Luis Cartier

Semaphorin CD100 from Activated T Lymphocytes Induces Process Extension Collapse in Oligodendrocytes and Death of Immature Neural Cells

An inappropriate cross talk between activated T lymphocytes infiltrating the CNS and neural cells can sustain the onset and progression of demyelination and axonal degeneration in neuroinflammatory diseases. To mimic this deleterious cross talk, we designed an experimental paradigm consisting of transient cocultures of T lymphocytes chronically activated by retrovirus infection (not virus productive) with human multipotent neural precursors or primary oligodendrocytes from rat brain. We showed that activated T lymphocytes induced apoptotic death of multipotent neural progenitors and immature oligodendrocytes after a progressive collapse of their process extensions. These effects were reminiscent of those induced by brain semaphorin on neural cells. Blockade by specific Abs of soluble CD100 (sCD100)/semaphorin 4D released by activated T cells, or treatment with rsCD100, demonstrated that this immune semaphorin has the ability to collapse oligodendrocyte process extensions and to trigger neural cell apoptosis, most likely through receptors of the plexin family. The specific presence of sCD100 in the cerebrospinal fluid and of CD100-expressing T lymphocytes in the spinal cord of patients suffering with neuroinflammatory demyelination pointed to the potential pathological effect of sCD100 in the CNS. Thus, our results show that CD100 is a new important element in the deleterious T cell-neural cell cross talk during neuroinflammation and suggest its role in demyelination or absence of remyelination in neuroinflammatory diseases including multiple sclerosis and human T lymphotropic virus type 1-associated myelopathy.

The presence of ancient human T-cell lymphotropic virus type I provirus DNA in an Andean mummy

The worldwide geographic and ethnic clustering of patients with diseases related to human T-cell lymphotropic virus type I (HTLV-I) may be explained by the natural history of HTLV-I infection. The genetic characteristics of indigenous people in the Andes are similar to those of the Japanese, and HTLV-I is generally detected in both groups. To clarify the common origin of HTLV-I in Asia and the Andes, we analyzed HTLV-I provirus DNA from Andean mummies about 1,500 years old. Two of 104 mummy bone marrow specimens yielded a band of human β-globin gene DNA 110 base pairs in length, and one of these two produced bands of HTLV-I-pX (open reading frame encoding p40x, p27x) and HTLV-I-LTR (long terminal repeat) gene DNA 159 base pairs and 157 base pairs in length, respectively. The nucleotide sequences of ancient HTLV-I-pX and HTLV-I-LTR clones isolated from mummy bone marrow were similar to those in contemporary Andeans and Japanese, although there was microheterogeneity in the sequences of some mummy DNA clones. This result provides evidence that HTLV-I was carried with ancient Mongoloids to the Andes before the Colonial era. Analysis of ancient HTLV-I sequences could be a useful tool for studying the history of human retroviral infection as well as human prehistoric migration.

Characteristic distribution of HTLV type I and HTLV type II carriers among native ethnic groups in South America.

To confirm the geographic and ethnic segregation of HTLV-I and HTLV-II carriers in native populations in South America, we have conducted a seroepidemiological study of native populations in South America, including HTLV-I carriers distributed among seven ethnic groups in the Andes highlands of Colombia, Peru, Bolivia, Argentina, and Chile, and two ethnic groups on Chiloe Island and Easter Island; and HTLV-II carriers distributed among seven ethnic groups of the lowlands along the Atlantic coast of Colombia, Orinoco, Amazon, and Patagonia, and one ethnic group on Chiloe Island. The incidence rate of HTLV-I and HTLV-II carriers varied among the ethnic groups, ranging from 0.8 to 6.8% for HTLV-I seropositivity and from 1.4 to 57.9% for HTLV-II seropositivity. A new HTLV-I focus was found among the Peruvian Aymara (1.6%), the Bolivian Aymara (5.3%) and Quechua (4.5%), the Argentine Puna (2.3%), and the Chilean Atacama (4.1%), while on HTLV-II focus was found among the Brazilian Kayapo (57.9%), the Paraguayan Chaco (16.4%), and the Chilean Alacalf (34.8%) and Yahgan (9.1%). The distribution of HTLV-I/II foci showed a geographic clustering of HTLV-I foci in the Andes highlands and of HTLV-II foci in the lowlands of South America. It was thus suggested that South American natives might be divided into two major ethnic groups by HTLV-I and HTLV-II carrier state.

