High-dose intravenous-penicillin in neurosyphilis: effect on intrathecal synthesis of IgG, IgM, IgA and IgD Hens MJ, Lolli F, Martin-Moro M, Gimknez-Roldhn S , Link H. High-dose intravenous-penicillin in neurosyphilis: effect on intrathecal synthesis of IgG, IgM, IgA and IgD. Acta Neurol Scand 1990: 82: 381-385. We evaluated occurrence and levels of intrathecal synthesis of IgG, IgM, IgA and IgD by calculating corresponding index values in 8 patients with definite and 4 with suspected or possible neurosyphilis, prior to, during and after high-dose intravenous penicillin therapy. Four patients with active neurosyphilis displayed intrathecal synthesis of IgG, IgM and IgA. Only 2 of them showed elevated IgD index, and both had taboparesis, pleocytosis and positive VDRL in CSF, and simultaneous elevation of the IgG, IgA and especially of the IgM indices. This suggests that intrathecal synthesis of IgD may occur in patients with severe CNS inflammation in response to diffuse CNS treponemal damage. Penicillin therapy incited transitory elevation of one or more of the immunoglobulin index values in most patients, possibly in response to massive treponemal lysis inside the CNS. Thereafler, the values became mostly normalized but exceptions occurred, including one patient who had elevated IgG and IgM index which persisted 3 1 months after therapy. Our data indicate that determinations of immunoglobulin indices performed on consecutive specimens from individual patients with neurosyphilis may be helpful in the evaluation of treatment.
Since Kabat et al. reported increased fraction of gammaglobulin in the cerebrospinal fluid (CSF) in neurosyphilis (l), there is cumulative evidence indicating that intrathecal synthesis of immunoglobulins of different isotypes occurs in this disease (2-4). Increased CSF IgG concentrations when expressed as% of total protein (2, 5), elevated CSF IgG index (6-lo), evidence of local synthesis of IgG specific for Treponema (T) pallidum (1 1-13), and oligoclonal IgG bands (6, 11, 14-17) which may contain antibodies specific for T. pallidum (6, 8, 13) have been reported. These bands may persist for months or even years after therapy (5, 6, 9). Similarly, local synthesis of IgM and IgA (2, 6, 18-21), and of T. pallidum-specific IgM and IgA has been described (13, 22). Persistent intrathecal synthesis of IgM and IgA has been described in 2 patients, lasting for 7 months and one year, respectively, following therapy (6). Information regarding intrathecal synthesis of IgD in this condition is scarce, but was reported by Out et al. in 40% of patients with neurosyphilis, as well as in a variety of other inflammatory neurological disorders (23). Treatment with penicillin in high dosage administered by the intravenous route is currently recommended to achieve effective concentrations in CSF (24). We now report the effect of this treatment
M. J. Hens’.’, F. Lolli’, M. Martin-Moro’, S. GimCnez-RoldBn*, H. Link’
’
Departments of Neurology, Karolinska Institutet, Huddinge University Hospital, Stockholm, Sweden, Hospital General Gregorio Maranbn, Madrid, Spain
Key words: neurosyphilis; cerebrospinal fluid; immunoglobulin; oligoclonal bands: treatment
H. Link, Department of
Neurology, Huddinge University Hospital, S-141 86 Huddinge, Sweden Accepted for publication July 16, 1990
strategy on the occurrence and levels of immunoglobulins belonging to different isotypes, including IgD, by calculating corresponding index values. Patients presenting with a “classical” clinical picture such as dementia paralytica or tabes dorsalis were compared with seropositive patients with organic central nervous system (CNS) disorders which may suggest neurosyphilis. Multiple specimens obtained from individual patients were examined in parallel in the same analysis to better identify possible changes of the intrathecal B cell response after institution of treatment. Material and methods
Twelve Spanish patients with positive fluorescence treponemal antibody absorption (FTA-abs) test in serum were included. Four of them had “classical” involvement of the brain or spinal cord indicative of neurosyphilis, i.e. dementia paralytica, tabes dorsalis, or a combination of both, in addition to CSF abnormalities including increased cell count and positive VDRL in CSF, thereby suggesting “active” syphilitic disease of the CNS (25). These patients are diagnosed as Definite Active Neurosyphilis (DANS) (patients- 1-4, Table 1). Four patients had “classical” neurological findings, 38 1
Hens et a]. irrespective of any previous attempts of treatment, but no CSF abnormalities, and it was therefore difficult to imply their clinical deterioration to persistent T. palladium in the CNS or to non-infective mechanisms (26, 27). We termed these patients Possibly Active Neurosyphilis (PANS) (patients 5-8, Table 1). A third subgroup was composed of four patients who had a reactive serum FTA-abs and sometimes VDRL as well, but normal CSF. All four patients displayed the recent onset of a neurological disorder such as epilepsy, ischaemic stroke or parkinsonism in addition to the positive serology for syphilis (28). These patients were considered as having Possible Neurosyphilis (PNS) (patients 9-12, Table 1). Table 2 summarizes the clinical diagnoses, serological reactivity, and CSF findings at admission for these three subgroups of patients. Informed written consent for this study was obtained from the patients or their relatives.
