Decoding the inflammatory signature of the major depressive episode: insights from peripheral immunophenotyping in active and remitted condition, a case-control study.

Depression is a prevalent and incapacitating condition with a significant impact on global morbidity and mortality. Although the immune system's role in its pathogenesis is increasingly recognized, there is a lack of comprehensive understanding regarding the involvement of innate and adaptive immune cells. To address this gap, we conducted a multicenter case-control study involving 121 participants matched for sex and age. These participants had either an active (or current) major depressive episode (MDE) (39 cases) or a remitted MDE (40 cases), including individuals with major depressive disorder or bipolar disorder. We compared these 79 patients to 42 healthy controls (HC), analyzing their immunological profiles. In blood samples, we determined the complete cell count and the monocyte subtypes and lymphocyte T-cell populations using flow cytometry. Additionally, we measured a panel of cytokines, chemokines, and neurotrophic factors in the plasma. Compared with HC, people endorsing a current MDE showed monocytosis (p = 0.001), increased high-sensitivity C-reactive protein (p = 0.002), and erythrocyte sedimentation rate (p = 0.003), and an altered proportion of specific monocyte subsets. CD4 lymphocytes presented increased median percentages of activation markers CD69+ (p = 0.007) and exhaustion markers PD1+ (p = 0.013) and LAG3+ (p = 0.014), as well as a higher frequency of CD4+CD25+FOXP3+ regulatory T cells (p = 0.003). Additionally, patients showed increased plasma levels of sTREM2 (p = 0.0089). These changes are more likely state markers, indicating the presence of an ongoing inflammatory response during an active MDE. The Random Forest model achieved remarkable classification accuracies of 83.8% for MDE vs. HC and 70% for differentiating active and remitted MDE. Interestingly, the cluster analysis identified three distinct immunological profiles among MDE patients. Cluster 1 has the highest number of leukocytes, mainly given by the increment in lymphocyte count and the lowest proinflammatory cytokine levels. Cluster 3 displayed the most robust inflammatory pattern, with high levels of TNFα, CX3CL1, IL-12p70, IL-17A, IL-23, and IL-33, associated with the highest level of IL-10, as well as ß-NGF and the lowest level for BDNF. This profile is also associated with the highest absolute number and percentage of circulating monocytes and the lowest absolute number and percentage of circulating lymphocytes, denoting an active inflammatory process. Cluster 2 has some cardinal signs of more acute inflammation, such as elevated levels of CCL2 and increased levels of proinflammatory cytokines such as IL-1ß, IFNγ, and CXCL8. Similarly, the absolute number of monocytes is closer to a HC value, as well as the percentage of lymphocytes, suggesting a possible initiation of the inflammatory process. The study provides new insights into the immune system's role in MDE, paving the ground for replication prospective studies targeting the development of diagnostic and prognostic tools and new therapeutic targets.

Immunological alterations in patients with current and lifetime suicide ideation and attempts: Examining the relationship with depressive symptoms.

Background: Suicidal ideation and attempt (SI/SA) have been associated with dysregulation of the immune response and inflammation. However, few studies have explored how innate and acquired cellular immunity impact on the peripheral immune response. Our study addresses this gap by examining the composition of peripheral immune cells and humoral markers among individuals with current SI/SA, individuals with a history of SI/SA, and healthy controls (HC). Additionally, we aim to explore whether depressive symptoms settle the relationship between inflammation and SI/SA. Methods: This is a multicenter case-control study that included 105 participants. Clinical and demographic characterists together with hemogram parameters, soluble pro and anti-inflamatory factors, and specific innate and adaptive immune cell populations were compared among patients with current SI/SA (n = 21), a history of lifetime SI/SA (n = 42), and HC (n = 42). Results: Patients with both current and lifetime SI/SA had a significant increase in the absolute count of monocytes and in the monocyte/lymphocyte ratio (MLR). Additionally, patients with current and lifetime SI/SA showed a significant increase in high-sensitivity C- reactive protein (hs-CRP), and patients with lifetime SI/SA also showed higher levels of Erythrocyte Sedimentation Rate (ESR). The cellular inflammatory status of patients with SI/SA was characterized by altered proportions of monocytes with higher levels of nonclassical and intermediate monocytes. No differences were observed in the number of lymphocytes and the proportion of CD4 and CD8 between patients and HC, but we found differences in markers of exhaustion of CD4 lymphocytes, with increased levels of Programmed cell death protein 1 (PD1) in Current SI/SA and Lymphocyte activation gene 3 (LAG3) in Current SI/SA and Lifetime SI/SA compared to HC. The plasmainflammatory status was marked by higher levels of soluble Triggering receptor expressed on myeloid cells 2 (sTREM2) in patients with lifetime SI/SA compared to HC. Finally, the multinomial analysis indicates that inflammation and depressive symptoms are independently associated with SI/SA. Conclusion: This study highlights the association of immunological alterations with SI/SA. Furthremore, SI/SA is independently influenced by depressive symptoms and inflammation. This may have important therapeutic implications, as in these patients, it may be necessary to treat the inflammatory process beyond treating the depressive symptoms.

