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Neurogenesis was believed to end after the period of embryonic development. However, the possibility of obtaining an expressive number of cells with functional neuronal characteristics implied a great advance in experimental research. New techniques have emerged to demonstrate that the birth of new neurons continues to occur in the adult brain. Two main rich sources of these cells are the subventricular zone (SVZ) and the subgranular zone of the hippocampal dentate gyrus (SGZ) where adult neural stem cells (aNSCs) have the ability to proliferate and differentiate into mature cell lines. The cultivation of neurospheres is a method to isolate, maintain and expand neural stem cells (NSCs) and has been used extensively by several research groups to analyze the biological properties of NSCs and their potential use in injured brains from animal models. Throughout this review, we highlight the areas where this type of cell culture has been applied and the advantages and limitations of using this model in experimental studies for the neurological clinical scenario.

To assess, in vitro, the influence of heat air treatment on cytotoxicity and degree of conversion (DC) of universal self-etch adhesives (Ambar Universal APS, Scotchbond Universal Adhesive, and Tetric N-Bond Universal) in an NIH/3T3 fibroblast cell culture. Samples were divided into three groups: 1) no heat treatment (control), 2) 37°C and 3) 60°C heat treatment before photopolymerization. Cytotoxicity was analyzed by MTT assay and the DC by FTIR. All adhesives heated at 60°C showed reduced cytotoxicity levels when compared with those heated at 37°C. In general, DC of Ambar Universal APS presented the highest DC than Scotchbond Universal Adhesive and Tetric N-Bond Universal, and the hot air treatment do not influence the conversion. Heat treatment at 60°C was able to reduce the cytotoxicity of universal self-etch adhesives, even, the heat treatment does not enhances the DC.

Temporal lobe epilepsy is the most common form of intractable epilepsy in adults. More than 30% of individuals with epilepsy have persistent seizures and have drug-resistant epilepsy. Based on our previous findings, treatment with bone marrow mononuclear cells (BMMC) could interfere with early and chronic phase epilepsy in rats and in clinical settings. In this pilocarpine-induced epilepsy model, animals were randomly assigned to two groups: control (Con) and epileptic pre-treatment (Ep-pre-t). The latter had status epilepticus (SE) induced through pilocarpine intraperitoneal injection. Later, seizure frequency was assessed using a video-monitoring system. Ep-pre-t was further divided into epileptic treated with saline (Ep-Veh) and epileptic treated with BMMC (Ep-BMMC) after an intravenous treatment with BMMC was done on day 22 after SE. Analysis of neurobehavioral parameters revealed that Ep-BMMC had significantly lower frequency of spontaneous recurrent seizures (SRS) in comparison to Ep-pre-t and Ep-Veh groups. Hippocampus-dependent spatial and non-spatial learning and memory were markedly impaired in epileptic rats, a deficit that was robustly recovered by treatment with BMMC. Moreover, long-term potentiation-induced synaptic remodeling present in epileptic rats was restored by BMMC. In addition, BMMC was able to reduce abnormal mossy fiber sprouting in the dentate gyrus. Molecular analysis in hippocampal tissue revealed that BMMC treatment down-regulates the release of inflammatory cytokine tumor necrosis factor-α (TNF-α) and Allograft inflammatory factor-1 (AIF-1) as well as the Rho subfamily of small GTPases [Ras homolog gene family member A (RhoA) and Ras-related C3 botulinum toxin substrate 1 (Rac)]. Collectively, delayed BMMC treatment showed positive effects when intravenously infused into chronic epileptic rats.