Penicillin therapy
All 12 patients received a course of high-dose penicillin by the intravenous route, either following the schedule recommended by Center for Disease Control, Atlanta, U.S.A. (24) or by Boudin et al. (29), which is commonly used in France (30), Morocco (31) and Spain (32). CSF and serum studies
Simultaneously taken serum and CSF specimens, the latter obtained by atraumatic lumbar puncture, were sampled from every patient prior to institution of therapy, during treatment and thereafter, as shown in Table 1. The samples were kept at - 70 C until use. Automated immunoprecipitation nephelometry was utilized to measure albumin and IgG concentrations in unconcentrated CSF and serum. O
Table 1. Effects of penicillin therapy on CSF variables in neurosyphilis. Patients No. 1-4 had definite active neurosyphilis (DANS), No. 5-8 had possibly active neurosyphilis (PANS), and No. 9-12 had possible neurosyphilis(PNS). Patient No. 1 and 2 had two courses(a and b)of penicilin treatment. TD = Tabes Dorsalis. DP = Dementia Paralytica. TP = Taboparesis. MVS = Meningovascular Syphilis. PK = Parkinsonism. Numbers within brackets denote upper reference values. ND = not done.
CSF indices for Patient No.
Diagnosis
1. a
DANS (TP)
Days after initiation of therapy Before 5
8 1. b
(5 months later)
2. a
DANS (DP)
2. b (10 months later)
3
DANS (TP)
4
DANS (TP)
5
PANS (TD)
6
PANS (TD)
7
PANS (TD)
8
PANS (MVS)
9
PNS (PK)
10
PNS (Stroke)
11 12
PNS (Epilepsy) PNS (Stroke)
Before 14 19 Before 23 Before 10 Before 6 10 31 months later Before 6 10 Before 23 Before 10 Before 21 Before 10 Before 7 Before 20 23 Before 6 Before 6
CSFiSerum albumin ratio
IgG (0.7)
IgM (0.06)
IgA (0.34)
IgD (0.50)
7.5 5.2 5.9 6.5 7.6 6.2 5.7 7.4 4.6 5.1 7.9 4.9 4.7 2.9
2.9 3.2 2.7 1.0 0.7 0.7 3.1 2.4 2.9 2.0 2.5 3.0 2,9 1.1
0.88 0.68 0.62 0.23 0.17 0.19 0.83 0.51 0.22 0.1 1 3.30 4.82 4.32 0.31
0.80 1.oo 1.07 0.46 0.54 0.46 0.41 0.36 0.31 0.22 0.54 0.50 0.48 ND
0.32 ND 0.32 0.33 ND 0.23 0.34 0.26 0.15 0.18 0.67 ND 1.04 ND
5.1 4.7 4.6 2.7 2.9 4.4 3.2 5.9 7.3 4.6 5.1 4.2 4.5 4.8 5.6 3.4 3.3 3.4 4.6 5.7
6.6 10,4 8.6 0.8 0.7 0.6 0.6 0.4 0.6 0.6 0.7 0.6 0.7 0.6 0.5 0.5 0.5 0.6 0.5 0.5
3.45 5.96 2.49 0.14 0.1 1 0.05 0.03 0.04 0.04 0.05 0.25 0.07 0.12 0.04 0.10 0.08 0.06 0.13 0.04 0.04
2.42 3.64 2.68 0.26 0.27 0.20 0.25 0.22 0.20 0.23 0.53 0.19 0.18 0.30 0.36 0.29 0.20 0.24 0.31 0.3 1
1.10 ND 1.33 0.22 0.21 0.05 0.1 1 0.00 0.00 0.26 0.35 0.17 0.22 0.09 ND 0.13 0.13 0.18 ND ND
'
' Upper reference levels fluctuate in relation to age, being 9.0 in all patients except No. 3, 9 and 10 where it is 7.2 (compare ref. 33). 382
Oligoclonal IgG bands in CSF
3 ND 3 2 ND 2 4 4 4 4
0 ND
0 ND
2 ND 2 3 0 0 0 0
0 0 0 0 0 0 ND 0
0 0 0 0
High-dose i.v.-penicillin in neurosyphilis
42-63 (64)
53-68 (58)
410
311
410
Clinical diagnosis Tabes dorsalis Dementia paralytica Taboparesis Meningovascular syphilis Stroke Late-onset epilepsy Parkinsonism
1 1 2 0 0 0 0
2
0 0
0 0 0
2 1 1
isoelectric focusing revealed 2-4 oligoclonal IgG bands restricted to CSF in 3 of these patients. Among the patients with PANS, only No. 5 hd increased IgG and IgM indices, and also oligoclonal IgG bands in CSF. In the group with PNS, borderline increase of the IgM index in Patient 9 was the only abnormality. Considering the CSF IgD index, Patients 3 and 4 (Table 1) with DANS had elevated values as a reflection of intrathecal IgD synthesis. Both had evidence of simultaneous intrathecal production of IgG, IgM and IgA; both were suffering from taboparesis and had in fact clinical evidence for the most severe and diffuse CNS involvement in our patient material, and also the highest IgM index values.
