Biomedical subjects
Publications and source records attributed to J Teixidó.
A compilation of combinatorial chemistry techniques applied to anti-HIV drug development is presented in this review. This synthetic strategy together with high throughput screening assays has allowed the discovery and optimization of novel lead anti-HIV compounds.
INTRODUCTION: Despite the interest generated by the increasing number of studies that measure Quality of Life among patients and caregivers, few of these studies measure the caregivers burden in Peritoneal Dialysis (PD). OBJECTIVE: The main target of this study was to create a burden measure questionnaire applicable amongst caregivers of PD patients. METHODOLOGY INCLUSION CRITERIA: 1) Patients had to be in PD treatment for more than 3 months; 2) Patients had to receive help with the PD treatment from a caregiver. The study was divided into 3 phases: 1st) design and use of the initial questionnaire; 2nd) a test-retest on a modified scale; and 3rd) to provide the questionnaire-3 to two collaborative centres with similar PD programs. Four groups of caregivers were established: A1:23, A2:17, B:7 and C:16 caregivers. We applied 5 scales (5): 1--Patient Dependence on caregiver, from caregivers' view (D); 2--Complete caregiver burden (CB), including 12 items which measure the caregivers' subjective burden, 3--Reduced caregiver burden (RB), as the one before but with only 8 items, 4--Repercussions on the caregiver (R), which measures objective burden; 5--Specific PD tasks (ST), a scale that measures the effort the tasks implied in the PD treatment represent for the caregiver. RESULTS: We studied 63 caregivers (table I): mean age: 53.43 (SD = 12.3); Sex: Females: 86.4%, Males: 13.6%, corresponding to 63 patients: mean age: 59.79 (SD = 15.9); Sex: Males: 80.3%, Females: 19.7%. Valuable results for reliability, unidimensionality, and discrimination were obtained in the 1st and 2nd phases, except for burden scale which was compound of two factors; then one of those factors was suppressed. In the 3rd phase, ANOVA did not show any differences between centres (table II). Consequently, all caregivers could be analysed together. Reliability results for each one of the third phase scales (table III) were: D: Cronbach alpha = 0,886; CB: alpha = 0,894; RB: alpha = 0,857; R: alpha = 0,892; ST: alpha = 0,62. Although the ST scale obtained an acceptable reliability, it was suppressed in the 3rd phase due to the low correlation with other scales and the fact that it was not applicable to all caregivers. Finally, a direct correlation was found between third phase scales (table IV): D-RB: r = 0.502, p < or = 0.001; D-R: r = 0.599, p < or = 0.001; RB-R: r = 0.775, p < or = 0.001. We must headlight that both Burden scales, and the Repercussion scale, obtained a direct correlation with the Dependency scale. CONCLUSION: A questionnaire has been created to measure burden and repercussions on caregivers of peritoneal dialysis patients. It can already be applied, as requirements of both reliability and validity are fulfilled. This questionnaire can be a useful tool to prevent caregivers' burnout.
During the last 12 years, an increasing frequency in condemnation of hunted red deer and wild boar carcasses due to the presence of tubercle-like lesions has been observed in Extremadura (Western Spain). Before 1993, tuberculosis was a very rare finding in hunted animals. The current tuberculosis regional prevalence in cattle approaches 0.4% after years of expensive test and slaughter campaigns. It is imperative to investigate the epidemiology of Mycobacterium bovis infection in red deer and wild boar in order to keep a good health status and to maintain the effectiveness of domestic species TB eradication programs. The present paper evaluates the problem in Sierra de San Pedro, estimating the prevalence of TB in wild boar and red deer, the main wild artiodactyls in the area, and domestic cattle since 1992-2004, by the use of a low-cost surveillance method based on detailed pathological inspection of hunted animal carcasses. Microbiology and molecular epidemiology studies on several M. bovis isolates from domestic and wild animals helped to define the interspecies contacts. These findings, as well as recent history of game estates management and descriptive epidemiology field work, throw light on the rise and maintenance of these epizootics.
