Determinants of HIV-1 CD4-Independent Brain Adaptation.

BACKGROUND: HIV-1 is known to adapt to the local environment in its usage of receptors, and it can become CD4 independent in the brain where the receptor is scarce. This adaptation is through amino acid variations, but the patterns of such variation are not yet well understood. Given that infection of long-lived CD4-low and CD4-negative cells in anatomical compartments such as the brain expands cell tropism in vivo and may serve as potential viral reservoirs that pose challenge for HIV eradication, understanding the evolution to CD4 independence and envelope conformation associated with infection in the absence of CD4 will not only broaden our insights into HIV pathogenesis but may guide functional cure strategies as well. METHODS: We characterize, by site-directed mutagenesis, neutralization assay, and structural analysis, a pair of CD4-dependent (cl2) and CD4-independent (cl20) envelopes concurrently isolated from the cerebral spinal fluid of an SHIV-infected macaque with neurological AIDS and with minimum sequence differences. RESULTS: Residues different between cl2 and cl20 are mapped to the V1V2 and surrounding regions. Mutations of these residues in cl2 increased its CD4 independence in infection, and the effects are cumulative and likely structural. CONCLUSIONS: Our data suggested that the determinants of CD4 independence in vivo mapped principally to V1V2 of gp120 that can destabilize the apex of the envelope spike, with an additional change in V4 that abrogated a potential N-linked glycan to facilitate movement of the V1V2 domain and further expose the coreceptor-binding site.

Central memory CD4+ T cells dominate the normal cerebrospinal fluid.

BACKGROUND: To use cerebrospinal fluid (CSF) immune phenotyping as a diagnostic and research tool, we have set out to establish reference values of white blood cell (WBC) subsets in CSF. METHODS: We assessed the absolute numbers and percentages of WBC subsets by 6-color flow cytometry in paired CSF and blood samples of 84 individuals without neurological disease who underwent spinal anaesthesia for surgery. Leukocyte (i.e., lymphocytes, granulocytes, and monocytes), lymphocyte (i.e., T [CD4(+) and CD8(+) ], NK, NKT and B cells), T cell (i.e., naïve, central memory, effector memory, and regulatory) and dendritic cell subsets (i.e., myeloid and plasmacytoid) were studied. RESULTS: CSF showed a predominance of T cells, while granulocytes, B and NK cells were relatively rare compared to blood. The majority of T cells in CSF consisted of CD4(+) T cells (∼70%), most of them (∼90%) with a central memory phenotype, while B cells were almost absent (<1%). Among the small population of dendritic cells in CSF, those of the myeloid subtype were more frequent than plasmacytoid dendritic cells (medians: 1.7% and 0.4% of leukocytes, respectively), whilst both subsets made up 0.2% of leukocytes in blood. CONCLUSIONS: This study reports reference values of absolute numbers and percentages of WBC subsets in CSF, which are essential for further investigation of the immunopathogenesis of neuro-inflammatory diseases. Furthermore, the relative abundance of CD4(+) T cells, mainly with a central memory phenotype, and the presence of dendritic cells in CSF suggests an active adaptive immune response under normal conditions in the central nervous system (CNS).

Flow cytometric analysis of T cell subsets in paired samples of cerebrospinal fluid and peripheral blood from patients with neurological and psychiatric disorders.

