Artículos (Citología e Histología Normal y Patológica)

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A mobile app using therapeutic exercise and education for self-management in patients with hand rheumatoid arthritis: a randomized controlled trial protocol
(BMC, 2020) Rodríguez Sánchez Laulhé, P.; Luque Romero, Luis Gabriel; Blanquero, Jesús; Suero Pineda, Alejandro; Biscarri Carbonero, Angela; Barrero García, Francisco José; Heredia Rizo, Alberto Marcos; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Universidad de Sevilla. Departamento de Fisioterapia; Universidad de Sevilla. CTS1110: Uncertainty, Mindfulness, Self, and Spirituality
Background: Therapeutic exercise is a safe and cost-effective approach to alleviate hand rheumatoid arthritis (RA)- related symptoms. This study aims to investigate the differences in self-management between a smartphone app (CareHand), using hand exercises and educational advices, compared with a standard approach, on hand overall function, pain intensity, stiffness, and grip and pinch strength in patients with hand RA. Methods: The project is a prospective, longitudinal, superiority, randomized controlled trial. Fifty-eight participants with hand RA will be randomly assigned into an experimental group (CareHand app) or a control group (conventional treatment). Control intervention involves a paper sheet with exercises and recommendations, and the experimental group includes the use of a smartphone app, which provides individualized exercise programs, self-management, and educational strategies to promote adherence to treatment. Both intervention protocols will last for 3 months. The principal investigator will conduct an educational session at baseline for all participants. Primary outcome comprises the overall hand function, assessed with the Michigan Hand Outcome Questionnaire (MHQ). Secondary outcomes include self-reported functional ability with the Quick DASH questionnaire, self-reported pain intensity and morning stiffness using a Visual Analogue Scale (VAS), and hand grip and pinch strength (dynamometer). Outcome measures will be collected at baseline, and at 1 month and 3-month follow-up. Discussion: This study will evaluate the effectiveness of a tele-rehabilitation tool, which uses exercise and self-management strategies, compared to a conventional approach, in patients with hand RA. The smartphone app will allow to monitor the patient’s status and to enhance patient-therapist communication. Some limitations may be related to the short follow-up duration and the lack of evaluation of psychosocial factors. Overall, this new way of promoting long-term effects in patients with a chronic rheumatic disease could be feasible and easy to implement in daily life clinical practice and current musculoskeletal care. Trial registration: ClinicalTrials.gov NCT04263974. Registered on 7 March 2020. Date of last update 15 April 2020. Ethics committee code: PI_RH_2018.
A new animal model of atrophy–hypertrophy complex and liver damage following Yttrium-90 lobar selective internal radiation therapy in rabbits
(Nature Publishing Group, 2022) Páramo, María; Santamaría, Eva; Idoate Gastearena, Miguel Ángel; Rodríguez Fraile, Macarena; Benito, Alberto; Collantes, María; Iñarrairaegui, Mercedes; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica
Lobar selective internal radiation therapy (SIRT) is widely used to treat liver tumors inducing atrophy of the treated lobe and contralateral hypertrophy. The lack of animal model has precluded further investigations to improve this treatment. We developed an animal model of liver damage and atrophy–hypertrophy complex after SIRT. Three groups of 5–8 rabbits received transportal SIRT with Yttrium 90 resin microspheres of the cranial lobes with different activities (0.3, 0.6 and 1.2 GBq), corresponding to predicted absorbed radiation dose of 200, 400 and 800 Gy, respectively. Another group received non-loaded microspheres (sham group). Cranial and caudal lobes volumes were assessed using CT volumetry before, 15 and 30 days after SIRT. Liver biochemistry, histopathology and gene expression were evaluated. Four untreated rabbits were used as controls for gene expression studies. All animals receiving 1.2 GBq were euthanized due to clinical deterioration. Cranial SIRT with 0.6 GBq induced caudal lobe hypertrophy after 15 days (median increase 34% -ns-) but produced significant toxicity. Cranial SIRT with 0.3 GBq induced caudal lobe hypertrophy after 30 days (median increase 82%, p = 0.04). No volumetric changes were detected in sham group. Transient increase in serum transaminases was detected in all treated groups returning to normal values at 15 days. There was dose-dependent liver dysfunction with bilirubin elevation and albumin decrease. Histologically, 1.2 GBq group developed permanent severe liver damage with massive necrosis, 0.6 and 0.3 GBq groups developed moderate damage with inflammation and portal fibrosis at 15 days, partially recovering at 30 days. There was no difference in the expression of hepatocyte function and differentiation genes between 0.3 GBq and control groups. Cranial SIRT with 0.3 GBq of 90Y resin microspheres in rabbits is a reliable animal model to analyse the atrophy–hypertrophy complex and liver damage without toxicity.