Focal cortical dysplasia (FCD) is a malformation of cortical development which is strongly associated with drug-refractory epilepsy. Certain studies have demonstrated an increase in mTOR signaling in patients with FCD on the basis of observation of phosphorylated molecules. The aim of the present study was to verify the differences in genes involved in cell proliferation, adhesion, and control of apoptosis during embryonic neurogenesis in iPSCs derived from the Focal Cortical Dysplasia. Fibroblasts were obtained from the skin biopsies of patients with FCD (n = 2) and controls (n = 2). iPSCs were generated by exposing the fibroblasts to viral vectors that contained the Yamanaka factors (OCT4, SOX2, KLF4, and c-MYC genes) responsible for promoving cell reprogramation. The fibroblasts and iPSCs were tested during different phases of neurodifferentiation for migration capacity and expression of the genes involved in the PI3K pathway. Fibroblasts of patients with FCD migrated with greater intensity during the first two time points of analyses. iPSCs did not exhibit any difference in cell migration between the groups. Fibroblasts, brain tissue, and iPSCs of the patients with FCD exhibited a significant reduction in the relative expression values of 4EBP-1. During neurodevelopment, the iPSCs from patients with FCD exhibited a reduction in the expression of cIAP-1, cIAP-2, PI3K, β-Catenin and 4EBP-1 gene. We suggest that the differences observed in the migration potential of adult cells and in the gene expression related to the fundamental processes involved in normal brain development during the neurodifferentiation process might be associated with cortical alteration in the patients with FCD.

The microdiscectomy used for the treatment of intervertebral disc disorders leaves an open incision in the annulus fibrosus that must be sealed to avoid re-herniation and other subsequent degenerations. In this study, we developed an injectable and in situ polymerizable polyurethane adhesive as a long-term post-surgical annulus fibrosus repair strategy. It was investigated the chemical structure of the urethane-based adhesive and its physico-chemical, viscoelastic, kinetic, and in vitro cytotoxic properties. The adhesive formulated from the polycarbonate diol with the highest molar mass was the one that exhibited a compressive behavior closest to the intervertebral disc outer region, and therefore, the most suitable for restoration. This adhesive showed 18-day stability under moisture and required a preparation time of 10 h at 60°C before use. The material also adhered covalently to gelatin (without catalyst or initiator) and positively impacted cell proliferation after its polymerization, which are essential requirements for clinical translation. These findings confirmed the ability of the polyurethane adhesive to act as an annulus fibrosus sealant, although further improvements in its formulation are necessary.

Undernourishment is a global issue, especially in developing countries, affecting newborns and children in a vulnerable period of brain development. Previous studies of undernourishment models suggested a relationship between undernourishment and epilepsy. The exposure to both undernourishment and recurrent seizures early in life appears to have detrimental effects on the developing brain. This study aims to investigate the neurobiological consequences of undernourishment and recurrent seizures exposure early in life, investigating Long-Term Potentiation (LTP) induction and gene expression of NMDA receptor subunits in the hippocampus during adulthood (P60). Animals were exposed to maternal deprivation protocol from P2 to P15 to control food intake in rat pups and Flurothyl-induced seizures from P7 to P10. Electrophysiological records of hippocampal slices were recorded and gene expression of NR1A, NR2A, NR2B, NR2C, NR2D and BDNF were investigated. Animals exposed to undernourishment or recurrent seizures failed to promote LTP after stimulation. Furthermore, seizure exposure early in life led to increased expression of hippocampal NR1A, NR2A, NR2B, NR2C and NR2D when compared to controls. Interestingly, when animals were exposed to undernourishment paradigm early in life, this upregulation of NDMA subunits was absent. In conclusion, our study showed impaired LTP after undernourishment and recurrent seizures early in life, together with differential expression of NDMA expression in the hippocampus during adulthood.

Despite global efforts to establish effective interventions for coronavirus disease 2019 (COVID-19) and its major complications, such as acute respiratory distress syndrome (ARDS), the treatment remains mainly supportive. Hence, identifying an effective and safe therapy for severe COVID-19 is critical for saving lives. A significant number of cell-based therapies have been through clinical investigation. In this study, we performed a systematic review of clinical studies investigating different types of stem cells as treatments for COVID-19 and ARDS to evaluate the safety and potential efficacy of cell therapy. The literature search was performed using PubMed, Embase, and Scopus. Among the 29 studies, there were eight case reports, five Phase I clinical trials, four pilot studies, two Phase II clinical trials, one cohort, and one case series. Among the clinical studies, 21 studies used cell therapy to treat COVID-19, while eight studies investigated cell therapy as a treatment for ARDS. Most of these (75%) used mesenchymal stem cells (MSCs) to treat COVID-19 and ARDS. Findings from the analyzed articles indicate a positive impact of stem cell therapy on crucial immunological and inflammatory processes that lead to lung injury in COVID-19 and ARDS patients. Additionally, among the studies, there were no reported deaths causally linked to cell therapy. In addition to standard care treatments concerning COVID-19 management, there has been supportive evidence towards adjuvant therapies to reduce mortality rates and improve recovery of care treatment. Therefore, MSCs treatment could be considered a potential candidate for adjuvant therapy in moderate-to-severe COVID-19 cases and compassionate use.

