Lack of association between parenchymal neurocysticercosis and HLA Class I and Class II antigens

Abstracts

Neurocysticercosis, caused by encysted larvae of the tapeworm Taenia solium, is the most common infection of the central nervous system and a major public health problem in many countries. Prevalence in the region of Curitiba, located in the southern Brazilian State of Paraná, is one of the highest in the world. The genetics of host susceptibility to neurocysticercosis (NCC) is still obscure. To investigate if major histocompatibility complex (MHC) genes influence individual susceptibility to NCC, we performed a case-control association analysis. Fifty-two Caucasoid patients and 149 matched controls were typed for antigens of the HLA-A, B, C, DR and DQ loci. All patients had computerized tomography and clinical features compatible with parenchymal NCC. Indirect immunofluorescence of cerebrospinal fluid showed that 19 (37%) of the patients presented anti-cysticercus antibodies at titers <FONT FACE="Symbol">³</font> 1:10. Frequencies of HLA specificities in the whole group of patients and in the subgroup with antibodies in cerebrospinal fluid were compared to those of the control group. No significant difference was found. These results do not support the hypothesis of HLA gene participation in susceptibility to parenchymal neurocysticercosis.

A neurocisticercose, causada pelo cisticerco, a larva do cestóide Taenia solium, é a infecção mais comum do sistema nervoso central e constitui importante problema de saúde pública em muitos países. A sua prevalência na região de Curitiba, localizada no Estado do Paraná, foi estimada em 9%, situando-se entre as mais elevadas do mundo. Os aspectos genéticos de suscetibilidade à neurocisticercose (NCC) ainda são pouco conhecidos. Com o objetivo de investigar se genes do MHC influenciam a suscetibilidade individual à NCC, realizamos uma análise de associação caso-controle. Cinqüenta e dois pacientes caucasóides e 149 indivíduos-controle pareados foram tipados para antígenos dos locos HLA-A, B, C, DR e DQ. Todos os pacientes apresentavam tomografia computadorizada e sinais clínicos compatíveis com NCC parenquimatosa. Imunofluorescência indireta do líquido céfalo-raquidiano mostrou que 19 (37%) pacientes apresentavam anticorpos anti-cisticerco com títulos <FONT FACE="Symbol">³</font>1:10. As freqüências de especificidades HLA no grupo total de pacientes e no subgrupo de pacientes que apresentaram anticorpos no líquor foram comparadas àquelas do grupo controle. Nenhuma diferença significativa foi detectada. Esses resultados não sustentam a hipótese de participação dos genes HLA na suscetibilidade à neurocisticercose parenquimatosa.

Lack of association between parenchymal neurocysticercosis and HLA Class I and Class II antigens

Eni Picchioni Bompeixe, Sonia Maria Correia Machado da Costa, Walter Oleschko Arruda and Maria Luiza Petzl-Erler

Departamento de Genética, UFPR, Caixa Postal 19071, 81531-990 Curitiba, PR, Brasil. Send correspondence to M.L.P.-E. Fax: [phone]. E-mail: [e-mail]

ABSTRACT

Neurocysticercosis, caused by encysted larvae of the tapeworm Taenia solium, is the most common infection of the central nervous system and a major public health problem in many countries. Prevalence in the region of Curitiba, located in the southern Brazilian State of Paraná, is one of the highest in the world. The genetics of host susceptibility to neurocysticercosis (NCC) is still obscure. To investigate if major histocompatibility complex (MHC) genes influence individual susceptibility to NCC, we performed a case-control association analysis. Fifty-two Caucasoid patients and 149 matched controls were typed for antigens of the HLA-A, B, C, DR and DQ loci. All patients had computerized tomography and clinical features compatible with parenchymal NCC. Indirect immunofluorescence of cerebrospinal fluid showed that 19 (37%) of the patients presented anti-cysticercus antibodies at titers ³ 1:10. Frequencies of HLA specificities in the whole group of patients and in the subgroup with antibodies in cerebrospinal fluid were compared to those of the control group. No significant difference was found. These results do not support the hypothesis of HLA gene participation in susceptibility to parenchymal neurocysticercosis.

