The small heat shock proteins, HSPB1 and HSPB5, interact differently with lipid membranes.

Increasing evidence shows that heat shock proteins (hsp) escape the cytosol gaining access to the extracellular environment, acting as signaling agents. Since the majority of these proteins lack the information necessary for their export via the classical secretory pathway, attention has been focused on alternative releasing mechanisms. Crossing the plasma membrane is a major obstacle to the secretion of a cytosolic protein into the extracellular milieu. Several mechanisms have been proposed, including direct interaction with the plasma membrane or their release within extracellular vesicles (ECV). HSPB1 (Hsp27), which belongs to the small hsp family, was detected within the membrane of ECV released from stressed HepG2 cells. To further investigate this finding, we studied the interaction of HSPB1 with lipid membranes using liposomes. We found that HSPB1 interacted with liposomes made of palmitoyl oleoyl phosphatidylserine (POPS), palmitoyl oleoyl phosphatidylcholine (POPC), and palmitoyl oleoyl phosphatidylglycerol (POPG), with different characteristics. Another member of the small hsp family, HSPB5 (αB-crystallin), has also been detected within ECV released from HeLa cells transfected with this gene. This protein was found to interact with liposomes as well, but differently than HSPB1. To address the regions interacting with the membrane, proteoliposomes were digested with proteinase K and the protected domains within the liposomes were identified by mass spectroscopy. We observed that large parts of HSPB1 and HSPB5 were embedded within the liposomes, particularly the alpha-crystallin domain. These observations suggest that the interaction with lipid membranes may be part of the mechanisms of export of these proteins.

Evidence against a role of P-glycoprotein in the clearance of the Alzheimer's disease Aß1-42 peptides.

It has been proposed that the amyloid-ß peptides (Aß) cause the neuronal degeneration in the Alzheimer's disease brain. An imbalance between peptide production at the neuronal level and their elimination across the blood-brain-barrier (BBB) results in peptide accumulation inside the brain. The identification and functional characterization of the transport systems in the BBB with the capacity to transport Aß is crucial for the understanding of Aß peptide traffic from the brain to the blood. In this context, it has been suggested that the P-glycoprotein (P-gp), expressed in endothelial cells of the BBB, plays a role in the elimination of Aß. However, there is little, if any, experimental evidence to support this; therefore, the aim of this investigation was to determine whether P-gp is capable of transporting Aß peptides. Our results show that ATPase activity measured in plasma membrane vesicles of K562 cells overexpressing P-gp is not increased by the presence of Aß42, suggesting that Aß42 is not a P-gp substrate. Similarly, P-gp of pirarubicin was unaffected by Aß42. Moreover, the overexpression of P-gp does not protect cells against Aß42 toxicity. Taken together, our results support the conclusion that Aß42 is not transported by P-gp.

Multidrug resistance (MDR) reversers: High activity and efficacy in a series of asymmetrical N,N-bis(alkanol)amine aryl esters.

As a continuation of our research on potent and efficacious P-gp-dependent multidrug resistance (MDR) reversers, several new N,N-bis(alkanol)amine aryl esters were designed and synthesized, varying the aromatic moieties or the length of the methylenic chain. The new compounds were tested on doxorubicin-resistant erythroleukemia K562 cells (K562/DOX) in the pirarubicin uptake assay, where most of the new compounds were shown to be active. In particular the asymmetrical compounds, characterized by two linkers of different length, generally showed fairly high activities as MDR reversers. Some selected compounds (isomers 15-17) were further studied by evaluating their doxorubicin cytotoxicity enhancement (reversal fold, RF) on the K562/DOX cell line. The results of both pharmacological assays indicate that compounds 16 (GDE6) and 17 (GDE19) could be interesting leads for the development of new P-gp dependent MDR modulators.

New structure-activity relationship studies in a series of N,N-bis(cyclohexanol)amine aryl esters as potent reversers of P-glycoprotein-mediated multidrug resistance (MDR).

As a continuation of previous research on a new series of potent and efficacious P-gp-dependent multidrug resistant (MDR) reversers with a N,N-bis(cyclohexanol)amine scaffold, we have designed and synthesized several analogs by modulation of the two aromatic moieties linked through ester functions to the N,N-bis(cyclohexanol)amine, aiming to optimize activity and to extend structure-activity relationships (SAR) within the series. This scaffold, when esterified with two different aromatic carboxylic acids, gives origin to four geometric isomers (cis/trans, trans/trans, cis/cis and trans/cis). The new compounds were tested on doxorubicin-resistant erythroleukemia K562 cells (K562/DOX) in the pirarubicin uptake assay. Most of them resulted in being potent modulators of the extrusion pump P-gp, showing potency values ([I](0.5)) in the submicromolar and nanomolar range. Of these, compounds 2b, 2c, 3d, 5a-d and 6d, showed excellent efficacy with a α(max) close to 1. Selected compounds (2d, 3a, 3b, 5a-d) were further studied to evaluate their doxorubicin cytotoxicity potentiation (RF) on doxorubicin-resistant erythroleukemia K562 cells and were found able to enhance significantly doxorubicin cytotoxicity on K562/DOX cells. The results of both pirarubicin uptake and the cytotoxicity assay, indicate that the new compounds of the series are potent P-gp-mediated MDR reversers. They present a structure with a mix of flexible and rigid moieties, a property that seems critical to allow the molecules to choose the most productive of the several binding modes possible in the transporter recognition site. In particular, compounds 5c and 5d, similar to the already reported analogous isomers 1c and 1d,(29) are potent and efficacious modulators of P-gp-dependent MDR and may be promising leads for the development of MDR-reversal drugs.