A Novel NFIX-STAT6 Gene Fusion in Solitary Fibrous Tumor: A Case Report
(MDPI, 2021-07-13) Moura, David; Díaz Martín, Juan; Bagué, Silvia; Orellana-Fernandez, Ruth; Sebio, Ana; Salguero-Aranda, Carmen; Martin-Broto, Javier; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica
Solitary fibrous tumor is a rare subtype of soft-tissue sarcoma with a wide spectrum of histopathological features and clinical behaviors, ranging from mildly to highly aggressive tumors. The defining genetic driver alteration is the gene fusion NAB2–STAT6, resulting from a paracentric inversion within chromosome 12q, and involving several different exons in each gene. STAT6 (signal transducer and activator of transcription 6) nuclear immunostaining and/or the identification of NAB2–STAT6 gene fusion is required for the diagnostic confirmation of solitary fibrous tumor. In the present study, a new gene fusion consisting of Nuclear Factor I X (NFIX), mapping to 19p13.2 and STAT6, mapping to 12q13.3 was identified by targeted RNA-Seq in a 74-year-old female patient diagnosed with a deep-seated solitary fibrous tumor in the pelvis. Histopathologically, the neoplasm did not display nuclear pleomorphism or tumor necrosis and had a low proliferative index. A total of 378 unique reads spanning the NFIXexon8–STAT6exon2 breakpoint with 55 different start sites were detected in the bioinformatic analysis, which represented 59.5% of the reads intersecting the genomic location on either side of the breakpoint. Targeted RNA-Seq results were validated by RT-PCR/ Sanger sequencing. The identification of a new gene fusion partner for STAT6 in solitary fibrous tumor opens intriguing new hypotheses to refine the role of STAT6 in the sarcomatogenesis of this entity.
A Prospective Cohort of SARS-CoV-2-Infected Health Care Workers: Clinical Characteristics, Outcomes, and Follow-up Strategy
(Oxford University Press, 2020-12-08) Nicolás, David; Camós-Carreras, Anna; Spencer, Felipe; Arenas, Andrea; Fernández Rodríguez, Ana; Llop, Lourdes; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica
Background During the coronavirus disease 2019 (COVID-19) outbreaks, health care workers (HCWs) are at a high risk of infection. Strategies to reduce in-hospital transmission between HCWs and to safely manage infected HCWs are lacking. Our aim was to describe an active strategy for the management of COVID-19 in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)–infected HCWs and investigate its outcomes. Methods A prospective cohort study of SARS-CoV-2-infected health care workers in a tertiary teaching hospital in Barcelona, Spain, was performed. An active strategy of weekly polymerase chain reaction screening of HCWs for SARS-CoV-2 was established by the Occupational Health department. Every positive HCW was admitted to the Hospital at Home Unit with daily assessment online and in-person discretionary visits. Clinical and epidemiological data were recorded. Results Of the 590 HCWs included in the cohort, 134 (22%) were asymptomatic at diagnosis, and 15% (89 patients) remained asymptomatic during follow-up. A third of positive cases were detected during routine screening. The most frequent symptoms were cough (68%), hyposmia/anosmia (49%), and fever (41%). Ten percent of the patients required specific treatment at home, while only 4% of the patients developed pneumonia. Seventeen patients required a visit to the outpatient clinic for further evaluation, and 6 of these (1%) required hospital admission. None of the HCWs included in this cohort required intensive care unit admission or died. Conclusions Active screening for SARS-CoV-2 among HCWs for early diagnosis and stopping in-hospital transmission chains proved efficacious in our institution, particularly due to the high percentage of asymptomatic HCWs. Follow-up of HCWs in Hospital at Home units is safe and effective, with low rates of severe infection and readmission.
