Telefarmacia como estrategia de dispensación coordinada entre Farmacia Hospitalaria y Atención Primaria / Telepharmacy as a strategy for co-ordinated dispensing between Hospital Pharmacy and Primary Care

A partir de la publicación de la Orden Ministerial SND/293/2020,del 25 de marzo, tras la declaración de la emergencia sanitariapor SARS-CoV-2 en España, se diseñaron una serie de medidasque garantizaran la restricción de movilidad de la población sinperjuicio del acceso a los medicamentos hospitalarios. Así, a raízde dicha publicación, en el Servicio de Farmacia Hospitalaria delHospital SAS La Línea se desarrolló un programa de Telefarmaciacon puntos de dispensación en los centros de Atención Primariaadscritos y una consulta telefónica de Atención Farmacéutica.Se realizaron un total de 1.007 dispensaciones en 301 pacientesdurante el periodo de estudio comprendido entre el 1 de julio al15 de noviembre de 2020. Así, al finalizar dicho periodo, 235fueron los pacientes que permanecieron incluidos en el programa de Telefarmacia con una edad media de 64 años y un54,5% (128) mujeres. A estos pacientes, se les enviaron por correo postal una encuesta de satisfacción que podrían devolvervoluntaria y anónimamente al Servicio de Farmacia. Se recibieronun total de 62 encuestas, mostrándose los pacientes satisfechosen un 96,77% (60) con el servicio de entrega, 90,32% (56) conel trato recibido, 87,10% (54) con la puntualidad y 98,39% (61)con las condiciones de conservación en el Centro de Salud.El alto grado de satisfacción de los pacientes encuestadosrefleja que el nuevo programa de Telefarmacia podrían responder a las necesidades individuales de los pacientes,siendo para ello fundamental la coordinación de todos losprofesionales implicados, así como la corresponsabilidad delos pacientes en el control de sus tratamientos.Con los resultados obtenidos en el presente estudio, parece justificado mantener en el futuro los centros de atención primariacomo punto de dispensación junto con un mayor desarrollo dela consulta no presencial de Atención Farmacéutica. (AU)

As of the publication of Ministerial OrderSND/293/2020, of March 25, after thedeclaration of the health emergency dueto SARS-CoV-2 in Spain, a series of measures were designed to guarantee therestriction of population mobility withoutprejudice to access to hospital medications. Thus, as a result of said publication,a telepharmacy program was developedat the clinical pharmacy service of theHospital SAS La Línea with dispensingpoints in the attached Primary Care centers and a Pharmaceutical Care Telephone Consultation.A total of 1,007 dispensations weremade in 301 patients during the studyperiod from July 1 to November 15,2020. Thus, at the end of that period,235 were the patients who remainedincluded in the Telepharmacy programwith a mean age 64 years and 54.5%(128) women. A satisfaction survey wassent to these patients by post, whichthey could return voluntarily and anonymously to the pharmacy service. A totalof 62 surveys were received, showing96.77% (60) satisfied patients with thedelivery service, 90.32% (56) with theattention received, 87.10% (54) withthe punctuality and 98.39% (61) withthe conditions of conservation in thehealth center.The high degree of satisfaction of thesurveyed patients reflects that the newTelepharmacy program could respondto the individual needs of the patients,being essential for this the coordinationof all the professionals involved, as wellas the co-responsibility of the patientsin the control of their treatments.With the results obtained in the present study, it seems justified in the future to maintain primary care centersas a dispensing point together with afurther development of the non-faceto-face consultation of PharmaceuticalCare. (AU)

Indirect comparison of biological treatments in refractory rheumatoid arthritis.