In this study we describe photodamaging and photokilling effects of palladium(II)-tetraphenylporphycene (PdTPPo) (previously incorporated into dipalmitoylphosphatidylcholine liposomes) on the human lung adenocarcinoma A-549 cell line. No dark cytotoxicity was found when the drug was applied at 10(-6) M or 5 x 10(-7) M for 1 or 18 h, respectively. After 1-h treatment with 10(-7) M or 5 x 10(-7) M PdTPPo followed by red light irradiation for variable times, dose-dependent lethal effects were observed in A-549 cells. Apoptosis was not found after the above photodynamic treatments or under even milder sublethal conditions. In contrast to HeLa cells subjected to PdTPPo photosensitization where either apoptosis or necrosis were induced, morphological analysis and electrophoretical DNA pattern of A-549 cells always revealed a clearly necrotic death mechanism. However, A-549 cells died by apoptosis after serum and L-glutamine deprivation, indicating that only the photodynamically induced apoptosis was inhibited. Immunofluorescent labeling revealed that microtubules and actin microfilaments were immediately and strongly damaged by photodynamic treatments with PdTPPo. No metaphase arrest and/or mitotic alterations were observed after phototreatments. Present results show that the cell type plays a fundamental role in relation to the apoptotic or necrotic response to photosensitization, and that cytoskeletal components are important targets implicated in cell death processes.
Chemokines are secreted into the tumor microenvironment by tumor-infiltrating inflammatory cells as well as by tumor cells. Chemokine receptors mediate agonist-dependent cell responses, including migration and activation of several signaling pathways. In the present study we show that several human melanoma cell lines and melanoma cells on macroscopically infiltrated lymph nodes express the chemokine receptors CXCR3 and CXCR4. Using the highly invasive melanoma cell line BLM, we demonstrate that the chemokine Mig, a ligand for CXCR3, activates the small GTPases RhoA and Rac1, induces a reorganization of the actin cytoskeleton, and triggers cell chemotaxis and modulation of integrin VLA-5- and VLA-4-dependent cell adhesion to fibronectin. Furthermore, the chemokine SDF-1alpha, the ligand of CXCR4, triggered modulation of beta(1) integrin-dependent melanoma cell adhesion to fibronectin. Additionally, Mig and SDF-1alpha activated MAPKs p44/42 and p38 on melanoma cells. Expression of functional CXCR3 and CXCR4 receptors on melanoma cells indicates that they might contribute to cell motility during invasion as well as to regulation of cell proliferation and survival.
We have examined the role of the small GTPase Rho and its downstream effector, the Rho-associated kinase (ROCK), in the control of the adhesive and signaling function of the lymphocyte function-associated antigen-1 (LFA-1) integrin in human T-lymphocytes. Inhibition of Rho (either by treatment with C3-exoenzyme or transfection with a dominant-negative form of Rho (N19Rho)) or ROCK (by treatment with Y-27632) results in the following: (a) partial disorganization and aggregation of cortical filamentous actin (F-actin); (b) induction of LFA-1-mediated cellular adhesion to the LFA-1 ligand intercellular adhesion molecule-1 (ICAM-1) through a mechanism involving clustering of LFA-1 molecules, rather than alterations in the level of expression or in the affinity state of this integrin; and (c) induction of cellular polarization and activation of the tyrosine kinase PYK2. Transfection of T-cells with a constitutively active form of Rho (V14Rho) blocks the clustering of LFA-1 on the membrane and the LFA-1-mediated activation of PYK2. Importantly, the activation of PYK2 caused by inhibition of Rho or ROCK takes place only when the T-cells are plated onto ICAM-1 but not when they are either prevented from interacting with ICAM-1 with anti-LFA-1 blocking antibodies or when they are plated on the nonspecific poly-l-lysine substrate. These results indicate that the small GTPase Rho regulates the tyrosine kinase PYK2 in T-cells through the F-actin-mediated control of the activity of the integrin LFA-1. These findings represent a novel paradigm for the regulation of the activity of a cytoplasmic tyrosine kinase by the small GTPase Rho.