A Quantitative Digital Analysis of Tissue Immune Components Reveals an Immunosuppressive and Anergic Immune Response with Relevant Prognostic Significance in Glioblastoma
(MDPI, 2022) Idoate Gastearena, Miguel Ángel; López Janeiro, Álvaro; Lecumberri Aznarez, Arturo; Arana Iñiguez, Iñígo; Guillén Grima, Francisco; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Instituto de Salud Carlos III; European Commission (EC). Fondo Europeo de Desarrollo Regional (FEDER)
Objectives: Immunostimulatory therapies using immune checkpoint blockers show clinical activity in a subset of glioblastoma (GBM) patients. Several inhibitory mechanisms play a relevant role in the immune response to GBM. With the objective of analyzing the tumor immune microenvi ronment and its clinical significance, we quantified several relevant immune biomarkers. Design: We studied 76 primary (non-recurrent) GBMs with sufficient clinical follow-up, including a subgroup of patients treated with a dendritic cell vaccine. The IDH-mutation, EGFR-amplification, and MGMT methylation statuses were determined. Several relevant immune biomarkers, including CD163, CD8, PD1, and PDL1, were quantified in representative selected areas by digital image analysis and semi quantitative evaluation. The percentage of each immune expression was calculated with respect to the total number of tumor cells. Results: All GBMs were wild-type IDH, with a subgroup of classical GBMs according to the EGFR amplification (44%). Morphologically, CD163 immunostained microglia and intratumor clusters of macrophages were observed. A significant direct correlation was found between the expression of CD8 and the mechanisms of lymphocyte immunosuppression, in such a way that higher values of CD8 were directly associated with higher values of CD163 (p < 0.001), PDL1 (0.026), and PD1 (0.007). In a multivariate analysis, high expressions of CD8+ (HR = 2.05, 95%CI (1.02–4.13), p = 0.034) and CD163+ cells (HR 2.50, 95%CI (1.29–4.85), p = 0.007), were associated with shorter survival durations. The expression of immune biomarkers was higher in the non-classical (non-EGFR amplified tumors) GBMs. Other relevant prognostic factors were age, receipt of the dendritic cell vaccine, and MGMT methylation status. Conclusions: In accordance with the inverse correlation between CD8 and survival and the direct correlation between effector cells and CD163 macrophages and immune-checkpoint expression, we postulate that CD8 infiltration could be placed in a state of anergy or lymphocytic inefficient activity. Furthermore, the significant inverse correlation between CD163 tissue concentration and survival explains the relevance of this type of immune cell when creating a strong immunosuppressive environment. This information may potentially be used to support the selection of patients for immunotherapy.
A Reassessment of the Barrier Effect of the Physis against Metaphyseal Osteosarcoma: A Comprehensive Pathological Study with Its Radiological and Clinical Follow-Up Correlations
(MDPI, 2022) Idoate Gastearena, Miguel Ángel; Dámaso Aqiuerreta, Jesús; Lamo Espinosa, José María; San-Julián, Mikel; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica
Osteosarcoma is a primary malignant bone tumor usually arising at the metaphysis of long bones, particularly around the knee. The physis has been regarded as a barrier capable of blocking tumor extension, thus allowing it to preserve their epiphysis and therefore improve functional results. With the objective of clarifying how effective the physis is as a barrier to tumor spread, a large series of skeletally immature patients with osteosarcoma were reviewed. From 452 metaphyseal osteosarcomas a selection of 282 cases in which the tumor was close or crossing the physis were carried out. This sub-sample was split into two groups according to the surgical treatment (epiphyseal preservation or not). The specimens obtained by resection were studied, and the physeal and metaphyseal areas were studied by multiple sections. Immunostaining against VEGF of physis was obtained in selected cases. In about half of the patients affected by metaphyseal malignant bone tumors, the growth plate and epiphysis were not compromised by the tumor. Three sequential invasive growth patterns of an osteosarcoma in its relationship with the physis could be distinguished. An intense angiogenesis and osteoclastic reaction could be observed in the growth plate in the free zone between the tumor and the physis. The local recurrence incidence was lower in the epiphyseal preservation treated patients than it was in the conventional treatment (8% vs. 12%). Most local recurrences appeared in the first 2 years. The overall survival of patients treated with epiphyseal preservation was better than that of the patients treated without preserving the epiphysis (73% vs. 59%; p = 0.03) at a mean follow-up of 18 years. We have described an angiogenic and osteoclastic reaction in the base of the growth plate in the proximity of the advance front of the tumor, which could facilitate the osteosarcoma invasion. It is also shown that the preoperative imaging method for examination is a valid approach for the decision to carry out epiphyseal preservation. Finally, we concluded that epiphyseal preservation combine.
Acute oxidant damage promoted on cancer cells by amitriptyline in comparison with some common chemotherapeutic drugs
(Wolters Kluwer Health, 2010) Cordero, Mario David; Sánchez-Alcázar, José Antonio; Bautista-Ferrufino, María Rosa; Carmona-López, María Inés; Illanes, Matilde; Alcudia Cruz, Ana; Miguel Rodríguez, Manuel de; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica
Oxidative therapy is a relatively new anticancer strategy based on the induction of high levels of oxidative stress, achieved by increasing intracellular reactive oxygen species (ROS) and/or by depleting the protective antioxidant machinery of tumor cells. We focused our investigations on the antitumoral potential of amitriptyline in three human tumor cell lines: H460 (lung cancer), HeLa (cervical cancer), and HepG2 (hepatoma); comparing the cytotoxic effect of amitriptyline with three commonly used chemotherapeutic drugs: camptothecin, doxorubicin, and methotrexate. We evaluated apoptosis, ROS production, mitochondrial mass and activity, and antioxidant defenses of tumor cells. Our results show that amitriptyline produces the highest cellular damage, inducing high levels of ROS followed by irreversible serious mitochondrial damage. Interestingly, an unexpected decrease in antioxidant machinery was observed only for amitriptyline. In conclusion, based on the capacity of generating ROS and inhibiting antioxidants in tumor cells, amitriptyline emerges as a promising new drug to be tested for anticancer therapy.
