Submandibular puncture with injection needle in mice: comparison between two different procedures

La punción del plexo venoso a nivel submandibular en ratones es una técnica aceptada para la obtención de volúmenes de sangre adecuados para análisis de rutina. Si bien se recomienda el uso de lancetas comerciales, en Argentina no están disponibles, por lo tanto pueden sustituirse por agujas de inyección. El objetivo de este trabajo fue comparar el uso de agujas 21G x 1? utilizando dos procedimientos de punción submandibular en ratones BALB/c de 8 semanas deedad: G1) con capuchón cortado a 10 mm (restringiendo la profundidad de la punción) y G2) sin capuchón para determinar: a) volumen de sangre obtenido, b) datos hematológicos y glucemia y c) daño tisular. El volumen de sangre obtenido por la punción submandibular con aguja sin capuchón fue significativamente mayor que el colectado con capuchón. Los valores hematológicos y la glucemia fueron normales para ambos procedimientos. No se observaron daños histológicos en ninguno de los procedimientos empleados. En conclusión, se recomienda el uso de aguja como procedimiento de refinamiento para la obtención de sangre submandibular dado que es económico, práctico, permite obtener un volumen sanguíneo moderado sin provocar daño tisular y asegura la recuperación de los animales.The puncture of the venous plexus in the submandibular region in mice is an accepted technique to obtain blood volumes suitable for routine clinical analysis. Although the use of commercial lancets is recommended, in Argentina they are not available and they could be replaced by injection needles. The aim of this study was to compare the use of 21G x 1" needles using two different submandibular puncture procedures in 8-week-old BALB/c mice: G1) with cap cut to 10 mm (restricting the depth of the puncture) and, G2) without cap to determine: a) blood volume obtained, b) hematological data and glycaemia and c) tissue damage. The blood volume obtained by submandibular needle puncture without cap was significantly higher than the one collected with cap. Hematological values and glycaemia were normal for both procedures. No histological damage was observed in any of the procedures used. In conclusion, it is recommended the use of needle without cap as a refining procedure to collect submandibular blood since it is cheapest, practical, allows to obtain moderate volumes no causing tissue damage and ensuring the animal recovery.Fil: Fernandez, Vanesa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; ArgentinaFil: Dominguez, Maria Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; ArgentinaFil: Garcia, Jorge P.. Universidad Nacional del Centro de la Provincia de Buenos Aires. Facultad de Ciencias Veterinarias. Departamento de Sanidad Animal y Medicina Preventiva. Laboratorio de Inmunoquímica y Biotecnología; ArgentinaFil: Estein, Silvia Marcela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; Argentin

Detection of equine herpesvirus 1 genome 1B in Argentina

To determine the genomic variation of equine herpesviruses (EHVs) isolated in Argentina between 1979 and the first half of 2004, DNA sequences from all 69 strains isolated were analysed. Sixty strains were recovered from aborted fetuses, one from leucocyte-rich plasma from a horse with respiratory signs and eight from cases of neonatal disease. The DNA was extracted from rabbit kidney epithelial (RK13) cells infected with each strain and digested with three restriction endonucleases (BamHI, Bg/II and KpnI). Two strains could be differentiated using BamHI restriction and were assigned to the EHV-1 1B prototype group. Only one of these two strains was typed EHV-1 1B with Bg/II. DNA digestion with KpnI was ineffective. The results obtained in this study demonstrate that the EHV-1 1B genome has been present in Argentina since at least 1996. The finding of two strains with this electropherotype suggests that there is genomic heterogeneity among Argentinian isolates.Facultad de Ciencias Veterinaria

Solvent-Free Microwave Extraction of Thymus mastichina Essential Oil: Influence on Their Chemical Composition and on the Antioxidant and Antimicrobial Activities

This article belongs to the Special Issue Antiradical, Chemopreventive and Antimicrobial Analysis of Bioactive SubstancesSolvent-free microwave extraction (SFME) is a combination of microwave heating and dry distillation performed at atmospheric pressure without the addition of water or organic solvents that has been proposed as a green method for the extraction of essential oils from aromatic and medicinal herbs. In this work, SFME and the conventional techniques of steam distillation (SD) and hydrodistillation (HD) were compared with respect to the extraction and antioxidant and antimicrobial activities of Thymus mastichina essential oil. The main constituent of essential oils obtained using different methods was 1,8-cineole (eucalyptol). The results showed that the essential oils extracted by means of SFME in 30 min were quantitatively (yield) and qualitatively (aromatic profile) similar to those obtained using conventional HD over 120 min. In addition, SFME generates less waste and less solvent, consumes less energy, and provides a higher yield for a shorter extraction time, which is advantageous for the extraction of the T. mastichina essential oil compared to SD. The antioxidant and antimicrobial activities of the T. mastichina essential oil obtained from either SFME or conventional extraction methods (SD or HD) showed a similar pattern. Large-scale experiments using this SFME procedure showed a potential industrial application.The authors acknowledge the Fundação para a Ciência e Tecnologia (FCT), Fundo Europeu de Desenvolvimento Regional (FEDER), and COMPETE 2020 for their financial support under theresearch project “The development of dermo-biotechnological applications using natural resourcesin the Beira and Serra da Estrela regions—DermoBio”, ref. SAICT-POL/23925/2016, presentedin the Notice for the Presentation of Applications No. 02/SAICT/2016—Scientific Research andTechnological Development Projects (IC & DT) in Co-Promotion (PDF) Solvent-Free Microwave Extraction of Thymus mastichina Essential Oil: Influence on Their Chemical Composition and on the Antioxidant and Antimicrobial Activities.info:eu-repo/semantics/publishedVersio

