Trans-[Pt(amine)Cl2(PPh3)] Complexes Target Mitochondria and Endoplasmic Reticulum in Gastric Cancer Cells.

Gastric cancer prognosis is still notably poor despite efforts made to improve diagnosis and treatment of the disease. Chemotherapy based on platinum agents is generally used, regardless of the fact that drug toxicity leads to limited clinical efficacy. In order to overcome these problems, our group has been working on the synthesis and study of trans platinum (II) complexes. Here, we explore the potential use of two phosphine-based agents with the general formula trans-[Pt(amine)Cl2(PPh3)], called P1 and P2 (with dimethylamine or isopropylamine, respectively). A cytotoxicity analysis showed that P1 and especially P2 decrease cell viability. Specifically, P2 exhibits higher activity than cisplatin in gastric cancer cells while its toxicity in healthy cells is slightly lower. Both complexes generate Reactive Oxygen Species, produce DNA damage and mitochondrial membrane depolarization, and finally lead to induced apoptosis. Thus, an intrinsic apoptotic pathway emerges as the main type of cell death through the activation of BAX/BAK and BIM and the degradation of MCL1. Additionally, we demonstrate here that P2 produces endoplasmic reticulum stress and activates the Unfolded Protein Response, which also relates to the impairment observed in autophagy markers such as p62 and LC3. Although further studies in other biological models are needed, these results report the biomolecular mechanism of action of these Pt(II)-phosphine prototypes, thus highlighting their potential as novel and effective therapies.

Optimizing the Delivery of mRNA to Mesenchymal Stem Cells for Tissue Engineering Applications.

Messenger RNA (mRNA) represents a promising therapeutic tool in the field of tissue engineering for the fast and transient production of growth factors to support new tissue regeneration. However, one of the main challenges to optimizing its use is achieving efficient uptake and delivery to mesenchymal stem cells (MSCs), which have been long reported as difficult-to-transfect. The aim of this study was to systematically screen a range of nonviral vectors to identify optimal transfection conditions for mRNA delivery to MSCs. Furthermore, for the first time, we wanted to directly compare the protein expression profile from three different types of mRNA, namely, unmodified mRNA (uRNA), base-modified mRNA (modRNA), and self-amplifying mRNA (saRNA) in MSCs. A range of polymer- and lipid-based vectors were used to encapsulate mRNA and directly compared in terms of physicochemical properties as well as transfection efficiency and cytotoxicity in MSCs. We found that both lipid- and polymer-based materials were able to successfully condense and encapsulate mRNA into nanosized particles (<200 nm). The overall charge and encapsulation efficiency of the nanoparticles was dependent on the vector type as well as the vector:mRNA ratio. When screened in vitro, lipid-based vectors proved to be superior in terms of mRNA delivery to MSCs cultured in a 2D monolayer and from a 3D collagen-based scaffold with minimal effects on cell viability, thus opening the potential for scaffold-based mRNA delivery. Modified mRNA consistently showed the highest levels of protein expression in MSCs, demonstrating 1.2-fold and 5.6-fold increases versus uRNA and saRNA, respectively. In summary, we have fully optimized the nonviral delivery of mRNA to MSCs, determined the importance of careful selection of the mRNA type used, and highlighted the strong potential of mRNA for tissue engineering applications.

Platinum iodido drugs show potential anti-tumor activity, affecting cancer cell metabolism and inducing ROS and senescence in gastrointestinal cancer cells.

Cisplatin-based chemotherapy has associated clinical disadvantages, such as high toxicity and resistance. Thus, the development of new antitumor metallodrugs able to overcome different clinical barriers is a public healthcare priority. Here, we studied the mechanism of action of the isomers trans and cis-[PtI2(isopropylamine)2] (I5 and I6, respectively) against gastrointestinal cancer cells. We demonstrate that I5 and I6 modulate mitochondrial metabolism, decreasing OXPHOS activity and negatively affecting ATP-linked oxygen consumption rate. Consequently, I5 and I6 generated Reactive Oxygen Species (ROS), provoking oxidative damage and eventually the induction of senescence. Thus, herein we propose a loop with three interconnected processes modulated by these iodido agents: (i) mitochondrial dysfunction and metabolic disruptions; (ii) ROS generation and oxidative damage; and (iii) cellular senescence. Functionally, I5 reduces cancer cell clonogenicity and tumor growth in a pancreatic xenograft model without systemic toxicity, highlighting a potential anticancer complex that warrants additional pre-clinical studies.