Laboratory abnormalities Positive serum FTA-abs’ CSF pleocytosis Range (per mm3) VDRL reactive CSF Range of titres
4 4 6-86 4 1/2-1/16
4 0 < -5 0 neg
4
Effects of therapy on immunoglobulin abnormalities
Table 2. Clinical and laboratory findings in 12 patients with neurosyphilis. DANS = Definite Active Neurosyphilis. PANS = Possibly Active Neurosyphilis. PNS = Possible Neurosyphilis. DANS
PANS
PNS
Number of patients
4
4
4
Age range (mean)
49-12 (58)
Sex (M/F)
1 0 1
0 0
0 < -5 0 neg
’ FTA-abs is equal to Fluorescence Treponemal Antibody absorption The blood-brain barrier was evaluated by determining the CSF/serum albumin ratio. Age-dependent reference values previously reported were used (33). To measure concentrations of IgM, IgA and IgD in CSF and serum, we used double antibody sandwich Enzyme Linked Immuno Sorbent Assay (ELISA) (34). The sera and CSF from individual patients were analyzed in parallel for protein levels. CSF IgG index equal to (CSF IgG/serum IgG) : (CSF albumin/serum albumin) was calculated (33). Our upper reference value is 0.7 and increased values are considered to reflect intrathecal IgG production. CSF indices for IgM, IgA and IgD were calculated as for IgG. Upper reference limits are 0.06 for IgM index, 0.34 for IgA index and 0.50 for IgD index, based on examination of patients with tension headache (34). Agarose isoelectric focusing followed by protein transfer to nitrocellulose membrane, immunostaining with double-antibody peroxidase labeling, and avidin-biotin amplification, was performed on unconcentrated CSF and diluted serum to detect oligoclonal IgG bands (35). Occurrence of two or more bands in CSF but not in corresponding serum, both run in parallel, was considered to reflect intrathecal synthesis of oligoclonal IgG. Results Pretreatment immunoglobulin abnormalities
All 4 patients with DANS had increased CSF IgG, IgM and IgA indices indicating intrathecal production of these immunoglobulins (Table 1). Agarose
Following initiation of therapy, DANS Patients 1,3, 4 displayed initial increase of the IgG index, Nos 3 and 4 had an initial elevation of the CSF IgM index and also of the IgD index, and Patients 1 and 4 of the IgA index; 8-23 days after initiation of therapy, when the final specimens were taken in Patients 1 , 2 and 4, most immunoglobulin index values had decreased, but in none of them had a previously elevated value become normalized. Patient 3 with DANS also examined 31 months after treatment, when he still displayed elevated IgG and IgM indices. IgA and IgD were not examined because of paucity of CSF. Patients 1, 2 and 4 had oligoclonal IgG bands in CSF, and the band patterns were unaffected 8-23 days after initiation of therapy. In the only patient with PANS (No. 5) who displayed evidence for intrathecal immunoglobulin synthesis in the form of elevated IgG and IgM indices, as well as oligoclonal IgG bands, both the IgG index and isoelectric focusing pattern normalized on re-examination about 3 weeks after initiation of treatment, while the IgM index was still elevated. However, PANS Patient 8 developed increased IgA and IgM indices following initiation of penicillin treatment, and an increase of the IgM index was similarly observed in 3 of the patients with PNS. DANS Patients 1 and 2 (Table 1) were treated again with high-dose penicillin 5 and 10 months, respectively, after the first course of therapy. Both had still elevated IgG and IgM index values as well as oligoclonal IgG bands, and one of them had also elevated IgA index. However, all values were lower when compared with those registered in relation to the first course of treatment. After a second course of penicillin treatment, both the IgG and IgM indices decreased further, but they were not fully normalized during the short follow-up periods. The IgA index remained elevated in one of these two patients. Only one of the 12 patients (No. 3, Table 1) 383
Hens et al. showed a mild derangement of the blood-brain barrier function as evaluated by the CSF/serum albumin ratio. This abnormality vanished early during therapy. Discussion
We have shown that penicillin treatment of patients with Definitive Active Neurosyphilis (DANS) may lead to further increase of intrathecal synthesis of IgG, IgA and IgM compared with pretreatment data. Furthermore, one patient with DANS had elevated IgG and IgM index values 31 months after treatment, reflecting persistence of the intrathecal B cell response. Taboparesis encountered in two patients was characterized by intrathecal production of IgG, IgM and IgD in parallel. These two patients also displayed the highest CSF IgM values which we registered. Elevated CSF IgG and IgM indices constituted in our series of patients the most sensitive markers for an intrathecal B cell response. Elevated values were present in some patients who otherwise lacked CSF abnormalities. Local synthesis of IgD as reflected by elevated IgD index was restricted to patients with the most severe clinical symptoms. Similar findings have been reported in other inflammatory nervous system disorders including