Características de los pacientes con enfermedad crónica avanzada incluidos en un programa de cuidados paliativos domiciliario / Characteristics of patients with advanced chronic disease included in a domiciliary palliative care program

OBJETIVO: Conocer las características sociodemográficas y clínicas, y los aspectos relativos a información de los pacientes no oncológicos (PNO) en situación de enfermedad avanzada-terminal incluidos en un programa de cuidados paliativos domiciliario. Identificar las variables que pudieran estar relacionadas con el fallecimiento en el domicilio. Material y MÉTODO: Estudio descriptivo retrospectivo mediante revisión de historias clínicas de los PNO atendidos por un Equipo de Soporte de Atención Paliativa Domiciliaria durante el periodo comprendido entre noviembre de 2009 y marzo de 2013. Se analizaron datos sociodemográficos y clínicos del paciente y del cuidador principal, los conocimientos sobre el diagnóstico y el pronóstico del paciente y la familia, las preferencias para el cuidado y el fallecimiento de ambos y el lugar del fallecimiento. Para el análisis de los datos se utilizó el paquete informático SPSS(R) v.21. RESULTADOS: Se revisaron 371 historias, de las cuales 249 cumplieron CRITERIOS DE INCLUSIÓN: Los PNO representaron el 36,19% de la actividad, con una media de edad de 81,4 años. La enfermedad más frecuente fue la demencia (22,1%). El 67,5% de los pacientes procedían de Atención Primaria y del medio residencial. El motivo de derivación fue en un 85,5% para el control de síntomas, siendo los más frecuentes astenia, disnea y dolor. Los pacientes presentaron una comorbilidad alta, frecuentes síndromes geriátricos, algún grado de deterioro cognitivo en un 65% y deterioro funcional importante (índice de Barthel 21,52, Palliative Performance Scale 38,18). La cuidadora principal fue mayoritariamente mujer, hija del paciente y con sobrecarga para los cuidados en el 55,4%. Del grupo sin afectación cognitiva, un 83% conocían el diagnóstico, un 30% el pronóstico, el 95% preferían el domicilio para los cuidados y mantenían esta preferencia para el fallecimiento el 78%. El 66% del total de los pacientes falleció en domicilio/residencia. El análisis de los posibles factores relacionados con la muerte en el domicilio mostró resultados significativos cuando el ámbito geográfico para el cuidado era el medio rural y el paciente había manifestado su preferencia por fallecer en el domicilio. También cuando el cuidador principal no mostraba sobrecarga y tenía una percepción subjetiva de buena salud. CONCLUSIONES: Se identificó una población de PNO en situación de enfermedad avanzada caracterizada por edad elevada, alta carga sintomática, importante comorbilidad, deterioro funcional y alta mortalidad. Más de la mitad de los cuidadores presentaron sobrecarga para los cuidados. Se trata, por tanto, de enfermos con necesidades similares a las de los pacientes oncológicos avanzados. La prevalencia elevada de deterioro cognitivo supuso que solo la mitad de los pacientes fueron capaces de participar en la información y en la toma de decisioneS

OBJECTIVE: To describe the socio-demographic and clinic characteristics and aspects related to information of non-oncological patients (NOP) with advanced disease, which are included in a palliative-home care support unit. To identify the variables that could be associated with the death of NOP at home. MATERIAL AND METHOD: A retrospective descriptive study was performed by reviewing the clinical histories of NOP attended by a Palliative-home Care Support Unit during the period from November 2009 to March 2013. An analysis was performed on the socio-demographic and clinical data of the patients and their main caregivers, as well as the analysed, diagnostic and prognostic information provided by the family and the patient, preferences about end of life care of both parties, and the place of death. The data analysis was performed with the computer pack SPSS(R) v.21. RESULTS: Of the 371 clinical histories reviewed, 249 patients met the inclusion criteria. The NOP represented the 36.19% of the activity, with a mean age of 81.4 years. The most common disease was dementia (22.1%). More than two-thirds (67.5%) of the patients came from Primary Care and residential homes for the elderly. The main reason for referral was to control symptoms in 85.5%, with the most frequent being asthenia, dyspnoea, and pain. A high comorbidity, geriatric symptoms, cognitive impairment was observed in 65%, and an important functional impairment (mean Barthel Index of 21.52, and Palliative Performance Scale score of 38.18). The majority of main caregivers were women, and the patient's daughter, and the burden of caring was identified in 55.4%. In the group without cognitive impairment, the diagnosis was known by 83%, and 30% knew the prognosis. Staying at home to receive end of life care was preferred by 95%, and 78% kept this preference for dying. Two-thirds (66%) of all the patients died at home or in residential homes for the elderly. The statistical analysis of the possible factors associated with dying at home showed a significant odds ratio when the patient lived in rural areas, and preferred home as the place of death. Another factor is when the main caregiver does not suffer care burden and has a subjective perception of good health. CONCLUSIONS: The predominance of NOP in end of life situations is characterised by advanced age, high comorbidity, increased symptomatic burden, significant overall functional impairment on admission, and high mortality. More than the half of the caregivers showed excessive burden of caring. In conclusion, they are patients with similar needs to those with advanced oncological disease. The high prevalence of cognitive impairment suggested that only half of the patients were able to participate in the information and in the advanced care planning

Co-culture of the 55-6 B cell hybridoma with the EL-4 thymoma cell. Effect on cell growth and monoclonal antibody production.