Administration of L-arginine reduces the delay of the healing process caused by Ibuprofen. Implication of COX and growth factors expression
(Histology and Histopathology, 2005) Sánchez Fidalgo, Susana; Martín Lacave, Inés María; Illanes, Matilde; Bruseghini, Leo; Esteras, Antonio; Motilva Sánchez, Virginia; Universidad de Sevilla. Medicina Preventiva y Salud Pública; Universidad de Sevilla. Citología e Histología Normal y Patológica; Universidad de Sevilla. Departamento de Farmacología
The objective of the present study has been to advance knowledge of the gastric role played by the amino acid L-Arginine (L-Arg) in the evolution of a chronic gastric ulcer. In order to clarify it, L-Arg alone or together with Ibuprofen have been administrated in an experimental acetic acid chronic ulcer, analysing characteristic parameters of an active curative process, such as PGE2 production, COX expression, and also angiogenesis, proliferation/apoptosis and growth factors expression. Our results reveal that L-Arg is favourable in the healing process improving the curative course. Ibuprofen caused a delay in ulcer healing, more evident 14 days after ulcer induction; COX-2 expression was increased at the 7th day although no signal of protein could be detected after 14 days; PGE2 production was inhibited in intact and ulcerated areas at both times assayed. In contrast, treatment with L-Arg reduced the delay of the lesion, the increment in COX-2 expression induced by Ibuprofen, and was able to maintain PGE2 levels similar to the control group after 14 days. Additionally, the histological study showed that the healing effects of L-Arg might be associated with an increased angiogenesis and FGF-2 expression. These actions could be considered key factors in the healing response associated with L-Arg administration. However, the proliferation study assayed with the PCNA-immunostaining method did not reveal significant differences, as the same as the apoptosis analysis. In conclusion, the coupling of L-Arg to Ibuprofen is an attractive alternative to Ibuprofen administration alone because it not only attenuates but also improves the evolution of chronic lesions through mechanisms that implicate endogenous PG and FGF-2-associated pathways, which allow an increase of angiogenesis process.
Advanced sporadic renal epithelioid angiomyolipoma: Case report of an extraordinary response to sirolimus linked to TSC2 mutation
(Springer Nature, 2018-05-15) Espinosa, Marta; Roldán-Romero, Juan María; Durán, Ignacio; Álava Casado, Enrique de; Apellaniz-Ruiz, María; Cascón, Alberto; Garrigós, Carmen; Robledo, Mercedes; Rodríguez-Antona, Cristina; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Universidad de Sevilla. Departamento de Farmacología; Ministerio de Economia, Industria y Competitividad (MINECO). España
Background: Renal epithelioid angiomyolipomas (EAML) are rare tumors with aggressive behavior. EAML can be sporadic or develop within the tuberous sclerosis complex syndrome, where mutations of TSC1 or TSC2 genes (critical negative regulators of mTOR Complex 1) result in an increased activation of mTOR pathway. Optimal EAML treatment, including mTOR inhibitors, remains undetermined. Case presentation: Here we present the case of a young adult with a renal EAML that after radical nephrectomy developed metastases, first in liver and then in lumbar vertebrae. After complete surgical resection of these lesions, liver recurrence was detected, this time with incomplete surgical resection. After finding a new liver lesion, systemic treatment with sirolimus started. The patient exhibited a complete and durable response to this drug, being disease free at the time of publication, after 36 months of treatment. Targeted next generation sequencing (NGS) of MTOR, TSC1 and TSC2 genes in the primary tumor, metastasis and blood of the patient, revealed one inactivating TSC2 mutation (c.2739dup; p.K914*) in the tumor cells. Immunohistochemistry revealed decreased TSC2 protein content and increased phospho-S6 in the tumor cells, demonstrating mTOR pathway activation. Conclusion: NGS on an EAML patient with an extraordinary response to sirolimus uncovered TSC2 inactivation as the mechanism for the response. This study supports NGS as a useful tool to identify patients sensitive to mTOR inhibitors and supports the treatment of malignant EAML with these drugs.