WHAT IS KNOWN AND OBJECTIVE: A number of biological treatments are available for rheumatoid arthritis. They are effective some patients but their comparative efficacy is inadequately evaluated. Our aim was to compare the efficacy of adalimumab, etanercept, infliximab, abatacept, tocilizumab, golimumab and certolizumab pegol in rheumatoid arthritis, refractory to disease-modifying antirheumatic drugs (DMARDs), through a systematic review of published trials. METHODS: As there were no direct comparisons, we searched for studies with similar characteristics to identify trials with results suitable for indirect comparison. Randomized, placebo-controlled pivotal clinical trials, with reported American College of Rheumatology ACR50 data at 24/30 weeks as efficacy endpoint, approved clinical doses and patients resistant to DMARDs who had not previously received other biological treatments were included. ACR50 was defined as the primary endpoint for the indirect comparison, with ACR20 and ACR70 as secondary endpoints. When two or more trials on one same drug were available, and a combined analysis was performed when appropriate. In the indirect comparison, the Bucher adjusted method was used with etanercept as reference drug. In the equivalence study, the equivalence window was a response efficacy difference of 15% between the alternatives. RESULTS AND DISCUSSION: Ten trials were found suitable for detailed analysis. In the clinical trials, all the biological drugs were seen to be more effective than placebo. Indirect comparison based on the ACR50 efficacy criterion all biological treatments showed similar results within the defined equivalence Δ value. The absolute efficacy difference (reduction of absolute risk, RAR) versus etanercept being 2·6% with adalimumab, 14% with infliximab, 11·6% with abatacept, 3% with tocilizumab, 12·4% with golimumab and 6·5% with certolizumab pegol. WHAT IS NEW AND CONCLUSION: The biological drugs used in rheumatoid arthritis are no different in efficacy. Their therapeutic positioning depends on their relative safety and convenience profiles.

Coordinación de asignaturas en el plan de estudios como actividad formativa del equipo docente multidisciplinar para la Licenciatura de Farmacia / Coordination of subjects in the program as a formative activity of the multidisciplinary educational team for the degree in Pharmacy

La Universidad de Granada, dentro del Plan Estratégico y el Contrato Programa 2007-2011 de lasUniversidades Públicas de Andalucía puso en marcha la convocatoria de apoyo a la formación delprofesorado principiante y mejora de la docencia por el Vicerrectorado para la Garantía de la Calidad(http://calidad.ugr.es/pages/secretariados/form_apoyo_calidad/apoyo_formacion_principiante/convocatoria). En el proyecto participan 15 profesores, 5 profesores experimentados y 10 profesores novelesde seis departamentos diferentes.. Entre los objetivos se incluye la optimización de la actividaddocente de profesorado principiante. En este sentido se estudió la situación en el nuevo plan de estudiode Grado en Farmacia de las asignaturas que dichos profesores impartimos, así como la relación entreellas y la posible existencia de solapamientos de contenidos docentes. Las fichas docentes muestranciertos solapamientos de competencias y carencias en algunos casos. Esto supuso una revisiónexhaustiva para llevar a cabo una correcta coordinación entre los profesores que le permita alalumnado un aprendizaje organizado y coherente(AU)

The University of Granada, inside the Strategic Plan and the Contract Program 2007-2011 of thePublic Universities of Andalusia started a public call to improve the formation of young lecturers andthe teaching activity(http://calidad.ugr.es/pages/secretariados/form_apoyo_calidad/apoyo_formacion_principiante/convocatoria). In the project there take part 15 lecturers, 5 experienced and 10 young lecturers from sixdifferent departments. Within the objectives it is included the optimisation of the teaching activity ofyoung lecturers. In this sense, it was studied the situation of the subjects that the above mentionedlecturers give within the new degree in Pharmacy, as well as the relation between they and the possibleexistence of any overlapping in the contents. The teaching contents showed the existence of certainoverlapping within competences and deficiencies in some cases. This supposed an exhaustive reviewto carry out the correct coordination between the different lecturers so that it will allow the organizedand coherent learning of the students(AU)

Evaluation of hepatic damage and local immune response in goats immunized with native glutathione S-transferase of Fasciola hepatica.

Worm burden, hepatic damage and local cellular and humoral immune responses were assessed in goats immunized with glutathione-S-transferase and challenged with Fasciola hepatica. Infected but unimmunized and uninfected control groups were also studied. Hepatic damage was evaluated grossly and microscopically. Local immune response was evaluated by (1) microscopical examination of hepatic lymph nodes (HLNs); (2) analysis of the distribution of CD2(+), CD4(+), CD8(+), T-cell receptor gammadelta(+) lymphocytes and immunoglobulin (Ig) G(+) plasma cells; and (3) investigation of the distribution of cells expressing interleukin (IL)-4 and interferon (IFN)-gamma in the hepatic inflammatory infiltrates and HLNs. Immunized animals did not have significant reduction in fluke number, but there was significant (P<0.05) reduction of fluke size relative to the control groups. The lesions in the two infected groups were similar and consisted of fibrous perihepatitis and white tortuous tracts, mainly involving the left hepatic lobe. Microscopical lesions were similar in both infected groups and were typical of chronic fascioliosis. These included portal fibrosis, inflammatory infiltration with plasma cells, formation of lymphoid follicles, accumulation of haemosiderin-laden macrophages and granulomatous foci. Both infected groups had a marked local immune response characterized by infiltration of CD2(+), CD4(+) and CD8(+) T lymphocytes, and IgG(+) plasma cells in hepatic lesions and in HLNs. There was no expression of IL-4 or INF-gamma by cells in the hepatic inflammatory infiltrate, but expression of INF-gamma in HLNs was much lower than that of IL-4, suggesting an immune response dominated by T helper 2 cells.