The cell growth and monoclonal antibody production of the 55-6 hybridoma cell co-cultured with the murine thymoma cell line EL-4 at different initial 55-6:EL-4 ratios were investigated. Both populations were seeded in co-culture without previous stimulation and therefore with low constitutive CD40 and CD40 ligand (CD154) expression levels, and in the absence of exogenous co-stimuli. Viable cell density and growth rate data seem to suggest a competition for nutrients, which is detrimental for both cells in terms of biomass production and also of growth rate for 55-6. Final concentrations of antibody and specific antibody production rates were affected by the initial 55-6:EL-4 ratio. The 4:1 ratio yielded the highest IgG2a concentration, whereas the highest specific antibody production rate was obtained at the 2:1 ratio. Changes mainly in CD154 and also in CD40 expression in co-cultures could suggest cross-talk between both populations. In conclusion, different types of interactions are probably present in this co-culture system: competition for nutrients, cognate interaction and/or autocrine or paracrine interactions that influence the proliferation of both cells and the hybridoma antibody secretion. We are hereby presenting a pre-scale-up process that could speed up the optimization of large-scale monoclonal antibodies production in bioreactors by emulating the in vivo cell-cell interaction between B and T cells without previous stimulation or the addition of co-stimulatory molecules.

Cytotoxicity of yessotoxin and okadaic acid in mouse T lymphocyte cell line EL-4.

Yessotoxin (YTX) and okadaic acid (OA), algal toxins accumulated in edible shellfish, were previously shown to induce a specific and reversible T Cell Receptor (TCR) down-regulation in T lymphocyte EL-4 cells, in a time and concentration-dependent manner, via protein kinase C (PKC) and serine/threonine protein phosphatase 2A (PP2A) activities. In this study we have evaluated the development of other signs of toxicity induced by low concentrations of YTX or OA for 3 days of treatment. Concentrations of YTX as low as 1 nM decreased a 35% the concentration of viable cells after 48 h exposure to the toxin, while concentrations as little as 5 nM YTX or OA were sufficient to induce membrane blebbing. The concentration of YTX that produced after 24 h of incubation a 50% reduction in maximum cell viability (EC5024) was approximately 46 nM, whereas with OA over 75% of the cells were still viable after exposure to 100 nM OA. According to our results, the cytoskeleton of EL-4 cells seems to be a cell component particularly sensitive to YTX and OA with disruption of F-actin cytoskeleton in these cells treated with concentrations of YTX or OA as low as 5 nM at 48 h incubation. Toxicity by YTX or OA involved typical hallmarks of apoptosis and an increase of reactive oxygen species (ROS) production. The cytotoxic effects of YTX and OA reported here, and the previously demonstrated potential of these toxins to regulate the activity of EL-4 cells through the regulation of TCR expression, rise reasonable concern about possible risks for human health associated to the chronic exposure to low amounts of YTX or OA itself or enhanced by the presence of other shellfish toxins specially by a population potentially at risk such as immunocompromised patients.

Human monoclonal antibodies to HIV-1 gp140 from mice bearing YAC-based human immunoglobulin transloci.

Mice carrying human immunoglobulin transloci were immunised with HIV-1 gp140 antigen to gain insight into the range and nature of human monoclonal antibodies (mAbs) that can be elicited from such humanised mice. Using five-feature mice that harbour YAC-based germline-configuration human IgM, Igκ and Igλ transloci in a mouse background disrupted for endogenous mouse IgH and Igκ expression, gp140-specific human IgM mAbs were readily elicited following serial immunisation. These mAbs were converted to human IgG1 format and were found to bind diverse epitopes within gp140, exhibiting high functional affinity for the antigen-typically in the nanomolar or sub-nanomolar range. The number of specific, stable hybridomas per mouse was, however, low (typically around five) with the hybridomas within individual mice often being clonally related. Nevertheless, different mice used B cell clones expressing varied V(D)J combinations, with affinity maturation through somatic hypermutation making a critical contribution. Thus, a wide range of distinct high-affinity mAbs can be obtained by immunising multiple animals. The results confirm the utility of the translocus-mouse approach and give insight into strategies for possible future improvement.

Effects of hydroxyurea on monoclonal antibody production induced by anti-mIgG and LPS stimulation on murine B cell hybridomas.

Chemical treatment with hydroxyurea (HU) has been selected as a simple and low cost strategy to generate a cell population enriched for the G1 phase. After the chemical treatment with HU, cells were stimulated with anti-mIgG to test if the positive effects of anti-mIgG on CD40 expression and specific IgG2a production rate were improved upon a cell population with a higher percentage of cells in G1 phase at the beginning of the cell culture. In addition, other treatments assayed in this work were the cell stimulation with Lipopolysaccharide (LPS) both before and after the HU treatment. It has been observed that the use of HU under conditions able to maintain the cells in viable state (0.1 mM for 20 h), has a negative effect on CD40 expression and specific IgG2a production rate induced by anti-mIgG. The positive effect of LPS on cell stimulation induced by anti-mIgG is reduced on cells treated with HU.