Dithiobiureas Palladium(II) complexes' studies: From their synthesis to their biological action.

Dithiobiureas coordination chemistry towards palladium (II) ions and their possible application is presented and discussed. 1,6-(4-Methoxyphenyl)-2,5-dithiobiurea and 1,6-(4-chlorophenyl)-2,5-dithiobiurea afford two Pd(II) complexes with the general formula [Pd2(H2L)Cl2(PPh3)2]. The metal ion forms one chelate ring with the dithiobiurea, and binds to a triphenylphosphine and an additional leaving group cisplatin like. One of the complexes (1) is endowed not only with stability in DMSO and aqua solutions containing a biological buffer but also with cytotoxicity versus gastric cancer cell lines. Complex 1 does not interact covalently to DNA models, neither activates p53 or Checkpoint Kinase 1 key proteins for DNA damage response. Thus, we propose that complex 1 exerts its action by activating Mitogen-Activated Protein Kinases [p38, Extracellular signal-regulated kinases (ERKs), c-Jun N-terminal kinases (JNKs)] as cell death inductors.

Self-amplifying RNA vaccine protects mice against lethal Ebola virus infection.

Emerging and re-emerging viruses, such as Zaire Ebola virus (EBOV), pose a global threat and require immediate countermeasures, including the rapid development of effective vaccines that are easy to manufacture. Synthetic self-amplifying RNAs (saRNAs) attend to these needs, being safe and strong immune stimulators that can be inexpensively produced in large quantities, using cell-free systems and good manufacturing practice. Here, the first goal was to develop and optimize an anti-EBOV saRNA-based vaccine in terms of its antigen composition and route of administration. Vaccinating mice with saRNAs expressing the EBOV glycoprotein (GP) alone or in combination with the nucleoprotein (NP) elicited antigen-specific immune responses. GP-specific antibodies showed neutralizing activity against EBOV. Strong CD4+ T cell response against NP and GP and CD8+ T cell response against NP were detected by ELISpot assays. Intramuscular vaccination with saRNAs conferred better immune response than intradermal. Finally, mice vaccinated in a prime-boost regimen with saRNAs encoding both GP and NP or with GP alone survived an EBOV infection. In addition, a single dose of GP and NP saRNAs was also protective against fatal EBOV infection. Overall, saRNAs expressing viral antigens represent a promising vaccine platform.

Immunogenicity of stabilized HIV-1 Env trimers delivered by self-amplifying mRNA.

Self-amplifying mRNA (saRNA) represents a promising platform for nucleic acid delivery of vaccine immunogens. Unlike plasmid DNA, saRNA does not require entry into the nucleus of target cells for expression, having the capacity to drive higher protein expression compared to mRNA as it replicates within the cytoplasm. In this study, we examined the potential of stabilized native-like HIV-1 Envelope glycoprotein (Env) trimers to elicit immune responses when delivered by saRNA polyplexes (PLXs), assembled with linear polyethylenimine. We showed that Venezuelan equine encephalitis virus (VEEV) saRNA induces a stronger humoral immune response to the encoded transgene compared to Semliki Forest virus saRNA. Moreover, we characterized the immunogenicity of the soluble and membrane-bound ConSOSL.UFO Env design in mice and showed a faster humoral kinetic and an immunoglobulin G (IgG)2a skew using a membrane-bound design. The immune response generated by PLX VEEV saRNA encoding the membrane-bound Env was then evaluated in larger animal models including macaques, in which low doses induced high IgG responses. Our data demonstrated that the VEEV saRNA PLX nanoparticle formulation represents a suitable platform for the delivery of stabilized HIV-1 Env and has the potential to be used in a variety of vaccine regimens.

Clinical characterization and outcomes of 85 patients with neurosarcoidosis.

