Clinical and laboratory features of HTLV-I asymptomatic carriers and patients with HTLV-I-associated myelopathy/tropical spastic paraparesis from the Brazilian Amazon

ABSTRACT Clinical and laboratory parameters including blood and cerebrospinal fluid (CSF) neopterin were investigated in human-T-lymphotropic-virus-type-I associated-myelopathy/tropical-spastic-paraparesis-HAM/TSP and in HTLV-I carriers. HAM/TSP (n = 11, 2 males/9 females, median age = 48 years), recently diagnosed HTLV-I carriers (n = 21, 15 females/6 males, median age = 44 years), healthy individuals (n = 20, 10 males/10 females, median age = 34.6 years) from the Brazilian Amazon (Manaus, Amazonas State) were investigated. Neopterin was measured (IBL ELISA Neopterin, Germany) in serum samples of all the participants, in CSF of 9 HAM/TSP patients as well as in 6 carriers. In HAM/TSP patients, CSF cell counts, protein and glucose were measured, the Osame’s motor-disability-score/OMDS was determined, and brain/spinal cord magnetic-resonance-imaging (MRI) was performed. HAM/TSP patients had normal CSF glucose, leukocyte counts; and normal protein levels predominated. Brain-MRI showed white-matter lesions in 7 out of 11 HAM/TSP patients. OMDS varied from 2-8: 9 were able to walk, 2 were wheel-chair-users. The median serum neopterin concentration in HAM/TSP patients was 6.6 nmol/ L; min. 2.8- max. 12.5 nmol/ L); was lower in carriers (4.3 nmol/L; min. 2.7- max. 7.2 nmol/ L) as well as in healthy participants (4.7 nmol/ L; min. 2.7- max. 8.0 nmol/ L) (p < 0.05). CSF neopterin concentrations in HAM/TSP patients were higher than in serum samples, and higher compared to carriers (p < 0.05). Carriers had similar serum-CSF neopterin concentrations compared to healthy participants. Variable clinical and laboratory profiles were seen in HAM/TSP patients, however our results support the neopterin measurement as a potential biomarker of disease activity.

Standardisation of Western blotting to detect HTLV-1 antibodies synthesised in the central nervous system of HAM/TSP patients

Intrathecal synthesis of human T-lymphotropic virus type 1 (HTLV-1) antibodies (Abs) represents conclusive evidence of a specific immune response in the central nervous system of HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients. Western blotting (WB) for HTLV Abs in serum is a confirmatory test for HTLV-1 infection. The aim of this study was to standardise the Western blot to demonstrate the intrathecal pattern of Abs against HTLV-1 proteins in HAM/TSP patients. Paired cerebrospinal fluid (CSF) and serum samples were selected from 20 patients with definite HAM/TSP, 19 HTLV-1 seronegative patients and two HTLV-1 patients without definite HAM/TSP. The presence of reactive bands of greater intensity in the CSF compared to serum (or bands in only the CSF) indicated the intrathecal synthesis of anti-HTLV-1 Abs. All definite HAM/TSP patients presented with an intrathecal synthesis of anti-HTLV-1 Abs; these Abs were not detected in the control patients. The most frequent intrathecal targets of anti-HTLV-1 Abs were GD21, rgp46-I and p24 and, to a lesser extent, p19, p26, p28, p32, p36, p53 gp21 and gp46. The intrathecal immune response against env (GD21 and rgp46-I) and gag (p24) proteins represents the most important humoral pattern in HAM/TSP. This response may be used as a diagnostic marker, considering the frequent association of intrathecal anti-HTLV-1 Ab synthesis with HAM/TSP and the pathogenesis of this neurological disease.