Alpha-catenin-dependent recruitment of the centrosomal protein cap350 to adherens junctions allows epithelial cells to acquire a columnar shape
(Public Library Science, 2015-03-12) Gavilán Dorronzoro, María de la Paz; Arjona, Marina; Zurbano, Ángel; Formstecher, Etienne; Martinez-Morales, Juan R.; Bornens, Michel; Rios, Rosa M.; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Junta de Andalucía; Gobierno de España
Abstract Epithelial morphogenesis involves a dramatic reorganisation of the microtubule cytoskele ton. How this complex process is controlled at the molecular level is still largely unknown. Here, wereport that the centrosomal microtubule (MT)-binding protein CAP350 localises at adherens junctions in epithelial cells. By two-hybrid screening, we identified a direct interaction of AP350with the adhesion protein α-catenin that was further confirmed by co-immunoprecipitation experiments. Block of epithelial cadherin (E-cadherin)-mediated cell-cell adhesion or α-catenin depletion prevented CAP350 localisation at cell-cell junctions. Knocking downjunction-located CAP350 inhibited the establishment of an apico-basal array of microtubules and impaired the acquisition of columnar shape in Madin-Darby canine kidney II (MDCKII) cells grown as polarised epithelia. Furthermore, MDCKII cystogenesis was also defective in junctional CAP350-depleted cells. CAP350-depleted MDCKII cysts were smaller and contained either multiple lumens or no lumen. Membrane polarity wasnotaffected, but cortical microtubule bundles did not properly form. Our results indicate that CAP350mayactasanadaptorbetweenadherensjunctions and microtubules, thus regulating epithelial differentiation and contributing to the definition of cell architecture. We also uncover a central role of α-catenin in global cytoskeleton remodelling, in which it acts not only on actin but also on MT reorganisation during epithelial morphogenesis.
ALPL-1 is a target for chimeric antigen receptor therapy in osteosarcoma
(Nature Publishing Group; Nature Portfolio, 2023-06-08) Mensali, Nadia; Köksal, Hakan; Joaquina, Sandy; Wernhoff, Patrik; Casey, Nicholas P.; Romecin, Paola; Álava Casado, Enrique de; Wälchli, Sébastien; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Instituto de Salud Carlos III; Universidad de Sevilla. CTS1035: Patología Molecular del Cáncer Sólido
Osteosarcoma (OS) remains a dismal malignancy in children and young adults, with poor outcome for metastatic and recurrent disease. Immunotherapies in OS are not as promising as in some other cancer types due to intra-tumor heterogeneity and considerable off-target expression of the potentially targetable proteins. Here we show that chimeric antigen receptor (CAR) T cells could successfully target an isoform of alkaline phosphatase, ALPL-1, which is highly and specifically expressed in primary and metastatic OS. The target recognition element of the second-generation CAR construct is based on two antibodies, previously shown to react against OS. T cells transduced with these CAR constructs mediate efficient and effective cytotoxicity against ALPL-positive cells in in vitro settings and in state-of-the-art in vivo orthotopic models of primary and metastatic OS, without unexpected toxicities against hematopoietic stem cells or healthy tissues. In summary, CAR-T cells targeting ALPL-1 show efficiency and specificity in treating OS in preclinical models, paving the path for clinical translation.
Amitriptyline down-regulates coenzyme Q⁠10 biosynthesis in lung cancer cells
(Elsevier, 2017) Ortiz, Tamara; Villanueva-Paz, Marina; Díaz-Parrado, Eduardo; Illanes, Matilde; Fernández-Rodríguez, Ana; Sánchez-Alcázar, José A.; Miguel Rodríguez, Manuel de; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica
Amitriptyline, a tricyclic antidepressant, has been proposed as an antitumoral drug in oxidative therapy. Its pro-apoptotic effects, mediated by high reactive oxygen species generation, have been already described. In this study we analysed the effect of amitriptyline on the biosynthesis of coenzyme Q⁠10 (CoQ), an essential component for electron transport and a potent membrane antioxidant involved in redox signaling. We treated H460 cells, a non-small-cell lung cancer cell line, with amitriptyline and we analysed CoQ levels by HPLC and CoQ biosynthesis rate, as well as the enzymes involved in CoQ biosynthesis by real-time PCR and Western blot. Amitriptyline treatment induced a dose-dependent decrease in CoQ levels in tumor cells. CoQ decreased levels were associated with down-regulation of the expression of COQ4 gene, as well as decreased Coq4 and Coq6 protein levels. Our Vndings suggest that the effect of amitriptyline on CoQ biosynthesis highlights the potential of this drug for antitumoral oxidative therapy.