Recent studies suggest inflammatory mechanisms involved in the pathogenesis of major psychiatric disorders (MPD). T cells play a major role during inflammation, but little is known about T cell subpopulations in the cerebrospinal fluid (CSF). We investigated the frequency of cells positive for the surface markers CD4, CD8, CD25, CD45, CD69, and CD127 in 45 paired cerebrospinal fluid (CSF) and peripheral blood (PB) samples by multiparameter flow cytometry from patients with MPD of the schizophrenic and affective spectrum with normal CSF cell counts and compared them with those from patients with non-inflammatory (NIND), chronic inflammatory (CIND) neurological disorders, and meningitis (MEN). In MEN patients, CD4+ cell frequency in PB, but not in CSF, was significantly increased as compared to CIND and NIND. No difference between patient groups was observed for CD8+. CD4+CD45RO+ double positive cells in PB were significantly lower in CIND than in MEN or NIND. The frequency of CD4+CD25+ cells in PB was significantly higher in MEN than in MPD or CIND. For CSF, the percentage of CD4+CD127(dim) cells was significantly lower in MEN than in MPD. CD4+CD127(dim) in PB and CSF showed overlapping characteristic clusters between MPD and CIND and MEN patients. Overall, the hypothesis of low degree inflammation in a subgroup of MPD is supported. The analysis of lymphocyte subsets in PB and CSF constitutes a novel promising tool to understand underlying pathomechanisms in psychiatric and neurological disorders on an individual case level.

[Soluble CD4 antigens in the cerebrospinal fluid as a marker of lymphocytic infiltration of the central nervous system]. / Solubilni CD4 antigeni u cerebrospinslnoj tecnosti kao marker limfocitne infiltracije centralnog nervnog sistema.

Early diagnosis of the central nervous system (CNS) infections is a precondition of their successful treatment. However, the essential standard examination of the cerebrospinal fluid (CSF) is sometimes neither specific enough to define their basic nature, nor sufficient to differentiate them from processes of non-infectious origin. Supposing that the released surface molecules of activated immunocompetent cells could better define the character of inflammatory reaction, the levels of soluble CD4 antigens (sCD4) were determined with enzyme-immunosorbent test in the CSF of the patients with various CNS diseases. In contrast to cerebrovascular insults, toxic-metabolic, and other conditions in control group, detectable sCD4 concentrations in acute encephalitis (24 +/- 11 U/ml) were verified at the beginning of the disease, being also present in cytologically diagnosed normal CSF findings. They were significantly higher (p<0.05) compared to acute serous meningitis (13.5 +/- 8 U/ml), while in purulent meningitis they were measurable only after the disease progression--in correlation with the disturbed brain system function. The obtained results suggested the significance of CD4 antigen levels in CSF as a sensitive and specific marker of lymphocytic infiltration of the brain parenchyma, the measurement of which could contribute to early identification of the CNS infections, better understanding of their pathogenesis, and the assessment of the actual level of the destruction of neurons.

Computerized reaction time battery versus a traditional neuropsychological battery: detecting HIV-related impairments.

In recent years, interest in the use of computerized neuropsychological (NP) assessment measures has increased. However, there are limited data regarding how performance on these measures relates to performance on more traditional, clinical instruments. In the present study, 82 HIV+ men, who were all believed on clinical grounds to have neurobehavioral impairment, completed a traditional NP battery (TNB) and the California Computerized Assessment Package (CalCAP, a collection of computerized reaction time tests). Summary scores based on a TNB, as well as those based on the CalCAP, demonstrated significant associations with both degree of immunosuppression (CD4 count) and detectable viral load in cerebrospinal fluid, but not with detectable viral load in plasma. Established norms on the TNB and CalCAP batteries resulted in classifying 57% and 49% of the HIV+ sample as impaired, respectively. When using the TNB as the "gold standard," impairment classifications based on CalCAP summary scores exhibited a sensitivity of 68% and a specificity of 77%. Overall agreement on impairment classifications between batteries was low (kappa = .44). Data from this study suggest that traditional NP batteries and computerized reaction time tests do not measure the same thing, and are not interchangeable in examining HIV-related NP impairments.

[Immune events in central nervous system of early and late onset Alzheimer's disease patients]. / Eventos inmunológicos en el sistema nervioso central de pacientes con enfermedad de Alzheimer temprana y tardía.