In vivo fluctuation of Tax, Foxp3, CTLA-4, and GITR mRNA expression in CD4+CD25+ T cells of patients with human T-lymphotropic virus type 1-associated myelopathy

HTLV-1 Tax expression exerts an inhibitory effect on the Foxp3 transcription factor in CD4+CD25+ T-regulatory cells (Treg). For a better understanding of the role of Tax mRNA in the gene expression of cellular markers we measured Tax, Foxp3, CTLA-4, GITR, TGF-β, and IL-10 mRNA in Treg cells of 50 patients with human T-lymphotropic virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP; 27 women and 23 men; mean age: 56.7 years). The control group consisted of 23 non-infected subjects (12 women and 11 men) with a mean age of 51.3 years. Real-time PCR was used to measure mRNA of Tax proteins and several cellular markers of Treg function. Determinations revealed a high level of Tax mRNA in HAM/TSP (124.35 copies/100 CD4+CD25+ T cells). Foxp3, GITR, and CTLA-4 mRNA levels were lower in HAM/TSP patients (mean ± SD, 22.07 ± 0.78, 9.63 ± 0.36, and 4.54 ± 0.39, respectively) than in non-infected controls (47.15 ± 12.94, 22.14 ± 1.91, and 21.07 ± 2.31). Both groups had similar levels of TGF-β and IL-10. An inverse relationship was found between Tax levels and Foxp3, CTLA-4, and GITR levels. Conversely, there was a direct correlation between levels of Foxp3, GITR, and CTLA-4. Disease severity and evolution time did not correlate with Tax or Foxp3 levels. The present results suggest that Tax and Foxp3 mRNA vary with the same degree of disease severity in HAM/TSP patients. Tax fluctuations may affect CTLA-4 and GITR expression via the Foxp3 pathway, causing virus-induced dysfunction of CD4+CD25+ T cells in HAM/TSP patients.

Motor behavioral abnormalities and histopathological findings of Wistar rats inoculated with HTLV-1-infected MT2 cells

The objective of the present study was to describe motor behavioral changes in association with histopathological and hematological findings in Wistar rats inoculated intravenously with human T-cell lymphotropic virus type 1 (HTLV-1)-infected MT2 cells. Twenty-five 4-month-old male rats were inoculated with HTLV-1-infected MT2 cells and 13 control rats were inoculated with normal human lymphocytes. The behavior of the rats was observed before and 5, 10, 15, and 20 months after inoculation during a 30-min/rat testing time for 5 consecutive days. During each of 4 periods, a subset of rats was randomly chosen to be sacrificed in order to harvest the spinal cord for histopathological analysis and to obtain blood for serological and molecular studies. Behavioral analyses of the HTLV-1-inoculated rats showed a significant decrease of climbing, walking and freezing, and an increase of scratching, sniffing, biting, licking, and resting/sleeping. Two of the 25 HTLV-1-inoculated rats (8 percent) developed spastic paraparesis as a major behavioral change. The histopathological changes were few and mild, but in some cases there was diffuse lymphocyte infiltration. The minor and major behavioral changes occurred after 10-20 months of evolution. The long-term observation of Wistar rats inoculated with HTLV-1-infected MT2 cells showed major (spastic paraparesis) and minor motor abnormalities in association with the degree of HTLV-1-induced myelopathy.

Aislamiento del virus linfotrópico de células T humano tipo I de un paciente con paraparesia espástica tropical: primer reporte en Cuba / Isolation of human T cell lymphotropic virus type 1 from a patient affected by tropical spastic paraparesis: first report in Cuba

Introducción: la paraparesia espástica tropical o mielopatía asociada a HTLV-I (PET/MAH) es una afección neurológica crónica etiológicamente ligada al virus linfotrópico de células T humano tipo I (HTLV-I). Objetivo: realizar la confirmación serológica de la infección y el aislamiento del virus linfotrópico de células T humano tipo I, a partir de las células mononucleares de sangre periférica, de una paciente con un cuadro clínico compatible con paraparesia espástica tropical. Métodos: las células mononucleares de sangre periférica se cocultivaron durante 35 d. La detección viral se realizó por inmunofluorescencia específica de membrana y reacción en cadena de la polimerasa para los genes tax, pol y gag. Resultados: se confirmó la presencia de anticuerpos contra el virus linfotrópico de células T humano tipo I en el suero. Se observó un incremento del número de células positivas por inmunofluorescencia indirecta en el monitoreo secuencial y se detectó la presencia de ADN proviral en las células mononucleares de sangre periférica. Conclusiones: los resultados evidenciaron el aislamiento del virus y corroboraron, por primera vez en Cuba, la asociación entre el virus linfotrópico de células T humano tipo I y paraparesia espástica tropical.