Integration of miRNA and mRNA expression profles reveals microRNA-regulated networks during muscle wasting in cardiac cachexia

Cardiac cachexia (CC) is a common complication of heart failure (HF) associated with muscle wasting and poor patient prognosis. Although different mechanisms have been proposed to explain muscle wasting during CC, its pathogenesis is still not understood. Here, we described an integrative analysis between miRNA and mRNA expression profiles of muscle wasting during CC. Global gene expression profiling identified 1,281 genes and 19 miRNAs differentially expressed in muscle wasting during CC. Several of these deregulated genes are known or putative targets of the altered miRNAs, including miR-29a-3p, miR-29b-3p, miR-210-5p, miR-214, and miR-489. Gene ontology analysis on integrative mRNA/miRNA expression profiling data revealed miRNA interactions affecting genes that regulate extra-cellular matrix (ECM) organization, proteasome protein degradation, citric acid cycle and respiratory electron transport. We further identified 11 miRNAs, including miR-29a-3p and miR-29b-3p, which target 21 transcripts encoding the collagen proteins related to ECM organization. Integrative miRNA and mRNA global expression data allowed us to identify miRNA target genes involved in skeletal muscle wasting in CC. Our functional experiments in C2C12 cells confirmed that miR-29b down-regulates collagen genes and contributes to muscle cell atrophy. Collectively, our results suggest that key ECM-associated miRNAs and their target genes may contribute to CC in HF

Lung Cytokines and Systemic Inflammation in Patients with COPD

Rationale: Chronic obstructive pulmonary disease (COPD) is characterized by lung and systemic inflammation. The role of cytokines in local and systemic inflammation in COPD is not well understood. This study aimed to compare plasma and bronchoalveolar lavage (BAL) fluid cytokine levels in COPD and non-COPD subjects with the intent of better understand their potential roles in driving local and systemic inflammation. Methods: This cross-sectional study analyzed data from 65 subjects: 31 with COPD confirmed by spirometry and 34 non-COPD controls. All subjects underwent spirometry, plasma sample collection, and bronchoscopy/BAL. Levels of 21 inflammatory cytokines were measured in the plasma (systemic inflammation) and BAL (lung inflammation) using a multiplex assay. Results:COPD subjects were overall older (median age 59 vs 36; p = Conclusion: Elevated levels of cytokines were identified in the plasma of COPD subjects when compared to controls, supporting the role of these mediators as one of the mechanisms of systemic inflammation in COPD. In contrast, lung cytokines were not elevated suggesting that inflammation in the setting of COPD may not originate and/or perpetuate in the lungs, or that the BAL fluid is not an optimal source of information when evaluating inflammation in COPD. Although the role of these cytokines remains uncertain, anti-cytokine therapy might modulate inflammation in COPD and perhaps improve outcomes

IND-Enabling Studies for a Clinical Trial to Genetically Program a Persistent Cancer-Targeted Immune System

PURPOSE: To improve persistence of adoptively transferred T-cell receptor (TCR)-engineered T cells and durable clinical responses, we designed a clinical trial to transplant genetically-modified hematopoietic stem cells (HSCs) together with adoptive cell transfer of T cells both engineered to express an NY-ESO-1 TCR. Here, we report the preclinical studies performed to enable an investigational new drug (IND) application. EXPERIMENTAL DESIGN: HSCs transduced with a lentiviral vector expressing NY-ESO-1 TCR and the PET reporter/suicide gene HSV1-sr39TK and T cells transduced with a retroviral vector expressing NY-ESO-1 TCR were coadministered to myelodepleted HLA-A2/Kb mice within a formal Good Laboratory Practice (GLP)-compliant study to demonstrate safety, persistence, and HSC differentiation into all blood lineages. Non-GLP experiments included assessment of transgene immunogenicity and in vitro viral insertion safety studies. Furthermore, Good Manufacturing Practice (GMP)-compliant cell production qualification runs were performed to establish the manufacturing protocols for clinical use. RESULTS: TCR genetically modified and ex vivo-cultured HSCs differentiated into all blood subsets in vivo after HSC transplantation, and coadministration of TCR-transduced T cells did not result in increased toxicity. The expression of NY-ESO-1 TCR and sr39TK transgenes did not have a detrimental effect on gene-modified HSC's differentiation to all blood cell lineages. There was no evidence of genotoxicity induced by the lentiviral vector. GMP batches of clinical-grade transgenic cells produced during qualification runs had adequate stability and functionality. CONCLUSIONS: Coadministration of HSCs and T cells expressing an NY-ESO-1 TCR is safe in preclinical models. The results presented in this article led to the FDA approval of IND 17471