INTRODUCTION

Cysticercosis, an infection caused by the larva of Taenia solium, is known as neurocysticercosis (NCC) when the cysticercus is lodged in the central nervous system. NCC is the most common central nervous system infection in developing countries as well as several developed nations. It is endemic in many parts of the world, including some regions in Latin America. In the Mexican population, autopsy studies have reported a prevalence of 3.8% in endemic areas (del Brutto and Sotelo, 1988). In Brazil, prevalence varies from one region to another, being 0.12%-9% in autopsies (Agapejev, 1996). Prevalence is highest in the south and the southeast. In the region of Curitiba, the main city of the southern Brazilian State of Paraná, it was estimated to be as high as 9.2% in 1993, based on computerized tomography (CT) (Antoniuk, 1994). This rate is among the highest in the world and, therefore, NCC is an important public health problem for this population.

The central nervous system is one of the major target organs of the cysticercus. The parasite may lodge in the parenchyma, subarachnoid space, ventricular system or spinal cord. Infection may remain asymptomatic or result in an extremely varied disease. The most common symptoms reported by patients are headache, convulsions, vomiting and behavioral alterations (Agapejev, 1996). Diagnosis, treatment and prognosis vary greatly according to the location, number and stage of the lesions. Pathogenesis and heterogeneous clinical outcome may be consequences of action of host and parasite variables (both genetic and non-genetic) and their interaction. Individual reactions against the parasite are very diverse. Development of lesions and the consequent outbreak of symptoms are mainly due to the host inflammatory response (Pittella, 1997). In some patients, there is evidence of an inflammatory reaction around the cysts, which eventually are destroyed. In other patients, the cysts may persist unchanged for many years (Dixon and Lipscomb, 1961; Noboa et al., 1988). Anti-cysticercus antibodies (mainly IgG) may be found in the serum and cerebrospinal fluid (CSF) of almost all patients with NCC (Corona et al., 1986).

Host variables, which may influence the course of infection, have been poorly investigated. It is unclear if host genes alter human susceptibility to the infection, or the form and severity of the disease. In mice, major histocompatibility complex (MHC) genes (or genes closely linked to this complex) were found to greatly influence growth of Taenia crassiceps. Further, gender was shown to play a role in susceptibility: female mice were significantly more susceptible than males (Sciutto et al., 1991). Human leukocyte antigens (HLA) were found on the surface of most damaged cysticerci in patients, but not on undamaged ones (Correa et al., 1986; Trejo et al., 1989). This led to the proposition that HLA molecules, when present on the cyst, are involved in host protection phenomena rather than in parasite evasion strategies (Correa et al., 1986). Significant associations between NCC and HLA antigens have been reported in a Mexican mestizo population (del Brutto et al. 1991). There are no studies regarding a possible correlation between NCC and HLA diversity in other populations. In the search for host factors contributing to the pathogenesis of NCC, and considering previous reports relating MHC genes to the disease, we tested the hypothesis of an association between HLA markers and parenchymal NCC in a Brazilian population from a high-incidence area in the State of Paraná.

MATERIAL AND METHODS

Patients and controls

Peripheral blood lymphocytes were obtained from 52 patients and 149 unrelated controls. All individuals were attended at the Federal University Hospital in Curitiba, Paraná. Controls were carefully matched to patients on the basis of ethnic background, gender and socioeconomic parameters. Fifty-nine controls were surveyed during the same period as the patients; the remaining 90 individuals had been analyzed previously in the same laboratory (Tsuneto et al., 1989; Petzl-Erler et al., 1991). All individuals enrolled in this study were Caucasoids residing in southern Brazil, mainly in the State of Paraná (81% of the patients and 93% of the controls). They were of low socioeconomic condition and most (84 and 85%, respectively) were rural workers.

Diagnosis was based on clinical criteria and CT. All 52 patients presented CT compatible with parenchymal NCC. Indirect immunofluorescence (IF) of CSF (Machado et al., 1973) showed that 19 patients were positive, presenting anti-cysticercus antibodies at titers ³ 1:10. According to the classification of van As and Joubert (1991), 8 of the patients presented the active form, 16 the inactive form, and 28 the mixed form. Clinical examination demonstrated that 46 patients had epilepsy, 26 complained of headache, 10 presented intracranial hypertension and 5 had other symptoms.

Male to female ratio was 1:1 (26 males and 26 females), which agrees with that reported for NCC by other authors (Trejo et al., 1989). Mean patient age was 32.4 ± 13.7 (range: 5 to 68 years) and, for the controls, it was 40.1 ± 25.8 (range: 19 to 70 years).

HLA typing

Class I typing was performed on T cells and class II typing on B cells obtained from heparinized venous blood. Standard microlymphocytotoxicity using trypan blue as the exclusion dye was used (Amos, 1976). The serum panel consisted of 238 well-defined antisera, including local reagents as well as those from other laboratories. Many were validated in the 9th and 10th International Histocompatibility Workshops. These antisera permitted identification of 50 HLA-A, -B and -C, 13 HLA-DR, and 3 HLA-DQ specificities.

