Discriminating aspects of global metabolism of neonatal cardiomyocytes from wild type and KO-CSRP3 rats using proton magnetic resonance spectroscopy of culture media samples.

Knockout of multifunction gene cysteine- and glycine-rich protein 3 (CSRP3) in cardiomyocytes (CMs) of mice leads to heart dilation, severely affecting its functions. In humans, CSRP3 mutations are associated with hypertrophic (HCM) and dilated cardiomyopathy (DCM). The absence of the CSRP3 expression produces unknown effects on in vitro neonatal CMs' metabolism. The metabolome changes in culture media conditioned by CSRP3 knockout (KO-CSRP3), and wild type (WT) neonatal cardiomyocytes were investigated under untreated or after metabolic challenging conditions produced by isoproterenol (ISO) stimulation, by in vitro high-resolution proton magnetic resonance spectroscopy (1H-MRS)-based metabolomics. Metabolic differences between neonatal KO-CSRP3 and WT rats' CMs were identified. After 72 h of culture, ISO administration was associated with increased CMs' energy requirements and increased levels of threonine, alanine, and 3-hydroxybutyrate in both neonatal KO-CSRP3 and WT CMs conditioned media. When compared with KO-CSRP3, culture media derived from WT cells presented higher lactate concentrations either under basal or ISO-stimulated conditions. The higher activity of ketogenic biochemical pathways met the elevated energy requirements of the contractile cells. Both cells are considered phenotypically indistinguishable in the neonatal period of animal lives, but the observed metabolic stress responses of KO-CSRP3 and WT CMs to ISO were different. KO-CSRP3 CMs produced less lactate than WT CMs in both basal and stimulated conditions. Mainly, ISO-stimulated conditions produced evidence for lactate overload within KO-CSRP3 CMs, while WT CMs succeeded to manage the metabolic stress. Thus, 1H-MRS-based metabolomics was suitable to identify early inefficient energetic metabolism in neonatal KO-CSRP3 CMs. These results may reflect an apparent lower lactate transport and consumption, in association with protein catabolism.

Chick Embryo Model for Homing and Host Interactions of Tissue Engineering-Purposed Human Dental Stem Cells.

Human dental stem cells (hDSC) have a potential for regenerative therapies and could differentiate in vitro into many tissues, such as dentin, nerve, and vascular endothelium. Gallus gallus domesticus developing fertilized egg or chick embryo is an experimental model absent of xenografts rejection, largely employed in replacement of mammal species in scientific research and preclinical studies to evaluate angiogenesis and vasculogenesis, tissue differentiation, and embryonic development. This multiscale research deals with the homing and cell signaling effects of a standardized hDSC toward the receptor tissues of G. gallus domesticus in ovo. The hDSC were obtained from the explantation from third molars, characterized by cell cytometry, and employed without any further purification procedure. Four experimental groups were studied, according to the kind of cell tracing strategy, named: Control, mCherry-labeled hDSC, QTracker-labeled hDSC, and QTracker-exposed controls. The eggs were kept in an incubator temperature of 37.6°C and humidity 86%, and the embryos were euthanized after 10 days of incubation. In vivo fluorescence and histological analysis were conducted. The fluorescence of the embryos inoculated with mCherry hDSC or the QTracker hDSC was associated with the bones and the beak tooth, and labeled cell islands could be localized in part of the samples. The inoculation of the QTracker probe resulted in proliferating bone tissue labeling. The hDSC inoculated groups presented cartilage plate hypertrophy and atypical morphology, meanwhile Control eggs were negative. The results demonstrated that hDSC can migrate to the cartilaginous tissues of the chick embryos, survive in this environment, implant, and interfere with the growth of developing bone.

Osteoinductive effects of preoperative dexamethasone in human dental pulp stem cells primary culture.

AIM: The use of dexamethasone (DEX) in mesenchymal cell culture induces osteoblastic differentiation and, consequently, formation of mineralized tissues. Tissue engineering proposes the development of therapeutic strategies aimed at structural and functional regeneration of biological tissues. In this sense, cell characterization in vitro is critical to ensure the development of such techniques. Our objective was to evaluate the osteoinductive effect of DEX administered as a preoperative medication in primary cell culture of human dental pulp stem cell. METHODOLOGY: Cells from the third molar pulp were divided into two experimental groups, each with two preoperative medication protocols used in dental practice and differentiated by the intake of DEX in one of them. The assessment of proliferation, differentiation and viability through trypan blue, methylthiazol tetrazolium, and von Kossa and alizarin red assays, respectively, were held within fixed intervals: 7, 14, 21 and 28 days. CONCLUSION: This study has shown that DEX may influence in vitro human dental pulp stem cell behavior.