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L Olivares

Publications and source records attributed to L Olivares.

At least 19 recordsLinked to original sources
PubMed2004

NK cells modulate the cytotoxic activity generated by Mycobacterium leprae-hsp65 in leprosy patients: role of IL-18 and IL-13.

Protection against intracellular pathogens such as Mycobacterium leprae is critically dependent on the function of NK cells at early stages of the immune response and on Th1 cells at later stages. In the present report we evaluated the role of IL-18 and IL-13, two cytokines that can influence NK cell activity, in the generation of M. leprae-derived hsp65-cytotoxic T lymphocytes (CTL) from peripheral blood mononuclear cells (PBMC) of leprosy patients. We demonstrated that IL-18 modulates hsp65-induced CTL generation and collaborates with IL-12 for this effect. In paucibacillary (PB) patients and normal controls (N) depletion of NK cells reduces the cytolytic activity. Under these conditions, IL-12 cannot up-regulate this CTL generation, while, in contrast, IL-18 increases the cytotoxic activity both in the presence or absence of NK cells. IL-13 down-regulates the hsp65-induced CTL generation and counteracts the positive effect of IL-18. The negative effect of IL-13 is observed in the early stages of the response, suggesting that this cytokine affects IFNgamma production by NK cells. mRNA coding for IFNgamma is induced by IL-18 and reduced in the presence of IL-13, when PBMC from N or PB patients are stimulated with hsp65. Neutralization of IL-13 in PBMC from multibacillary (MB) leprosy patients induces the production of IFNgamma protein by lymphocytes. A modulatory role on the generation of hsp65 induced CTL is demonstrated for IL-18 and IL-13 and this effect takes place through the production of IFNgamma.

Adult↗
PubMed1997

Analysis of the transmembrane topology of the glycine transporter GLYT1.

A theoretical 12-transmembrane segment model based on the hydrophobic moment has been proposed for the transmembrane topology of the glycine transporter GLYT1 and all other members of the sodium- and chloride-dependent transporter family. We tested this model by introducing N-glycosylation sites along the GLYT1 sequence as reporter for an extracellular localization and by an in vitro transcription/translation assay that allows the analysis of the topogenic properties of different segments of the protein. The data reported herein are compatible with the existence of 12 transmembrane segments, but support a rearrangement of the first third of the protein. Contrary to prediction, hydrophobic domain 1 seems not to span the membrane, and the loop connecting hydrophobic domains 2 and 3, formerly believed to be intracellular, appears to be extracellularly located. In agreement with the theoretical model, we provide evidence for the extracellular localization of loops between hydrophobic segments 5 and 6, 7 and 8, 9 and 10, and 11 and 12.

Amidohydrolases↗
PubMed1995

Regional distribution and developmental variation of the glycine transporters GLYT1 and GLYT2 in the rat CNS.

The high-affinity glycine transporter in neurons and glial cells is the primary means of inactivating synaptic glycine. Previous molecular cloning studies have indicated heterogeneity of glycine transporters in the CNS. Here the distribution of glycine transporter GLYT1 and GLYT2 transcripts and proteins in different regions and developmental stages of the rat brain were analysed by Northern, Western and in situ hybridization techniques. Sequence-specific riboprobes and two specific antibodies raised against fusion proteins were used, containing either 76 or 193 amino acids of the C or N terminus of the GLYT1 and GLYT2 transporters respectively. High levels of GLYT1 transcripts were found in the spinal cord, brainstem and cerebellum, and moderate levels in forebrain regions such as the cortex or hippocampus. GLYT2 transcripts are restricted to the spinal cord, brainstem and cerebellum. The onset of both GLYT1 and GLYT2 expression in the brainstem occurred in late fetal life, and full expression of these proteins was observed before weaning. There was a stepwise increase in the levels of mRNA and protein for these two transporters, reaching a maximum by the second postnatal week, followed by a slight decrease until adult values were reached by the fourth postnatal week. These data reveal interesting parallelism between the distribution of different glycine transporters and glycine receptor subunits, and suggest discrete roles for distinct glycine transporters.

Aging↗
PubMed1995

The role of N-glycosylation in the targeting and activity of the GLYT1 glycine transporter.