Statistical analysis

Antigen frequencies were obtained by dividing the number of individuals positive for each given specificity by the total number of patients (or controls) typed. To check for associations between the disease and HLA, antigen frequencies of patients and controls were compared in a 2 x 2 contingency table for each HLA specificity. Significance of the differences was estimated by Fisher's exact test.

RESULTS

The comparison of antigen frequencies (Table I) of the control and patient groups, and between the same controls and IF-positive patients by Fisher's exact test showed no significant difference at the 5% probability level. Thus, the data do not support the hypothesis of association between HLA and NCC or presence of antibodies to the cysticercus in CSF.

A separate analysis of the patient subgroup with negative serology was not performed, since this is a heterogeneous group which, in addition to individuals who did not generate an antibody response, may include individuals with a low antibody titer as well as those who underwent seroconversion to negative (Garcia et al., 1997).

The antigen frequencies given (Table I) can be used as reference for the Caucasoid population of the State of Paraná, since there are no published reports on HLA class I and II antigen frequencies of this population.

DISCUSSION

The results of this study do not sustain the hypothesis of MHC diversity participating in the variation of susceptibility to parenchymal neurocysticercosis. Evidently, as discussed below, this does not exclude a role of the tested MHC molecules in the immune response to the cysticercus. However, individuals with different HLA molecules do not seem to differ in their capability to escape disease.

Although in mice MHC genes were found to influence the growth of Taenia crassiceps (Sciutto et al., 1991), both the host and the parasite species were different in that experimental model, and the location of the parasite was the peritoneal cavity. Further, human intra-specific genetic diversity is much different than that between congenic laboratory mice differing just in their MHC. In addition, as Sorci et al. (1997) pointed out, the finding of genetic variability of susceptibility to parasites in homogeneous environments (such as those created in the laboratory) does not necessarily imply that the same amount of genetic variation is actually expressed in variable environments. Heritability estimates of various phenotypic traits are higher when measured in the laboratory as compared to estimates from the field for the same species. Results obtained from studies of a single host strain may not apply to others, and extrapolation to different host species may be particularly difficult.

A positive association between parenchymal NCC and specificity A28, and a negative association with DQ2, has been reported for a Mexican population (del Brutto et al., 1991). It should be considered that antigens A28 and DQ2 are, on average, respectively more frequent and less frequent among Amerindians (which contributed much to the Mexican mestizo population) than in Europeans (Kostyu and Amos, 1981; Petzl-Erler et al., 1997), which contributed most to our population's gene pool. Population stratification and sampling differences may explain the apparently conflicting results of the previous (del Brutto et al., 1991) and the present studies. Thus, for instance, if factors other than HLA but associated to certain HLA alleles (or phenotypes) modulate susceptibility to NCC, and if these factors are unevenly distributed in the different ethnic groups, a secondary association with HLA might result from the analysis of stratified admixed populations, even if patients and controls seemingly are well matched. Furthermore, a spurious association may result if the control and patient groups are not perfectly matched for ethnic origin. An alternative explanation for the discrepant results, which considers the strong linkage disequilibrium among MHC genes, is that the susceptibility gene, whose alleles control susceptibility/resistance to NCC, has not been identified, and that alleles associated with increased and decreased susceptibility to NCC are associated with A28 and DQ2 in Mexicans but not Brazilians. In this case, however, rather than finding an absence of associations, we would expect the disease to be correlated with different HLA antigens in our population.

The possibility that high-resolution HLA typing at the DNA level would reveal any association between HLA and NCC is unlikely. Although this cannot be excluded, we consider this possibility remote based on known HLA-disease associations and the relationship between HLA serological specificities and HLA alleles in populations (compared with this relationship in the species as a whole). Thus, the known associations are significant when HLA typing is performed at both the serological (or low resolution DNA) and high-resolution DNA levels, even if only one or some of the alleles encoding the HLA antigens are associated with the disease. Examples are the positive associations between ankylosing spondylites and HLA-B27 (López-Larrea et al., 1995; Thorsby, 1997), insulin-dependent (type 1) diabetes mellitus and DQ8 and DQ2 (Thorsby 1997), endemic pemphigus foliaceus and DR1 (Petzl-Erler and Santamaria, 1989; Moraes et al., 1997), among many others. This is not unexpected if population genetics is considered. The many alleles corresponding to some of the HLA antigens occur in the human species as a whole, and are unevenly distributed among populations. The majority of the alleles described are extremely rare. These rare alleles have been detected in only one or a few individuals of a population. Each population bears just some of the alleles encoding a given antigen, and just one or a few of these occur at a reasonably high frequency. In particular, antigens A28 and DQ2, which frequencies have been reported to be respectively increased and decreased in Mexican NCC patients, present a restricted genetic diversity in the human species as a whole (Bodmer et al., 1997), and only one allele predominates, even in populations of admixed ethnic background like the Brazilian and Mexican populations (Petzl-Erler et al. 1997). If an association between these HLA variants and NCC occurred in the population analyzed in the present work, it would have been easily detected in this study.