The pandemic experienced in recent months has raised questions that should be investigate in the clinical practice. Transplacental transmission of SARS-CoV-2 and the consequences to the fetus and newborn have called attention due to the increasing number of infections, contradicting previous evidences that there was no possibility of coronavirus transmission from the mother to the fetus. In this work we showed three cases of pregnant women with positive SARS-CoV-2 antibodies serology on admission in Naval Hospital Marcílio Dias (HNMD), Rio de Janeiro, Brazil. Samples of umbilical cord blood was double positive (IgM and IgG) for one patient, double negative for one patient and positive for IgG and negative for IgM for third patient. Maternal and neonatal nasopharyngeal swab samples analyzed by PCR for SARS-CoV-2 was positive for two maternal patients and negative for all newborns tested. It was possible to detect the SARS-CoV-2 in amniotic fluid and umbilical cord blood using the nested-PCR technics, thus being successfully evidenced transplacental transmission. We suggested that nasopharyngeal swab PCR test of neonates does not have a correlation with vertical transmission and thus, this molecular test is not useful for investigation of transplacental infection.

Induced pluripotent stem cells and cerebral organoids enable a better understanding of the pathophysiology, mechanisms of action, identification of therapeutic targets, and new drugs and alternatives for the treatment of several pathologies, including profiling Zika virus infection. Those allow us to observe aspects of the disease pathogenesis that would be very difficult or impossible when compared with other study models, such as postmortem tissue and animal model. This chapter aims to address these two main types of disease modeling tools associated with the several findings applied to Zika virus infection.

Background. Direct pulp capping is a method designed to preserve the exposed dental pulp. Due to good biological, physical, and mechanical properties, new versions of calcium silicate-based materials have been developed as pulp capping materials. The present study aimed to evaluate the cytotoxic effects of four calcium silicate-based pulp capping materials, of which the Bio-C Repair Íon+ is still in an experimental phase. Methods. Biodentine, MTA Repair HP, Bio-C Repair, and Bio-C Repair Íon+ cements were dispensed in a metallic matrix to produce 125-mm3 specimens, which were immersed in Dulbecco’s Modified Eagle Medium (DMEM) to obtain extracts. NIH 3T3 cells were cultured and exposed to the extracts for 24 hours and seven days. Cell viability was assessed by the methyl tetrazolium test (MTT). The mean values for the experimental and control groups (without treatment) were compared by analysis of variance (ANOVA) and post hoc Tukey tests, considering a significance level of 5%. Results. All the tested materials demonstrated a reduction in cell viability (P

Background: Pregnant women are susceptible to the novel coronavirus (SARS-CoV-2) and the consequences on the fetus are still uncertain. Here, we present a case of a pregnant woman with subclinical hypothyroidism and PAI-1 4G/5G mutation who was infected with SARS-CoV-2 at the end of the third trimester of pregnancy. Methods: nested PCR were performed to detect the virus, followed by ssDNA sequencing. Results: transplacental transmission of SARS-CoV-2 can cause placental inflammation, ischemia and neonatal viremia, with complications such as preterm labor and damage to the placental barrier in patients with PAI-1 4G/5G mutation. Conclusion: we show the possibility of transplacental transmission of SARS-CoV-2 infection during the last weeks of pregnancy.

