Beatriz Hernández-Téllez

GABA (A) receptor subunits RNA expression in mice peritoneal macrophages modulate their IL-6/IL-12 production

The expression and functionality of the gamma aminobutyric acid A receptor (GABA(A)R) in mice macrophages was explored. Reverse Transcriptase-Polymerase Chain Reaction showed that macrophages express the GABA(A)R RNA for the alpha1, alpha2, beta3 and delta subunits, but not for the alpha5, beta1, beta2 and gamma3 subunits. The expression of alpha1 subunit was also revealed by confocal microscopy. LPS-stimulated macrophages significantly reduced their in vitro production of IL-6 and IL-12 after GABA adding to the culture medium. These results showed that BALB/c mice peritoneal macrophages express a functional subset of GABA(A)R subunits.

Inhaled vanadium pentoxide decrease gamma-tubulin of mouse testes at different exposure times

Vanadium is an important environmental and industrial pollutant whose concentrations have increased in the last decades. Due to its status as reproductive toxicant and a microtubule damaging agent, the present study investigated by immunohistochemistry the effect of the inhalation of vanadium pentoxide on gamma-tubulin within somatic and testicular germ cells. Male mice inhaled vanadium pentoxide (V2O5) (0.02 M) 1 h/twice a week for 12 weeks. Our results demonstrated that vanadium accumulates in the testes starting with the initial inhalation (24 h), and this pattern remained until the last week of treatment. In general, vanadium was capable of significantly decreasing the percentage of gamma-tubulin in all analyzed testicular cells (Sertoli, Leydig and germ cells) starting with the first week of treatment. For all cell types studied, regression analysis revealed a negative and significant relationship between the percentage of immunopositive cells to gamma-tubulin and exposure time, showing a time dependent response in all cases. Our findings suggest that alterations on this protein might imply changes in microtubule-involved function such as cell division, which in the testes might lead to damage in the spermatogenesis, leading probably to infertility.

GK-1 Improves the Immune Response Induced by Bone Marrow Dendritic Cells Loaded with MAGE-AX in Mice with Melanoma

The aim of dendritic cell (DC) vaccination in cancer is to induce tumor-specific effector T cells that may reduce and control tumor mass. Immunostimulants that could drive a desired immune response are necessary to be found in order to generate a long lasting tumor immune response. GK-1 peptide, derived from Taenia crassiceps, induces not only increase in TNFα, IFNγ, and MCP-1 production in cocultures of DCs and T lymphocytes but also immunological protection against influenza virus. Moreover, the aim of this investigation is the use of GK-1 as a bone marrow DCs (BMDCs) immunostimulant targeted with MAGE antigen; thus, BMDC may be used as immunotherapy against murine melanoma. GK-1 induced in BMDCs a meaningful increment of CD86 and IL-12. In addition, the use of BMDCs TNFα/GK-1/MAGE-AX induced the highest survival and the smallest tumors in mice. Besides, the treatment helped to increase CD8 lymphocytes levels and to produce IFNγ in lymph nodes. Moreover, the histopathological analysis showed that BMDCs treated with GK-1/TNFα and loaded with MAGE-AX induced the apparition of more apoptotic and necrotic areas in tumors than in mice without treatment. These results highlight the properties of GK-1 as an immunostimulant of DCs and suggest as a potential candidate the use of this immunotherapy against cancer disease.

Neurons and satellite glial cells in adult rat lumbar dorsal root ganglia express connexin 36

Previous studies have shown that following peripheral nerve injury there was a downregulation of the gap junction protein connexin 36 (Cx36) in the spinal cord; however, it is not known whether Cx36 protein is expressed in the dorsal root ganglia (DRGs), nor if its levels are altered following peripheral nerve injuries. Here we address these aspects in the adult rat lumbar DRG. Cx36 mRNA was detected using qRT-PCR, and Cx36 protein was identified in DRG sections using immunohistochemistry (IHC) and immunofluorescence (IF). Double staining revealed that Cx36 co-localizes with both anti-β-III tubulin, a neuronal marker, and anti-glutamine synthetase, a satellite glial cell (SGC) marker. In neurons, Cx36 staining was mostly uniform in somata and fibers of all sizes and its intensity increased at the cell membranes. This labeling pattern was in contrast with Cx36 IF dots mainly found at junctional membranes in islet beta cells used as a control tissue. Co-staining with anti-Cx43 and anti-Cx36 showed that whereas mostly uniform staining of Cx36 was found throughout neurons and SGCs, Cx43 IF puncta were localized to SGCs. Cx36 mRNA was expressed in normal lumbar DRG, and it was significantly down-regulated in L4 DRG of rats that underwent sciatic nerve injury resulting in persistent hypersensitivity. Collectively, these findings demonstrated that neurons and SGCs express Cx36 protein in normal DRG, and suggested that perturbation of Cx36 levels may contribute to chronic neuropathic pain resulting from a peripheral nerve injury.