bacterial meningitis and Lyme neuroborreliosis (23, 34). Intrathecal IgA production was also found to be common in our patients with neurosyphilis, unlike blood-brain barrier derangement which was evident in only one of the 12 patients. Its early correction under high-dose i.v. penicillin therapy is not an unexpected finding, provided that even i.m. penicillin schedules, which do not attain CSF treponemicidal concentrations, can normalize both CSF cell count and protein content (36-38). Our findings suggest that blood-brain barrier correction under therapy probably constitutes the earliest marker of CSF normalization, followed in time by a decrease in the intrathecal synthesis of immunoglobulins, a sequence of events already suggested by others (6, ll). Neurosyphilis, “the great imitator” (9), may present clinically in many ways, and seropositivity in individuals suffering from common neurological disorders as stroke or epilepsy may occasionally rise doubts regarding the significance of their serological findings. Four such patients in our series with normal CSF except a borderline increase of the CSF IgM index in one of them, were empirically submitted to penicillin therapy. Interestingly, two patients (No. 10 and 11) developed abnormal IgM index just while receiving treatment. The B cell response against T. pallidum starts with the synthesis of IgM antibodies (40),and intrathecal synthesis of previously undetected specific IgM antibodies has 384
been observed in response to therapy (41). Normal CSF has been reported in patients with neurosyphilis, and spirochetes have been identified in the CSF of patients with otherwise normal cell count and non-reactive VDRL (25, 42). The intrathecal synthesis of IgM, and its increase observed in our patients shortly after initiation of penicillin therapy, could be related to the lysis of dormant treponemes, followed by activation of the immune system and subsequent antibody production (2 l), in a similar way as has been described in cerebral cysticercosis during therapy with praziquantel (43). Documentation of appearance of intrathecal IgM production after treatment could be useful for differentiating symptomatic neurosyphilis, even when the clinical picture is atypical, from asymptomatic neurosyphilis (44). Our data documented that an intrathecal immune response may persist for nearly 3 years in patients whose neurosyphilis had strictly been treated according to current recommendations (24). Similar findings in a patient with clinically suspected neurosyphilis, do therefore not distinguish active disease from neurological sequelae in individuals who already had received adequate therapy in the past. Assuming that high-dose i.v. penicillin, such as used in this series, is curative, our findings appear in keeping with the concept of “CSF biologic scar” (1 1). However, in view of the fact that seropositive individuals with CNS disease but otherwise normal CSF show intrathecal synthesis of IgM just while receiving high-dosage penicillin, thereby indirectly suggesting that T. pallidum was present in their CNS, we would recommend repeated treatment and IgM index determinations in individuals suspected to have received inadequate therapy in the past or showing progressive neurological deterioration. Acknowledgements We thank Mr J Garcia Gutierrez for skilful technical assistance and Ms Yvonne Nilsson for excellent secretarial help. Dr M Hens was recipient of a grant from Margit and Folke Foundation. Dr F Lolli was recipient of a grant from the Italian Multiple Sclerosis Association (AISM).
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High-dose i.v.-penicillin in neurosyphilis 4. LATERRE EC, CALLEWAERT A, HEREMANS JF, SFAELLO Z. Electrophorectic morphology of gamma globulins in cerebrospinal fluid of multiple sclerosis and other diseases of the nervous system. Neurology 1970: 20: 982-990. 5. OXELIUS V, RORSMAN H, LAURELL A. Immunoglobulins of cerebrospinal fluid in syphilis. Br J Vener Dis 1969: 45: 121-1 25. N, KAM-HANSEN S, LINK H, 6. STRANDBERG-PEDERSEN MAVRAM. Specificity of immunoglobulins synthesized within the central nervous system in neurosyphilis. APMIS 1982: 90 (C): 97-104. 7. HOLMES MD, BRANT-ZAWADZKI MM, SIMONRP. Clinical features of meningovascular syphilis. Neurology 1984: 34: 553-556. 8. VARTDAL F, VANDVIK B, MICHAELSEN TE, LOE K, NORRBYE. Neurosyphilis: intrathecal synthesis of oligoclonal antibodies to Treponema pallidum. Ann Neurol 1982: 11: 35-40. P, POLONIM, ITTERIO A, CAMANA c , 9. MAZZARELLO CERONIC. Cerebrospinal fluid IgG changes in neurosyphilis after high-dose penicillin G treatment. Arch Neurol 1987: 44: 249. 10. KINNUNEN E, HILLBOMM. The significance of cerebrospinal fluid routine screening for neurosyphilis. J Neurol Sci 1986: 75: 205-21 1. 11. PRANGE HW, MOSKOPHIDIS M, SCHIPPER HI, M ~ ~ L LF. ER Relationship between neurological features and intrathecal synthesis of IgG antibodies to Treponema pallidum in untreated and treated human neurosyphilis. J Neurol 1983: 230: 241-252. E RMOSKOPHIDIS M. Estimation of the local pro12. M ~ ~ L LF, duction of antibodies to Treponema pallidum in the central nervous system of patients with neurosyphilis. Br J Vener Dis 1983: 59: 80-84. 