Amitriptyline induces mitophagy that precedes apoptosis in human HepG2 cells
(2016-08-07) Villanueva-Paz, Marina; Cordero, Mario D.; Pavón, Ana Delgado; Castejón Vega, Beatriz; Cotán, David; De la Mata, Mario; Oropesa-Ávila, Manuel; Alcocer-Gómez, Elísabet; Muntané Relat, Jordi; Miguel Rodríguez, Manuel de; Sánchez-Alcázar, José Antonio; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Universidad de Sevilla. Departamento de Fisiología Médica y Biofísica; Universidad de Sevilla. Departamento de Psicología Experimental
Systemic treatments for hepatocellular carcinoma (HCC) have been largely unsuccessful. This study investigated the antitumoral activity of Amitriptyline, a tricyclic antidepressant, in hepatoma cells. Amitriptyline-induced toxicity involved early mitophagy activation that subsequently switched to apoptosis. Amitriptyline induced mitochondria dysfunction and oxidative stress in HepG2 cells. Amitriptyline specifically inhibited mitochondrial complex III activity that is associated with decreased mitochondrial membrane potential (∆Ψm) and increased reactive oxygen species (ROS) production. Transmission electron microscopy (TEM) studies revealed structurally abnormal mitochondria that were engulfed by double-membrane structures resembling autophagosomes. Consistent with mitophagy activation, fluorescence microscopy analysis showed mitochondrial Parkin recruitment and colocalization of mitochondria with autophagosome protein markers. Pharmacological or genetic inhibition of autophagy exacerbated the deleterious effects of Amitriptyline on hepatoma cells and led to increased apoptosis. These results suggest that mitophagy acts as an initial adaptive mechanism of cell survival. However persistent mitochondrial damage induced extensive and lethal mitophagy, autophagy stress and autophagolysome permeabilization leading eventually to cell death by apoptosis. Amitriptyline also induced cell death in hepatoma cells lines with mutated p53 and non-sense p53 mutation. Our results support the hypothesis that Amitriptyline-induced mitochondrial dysfunction can be a useful therapeutic strategy for HCC treatment, especially in tumors showing p53 mutations and/or resistant to genotoxic treatments.
Amoxicillin and Clarithromycin Mucoadhesive Delivery System for Helicobacter pylori Infection in a Mouse Model: Characterization, Pharmacokinetics, and Efficacy
(MDPI, 2021-01) Villegas Lama, Isabel; Rosillo Ramírez, María de los Ángeles; Alarcón de la Lastra Romero, Catalina; Vázquez Román, María Victoria; Llorente, María; Sánchez, Susana; Fernández Campos, Francisco; Universidad de Sevilla. Departamento de Farmacología; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Ministerio de Ciencia e Innovación (MICIN). España
Helicobacter pylori is the main pathogen responsible for gastric ulcers and a predisposing factor of stomach cancer. Although current treatment is usually successful, it requires high doses and frequent administration. An innovative mucoadhesive system (Mucolast®) loaded with amoxicillin and clarithromycin is proposed to improve the efficacy of treatment against H. pylori. The drug product was optimized based on its viscoelastic properties to obtain long-term stability of the vehicle. The drug release mechanisms were different for both antibiotics based on their solubilization status. A systemic and stomach pharmacokinetic profile was obtained after three different doses were administered to mice, obtaining similar systemic exposure levels but an increase in drug concentration in the stomach. The efficacy results in mice infected with H. pylori also demonstrated the superiority of the antibiotics when administered in Mucolast®, as shown by the bacterial count in stomach tissue and under histopathological and biochemical evaluation. The proposed treatment was efficacious and safe and is presented as a realistic alternative to current treatment options to improve patient compliance and to reduce bacterial resistance.
An Immunohistochemical Profile Predictive of Recurrence and/or Mortality in Patients with Non-Special Type Invasive Mammary Carcinoma
(Sci Printers & Public Inc, 2016) Macías García, Laura; Robles-Frias, A.; García Gutiérrez, Jorge; González Campora, Ricardo; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Universidad de Sevilla. Departamento de Lenguajes y Sistemas Informáticos
OBJECTIVE: To identify and deine an immunohistochemical proile predictive of recurrence and/or mortality in patients with non-special type (NST) invasive mammary carcinoma. STUDY DESIGN: We identiied 234 patients diagnosed with NST invasive mammary carcinoma. They were followed up for at least 5 years. Tissue microarrays were constructed for immunohistochemical examination with different antibodies. RESULTS: Signiicant correlations were observed between tumor recurrence and expression of CK 5/6, fascin, FOXA1, Ki67, PR, and TOP2A, and between mortality and CDK-2 expression. Taking both disease outcomes together, signiicant correlations were observed with ER, p53, and the basal-type molecular phenotype (CK5/6). Multivariate analysis highlighted a signiicant correlation between tumor recurrence and coexpression of TOP2A/ER and between mortality and coexpression of C-KIT/ER. CONCLUSION: The present study suggests that expression of ER, PR, FOXA1, C-KIT, and TOPO2A constitutes a favorable immunohistochemical proile, while expression of CK5/6, CDK2, fascin, Ki67, and p53 may be regarded as unfavorable, since individual expression of these markers is associated with a better or worse prognosis, respectively. The immunohistochemical signature proposed here enables the identiication of breast cancer patients with a high or low risk of recurrence
