Central nervous system infections in a tropical area: influence of emerging and rare infections.

BACKGROUND AND PURPOSE: The frequency of infectious encephalitis and the distribution of causative pathogens in tropical areas are poorly known and may be influenced by emerging and rare infections. The aim was to characterize a large series of acute infectious encephalitis and myelitis in immunocompetent patients from the Caribbean island of Guadeloupe identifying clinical, biological and radiological features according to pathogens. METHODS: Using a hospital database, detailed information on a comprehensive series of immunocompetent patients with acute infectious myelitis and encephalitis over the 2012-2018 period was retrospectively collected. RESULTS: From 259 suspected cases with acute central nervous system infection, 171 cases were included for analysis, comprising 141 encephalitis, 22 myelitis and eight encephalomyelitis. The annual incidence peaked at 15.0/100 000 during the Zika 2016 outbreak. Children accounted for 22.2% of cases. Eight adults died during hospital stay, all encephalitis. Seventeen infectious agents, two of which had never been described in Guadeloupe so far, were identified in 101 cases (59.1%), including 35 confirmed cases (34.7%), 48 probable cases (47.5%), 15 possible cases (14.9%) and three clinical cases (3.0%). The most frequent etiologic agents were Zika virus in 23 cases (13.5%), herpes simplex in 12 (7.0%), varicella zoster virus in 11 (6.4%), dengue virus in 11 (6.4%) and leptospirosis in 11 (6.4%). CONCLUSIONS: The Zika outbreak had a major influence on the annual incidence of acute central nervous system infection. Acute neuroleptospirosis is over-represented in our series. Further efforts are mandatory to develop new diagnostic tools for pathogen profiling.

A role for CXCR4 signaling in survival and migration of neural and oligodendrocyte precursors.

Oligodendrocyte development is controlled by a number of survival and migratory factors. The present study shows that signaling of CXCR4 receptor by the chemokine CXCL12 regulates survival and migration of neural precursors (NP) as well as oligodendrocyte progenitors (OP). CXCR4 is expressed by E14 striatal NP and OP generated by neurospheres. In CXCR4-defective mice, the number of NP in neurosphere outgrowth was twofold less than in wild-type (WT) mice; NP radial cell migration was also decreased. In contrast, the addition of CXCL12 to WT NP increased radial migration from the sphere in a dose-dependent manner with a maximal response at 200 nM. When oligodendrocytes differentiated in neurosphere outgrowth, CXCR4 was downregulated. OP isolated from newborn brain coexpressed CXCR4 with platelet-derived growth factor receptor-alpha (PDGFR alpha) or chondroitin sulfate proteoglycan; receptor expression also decreased during differentiation in vitro. Neonatal OP showed a peak migratory response to 20 nM of CXCL12 in chemotactic chambers, a migration inhibited by a CXCR4 antagonist and anti-CXCL12 antibody. In the embryonic spinal cord, the number of OP-expressing PDGFR alpha was reduced more than twofold in CXCR4-defective mice compared with WT and the ratio of ventral to dorsal OP was significantly increased. This indicates a defect in OP survival and their dorsal migration from the ventral cord region, probably because CXCR4(-/-) OP are unable to respond to CXCL12 made by vascular endothelia and the pia mater. We propose that CXCR4 signaling regulate survival and outward chemotactic migration of OP during embryonic and postnatal CNS development.

Multiple sclerosis-associated retrovirus particles cause T lymphocyte-dependent death with brain hemorrhage in humanized SCID mice model.

