Papers by Valentina calamia
Supplemental material, suppl_table_2 for Predictive modeling of therapeutic response to chondroit... more Supplemental material, suppl_table_2 for Predictive modeling of therapeutic response to chondroitin sulfate/glucosamine hydrochloride in knee osteoarthritis by Francisco J. Blanco, María Camacho-Encina, Lucía González-Rodríguez, Ignacio Rego-Pérez, Jesús Mateos, Patricia Fernández-Puente, Lucía Lourido, Beatriz Rocha, Florencia Picchi, María T. Silva-Díaz, Marta Herrero, Helena Martínez, Josep Verges, Cristina Ruiz-Romero and Valentina Calamia in Therapeutic Advances in Chronic Disease
Supplemental material, suppl_table_1 for Predictive modeling of therapeutic response to chondroit... more Supplemental material, suppl_table_1 for Predictive modeling of therapeutic response to chondroitin sulfate/glucosamine hydrochloride in knee osteoarthritis by Francisco J. Blanco, María Camacho-Encina, Lucía González-Rodríguez, Ignacio Rego-Pérez, Jesús Mateos, Patricia Fernández-Puente, Lucía Lourido, Beatriz Rocha, Florencia Picchi, María T. Silva-Díaz, Marta Herrero, Helena Martínez, Josep Verges, Cristina Ruiz-Romero and Valentina Calamia in Therapeutic Advances in Chronic Disease
Supplemental material, supplementary_figures for Predictive modeling of therapeutic response to c... more Supplemental material, supplementary_figures for Predictive modeling of therapeutic response to chondroitin sulfate/glucosamine hydrochloride in knee osteoarthritis by Francisco J. Blanco, María Camacho-Encina, Lucía González-Rodríguez, Ignacio Rego-Pérez, Jesús Mateos, Patricia Fernández-Puente, Lucía Lourido, Beatriz Rocha, Florencia Picchi, María T. Silva-Díaz, Marta Herrero, Helena Martínez, Josep Verges, Cristina Ruiz-Romero and Valentina Calamia in Therapeutic Advances in Chronic Disease
Mitochondria are involved in many cellular processes; mi-tochondrial dysfunctions have been assoc... more Mitochondria are involved in many cellular processes; mi-tochondrial dysfunctions have been associated with apo-ptosis, aging, and a number of pathological conditions, in-cluding osteoarthritis (OA). Mitochondrial proteins are attractive targets for the study of metabolism of the chon-drocyte, the unique cell type present in mature cartilage, and its role in tissue degradation. Using a proteomics ap-proach based on two-dimensional DIGE and MALDI-TOF/ TOF mass spectrometric identification of mitochondria-enriched protein fractions from human articular chondro-cytes, we analyzed mitochondrial protein changes that are characteristic of OA chondrocytes. A total of 73 protein forms were unambiguously identified as significantly al-tered in OA; 23 of them have been previously described as mitochondrial. An extensive statistical and cluster analysis of the data revealed a mitochondrial protein profile charac-teristic for OA. This pattern includes alterations in energy production, maintenan...
Mesenchymal stem cells (MSCs) were isolated from umbilical cord stromal tissues and differentiate... more Mesenchymal stem cells (MSCs) were isolated from umbilical cord stromal tissues and differentiated towards chondrocyte-like cells through spheroids. RT-PCR, immunohistochemistry, flow cytometry, and secretome analysis were done to test this new model of chondrogenesis (Stem Cell & Dev. 2010). In this work we have studied proteome of this differentiation by DIGE analysis from 12 donors and 6 check points 4,7,14,28 and 46 days into chondrogenic medium. Protein identification was realized by MALDI-TOF/TOF mass spectrometry. 1370 spots were seen in the gel DIGE and 97 spots were modulated considering p≤ 0.01 statistically significant and were identified. 35 proteins were down-regulated and 62 were up-regulated in all time points studied compared with MSCs no differentiated. The modulated proteins were also classified in different groups according cellular function, the most of the proteins were involved in cell organization (45%) followed by redox and stress function (19%), metabolism (...
