Results Regarding Self Renewal and Differentiation of Cells Composing Inner Ear Derived Spheres

The purpose of the ongoing research is to improve our current skills and knowledge in stem cell isolation, cultivation and differentiation from the vestibular epithelia of young mice. We harvested utricles and sacculi from 7 days old mice. Obtained cells were cultivated at 37ºC and 5% CO2 in DMEM with F12 Nutrient mixture, B27, N2 supplement, IGF-1 and EGF. Sphere pluripotency was established with Nanog and Oct-4 stem cell markers. We mechanically dissociated primary spheres and cultivated them. Cells were characterized by immunofluorescence and immunohistochemistry for myosinVIIA (hair cell marker), nestin (intermediate filament VI marker) and beta III tubulin. We proved that vestibular epithelia contains pluripotent stem cells which formed spheres. Sphere-derived cells’ pluripotency was demonstrated by the expression of nanog, oct 4 and nestin markers. Also, after sphere dissociation we obtained a higher number of spheres beeing pluripotent and capable of self-renewal. We obtained through differentiation different cell types including neuron like-cells which were positive for myosin VIIA, nestin and beta III tubulin. Utricular epithelia of seven days old mice contains sufficient pluripotent stem cells which generate spheres. Cells obtained from utricular epithelia are pluripotent because they express nanog, oct 4, nesThe purpose of the ongoing research is to improve our current skills and knowledge in stem cell isolation, cultivation and differentiation from the vestibular epithelia of young mice. We harvested utricles and sacculi from 7 days old mice. Obtained cells were cultivated at 37ºC and 5% CO2 in DMEM with F12 Nutrient mixture, B27, N2 supplement, IGF-1 and EGF. Sphere pluripotency was established with Nanog and Oct-4 stem cell markers. We mechanically dissociated primary spheres and cultivated them. Cells were characterized by immunofluorescence and immunohistochemistry for myosinVIIA (hair cell marker), nestin (intermediate filament VI marker) and beta III tubulin. We proved that vestibular epithelia contains pluripotent stem cells which formed spheres. Sphere-derived cells’ pluripotency was demonstrated by the expression of nanog, oct 4 and nestin markers. Also, after sphere dissociation we obtained a higher number of spheres beeing pluripotent and capable of self-renewal. We obtained through differentiation different cell types including neuron like-cells which were positive for myosin VIIA, nestin and beta III tubulin. Utricular epithelia of seven days old mice contains sufficient pluripotent stem cells which generate spheres. Cells obtained from utricular epithelia are pluripotent because they express nanog, oct 4, nestin, characteristic for cell progenitors

Unique Growth Pattern Presentation of a Papillary Renal Cell Carcinoma

Papillary renal cell carcinoma (PRCC) is defined by the WHO 2022 classification as a malignant tumor derived from the renal tubular epithelium. However, the WHO 2016 classification subdivided PRCC into two types, with type 1 PRCC showing papillae covered by a single layer of neoplastic cells, and type II PRCC, which can show multiple types of histologies and is more aggressive. The WHO 2022 classification eliminated the subcategorization of PRCC. Here, we present a histopathological case study with a 4-year follow-up diagnosed in 2018 as type I PRCC (WHO 2016) with intra-pyelocalyceal growth pattern in a 59-year-old male patient with a history of Type II diabetes mellitus, left-sided renal–ureteral lithiasis, and benign hypertrophy of the prostate. Microscopically the tumor was composed of small cuboidal cells with inconspicuous nucleoli, arranged on a single layer of tubulo-papillary cores, and scant, foamy macrophages. The tumor had a non-infiltrative, expansive pyelocalyceal growth pattern. Immunohistochemically (IHC), the tumor cells were CK7-intense and diffusely positive, and stained granular for AMACR. Next-generation sequencing (NGS) was performed for the tumor and the normal adjacent tissue for in-depth pathological characterization. To our knowledge, this is the first reported case where a PRCC displays this unique intra-pyelocalyceal growth pattern, mimicking a urothelial cell carcinoma of the renal pelvis system

TRIAL TO CULTIVATE AND ISOLATE NEURONAL LIKE CELLS FROM 7 DAYS OLD MICE BRAIN

role to replace lost cells due to physiological turnover, injury, or disease and tosupport cell genesis contributing to the cell number homeostasis. Long time it wasthought that adult mammalian central nervous system doesn't possess any or fewregenerative capacity. Nowadays it was demonstrated that also in the brain thereare stem cells which have the capacity to differentiate into astocytes,oligodendrocytes and neurons.In few degenerative diseases the stem cells lose the regenerative capacity withconsequences in diminishing and loss of functional capacity. Stem cell therapyrepresents a novel and promising therapeutic approach to treatment of a variety ofdegenerative disease as multiple sclerosis. For this it is necessary that a efficientstem cell source can be found and secondary to be proven that these transplantablecells have differential potential into neural tissue.In order to be able to possess a stem cell source capable to build an implant it isnecessary to know the cultivation technology and also the instruments to prove theircapacity to differentiate into specific cells of the nervous system. These were themotives that enabled us to to try to harvest, cultivate and differentiate stem cellsfrom the murine central nervous system