Phylogenetic analysis has demonstrated that the etiologic agent of the 2020 pandemic outbreak is a betacoronavirus named SARS-CoV-2. For public health interventions, a diagnostic test with high sensitivity and specificity is required. The gold standard protocol for diagnosis by the Word Health Organization (WHO) is RT-PCR. To detect low viral loads and perform large-scale screening, a low-cost diagnostic test is necessary. Here, we developed a cost-effective test capable of detecting SARS-CoV-2. We validated an auxiliary protocol for molecular diagnosis with the SYBR Green RT-PCR methodology to successfully screen negative cases of SARS-CoV-2. Our results revealed a set of primers with high specificity and no homology with other viruses from the Coronovideae family or human respiratory tract pathogenic viruses, presenting with complementarity only for rhinoviruses/enteroviruses and Legionella spp. Optimization of the annealing temperature and polymerization time led to a high specificity in the PCR products. We have developed a more affordable and swift methodology for negative SARS-CoV-2 screening. This methodology can be applied on a large scale to soften panic and economic burden through guidance for isolation strategies.

Infection with Zika virus (ZIKV) was recently demonstrated to be associated with damage to the central nervous system, especially microcephaly and the Guillain-Barré syndrome. This finding had alarmed public health agencies and mobilized institutions around the world to search for more information about the virus, its effects, pathophysiological mechanisms, and potential immunizations and treatments. Given the increasing interest in using iPSCs and cerebral organoids to model the congenital infection and neuropathogenesis induced by ZIKV, the aim of this review was to present an up-to-date summary of the publications on the association of ZIKV with microcephaly, using iPSCs and organoids. According to our review, the number of studies has decreased concomitantly with a decrease in the number of cases. The presence of subclinical lesions at birth, which may eventually present cognitive or behavioral problems in the future, suggests that persistent research efforts on the virus should be undertaken by the global health community till the threat is completely wiped out.

Focal cortical dysplasia (FCD) is the most commonly encountered developmental malformation that causes refractory epilepsy. Focal cortical dysplasia type 2 is one of the most usual neuropathological findings in tissues resected therapeutically from patients with drug-resistant epilepsy. Unlike other types of FCD, it is characterized by laminar disorganization and dysplastic neurons, which compromise the organization of the six histologically known layers in the cortex; the morphology and/or cell location can also be altered. A comprehensive review about the pathogenesis of this disease is important because of the necessity to update the results reported over the past years. Here, we present an updated review through Pubmed about the mammalian target of rapamycin (MTOR) pathway in FCD type 2. A wide variety of aspects was covered in 44 articles related to molecular and cellular biology, including experiments in animal and human models. The first publications appeared in 2004, but there is still a lack of studies specifically for one type of FCD. With the advancement of techniques and greater access to molecular and cellular experiments, such as induced pluripotent stem cells (iPSCs) and organoids, it is believed that the trend is increasing the number of publications contributing to the achievement of new discoveries.

Focal cortical dysplasia (FCD) is caused by numerous alterations, which can be divided into abnormalities of the cortical architecture and cytological variations; however, the exact etiology of FCD remains unknown. The generation of induced pluripotent stem cells (iPSCs) from the cells of patients with neurological diseases, and their subsequent tissue‑specific differentiation, serves as an invaluable source for testing and studying the initial development and subsequent progression of diseases associated with the central nervous system. A total of 2 patients demonstrating seizures refractory to drug treatment, characterized as FCD Type IIb, were enrolled in the present study. Fibroblasts were isolated from residual skin fragments obtained from surgical treatment and from brain samples obtained during surgical resection. iPSCs were generated following exposure of fibroblasts to viral vectors containing POU class 5 homeobox 1 (OCT4), sex determining region Y‑box 2 (SOX2), Kruppel‑like factor 4 and c‑MYC genes, and were characterized by immunohistochemical staining for the pluripotent markers homeobox protein NANOG, SOX2, OCT4, TRA1‑60 and TRA1‑81. The brain samples were tested with antibodies against protein kinase B (AKT), phosphorylated‑AKT, mechanistic target of rapamycin (mTOR) and phosphorylated‑mTOR. Analysis of the AKT/mTOR pathway revealed a statistically significant difference between the cerebral tissues of the two patients, which were of different ages (45 and 12 years old). Clones with the morphological features of embryonic cells were detected on the 13th day and were characterized following three subcultures. The positive staining characteristics of the embryonic cells confirmed the successful generation of iPSCs derived from the patients' fibroblasts. Therefore, the present study presents a method to obtain a useful cellular source that may help to understand embryonic brain development associated with FCD.