La artrosis es la patologia reumatica mas frecuente. Esta relacionada con la edad y se caracteriz... more La artrosis es la patologia reumatica mas frecuente. Esta relacionada con la edad y se caracteriza principalmente por la degradacion del cartilago. Pese a su gran prevalencia, poco se sabe de los mecanismos moleculares por los que se produce la degradacion del tejido articular. Datos previos de nuestro grupo sugieren que fumar puede ser un factor protector contra la artrosis. Por tanto, en este trabajo nos hemos propuesto evaluar desde un punto de vista proteomico el posible efecto antiinflamatorio o anabolico de la nicotina en la patologia articular. Para ello, condrocitos de donantes normales y artrosicos se cultivaron en medio DMEM con lisina y arginina marcada (condicion “heavy”) o con estos aminoacidos no marcados (condicion “light”) para realizar un estudio proteomico mediante SILAC (stable isotope labelling by amino acids in cell culture). Tras el marcaje, los condrocitos fueron estimulados con nicotina (50nM), IL1β (una citoquina inflamatoria) o la combinacion de ambos compu...
Recientes trabajos realizados por nuestro grupo han demostrado la utilidad de las técnicas proteó... more Recientes trabajos realizados por nuestro grupo han demostrado la utilidad de las técnicas proteómicas para la evaluación de diferentes fármacos empleados en el tratamiento de la artrosis (OA), la patología articular más frecuente hoy en día . En la articulación artrósica, las proteínas secretadas por los condrocitos (las células del cartílago más directamente implicadas en el desarrollo de esta enfermedad) pueden llegar al torrente sanguíneo; de ahí su posible utilidad como marcadores no invasivos para monitorizar y personalizar los tratamientos anti-OA a largo plazo. Por este motivo, el objetivo de este trabajo es comparar dos técnicas proteómicas cuantitativas (iTRAQ e SILAC) para el estudio del secretoma de los condrocitos y poder evaluar la mejor opción para abordar un estudio farmacoproteómico en este campo. Para ello, se utilizaron condrocitos articulares humanos obtenidos a partir de biopsias de donantes artrósicos. Las células fueron cultivadas en un medio estándar (iTRAQ) ...
Osteoarthritis and Cartilage Open
Osteoarthritis and Cartilage
Molecular & Cellular Proteomics
In osteoarthritis (OA), impairment of cartilage regeneration can be related to a defective chondr... more In osteoarthritis (OA), impairment of cartilage regeneration can be related to a defective chondrogenic differentiation of mesenchymal stromal cells (MSCs). Therefore, understanding the proteomic- and metabolomic-associated molecular events during the chondrogenesis of MSCs could provide alternative targets for therapeutic intervention. Here, a SILAC-based proteomic analysis identified 43 proteins related with metabolic pathways whose abundance was significantly altered during the chondrogenesis of OA human bone marrow MSCs (hBMSCs). Then, the level and distribution of metabolites was analyzed in these cells and healthy controls by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI), leading to the recognition of characteristic metabolomic profiles at the early stages of differentiation. Finally, integrative pathway analysis showed that UDP-glucuronic acid synthesis and amino sugar metabolism were downregulated in OA hBMSCs during chondrogenesis compare...
Therapeutic Advances in Chronic Disease
Background: In the present study, we explored potential protein biomarkers useful to predict the ... more Background: In the present study, we explored potential protein biomarkers useful to predict the therapeutic response of knee osteoarthritis (KOA) patients treated with pharmaceutical grade Chondroitin sulfate/Glucosamine hydrochloride (CS+GH; Droglican, Bioiberica), in order to optimize therapeutic outcomes. Methods: A shotgun proteomic analysis by iTRAQ labelling and liquid chromatography–mass spectrometry (LC-MS/MS) was performed using sera from 40 patients enrolled in the Multicentre Osteoarthritis interVEntion trial with Sysadoa (MOVES). The panel of proteins potentially useful to predict KOA patient’s response was clinically validated in the whole MOVES cohort at baseline ( n = 506) using commercially available enzyme-linked immunosorbent assays kits. Logistic regression models and receiver-operating-characteristics (ROC) curves were used to analyze the contribution of these proteins to our prediction models of symptomatic drug response in KOA. Results: In the discovery phase ...