Protective effects of Rosmarinus tomentosus ethanol extract on thioacetamide-induced liver cirrhosis in rats.

The capability of an ethanol extract of Rosmarinus tomentosus to protect rat liver in an experimental model of cirrhosis induced by thioacetamide (TAA) has been evaluated. Four groups of rats were used: Two of them received 300 mg TAA/l in the drinking water for 3 months while the other two, which served as controls, were given water ad libitum. During the same period and for each one of the treatments, one group received a semi-purified (SP) diet and the other one was fed the same diet supplemented with 1% of the dry residue obtained from R. tomentosus ethanol extract (SP+E). There was a significant reduction of TAA toxicity in rats fed the SP+E diet, as assessed by plasma and liver biochemical markers, and by liver histopathology. Plasma total protein concentration was restored, urea concentration and plasma alkaline phosphatase and gamma-glutamyl-transferase activities were reduced. A significant correction of plasma fatty acids concentrations was also evident. Hepatic alkaline phosphatase and gamma-glutamyl-transferase activities were significantly reduced in animals fed SP+E diet and glucose-6-phosphatase activity was significantly enhanced. The results suggest that R. tomentosus ethanol extract administered in the diet affords protection against TAA-induced cirrhosis, preventing most of the histological changes and functionality alterations own to this experimental pathology.

Antibacterial, antiprotozoal and antioxidant activity of five plants used in Izabal for infectious diseases.

Methanol and aqueous extracts from fi ve plant species, used in traditional medicine in Guatemala for the treatment of microbial infections, were tested in vitro for their ability to scavenge DPPH, OH(.) and O(2) (-) radicals and to inhibit lipoperoxidation (LPO) in order to establish a relationship between their antioxidant activities and their effects against infectious agents. Acalypha guatemalensis, Ocimum micranthum and Smilax spinosa possessed a significant activity against both the three free radicals assayed and LPO; Guazuma ulmifolia showed effects against DPPH and OH(.). Piper auritum showed no activity. These extracts were also evaluated for antibacterial and antiprotozoal activities. A. guatemalensis showed activity against Pseudomonas aeruginosa; S. spinosa was active against Salmonella typhi, and A. guatemalensis, and S. spinosa against Trypanosoma cruzi or Leishmania spp.

Effects of chronic quercetin treatment on hepatic oxidative status of spontaneously hypertensive rats.

The effects of chronic administration of an oral daily dose of quercetin (10 mg Kg(-1)), the most abundant dietary flavonoid, were investigated on hepatic oxidative status in spontaneously hypertensive rats and normotensive Wistar Kyoto rats. Decreased liver glutathione peroxidase activity, increased liver total glutathione levels and increased both hepatic and plasmatic malondialdehyde concentrations were observed in spontaneously hypertensive rats when compared to Wistar Kyoto rats. In spontaneously hypertensive rats, treatment with quercetin for 5 weeks reduced blood pressure, increased glutathione peroxidase activity and reduced both plasma and hepatic malondialdehyde levels. However, none of these effects were observed in Wistar Kyoto rats. In conclusion, quercetin shows both antihypertensive and antioxidant properties in this model of genetic hypertension.

Src and Pyk2 mediate G-protein-coupled receptor activation of epidermal growth factor receptor (EGFR) but are not required for coupling to the mitogen-activated protein (MAP) kinase signaling cascade.