13. VARTDALF, VANDVIKB. Characterization of classes of intrathecally synthesized antibodies by imprint immunofixation of electrophoretically separated sera and cerebrospinal fluids. APMIS 1983: 91 (C): 69-75. 14. CHU AB, SEVERJL, MADDENDL, et al. Oligoclonal IgG bands in cerebrospinal fluid in various neurological diseases. Ann Neurol 1983: 13: 434-439. 15. MILLERJR, BURKEAM, BEVERCT. Occurrence of oligoclonal bands in multiple sclerosis and other CNS diseases. Ann Neurol 1983: 13: 53-58. 16. GEORGEPM, LORIERAM, DONALDSON IMG. An evaluation of cerebrospinal fluid oligoclonal banding confirmed by immunofixation on agarose gel. J Neurol Neurosurg Psychiatry 1983: 46: 500-504. 17. XuX, MCFARLIN DE. Oligoclonal bands in CSF: twins with MS. Neurology 1984: 34: 769-774. 18. SINDICCJM, CAMBIASO CL, DEPRE A, LATERREEC, MASSONPL. The concentration of IgM in the cerebrospinal fluid of neurological patients. J Neurol Sci 1982: 55: 339-350. 19. SINDICCJM, DELACROIX DL, VAERMAN JP, LATERRE EC, MASSONPL. Study of IgA in the cerebrospinal fluid of neurological patients with special reference to size, subclass and local production. J Neuroimmunol 1984: 7: 65-75. EJ. Oligoclonal 20. KEIR G, WALKERRWH, THOMPSON immunoglobulin M in cerebrospinal fluid from multiple sclerosis patients. J Neurol Sci 1982: 57: 281-285. 21. SHARIEFMK, THOMPSON EJ. Immunoglobulin M in the cerebrospinal fluid: an indicator of recent immunological stimulation. J Neurol Neurosurg Psychiatry 1989: 52: 949-953. 22. MOLLERF, MOSKOPHIDIS M, PRANCE HW. Demonstration of locally synthesized immunoglobulin M antibodies of Treponemapallidum in the central nervous system of patients
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with untreated nsuros;.?hilis. J Neuroimmunol 1984: 7: 43-54. OUTTA, HISCHE€.%.I AS \VALBEEK HK, VAN DER HELM HJ. Immunoglobulin D in cerebrospinal fluid. Clin Chim Acta 1987: 165: 259-293. Centers for Disease Control. 1985 STD treatment guidelines. Atlanta: Massachusetts Med SOC,1985. SIMONRP. Neuros>philis. Ann Neurol 1985: 42: 606-613. GIM~NEZ-ROLDSN S , BENITOC, MARTfN M. Dementia paralytica: deterioration from communicating [?I hydrocaphalus. J Neurol Neurosurg Psychiatry 1979: 42: 501-508. GIM~NEZ-ROLDAN S , MARTfN M. Tabetic lightning pains: high-dosage intravenous penicillin versus carbamazepine therapy. Eur Neurol 1981: 20: 424-428. HOOSHMAND H, ESCOBAR MR, KOPFSW. Neurosyphilis. A study of 241 patients. JAMA 1972: 219: 726-729. BOUDING. Aspects actuels de la syphilis nerveuse et de son traitement. Rev Pract 1966: 16: 3099-3106. MICHEL D, BLANC A, LAURENT B, FOYATIERN, PORTAFAIX M. Etude biologique, psychomktrique et tomodensitomktrique de la neurosyphilis traitke. Rev Neurol (Paris) 1983: 139: 737-744. EM ALAOUI-FARIS M, CHKILIT, MEJEDELA et al. Les complications neurologiques de la syphilis. Etude de cinquante-trois cas. Sem HBp Paris 1986: 62: 3511-3515. MARTfN MOROM, BARTOLOMB P, DE ANDRBSc et al. Penicilina intravenosa en el tratamiento de la neurosifilis. Ann Neurobiol 1982: 45: 29-42. TIBBLING G, LINKH, OHMAN S . Principles of albumin and IgG analyses in neurological disorders. I. Establishment of reference values. Scand J Clin Lab Invest 1977: 37: 385-390. LOLLIF, HALAWA I, LINKH. Intrathecal synthesis of IgG, IgA, IgM and IgD in untreated multiple sclerosis and controls. Acta Neurol Scand 1989: 80: 238-247. OLSSONT, KOSTULAS V, LINKH. Improved detection of oligoclonal IgG in cerebrospinal fluid by isoelectric focusing in agarose, double-antibody peroxidase labelling, and avidin-biotin amplification. Clin Chem 1984: 30: 1246-1249. MOHRJA, GRIFFITHS W, JACKSONR, SAADAH H, BIRDP, RIDDLEJ. Neurosyphilis and penicillin levels in cerebrospinal fluid. JAMA 1976: 236: 2208-2209. TRAMONT EC. Persistence of Treponemapallidum following penicillin-G therapy. JAMA 1976: 236: 2206-2207. LOWHAGEN GB, ANDERSSON M, BLOMSTRAND C, ROUPE G. Central nervous system involvement in early syphilis. Part I. Intrathecal immunoglobulin production. Acta Derm Venereol (Stockh) 1983: 63: 409-417. KOFFMAN0. The changing pattern of neurosyphilis. Can Med Assoc J 1956: 74: 807-812. MOSKOPHIDISM, M ~ ~ L L EF.R Molecular analysis of immunoglobulins M and G immune response to protein antigens of Treponema pallidum in human syphilis. Infect Immun 1984: 43: 127-132. BEHRW, BARNERT J, WIENBECKM. Diagnostische probleme bei der Neurosyphilis. Dtsch Med Wochenschr 1988: 113: 1718-1722. JAFFEHW, KABINSSA. Examination of cerebrospinal fluid in patients with syphilis. Rev Infect Dis 1982: 4 (Suppl): S 842- S 847. ESTAROLB, JUAREZ H, IRIGOYENMDC, GONZALESBARRANCO D, CORONAT. Humoral immune response in patients with cerebral parenchymal cysticercosis treated with praziquantel. J Neurol Neurosurg Psychiatry 1989: 52: 254-257. LEE JB, FARSHYCE, HUNTEREF, HAMBIEEA, WOBIG GH, LARSENSA. Detection of immunoglobulin M in cerebrospinal fluid from syphilis patients by enzyme-linked immunosorbent assay. J Clin Microbiol 1986: 24: 736-740.