INTRODUCTION: Alzheimer s disease (AD) is a progressive degenerative disease affecting a significant proportion of the elderly population. The disease is characterized clinically by a progressive loss of memory function and mental impairment associated with the presence of degenerative well known pathological lesions. Although, the pathogenesis of AD is unclear; several reports indicate the involvement of immune factors. PATIENTS AND METHODS: This paper evaluates some cerebrospinal fluid immune markers from 21 patients with early and late AD and 20 age matched non-demented subjects. The analytical method included the evaluation of T cell subpopulations (using AcMc CD2, CD4, CD8) and activated T cells (AcMc HLA-DR and CD25) from CSF and peripheral blood by immunocytochemical techniques on a fixed cell slide as described by Bernd. The lymphocyte phenotype expressed as a percentage of positively stained cells for each cell surface marker evaluated. RESULTS: Some significant differences were observed for T cell subpopulations from different compartments, between the different AD groups and the controls (p< 0.05). Nevertheless, the most significant differences were found in the activated T cells from cerebrospinal fluid between AD groups and controls (p< 0.01). CONCLUSIONS: These results support the theory of neuroimmune dysregulation, probably involved in the progressive neurodegeneration and dementia in some AD.

Leukocyte trafficking in free-flowing cerebrospinal fluid of normal rhesus macaques (Macaca mulatta).

Cellular components in free-flowing cerebrospinal fluid (CSF) of normal rhesus macaques were characterized. Microscopic counting enumerated the total number of leukocytes, percentage of polymorphonuclear cells (PMN), leukocytes with nonspecific esterase (NSE), and those reducing nitro blue tetrazolium (NBT). Flow cytometric analysis further identified CD4, CD8, CD14, and CD20 positive leukocytes. These experiments established reliable techniques for evaluating cellular components in CSF from rhesus macaques and documented the difference in the CD4/CD8 ratio between peripheral blood (PB) and CSF compartments under normal physiological conditions.

Significance of elevated levels of soluble factors in the cerebrospinal fluid in patients with adult T-cell leukemia.

Although it has been recognized previously that several markers are present in the cerebrospinal fluid (CSF), their clinical usefulness of these markers in the diagnosis of malignant lymphoma infiltrating to the CNS has not yet been established. In order to determine their diagnostic usefulness as markers of meningeal infiltration by lymphoma cells in patients with adult T-cell leukemia (ATL), we measured some soluble factors in the CSF of patients with ATL and non-ATL patients. Soluble CD4 (sCD4) was highly elevated in all patients with ATL and meningeal infiltration. The CSF level of the soluble interleukin-2 receptor (sIL-2R; sCD25) was markedly elevated in 13 (72.2%) of 18 patients with ATL and meningeal infiltration. Levels of sCD4 and sCD25 in the CSF of patients with ATL and meningeal infiltration were significantly higher than in non-ATL patients (p < .01 and p < .001, respectively). These findings indicate that levels of sCD4 and sCD25 in the CSF are probably associated with meningeal infiltration by leukemia cells expressing CD4 and CD25 on surface membranes. CSF levels of sCD4 in 14 (60.9%) of 23 ATL patients and sCD25 in 13 (72.2%) of 18 ATL patients without meningeal infiltration were moderately elevated. These findings suggest that a small number of leukemic cells which were not detected by conventional CSF examination may have infiltrated the meninges in these patients. Sequential measurements of sCD4 and sCD25 in CSF obtained from patients with meningeal infiltration by leukemic cells showed that sCD4 and sCD25 levels reflected the activity of leukemic meningitis and correlated with the number of cells in CSF. However, the levels of sCD4 in CSF did not fall below the limit of detection even when the number of cells in CSF became normal. It is thought that the level of sCD4 in CSF is a more sensitive marker for detecting the infiltration of leukemic cells in CSF than the number of cells present in the CSF considering the clinical course of two patients with acute type ATL. Therefore, ATL patients with meningeal infiltration should receive treatments until sCD4 levels become normal and not just until the number of cells become normal. Our results also suggest that measurement of CSF levels of sCD4 and sCD25 is useful for the differential diagnosis of aseptic meningitis and meningeal infiltration by leukemic cells in patients with smoldering ATL. We conclude that measurement of soluble factors in CSF plays an important role in diagnosis, prophylaxis and treatment of meningitis in patients with ATL.