International Immunopharmacology
Selective cyclooxigenase (COX)-2 inhibitors were developed to prevent traditional non-steroidal a... more Selective cyclooxigenase (COX)-2 inhibitors were developed to prevent traditional non-steroidal anti-inflammatory drugs (tNSAIDs) gastro-intestinal adverse effects. VA692, a recently disclosed selective COX-2 inhibitor, structurally related to well-known marketed coxibs, showed anti-inflammatory, and anti-nociceptive properties. The aim of this study was to analyze the anti-inflammatory effect of VA692, in comparison with celecoxib. At this purpose we evaluated the pro-inflammatory cytokines and anti-oxidant enzymes gene expression, apoptosis and ROS production, and PGE release in chondrocytes (both primary cultures and immortalized T/C-28a2 cell line) treated with the two drugs. Furthermore, a proteomic analysis has been performed in T/C-28a2 cell line to evaluate modifications in their proteomic profile following drug treatment in presence of IL-1β. Our results demonstrated the anti-inflammatory effect of the novel synthesized VA692, and confirmed those of celecoxib, in counteracting the stimulus of IL-1β in both osteoarthritic (OA) chondrocytes and T/C-28a2 cell line. Furthermore, the data underlined the possible anti-apoptotic and anti-oxidant role of VA692, implying its regulation in superoxide anion production as indicated by the modulation of anti-oxidant enzymes. The proteomic analysis provides new information about the effect of VA692 on human T/C-28a2 intracellular proteome, demonstrating the usefulness of this approach in the identification and quantifications of several proteins. Modulation of some proteins such as Hsp90 and SOD by VA692 could explain its role in the therapeutic approach of OA. Based on our results, we can affirm that VA692 has more beneficial effect compared with celecoxib particularly regarding the modulation of oxidant/anti-oxidant system and proteome profile of human articular chondrocytes.
Osteoarthritis and Cartilage
Abstracts Accepted for Publication
Background: The Leeds Assessment of Neuropathic Symptoms (LANSS) and the painDETECT questionnaire... more Background: The Leeds Assessment of Neuropathic Symptoms (LANSS) and the painDETECT questionnaire (PDQ) are two validated screening tools for neuropathic pain (NP). Recent evidence reported a low level of agreement between these tests in knee Osteoarthritis. Several studies have recently applied the PDQ in Rheumatoid Arthritis (RA), suggesting a NP component in these patients, although the application and performance comparison with LANSS is yet to be studied. Objectives: Evaluate PDQ and LANSS performance for NP classification and investigate its optimal cutoff points in a RA cohort. Methods: Observational, cross-sectional study was designed including RA patients followed at our Rheumatology department. Patients with diagnosed neuropathy or non-RA risk factors for NP were excluded. Selected patients were evaluated in a medical visit where LANSS and PDQ were applied. Agreement between the two questionnaires was evaluated using kappa coefficient analysis. Receiver operating characteristic (ROC) analysis was performed using each tool as gold-standard and cutoff points to optimize agreement were investigated. Nonconcordant patients were compared with concordant patients using parametric and non-parametric tests. Significance level was set as <0.05. Results: 112 RA patients were included, 86 (77%) were females, with a mean (SD) age of 55.1 (10.8) years and median disease duration of 13 years (range: 2-41). 102 (91%) were treated with DMARDs and 42% with a biologic DMARD. 45 (40%) patients had NP applying the LANSS (≥12) and 28% had NP in the PDQ (19 possible and 12 likely; no demographic or clinical significant differences were found between these two groups). 