Peripheral blood cellular immunophenotype in suicidal ideation, suicide attempt, and suicide: a systematic review and meta-analysis.

Previous meta-analyses have documented the association of immune-inflammatory pathways with the pathophysiology of Major Depressive Episode (MDE), as reflected by alterations in peripheral blood immune cell counts. However, it remains unclear whether these immunological changes are distinct in individuals experiencing suicidal ideation (SI) or suicidal behavior (SB), beyond the context of an MDE. This systematic review and meta-analysis aimed to examine peripheral immune cell profiles across samples with SI/SB and compare them to healthy controls or patients with MDE. A systematic literature search was conducted in MEDLINE, Embase, and PsycINFO for articles published from inception until June 12, 2023. Two independent reviewers screened the articles for inclusion, extracted data, and assessed the risk of bias using the Newcastle-Ottawa scale. Meta-analyses were performed using a random-effects model to calculate standardized mean differences (SMDs) and 95% confidence intervals (CIs) for immune cell counts or ratios between groups with and without SI/SB. Heterogeneity across studies was assessed using the restricted maximum-likelihood estimator for tau statistic and I2-statistic and tested by the Q test. Publication bias was evaluated using the Egger´s test and funnel plots. Meta-regression analyses were conducted to explore the potential moderating effects of age, gender, current or lifetime SI/SB, and the type of self-harming behavior (SI or SB). The study was registered with PROSPERO (CRD42023433089). The systematic review included 30 studies, with data from 19 studies included in the meta-analyses comprising 139 unique comparisons. Eleven different cell populations or ratios were included, comprising 1973 individuals with SI/SB and 5537 comparison subjects. White blood cell (WBC) and neutrophil counts were higher in individuals with SI/SB than in controls (WBC: SMD = 0.458; 95% CI = 0.367-0.548; p value ≤ 0.001; I2 = 0.002% and; Neutrophils: SMD = 0.581; 95% CI = 0.408-0.753; p < 0.001), indicating an inflammatory process. The neutrophil-to-lymphocyte ratio (NLR) emerged as a potential marker, demonstrating a notable elevation in individuals with SI/SB (SMD = 0.695; 95% CI = 0.054-1.335; p value = 0.033; I2 = 94.281%; Q test p value ≤ 0.001). The elevated NLR appears to be primarily driven by the increase in neutrophil counts, as no significant differences were found in lymphocyte counts between groups. Comparisons among participants with and without SI/SB and depression revealed similar trends with increased NLR, monocyte-to-lymphocyte ratio (MLR), and platelet-to-lymphocyte ratio (PLR) observed in depressed individuals with SI/SB compared to those without SI/SB. Broad alteration in the peripheral immune cell populations and their ratios were observed in individuals with SI/SB, indicating an immune activation or dysfunction. Notably, these immunological changes were also evident when comparing MDE individuals with and without SI/SB, suggesting that such immune dysfunction associated with suicidality cannot be solely attributed to or explained by depressive symptoms. The NLR, MLR, and PLR ratios, in combination with novel immune cellular and protein biomarkers, open new avenues in understanding the immunological underpinnings of SI/SB. These findings highlight the potential utility of immune markers as part of a multi-modal approach for risk stratification and therapeutic monitoring in SI/SB.

A randomized 3-month, parallel-group, controlled trial of CALMA m-health app as an adjunct to therapy to reduce suicidal and non-suicidal self-injurious behaviors in adolescents: study protocol.