A retroviral element (multiple sclerosis-associated retrovirus, MSRV) defining a family of genetically inherited endogenous retroviruses (human endogenous retrovirus type W, HERV-W) has been characterized in cell cultures from patients with multiple sclerosis. Recently, MSRV retroviral particles or the envelope recombinant protein were shown to display superantigen activity in vitro, but no animal model has yet been set up for studying the pathogenicity of this retrovirus. In the present study, the pathogenicity of different sources of MSRV retroviral particles has been evaluated in a hybrid animal model: severe combined immunodeficiency (SCID) mice grafted with human lymphocytes and injected intraperitoneally with MSRV virion or mock controls. MSRV-injected mice presented with acute neurological symptoms and died within 5 to 10 days post injection. Necropsy revealed disseminated and major brain hemorrhages, whereas control animals did not show abnormalities (P <.001). In ill animals, reverse transcriptase-polymerase chain reaction (RT-PCR) analyses showed circulating MSRV RNA in serum, whereas overexpression of proinflammatory cytokines such as tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma was evidenced in spleen RNA. Neuropathological examination confirmed that hemorrhages occurred prior to death in multifocal areas of brain parenchyma and meninges. Further series addressed the question of immune-mediated pathogenicity, by inoculating virion to SCID mice grafted with total and T lymphocyte-depleted cells in parallel: dramatic and statistically significant reduction in the number of affected mice was observed in T-depleted series (P <.001). This in vivo study suggests that MSRV retroviral particles from MS cultures have potent immunopathogenic properties mediated by T cells compatible with the previously reported superantigen activity in vitro, which appear to be mediated by an overexpression of proinflammatory cytokines.

Developmental pattern of expression of the alpha chemokine stromal cell-derived factor 1 in the rat central nervous system.

Stromal cell-derived factor 1 (SDF-1) is an alpha-chemokine that stimulates migration of haematopoietic progenitor cells and development of the immune system. SDF-1 is also abundantly and selectively expressed in the developing and mature CNS, as we show here. At embryonic day 15, SDF-1 transcripts were detected in the germinal periventricular zone and in the deep layer of the forming cerebral cortex. At birth, granule cells in the cerebellum and glial cells of the olfactory bulb outer layer showed an SDF-1 in situ hybridization signal that decreased progressively within the next 2 weeks. In other regions such as cortex, thalamus and hippocampus, SDF-1 transcripts detected at birth progressively increased in abundance during the postnatal period. SDF-1 protein was identified by immunoblot and/or immunocytochemistry in most brain regions where these transcripts were detected. SDF-1 was selectively localized in some thalamic nuclei and neurons of the fifth cortical layer as well as in pontine and brainstem nuclei which relay the nociceptive response. The presence of SDF-1 transcripts in cerebellar granule cells was correlated with their migration from the external to the inner granular layers with disappearance of the signal when migration was completed. In contrast, SDF1 mRNA signal increased during formation of the hippocampal dentate gyrus and stayed high in this region throughout life. The selective and regulated expression of SDF-1 in these regions suggests a role in precursor migration, neurogenesis and, possibly, synaptogenesis. Thus this alpha chemokine may be as essential to nervous system function as it is to the immune system.

Similar subcortical pattern of cognitive impairment in AIDS patients with and without dementia.

The aim of this study was to develop a series of neuropsychological tests that define the cortical and subcortical features of cognitive impairment and the characteristics of memory in demented and mildly cognitively impaired AIDS patients. We attempted to establish a usable method to assess and determine the type and degree of cognitive impairment in individual AIDS patients. We examined 53 patients without central nervous system opportunistic infections. A short battery included two scales of global efficiency (the Mattis dementia rating scale and the Mini Mental State Examination), a psychomotor speed test, an executive control assessment and explicit memory evaluation. Patients were categorized into four groups based on their score on both the Mattis dementia rating scale and the DSM-IV criteria: (1) asymptomatic; (2) having AIDS without cognitive impairment; (3) having AIDS with mild cognitive impairment; and (4) having AIDS dementia. Patients with mildly impaired cognition demonstrated slowed thinking, abnormal initiation and conceptualization, and memory impairment. AIDS dementia patients had slower motor activity and memory recall was more severely affected. The short neuropsychological battery was able to characterize modified cognitive performances in both severely and mildly cognitively impaired AIDS patients. The subcortical pattern of the memory disorder was obvious, regardless of the degree of cognitive impairment.

Differential signalling of the chemokine receptor CXCR4 by stromal cell-derived factor 1 and the HIV glycoprotein in rat neurons and astrocytes.