To analyze the frequency and clinical phenotype of neurosarcoidosis (NS) in one of the largest nationwide cohorts of patients with sarcoidosis reported from southern Europe. NS was evaluated according to the Diagnostic Criteria for Central Nervous System and Peripheral Nervous System Sarcoidosis recently proposed by Stern et al. Pathologic confirmation of granulomatous disease was used to subclassify NS into definite (confirmation in neurological tissue), probable (confirmation in extraneurological tissue) and possible (no histopathological confirmation of the disease). Of the 1532 patients included in the cohort, 85 (5.5%) fulfilled the Stern criteria for NS (49 women, mean age at diagnosis of NS of 47.6 years, 91% White). These patients developed 103 neurological conditions involving the brain (38%), cranial nerves (36%), the meninges (3%), the spinal cord (10%) and the peripheral nerves (14%); no patient had concomitant central and peripheral nerve involvements. In 59 (69%) patients, neurological involvement preceded or was present at the time of diagnosis of the disease. According to the classification proposed by Stern et al., 11 (13%) were classified as a definite NS, 61 (72%) as a probable NS and the remaining 13 (15%) as a possible NS. In comparison with the systemic phenotype of patients without NS, patients with CNS involvement presented a lower frequency of thoracic involvement (82% vs 93%, q = 0.018), a higher frequency of ocular (27% vs 10%, q < 0.001) and salivary gland (15% vs 4%, q = 0.002) WASOG involvements. In contrast, patients with PNS involvement showed a higher frequency of liver involvement (36% vs 12%, p = 0.02) in comparison with patients without NS. Neurosarcoidosis was identified in 5.5% of patients. CNS involvement prevails significantly over PNS involvement, and both conditions do not overlap in any patient. The systemic phenotype associated to each involvement was clearly differentiated, and can be helpful not only in the early identification of neurological involvement, but also in the systemic evaluation of patients diagnosed with neurosarcoidosis.

Chemical and Biological Evaluation of Thiosemicarbazone-Bearing Heterocyclic Metal Complexes.

Thiosemicarbazones (TSCNs) constitute a broad family of compounds (R1R2C=N-NH-C(S)- NR3R4), particularly attractive because many of them display some biological activity against a wide range of microorganisms and cancer cells. Their activity can be related to their electronic and structural properties, which offer a rich set of donor atoms for metal coordination and a high electronic delocalization providing different binding modes for biomolecules. Heterocycles such as pyrrole, imidazole and triazole are present in biological molecules such as Vitamine B12 and amino acids and could potentially target multiple biological processes. Considering this, we have explored the chemistry and biological properties of thiosemicarbazones series and their complexes bearing heterocycles such as pyrrole, imidazole, thiazole and triazole. We focus at the chemistry and cytotoxicity of those derivatives to find out the structure activity relationships, and particularly we analyzed those examples with the TSCN units in which the mechanism of action information has been profoundly studied and pathways determined, to promote future studies for heterocycle derivatives.

New Platinum(II) Triazole Thiosemicarbazone Complexes: Analysis of Their Reactivity and Potential Antitumoral Action.

The synthesis and characterization of three new platinum complexes, with 3,5-diacetyl-1,2,4-triazole bis(4-N-isopropylthiosemicarbazone) as a ligand, are reported. The specific conditions under which solvent coordination takes place are reported and the X-ray structure of the complex with one solvent molecule of dimethyl sulfoxide is resolved. Analysis of the reactivity of these platinum compounds aids in finding the best solution profile for biological investigations. Then, the interactions of the complexes with biological models, such as calf-thymus DNA, are studied by using UV spectroscopy and tracking the changes in electrophoretic mobility produced in the supercoiled plasmid DNA model. Initial screening of these potential antitumoral compounds indicates possible selective antitumoral action.

Side effects, complications and outcome of thoracoscopic sympathectomy for palmar and axillary hyperhidrosis in 406 patients.