We recently described the syntheses of 12a-c, 4-amino-7-oxo substituted analogues of 5-deaza-5,6,7,8-tetrahydrofolic acid (5-DATHF), and 5,10-dideaza-5,6,7,8-tetrahydrofolic acid (DDATHF), in six steps from commercially available p-substituted methyl benzoates in 20-27% overall yields. Such analogues were tested in vitro against CCRF-CEM leukemia cells and showed that they are completely devoid of any activity, the IC(50) being higher than 20 microg/mL for all cases. To clarify if the presence of the carbonyl group in position C7, the distinctive feature of our synthetic methodology, is the reason for this lack of activity, we have now obtained the 7-oxo substituted analogues of 5-DATHF and DDATHF, 18a-c, in 10-30% overall yield. Testing of 18a-c in vitro against CCRF-CEM leukemia cells revealed that these compounds are totally inactive. A molecular modeling study of 18b inside the active site of the complex E. coliGARTFase-5-DATHF-GAR pointed to an electronic repulsion between the atoms of the 7-oxo group and the carbonyl group of Arg90 as a possible explanation for the inactivity of 18a-c.
The chemokine stromal cell-derived factor-1alpha (SDF-1alpha) and its G-protein-linked receptor CXCR4 are involved in hematopoietic progenitor cell and lymphocyte migration. The integrin VLA-4 is a cell adhesion receptor for CS-1/fibronectin and VCAM-1 and constitutes one of the main adhesion receptors mediating myeloma cell adhesion to bone marrow (BM) stroma in multiple myeloma (MM). It is shown here that MM CD38(hi)CD45RA(-) BM cells and myeloma-derived cell lines expressed CXCR4 and displayed a moderate chemotactic response to SDF-1alpha. Because cell migration in response to SDF-1alpha might require a dynamic regulation of integrin function, it was investigated whether SDF-1alpha can modulate VLA-4 function on myeloma cells. SDF-1alpha rapidly and transiently up-regulated VLA-4-mediated myeloma cell adhesion to both CS-1/fibronectin and VCAM-1, which was inhibited by pertussis toxin and cytochalasin D, indicating the involvement of G(i) protein downstream signaling and an intact cytoskeleton. Modulation of VLA-4-dependent myeloma cell adhesion by SDF-1alpha could contribute to the trafficking and localization of these cells in the BM microenvironment.
Hydrolysis of the amino groups in condensed 2,4-diaminopyrimidine systems (1) has been used as a common method for the synthesis of oxo-substituted pyrimidines. In particular, the treatment with 6 M HCl usually yields exclusively the 2-amino-4-oxopyrimidine isomer (2). During our work, we found that the hydrolysis of the amino groups present in some condensed 2,4-diaminopyrimidine systems unexpectedly afforded exclusively the 4-amino-2-oxopyrimidine isomer (3). In this paper, we present the experimental work and ab initio calculations carried out to understand this discrepancy. As a part of such study, eight compounds containing a 2,4-diaminopyrimidine moiety were calculated in gas phase and in aqueous solution, and some acid hydrolyses were reexamined. Results showed that the presence of an electron-donating nitrogen linked to C6 of the 2,4-diaminopyrimidine ring changes the preferred hydrolysis site to yield the 4-amino-2-oxopyrimidine isomer.
Stromal cell-derived factor-1alpha (SDF-1alpha) is a potent chemoattractant for hematopoietic progenitor cells (HPC), suggesting that it could play an important role during their migration within or to the bone marrow (BM). The integrin VLA-4 mediates HPC adhesion to BM stroma by interacting with CS-1/fibronectin and VCAM-1. It is required during hematopoiesis and homing of HPC to the BM. As HPC migration in response to SDF-1alpha might require dynamic regulation of integrin function, we investigated if SDF-1alpha could modulate VLA-4 function on BM CD34(hi) cells.CD34(hi) BM cells and hematopoietic cell lines were tested for the effect of SDF-1alpha on VLA-4-dependent adhesion to CS-1/fibronectin and VCAM-1, as well as to BM stroma. CD34(hi) BM cells that adhered to VLA-4 ligands after SDF-1alpha treatment were characterized in colony-forming and long-term culture-initiating cell (LTC-IC) assays.SDF-1alpha rapidly (1 minute) and transiently upregulated the adhesion of CD34(hi) BM cells and hematopoietic cell lines to both CS-1/fibronectin and VCAM-1, and to BM stromal cells. The upregulation of VLA-4-dependent cell adhesion by SDF-1alpha targeted primitive LTC-IC as well as committed CD34(hi) cells. SDF-1alpha-triggered enhancement in VLA-4 function was inhibited by pertussis toxin (PTx) and cytochalasin D, indicating the involvement of G(i) protein downstream signaling and an intact cytoskeleton. Instead, activation of p44/42 MAP kinases by SDF-1alpha did not functionally correlate with enhancement of VLA-4-dependent cell adhesion. Modulation of VLA-4-mediated CD34(hi) BM cell adhesion by SDF-1alpha could play a key role in their migration within and to the BM and therefore influence their proliferation and differentiation.