Background: Tropical spastic paraparesis or HTLV-I-associated myelopathy (TSP/HAM) is a chronic neurological disease etiologically linked to human T-cell lymphotropic virus type I (HTLV-I). Objective: to serologically confirm infection by and isolation of HTLV-I from the peripheral blood mononuclear cells from a patient who presented with a clinical picture similar to that of tropical spastic paraparesis. Methods: peripheral blood mononuclear cells were co-cultured for 35 days. Viral detection by membrane- specific immunofluorescence and polymerase chain reaction of genes tax, pol and gag were performed. Results: The presence of antibody to human T cell lymphotropic virus type I in serum was confirmed. Indirect immunofluorescence in the sequential monitoring made it possible to observe an increase of the number of positive cells whereas proviral DNA was detected in the peripheral blood mononuclear cells. Conclusions: these results support the evidence of viral isolation and confirmed, for first time in Cuba, the association between HTLV-I and TSP.

Avaliação do desempenho de parâmetros imunológicos como indicadores de progressão clínica da infecção crônica pelo HTLV-1 / Evaluation of the performance of immunological parameters as indicators for clinical progression of chronic HTLV-1 infection

Neste estudo, foi avaliado o desempenho isolado e combinado de parâmetros laboratoriais, percentual de linfócitos B ( por centoLB), a razão entre células T/B e o por centoCD8+HLA-DR+/CD8+, na identificação de indivíduos assintomáticos-AS ou portadores de HAM/TSP-HT numa população de casos soropositivos para HTLV-1. índices expressos em porcentagem demonstram que cada parâmetro, isoladamente, apresenta desempenho moderado, com co-negatividade=83 por cento e 91 por cento para por centoLB e razão entre células T/B, respectivamente e co-positividade=78 por cento para por centoCD8+HLA-DR+/CD8+. A análise combinada ( por centoCD8+HLA-DR+/CD8+ e razão T/B) não revelou ganho significativo no desempenho (co-positividade=75 por cento, co-negatividade=74 por cento). A análise das razões de verossimilhança em diferentes faixas de valores, para os parâmetros isolados, revelou que um indivíduo soropositivo para HTLV-1 com por centoLB<7 por cento, razão entre células T/B>11 e por centoCD8+HLA-DR+/CD8+>70 por cento possui, respectivamente, 11, 19 e quase 10 vezes mais chances de pertencer ao grupo HT. Portanto, recomenda-se o uso desses indicadores fenótipos na propedêutica laboratorial complementar de monitoração da progressão clínica da infecção crônica pelo HTLV-1.

This study evaluated the performance of single and combined laboratory parameters, B-lymphocyte percentages ( percentLB), T/B cell ratio and percentCD8+HLA-DR+/CD8+, to differentiate asymptomatic cases (AS) from HAM/TSP patients (HT) within a population of HTLV-1 seropositive cases. Percentage indices demonstrated that each parameter alone presented moderate performance, with co-negativity of 83 and 91 percent for percentLB and T/B cell ratio, respectively, and co-positivity of 78 percent for percentCD8+HLA-DR+/CD8+. Combined analysis ( percentCD8+HLA-DR+/CD8+ and T/B cell ratio) did not show any substantial performance enhancement (co-positivity = 75 percent and co-negativity = 74 percent). Likelihood ratio analysis using different value ranges for the separate parameters revealed that HTLV-1 seropositive cases with percentLB<7 percent, T/B cell ratio>11 and percentCD8+HLA-DR+/CD8+>70 percent would have, respectively, 11, 19 and 10 times greater chance of belonging to the HT group. Therefore, use of these phenotypic indicators as complementary laboratory methods for monitoring the clinical progression of chronic HTLV-1 infection is recommended.