In cellular therapy models, the establishment of accurate methods to track the homing, cell fate and survival are fundamental to ensure safety and efficacy. The PCR is used to detect donor cells DNA in xenotransplant models. Five different concentrations of EGFP+ DNA from C57/BL6 mouse muscle tissue: 1,000 pg/μL, 500 pg/μL, 100 pg/μL, 10 pg/μL e 1 pg/μL were used to contaminate 10 ng/μL of DNA Wistar rat. Polymerase chain reaction was performed using 2 μL of each sample. EGFP+DNA were detected in samples containing 2,000 pg, 1,000 pg, 200 pg and 20 pg. Samples with 2 pg EGFP+DNA did not show signal. Here we demonstrate an alternative to proving the presence or confirm the absence, of exogenous DNA in organs and tissues. The concentration curve for cell migration detection by real time PCR will allow the use in systemically transplanted cells in animal models subjected to cell therapy.

The pathogenesis of endometriosis is not clear; however, microRNAs (miRNAs/miRs) are involved in the pathogenesis. miRNAs are short noncoding RNAs involved in post-transcriptional regulation of gene expression by silencing the expression of target genes. The expression of miR-135a/b is associated with endometrial receptivity and implantation; the expression is also associated with the expression of certain genes, including homeobox protein Hox-A10 (HOXA-10). The present study investigated the expression of miR-135a/b in eutopic and ectopic endometrium tissues throughout the different phases of the menstrual cycle. Samples of ectopic endometriosis lesions and eutopic endometrium tissue from 23 patients who underwent laparoscopic surgery were obtained and analyzed. miRNA was extracted and the expression levels of miR-135a/b were determined by reverse transcription quantitative polymerase chain reaction assays using U6 as a housekeeping control. The expression levels of miR-135a and miR-135b in endometriosis lesions were decreased compared with the levels in endometrium tissue. However, miR-135a/b expression levels were increased in the secretory phase compared with the proliferative phase in endometriosis lesions. The increased expression of miR-135a/b during the secretory phase compared with the proliferative phase suggested that these genes serve a determinant role in the homeostasis of reproductive tissue. Therefore, the expression of genes may affect endometrial functioning, impairing embryo implantation.

Introduction Multiple Sclerosis (MS) is a chronic, autoimmune, demyelinating disease of the central nervous system (CNS). Currently, several protocols are described for the different phases of MS. In this longitudinal study, we aim to quantify the concentration of plasma cytokines of MS patients treated with Fingolimod alone or after Glatiramer Acetate (GA) or Interferon-beta (IFN-β), in order to compeer both treatments and describes if it is possible to use them as biomarkers. Objective Compare the two different types of drug treatment and describes possible immune biomarkers in RRMS patients treated with Fingolimod alone or after GA or IFN-β. Materials and methods This is a controlled, non-randomized clinical trial. Plasma concentrations of IL-31, sCD40L and nine others cytokines were evaluated in two groups of patients with a one-year follow-up. Group 1 (n = 12): RRMS patients treated with GA or IFN-β for at least six months before the study who changed therapy to Fingolimod after six months, and Group 2 (n = 12): naive RRMS patients who started treatment with Fingolimod. We used ANOVA two-way to analyze the cytokines and Spearman coefficient to evaluate the correlation. Results Although Group 2 started with a greater number of relapses per disease duration, Fingolimod treatment was effective in decreasing this parameter, as well as EDSS over 12 months. However, the treatment with GA or IFN-β on Group 1 showed a tendency to increase the number of relapses after 6 months of follow-up, which decrease when the therapy was changed to Fingolimod. After the evaluation of 11 cytokines in one year, we found that IL-31 and sCD40L were the biomarkers that demonstrated a more difference when compared to the classical ones, following the clinical pattern over the treatment period. Conclusions Our study describes the existence of two promising plasmatic biomarkers (IL-31 and sCD40L), which reduced plasmatic levels in RRMS patients followed the treatment time of Fingolimod, despite that more studies are needed to prove their efficiency.