Background: Suicidal and non-suicidal self-injurious behaviors are among the leading causes of death and injury in adolescents and youth worldwide. Mobile app development could help people at risk and provide resources to deliver evidence-based interventions. There is no specific application for adolescents and young people available in Spanish. Our group developed CALMA, the first interactive mobile application with the user in Spanish, which provides tools based on Dialectical Behavioral Therapy to manage a crisis of suicidal or non-suicidal self-directed violence with the aim of preventing suicide in adolescents and youth. Methods: To test the effectiveness, safety and level of engagement of the CALMA app in people aged 10 to 19 who are treated in mental health services of two public hospitals, we will conduct a parallel-group, two-arm randomized controlled trial. Participants will be assessed face-to-face and via video call at four timepoints: day-0 (baseline), day-30, day-60, and day-90. A total of 29 participants per group will be included. Change in the frequency of suicidal and non-suicidal self-injurious behaviors will be compared between groups, as well as the level of emotional dysregulation, level of app engagement and time of psychiatric admission during the follow-up period. Discussion: This study is particularly relevant to young people given their widespread use of mobile technology, while there are currently no available smartphone app-based self-guided psychological strategies in Spanish that attempt to reduce suicidal behavior in adolescents who are assisted in the public health sector from low and middle-income countries in Latin America. Clinical trial registration: https://clinicaltrials.gov/, NCT05453370.

CALMA, a Mobile Health Application, as an Accessory to Therapy for Reduction of Suicidal and Non-Suicidal Self-Injured Behaviors: A Pilot Cluster Randomized Controlled Trial.

The present study was aim to evaluate the acceptability and preliminary effectiveness of a mobile-health application, CALMA, based on dialectical behavioral therapy skills. CALMA was proposed as an adjunct to therapy for the reduction of non-suicidal and suicidal self-injury behaviors. A parallel design was used to compare the intervention group (DBT + CALMA) and the comparison group (DBT). Patients were randomized based on their groups following a randomized cluster design. After treatment, patients completed a follow-up assessment four weeks later. Each cluster was represented by a different DBT Skills Training Group offered weekly at FORO Foundation for Mental Health. Six DBT Skills Training groups were randomized to the intervention or comparison group. Twenty-one individuals met inclusion criteria and eighteen were included in the analysis. Acceptability was measured with the User Experience Questionnaire short version (UEQ-s) and preliminary efficacy with the Self-injurious thoughts and behaviors interview (SITBI). CALMA shows good acceptability to be used as an adjunct to therapy specific to the reduction of suicidal and non-suicidal self-directed violence. Evidence for preliminary efficacy included a high probability of decreased in more behaviors evaluated with the SITBI pre- and post-intervention for the group that received CALMA as compared to the comparison group. Our study provides initial evidence for the effectiveness and acceptability of CALMA. Future studies scaling up this intervention in a larger number of participants are necessary. CALMA may be especially useful in low and middle-income Latin American countries to improve access to evidence-based interventions openly available in Spanish and free to download.HighlightsCALMA is a DBT-based app aimed to reduce non-suicidal and suicidal self-directed violence.The app showed good acceptability by usersA high probability of decrease in suicidal ideation, suicidal plan, suicidal gesture, thoughts about NSSI and NSSI pre- and post-intervention was observed.

Comparison of traditional model-based statistical methods with machine learning for the prediction of suicide behaviour.

BACKGROUND: Despite considerable research efforts during the last five decades, the prediction of suicidal behaviour (SB) using traditional model-based statistical has been weak. This marks the need to explore new statistical methods. OBJECTIVE: To compare the performance of Cox regression models versus Random Survival Forest (RSF) to predict SB. METHODS: Using a data set of more than 300 high-risk suicidal patients from a multicenter prospective cohort study, we compare Cox regression models with RSF to address predictors of time to suicide reattempt. Cross-validation was used to assess model prediction performance, including the area under the receiver operator curve (AUC), precision, Integrated Brier Score (IBS), sensitivity, and specificity. RESULTS: A variant of the RSF denominated the RSFElimin, in which irrelevant predictor variables were eliminated from the model, presented the best accuracy, sensitivity, AUC and IBS. At the same time, the sensitivity of this method was slightly lower than that obtained with the Cox regression model with all predictor variables (CoxComp). CONCLUSION: The RSF, a machine learning model, seems more sensitive and precise than the traditional Cox regression model in predicting suicidal behaviour.

The history of multiple adverse childhood experiences in patients with schizophrenia is associated with more severe symptomatology and suicidal behavior with gender-specific characteristics.

