Anacardium occidentale Bark as an Antidiabetic Agent.

Anacardium occidentale L. is used throughout the world to treat type 2 diabetes. In Portugal, a traditional herbal preparation made with stem bark of this species (AoBTHP) has been used for more than 30 years to treat this pathology. The AoBTHP was standardized on total phenolic content, and its hypoglycemic activity was assessed using db/db mice (n = 26) for 92 days. Three doses (40.2, 71.5, and 127.0 mg/kg/day, per os) were tested, and glibenclamide (5 mg/kg/day) was used as positive control. During the study, glycemia was measured under non-fasting or fasting states. In sequence, thin-layer chromatography bioautographic assays were used for the detection of possible alpha- and beta-glucosidase inhibitors. A significant hypoglycemic effect in fasting glycemia in days 31 and 57 was observed with the three tested doses. The 71.5 mg/kg and 127.0 mg/kg AoBTHPs significantly reduced non-fasting glycemia on day 24. The highest dose showed the most significant hypoglycemic effect. Gallic acid was identified as the major alpha- and beta-glucosidase inhibitor. The 127 mg/kg/day AoBTHP dose showed a greater glucose-lowering effect than glibenclamide. For the first time, a standardized AoBTHP was tested using an in vivo diabetes model, and its usage was preclinically validated for type 2 diabetes treatment. The hypoglycemic activity of an AoBTHP can be related to the presence of alpha- and beta-glucosidase inhibitors, such as gallic acid, but other mechanisms can also be involved.

Canine multicentric lymphoma exhibits systemic and intratumoral cytokine dysregulation.

Non-Hodgkin Lymphoma (NHL) is among the most common neoplasias in dogs and humans. Owing to remarkable similarities with its human counterpart, the canine lymphoma (cNHL) model has been proposed as a powerful framework for rapid and clinically relevant translation of novel immunotherapies. However, the establishment of cNHL as a predictive preclinical model has been hampered by the limited characterization of the canine immune system. Cytokines are key players of the interaction between tumor and its microenvironment. In human NHL, multiple cytokines have been linked to the development of lymphoma and are relevant biomarkers for treatment response and prognosis. In contrast, few studies have investigated cytokines in cNHL. Within this context, this study aimed to investigate cytokine regulation in cNHL. A multicentric cNHL biobank was successfully constructed. Cytokine mRNA profiles in tumor tissue and circulating PBMC were analyzed by qRT-PCR and compared to a healthy control group. Specific primers were used to evaluate Th1, Th2 and Th17 responses. Systemic cytokine concentrations were measured using a commercial canine multiplex assay which included IL-2, IL6, IL-10 and TNF-α, and compared to a healthy control group. Our results demonstrated a dysregulation of cytokine mRNA expression, representative of the tumor microenvironment and systemic response in cNHL. Intratumoral cytokine response revealed a significant downregulation of humoral and Th1 responses. The systemic response demonstrated a distinct mRNA pattern, however immunosuppression also prevailed. Cytokine serum quantification showed a significant increase of IL-10 concentration in cNHL. Significant differences in hematological parameters were described and a correlation between IL-6 protein serum levels and neutrophil count was shown. Finally, data analysis demonstrated that baseline pretreatment IFN-γ tissue mRNA levels were correlated to survival outcome, predicting a favorable response to chemotherapy. Altogether, these results revealed that cNHL presents a local and systemic dysregulation in cytokine response. By confirming and extending previous research, our work contributed for the evaluation of potential cytokine candidates for diagnostic, prognostic purposes and therapeutic intervention, therefore adding value to comparative oncology.

The histone deacetylase inhibitor panobinostat is a potent antitumor agent in canine diffuse large B-cell lymphoma.

Non-Hodgkin lymphoma (NHL) is one of the most common causes of cancer-related death in the United States and Europe. Although the outcome of NHL patients has improved over the last years with current therapies, the rate of mortality is still high. A plethora of new drugs is entering clinical development for NHL treatment; however, the approval of new treatments remains low due in part to the paucity of clinically relevant models for validation. Canine lymphoma shares remarkable similarities with its human counterpart, making the dog an excellent animal model to explore novel therapeutic molecules and approaches. Histone deacetylase inhibitors (HDACis) have emerged as a powerful new class of anti-cancer drugs for human therapy. To investigate HDACi antitumor properties on canine diffuse large B-cell lymphoma, a panel of seven HDACi compounds (CI-994, panobinostat, SBHA, SAHA, scriptaid, trichostatin A and tubacin) was screened on CLBL-1 canine B-cell lymphoma cell line. Our results demonstrated that all HDACis tested exhibited dose-dependent inhibitory effects on proliferation of CLBL-1 cells, while promoting increased H3 histone acetylation. Amongst all HDACis studied, panobinostat proved to be the most promising compound and was selected for further in vitro and in vivo evaluation. Panobinostat cytotoxicity was linked to H3 histone and α-tubulin acetylation, and to apoptosis induction. Importantly, panobinostat efficiently inhibited CLBL-1 xenograft tumor growth, and strongly induced acetylation of H3 histone and apoptosis in vivo. In conclusion, these results provide new data validating HDACis and, especially, panobinostat as a novel anti-cancer therapy for veterinary applications, while contributing to comparative oncology.

Acellular Urethra Bioscaffold: Decellularization of Whole Urethras for Tissue Engineering Applications.

Patients with stress urinary incontinence mainly suffer from malfunction of the urethra closure mechanism. We established the decellularization of porcine urethras to produce acellular urethra bioscaffolds for future tissue engineering applications, using bioscaffolds or bioscaffold-derived soluble products. Cellular removal was evaluated by H&E, DAPI and DNA quantification. The presence of specific ECM proteins was assessed through immunofluorescence staining and colorimetric assay kits. Human skeletal muscle myoblasts, muscle progenitor cells and adipose-derived stromal vascular fractions were used to evaluate the recellularization of the acellular urethra bioscaffolds. The mechanochemical decellularization system removed ~93% of tissue's DNA, generally preserving ECM's components and microarchitecture. Recellularization was achieved, though methodological advances are required regarding cell seeding strategies and functional assessment. Through microdissection and partial digestion, different urethra ECM-derived coating substrates were formulated (i.e. containing smooth or skeletal muscle ECM) and used to culture MPCs in vitro. The skeletal muscle ECM substrates enhanced fiber formation leading to the expression of the main skeletal muscle-related proteins and genes, as confirmed by immunofluorescence and RT-qPCR. The described methodology produced a urethra bioscaffold that retained vital ECM proteins and was liable to cell repopulation, a crucial first step towards the generation of urethra bioscaffold-based Tissue Engineering products.