385
Since Kabat et al. reported increased fraction of gammaglobulin in the cerebrospinal fluid (CSF) in neurosyphilis (l), there is cumulative evidence indicating that intrathecal synthesis of immunoglobulins of different isotypes occurs in this disease (2-4). Increased CSF IgG concentrations when expressed as% of total protein (2, 5), elevated CSF IgG index (6-lo), evidence of local synthesis of IgG specific for Treponema (T) pallidum (1 1-13), and oligoclonal IgG bands (6, 11, 14-17) which may contain antibodies specific for T. pallidum (6, 8, 13) have been reported. These bands may persist for months or even years after therapy (5, 6, 9). Similarly, local synthesis of IgM and IgA (2, 6, 18-21), and of T. pallidum-specific IgM and IgA has been described (13, 22). Persistent intrathecal synthesis of IgM and IgA has been described in 2 patients, lasting for 7 months and one year, respectively, following therapy (6). Information regarding intrathecal synthesis of IgD in this condition is scarce, but was reported by Out et al. in 40% of patients with neurosyphilis, as well as in a variety of other inflammatory neurological disorders (23). Treatment with penicillin in high dosage administered by the intravenous route is currently recommended to achieve effective concentrations in CSF (24). We now report the effect of this treatment
M. J. Hens’.’, F. Lolli’, M. Martin-Moro’, S. GimCnez-RoldBn*, H. Link’
’
Departments of Neurology, Karolinska Institutet, Huddinge University Hospital, Stockholm, Sweden, Hospital General Gregorio Maranbn, Madrid, Spain
Key words: neurosyphilis; cerebrospinal fluid; immunoglobulin; oligoclonal bands: treatment
H. Link, Department of
Neurology, Huddinge University Hospital, S-141 86 Huddinge, Sweden Accepted for publication July 16, 1990
strategy on the occurrence and levels of immunoglobulins belonging to different isotypes, including IgD, by calculating corresponding index values. Patients presenting with a “classical” clinical picture such as dementia paralytica or tabes dorsalis were compared with seropositive patients with organic central nervous system (CNS) disorders which may suggest neurosyphilis. Multiple specimens obtained from individual patients were examined in parallel in the same analysis to better identify possible changes of the intrathecal B cell response after institution of treatment. Material and methods
Twelve Spanish patients with positive fluorescence treponemal antibody absorption (FTA-abs) test in serum were included. Four of them had “classical” involvement of the brain or spinal cord indicative of neurosyphilis, i.e. dementia paralytica, tabes dorsalis, or a combination of both, in addition to CSF abnormalities including increased cell count and positive VDRL in CSF, thereby suggesting “active” syphilitic disease of the CNS (25). These patients are diagnosed as Definite Active Neurosyphilis (DANS) (patients- 1-4, Table 1). Four patients had “classical” neurological findings, 38 1
Hens et a]. irrespective of any previous attempts of treatment, but no CSF abnormalities, and it was therefore difficult to imply their clinical deterioration to persistent T. palladium in the CNS or to non-infective mechanisms (26, 27). We termed these patients Possibly Active Neurosyphilis (PANS) (patients 5-8, Table 1). A third subgroup was composed of four patients who had a reactive serum FTA-abs and sometimes VDRL as well, but normal CSF. All four patients displayed the recent onset of a neurological disorder such as epilepsy, ischaemic stroke or parkinsonism in addition to the positive serology for syphilis (28). These patients were considered as having Possible Neurosyphilis (PNS) (patients 9-12, Table 1). Table 2 summarizes the clinical diagnoses, serological reactivity, and CSF findings at admission for these three subgroups of patients. Informed written consent for this study was obtained from the patients or their relatives.
Penicillin therapy
All 12 patients received a course of high-dose penicillin by the intravenous route, either following the schedule recommended by Center for Disease Control, Atlanta, U.S.A. (24) or by Boudin et al. (29), which is commonly used in France (30), Morocco (31) and Spain (32). CSF and serum studies
Simultaneously taken serum and CSF specimens, the latter obtained by atraumatic lumbar puncture, were sampled from every patient prior to institution of therapy, during treatment and thereafter, as shown in Table 1. The samples were kept at - 70 C until use. Automated immunoprecipitation nephelometry was utilized to measure albumin and IgG concentrations in unconcentrated CSF and serum. O
Table 1. Effects of penicillin therapy on CSF variables in neurosyphilis. Patients No. 1-4 had definite active neurosyphilis (DANS), No. 5-8 had possibly active neurosyphilis (PANS), and No. 9-12 had possible neurosyphilis(PNS). Patient No. 1 and 2 had two courses(a and b)of penicilin treatment. TD = Tabes Dorsalis. DP = Dementia Paralytica. TP = Taboparesis. MVS = Meningovascular Syphilis. PK = Parkinsonism. Numbers within brackets denote upper reference values. ND = not done.
CSF indices for Patient No.