An inducible ectopic expression system of EWSR1-FLI1 as a tool for understanding Ewing sarcoma oncogénesis
(Public Library of Science, 2020) García-Domínguez, Daniel J.; Hontecillas-Prieto, Lourdes; Andrés León, Eduardo; Sánchez-Molina, Sara; Rodríguez-Núñez, Pablo; Morón, Francisco J.; Hajji, Nabil; Mackintosh, Carlos; Álava Casado, Enrique de; Universidad de Sevilla. Departamento de Bioquímica Médica y Biología Molecular e Inmunología; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Asociacion Espanola Contra el Cancer (AECC); CIBERONC; Consejeria de Salud, Junta de Andalucia; European Commission (FP7HEALTH-2011-two-stage); Ministry of Economy and Competitiveness of Spain-FEDER (CIBERONC)
The presence of the chimeric EWSR1-FLI1 oncoprotein is the main and initiating event defining Ewing sarcoma (ES). The dysregulation of epigenomic and proteomic homeostasis induced by the oncoprotein contributes to a wide variety of events involved in oncogenesis and tumor progression. Attempts at studying the effects of EWSR1-FLI1 in non-tumor cells to understand the mechanisms underlying sarcomagenesis have been unsuccessful to date, as ectopic expression of EWSR1-FLI1 blocks cell cycle progression and induces apoptosis in the tested cell lines. Therefore, it is essential to find a permissive cell type for EWSR1-FLI1 expression that allows its endogenous molecular functions to be studied. Here we have demonstrated that HeLa cell lines are permissive to EWSR1-FLI1 ectopic expression, and that our model substantially recapitulates the endogenous activity of the EWSR1-FLI1 fusion protein. This model could contribute to better understanding ES sarcomagenesis by helping to understand the molecular mechanisms induced by the EWSR1-FLI1 oncoprotein.
An international working group consensus report for the prioritization of molecular biomarkers for Ewing sarcoma
(Nature Research, 2022-09-17) Shulman, David S.; Whittle, Sarah B.; Surdez, Didier; Bailey, Kelly M.; Álava Casado, Enrique de; Yustein, Jason T.; Grohar, Patrick J.; Citología e Histología Normal y Patológica
The advent of dose intensified interval compressed therapy has improved event-free survival for patients with localized Ewing sarcoma (EwS) to 78% at 5 years. However, nearly a quarter of patients with localized tumors and 60–80% of patients with metastatic tumors suffer relapse and die of disease. In addition, those who survive are often left with debilitating late effects. Clinical features aside from stage have proven inadequate to meaningfully classify patients for risk-stratified therapy. Therefore, there is a critical need to develop approaches to risk stratify patients with EwS based on molecular features. Over the past decade, new technology has enabled the study of multiple molecular biomarkers in EwS. Preliminary evidence requiring validation supports copy number changes, and loss of function mutations in tumor suppressor genes as biomarkers of outcome in EwS. Initial studies of circulating tumor DNA demonstrated that diagnostic ctDNA burden and ctDNA clearance during induction are also associated with outcome. In addition, fusion partner should be a pre-requisite for enrollment on EwS clinical trials, and the fusion type and structure require further study to determine prognostic impact. These emerging biomarkers represent a new horizon in our understanding of disease risk and will enable future efforts to develop risk-adapted treatment.
Anti-inflammatory effects of rosmarinic acid-loaded nanovesicles in acute colitis through modulation of NLRP3 inflammasome
(MDPI, 2021-01-26) Marinho, Sonia; Illanes, Matilde; Ávila Román, Francisco Javier; Motilva Sánchez, Virginia; Talero Barrientos, Elena Mª; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Universidad de Sevilla. Departamento de Farmacología, Pediatría y Radiología
Ulcerative colitis (UC), one of the two main types of inflammatory bowel disease, has no effective treatment. Rosmarinic acid (RA) is a polyphenol that, when administered orally, is metabolised in the small intestine, compromising its beneficial effects. We used chitosan/nutriosecoated niosomes loaded with RA to protect RA from gastric degradation and target the colon and evaluated their effect on acute colitis induced by 4% dextran sodium sulphate (DSS) for seven days in mice. RA-loaded nanovesicles (5, 10 and 20 mg/kg) or free RA (20 mg/kg) were orally administered from three days prior to colitis induction and during days 1, 3, 5 and 7 of DSS administration. RA-loaded nanovesicles improved body weight loss and disease activity index as well as increased mucus production and decreased myeloperoxidase activity and TNF-_ production. Moreover, RA-loaded nanovesicles downregulated protein expression of inflammasome components such as NLR family pyrin domain-containing 3 (NLRP3), adaptor protein (ASC) and caspase-1, and the consequent reduction of IL-1_ levels. Furthermore, nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) protein expression increased after the RA-loaded nanovesicles treatment However, these mechanistic changes were not detected with the RA-free treatment. Our findings suggest that the use of chitosan/nutriose-coated niosomes to increase RA local bioavailability could be a promising nutraceutical strategy for oral colon-targeted UC therapy.