Structural Effect of Different EDC Crosslinker Concentration in Gelatin- Hyaluronic Acid Scaffolds

Introduction: Gelatin and hyaluronic acid are two biopolymers with different applications in tissue engineering. They may be employed to construct diverse scaffolds that allow cells to differentiate and proliferate on them. In order to obtain the best functional and mechanical conditions in scaffolds, they must be crosslinked to form covalent links between gelatin and hyaluronic acid. The crosslinker 1-ethy-3-(3-dymethylaminopropyl) carbodiimide hydrochloride (EDC) is a compound widely used due to its low cytotoxicity. Besides, the concentration of the crosslinker may modify the physical properties and morphological characteristics of scaffolds when it forms covalent links between biopolymers, helping to construct different kinds of scaffolds used for developing soft tissues. However, the development of scaffolds made of gelatin and hyaluronic acid crosslinked with EDC has been poorly studied. In addition, the concentrations used for crosslinking gelatin and hyaluronic acid are contradictory. Therefore, the aim of this study was to analyze the structure of gelatin/hyaluronic acid scaffolds crosslinked with EDC. Methods: Gelatin-hyaluronic acid scaffolds were prepared by direct freeze-drying. Afterwards, They were crosslinked with different concentrations of EDC (6, 30, 50 and 60 mM) for 12 h. Results: This research has demonstrated that the gelatin/hyaluronic acid scaffolds crosslinked with the highest concentrations of the crosslinker had fewer water concentration absorbed, pore size diminished and pore number increased in comparison with control groups. Despite scaffolds composition has not changed in any of the concentrations, the bone marrow mesenchymal cells mortality percentage increased when cells were placed on the scaffolds of concentration 60 mM, perhaps for the residual 1-ethy-3-(3-dymethylaminopropyl) carbodiimide hydrochloride found in the scaffolds. Conclusion: Our results revealed that different EDC concentrations may modify the physical and biological characteristics of gelatin/hyaluronic acid scaffolds; as a result, the scaffolds obtained may be used for the manufacture of different tissues in regenerative medicine.

Quantitative multiplexed proteomics of Taenia solium cysts obtained from the skeletal muscle and central nervous system of pigs

In human and porcine cysticercosis caused by the tapeworm Taenia solium, the larval stage (cysts) can infest several tissues including the central nervous system (CNS) and the skeletal muscles (SM). The cyst’s proteomics changes associated with the tissue localization in the host tissues have been poorly studied. Quantitative multiplexed proteomics has the power to evaluate global proteome changes in response to different conditions. Here, using a TMT-multiplexed strategy we identified and quantified over 4,200 proteins in cysts obtained from the SM and CNS of pigs, of which 891 were host proteins. To our knowledge, this is the most extensive intermixing of host and parasite proteins reported for tapeworm infections.Several antigens in cysticercosis, i.e., GP50, paramyosin and a calcium-binding protein were enriched in skeletal muscle cysts. Our results suggested the occurrence of tissue-enriched antigen that could be useful in the improvement of the immunodiagnosis for cysticercosis. Using several algorithms for epitope detection, we selected 42 highly antigenic proteins enriched for each tissue localization of the cysts. Taking into account the fold changes and the antigen/epitope contents, we selected 10 proteins and produced synthetic peptides from the best epitopes. Nine peptides were recognized by serum antibodies of cysticercotic pigs, suggesting that those peptides are antigens. Mixtures of peptides derived from SM and CNS cysts yielded better results than mixtures of peptides derived from a single tissue location, however the identification of the ‘optimal’ tissue-enriched antigens remains to be discovered. Through machine learning technologies, we determined that a reliable immunodiagnostic test for porcine cysticercosis required at least five different antigenic determinants.

Corrigendum to “GK-1 Improves the Immune Response Induced by Bone Marrow Dendritic Cells Loaded with MAGE-AX in Mice with Melanoma”

In the paper titled “GK-1 Improves the Immune Response Induced by Bone Marrow Dendritic Cells Loaded with MAGE-AX in Mice with Melanoma” [1] the phrase “Recently, it has been reported that subcutaneous administration of GK-1 in mice with melanoma induces regression of the tumor mass and increases survival” should be as follows: “Recently, our group has shown that subcutaneous administration of GK-1 and DCs in mice with melanoma induces regression of the tumor mass and increases survival (unpublished data).”

Connexin 30.2 is expressed in exocrine vascular endothelial and ductal epithelial cells throughout pancreatic postnatal development

Previously we have demonstrated that the GJ protein connexin 30.2 (Cx30.2) is expressed in pancreatic beta cells and endothelial cells (ECs) of the islet. In the present study, we address whether Cx30.2 is expressed in the exocrine pancreas, including its vascular system. For this, adult mouse pancreatic sections were double labeled with specific antibodies against Cx30.2 and CD31, an endothelial cell marker, or with anti-α-actin smooth muscle, a smooth muscle cell (SMC) marker or anti-mucin-1, a marker of epithelial ductal cells, using immunofluorescence (IF) studies. Cx30.2-IF hot spots were found at junctional membranes of exocrine ECs and SMCs of blood vessels. Furthermore, Cx30.2 was localized in mucin-1 positive cells or epithelial ductal cells. Using immunohistochemistry (IHC) studies, it was found that in vessels and ducts of different diameters, Cx30.2 was also expressed in these cell types. In addition, it was found that Cx30.2 is already expressed in these cell types in pancreatic sections of 3, 14 and 21 days postpartum. Moreover, this cell specific pattern of expression was also found in the adult rat, hamster and guinea pig pancreas. Expression of Cx30.2 mRNA and protein in the pancreas of all these species was confirmed by RT-PCR and Western blot studies. Overall, our results suggest that intercellular coupling mediated by Cx30.2 intercellular channels may synchronize the functional activity of ECs and SMCs of vascular cells, as well as of epithelial ductal cells after birth.