The epidermal growth factor receptor (EGFR) and the non-receptor protein tyrosine kinases Src and Pyk2 have been implicated in linking a variety of G-protein-coupled receptors (GPCR) to the mitogen-activated protein (MAP) kinase signaling cascade. In this report we apply a genetic strategy using cells isolated from Src-, Pyk2-, or EGFR-deficient mice to explore the roles played by these protein tyrosine kinases in GPCR-induced activation of EGFR, Pyk2, and MAP kinase. We show that Src kinases are critical for activation of Pyk2 in response to GPCR-stimulation and that Pyk2 and Src are essential for GPCR-induced tyrosine phosphorylation of EGFR. By contrast, Pyk2, Src, and EGFR are dispensable for GPCR-induced activation of MAP kinase. Moreover, GPCR-induced MAP kinase activation is normal in fibroblasts deficient in both Src and Pyk2 (Src-/-Pyk2-/- cells) as well as in fibroblasts deficient in all three Src kinases expressed in these cells (Src-/-Yes-/-Fyn-/- cells). Finally, experiments are presented demonstrating that, upon stimulation of GPCR, activated Pyk2 forms a complex with Src, which in turn phosphorylates EGFR directly. These experiments reveal a role for Src kinases in Pyk2 activation and a role for Pyk2 and Src in tyrosine phosphorylation of EGFR following GPCR stimulation. In addition, EGFR, Src family kinases, and Pyk2 are not required for linking GPCRs with the MAP kinase signaling cascade.

The induction of the human hepatic CYP2E1 gene by interleukin 4 is transcriptional and regulated by protein kinase C.

Cytochrome P4502E1 (CYP2E1) plays a key role in the metabolism of numerous drug substrates, mostly in mammalian liver. Both the apoprotein and mRNA levels are increased in response to interleukin 4 (IL-4) in primary human hepatocyte cultures. We developed a human hepatoma cell model that faithfully reproduces the responsiveness of the CYP2E1 gene to IL-4 at least in part through transcriptional activation, upon treatment with 150 U/ml of IL-4. As expected, IL-4 induced tyrosine phosphorylation of the STAT6 transcription factor, an effect prevented by the tyrosine kinase inhibitor tyrphostin A25. However, this inhibitor as well as genistein (another inhibitor of tyrosine kinases) had no effect on the IL-4 induction of CYP2E1. Similarly, protein kinase A activators (forskolin and dibutyryl-cAMP) and inhibitor (H89) did not influence the response to IL-4. However, PKC inhibitors (H7 and calphostin C) strongly blocked any induction of the gene, as well as the IL-4-dependent translocation of PKCS. Taken together, our results show that IL-4 coordinately induces CYP2E1 transcription, mRNA and apoprotein levels in human hepatoma cells in a PKC-dependent manner, potentially through the activity of the PKCzeta isoform.

Antihepatotoxic activity of Rosmarinus tomentosus in a model of acute hepatic damage induced by thioacetamide.

R. tomentosus is a vegetal species closely related to the culinary rosemary (R. officinalis), a plant reported to contain antihepatotoxic agents. A dried ethanol extract of the aerial parts of Rosmarinus tomentosus (Lamiaceae) and its major fraction separated by column chromatography (fraction F19) were evaluated for antihepatotoxic activity in rats with acute liver damage induced by a single oral dose of thioacetamide. Silymarin was used as a reference antihepatotoxic substance. Pre-treatment with R. tomentosus ethanol extract, fraction F19 or silymarin significantly reduced the impact of thioacetamide toxicity on plasma protein and urea levels as well as on plasma aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase and gamma-glutamyl transpeptidase activities compared with thioacetamide-treated animals (group T). Pre-treatment with R. tomentosus ethanol extract significantly reduced the impact of thioacetamide damage on alkaline phosphatase and gamma-glutamyl transpeptidase activities compared with group T. Silymarin administration significantly reduced alkaline phosphatase and gamma-glutamyl transpeptidase activities compared with group T. Fraction F19 administration reduced only alkaline phosphatase activity compared with group T. According to these data, R. tomentosus extract shows promising antihepatotoxic activity, suggesting the need to isolate the chemical principles responsible for this activity and to study this activity in a model of thioacetamide-induced cirrhosis.

Hepatotoxicity of tacrine: occurrence of membrane fluidity alterations without involvement of lipid peroxidation.

