Transformation and tumorigenicity testing of simian cell lines and evaluation of poliovirus replication

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

Detection of pathogens from environmental samples for the evaluation of the health status of laboratory animals

Pathogens present in the environment are the biggest source of diseases and epidemics in the breeding of laboratory animals. The presence of microorganisms, in fact, can critically influence the animal health status and consequently the validity and reproducibility of experimental data. In accordance with the 3Rs principle, this study fits into the Refinement and Reduction concepts. The development of a health surveillance plan on environmental material from animal housing would have an important impact not only on maintaining an adequate state of health and on the generation of quality experimental data, but also on reducing the number of animals to be sacrificed.</jats:p

Health monitoring program for the control of laboratory animal diseases

Pathogens present in the environment are the biggest source of diseases and epidemics in the breeding of laboratory animals. In fact, the presence of microorganisms can critically influence the animal health status and, consequently, the validity and reproducibility of experimental data. In accordance with the 3Rs principle (Refinement, Reduction, Replacement), this study is part of the Refinement concept. The FELASA guidelines, formulated with the aim of guaranteeing the best animal health state, are a valid support for researchers. In this preliminary study, health-monitoring program was carried out within the breeding of laboratory animals in IZSLER facility. The main murine viruses were analyzed through molecular biology techniques (PCR, RT-PCR) and enzyme immunoassays (indirect ELISA). The established surveillance program steadily guarantees animal health and ensures the most controlled environmental and sanitary conditions. Further investigations will be needed to develop virus control strategies.</jats:p

Interplay between the intracellular energy sensor AMP-activated protein kinase (AMPK) and the estrogen receptor activities in regulating rat pituitary tumor cell (GH3) growth in vitro

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

Cell lines used in the study.

<p>Cell lines used in the study.</p

Susceptibility of cell lines to <i>Poliovirus</i>.

Attenuated Poliovirus Sabin LsC 2ab Sabin type 1 (Panel A), P712 Ch 2ab Sabin type 2 (B) and Leon 12alb Sabin type 3 (C) infectious titers obtained on the indicated cell lines. Data are reported as TCID50/ml log10 mean ± standard error of the mean. Asterisk indicates a P value < 0.05 significance between the two data sets marked by the line.</p

Cell line quality and tumorigenicity parameters.

<p>Cell line quality and tumorigenicity parameters.</p