Strengths and vulnerabilities of a sample of gay and bisexual male adolescents in Puerto Rico

La adolescencia es un proceso de desarrollo de múltiples dimensiones que incluyen los significados sociales que se le atribuyen colocando a la juventud en un limbo social; no son niños o niñas pero todavia no son adultos. Es importante contextualizar las dificultades que enfrentan los adolescentes gay debido a su identidad sexual, su vulnerabilidad social asociada a su particular etapa de desarrollo y su etnicidad. Exploramos las vulnerabilidades y fortalezas de una muestra de jóvenes adolescentes gay puertorriqueños. Participaron 61 jóvenes gay y bisexuales de alto nivel educativo, que residían en Puerto Rico. Examinamos los niveles de depresión, el apoyo social percibido, el uso de alcohol y drogas y la conducta sexual. Los resultados demuestran que el 45 por cento de los participantes informaron altos niveles de depresión. Sin embargo los participantes también mostraron un bajo consumo de alcohol y drogas, poca o ninguna actividad sexual de riesgo y una gran satisfacción con el apoyo social recibido. Los participantes mostraron gran resiliencia asociada a sus redes de apoyo social, el uso consistente de protección en las conductas sexuales de alto riesgo y capacidad para integrar su orientación sexual en su desarrollo personal en una sociedad latina y heterosexista. Apesar de que los resultados no pueden generalizarse a toda la población de jóvenes adolescentes gay en Puerto Rico, esta información es útil para apoyar la necesidad de intervenciones a nivel de comunidad que manejen las fortalezas de esta población

Strengths and vulnerabilities of a sample of gay and bisexual male adolescents in Puerto Rico

La adolescencia es un proceso de desarrollo de múltiples dimensiones que incluyen los significados sociales que se le atribuyen colocando a la juventud en un limbo social; no son niños o niñas pero todavia no son adultos. Es importante contextualizar las dificultades que enfrentan los adolescentes gay debido a su identidad sexual, su vulnerabilidad social asociada a su particular etapa de desarrollo y su etnicidad. Exploramos las vulnerabilidades y fortalezas de una muestra de jóvenes adolescentes gay puertorriqueños. Participaron 61 jóvenes gay y bisexuales de alto nivel educativo, que residían en Puerto Rico. Examinamos los niveles de depresión, el apoyo social percibido, el uso de alcohol y drogas y la conducta sexual. Los resultados demuestran que el 45 por cento de los participantes informaron altos niveles de depresión. Sin embargo los participantes también mostraron un bajo consumo de alcohol y drogas, poca o ninguna actividad sexual de riesgo y una gran satisfacción con el apoyo social recibido. Los participantes mostraron gran resiliencia asociada a sus redes de apoyo social, el uso consistente de protección en las conductas sexuales de alto riesgo y capacidad para integrar su orientación sexual en su desarrollo personal en una sociedad latina y heterosexista. Apesar de que los resultados no pueden generalizarse a toda la población de jóvenes adolescentes gay en Puerto Rico, esta información es útil para apoyar la necesidad de intervenciones a nivel de comunidad que manejen las fortalezas de esta población (AU)

Evaluation of an HIV/AIDS prevention intervention targeting Latino gay men and men who have sex with men in Puerto Rico.

This article presents the evaluation of an HIV prevention model and intervention targeting Latino gay males and men who have sex with men in Puerto Rico. Based on the health belief model in combination with concepts of self-efficacy, cognitive theory, and community development, a series of workshops were developed to enhance safer sex behavior among participants. A total of 587 men recruited by availability in gay meeting places and through peer referral participated in the intervention between 1992 and 1995. Participants engaged in a 3-hour small group meeting and four 3-hour workshops as part of the intervention. Pretests and posttests were administered to measure knowledge, attitudes, and behaviors related to HIV infection; a risk index was calculated to measure the level of behavior risk for HIV infection. Participants demonstrated lower risk indexes after the intervention. This model represents a culturally competent and relevant intervention for similar communities.