The current study aims to compare the prevalence of adverse childhood experiences (ACEs) between patients with schizophrenia and non-psychiatric control group, and to analyze the association of having suffered multiple ACEs with clinical symptoms of schizophrenia and suicidal behavior. A multicenter, cross-sectional study was conducted across three facilities in Buenos Aires, Argentina. One-hundred patients with schizophrenia and 50 healthy subjects were assessed with the Adverse Childhood Experiences questionnaire (ACE-Q), the Positive and Negative Syndrome Scale (PANSS), and the Columbia-Suicide Severity Rating Scale (C-SSRS). We observed that the prevalence of at least one ACE in schizophrenic patients was almost double in comparison with the non-psychiatric control group. Multiple ACEs were associated with persistent auditory hallucinations and lower negative symptoms in both sexes. Higher frequency of death ideation and a higher number of suicide attempts were reported among women. The strength of this study is the possibility of comparing the presence of ACEs between schizophrenic patients and non-psychiatric control using the same questionnaire in an under-reported sample of low socio-economic patients assisted in public hospitals. A limitation is that the history of ACEs relied on the retrospective assessment of childhood experiences, and adults could over-report ACEs because of recall bias.

[Is it necessary to have advanced statistical knowledge to access medical literacy?] / ¿Es necesario tener conocimientos avanzados de estadística para acceder a las publicaciones médicas?

The introduction of statistical analysis in biomedical journals should be attributed to Dunn and Greenwood in the 1930s, who reflect the concepts of statistical analysis and interpretation. The aim of this study was to analyze the use of different statistical tests and the level of accessibility analysis-dependent and article-dependent of the original articles published in the journal Medicina (B Aires) in the period 2008-2017. A bibliometric, descriptive, observational, cross-sectional study was carried out, in which the original articles were evaluated according to the scale proposed by Mora Ripoll et al. From the 301 articles in Spanish reviewed, 96% presented statistical analyses. Among the 215 articles that used some inferential statistics technique, 49.7% used the chi-square test, z tests for proportions, the Fisher exact test or the McNemar test; 29.2% used student t-tests and z-tests, including the use of these techniques or the use of confidence intervals for medium-conformance contrasts, and/or median-homogeneity (two samples), in paired or independent samples. The results indicate that a reader who knows about the tests included in Level II will have statistical access to 75% of the original articles published. It was found that it is not necessary to have advanced knowledge of statistics to access most publications, but it is important that these contents are developed with emphasis on statistical reasoning over the application and use of software or the choice of the appropriate test.

La introducción de análisis estadísticos en revistas biomédicas debe atribuirse a Dunn y a Gree wood en la década del 30, quienes reflejan los conceptos del análisis e interpretación estadística. El objetivo del trabajo fue analizar la utilización de distintas pruebas estadísticas y el grado de accesibilidad análisis-dependiente y artículo-dependiente de los artículos originales publicados en la revista Medicina (B Aires) en el período 2008-2017. Se realizó un estudio bibliométrico, descriptivo, observacional, de corte transversal, en el cual se incluyeron los artículos originales que fueron evaluados de acuerdo a la escala propuesta por Mora Ripoll y col. Se revisaron 301 artículos en idioma español, el 96% presentó análisis estadísticos. Entre los 215 artículos que emplearon alguna técnica de estadística inferencial, el 49.7% utilizó la prueba chi-cuadrado, pruebas z para proporciones, el test exacto de Fisher o prueba de McNemar. El 29.2% empleó pruebas t de student y pruebas z, incluyendo el uso de estas técnicas o de intervalos de confianza para contrastes de conformidad de medianas, y/o homogeneidad de medianas (dos muestras), en muestras apareadas o independientes. Los resultados indican que un lector conocedor de las pruebas incluidas en el Nivel II tendrá acceso estadístico al 75% de los artículos originales publicados. Se comprobó que no es necesario tener conocimientos avanzados de estadística para acceder a la mayoría de las publicaciones, pero sí es importante que estos contenidos sean desarrollados haciendo hincapié en el razonamiento estadístico por sobre la aplicación y utilización de software o la elección del test adecuado.

¿ Es necesario tener conocimientos avanzados de estadística para acceder a las publicaciones módicas? / Is it necessary to have advanced statistical knowledge to access medical literacy?

