Zika virus (ZIKV) infection in utero might lead to microcephaly and other congenital defects. Since no specific therapy is available thus far, there is an urgent need for the discovery of agents capable of inhibiting its viral replication and deleterious effects. Chloroquine is widely used as an antimalarial drug, anti-inflammatory agent, and it also shows antiviral activity against several viruses. Here we show that chloroquine exhibits antiviral activity against ZIKV in Vero cells, human brain microvascular endothelial cells, human neural stem cells, and mouse neurospheres. We demonstrate that chloroquine reduces the number of ZIKV-infected cells in vitro, and inhibits virus production and cell death promoted by ZIKV infection without cytotoxic effects. In addition, chloroquine treatment partially reveres morphological changes induced by ZIKV infection in mouse neurospheres.

Astrocytes, members of the glial family, interact through the exchange of soluble factors or by directly contacting neurons and other brain cells, such as microglia and endothelial cells. Astrocytic projections interact with vessels and act as additional elements of the Blood Brain Barrier (BBB). By mechanisms not fully understood, astrocytes can undergo oncogenic transformation and give rise to gliomas. The tumors take advantage of the BBB to ensure survival and continuous growth. A glioma can develop into a very aggressive tumor, the glioblastoma (GBM), characterized by a highly heterogeneous cell population (including tumor stem cells), extensive proliferation and migration. Nevertheless, gliomas can also give rise to slow growing tumors and in both cases, the afflux of blood, via BBB is crucial. Glioma cells migrate to different regions of the brain guided by the extension of blood vessels, colonizing the healthy adjacent tissue. In the clinical context, GBM can lead to tumor-derived seizures, which represent a challenge to patients and clinicians, since drugs used for its treatment must be able to cross the BBB. Uncontrolled and fast growth also leads to the disruption of the chimeric and fragile vessels in the tumor mass resulting in peritumoral edema. Although hormonal therapy is currently used to control the edema, it is not always efficient. In this review we comment the points cited above, considering the importance of the BBB and the concerns that arise when this barrier is affected.

IMPORTANCE Recent studies have reported an increase in the number of fetuses and neonates with microcephaly whose mothers were infected with the Zika virus (ZIKV) during pregnancy. To our knowledge, most reports to date have focused on select aspects of the maternal or fetal infection and fetal effects. OBJECTIVE To describe the prenatal evolution and perinatal outcomes of 11 neonates who had developmental abnormalities and neurological damage associated with ZIKV infection in Brazil. DESIGN, SETTING, AND PARTICIPANTS We observed 11 infants with congenital ZIKV infection from gestation to 6 months in the state of Paraíba, Brazil. Ten of 11 women included in this study presented with symptoms of ZIKV infection during the first half of pregnancy, and all 11 had laboratory evidence of the infection in several tissues by serology or polymerase chain reaction. Brain damage was confirmed through intrauterine ultrasonography and was complemented by magnetic resonance imaging. Histopathological analysis was performed on the placenta and brain tissue from infants who died. The ZIKV genome was investigated in several tissues and sequenced for further phylogenetic analysis. MAIN OUTCOMES AND MEASURES Description of the major lesions caused by ZIKV congenital infection. RESULTS Of the 11 infants, 7 (63.6%) were female, and the median (SD) maternal age at delivery was 25 (6) years. Three of 11 neonates died, giving a perinatal mortality rate of 27.3%. The median (SD) cephalic perimeter at birth was 31 (3) cm, a value lower than the limit to consider a microcephaly case. In all patients, neurological impairments were identified, including microcephaly, a reduction in cerebral volume, ventriculomegaly, cerebellar hypoplasia, lissencephaly with hydrocephalus, and fetal akinesia deformation sequence (ie, arthrogryposis). Results of limited testing for other causes of microcephaly, such as genetic disorders and viral and bacterial infections, were negative, and the ZIKV genome was found in both maternal and neonatal tissues (eg, amniotic fluid, cord blood, placenta, and brain). Phylogenetic analyses showed an intrahost virus variation with some polymorphisms in envelope genes associated with different tissues. CONCLUSIONS AND RELEVANCE Combined findings from clinical, laboratory, imaging, and pathological examinations provided a more complete picture of the severe damage and developmental abnormalities caused by ZIKV infection than has been previously reported. The term congenital Zika syndrome is preferable to refer to these cases, as microcephaly is just one of the clinical signs of this congenital malformation disorder.

Zika virus (ZIKV) has been associated to central nervous system (CNS) harm, and virus was detected in the brain and cerebrospinal fluids of microcephaly and meningoencephalitis cases. However, the mechanism by which the virus reaches the CNS is unclear. Here, we addressed the effects of ZIKV replication in human brain microvascular endothelial cells (HBMECs), as an in vitro model of blood brain barrier (BBB), and evaluated virus extravasation and BBB integrity in an in vivo mouse experimental model. HBMECs were productively infected by African and Brazilian ZIKV strains (ZIKVMR766 and ZIKVPE243), which induce increased production of type I and type III IFN, inflammatory cytokines and chemokines. Infection with ZIKVMR766 promoted earlier cellular death, in comparison to ZIKVPE243, but infection with either strain did not result in enhanced endothelial permeability. Despite the maintenance of endothelial integrity, infectious virus particles crossed the monolayer by endocytosis/exocytosis-dependent replication pathway or by transcytosis. Remarkably, both viruses' strains infected IFNAR deficient mice, with high viral load being detected in the brains, without BBB disruption, which was only detected at later time points after infection. These data suggest that ZIKV infects and activates endothelial cells, and might reach the CNS through basolateral release, transcytosis or transinfection processes. These findings further improve the current knowledge regarding ZIKV dissemination pathways.