BACKGROUND: Thoracic sympathectomy (TS) is the treatment of choice for severe primary hyperhidrosis. However, complications, side effects and satisfaction have not been well defined. OBJECTIVE: To analyze the complications, side effects, satisfaction degree and quality of life of patients after TS for primary upper limb hyperhidrosis. METHODS: One-year follow-up after 406 consecutive TS for primary upper limb hyperhidrosis. RESULTS: Bilateral TS was completed in all patients. Complications arose in 23 cases (5.6%), with pneumothorax being the most frequent. The success rate after discharge, 6 and 12 months was respectively, 100%, 98.1% and 96.5% for palmo-axillary hyperhidrosis; 100%, 99.3% and 97.8% for isolated palmar hyperhidrosis and 100%, 85.7% and 71.4% for isolated axillary hyperhidrosis. No persistence of hyperhidrosis was observed. Global recurrence was 3.7% (28.5% axillary hyperhidrosis group). Compensatory sweating (CS) appeared in 55% and was not related to the extension of the TS. Being female was a predisposing factor of CS (p<0.004). Excessive dryness appeared at 9% and was associated with extensive TS (P<0.001). Plantar hyperhidrosis improved at 33.6%, worsened at 10% and remained stable during the follow-up. Satisfaction degree decreased with the passage of time and was associated with recurrence. Quality of life was excellent at discharge, 6 and 12 month in 100%, 100% and 97%, respectively. CONCLUSIONS: Pneumothorax is the most frequent complication of TS. CS is the main and undesirable side effect, appears with the passage of time, and is not related to the extension of TS. Being female is the only predictor factor of suffering CS. Plantar hyperhidrosis improves initially, although tends to reappear. Excessive dryness appears in extensive TS and does not improve over time. Postoperative satisfaction degree is high but decreases over time owing to the appearance of recurrence. Effectiveness and the absence of CS determine an excellent quality of life. Six percent of the patients regret the surgery because of severe CS. Informing patients of possible side effects before TS is essential.

[Indicators of severity in chest trauma]. / Indicadores de gravedad en los traumatismos torácicos.

OBJECTIVE: We undertook a review of patients with chest trauma attended between January 1992 and June 2005 in order to establish severity criteria in these cases. PATIENTS AND METHODS: During the study period, 1,772 cases (1,346 [76%] males) were treated, with ages ranging from 7 to 98 years (mean, 46.4 years). The Revised Trauma Score (RTS) was calculated and the following variables were also studied as potential indicators of severity: age, extent of the injury, number of rib fractures, presence of lung contusion, hemothorax, cardiorespiratory repercussions, and need for mechanical ventilation. RESULTS: At the time of admission, 84.4% of patients presented only symptoms related to the injury, with no general repercussions, and 66.7% had an RTS of 12. The number of rib fractures was a reliable indicator of severity, as was the presence of multiple injuries, lung contusion, need for mechanical ventilation, and cardiorespiratory repercussions. Neither age nor presence of hemothorax was found to be an indicator of severity. Pleural drainage was performed in 756 cases and was effective in 670 (88.6%). CONCLUSIONS: There are a number of indicators of severity in chest trauma, related more closely to the type and repercussions of the trauma than to the age of the patient. There is a high incidence of fluid or gas accumulation in the pleural space, though this can be easily managed by pleural drainage, which constitutes the main therapeutic procedure in chest trauma.

Indicadores de gravedad en los traumatismos torácicos / Indicators of severity in chest trauma

Objetivo: Revisamos la experiencia en traumatismos torácicos (TT) atendidos entre enero de 1992 y junio de 2005 con la intención de establecer criterios de gravedad en ellos. Pacientes y métodos: Durante dicho período se atendieron 1.772 casos con edades comprendidas entre los 7 y 98 años (1.346 varones; 76%) y con una media de edad de 46,4 años. Se aplicó la escala Revised Trauma Score (RTS) y se tuvieron en cuenta, como indicadores, la edad, el grado de traumatismo, el número de fracturas costales, la presencia de contusión pulmonar, hemotórax, la repercusión cardiorrespiratoria y la necesidad de ventilación mecánica. Resultados: En el momento del ingreso el 84,4% tenía únicamente síntomas relacionados con el traumatismo, sin repercusión general, y el 66,7% presentaba un índice RTS de 12. El número de fracturas costales fue un marcador eficaz de gravedad, al igual que la presencia de politraumatismo, contusión pulmonar, necesidad de ventilación mecánica y repercusión cardiorrespiratoria. La edad no se reveló como un índice de gravedad, y tampoco la presencia de hemotórax. En 756 ocasiones se realizó drenaje pleural, que fue resolutivo en 670 (88,6%). Conclusiones: Los TT tienen una serie de indicadores de gravedad, más relacionados con el tipo e impacto del traumatismo que con la edad del paciente. El síndrome de ocupación pleural es un problema de mucha incidencia, aunque de resolución fácil con un drenaje pleural. Éste constituye la principal arma terapéutica en los TT