Using Cluster Analysis of Cytokines to Identify Patterns of Inflammation in Hospitalized Patients with Community-acquired Pneumonia:A Pilot Study

Purpose: Patients with severe community-acquired pneumonia (CAP) are believed to have an exaggerated inflammatory response to bacterial infection. Therapies aiming to modulate the inflammatory response have been largely unsuccessful, perhaps reflecting that CAP is a heterogeneous disorder that cannot be modulated by a single anti-inflammatory approach. We hypothesize that the host inflammatory response to pneumonia may be characterized by distinct cytokine patterns, which can be harnessed for personalized therapies. Methods: Here, we use hierarchical cluster analysis of cytokines to examine if patterns of inflammatory response in 13 hospitalized patients with CAP can be defined. This was a secondary data analysis of the Community-Acquired Pneumonia Inflammatory Study Group (CAPISG) database. The following cytokines were measured in plasma and sputum on the day of admission: interleukin (IL)-1β, IL-1 receptor antagonist (IL-1ra), IL-6, CXCL8 (IL-8), IL-10, IL-12p40, IL-17, interferon (IFN)γ, tumor necrosis factor (TNF)α, and CXCL10 (IP-10). Hierarchical agglomerative clustering algorithms were used to evaluate clusters of patients within plasma and sputum cytokine determinations. Results: A total of thirteen patients were included in this pilot study. Cluster analysis identified distinct inflammatory response patterns of cytokines in the plasma, sputum, and the ratio of plasma to sputum. Conclusions: Inflammatory response patterns in plasma and sputum can be identified in hospitalized patients with CAP. Characterization of the local and systemic inflammatory response may help to better discriminate patients for enrollment into clinical trials of immunomodulatory therapies

Chromenopyrazole, a Versatile Cannabinoid Scaffold with in Vivo Activity in a Model of Multiple Sclerosis

A combination of molecular modeling and structure activity relationship studies has been used to fine-tune CB2 selectivity in the chromenopyrazole ring, a versatile CB1/CB2 cannabinoid scaffold. Thus, a series of 36 new derivatives covering a wide range of structural diversity has been synthesized, and docking studies have been performed for some of them. Biological evaluation of the new compounds includes, among others, cannabinoid binding assays, functional studies, and surface plasmon resonance measurements. The most promising compound [43 (PM226)], a selective and potent CB2 agonist isoxazole derivative, was tested in the acute phase of Theiler's murine encephalomyelitis virus-induced demyelinating disease (TMEV-IDD), a well established animal model of primary progressive multiple sclerosis. Compound 43 dampened neuroinflammation by reducing microglial activation in the TMEV

The Role of Early Pregnancy Maternal pGCD59 Levels in Predicting Neonatal Hypoglycemia-Subanalysis of the DALI Study

CONTEXT: Neonatal hypoglycaemia (NH) is the most common metabolic problem in infants born of mothers with gestational diabetes. Plasma glycated CD59 (pGCD59) is an emerging biomarker that has shown potential in identifying women at risk of developing gestational diabetes. The aim of this study was to assess the association between early maternal levels of pGCD59 and NH. OBJECTIVE: The aim of this study was to assess the association between early pregnancy maternal levels of plasma glycated CD59 (pGCD59) and neonatal hypoglycemia (NH). METHODS: This is an observational study of pregnant women with a prepregnancy body mass index (BMI) greater than or equal to 29 screened for eligibility to participate in the Vitamin D and Lifestyle Intervention for Gestational Diabetes (DALI) trial. This analysis included 399 pregnancies. Levels of pGCD59 were measured in fasting maternal samples taken at the time of a 75-g, 2-hour oral glucose tolerance test performed in early pregnancy (< 20 weeks). NH, the study outcome, was defined as a heel-prick capillary glucose level of less than 2.6 mmol/L within 48 hours of delivery. RESULTS: We identified 30 infants with NH. Maternal levels of pGCD59 in early pregnancy were positively associated with the prevalence of NH (one-way analysis of variance, P < .001). The odds of NH were higher in infants from mothers in tertile 3 of pGCD59 levels compared to those from mothers in tertile 1 (odds ratio [OR]: 2.41; 95% CI, 1.03-5.63). However, this was attenuated when adjusted for maternal BMI (OR: 2.28; 95% CI, 0.96-5.43). The cross-validated area under the curve (AUC) was 0.64 (95% CI, 0.54-0.74), and adjusted for maternal BMI, age, and ethnicity, the AUC was 0.70 (95% CI, 0.56-0.78). CONCLUSION: Although pGCD59 levels in early pregnancy in women with BMI greater than or equal to 29 are associated with NH, our results indicate that this biomarker by itself is only a fair predictor of NH