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1.
New Microbiol ; 47(1): 38-46, 2024 May.
Artículo en Inglés | MEDLINE | ID: mdl-38700882

RESUMEN

The shortage of organs for human transplantation is a topic of extreme interest, and xenotransplantation with porcine organs has been recognized as a promising solution. However, the potential spillover linked to infectious agents present in pigs remains a concern. Among these, Pig Endogenous Retroviruses (PERVs), whose proviral DNAs are integrated in the genome of all pig breeds, represent an extremely important biological risk. This study aims to evaluate PERVs distribution in several swine cell lines and samples of domestic and feral pigs. Moreover, the capacity of PERVs to infect human and non-human primate cells and to integrate in the cellular genome was tested by Real-Time PCR and by Reverse Transcriptase assay. Results indicated a widespread diffusion of PERVs both in cell lines and samples analysed: the viral genome was found in all the established cell lines, in 40% of the primary cell lines and in 60% of the tissue samples tested. The assays indicated that the virus can be transmitted from porcine to human cells: in the specific case, infected NSK and NPTr cells allow passage to human 293 and MRC-5 cells with active production of the virus demonstrable via PCR and RT assay. In light of these aspects and also the lack of studies on PERVs, it appears clear that there are still many questions to be clarified, also by means of future studies, before xenotransplantation can be considered microbiologically safe.


Asunto(s)
Retrovirus Endógenos , Animales , Retrovirus Endógenos/genética , Retrovirus Endógenos/aislamiento & purificación , Porcinos , Humanos , Línea Celular , Infecciones por Retroviridae/veterinaria , Infecciones por Retroviridae/virología , Infecciones por Retroviridae/transmisión
2.
Front Physiol ; 13: 836480, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35936888

RESUMEN

Recently, research is undergoing a drastic change in the application of the animal model as a unique investigation strategy, considering an alternative approach for the development of science for the future. Although conventional monolayer cell cultures represent an established and widely used in vitro method, the lack of tissue architecture and the complexity of such a model fails to inform true biological processes in vivo. Recent advances in cell culture techniques have revolutionized in vitro culture tools for biomedical research by creating powerful three-dimensional (3D) models to recapitulate cell heterogeneity, structure and functions of primary tissues. These models also bridge the gap between traditional two-dimensional (2D) single-layer cultures and animal models. 3D culture systems allow researchers to recreate human organs and diseases in one dish and thus holds great promise for many applications such as regenerative medicine, drug discovery, precision medicine, and cancer research, and gene expression studies. Bioengineering has made an important contribution in the context of 3D systems using scaffolds that help mimic the microenvironments in which cells naturally reside, supporting the mechanical, physical and biochemical requirements for cellular growth and function. We therefore speak of models based on organoids, bioreactors, organ-on-a-chip up to bioprinting and each of these systems provides its own advantages and applications. All of these techniques prove to be excellent candidates for the development of alternative methods for animal testing, as well as revolutionizing cell culture technology. 3D systems will therefore be able to provide new ideas for the study of cellular interactions both in basic and more specialized research, in compliance with the 3R principle. In this review, we provide a comparison of 2D cell culture with 3D cell culture, provide details of some of the different 3D culture techniques currently available by discussing their strengths as well as their potential applications.

3.
Res Vet Sci ; 152: 134-149, 2022 Dec 20.
Artículo en Inglés | MEDLINE | ID: mdl-35969916

RESUMEN

Nanoscience and nanotechnologies have recently gained importance in several fields, such as industry and medicine. A big issue of the increasing application of nanomaterials is the poor literature regarding their potential toxicity in humans and animals. Recently, adult stem cells have been proposed as putative targets of nanoparticles (NPs). This study aims to investigate the effects of zerovalent-metallic NPs on isolated and amplified equine Adipose tissue derived Mesenchymal Stem Cells (eAdMSCs). Cells were treated with Cobalt (Co-), Iron (Fe-), and Nickel (Ni-) nanoparticles (NPs) at different concentrations and were characterized for the cytotoxic and genotoxic effects of exposure. Treatment with NPs resulted in reduced cell viability and proliferative capability in comparison with untreated cells. However, this did not influence eAdMSCs potency, as treated cells were able to differentiate towards the adipogenic and osteogenic lineages. Ni- and Fe-NPs showed cytoplasmic localization, while Co-NPs entered the nucleus and mitochondria, suggesting a potential genotoxic activity. Regarding p53 expression, it was enhanced in the first 48 h after treatments, with a drastic reduction of expression within 72 h. Higher p53 expression was reported in the case of Co-NP treatment, suggesting the tumorigenic potential of these NPs. Telomerase activity was enhanced by Fe- and Ni-NP treatments in a concentration- and time-dependent way. This was not true for Co-NP treated samples, suggesting a reduced replicative capacity of eAdMSCs upon Co-NP exposure. The present study is a preliminary investigation of the influence exerted by NPs on eAdMSC physiological activity in terms of cytotoxic and genotoxic effects. The present results revealed eAdMSC physiology to be strongly influenced by NPs in a dose-, time- and NP-dependent way.