La introducció;n de análisis estadísticos en revistas biomó;©dicas debe atribuirse a Dunn y a Greewood en la dó;©cada del 30, quienes reflejan los conceptos del análisis e interpretació;n estadística. El objetivo del trabajo fue analizar la utilizació;n de distintas pruebas estadísticas y el grado de accesibilidad análisis-dependiente y artículo-dependiente de los artículos originales publicados en la revista Medicina (B Aires) en el período 2008-2017. Se realizó; un estudio bibliomó;©trico, descriptivo, observacional, de corte transversal, en el cual se incluyeron los artículos originales que fueron evaluados de acuerdo a la escala propuesta por Mora Ripoll y col. Se revisaron 301 artículos en idioma espaó;±ol, el 96% presentó; análisis estadísticos. Entre los 215 artículos que emplearon alguna tó;©cnica de estadística inferencial, el 49.7% utilizó; la prueba chi-cuadrado, pruebas z para proporciones, el test exacto de Fisher o prueba de McNemar. El 29.2% empleó; pruebas t de student y pruebas z, incluyendo el uso de estas tó;©cnicas o de intervalos de confianza para contrastes de conformidad de medianas, y/o homogeneidad de medianas (dos muestras), en muestras apareadas o independientes. Los resultados indican que un lector conocedor de las pruebas incluidas en el Nivel II tendrá acceso estadístico al 75% de los artículos originales publicados. Se comprobó; que no es necesario tener conocimientos avanzados de estadística para acceder a la mayoría de las publicaciones, pero sí es importante que estos contenidos sean desarrollados haciendo hincapí en el razonamiento estadístico por sobre la aplicació;n y utilizació;n de software o la elecció;n del test adecuado.

The introduction of statistical analysis in biomedical journals should be attributed to Dunn and Greenwood in the 1930s, who reflect the concepts of statistical analysis and interpretation. The aim of this study was to analyze the use of different statistical tests and the level of accessibility analysis-dependent and article-dependent of the original articles published in the journal Medicina (B Aires) in the period 2008-2017. A bibliometric, descriptive, observational, cross-sectional study was carried out, in which the original articles were evaluated according to the scale proposed by Mora Ripoll et al. From the 301 articles in Spanish reviewed, 96% presented statistical analyses. Among the 215 articles that used some inferential statistics technique, 49.7% used the chi-square test, z tests for proportions, the Fisher exact test or the McNemar test; 29.2% used student t-tests and z-tests, including the use of these techniques or the use of confidence intervals for medium-conformance contrasts, and /or median-homogeneity (two samples), in paired or independent samples. The results indicate that a reader who knows about the tests included in Level II will have statistical access to 75% of the original articles published. It was found that it is not necessary to have advanced knowledge of statistics to access most publications, but it is important that these contents are developed with emphasis on statistical reasoning over the application and use of software or the choice of the appropriate test.

Interaction between prospective risk factors in the prediction of suicide risk.

BACKGROUND: To meet the goal of preventing suicide the most important thing is to know the risk factors of suicidal behavior and understand their interaction. AIMS: The current study aims to evaluate prospective predictors and the interaction between factors for suicide and suicide re-attempts in high-risk, suicidal patients during a 24 month prospective follow-up period. METHODS: A multicenter prospective cohort study was designed to compare data obtained from 324 patients admitted to the emergency department for current suicidal ideation or a recent suicide attempt. Participants were clinically evaluated at baseline and follow-up every 6 months to assess any unfavorable events (suicide or a suicide attempt). To estimate the rate of unfavorable events, the Kaplan-Meier method was used and Cox Proportional Hazards Regression Model was employed to examine predictors of suicide and suicide reattempt. RESULTS: The incidence of a new suicide attempt was 26,000 events/100,000 persons-years. The incidence of death by suicide was 1110 events/100,000 person-year. The most reliable predictors of unfavorable events were being women, previous suicide attempts, younger age, and childhood sexual abuse. Findings revealed an interaction between childhood sexual abuse and low psychosocial functioning that increased the risk of an unfavorable event. CONCLUSION: The risk of suicide re-attempts and suicide in the current 2-year follow-up was high. There was an interaction between low psychosocial functioning and childhood sexual abuse. This evidence should be taken into account for the evaluation and planning of preventive strategies.

Prevalence of potentially inappropriate medications in older adults in Argentina using Beers criteria and the IFAsPIAM List.