Tacrine (THA), used in the treatment of Alzheimer's disease, is known to induce hepatotoxicity, the mechanisms of which remain to be fully established. We have previously shown that THA reduced intracellular glutathione concentration in rat hepatocytes in primary culture, thus pointing to a possible role for oxidative stress in THA toxicity. To test this, the effects of antioxidant molecules, namely, the flavonoids silibinin, silibinin dihydrogensuccinate, and silymarin, were evaluated on the toxicity of THA in cultured rat hepatocytes. This toxicity was investigated after a 24-h treatment over a concentration range from 0 to 1 mM, in the presence or absence of antioxidant (1 and 10 microM). We found that simultaneous treatment of hepatocytes with any of the antioxidants and THA remained ineffective on the lactate dehydrogenase release induced by THA. Then, the production of lipid-derived radicals (to estimate lipid peroxidation) was measured in THA (0.05-0.50 mM)-treated cells using a spin-trapping technique coupled to electron paramagnetic resonance (EPR) spectroscopy. No increase of the EPR signal was observed over the period of 30 min to 24 h. In contrast, treatment of cells with the spin label 12-doxyl stearic acid followed by EPR spectroscopy showed that THA (0.05 and 0.25 mM) rapidly increased hepatocyte membrane fluidity. Extracellular application of GM1 ganglioside (60 microM) both reversed this increase in fluidity and partially reduced lactate dehydrogenase release on THA exposure. In conclusion, this work indicates that early alterations of membrane fluidity, not resulting from lipid peroxidation, are likely to play an important role in the development of THA toxicity.

Regulation of phenobarbital induction of the cytochrome P450 2b9/10 genes in primary mouse hepatocyte culture. Involvement of calcium- and cAMP-dependent pathways.

Phenobarbital (PB) has long been known as an inducer of drug-metabolizing enzymes in liver, but the molecular mechanism underlying this induction is still poorly understood. Using primary mouse hepatocyte culture, we have investigated the possible involvement of different regulatory pathways in PB action, by exposing PB-treated cells to various protein kinase/phosphatase modulators. Our results showed a negative role of the cAMP-dependent pathway, as treatment with cAMP-dependent protein kinase (PKA) activators (10 microM dibutyryl-cAMP and 50 microM forskolin) dramatically inhibited PB-induced Cyp2b9/10 mRNA accumulation, whereas PKA inhibitor potentiated the PB responsiveness of this gene. The cGMP-dependent protein kinase (PKG) seems to play a positive role as PKG inhibitor reduced the PB-induced level of Cyp2b9/10 mRNA. We also obtained two lines of evidence for the involvement of Ca2+ in modulating PB action. Firstly, measurements of intracellular Fura-2 fluorescence ratio in murine hepatocytes showed that long-term PB incubation (24 and 48 h) led to a significant increase of [Ca2+]i. Secondly, treatment with an intracellular Ca2+ chelator (BAPTA-AM) nearly completely abolished PB-induced Cyp2b9/10 expression. Ca2+ thus appeared to mediate PB action likely via Ca2+/calmodulin-dependent protein kinase II, as KN62, a specific inhibitor of this enzyme, also dramatically inhibited PB induction of the Cyp2b9/10 genes.

Regulation of phenobarbital-induction of CYP2B and CYP3A genes in rat cultured hepatocytes: involvement of several serine/threonine protein kinases and phosphatases.

We investigated the involvement of diverse protein kinases and phosphatases in the transduction pathways elicited by phenobarbital (PB), a well-known inducer of some hepatic cytochromes P450 (CYP). Different inhibitors or activators of protein kinases or phosphatases were assessed for their ability to modulate PB-induction of CYP2B and CYP3A mRNA expression. Rat hepatocytes in primary culture were treated with the test compounds one hour prior to, and then continuously, in the absence or presence of 1 mmol/L PB for 24 h. By northern blot analysis of CYP2B1/2 and 3A1/2 gene expression, we first confirmed the negative role of the adenosine 3':5' cyclic monophosphate (cAMP)/protein kinase A pathway and the positive role of some serine/threonine protein phosphatases in the mechanism of PB-induction. The present data further suggested that Ca2+/calmodulin-dependent protein kinases II (independently of Ca2+) and extracellular signal-regulated kinases 1/2 (ERK1/2) might function respectively as positive and negative regulator in the PB-induction of CYP2B and CYP3A. In contrast, protein kinases C and phosphatidylinositol-3-kinase did not appear to be involved, while the role of tyrosine kinases remained unclear. We conclude that a complex network of phosphorylation/dephosphorylation events might be crucial for PB-induction of rat CYP2B and CYP3A.