Enhanced monoclonal antibody production in hybridoma cells by LPS and Anti-mIgG.

This paper reports on a methodology for increasing proliferation and monoclonal antibody (mAb) production in hybridoma cultures. The 55-6 murine B cell hybridoma line (CD40 and CD19-deficient expression) was treated with increasing concentrations of lipopolysaccharide (LPS). Expression of CD69, CD40, and CD19 surface antigens on 55-6 cells did not show significant changes from untreated cells. The specific growth rate decreased at higher concentrations of LPS, but the monoclonal antibody production rate was highest at the highest LPS concentration assayed. These data are in agreement with the lowest growth rate found at this concentration of LPS. Furthermore, cells were cultured with anti-mouse surface immunoglobulin G antibody (anti-mIgG) plus LPS to find out whether LPS-derived signals and anti-mIgG stimuli are synergistic. CD69, CD40, and CD19 expression was greater than for either untreated cells (control culture) or cells stimulated with LPS alone. Moreover, LPS stimulation in combination with anti-mIgG enhanced both the growth rate and IgG2a production over the control culture and cells stimulated with LPS alone. Maximum antibody concentration increased almost 500% compared to the control and about 100% with respect to culture stimulated with LPS alone. The maximum specific IgG2a production rate was about 300% higher than in the control culture and about 30% higher than in culture stimulated only with LPS.

Effects of synchronization on CD40 expression and antibody production in hybridoma cells stimulated with anti-mIgG.

In previous experiments with the 55-6 hybridoma cell line, we showed that cell stimulation with anti-mouse surface immunoglobulin G antibody (anti-mIgG) increased both CD40 expression and specific monoclonal antibody (mAb) production rate. Cell preincubation with lipopolysaccharide (LPS) prior to anti-mIgG stimulation enhanced these results. Moreover, the expression of both CD40 and surface immunoglobulin G (sIgG) were higher for cells in the G1 phase of the cell cycle. Therefore, to determine the relationship between cell cycle position, CD40 expression, and mAb productivity, in this work cells were synchronized in the G1 phase by thymidine block. In addition, synchronized cells were subjected to different treatments with anti-mIgG. Although synchronized cells showed a slight increase in both CD40 expression and maximum specific growth rate (mu max) compared with unsynchronized cells, specific productivity did not show significant changes. However, the stimulation of synchronized cells with anti-mIgG increased over 65% the expression of CD40 and over 50% the specific productivity in comparison with that obtained on unsynchronized cells after anti-mIgG stimulation. These data improved additionally over 15 and 60%, respectively, by adding 2 mM thymidine to the culture medium. These results suggest that the effect of the positive association between G1 phase, CD40 expression, and specific productivity is subordinated to the effect of anti-mIgG stimulation, which is enhanced by increasing the percentage of cells on the G1 phase of the cell cycle. Contrary to expectations, LPS preincubation of synchronized cells prior to anti-mIgG stimulation did not increase the specific productivity in comparison with non-preincubated cells, and the expression of CD40 was minor compared to that on non-preincubated cells.

Induction of CD40 expression and enhancement of monoclonal antibody production on murine B cell hybridomas by cross-linking of IgG receptors.

The 55-6 murine B cell hybridoma line not constitutively expressing CD40 was treated with increasing amounts of intact anti-mouse surface immunoglobulin G antibody (anti-mIgG) either not preincubated or preincubated for 48 h with lipopolysaccharide (LPS). In vitro, cross-linking of surface immunoglobulin G (sIgG) with the whole molecule of anti-IgG antibodies induced the expression of CD69, CD40, and CD19 surface antigens on 55-6 cells. The effect of sIgG ligation was dose-dependent, and preincubation with LPS enhanced their responsiveness to anti-mIgG stimulation. The expression of these surface molecules reached the maximum value during the first part of the cell cycle, corresponding to the position of the G1 peak of the DNA distribution. Stimulation of cells with anti-mIgG did not induce changes either in the number of viable cells or in the fraction of cells undergoing proliferation (mitosis). However, preincubation of 55-6 cells with LPS for 48 h before stimulation with anti-mIgG increased both the maximum specific growth rate (micromax) and the percentage of cells in the G2/M phase, in comparison with non-preincubated cells. Moreover, on cells preincubated with LPS prior to anti-mIgG treatment, specific IgG2a production rate was enhanced significantly compared to that obtained in control cultures. The correlation between the antibody production rate and the amount of IgG that is detectable on the cell surface was analyzed by flow cytometry. A good correlation between secreted and surface IgG was observed, and the results of cell cycle analyses demonstrated that the 55-6 hybridoma cell line has a substantially higher sIgG content in G1 phase.

Nefropatía diabética / Diabetic nephropathy

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