Diagnosis
1. a
DANS (TP)
Days after initiation of therapy Before 5
8 1. b
(5 months later)
2. a
DANS (DP)
2. b (10 months later)
3
DANS (TP)
4
DANS (TP)
5
PANS (TD)
6
PANS (TD)
7
PANS (TD)
8
PANS (MVS)
9
PNS (PK)
10
PNS (Stroke)
11 12
PNS (Epilepsy) PNS (Stroke)
Before 14 19 Before 23 Before 10 Before 6 10 31 months later Before 6 10 Before 23 Before 10 Before 21 Before 10 Before 7 Before 20 23 Before 6 Before 6
CSFiSerum albumin ratio
IgG (0.7)
IgM (0.06)
IgA (0.34)
IgD (0.50)
7.5 5.2 5.9 6.5 7.6 6.2 5.7 7.4 4.6 5.1 7.9 4.9 4.7 2.9
2.9 3.2 2.7 1.0 0.7 0.7 3.1 2.4 2.9 2.0 2.5 3.0 2,9 1.1
0.88 0.68 0.62 0.23 0.17 0.19 0.83 0.51 0.22 0.1 1 3.30 4.82 4.32 0.31
0.80 1.oo 1.07 0.46 0.54 0.46 0.41 0.36 0.31 0.22 0.54 0.50 0.48 ND
0.32 ND 0.32 0.33 ND 0.23 0.34 0.26 0.15 0.18 0.67 ND 1.04 ND
5.1 4.7 4.6 2.7 2.9 4.4 3.2 5.9 7.3 4.6 5.1 4.2 4.5 4.8 5.6 3.4 3.3 3.4 4.6 5.7
6.6 10,4 8.6 0.8 0.7 0.6 0.6 0.4 0.6 0.6 0.7 0.6 0.7 0.6 0.5 0.5 0.5 0.6 0.5 0.5
3.45 5.96 2.49 0.14 0.1 1 0.05 0.03 0.04 0.04 0.05 0.25 0.07 0.12 0.04 0.10 0.08 0.06 0.13 0.04 0.04
2.42 3.64 2.68 0.26 0.27 0.20 0.25 0.22 0.20 0.23 0.53 0.19 0.18 0.30 0.36 0.29 0.20 0.24 0.31 0.3 1
1.10 ND 1.33 0.22 0.21 0.05 0.1 1 0.00 0.00 0.26 0.35 0.17 0.22 0.09 ND 0.13 0.13 0.18 ND ND
'
' Upper reference levels fluctuate in relation to age, being 9.0 in all patients except No. 3, 9 and 10 where it is 7.2 (compare ref. 33). 382
Oligoclonal IgG bands in CSF
3 ND 3 2 ND 2 4 4 4 4
0 ND
0 ND
2 ND 2 3 0 0 0 0
0 0 0 0 0 0 ND 0
0 0 0 0
High-dose i.v.-penicillin in neurosyphilis
42-63 (64)
53-68 (58)
410
311
410
Clinical diagnosis Tabes dorsalis Dementia paralytica Taboparesis Meningovascular syphilis Stroke Late-onset epilepsy Parkinsonism
1 1 2 0 0 0 0
2
0 0
0 0 0
2 1 1
isoelectric focusing revealed 2-4 oligoclonal IgG bands restricted to CSF in 3 of these patients. Among the patients with PANS, only No. 5 hd increased IgG and IgM indices, and also oligoclonal IgG bands in CSF. In the group with PNS, borderline increase of the IgM index in Patient 9 was the only abnormality. Considering the CSF IgD index, Patients 3 and 4 (Table 1) with DANS had elevated values as a reflection of intrathecal IgD synthesis. Both had evidence of simultaneous intrathecal production of IgG, IgM and IgA; both were suffering from taboparesis and had in fact clinical evidence for the most severe and diffuse CNS involvement in our patient material, and also the highest IgM index values.
Laboratory abnormalities Positive serum FTA-abs’ CSF pleocytosis Range (per mm3) VDRL reactive CSF Range of titres
4 4 6-86 4 1/2-1/16
4 0 < -5 0 neg
4
Effects of therapy on immunoglobulin abnormalities
Table 2. Clinical and laboratory findings in 12 patients with neurosyphilis. DANS = Definite Active Neurosyphilis. PANS = Possibly Active Neurosyphilis. PNS = Possible Neurosyphilis. DANS
PANS
PNS
Number of patients
4
4
4
Age range (mean)
49-12 (58)
Sex (M/F)
1 0 1
0 0
0 < -5 0 neg
’ FTA-abs is equal to Fluorescence Treponemal Antibody absorption The blood-brain barrier was evaluated by determining the CSF/serum albumin ratio. Age-dependent reference values previously reported were used (33). To measure concentrations of IgM, IgA and IgD in CSF and serum, we used double antibody sandwich Enzyme Linked Immuno Sorbent Assay (ELISA) (34). The sera and CSF from individual patients were analyzed in parallel for protein levels. CSF IgG index equal to (CSF IgG/serum IgG) : (CSF albumin/serum albumin) was calculated (33). Our upper reference value is 0.7 and increased values are considered to reflect intrathecal IgG production. CSF indices for IgM, IgA and IgD were calculated as for IgG. Upper reference limits are 0.06 for IgM index, 0.34 for IgA index and 0.50 for IgD index, based on examination of patients with tension headache (34). Agarose isoelectric focusing followed by protein transfer to nitrocellulose membrane, immunostaining with double-antibody peroxidase labeling, and avidin-biotin amplification, was performed on unconcentrated CSF and diluted serum to detect oligoclonal IgG bands (35). Occurrence of two or more bands in CSF but not in corresponding serum, both run in parallel, was considered to reflect intrathecal synthesis of oligoclonal IgG. Results Pretreatment immunoglobulin abnormalities
All 4 patients with DANS had increased CSF IgG, IgM and IgA indices indicating intrathecal production of these immunoglobulins (Table 1). Agarose
Following initiation of therapy, DANS Patients 1,3, 4 displayed initial increase of the IgG index, Nos 3 and 4 had an initial elevation of the CSF IgM index and also of the IgD index, and Patients 1 and 4 of the IgA index; 8-23 days after initiation of therapy, when the final specimens were taken in Patients 1 , 2 and 4, most immunoglobulin index values had decreased, but in none of them had a previously elevated value become normalized. Patient 3 with DANS also examined 31 months after treatment, when he still displayed elevated IgG and IgM indices. IgA and IgD were not examined because of paucity of CSF. Patients 1, 2 and 4 had oligoclonal IgG bands in CSF, and the band patterns were unaffected 8-23 days after initiation of therapy. In the only patient with PANS (No. 5) who displayed evidence for intrathecal immunoglobulin synthesis in the form of elevated IgG and IgM indices, as well as oligoclonal IgG bands, both the IgG index and isoelectric focusing pattern normalized on re-examination about 3 weeks after initiation of treatment, while the IgM index was still elevated. However, PANS Patient 8 developed increased IgA and IgM indices following initiation of penicillin treatment, and an increase of the IgM index was similarly observed in 3 of the patients with PNS. DANS Patients 1 and 2 (Table 1) were treated again with high-dose penicillin 5 and 10 months, respectively, after the first course of therapy. Both had still elevated IgG and IgM index values as well as oligoclonal IgG bands, and one of them had also elevated IgA index. However, all values were lower when compared with those registered in relation to the first course of treatment. After a second course of penicillin treatment, both the IgG and IgM indices decreased further, but they were not fully normalized during the short follow-up periods. The IgA index remained elevated in one of these two patients. Only one of the 12 patients (No. 3, Table 1) 383