Intracellular pH alterations induced by tacrine in a rat liver biliary epithelial cell line.

1. The effects of tacrine (THA) on intracellular pH (pH(i)) were examined in a rat liver biliary epithelial cell line (RLEC) in HEPES-buffered medium. pH(i) was recorded using the pH-sensitive fluoroprobe, carboxy-SNARF-1 (carboxy-seminaphtorhodafluor). 2. In the steady state, short-term exposures to THA resulted in alkalinization and re-acidification at 0.1 and 0.25 mM. Following a 24 h-treatment, no significant difference in pH(i) could be detected at 0.1 and 0.25 mM THA, whereas at 0.05 mM, pH(i) was slightly more acid (7.17+/-0. 02, n=16 versus 7.21+/-0.02, n=24 [control]). 3. In control and short-term treated cells, intracellular intrinsic buffering power (beta(i)) increased roughly linearly as pH(i) decreased. This dependence was not seen following long-term treatment. In all cases, beta(i) was increased by THA (by 1.6 to 3.5 fold). 4. Following an acid load (induced by 20 mM NH(4)Cl removal), pH(i) recovery in RLEC relied upon Na(+)/H(+) exchange. A short-term treatment (0.25 mM THA) did not affect total acid extrusion. In contrast, a 24 h-treatment with 0.05 mM THA reduced it (by approximately 36% at a pH(i) of 6.73) while at 0.25 mM, a large increase was detected (by approximately 109% at a pH(i) of 6.75). In Na(+)-free medium, THA (0. 25 mM) still induced an alkalinization in the steady state. Following an acid load, THA stimulated a Na(+)-independent acid efflux in a dose-dependent manner, inhibitable by alpha-cyano-4-hydroxy cinnamate (CHC, 4 mM) but not by quercetin (0. 125 mM). 6. In conclusion, this work demonstrates that THA affects pH(i) in RLEC, through a decrease in Na(+)/H(+) exchange and an increase in beta(i). Stimulation of a CHC-inhibitable, Na(+)-independent acid efflux is also detected.

Involvement of cyclic nucleotide- and calcium-regulated pathways in phenobarbital-induced cytochrome P-450 3A expression in mouse primary hepatocytes.

Several of the hepatic microsomal cytochromes P-450 (CYP) including CYP3A are inducible by phenobarbital (PB). However, the intracellular pathways involved in the action of PB on CYP3A remain poorly known. With the aim to unravel some of the main aspects of PB signaling, we first devised a simple model of mouse cultured primary hepatocytes in which CYP3A mRNA and protein were strongly induced by PB in the absence of dexamethasone and were at maximum levels after a 48-h treatment with a 2-mM dose of PB. Under these culture conditions, we studied the effects of inhibitors and activators of different protein kinases or phosphatases on CYP3A mRNA and protein induction by PB. CYP3A-induced expression was inhibited by activators of cyclic AMP-dependent protein kinase (PKA) (dibutyryl-cyclic AMP and forskolin) whereas inhibition of PKA by PKA inhibitor enhanced induction. 8-br-cGMP produced effects similar to the activators of PKA, and so did the specific inhibitor of cGMP-dependent protein kinase, beta-phenyl-1, N(2)-etheno-8-bromoguanosine-3,5'-cyclic monophosphorothioate, Rp-isomer (Rp-8-Br-PET-cGMPS). Inhibition of Ca(2+)/calmodulin-dependent protein kinase by KN-62 or the intracellular Ca(2+) chelator BAPTA-AM produced an inhibition of CYP3A induction by PB. Specific inhibitors of protein kinase C, mitogen-activated protein kinase kinase, phosphatidylinositol-3-kinase, or serine/threonine phosphatase did not produce any effect. Taken together, our results suggest that CYP3A induction by PB is regulated positively by calmodulin-dependent protein kinase and cGMP-dependent protein kinase, and negatively by PKA in mouse hepatocytes in primary culture.

The proto-oncogene c-Cbl is a negative regulator of DNA synthesis initiated by both receptor and cytoplasmic tyrosine kinases.