Clinical and neuropathological study of six patients with spastic paraparesis associated with HTLV-I : An axomyelinic degeneration of the central nervous system

Between 1990 to 1994, 6 TSP/HAM patients, 3 women and 3 men with an average age of 57.1 years (39 to 76 years old), who died in the Salvador Hospital were submitted to postmortem examination. The mean time of paraparesis was 7 years (3 to 17 years), and 2 patients had pseudobulbar signs. Three cases had macroscopic atrophy of the spinal cord. Histologically, all cases had lesions in the pyramidal tracts and 4 cases showed somatotopic lesions of the Golls tracts which followed a “dying back” ascendant and descendant distribution, respectively. In 2 cases, both of which had intellectual impairment, demyelination of the subcortical and parathalamic areas was observed without U fiber involvement. Abnormal vessels with gross thickening of the adventitia, many of them with lymphocytic cuffs, were seen everywhere, especially in the spinal cord, brain stem, midbrain and meninges, but no relation between these findings and the parenchymal lesions was observed. Also, in the cases with posterior column involvement, neuronal changes and proliferation of satellite cells in the dorsal ganglia were found. All cases showed histological sialoadenitis and none had inflammatory muscle changes. We conclude that the lesions affected the neuraxis in a systemic axial fashion as in degenerative diseases, and did not seem to be secondary to vascular or inflammatory abnormalities.

Alteration of Blood-Brain Barrier Integrity by Retroviral Infection

The blood–brain barrier (BBB), which forms the interface between the blood and the cerebral parenchyma, has been shown to be disrupted during retroviral-associated neuromyelopathies. Human T Lymphotropic Virus (HTLV-1) Associated Myelopathy/Tropical Spastic Paraparesis (HAM/TSP) is a slowly progressive neurodegenerative disease associated with BBB breakdown. The BBB is composed of three cell types: endothelial cells, pericytes and astrocytes. Although astrocytes have been shown to be infected by HTLV-1, until now, little was known about the susceptibility of BBB endothelial cells to HTLV-1 infection and the impact of such an infection on BBB function. We first demonstrated that human cerebral endothelial cells express the receptors for HTLV-1 (GLUT-1, Neuropilin-1 and heparan sulfate proteoglycans), both in vitro, in a human cerebral endothelial cell line, and ex vivo, on spinal cord autopsy sections from HAM/TSP and non-infected control cases. In situ hybridization revealed HTLV-1 transcripts associated with the vasculature in HAM/TSP. We were able to confirm that the endothelial cells could be productively infected in vitro by HTLV-1 and that blocking of either HSPGs, Neuropilin 1 or Glut1 inhibits this process. The expression of the tight-junction proteins within the HTLV-1 infected endothelial cells was altered. These cells were no longer able to form a functional barrier, since BBB permeability and lymphocyte passage through the monolayer of endothelial cells were increased. This work constitutes the first report of susceptibility of human cerebral endothelial cells to HTLV-1 infection, with implications for HTLV-1 passage through the BBB and subsequent deregulation of the central nervous system homeostasis. We propose that the susceptibility of cerebral endothelial cells to retroviral infection and subsequent BBB dysfunction is an important aspect of HAM/TSP pathogenesis and should be considered in the design of future therapeutics strategies.

Familial Creutzfeldt-Jakob disease in Chile is associated with the codon 200 mutation of the PRNP amyloid precursor gene on chromosome 20

We have found the codon 200Lys mutation in 6 Chilean CJD families, including a family in the rural case cluster in Chillán. Thus, all 3 of the known clusters of CJD, in Slovakia, Libyan-born Israeli Jews, and Chile, are linked to the presence of the same mutation. The phenotypic features of the disease in these families are similar to those reported for other clustered or individual families elsewhere in the world. The heterogeneous genetic composition of the Chilean population interpreted in light of historical migration patterns suggests that the mutation may have entered Chile by Jewish emigration from Spain.

Molecular phylogeny of human T-cell leukemia virus type I and II of amerindians in Colombia and Chile

Six human T-cell leukemia virus type I (HTLV-I) and eight human T-cell leukemia virus type II (HTLV-II) cases newly isolated from the South American countries of Colombia and Chile were analyzed together with the two Amerindian HTLV-I isolates previously reported. All of the HTLV-I isolates belonged to the transcontinental subgroup of the “cosmopolitan” group, and Colombian isolates, including those from native Amerindians and Negroes, formed a single tight cluster within this subgroup. The transcontinental subgroup consisted of isolates from various regions such as the Caribbean basin, India, Iran, South Africa, Sakhalin, and Japan, and included isolates from the “Ainu” and “Okinawa” people, regarded as relatively pure Japanese descended from the prehistoric “Jomon” period which began more than 10,000 years ago. This implied a dissemination of the subgroup associated with the movement of human beings in ancient times. On the other hand, all of the HTLV-II isolates from native Amerindians in Colombia and Chile belonged to the HTLV-IIb subtype which has previously been reported to be mainly endemic in certain populations of native Amerindians. The southernmost isolate from Chile, showing wide distribution of the llb subtype in native South Amerindians and largest heterogeneity of the subtype in Colombian isolates, supported the idea that the HTLV-IIb subtype has been endemic for a long time in native Indians of South America.