HTLV-I negative tropical spastic paraparesis: a scientific challenge

We reviewed the historical, clinical and etiological aspects of the progressive chronic spastic myelopathies of unknown etiology, disserting on the clinical similarities between HTLV-I seropositive and seronegative tropical spastic paraparesis (TSP), as well as focusing on the PCR studies of the seronegative TSP

Evaluation of serum interlekin - 2 Roc in H A M / TSP patients and HTLV - 1 carriers in Mashhad - Iran

Seropositive rate of human T-cell leukemia virus type 1 [HTLV-1] in Mashhad [a city in the north east of Iran] is 2.3% which is a new endemic area in the world. Less than 2% of these HTLV-1 carriers suffer from HTLV-1 associated myelopathy/tropical spastic paraparesis [HAM/TSP]. Twenty serum samples from HAM/TSP patients, 44 from HTLV-1 carriers and 20 from healthy individuals were tested for serum IL-2R[alpha] by ELISA method. The mean soluble interleukin-2 receptor alpha [sIL-2R[alpha] in HAM/TSP patients, HTLV-1 carriers and healthy subjects was 1420, 1460 and 374 pg/ml respectively. This study revealed that the level of sIL-2R[alpha] in HAM/TSP patients and HTLV -1 carriers was significantly more than healthy subjects [P<0.05], but no significant difference between HTLV-1 carriers and HAM/TSP patients was found

The value of csf analysis for the differential diagnosis of HTLV-I associated myelopathy and multiple sclerosis

Amostras do líquido cefalorraquidiano (LCR) e soro de 17 pacientes brasileiros com HAM/TSP, seis com esclerose múltipla e seis com epilepsia idiopática (controle nao-inflamatório) foram analisadas para a presença de anticorpos para o vírus de sarampo, rubéola, varicela zoster e herpes simples pelo método de ELISA. Todos os casos de HAM/TSP e esclerose múltipla tinham resposta imune poliespecífica intratecal para sarampo e rubéola. Anticorpos específicos para sarampo e rubéola (resposta MRZ) foram observados em todos os pacientes com esclerose múltipla, mas nao nos controles com epilepsia idiopática. A relevância das respostas poliespecífica e monoespecífica é discutida para essas doenças neurológicas crônicas.

Pulmonary involvement in patients with HTLV-I associated myelopathy

A frequência do comprometimento pulmonar em pacientes com doença neurológica associada ao primeiro vírus linfotrópico humano de células T(HTLV-I) tem sido demonstrado por vários autores na Africa, Asia e América Latina. Com o objetivo de estudar o envolvimento pulmonar em pacientes com mielopatia por HTLV-I (HAM) estudamos o lavado bronco-alveolar (LBA) de pacientes com HAM e 13 pacientes com mielopatias de outras etiologias. A contagem diferencial das células do LBA de pacientes com HAM demonstrou percentual de Linfócitos maior que 20 em 18 (82 por cento) dos pacientes enquanto aqueles com mielopatias de outra natureza mostraram linfocitose no LBA em apenas 2 (15 por cento). Concluímos que o pulmao se constitui em importante órgao para a patogênese de HAM.

HTLV-I associated myelopathy in Porto Alegre (Southern Brazil)

Mielotapia associada ao HTLV-I/paraparesia espástica tropical (TSP/HAM) tem sido descrita de forma crescente em praticamente todas as regioes do Brasil. Relatam-se cinco casos confirmados e documentados de pacientes com TSP/HAM no Rio Grande do Sul, observando-se em todos paraparesia espática, bexiga neutrogênica e distúrbios sensitivos superficiais e/ou profundos de graus variáveis. Em um caso denotou-se padrao em surto e remissao, caracterizado por ataxia cerebelar (sugestivo de esclerose múltipla). Os cinco pacientes foram submetidos a investigaçao clínica, laboratorial, urodinâmica, neurofisiológica e de neuroimagem. O objetivo de presente trabalho foi apresentar a regiao sul do Brasil como uma área de significativa endemicidade de infecçao por HTLV-I/II (0,42 por cento de prevalência entre doadores de sangue), bem como evidenciar a existência de pacientes com doença neurológica associada a este retrovírus.

Evidence of preferential female prevalence of HTLV-I associated tropical spastic paraparesis in Bahia, Brazil

Com o objetivo de avaliar a prevalência de infecçäo pelo HTLV-I e sua associaçäo a paraparesia espática tropical na Bahia, foram estudadas amostras de líquido céfalorraqueano (LCR) e/ou soro de pacientes com esta doença e de outros grupos de indivíduos. Os resultados indicam que mulheres säo mais afetadas pela doença associada à infecçäo pelo HTLV-I que os homens. Algumas alteraçöes do exame do LCR podem sugerir infecçäo pelo HTLV-I em pacientes com paraparesia espática tropical