82 (73%) patients had concordant NP classification (59 negative, 23 positive) by the two tests. Concordant group had significantly superior median disease duration and inferior LANSS scores compared to non-concordant group (14 vs 12 years and 8 vs 13, respectively, p<0.05) with no other significant differences found. A moderate agreement (κ=0.41) and linear correlation (r=0.58, p<0.001) were observed between the two tests. In the ROC curve analysis, PDQ (≥13) showed an area under the curve (AUC) of 0.80, 95% CI [0.72-0.88] with a sensitivity and specificity of 51% and 88%, respectively, using LANSS as gold standard. LANSS (≥12) had an AUC of 0.80, 95% CI [0.71-0.90] and a sensitivity and specificity of 74% and 73%, respectively, using PDQ as gold standard. After ROC curve analysis, optimal cutoff for PDQ was 10, showing greater sensitivity (69%) but lower specificity (79%) with a slight increase in the agreement between the tests (κ=0.48). For the LANSS, the optimal cutoffs were the previous value or 13 (sensitivity 68% and specificity 78%) with a modest gain in the agreement (κ=0.42). Correction for both cutoff points resulted in a more substantial increase in agreement level (κ=0.51). Conclusions: In this study, LANSS and PDQ had a moderate level of agreement, possibly because they capture different dimensions of NP. New possible cutoffs were studied to increase agreement between the tests. Further studies with other conditions and a validated gold-standard for NP are needed to confirm this data.
Journal of proteome research, Jan 4, 2017
Carcinoma, the most common type of cancer, arises from epithelial cells. The transition from aden... more Carcinoma, the most common type of cancer, arises from epithelial cells. The transition from adenoma to carcinoma is associated with the loss of E-cadherin and, in consequence, the disruption of cell-cell contacts. E-cadherin is a tumour suppressor and it is down-regulated during epithelial-to-mesenchymal transition (EMT), indeed its loss is a predictor of poor prognosis. Hakai is an E3 ubiquitin-ligase protein that mediates E-cadherin ubiquitination, endocytosis and finally degradation, leading the alterations of cell-cell contacts. Although E-cadherin is the most established substrate for Hakai activity, other regulated molecular targets for Hakai may be involved in cancer cell plasticity during tumour progression. In this work we employed an iTRAQ approach to explore novel molecular pathways involved in Hakai-driven EMT during tumour progression. Our results show that Hakai may have an important influence on cytoskeleton-related proteins, extracellular exosome-associated proteins...
Journal of Proteomics, 2017
The methods currently available for the diagnosis and monitoring of osteoarthritis (OA) are very ... more The methods currently available for the diagnosis and monitoring of osteoarthritis (OA) are very limited and lack sensitivity. Being the most prevalent rheumatic disease, one of the most disabling pathologies worldwide and currently untreatable, there is a considerable interest pointed in the verification of specific biological markers for improving its diagnosis and disease progression studies. Considering the remarkable development of targeted proteomics methodologies in the frame of the Human Proteome Project, the aim of this work was to develop and apply a MRM-based method for the multiplexed analysis of a panel of 6 biomarker candidates for OA encoded by the Chromosome 16, and another 8 proteins identified in previous shotgun studies as related with this pathology, in specimens derived from the human joint and serum. The method, targeting 35 different peptides, was applied to samples from human articular chondrocytes, healthy and osteoarthritic cartilage, synovial fluid and serum. Subsequently, a verification analysis of the biomarker value of these proteins was performed by single point measurements on a set of 116 serum samples, leading to the identification of increased amounts of Haptoglobin and von Willebrand Factor in OA patients. Altogether, the present work provides a tool for the multiplexed monitoring of 14 biomarker candidates for OA, and verifies for the first time the increased amount of two of these circulating markers in patients diagnosed with this disease. We have developed an MRM method for the identification and relative quantification of a panel of 14 protein biomarker candidates for osteoarthritis. This method has been applied to analyze human articular chondrocytes, articular cartilage, synovial fluid, and finally a collection of 116 serum samples from healthy controls and patients suffering different degrees of osteoarthritis, in order to verify the biomarker usefulness of the candidates. HPT and VWF were validated as increased in OA patients.
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Papers by Valentina calamia