CXCR4 is the Gi protein-linked seven-transmembrane receptor for the alpha chemokine stromal cell-derived factor 1 (SDF-1), a chemoattractant for lymphocytes. This receptor is highly conserved between human and rodent. CXCR4 is also a coreceptor for entry of human immunodeficiency virus (HIV) in T cells and is expressed in the CNS. To investigate how these CXCR4 ligands influence CNS development and/or function, we have examined the expression and signalling of this chemokine receptor in rat neurons and astrocytes in vitro. CXCR4 transcripts and protein are synthesized by both cell types and in E15 brain neuronal progenitors. In these progenitors, SDF-1, but not gp120 (the HIV glycoprotein), induced activation of extracellular signal regulated kinases (ERKs) 1/2 and a dose-dependent chemotactic response. This chemotaxis was inhibited by Pertussis toxin, which uncouples Gi proteins and the bicyclam AMD3100, a highly selective CXCR4 antagonist, as well as by an inhibitor of the MAP kinase pathway. In differentiated neurons, both SDF-1 and the glycoprotein of HIV, gp120, triggered activation of ERKs with similar kinetics. These effects were significantly inhibited by Pertussis toxin and the CXCR4 antagonist. Rat astrocytes also responded to SDF-1 signalling by phosphorylation of ERKs but, in contrast to cortical neurons, no kinase activation was induced by gp120. Thus neurons and astrocytes can respond differently to signalling by SDF-1 and/or gp120. As SDF-1 triggers directed migration of neuronal progenitors, this alpha chemokine may play a role in cortex development. In differentiated neurons, both natural and viral ligands of CXCR4 activate ERKs and may therefore influence neuronal function.

[Diseases transmitted by non-conventional agents ("prions"): nosology and diagnosis]. / Les maladies à agents transmissibles non conventionnels ("Prions"): nosologie et diagnostic.

Transmissible non conventional agents are currently called "Prions". This is not a neutral terminology: the attractive Prion hypothesis (the infectious agent being a protein able to replicate in the absence of DNA or RNA) due to Stanley Prusiner is the prevalent one, and has shown to be heuristic, but has not been formally proven and does not easily explain all the data, unless modified and expanded. No simple account has been given for the very unusual physical, chemical, and biological properties of non conventional agents. These infectious agents are associated with degenerative diseases of the nervous system that are either the consequence of a genetic mutation or develop spontaneously in apparently normal individuals, and then can be transmitted to various susceptible hosts, including man. Thus, non conventional agents cannot be considered only as fascinating biological enigmas. They constitute a challenge for public health. The changing characteristics of prion-associated diseases has led to a renewing of their clinical and neuropathological diagnostic criteria. A brief survey of the nosology and neuropathology of prions diseases, with emphasis on new data and on difficulties, is provided. A simple classification based on the familial, sporadic or infectious variety of the disease is suggested. Familial diseases can be named according to the genetic disorder. Sporadic and infectious diseases can be classified following the main clinical symptoms and signs, and the presence or absence of amyloid plaques in the brain, until new tools (analysis of the glycosylation pattern of PrP, strain recognition) allow a more precise nomenclature. The new epidemiology of Prion disorders allowed by these new approaches relies on a full study of Prion diseases affected patients, which necessarily involves their genetic study, and the analysis of brain tissue. This, for practical and ethical reasons, is better achieved by autopsy.

Obtention and characterization of primary astrocyte and microglial cultures from adult monkey brains.

Simple methods for obtention of primary cultures of isolated astrocytes and microglia from adult simian brain have been developed. Characterization of these two glial cell populations were performed by morphological observations and by immunocytochemistry. The astroglial cultures were obtained by an indirect method. After L-leucine methyl-ester treatment and trypsinizations, more than 99% of cells expressed glial fibrillary acidic protein (GFAP), whereas no macrophages or microglia could be detected. Likely, the 1% remaining cells were immature astrocytes or cells that lost their GFAP expression. Cultured simian astrocytes expressed vimentin, laminin, and fibronectin. We also found a constitutively low expression of major histocompatibility complex (MHC) class II by cultured astrocytes which was significantly enhanced by lipopolysaccharide (LPS), interferon gamma (IFN-gamma), or tumor necrosis factor alpha (TNF-alpha) treatments. Microglial cultures were obtained by a direct method of isolation using Percoll gradient separations and compared to simian monocyte-derived macrophages or alveolar macrophages. Microglial cells differed from macrophages by their proliferation upon granulocyte-macrophage colony stimulating factor (GM-CSF) treatment and by their typical morphology when observed by scanning electron microscopy. As macrophages, they expressed in vitro CD68, CD64, CD14, CD11b, MHC class II, and fibronectin. However, contrary to macrophages, simian cultured microglia expressed laminin. This observation suggests that microglia represent a new potential source of this extracellular matrix protein in the brain.