Objective: We undertook a review of patients with chest trauma attended between January 1992 and June 2005 in order to establish severity criteria in these cases. Patients and methods: During the study period, 1772 cases (1346 [76%] males) were treated, with ages ranging from 7 to 98 years (mean, 46.4 years). The Revised Trauma Score (RTS) was calculated and the following variables were also studied as potential indicators of severity: age, extent of the injury, number of rib fractures, presence of lung contusion, hemothorax, cardiorespiratory repercussions, and need for mechanical ventilation. Results: At the time of admission, 84.4% of patients presented only symptoms related to the injury, with no general repercussions, and 66.7% had an RTS of 12. The number of rib fractures was a reliable indicator of severity, as was the presence of multiple injuries, lung contusion, need for mechanical ventilation, and cardiorespiratory repercussions. Neither age nor presence of hemothorax was found to be an indicator of severity. Pleural drainage was performed in 756 cases and was effective in 670 (88.6%). Conclusions: There are a number of indicators of severity in chest trauma, related more closely to the type and repercussions of the trauma than to the age of the patient. There is a high incidence of fluid or gas accumulation in the pleural space, though this can be easily managed by pleural drainage, which constitutes the main therapeutic procedure in chest trauma

Elastofibroma of the thoracic wall.

Soft tissue tumors of the chest wall are rare. Between 1998 and 2007 we treated eight cases of elastofibroma of the thoracic wall, an infrequent primary tumor of the chest. Seven females and one male between 44 and 62 years presented with dorsal subscapular tumors of months and even years of evolution. One case was a relapse from previous interventions and in three cases the tumor was bilateral. A surgical excision was performed in all cases, confirming the source of the tumor. There were no postoperative complications or relapses. We concluded that elastofibroma is a tumor that appears most frequently in middle aged women, and that diagnosis can be established through the use of imaging and fine needle aspiration biopsy. Given its benign character and slow growth, in cases where it is asymptomatic, its evolution can be controlled without surgical intervention.

Dirofilariasis pulmonar. Una causa de nódulo pulmonar solitario / Pulmonary dirofilariasis. A cause of solitary pulmonary nodule

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Ecografia intraoperatoria en los tumores hepáticos / Intraoperative ecography in hepatic tumors

Luego de una introducción donde se hace una breve reseña, histórica, se presentan 3 casos en los que se realizó ecografía intraoperatoria en la División Cirugía General del Hospital Pirovano. Se hace referencia a la metodología exploratoria durante la realización de la ecografía peroperatoria, y a la experiencia de autores extrajeros que avalan el trabajo. Concluyendo que el método es sumamente apto para el diagnóstico de pequeños tumores intrahepáticos no diagnósticados preoperatoriamente por otros estudios por imágenes y no palpables ni visibles en el acto quirúrgico (AU)

Ecografia intraoperatoria en los tumores hepáticos / Intraoperative ecography in hepatic tumors

Luego de una introducción donde se hace una breve reseña, histórica, se presentan 3 casos en los que se realizó ecografía intraoperatoria en la División Cirugía General del Hospital Pirovano. Se hace referencia a la metodología exploratoria durante la realización de la ecografía peroperatoria, y a la experiencia de autores extrajeros que avalan el trabajo. Concluyendo que el método es sumamente apto para el diagnóstico de pequeños tumores intrahepáticos no diagnósticados preoperatoriamente por otros estudios por imágenes y no palpables ni visibles en el acto quirúrgico