In multiple myeloma, the malignant plasma cells in the bone marrow attach to stromal elements using several adhesion receptors. The integrin VLA-4 plays a major role in mediating myeloma cell adhesion to the stroma, by interacting with its ligands VCAM-1 and fibronectin. VLA-4, as well as other integrins, can be expressed in different states of activation, which convey different levels of adhesion. Modulation of VLA-4 adhesive activity by external stimuli has been demonstrated for hematopoietic progenitor cells and lymphocytes, and can represent a potential mechanism contributing to the localization of myeloma cells in the bone marrow. In this review we have summarized data on the characterization of VLA-4-mediated myeloma cell adhesion, and we present potential mechanisms of modulation of VLA-4 activity. In addition, we also speculate on the signalling generated upon interaction of myeloma VLA-4 with its ligands on the survival of these cells in the bone marrow.
Dendritic cells (DC) are highly specialized APC that are critical for the initiation of T cell-dependent immune responses. DC exert a sentinel function while immature and, after activation by inflammatory stimuli or infectious agents, mature and migrate into lymphoid organs to prime T cells. We have analyzed integrin expression on monocyte-derived DC (MDDC) and found that expression of CD49d integrins (CD49d/CD29 and CD49d/beta7) was induced/up-regulated during TNF-alpha- or LPS-initiated MDDC maturation, reflecting the induction/up-regulation of CD49d and beta7 mRNA. CD49d mRNA steady-state level increased more than 10 times during maturation, with the highest levels observed 24 h after TNF-alpha treatment. CD49d integrin expression conferred mature MDDC with an elevated capacity to adhere to the CS-1 fragment of fibronectin, and also mediated transendothelial migration of mature MDDC. Up-regulation of CD49d integrin expression closely paralleled that of the mature DC marker CD83. CD49d integrin expression was dependent on cell maturation, as its induction was abrogated by N:-acetylcysteine, which inhibits NF-kappaB activation and the functional and phenotypic maturation of MDDC. Moreover, CD49d integrin up-regulation and MDDC maturation were prevented by SB203580, a specific inhibitor of p38 mitogen-activated protein kinase, but were almost unaffected by the mitogen-activated protein/extracellular signal-related kinase kinase 1/2 inhibitor PD98059. Our results support the existence of a link between functional and phenotypic maturation of MDDC and CD49d integrin expression, thus establishing CD49d as a maturation marker for MDDC. The differential expression of CD49d on immature and mature MDDC might contribute to their distinct motility capabilities and mediate mature DC migration into lymphoid organs.