Asunto(s)
Células Madre Mesenquimatosas , Nanopartículas del Metal , Nanopartículas , Humanos , Caballos , Animales , Proteína p53 Supresora de Tumor , Nanopartículas del Metal/toxicidad , Supervivencia Celular , Hierro
4.
PLoS One ; 12(1): e0169391, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-28046048

RESUMEN

The key role of cell cultures in different scientific fields is worldwide recognized, both as in vitro research models alternative to laboratory animals and substrates for biological production. However, many safety concerns rise from the use of animal/human cell lines that may be tumorigenic, leading to potential adverse contaminations in cell-derived biologicals. In order to evaluate the suitability of 13 different cell lines for Poliovirus vaccine production, safety and quality, in vitro/in vivo tumorigenicity and Poliovirus propagation properties were evaluated. Our results revealed that non-human primate cell lines CYNOM-K1, FRhK-4, 4MBr-5 and 4647 are free of tumorigenic features and represent highly susceptible substrates for attenuated Sabin Poliovirus strains. In particular, FRhK-4 and 4647 cell lines are characterized by a higher in vitro replication, resulting indicated for the use in large-scale production field.


Asunto(s)
Carcinogénesis/patología , Transformación Celular Neoplásica/patología , Poliovirus/fisiología , Replicación Viral , Animales , Bioensayo , Línea Celular , Primates
5.
Pituitary ; 17(3): 203-9, 2014 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-23649927

RESUMEN

Estrogen receptor α has a role in regulating rat somatolactotroph tumor cell growth (GH3 cells). AMP-activated protein kinase (AMPK) is a metabolic checkpoint which is able to negatively regulate intracellular signaling downstream of growth factors receptors in conditions increasing cellular AMP levels. We have recently reported on the role of AMPK activation in affecting viability and proliferation of GH3 cells. In the present study, we investigated the interplay between ER- and AMPK-pathways. Results can be regarded as relevant to the development of novel multi-targeted pharmacological therapies against pituitary tumors. We confirmed that estradiol (E2) and the ER antagonist fulvestrant exert stimulatory and inhibitory effects, respectively on GH3 cell growth in a competitive manner. The upstream kinase LKB1 is known to phosphorylate and activate AMPK. Here we showed that neither E2 nor fulvestrant caused a downregulation of LKB1 expression and phospho-AMPK levels in GH3 cells. Actually, fulvestrant strongly reduced the phosphorylation of ACC, which is a direct target of AMPK and a known index of AMPK activity. 2-deoxyglucose, a compound reducing glucose utilization, caused an increase in AMPK activity vs baseline and was able to hinder the stimulatory effect of E2 on cell viability, confirming that the exposure of GH3 cells to estrogens does not prevent them from being responsive to the inhibitory activity of compounds activating AMPK. Finally, the AMPK activator AICAR (AMP analog) did not cause further decrease in cell viability in the course of co-treatments with fulvestrant versus fulvestrant alone, in agreement with impaired phospho-AMPK activity in the presence of the anti-estrogen.


Asunto(s)
Proteínas Quinasas Activadas por AMP/fisiología , Neoplasias Hipofisarias/patología , Receptores de Estrógenos/fisiología , Quinasas de la Proteína-Quinasa Activada por el AMP , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Antimetabolitos/farmacología , Línea Celular Tumoral , Desoxiglucosa/farmacología , Estradiol/análogos & derivados , Estradiol/farmacología , Antagonistas del Receptor de Estrógeno/farmacología , Fulvestrant , Neoplasias Hipofisarias/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/fisiología , Ratas , Receptores de Estrógenos/metabolismo
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