Human immunodeficiency virus type 1 DNA and RNA load in brains of demented and nondemented patients with acquired immunodeficiency syndrome.

The relationship between dementia and human immunodeficiency virus type 1 (HIV-1) cerebral load is not clearly understood. We used immunohistochemistry and competitive polymerase chain reaction to evaluate the density ofgp 41 immunostained cells and the amount of HIV-1 DNA and RNA in the midfrontal gyrus of 21 HIV-1 infected patients, nine of whom were demented. The amounts of HIV-1 DNA and RNA, and the density of gp 41-positive cells were significantly linked. In this small series of cases, (1) although as a mean, there was a larger viral load in demented patients than in nondemented, this did not reach the significance level (2) discrepancies appeared in the population under study, some demented patients having low viral loads.

Inconstant apolipoprotein E (ApoE)-like immunoreactivity in amyloid beta protein deposits: relationship with APOE genotype in aging brain and Alzheimer's disease.

The aim of this study was to analyze the relationship between apolipoprotein E (ApoE) and amyloid beta-protein (A beta) deposits in 29 brains from a series of elderly women, whose intellectual status had been prospectively assessed by the Blessed test score (BTS). In three cases the APOE genotype was epsilon 3/4, in one epsilon 2/2, in 25 epsilon 3/3. ApoE-like immunoreactive lesions resembling a classic senile plaque (ApoE-SP) or a diffuse deposit (ApoE-DD) were quantitatively assessed in the first temporal gyrus (Brodmann's area 22). The density of ApoE-positive deposits (ApoE-SP+ApoE-DD) was correlated with BTS (negative correlation R2 = 0.36, P = 0.0006). The correlation of the density of ApoE-DD with BTS (R2 = 0.26, P = 0.0051) was weaker than that of ApoE-SP (R2 = 0.50, P < 0.0001). The density of ApoE deposits was correlated with that of A beta-immunoreactive lesions (positive correlation R2 = 0.57, P < 0.0001). ApoE immunohistochemistry labeled fewer deposits than A beta immunohistochemistry but showed as many senile plaques (ApoE-SP) as tau immunohistochemistry and the Gallyas and Bodian techniques. Two of the three cases with the epsilon 3/4 allele had Alzheimer's disease (AD) with numerous ApoE-immunoreactive senile plaques. In contrast, the brain of the third epsilon 3/4 case, whose intellectual status was normal, contained numerous A beta deposits but lacked ApoE-like immunoreactivity. The presence of ApoE may, thus, not be a prerequisite for A beta deposition, even in cases with the epsilon 3/4 allele. The density of ApoE-SP peaked in layer III, whereas that of ApoE-DD was maximal in deeper cortical layers. This contrast in the laminar distribution indicates that only some ApoE-DD may evolve into ApoE-SP. ApoE deposition was linked to the intellectual decline and was constant in the most severely demented patients, in whom numerous ApoE-positive (and A beta-positive) neuritic plaques were surrounded by dystrophic neurites. ApoE may accumulate in neuritic senile plaques during the progression of AD.

Coupling of ETB endothelin receptor to mitogen-activated protein kinase stimulation and DNA synthesis in primary cultures of rat astrocytes.