Control Strategies for Prevention of Undesirable Traits in Cattle - Review

Cattle genome sequencing has a major impact on livestock breeding and provides useful information in the field biotechnology that can ultimately lead to the development of several techniques for early molecular diagnosis. These developments within molecular genetics have made possible the rapid and effective detection of lethal or mutant alleles associated with a disease-specific phenotype by DNA isolation followed by in vitro amplification techniques and enzymatic restriction. DNA tests are currently in use for the detection of heterozygotes and diagnosis of genetic diseases such as bovine leukocyte adhesion deficiency (BLAD), deficiency of uridine monophosphate synthase (DUMPS) and bovine citrullinemia. These genetic defects are autosomal recessive disorders which were widespread trough advanced reproductive technologies, including artificial insemination and multiple ovulation embryo transfer, as result of selection focused on desirable traits carried by elite bulls with high genetic merit. In order to prevent the risk of dissemination of inherited defects in dairy herds and to decrease the economic losses all bulls have to be tested, although the prevalence of the above mentioned recessive alleles is low. Therefore, it is advisable to use molecular markers to assess bovine health and to implement such programs for the purpose of monitoring hereditary diseases in cattle

PLURIPOTENT STEM CELLS FROM THE ADULT MOUSE UTRICLE

Researchers discovered that cochlear epithelia in mice especially the vestibular one, contains stem cells that have the capacity to differentiate in sensorial auditory hair cell progenitors specific to the organ. They are reduced in number as the animal progresses in age. This process leads to a loss in the regenerative and proliferative potential of sensorial inner ear epithelia secondary to different injuries. Isolation, cultivation and than in vitro differentiation of vestibular stem cells could become a regenerative implant for acquired hearing loss. These were the motives that determined us to try to isolate, cultivate and finally differentiate vestibular stem cells from vestibular epithelia.Utricles from 7 days old mice NMRI were harvested, the otolites were removed, the utricles were trypsinized in order to isolate cells. Obtained cells were cultivated at 37ºC and 5% CO2 in DMEM with F12 Nutrient mixture, B27, N2 supplement. Pluripotency of obtained spheres was established with the help of stem cell markers Nanog and Oct-4. For identification of progenitor cells we used the marker, which reveals the gene which encodes the protein nestin. In all experiments we obtained floating colonies called spheres, formed by mitotic multiplying. For testing the pluripotency of spheres we used Nanog and Oct-4, two transcription factors that are expressed at high levels in stem cells and that we found to be expressed in our spheres. The presence of nestin mRNA in cells composing the spheres showed that these progressed to a progenitor cell stage.We concluded that utricular epithelia in 7 days old mice contains sufficient stem cells that can be cultivated and that can be later differentiated

Innovative Biotechnology for Generation of Cardiac Tissue

Heart transplantation remains the only curative treatment for end-stage heart failure. This life-saving option continues to be limited by the low number of organ donors, graft rejection and adverse effects of immunosuppressants. Engineering bioartificial hearts from acellular native-derived scaffolds and stem cells has gained attention because of its potential to overcome these limitations. In this study, rat hearts (n = 20) were decellularized by means of coronary perfusion with 1% sodium dodecyl sulfate (SDS) in a modified Langendorff device. The electrical field behavior of the SDS molecule was studied and it was assumed that when applying an alternating current, the exposure time of the tissue to the detergent might decrease. To repopulate the decellularized extracellular matrix (ECM), human mesenchymal stem cells (hMSCs) were used, induced to differentiate into cardiomyocytes (CMs) with 5-azacytidine (5-aza). The results showed no cellular debris and an intact ECM following decellularization. Decellularization in the presence of an electric field proved to be faster, decreasing the potential risk of ECM damage due to the detergent. After cell seeding and culturing of eight scaffolds with hMSCs, the recellularization process was analyzed using optic microscopy (OM), which showed cells suggestive for CMs. This study presents a novel and efficient decellularization protocol using an electric field and suggests that hMSCs can be useful in the generation of a bioartificial heart

3D Bioprinting of Model Tissues That Mimic the Tumor Microenvironment

The tumor microenvironment (TME) influences cancer progression. Therefore, engineered TME models are being developed for fundamental research and anti-cancer drug screening. This paper reports the biofabrication of 3D-printed avascular structures that recapitulate several features of the TME. The tumor is represented by a hydrogel droplet uniformly loaded with breast cancer cells (106 cells/mL); it is embedded in the same type of hydrogel containing primary cells—tumor-associated fibroblasts isolated from the peritumoral environment and peripheral blood mononuclear cells. Hoechst staining of cryosectioned tissue constructs demonstrated that cells remodeled the hydrogel and remained viable for weeks. Histological sections revealed heterotypic aggregates of malignant and peritumoral cells; moreover, the constituent cells proliferated in vitro. To investigate the interactions responsible for the experimentally observed cellular rearrangements, we built lattice models of the bioprinted constructs and simulated their evolution using Metropolis Monte Carlo methods. Although unable to replicate the complexity of the TME, the approach presented here enables the self-assembly and co-culture of several cell types of the TME. Further studies will evaluate whether the bioprinted constructs can evolve in vivo in animal models. If they become connected to the host vasculature, they may turn into a fully organized TME