SARS-CoV-2 infection and replication kinetics in different human cell types: The role of autophagy, cellular metabolism and ACE2 expression.

AIMS: This study evaluated SARS-CoV-2 replication in human cell lines derived from various tissues and investigated molecular mechanisms related to viral infection susceptibility and replication. MAIN METHODS: SARS-CoV-2 replication in BEAS-2B and A549 (respiratory tract), HEK-293 T (kidney), HuH7 (liver), SH-SY5Y (brain), MCF7 (breast), Huvec (endothelial) and Caco-2 (intestine) was evaluated by RT-qPCR. Concomitantly, expression levels of ACE2 (Angiotensin Converting Enzyme) and TMPRSS2 were assessed through RT-qPCR and western blot. Proteins related to autophagy and mitochondrial metabolism were monitored in uninfected cells to characterize the cellular metabolism of each cell line. The effect of ACE2 overexpression on viral replication in pulmonary cells was also investigated. KEY FINDINGS: Our data show that HuH7, Caco-2 and MCF7 presented a higher viral load compared to the other cell lines. The increased susceptibility to SARS-CoV-2 infection seems to be associated not only with the differential levels of proteins intrinsically related to energetic metabolism, such as ATP synthase, citrate synthase, COX and NDUFS2 but also with the considerably higher TMPRSS2 mRNA expression. The two least susceptible cell types, BEAS-2B and A549, showed drastically increased SARS-CoV-2 replication capacity when ACE2 was overexpressed. These modified cell lines are relevant for studying SARS-CoV-2 replication in vitro. SIGNIFICANCE: Our data not only reinforce that TMPRSS2 expression and cellular energy metabolism are important molecular mechanisms for SARS-CoV-2 infection and replication, but also indicate that HuH7, MCF7 and Caco-2 are suitable models for mechanistic studies of COVID-19. Moreover, pulmonary cells overexpressing ACE2 can be used to understand mechanisms associated with SARS-CoV-2 replication.

The impact of the genetic background in a patient with papillary thyroid cancer and familial adenomatous polyposis.

Thyroid cancer is the most common endocrine malignancy, and papillary thyroid carcinoma (PTC) is the main subtype. The cribriform morular variant is a histological phenotype of PTC characterized by its relationship with familial adenomatous polyposis (FAP). Description of the case: We report the genetic assessment of a 20-year-old female patient diagnosed with a cribriform-morular variant of PTC and FAP. We aimed to assess the genetic background of the reported patient, looking for variants that would help us explain the predisposition to tumorigenesis. Genomic DNA was extracted from peripheral blood lymphocytes, and whole exome sequencing was performed. We applied an overrepresentation and gene-set enrichment analysis to look for an accumulation of effects of variants in multiple genes at the genome. We found an overrepresentation of single nucleotide variants (SNVs) in extracellular matrix interactions and cell adhesion genes. Underrepresentation of SNVs in genes related to the regulation of autophagy and cell cycle control was also observed. We hypothesize that the package of alterations of our patient may help to explain why she presented colonic manifestations and thyroid cancer. Our findings suggest that multiple variants with minor impact, when considered together, may be helpful to characterize one particular clinical condition.

Evaluation of neuroglobin and cytoglobin expression in adult rats exposed to silver nanoparticles during prepubescence.

Silver nanoparticles (AgNPs) are clusters of silver atoms with diameters that range from 1 to 100 nm. Due to the various shapes and large surface areas, AgNPs have been employed in the food and textile industries and medical fields. Therefore, because of the widespread use of these compounds, the aim of this study was to evaluate the effect of AgNP exposure on the gene and protein expression levels of Neuroglobin (Ngb) and Cytoglobin (Cygb), in the rat cortex, hippocampus and cerebellum. Post-natal day (PND) 21 male Wistar rats were randomly divided into three groups. One group received 15 µg/kg body weight of AgNP by gavage another group received 30 µg/kg and the control group that received saline, from PND23 to PND58. On PND102 the animals were euthanized and the cortex, hippocampus and cerebellum were isolated and evaluated for gene and protein expression levels of Nbg and Cygb. The results demonstrated that the 30 µg/kg AgNP group displayed increased gene and protein expression of Cygb in the cortex. In the Hippocampus, AgNP exposure did not modulate gene or protein expression levels of Ngb and Cygb. In cerebellum the Ngb gene and protein expression was increased with both doses of AgNP. AgNP exposure during prepubescence can modulate the gene and protein expression levels of Ngb and Cygb in adulthood. Furthermore, the observed modulation was specific to the cerebellum, and cortex, and was dose dependent.