To elucidate the role of N-glycosylation in the function of the high affinity glycine transporter GLYT1, we have investigated the effect of the glycosylation inhibitor tunicamycin as well as the effect of the disruption of the putative glycosylation sites by site-directed mutagenesis. SDS-polyacrylamide gel electrophoresis of proteins from GLYT1-transfected COS cells reveals a major band of 80-100 kDa and a minor one of 57 kDa. Treatment with tunicamycin produces a 40% inhibition in transport activity and a decrease in the intensity of the 80-100-kDa band, whereas the 57-kDa band decreases in size to yield a 47-kDa protein corresponding to the unglycosylated form of the transporter. Simultaneous mutation of Asn-169, Asn-172, Asn-182, and Asn-188 to Gln also produces the 47-kDa form of the protein, indicating that there are no additional sites for N-glycosylation. Progressive mutation of the potential glycosylation sites produces a progressive decrease in transport activity and in size of the protein, indicating that the four putative glycosylation sites are actually glycosylated. N-Glycosylation of the GLYT1 is not indispensable for the transport activity itself, as demonstrated by enzymatic deglycosylation of the transporter. Analysis of surface proteins by biotinylation and by immunofluorescence demonstrates that a significant portion of the unglycosylated GLYT1 mutant remains in the intracellular compartment. This suggests that the carbohydrate moiety of glycine transporter GLYT1 is necessary for the proper trafficking of the protein to the plasma membrane.

Amino Acid Transport Systems, Neutral↗
PubMed1995

Regulation by phorbol esters of the glycine transporter (GLYT1) in glioblastoma cells.

The high-affinity glycine transporter in neurons and glial cells is the primary means of inactivating synaptic glycine. The effects of 12-O-tetradecanoylphorbol ester (TPA), a potent activator of protein kinase C (PKC), on the high-affinity Na(+)-dependent glycine transport were investigated in C6 cells, a cell line of glial origin. Incubation of C6 cells with TPA led to concentration- and time-dependent decrease in the glycine transport that could be completely suppressed by the addition of the PKC inhibitor staurosporine. The TPA effect could be mimicked by oleoylacetylglycerol and exogenous phospholipase C. Northern and Western blot analysis indicate that C6 cells express the GLYT1 glycine transporter. Incubation of COS cells transiently transfected with a full-length clone of the GLYT1 transporter in the presence of TPA, produces a decrease in glycine uptake.

Alkaloids↗
PubMed1995

Glycine transporters are differentially expressed among CNS cells.

Glycine is the major inhibitory neurotransmitter in the spinal cord and brainstem and is also required for the activation of NMDA receptors. The extracellular concentration of this neuroactive amino acid is regulated by at least two glycine transporters (GLYT1 and GLYT2). To study the localization and properties of these proteins, sequence-specific antibodies against the cloned glycine transporters have been raised. Immunoblots show that the 50-70 kDa band corresponding to GLYT1 is expressed at the highest concentrations in the spinal cord, brainstem, diencephalon, and retina, and, in a lesser degree, to the olfactory bulb and brain hemispheres, whereas it is not detected in peripheral tissues. Pre-embedding light and electron microscopic immunocytochemistry show that GLYT1 is expressed in glial cells around both glycinergic and nonglycinergic neurons except in the retina, where it is expressed by amacrine neurons, but not by glia. The expression of a 90-110 kDa band corresponding to GLYT2 is restricted to the spinal cord, brain-stem, and cerebellum; in addition, very low levels occur in the diencephalon. GLYT2 is found in presynaptic elements of neurons thought to be glycinergic. However, in the cerebellum, GLYT2 is expressed both in terminal boutons and in glial elements. The physiological consequences of the regional and cellular distributions of these two proteins as well as the possibility of the existence of an unidentified neuronal form of GLYT1 are discussed.

Amino Acid Sequence↗
PubMed1994

Carboxyl terminus of the glycine transporter GLYT1 is necessary for correct processing of the protein.

The high affinity glycine transporter in neurons and glial cells is the primary means of inactivating synaptic glycine. To understand the structure-function relationships, especially the role of the intracellular carboxyl- and amino-terminal domains, we have modified the glycine transporter GLYT1 by using a polymerase chain reaction-based mutagenesis approach. Deletion of the first 30 amino acids of the amino terminus does not alter transport of glycine. Truncation of the last 34 amino acids of the carboxyl terminus did not impair glycine transport, but progressively more extensive deletions produced a progressive decrease in transport activity. All the fully active or partially active forms of the transporter retain the characteristic sodium and chloride dependence of the wild type. When the nonfunctional mutants of the carboxyl terminus were examined by an immunofluorescence technique, the carrier was no longer found in the membrane. This suggests that the carboxyl terminus of GLYT1 may be involved in the membrane insertion process. Moreover, the transporter that is not fully processed is not functional, because transport activity cannot be rescued in a solubilization-reconstitution experiment.