Video-microscopy in combination with digital image processing was used to analyze dynamic processes associated to the life cycle of Giardia lamblia trophozoites. These parasites swim and attach to the epithelial cells, producing the disease known as Giardiasis. Giardia is a multiflagellar cell, presenting 4 pairs of flagella. With the use of analogue and digital tools, we observed that in cells attached to glass slides only 2 of the 4 pairs present active beating (wave propagation). The frequency observed was 17-18 Hz to the anterior and 8-11 Hz to the ventral flagella. These data resulted from several hours of recording using both analogue video and high-speed digital camera. The caudal pair did not show active beating patterns and the same holds true for the posterior one. In this latter pair, oscillations were observed, but they were always associated to the transit of the wave produced by the ventral pair. The analysis performed with free moving cells showed that during its forward dislocation, Giardia lamblia presented either a lateral rocking or a complete rotational (tumbling) movement around its longitudinal axis. A dislocation of the caudal region of the cell both in the lateral and dorso-ventral direction was observed. This movement was completely independent from the flagellar beating and it is likely to be produced by a microtubular complex located in the caudal portion of the cell. The adhesion process of Giardia lamblia was also followed by video-microscopy and the data showed that the ventral disk had an active participation in this process.

Zika virus (ZIKV) causes significant public health concerns because of its association with congenital malformations, neurological disorders in adults, and, more recently, death. Considering the necessity to mitigate ZIKV-associated diseases, antiviral interventions are an urgent necessity. Sofosbuvir, a drug in clinical use against hepatitis C virus (HCV), is among the FDA-approved substances endowed with anti-ZIKV activity. In this work, we further investigated the in vivo activity of sofosbuvir against ZIKV. Neonatal Swiss mice were infected with ZIKV (2 × 107 PFU) and treated with sofosbuvir at 20 mg/kg/day, a concentration compatible with pre-clinical development of this drug. We found that sofosbuvir reduced acute levels of ZIKV from 60 to 90% in different anatomical compartments, such as the blood plasma, spleen, kidney, and brain. Early treatment with sofosbuvir doubled the percentage and time of survival of ZIKV-infected animals. Sofosbuvir also prevented the acute neuromotor impairment triggered by ZIKV. In the long-term behavioural analysis of ZIKV-associated sequelae, sofosbuvir prevented loss of hippocampal- and amygdala-dependent memory. Our results indicate that sofosbuvir inhibits ZIKV replication in vivo, which is consistent with the prospective necessity of antiviral drugs to treat ZIKV-infected individuals.

Sox2 is an important transcriptional regulator in embryonic and adult stem cells. Recently, Sox2 was identified as an oncogene in many endodermal cancers, including colon cancer. There is great interest in how Sox2 cooperates with other transcription factors to regulate stem cell renewal, differentiation, and reprogramming. However, we still lack a general understanding of Sox2 transcriptional action. To determine transcriptional partners of Sox2 in adult cells, we generated mice where gene expression could be induced by an externally applied stimulus. We analyzed the consequences in the intestine where cell turnover is rapid. Sox2 expression, but not Oct4, specifically increased the numbers of stem cells and repressed Cdx2, a master regulator of endodermal identity. In vivo studies demonstrated that Sox21, another member of the SoxB gene family, was a specific, immediate, and cell-autonomous target of Sox2 in intestinal stem cells. In vitro experiments showed that Sox21 was sufficient to repress Cdx2 in colon cancer cells and in pluripotent stem cells. Sox21 was also specifically induced by Sox2 in fibroblasts and inhibition of Sox21 blocked reprogramming to the pluripotent state. These results show that transcriptional induction of Sox21 is a rapid and general mediator of the effects of Sox2 on cell identity in a wide range of cell types.

BackgroundThe ability to grow a uniform cell type from the adult central nervous system (CNS) is valuable for developing cell therapies and new strategies for drug discovery. The adult mammalian brain is a source of neural stem cells (NSC) found in both neurogenic and non-neurogenic zones but difficulties in culturing these hinders their use as research tools [1], [2], [3], [4], [5], [6].Methodology/Principal FindingsHere we show that NSCs can be efficiently grown in adherent cell cultures when angiogenic signals are included in the medium. These signals include both anti-angiogenic factors (the soluble form of the Notch receptor ligand, Dll4) and pro-angiogenic factors (the Tie-2 receptor ligand, Angiopoietin 2). These treatments support the self renewal state of cultured NSCs and expression of the transcription factor Hes3, which also identifies the cancer stem cell population in human tumors. In an organotypic slice model, angiogenic factors maintain vascular structure and increase the density of dopamine neuron processes.Conclusions/SignificanceWe demonstrate new properties of adult NSCs and a method to generate efficient adult NSC cultures from various central nervous system areas. These findings will help establish cellular models relevant to cancer and regeneration.