saunders M, Knowles M, Chambers MS, et al. Cellular and humoral responses to measles in subacute sclerosing panencephalitis. Lancet 1969; 1:72-4. 15. Mizutani H, Mizutani H, Saito S, et al. Cellular hypersensitivity in subacute sclerosing panencephalitis. JAMA 1971;216: 1201-2. 16. Thurman GB, Ahmed A, Strong DM, et al. Lymphocyte activation in subacute sclerosing panencephaJitis virus and cytomegalovirus infections. J Exp Med 1973; 138:839-46 . 17. Valdimarsson H, Agnarsdottir G, Lachmann PJ. Cellular immunity in subacute sclerosing panencephalitis. Proc Roy Med 1974 ;67:1125-9 . 18. Livni E, Kott E, Ranon Y, et al. Cell-mediated immunity in subacute sclerosing panencephalitis. IsrJ Med Sci 1976;12:1183-8. 19. Sheremata W, Sazant A, Watters G. Subacu te sclerosing panencephalitis and multiple sclerosis: in vitro measles immunity and sensitization to myelin basic protein. Can Med Assoc J 1978; 118:509-13. 20. Freeman JM . The clinical spectrum and early diagnosis of Dawson encephalitis (with preliminary notes on treatment). J Pediatr 1969 ; 75 :590-605 . 21. Jondal M, Holm G, Wigzell H. Surface markers on human T and B lymphocytes. J Exp Med 1972;136 :207-15. 22. Clausen E. Tuberculin-induced migration inhibition of human peripheral leucocytes in agarose medium. Acta Allergol 1971;26:56-80. 23. Ahmed A, Strong DM, Sell KW, et al. Demonstration of a blocking factor in the plasma and spinal fluid of patients with subacute sclerosing panencephalitis. J Exp Med 1974; 139:902-24. 24. Yamanaka T, Chiba S, Nakao T, et al. Cell mediated immunity to measles virus in subacute sclerosing panencephalitis. Tohokll J Exp Med (Sendai) 1977;122:175-81. 25. Derakhshan I, Massoud A, Foroozanfar N, et al. Subacute sclerosing panencephalitis. Clinical and immunologic study of 23 patients. Neurology 1981 ;31: 177-80. 26. Aysun S, Ersoy F, Sanal G, et a1. Histo compatibility antigens in subacute sclerosing panencephalitis. Arch Neurol 1983;40:38.