Astrocytes have been shown to express endothelin (ET) receptors functionally coupled, via different heterotrimeric G proteins, to several intracellular pathways. To assess the relative contribution of each subtype in the astrocytic responses to ET-1, effects of BQ123, an antagonist selective for the ET receptor subtype A (ETA-R), and IRL1620, an agonist selective for the ET receptor subtype B (ETB-R), were investigated in primary cultures of rat astrocytes. Binding experiments indicated that the ETB-R is the predominant subtype in these cells. Inhibition of forskolin-stimulated cyclic AMP production was observed under. ETB-R stimulation. Bordetella pertussis toxin (PTX) pretreatment completely abolished this effect, indicating that this pathway is coupled to the ETB-R via Gi protein. Increases of tyrosine phosphorylation of cellular proteins, stimulation of mitogen-activated protein kinase (MAPK), and DNA synthesis were also found to be mediated by the ETB-R, but through PTX-insensitive G protein. IRL1620-induced MAPK activation involved the adapter proteins Shc and Grb2 and the serine/threonine kinase Raf-1. This study reveals that the various effects of ET-1 in astrocytes are mediated by the ETB-R, which couples to multiple signaling pathways including the MAPK cascade.

Prion protein gene expression in cultured astrocytes treated by recombinant growth hormone and insulin-like growth factor.

Cases of Creutzfeldt-Jakob disease recently occurred after long treatments with pituitary extracted human growth hormone (GH). Prion protein (PrP) and glial fibrillary acidic protein (GFAP), an astrocyte-specific marker, both accumulate in the central nervous systems of infected individuals during transmissible subacute spongiform encephalopathies (TSSE). PrP expression has been linked to susceptibility to and development of these diseases. We have investigated the effects of recombinant growth hormone, its main biological effector, insulin-like growth factor 1 (IGF-1), and two well known mitogens, epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF), on PrP and GFAP mRNA levels in primary cultures of murine astrocytes using Northern blot quantitation. After 48 h of exposure to these soluble factors, PrP mRNA levels remained unchanged relative to controls in all growth factor-treated cultures, whereas GFAP mRNA concentrations decreased markedly in EGF- and bFGF-treated cultures. These results suggest that GH and IGF-1 do not modulate PrP gene expression in astrocytes.

Pharmacological studies of a new derivative of amphotericin B, MS-8209, in mouse and hamster scrapie.

Transmissible subacute spongiform encephalopathies (TSSE) are neurodegenerative diseases characterized by the presence of a modified, partially proteinase-resistant host protein, PrPSc, which accumulates in the brains of infected individuals. Recently it has been reported that amphotericin B (AmB) treatment of hamsters infected with scrapie strain 263K prolongs the incubation period of the disease, and dissociates in vivo replication of the scrapie agent from PrPSc accumulation. We report here on data obtained after treatment with AmB and one of its derivatives, MS-8209, in experimental scrapie of mouse and hamster. Treatment was carried out by the intraperitoneal route 6 days per week, at three different dosages initiated at the time of infection. Two regimens were used: during the early time of infection or throughout the experimental infection. Results indicate that MS-8209 was as efficient as AmB in prolonging the incubation time and decreasing PrPSc accumulation in the hamster scrapie model. A dose-dependent response was observed in mice treated early after experimental infection. At a dose of 2.5 mg/kg, MS-8209 significantly prolonged the incubation period (by 11.9%). In long-term treatment of mice, MS-8209 and AmB markedly reduced PrPSc levels in the preclinical stage of the disease. These data demonstrate that the effect of AmB is not restricted to one model (hamster-263K). This regimen leads to an inversion of the PrPSc to proteinase-sensitive protein (PrPSens) ratio, suggesting PrPSens (presumably cellular PrPC) accumulation occurs before its conversion into PrPSc. As it has been shown that AmB does not modify the infectivity titre, we conclude that the drugs could act by inhibiting either the interaction of the scrapie agent with PrPSens during the early times of infection or the conversion of PrPSens into PrPSc.

Astrocyte gene expression in experimental mouse scrapie.