In C. elegans, genetic and biochemical data indicate that the Cbl homolog Sli-1 attenuates Let-23 (EGFR) signaling. To investigate whether c-Cbhl might have a role in mammalian growth factor-mediated mitogenic signaling, we microinjected NIH3T3 mouse fibroblasts with expression plasmids encoding wt and G306ECbl (a 'loss of function' mutant identified in C. elegans). We observed inhibition of PDGF BB- and EGF-induced DNA synthesis by wt Cbl but not the mutant. Microinjection of two different affinity purified polyclonal antisera against Cbl boosted a suboptimal PDGF-stimulated mitogenic response. The inhibition of both PDGF BB- and EGF-induced DNA synthesis by wt Cbl was reversed by co-expression with Myc but not with Fos. DNA synthesis initiated by constitutively activated Src was also blocked by Cbl expression, but curiously by the G306E mutant as well. These data are all consistent with the proposition that Cbl negatively affects mitogenic signaling in mammalian fibroblasts.

[Do the complaints of primary care users vary with the care model?]. / Son diferentes las quejas de los usuarios de atención primaria según el modelo de atención?

OBJECTIVES: 1. To compare the traditional model (TM) and Primary Care teams (PCTs) on the index of complaints. 2. To determine whether the reasons for complaints vary with the care model. DESIGN: An observational, cross-sectional study. SETTING: Primary Care Area 4, INSALUD, Madrid, covering 526,987 inhabitants. PARTICIPANTS: The complaints presented at PC Centres during 1993 were included. Those presented for reasons not concerning the centre were excluded. In all, 448 complaints were studied. MEASUREMENTS AND MAIN RESULTS: Significant differences were found in the general index of complaints according to the care model. Statistically significant differences were detected between PCT and the TM for complaints due to disagreement with the treatment and disagreement with the follow-up procedures. CONCLUSIONS: Since the indicators of prescription profiles and the following of procedures are better in PCTs than in the TM, perhaps patients should be encouraged to participate more in the taking of clinical decisions. This would help the introduction of improvements both in the technical quality of treatment and in the follow-up of chronic diseases to be perceived as such by patients. At present, as this study suggests, they are causes of greater dissatisfaction.

The adaptor protein Nck links receptor tyrosine kinases with the serine-threonine kinase Pak1.

Nck is an adaptor protein composed of a single SH2 domain and three SH3 domains. Upon growth factor stimulation, Nck is recruited to receptor tyrosine kinases via its SH2 domain, probably initiating one or more signaling cascades. In this report, we show that Nck is bound in living cells to the serine-threonine kinase Pak1. The association between Nck and Pak1 is mediated by the second SH3 domain of Nck and a proline-rich sequence in the amino terminus of Pak1. We also show that Pak1 is recruited by activated epidermal growth factor (EGF) and platelet-derived growth factor receptors. Moreover, Pak1 kinase activity is increased in response to EGF in HeLa cells transfected with human Pak1, and the kinase activity was enhanced when Nck was co-transfected. It is concluded that Nck links receptor tyrosine kinases with Pak1 and is probably involved in targeting and regulation of Pak1 activity.

Tyrosine phosphorylation of the c-cbl proto-oncogene protein product and association with epidermal growth factor (EGF) receptor upon EGF stimulation.

The murine retroviral oncogene v-cbl induces pre-B cell lymphomas and myelogenous leukemias. The protein product of the mammalian c-cbl proto-oncogene is a widely expressed cytoplasmic 120-kDa protein (p120cbl) whose normal cellular function has not been determined. Here we show that upon stimulation of human epidermal growth factor (EGF) receptor, p12ocbl becomes strongly tyrosine-phosphorylated and associates with activated EGF receptor in vivo. A GST fusion protein containing amino acids 1-486 of p120cbl, including a region highly conserved in nematodes, binds directly to the autophosphorylated carboxyl-terminal tail of the EGF receptor. Platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), or nerve growth factor (NGF) stimulation also results in tyrosine phosphorylation of p120cbl. Recent genetic studies in Caenorhabditis elegans indicate that Sli-1, a p120cbl homologue, plays a negative regulatory role in control of the Ras signaling pathway initiated by the C. elegans EGF receptor homologue. Our results indicate that p120cbl is involved in an early step in the EGF signaling pathway that is conserved from nematodes to mammals.