Hens et al. showed a mild derangement of the blood-brain barrier function as evaluated by the CSF/serum albumin ratio. This abnormality vanished early during therapy. Discussion
We have shown that penicillin treatment of patients with Definitive Active Neurosyphilis (DANS) may lead to further increase of intrathecal synthesis of IgG, IgA and IgM compared with pretreatment data. Furthermore, one patient with DANS had elevated IgG and IgM index values 31 months after treatment, reflecting persistence of the intrathecal B cell response. Taboparesis encountered in two patients was characterized by intrathecal production of IgG, IgM and IgD in parallel. These two patients also displayed the highest CSF IgM values which we registered. Elevated CSF IgG and IgM indices constituted in our series of patients the most sensitive markers for an intrathecal B cell response. Elevated values were present in some patients who otherwise lacked CSF abnormalities. Local synthesis of IgD as reflected by elevated IgD index was restricted to patients with the most severe clinical symptoms. Similar findings have been reported in other inflammatory nervous system disorders including bacterial meningitis and Lyme neuroborreliosis (23, 34). Intrathecal IgA production was also found to be common in our patients with neurosyphilis, unlike blood-brain barrier derangement which was evident in only one of the 12 patients. Its early correction under high-dose i.v. penicillin therapy is not an unexpected finding, provided that even i.m. penicillin schedules, which do not attain CSF treponemicidal concentrations, can normalize both CSF cell count and protein content (36-38). Our findings suggest that blood-brain barrier correction under therapy probably constitutes the earliest marker of CSF normalization, followed in time by a decrease in the intrathecal synthesis of immunoglobulins, a sequence of events already suggested by others (6, ll). Neurosyphilis, “the great imitator” (9), may present clinically in many ways, and seropositivity in individuals suffering from common neurological disorders as stroke or epilepsy may occasionally rise doubts regarding the significance of their serological findings. Four such patients in our series with normal CSF except a borderline increase of the CSF IgM index in one of them, were empirically submitted to penicillin therapy. Interestingly, two patients (No. 10 and 11) developed abnormal IgM index just while receiving treatment. The B cell response against T. pallidum starts with the synthesis of IgM antibodies (40),and intrathecal synthesis of previously undetected specific IgM antibodies has 384
been observed in response to therapy (41). Normal CSF has been reported in patients with neurosyphilis, and spirochetes have been identified in the CSF of patients with otherwise normal cell count and non-reactive VDRL (25, 42). The intrathecal synthesis of IgM, and its increase observed in our patients shortly after initiation of penicillin therapy, could be related to the lysis of dormant treponemes, followed by activation of the immune system and subsequent antibody production (2 l), in a similar way as has been described in cerebral cysticercosis during therapy with praziquantel (43). Documentation of appearance of intrathecal IgM production after treatment could be useful for differentiating symptomatic neurosyphilis, even when the clinical picture is atypical, from asymptomatic neurosyphilis (44). Our data documented that an intrathecal immune response may persist for nearly 3 years in patients whose neurosyphilis had strictly been treated according to current recommendations (24). Similar findings in a patient with clinically suspected neurosyphilis, do therefore not distinguish active disease from neurological sequelae in individuals who already had received adequate therapy in the past. Assuming that high-dose i.v. penicillin, such as used in this series, is curative, our findings appear in keeping with the concept of “CSF biologic scar” (1 1). However, in view of the fact that seropositive individuals with CNS disease but otherwise normal CSF show intrathecal synthesis of IgM just while receiving high-dosage penicillin, thereby indirectly suggesting that T. pallidum was present in their CNS, we would recommend repeated treatment and IgM index determinations in individuals suspected to have received inadequate therapy in the past or showing progressive neurological deterioration. Acknowledgements We thank Mr J Garcia Gutierrez for skilful technical assistance and Ms Yvonne Nilsson for excellent secretarial help. Dr M Hens was recipient of a grant from Margit and Folke Foundation. Dr F Lolli was recipient of a grant from the Italian Multiple Sclerosis Association (AISM).
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