Another aspect to consider is that Taenia solium intra-specific genetic variability evidently exists and is geographically stratified (McManus et al., 1989). However, T. solium diversity remains largely unknown, as is the case for most parasites. A better knowledge of parasitic variability would facilitate the search for interactions of host and parasite factors related to disease. Without detailed knowledge of host and parasite genetics, the outcome of their interaction is difficult to predict.

The lack of association between NCC and HLA reported here is not surprising if the antigenic complexity of the parasite and the known antigen presenting function of MHC class I and class II molecules are considered. This function implies an important role of these molecules in the immune response to parasites, including Taenia sp., but does not necessarily result in associations between HLA variants and disease outcome. Due to the extreme antigenic complexity of parasites, especially metazoan parasites, and the promiscuity of peptide-MHC molecule interaction, many HLA molecule variants are expected to present immunogenic peptides of the parasite to T cells. The products of individual HLA alleles restrict the response to defined sets of peptides (Matsumara et al., 1992), not to complex parasites. This could be the reason why no convincing associations between HLA alleles and infectious diseases have been found. Whenever associations between HLA antigens and infectious diseases have been reported, they occur with a certain form or manifestation of the disease, which most likely depends on an immune response to particular parasite antigens (e.g., the mucocutaneous form of leishmaniasis (Petzl-Erler et al., 1991) and the severe form of Plasmodium falciparum malaria (Hill et al., 1991)). In this study, only patients presenting parenchymal NCC were investigated, and the absence of association indicates that parenchymal NCC most likely is not an immunogenetic distinct form of the disease. Little is known about the specific immune response to the cysticercus. A better delineation of the spectrum of immune responses to Taenia solium and its larva is needed to evaluate the possible role of the diversity of MHC (and other) genes on the course of the infection and disease. If individuals differ in their degree of resistance against this infection and the immunopathology of cysticercosis turns out to be heterogeneous, a new search for genes controlling this diversity should be performed.

ACKNOWLEDGMENTS

We thank Dr. Bento Arce-Gomez, Maria da Graça Bicalho de Lacerda, and Noemi Farah Pereira for valuable comments and suggestions. We are grateful to the medical doctors from the Departments of Neurosurgery and Neurology of the Hospital de Clínicas, Universidade Federal do Paraná, for their essential assistance. This work was supported by CNPq.

RESUMO

A neurocisticercose, causada pelo cisticerco, a larva do cestóide Taenia solium, é a infecção mais comum do sistema nervoso central e constitui importante problema de saúde pública em muitos países. A sua prevalência na região de Curitiba, localizada no Estado do Paraná, foi estimada em 9%, situando-se entre as mais elevadas do mundo. Os aspectos genéticos de suscetibilidade à neurocisticercose (NCC) ainda são pouco conhecidos. Com o objetivo de investigar se genes do MHC influenciam a suscetibilidade individual à NCC, realizamos uma análise de associação caso-controle. Cinqüenta e dois pacientes caucasóides e 149 indivíduos-controle pareados foram tipados para antígenos dos locos HLA-A, B, C, DR e DQ. Todos os pacientes apresentavam tomografia computadorizada e sinais clínicos compatíveis com NCC parenquimatosa. Imunofluorescência indireta do líquido céfalo-raquidiano mostrou que 19 (37%) pacientes apresentavam anticorpos anti-cisticerco com títulos ³1:10. As freqüências de especificidades HLA no grupo total de pacientes e no subgrupo de pacientes que apresentaram anticorpos no líquor foram comparadas àquelas do grupo controle. Nenhuma diferença significativa foi detectada. Esses resultados não sustentam a hipótese de participação dos genes HLA na suscetibilidade à neurocisticercose parenquimatosa.

(Received March 5, 1998)

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Publication Dates

• Publication in this collection
  02 June 1999
• Date of issue
  Mar 1999

History

• Received
  05 Mar 1998