Anatomical specificity of the brain in the modulation of Neuroglobin and Cytoglobin genes after chronic bisphenol a exposure.

The aim of this study was to investigate the influence of Bisphenol A (BPA) exposure on Neuroglobin (Ngb) and Cytoglobin (Cygb) as well as oxidative stress gene expression in the cerebellum, hippocampus, hypothalamus and cortex. Male Wistar rats were randomly divided into 3 groups: Control and two groups receiving 2 different daily BPA dosages, 5 or 25 mg/kg from postnatal day 50 (PND50) through PND90 and they were euthanized at PND105. In the cortex, we found an increase in Ngb gene expression and also in superoxide dismutase 1 and Catalase (Cat). In the cerebellum, we found an increase in Ngb and Cat, in the hypothalamus, there was a decrease in Cygb and an increase in glutathione peroxidase and Cat and in hypoxia-inducible factor 1 alpha (Hif1α) at the low dosage and a decrease in Hif1α at the high BPA dosage. Finally, in the hippocampus, we observed a decrease in Ngb and Cygb and an increase in Hif1α. In summary, BPA promotes the modulation of both Ngb and Cygb, but such changes occur by different mechanisms depending on the exposure dose and anatomical area.

Antidepressant behavior in thyroidectomized Wistar rats is induced by hippocampal hypothyroidism.

Thyroidectomy is a surgical procedure indicated in cases of several maligned or benign thyroid diseases, thus, the aim of our study was to verify how the hypothyroidism induced by thyroidectomy influences behavioral parameters and its relation to thyroid hormones metabolism and neurogenesis at hippocampus. For this purpose, Adult male Wistar rats underwent to thyroidectomy to induce hypothyroidism. Behavioral tests, the thyroid profile and hippocampal gene expression were evaluated in control and in thyroidectomized animals. It was observed that thyroidectomized group had a significant increasing in serum thyroid-stimulating hormone (TSH) and a decreasing in thyroxine (T4) levels as well as in triiodothyronine (T3) serum level. It was also observed reduction of the monocarboxylate transporter 8 (Mct8), thyroid hormone receptor alfa (Trα1), deiodinase type 2 (Dio2), ectonucleotide pyrophosphatase/phosphodiesterase 2 (Enpp2) and brain-derived neurotrophic factor (Bdnf) mRNA expression in hippocampus of thyroidectomized animals. In the forced swimming test, it was verified that thyroidectomy promotes a decrease in time of immobility and climbing when compared with the control group. In summary, we demonstrated that antidepressant behavior in thyroidectomized Wistar rats is induced by hippocampal hypothyroidism. This effect could be associated to an impaired neuronal activity in acute stress response as it is observed in forced swimming paradigm.

Thyroid hormone modulates neuroglobin and cytoglobin in rat brain.

Thyroid hormones (THs) are essential and crucial for brain development, playing a role in growth and differentiation. Two globins named neuroglobin (Ngb) and cytoglobin (Cygb) are located in the brain, and each one has different distribution and function: They seem to have similar action by providing O(2) for respiratory chain, and detoxification of reactive oxygen species (ROS) and nitric oxide (NO) protecting tissues against irreversible lesions. We aimed to investigate the influence of thyroid state in Ngb and Cygb metabolism in different brain regions and evaluate their responses in cerebellum, hippocampus and cerebral cortex (hereafter called as cortex) after supraphysiological doses at different time points of TH administration. Experiments were carried out in rats, divided in eight experimental groups Control (C), thyroidectomy (Tx), and thyroidectomy treated with jugular intravenous injection (i.v). T3 (100 µl/100 g) injection and sacrificed after 30, 60, 120 min and 6, 12 and 24 h. In cortex, we found increase in Ngb gene and protein expression in different time points compared to C group, however Cygb gene and protein expression were decreased. In hippocampus, Ngb and Cygb protein expression increased 24 h after i.v. T3 injection in comparison to Tx. In cerebellum, we found increased Ngb gene expression after 120 min, 6, 12 and 24 h after T3 administration compared to Tx, and in contrast, protein expression was found to be significantly increased only 12 and 24 h compared to Tx. Ngb and Cygb expression in brain is influenced by thyroid hormone state both by its lack or excess.