T cell subsets in peripheral blood and cerebrospinal fluid from children with aseptic meningitis.

T cell subsets in peripheral blood (PB) and cerebrospinal fluid (CSF) obtained from patients with aseptic meningitis were studied using quantitative two-color fluorescence analysis with a flow cytometer. The percentage of HLA-DR+/CD3+ lymphocytes (activated T cells) in CSF was significantly increased in the recovery phase when compared to the acute phase, while no significant change in the activated T cells in PB was observed. More interestingly, CD4+ T lymphocytes in CSF were increased in the acute phase and subsequently decreased in the recovery phase. Instead, CD8+ T lymphocytes gradually accumulated into the CSF in the recovery phase, resulting in a successive decrease in the CD4/CD8 ratio. On the other hand, the CD4/CD8 ratio in PB remained normal during the course of aseptic meningitis. The present results suggest that T lymphocytes (CD4+ subset in the acute phase and CD8+ in the recovery phase) could be infiltrated and further activated at the site of inflammation, possibly in the subarachnoid space in the patients with aseptic meningitis.

Elevated levels of soluble factors in the cerebrospinal fluid in patients with adult T-cell leukaemia complicated with meningeal infiltration.

We measured some soluble factors in the cerebrospinal fluid (CSF) of patients with adult T-cell leukaemia (ATL) complicated with meningeal infiltration. Interleukin-4 (IL-4) was not detectable in the CSF of all cases with meningeal infiltration. Interleukin-6 (IL-6) was detected in a few patients with ATL. Measurement of IL-4 and IL-6 in CSF had no diagnostic value for meningeal infiltration of ATL cells. Soluble CD4 (sCD4) was highly elevated in all ATL patients with meningeal infiltration. Soluble interleukin-2 receptor (sIL-2R; sCD25) in CSF was markedly elevated in 13/18 ATL patients (72.2%) with meningeal infiltration. Levels of sCD4 and sCD25 in the CSF of ATL patients with central nervous system (CNS) symptoms were significantly higher than those of non-ATL patients with CNS symptoms. These observations indicate that sCD4 and sCD25 in the CSF are probably associated with meningeal infiltration of leukaemia cells that expressed CD4 and CD25 on the surface membrane and new markers for the meningeal infiltration of ATL cells.

Elevated soluble CD4 levels in the cerebrospinal fluid in patients with adult T-cell leukemia.

Levels of the soluble form of the leukocyte surface antigen CD4 (sCD4) were measured by enzyme linked immunosorbent assay (ELISA) in the cerebrospinal fluid (CSF) of patients with adult T-cell leukemia (ATL) and other malignant and non-malignant diseases. All patients with ATL and meningeal infiltration had markedly elevated levels of sCD4 in the CSF (53.7 +/- 34.9 U/ml). ATL patients without CSF pleocytosis often had elevated levels of sCD4 (15.1 +/- 9.2 U/ml). Non-ATL patients with CSF pleocytosis had elevated levels of sCD4 (23.3 +/- 12.2 U/ml) and those without CSF pleocytosis also showed elevation of sCD4 levels (16.8 +/- 9.3 U/ml). However, the mean levels of sCD4 in CSF from these patients were significantly lower than ATL patients with meningeal infiltration. Soluble CD4 in the CSF from healthy volunteers were below the detectable limit. We conclude that meningeal infiltration of CD4(+) ATL cells is strongly associated with elevated sCD4 levels in CSF, and some part of sCD4 in CSF may be originated from the native cells in the CNS as a response of inflammatory stimulations. Therefore, measurement of sCD4 may be useful in the diagnosis of meningeal infiltration and/or meningeal irritation in patients with ATL.