The integrin alpha(4)beta(7) is the cell adhesion receptor for the mucosal vascular addressin MAdCAM-1, and this interaction is dominant in lymphocyte homing to Peyer's patch high endothelial venules, and plays key roles in lymphocyte recruitment at sites of inflammation. To identify alpha(4) subunit amino acids important for alpha(4)beta(7)/MAdCAM-1 interaction, we expressed mutant alpha(4) and wild type beta(7) chains in K562 cells and analyzed the effect of the mutations on cell adhesion to a soluble MAdCAM-1 (sMAdCAM-1-Ig). Transfectants expressing mutated alpha(4) at Tyr(187) displayed a substantial decrease in adhesion to this ligand, which was associated with a reduced alpha(4)beta(7)/sMAdCAM-1-Ig interaction, as determined by soluble binding assays. Addition of Mn(2+) to the adhesion assays did not restore the impaired adhesion. Mutations at alpha(4) Gln(152)Asp(153) also affected transfectant adhesion to sMAdCAM-1-Ig, but did not involve an alteration of alpha(4)beta(7)/MAdCAM-1 binding, and adhesion was restored by Mn(2+). Instead, mutations at alpha(4) Asn(123)Glu(124) did not affect this adhesion. Mutation of alpha(4) Tyr(187) abolished alpha(4)beta(7)-mediated cell adhesion to CS-1/fibronectin, an additional ligand for alpha(4)beta(7), while alpha(4) Gln(152)Asp(153) transfectant mutants showed a reduced adhesion. These results identify alpha(4) Tyr(187) as a key residue during receptor alpha(4)beta(7)/ligand interactions, indicating that it plays important roles in alpha(4)beta(7)-mediated leukocyte adhesion, and provide a potential target for therapeutic intervention in several inflammatory pathologies.
Treatment of 2, solid supported synthetic equivalent of 3-formylchromone (4), and ethyl acetoacetate affords the salicylate structure 8 instead of the previously reported isophtalate 7. This is the first formation of a salicylate by a double carbonyl condensation of a malondialdehyde moiety ever reported.
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In this study we describe photokilling properties and effects on the mitotic index (MI) of cultured HeLa cells induced by palladium(II)-tetraphenylporphycene (PdTPP(0)). The drug was synthesized by refluxing tetraphenylporphycene (TPP(0)) and PdCl2 in dimethylformamide, followed by evaporation and purification by chromatography. Cells were treated with different concentrations of PdTPPo incorporated into dipalmitoylphosphatidylcholine liposomes, and red light irradiation (lambda > 600 nm) was performed at 21 mW/cm2. No dark toxicity was found when the drug was applied under our experimental conditions. Using lethal (LD(100)) treatments, cells showed the immediate occurrence of bubbles on the plasma membrane, whereas homogeneous nuclear condensation and loss of cytoplasm appeared 3-24 h later. An increased MI was found 6-8 h after sublethal LD(25) and LD(40) treatments as well as a high proportion of abnormal metaphases with altered spindle microtubules. Chromatin condensation and fragmentation were observed 8 h after LD(75) treatments. These results show that in comparison with TPP(0), the new sensitizer PdTPPo has more efficient photokilling properties which could be very valuable for the photodynamic therapy of cancer.
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The integrin VLA-4 mediates attachment of myeloma cells to multiple myeloma (MM) bone marrow stroma. The alternatively-spliced CS-1 region of fibronectin (FN) and VCAM-1 are main ligands for VLA-4 and are both expressed on MM stroma. The H1 region is present in all FN isoforms and represents an additional binding site for VLA-4. We employed FN fragments FN-H89 and FN-H0, that contain either the CS-1 and H1, or only the H1 sites, respectively, as well as soluble VCAM-1 (sVCAM-1), to characterize VLA-4-mediated adhesion pathways used by myeloma cells to attach to MM stroma. CD38highCD45RA- cells from MM bone marrow, and the myeloma-derived cell lines NCI-H929, IM-9 and RPMI 8226, specifically adhered, by different degrees, to FN-H89, FN-H0 and sVCAM-1, and their VLA-4-dependent adhesion was substantially up-regulated by the anti-beta1 antibody TS2/16, which increases the affinity of VLA-beta1 integrins. Furthermore, VLA-4 function on NCI-H929 cells was enhanced by TS2/16 during adhesion to MM stroma. The alpha4beta7 integrin mediated a small portion of myeloma cell line adhesion to FN-H89, mainly upon integrin activation with Mn2+. These results indicate that myeloma cells use VLA-4 to interact with CS-1/FN, H1/FN and VCAM-1 on MM stroma, and that its function can be potentially up-regulated, enabling higher degrees of cell adhesion to these VLA-4 ligands, which might influence myeloma cell localization in the bone marrow.