Amino Acid Transport Systems, Neutral↗
PubMed1992

L-glutamate transporter derived from mRNAs of primary glial cultures: expression in Xenopus laevis oocytes.

A high-affinity sodium-dependent L-glutamate transporter was expressed in Xenopus oocytes after microinjection of poly(A)+ RNA from primary astrocyte cultures from rat brain cortex. mRNA-induced L-glutamate transport was saturable by substrate and shows kinetic features similar to those found in intact glial cell preparations. L-Glutamate accumulation was prevented by rising the external K+ concentration or by coincubation with L-, D-aspartate or D-glutamate. After fractionation by sucrose density gradient, the mRNA encoding for the expressed L-glutamate transporter from glial cells was found in fractions containing messages of 2.05-2.9 kilobases (kb) in length.

Alkaline Phosphatase↗
PubMed1988

Two years of cyclophosphamide and 5-fluorouracil as adjuvant chemotherapy for stage II and III breast carcinoma.

The results after 6 years of a prospective clinical trial of adjuvant chemotherapy with a regimen of two drugs--cyclophosphamide and 5-fluorouracil (CF)-- for 2 years in 97 women with stage II or III breast cancer are reported. Eligible patients were free from distant metastases. All patients began adjuvant therapy within 4 weeks of surgery; therapy consisted of radical, modified, or extended radical mastectomy. No postoperative radiotherapy was given. The results are compared with a historical control group from previous consecutive patients treated by surgery alone. Patients were stratified by age (younger than 50 or older than or equal to 50) and nodal status (one to three positive axillary nodes vs. four or more positive nodes). The estimated 6-year survival was 60% for CF patients vs. 31% for control patients (P = 0.001). The estimated 6-year disease-free survival was 53.6 and 30.3% for CF and control, respectively (P = 0.007). There was a trend toward longer disease-free survival (DFS) and survival (S) in patients treated with CF, but this was not significant in all the subgroups. Disease-free survival was statistically significant in the subgroup of women greater than or equal to 50 years old with one to three positive nodes (P = 0.038); survival in the patients less than or equal to 49 years old with four or more positive nodes (P = 0.0036); and in patients greater than or equal to 50 years old with one to three lymph nodes involvement (P = 0.038).

Adult↗
PubMed1988

[Cryoglobulinemia. Study of 3 cases (2 essential and 1 secondary)].

There patients with diverse skin manifestations of cryoglobulinemia have been investigated. Case number one. An essential mixed cryoglobulinemia with purpuric cutaneous lesions, rheumatoid factor, asthenia and chronic glomerulonephritis. Case number two. A secondary mixed cryoglobulinemia (chronic lymphatic leukemia), with purpuric lesions. Case number three. An essential mixed cryoglobulinemia, with extensive and deep necrotic lesions that necessitated the amputation of the left hand; severe hepatic, renal, and splenic damage by deposit of cryoglobulin. Clinical features, etiopathogenesis and therapeutic possibilities are commented on.

Adult↗
PubMed1979

Carcinoma of the penis: analysis of 192 consecutive cases at the Instituto Nacional de Enfermedades Neoplasicas.

Between 1952 and 1976 192 consecutive cases of penile epidermoid carcinoma were seen at the Instituto Nacional de Enfermedades Neoplascias of Lima, Peru. The mean age when the disease developed was 60.5 years, with a peak incidence between 60 and 64 years (32 patients). No correlation was observed between extensive lesions of the shaft of the penis to high pathological staging. Clinical examination of the inguino-crural nodes is not a good criterion for staging. We wait six weeks after eradication of primary lesion before lymphadenectomy. No correlation exists between grade and pathological stage. When no lymph node was positive, the overall survival rate over five years was 90.69%; when lymph nodes were metastasized there was an overall survival rate over five years of only 9.39 percent. The coefficient of cancer versus noncancer cause of death was 1.25 for pathological stage I, 3.09 for pathological stage II, 4.83 for pathological stage III, and 10.000 for pathological stage IV. Our patients did have advanced disease, as 57.14% of deaths occurred at two years and 25.21% more at five years.

Adult↗