Anti-obesogenic effect of lupin-derived protein hydrolysate through modulation of adiposopathy, insulin resistance and gut dysbiosis in a diet-induced obese mouse
(Elsevier, 2024) Ponce España, Eduardo; Cruz Chamorro, Iván; Santos Sánchez, Guillermo; Álvarez López, Ana Isabel; Fernández Santos, José María; Pedroche, Justo; Millán Linares, María del Carmen; Bejarano Hernando, Ignacio; Lardone, Patricia Judith; Carrillo Vico, Antonio; Universidad de Sevilla. Departamento de Bioquímica Médica y Biología Molecular e Inmunología; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica
The prevalence of obesity is increasingly widespread, resembling a global epidemic. Lifestyle changes, such as consumption of high-energy-dense diets and physical inactivity, are major contributors to obesity. Common features of this metabolic pathology involve an imbalance in lipid and glucose homeostasis including dyslipidemia, insulin resistance and adipose tissue dysfunction. Moreover, the importance of the gut microbiota in the development and susceptibility to obesity has recently been highlighted. In recent years, new strategies based on the use of functional foods, in particular bioactive peptides, have been proposed to counteract obesity outcomes. In this context, the present study examines the effects of a lupin protein hydrolysate (LPH) on obesity, dyslipidemia and gut dysbiosis in mice fed a high-fat diet (HFD). After 12 weeks of LPH treatment, mice gained less weight and showed decreased adipose dysfunction compared to the HFD-fed group. HFD-induced dyslipidemia (increased triglycerides, cholesterol and LDL concentration) and insulin resistance were both counteracted by LPH consumption. Discriminant analysis differentially distributed LPH-treated mice compared to non-treated mice. HFD reduced gut ecological parameters, promoted the blooming of deleterious taxa and reduced the abundance of commensal members. Some of these changes were corrected in the LPH group. Finally, correlation analysis suggested that changes in this microbial population could be responsible for the improvement in obesity outcomes. In conclusion, this is the first study to show the effect of LPH on improving weight gain, adiposopathy and gut dysbiosis in the context of diet-induced obesity, pointing to the therapeutic potential of bioactive peptides in metabolic diseases.
Apoptotic microtubules delimit an active caspase free area in the cellular cortex during the execution phase of apoptosis
(NATURE PUBLISHING GROUP, 2013) Oropesa-Avila, M.; Fernandez-Vega, A.; Mata Fernández, María de la; Maraver, J. G.; Cordero, Mario D.; Cotán, D.; Miguel Rodríguez, Manuel de; Sánchez-Alcázar, José Antonio; Universidad de Sevilla. Departamento de Citología e Histología Normal y Patológica; Ministerio de Sanidad. España; Servicio Andaluz de Salud Junta de Andalucía
Apoptotic microtubule network (AMN) is organized during apoptosis, forming a cortical structure beneath plasma membrane, which has an important role in preserving cell morphology and plasma membrane permeability. The aim of this study was to examine the role of AMN in maintaining plasma membrane integrity during the execution phase of apoptosis. We demonstrated in camptothecin-induced apoptosis in H460 cells that AMN delimits an active caspase free area beneath plasma membrane that permits the preservation of cellular cortex and transmembrane proteins. AMN depolymerization in apoptotic cells by a short exposure to colchicine allowed active caspases to reach the cellular cortex and cleave many key proteins involved in plasma membrane structural support, cell adhesion and ionic homeostasis. Cleavage of cellular cortex and plasma membrane proteins, such as a-spectrin, paxilin, focal adhesion kinase (FAK), E-cadherin and integrin subunit b4 was associated with cell collapse and cell detachment. Otherwise, cleavage-mediated inactivation of calcium ATPase pump (PMCA-4) and Naþ/Ca2þ exchanger (NCX) involved in cell calcium extrusion resulted in calcium overload. Furthermore, cleavage of Naþ/Kþ pump subunit b was associated with altered sodium homeostasis. Cleavage of cell cortex and plasma membrane proteins in apoptotic cells after AMN depolymerization increased plasma permeability, ionic imbalance and bioenergetic collapse, leading apoptotic cells to secondary necrosis. The essential role of caspase-mediated cleavage in this process was demonstrated because the concomitant addition of colchicine that induces AMN depolymerization and the pan-caspase inhibitor z-VAD avoided the cleavage of cortical and plasma membrane proteins and prevented apoptotic cells to undergo secondary necrosis. Furthermore, the presence of AMN was also critical for proper phosphatidylserine externalization and apoptotic cell clearance by macrophages. These results indicate that AMN is essential to preserve an active caspase free area in the cellular cortex of apoptotic cells that allows plasma membrane integrity during the execution phase of apoptosis.

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