Background Medications in which the risk of adverse events exceeds the expectations of clinical benefits are called potentially inappropriate medications (PIMs). To identify the use of PIMs in elderly patients, the most commonly used tool are the Beers criteria, developed for the population of the United States. Recently, a consensus panel of Argentine experts developed the first Latin American tool, called the IFAsPIAM List. Objective The present study aimed to identify PIM prescriptions in elderly outpatients, to estimate the prevalence of PIMs, and to evaluate their possible relation with polypharmacy and gender and age of the patients. Also, we aimed to compare the results obtained by using the Beers criteria and the IFAsPIAM List. Setting Ten community pharmacies of Rosario, Santa Fe, Argentina. Methods A cross-sectional observational study was conducted between February and September 2015. Data were acquired from 56,952 prescriptions prescribed to 2231 patients aged 65 years old or older. To detect the use of PIMs, we used two tools: the Beers criteria and the IFAsPIAM List. Main outcome measure The prevalence of PIM use according to the Beers criteria and the IFAsPIAM List. Results The monthly average of medications dispensed per patient was 4.35 ± 2.18 and 42.27% of the patients presented major polypharmacy. The prevalence of PIMs was 72.75% according to the Beers criteria and 71.13% according to the IFAsPIAM List (Kappa coefficient k = 0.72), and was significantly higher in patients with major polypharmacy, older than 75 years old, and females. The most frequent PIMs prescribed were anxiolytics, analgesics and antipsychotics. Conclusions The IFAsPIAM List is an effective tool to evaluate the prescription of PIMs in the elderly. The results showed a high prevalence of PIMs with a multicausal origin and directly associated with polypharmacy. As clarified by the authors of the IFAsPIAM List, the criteria specified in the list do not substitute the clinical evaluation of each patient.

Potentially Inappropriate Medications in Elderly Ambulatory Patients: A Comparative Study between a Primary Health Care Center and a Community Pharmacy.

OBJECTIVE: This study aims to compare, both qualitatively and quantitatively, the medication dispensed to elderly patients in a primary health care center (PHC) and a community pharmacy (CP) in Argentina and to identify the prescription of potentially inappropriate medications (PIMs). METHODS: A cross-sectional observational study. Data were acquired from 886 prescriptions in the PHC and 2368 in the CP between February and April 2015. Dispensed medications were coded according to the Anatomical, Therapeutic, and Chemical (ATC) classification system. The frequency of prescriptions for each of them was determined. The number and monthly average of drugs dispensed were calculated for each patient. The use of PIMs was identified using Beers Criteria. RESULTS: In both institutions, the means of medications dispensed per individual and month were similar: 3.69 ± 1.93 in the PHC and 3.46 ± 2.18 in the CP. Most of the medications corresponded to cardiovascular system agents. In the CP, 111 prescriptions (4.69%) dispensed to 51 patients (19.39%) were identified as PIMs. In the PHC, 72 prescriptions (8.13%) dispensed to 27 patients (28.42%) were identified as PIMs. In patients with major polymedication the possibility of consuming these drugs was 2.55 times higher in the CP and 2.60 times higher in the PHC. The percentage of PIM prescriptions was significantly higher in the PHC, although the percentage of patients receiving them did not differ significantly. CONCLUSIONS: The prevalence of PIMs found in this population is relevant enough to implement measures that address the problem in an integral way, to improve the quality of prescriptions and the health outcomes of patients.

Assessment of polypharmacy in elderly patients by using data from dispensed medications in community pharmacies: analysis of results by using different methods of estimation.

Background Estimating the prevalence of polypharmacy is essential for the evaluation of public health. Many different methodologies are used to determine the number of drugs used by a patient. Objective To analyse and compare three different methods (simultaneous, cumulative and continuous medication) to determine the number of drugs used by a patient, to estimate the prevalence of polypharmacy and to evaluate the possible association between polypharmacy and the gender and age of patients. Method Cross-sectional observational study carried out between April and September 2015. Data were acquired from prescriptions corresponding to 3972 patients aged 65 years old or older in ten community pharmacies in Argentina. Results The prevalence of polypharmacy varied significantly according to the method used. Major polypharmacy (use of five or more drugs) was detected in 20.5-47.1% of the patients. The association between gender, age and polypharmacy was statistically significant only when using the continuous medication method. The prevalence of minor polypharmacy (use of two to four drugs) was similar with the three methods. Conclusion These results contribute to deciding which is the best method to determine polypharmacy according to the objective of future studies and considering the advantages and disadvantages of each of them.