Thyrotoxicosis Involves ß2-Adrenoceptor Signaling to Negatively Affect Microarchitecture and Biomechanical Properties of the Femur.

Background: Thyrotoxicosis increases bone turnover, resulting in net bone loss. Sympathetic nervous system (SNS) activation, via ß2-adrenoceptor (ß2-AR) signaling, also has osteopenic effects. Because thyroid hormones (TH) interact with the SNS to regulate several physiological processes, we hypothesized that this interaction also occurs to regulate bone mass. Previous studies support this hypothesis, as α2-AR knockout (KO) mice are less susceptible to thyrotoxicosis-induced osteopenia. Here, we evaluated whether TH-SNS interactions in bone involve ß2-AR signaling. Methods: Thyrotoxicosis was induced in 120-day-old female and male mice with ß2-AR gene inactivation (ß2-AR-/-) by daily treatment with supraphysiological doses of triiodothyronine (T3) for 12 weeks. The impact of thyrotoxicosis on femoral bone microarchitecture, remodeling, fracture risk, and gene expression of the receptor activator of nuclear factor-kappa-B (RANK)-RANK ligand (RANKL)-osteoprotegerin (OPG) pathway was evaluated. In addition, the effect of the ß2-AR-specific agonist clenbuterol (CL) on cAMP accumulation was determined in osteoblastic (MC3T3-E1) cells treated with T3 and/or 17ß-estradiol (E2). Results: Thyrotoxicosis negatively affected trabecular bone microarchitecture in wild-type (WT) females, but this effect was milder or nonexistent in ß2-AR-/- animals, whereas the opposite was seen in males. T3 treatment increased the femoral RANKL/OPG mRNA ratio and the endosteal perimeter and medullary area of the diaphysis in WT females and males, but not in ß2-AR-/- mice, suggesting that T3 promotes endosteal resorption in cortical bone, in a mechanism that involves ß2-AR signaling. T3 treatment increased endocortical mineral apposition rate only in WT females but not in ß2-AR-/- mice, suggesting that TH also induce bone formation in a ß2-AR signaling-dependent mechanism. T3 treatment decreased femoral resistance to fracture only in WT females, but not in KO mice. E2 and CL similarly increased cAMP accumulation in MC3T3-E1 cells; whereas T3 alone had no effect, but it completely blocked E2-stimulated cAMP accumulation, suggesting that some T3 effects on bone may involve E2/cAMP signaling in osteoblasts. Conclusions: These findings sustain the hypothesis that T3 interacts with the SNS to regulate bone morphophysiology in a ß2-AR signaling-dependent mechanism. The data also reveal sex as an important modifier of skeletal manifestations of thyrotoxicosis, as well as a modifier of the TH-SNS interactions to control bone microarchitecture, remodeling, and resistance to fracture.

Characterization of putative haematopoietic cells from bovine yolk sac.

The yolk sac is an extra-embryonic membrane that plays an important role in early embryonic survival. It is the production site for blood cells during embryonic mammalian development and is a likely source of stem cells. The aim of this study was to identify and characterize the putative haematopoietic cells from the yolk sac of bovine embryos at different stages of gestation. The yolk sac regresses according to gestational age and embryos are characterized into groups (I-V) according to the crown-rump measurement. Groups I-III survived in culture longer and exhibited the formation of cell clusters, whereas groups IV and V could not be maintained in culture for an extended period of time. Flow-cytometry analysis revealed that groups I-III had similar characteristics, including high expression levels of the haematopoietic markers CD34, CD90 and CD117. In groups IV and V, decreases were observed in the expression levels of CD117 and CD34. Cells were found to be capable of survival post-cryopreservation and exhibited varying abilities to form colonies in a methylcellulose matrix, depending on gestational age. Cytological analysis revealed the presence of blood cells (lymphocytes and monocytes). Quantitative PCR analysis demonstrated the presence of the haematopoietic progenitor genes GATA3 and LMO2, but not RUNX1. Thus, we have successfully isolated and characterized haematopoietic cells from the bovine embryo yolk sac at varying gestational ages. This study is crucial for the understanding of the development of the haematopoietic system and the embryonic function of this organ. Copyright © 2015 John Wiley & Sons, Ltd.

Regulation of neural stem cells by choroid plexus cells population.

The choroid plexus is a tissue on the central nervous system responsible for producing cerebrospinal fluid, maintaining homeostasis and neural stem cells support; though, all of its functions still unclear. This study aimed to demonstrate the niches of choroid plexus cells for a better understanding of the cell types and functions, using the porcine as the animal model. The collected material was analyzed by histology, immunohistochemistry, and cell culture. The cell culture was characterizated by immunocytochemistry and flow cytometry. Our results showed OCT-4, TUBIII, Nestin, CD45, CD73, CD90 positive expression and GFAP, CD105 negative expression, also methylene blue histological staining confirmed the presence of telocytes cells. We realized that the choroid plexus is a unique and incomparable tissue with different niches of cells as pluripotent, hematopoietic, neuronal progenitors and telocyte cells, which provide its complexity, differentiated functionality and responsibility on brain balance and neural stem cells regulation.

Alterações morfofuncionais renais em cães Golden Retriever Distróficos (GRMD) / Kidneys morphological changes in Golden Retriever Muscular Dystrophy dogs (GRMD)

A Golden Retriever Muscular Dystrophy (GRMD) é geneticamente homóloga à distrofia muscular de Duchenne (DMD) que acomete seres humanos. É uma doença genética que gera degeneração progressiva da musculatura esquelética. Considerando-se as intensas alterações musculares, é natural pensar em uma possível lesão renal decorrente da intensa lesão muscular. Foram avaliados seis cães machos da raça Golden Retriever afetados pela distrofia muscular (GRMD) e três cães machos clinicamente sadios. A concentração de creatinina foi determinada e as proteínas urinárias foram avaliadas por eletroforese em gel de poliacrilamida. Os resultados mostraram que a proteinúria patológica não está diretamente associada à Distrofia Muscular de Duchenne, porém diversos parâmetros apresentaram concentrações aumentadas para animais afetados, como a razão proteína/albumina, que foi maior em cães distróficos, podendo ser indício de microalbuminúria e conseqüente lesão renal precoce. Estes resultados visam embasar avaliações clínicas e futuros estudos considerando-se as patologias decorrentes ou associadas a esta doença genética.

The Golden Retriever Muscular Dystrophy (GRMD) is genetically homologous to Duchenne muscular dystrophy (DMD) that affects humans. It is a genetic disease that causes progressive degeneration of skeletal muscle. Considering the intense muscle changes, it is natural to think in possible kidney damage caused by intense muscle injury. We evaluated six male Golden Retriever dogs affected by Duchenne muscular dystrophy (GRMD) and three clinically healthy male dogs. The urinary proteins and creatinine concentration were determined. The proteins were analyzed by polyacrylamide gel electrophoresis. The results showed that pathological proteinuria is not directly associated with Duchenne muscular dystrophy, but several parameters showed increased concentrations for affected animals, as the ratio protein/albumin, which was higher in dystrophic dogs, probably a consequence of microalbuminuria a sign of early kidney damage. These results aim to base future studies and clinical evaluations considering the pathologies arising from or associated with this genetic disease.