T Lymphocytes Activated by Persistent Viral Infection Differentially Modify the Expression of Metalloproteinases and Their Endogenous Inhibitors, TIMPs, in Human Astrocytes: Relevance to HTLV-I-Induced Neurological Disease

Activation of T lymphocytes by human pathogens is a key step in the development of immune-mediated neurologic diseases. Because of their ability to invade the CNS and their increased secretion of proinflammatory cytokines, activated CD4+ T cells are thought to play a crucial role in pathogenesis. In the present study, we examined the expression of inflammatory mediators the cytokine-induced metalloproteinases (MMP-2, -3, and -9) and their endogenous inhibitors, tissue inhibitors of metalloproteinases (TIMP-1, -2, and -3), in human astrocytes in response to activated T cells. We used a model system of CD4+ T lymphocytes activated by persistent viral infection (human T lymphotropic virus, HTLV-I) in transient contact with human astrocytes. Interaction with T cells resulted in increased production of MMP-3 and active MMP-9 in astrocytes despite increased expression of endogenous inhibitors, TIMP-1 and TIMP-3. These data suggest perturbation of the MMP/TIMP balance. These changes in MMP and TIMP expression were mediated, in part, by soluble factors (presumably cytokines) secreted by activated T cells. Integrin-mediated cell adhesion is also involved in the change in MMP level, since blockade of integrin subunits (α1, α3, α5, and β1) on T cells resulted in less astrocytic MMP-9-induced expression. Interestingly, in CNS tissues from neurological HTLV-I-infected patients, MMP-9 was detected in neural cells within the perivascular space, which is infiltrated by mononuclear cells. Altogether, these data emphasize the importance of the MMP-TIMP axis in the complex interaction between the CNS and invading immune cells in the context of virally mediated T cell activation.

Dynamin 2 Mutations Associated With Human Diseases Impair Clathrin-Mediated Receptor Endocytosis

Dynamin 2 (DNM2) is a large GTPase involved in the release of nascent vesicles during endocytosis and intracellular membrane trafficking. Distinct DNM2 mutations, affecting the middle domain (MD) and the Pleckstrin homology domain (PH), have been identified in autosomal dominant centronuclear myopathy (CNM) and in the intermediate and axonal forms of the Charcot‐Marie‐Tooth peripheral neuropathy (CMT). We report here the first CNM mutation (c.1948G>A, p.E650 K) in the DNM2 GTPase effector domain (GED), leading to a slowly progressive moderate myopathy. COS7 cells transfected with DNM2 constructs harboring a disease‐associated mutation in MD, PH, or GED show a reduced uptake of transferrin and low‐density lipoprotein (LDL) complex, two markers of clathrin‐mediated receptor endocytosis. A decrease in clathrin‐mediated endocytosis was also identified in skin fibroblasts from one CNM patient. We studied the impact of DNM2 mutant overexpression on epidermal growth factor (EGF)‐induced extracellular signal‐regulated kinase 1 (ERK1) and ERK2 activation, known to be an endocytosis‐ and DNM2‐dependent process. Activation of ERK1/2 was impaired for all the transfected mutants in COS7 cells, but not in CNM fibroblasts. Our results indicate that impairment of clathrin‐mediated endocytosis may play a role in the pathophysiological mechanisms leading to DNM2‐related diseases, but the tissue‐specific impact of DNM2 mutations in both diseases remains unclear. Hum Mutat 30:1–9, 2009.

Ancient HTLV type 1 provirus DNA of Andean mummy.

The worldwide geographic and ethnic clustering of patients with diseases related to human T cell lymphotropic virus type 1 (HTLV-1) may be explained by the natural history of HTLV-1 infection. The genetic characteristics of indigenous people in the Andes are similar to those of the Japanese, and HTLV-1 is generally detected in both groups. To clarify the common origin of HTLV-1 in Asia and the Andes, we analyzed HTLV-1 provirus DNA from Andean mummies about 1500 years old. Two of 104 mummy bone marrow specimens yielded a band of human beta-globin gene DNA 110 base pairs in length, and one of these two produced bands of HTLV-1-pX (open reading frame encoding p(40x), p(27x)) and HTLV-1-LTR (long terminal repeat) gene DNA 159 base pairs and 157 base pairs in length, respectively. The nucleotide sequences of ancient HTLV-1-pX and HTLV-1-LTR clones isolated from mummy bone marrow were similar to those in contemporary Andeans and Japanese, although there was microheterogeneity in the sequences of some mummy DNA clones. This result provides evidence that HTLV-1 was carried with ancient Mongoloids to the Andes before the Colonial era. Analysis of ancient HTLV-1 sequences could be a useful tool for studying the history of human retroviral infection as well as human prehistoric migration.