The biological hallmark of transmissible spongiform encephalopathies is a significant accumulation, in brain, of the scrapie prion protein (PrPsc), often associated with an increased glial fibrillary acidic protein (GFAP) expression. This study was focused on astrocyte gene expression during scrapie development over a period of 172 days in intracerebrally inoculated newborn mice. The levels of expression of PrP and two specific astrocyte proteins, -GFAP and glutamine synthetase (GS)-, were investigated by Western and Northern blots. In brain, a 10-fold increased expression of GFAP mRNAS was demonstrated from 112 days post-inoculation to 172 days, whereas the "upregulation" of GS mRNAs was two-fold. GFAP was observed to increase 10- to 20-fold in scrapie-infected brain from day 112 to day 172, while PrP showed a three- to four-fold elevation. Both proteins were found in greater amount in the frontal cortex and cerebellum of animals with clinical scrapie than in those given an injection of normal brain. PrPsc was detected in scrapie brain from day 84 after inoculation, and thereafter increased about 20-fold until day 172. On the other hand, the concentration of glutamine synthetase remained constant in brain throughout the scrapie disease. To conclude, these results show that GFAP and GS mRNAs are differently upregulated in brain in the scrapie mouse model.

Modulation of prion protein gene expression by growth factors in cultured mouse astrocytes and PC-12 cells.

The present study was performed on primary cultures of mouse astrocytes and cultures of rat pheochromocytoma PC-12 in order to investigate the regulation of the prion protein (PrP) gene expression in relation to proliferation and differentiation. Treatment of PC-12 cells with interleukin-6 (IL-6) and beta-nerve growth factor (NGF) resulted in induction of neuronal differentiation. Northern blot analysis demonstrated a 4-fold increase of PrP mRNA in relation to cellular differentiation, after 7 days of treatment with either of the two factors. In astrocytes, PrP and glial fibrillary acidic protein (GFAP) mRNA levels were found to be regulated in a similar manner during development in vitro. A 3-fold increase of their mRNAs was observed from 5 to 14 days of culture (proliferation period). Then, their gene expressions showed a slight decrease from 14 to 28 days (maturation period). Treatment of astrocytes with IL-6, basic fibroblast growth factor (bFGF), and epidermal growth factor (EGF) appeared to markedly down-regulate the expression of GFAP mRNAs, which might reflect cell maturation. In contrast, they had no significant effect on the expression of PrP gene. These results suggest that the PrP gene expression is differently regulated in neural cells. In neuronal cells, it is mainly associated with differentiation. On the other hand, in astrocytes, the PrP mRNA level seems to be not only related to the proliferation and differentiation stages.

Variations in prion protein and glial fibrillary acidic protein mRNAs in the brain of scrapie-infected newborn mouse.

To begin to understand the molecular basis of cases of Creutzfeldt-Jakob disease recently described in young children, the expression of prion protein and glial fibrillary acidic protein (GFAP) mRNAs was investigated during the development of the brain of scrapie-infected newborn mice. Changes in the time course of expression were identified by Northern blot quantification between days 1 and 172. Although scrapie-infected and control animals showed no detectable changes in brain development (first 56 days of life), GFAP mRNAs were found to increase significantly as early as day 84. A 10-fold increase in the level of GFAP mRNA was observed in brain between day 112 and death (day 172).

Regulation of the glial fibrillary acidic protein, beta actin and prion protein mRNAs during brain development in mouse.

Developmental regulation in mRNAs of three brain proteins has been investigated by Northern blot evaluation in C57BL/6 mice. The mRNAs of two cytoskeletal components, glial fibrillary acidic protein (GFAP) and beta actin, varied significantly, and differently, during brain development (0-56 days postnatal). The beta actin mRNAs peaked at day 1 after a slight increase, then dropped rapidly during the first 15 days postnatal, and thereafter remained at a level which was strictly maintained throughout development and adulthood. Conversely, the GFAP mRNAs increased during the first two weeks after birth (astroglial proliferation), and then slightly declined until the adult stage (astroglial cell differentiation). The prion protein (PrP) mRNAs were detectable as soon as birth, and increased 4-fold during brain maturation. Then, during the adult life, the GFAP and PrP mRNAs did not change markedly. Nevertheless, slight but significant increases in the mRNA levels of both GFAP and PrP were observed at older stages (360 days). These results are analysed in the light of the implications of PrP and GFAP in scrapie infection models.