Malformations of cortical development (MCDs) include many different Central Nervous System (CNS) disorders related to a complex process of cortex formation. In children with refractory epilepsy to drug treatment undergoing surgery, focal cortical dysplasia (FCD), one of the MCDs, is considered the most common structural brain lesion found. This study aimed to study the possible alterations in neural differentiation process of human induced pluripotent stem cells (hiPSCs) related to migration and synaptic aspects from fibroblasts of two individuals affected by FCD type IIb (45-year-old male and 12-year-old female) and normal individuals. At the days 14th, 22nd and 35th, hiPSCs were neural differentiated and analyzed. Using qRT-PCR approach, the expression of 9 genes associated with synaptic and neural migration were quantified. Diagnostic of both patients was consistent with FCD type IIb. Our results showed that in all processes and groups, individuals with dysplasia presented alterations in most part of the genes in relation to control individuals. According to our results, it is suggested that the different expressions are mainly involved in alterations of the expression of receptors and capture sites, timing, coupling of synaptic vesicles with the presynaptic membrane, regulation of ion channel and synaptic exocytosis, imbalance of the apoptosis process and abnormal microtubules that may also contribute to delays in synaptogenesis. Thus, brain formation with dysplasia is probably influenced by these genes studied.

The human umbilical cord blood (HUCB) is an excellent source of adult stem cells, having the benefit of being younger than the bone marrow stem cells. The role of stem cells in the lesion repair mechanism is still being studied. We evaluated the capability of HUCB to interfere into the fibroblast dedifferentiation plasticity through cocultivation. Direct and indirect cocultures were maintained for 24, 48, and 72 hours. Coculture viability was evaluated by MTT assay. The messenger RNA was extracted, and the expression of p16 and p21 genes was estimated by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). The direct or indirect contact did not interfere with fibroblast cell viability. However, these direct and indirect contacts reduced the expression of p16 and p21 genes. A sigmoidal curve was applied to adjust gene expression against time, and a mathematical function was established for gene expression according to cell culture type. These results suggest that the differentiated cells were influenced by immature cells (HUCB) either by the direct contact or by signaling molecules, which alter their behavior and plasticity. Therefore our data may contribute to paracrine effects other than the commonly known to be responsible for the repair of lesions in stem cell therapy.

Focal Cortical Dysplasia (FCD), a common type of Malformations of cortical development, may result from an early disturbance in the migration and final arrangement of the cortical architecture of immature neurons. FCD type II is now known to be due to a post-zygotic somatic mutation that involves the mTOR and AKT pathways. The aim of the present study was to investigate the possible differences in neurogenesis and neurodifferentiation of iPSCs (induced pluripotent stem cells) from fibroblasts of individuals affected by FCD type II (2) and normal individuals (2). iPSCs were generated from skin fibroblasts of FCD individuals and healthy individuals. The reprogramming was done through the fibroblasts exposure to viral vectors containing the OCT4, KLF4, SOX2, and c-MYC genes and the clones were characterized by immunohistochemistry. iPSCs were neurodifferentiated and analyzed at the 14th, 22nd and 35th days. We also analyzed the cerebral cortex tissue, fibroblasts and iPSCs cells from the individuals. Through qRT-PCR, the expression of 4 genes involved in Notch signaling process were quantified. In general, individuals with dysplasia presented increase and decrease in the relative quantification in the most genes analyzed compared to control individuals in all processes and study groups. We suggest that, during embryonic neurogenesis, the neural precursor cells of FCD type II individuals present increase and decrease in gene expression in the Notch signaling pathway causing cortical formation disorders and can be seen as a candidate for the developmental changes observed in the cerebral cortex of individuals with FCD type II. This altered gene expression may be related to brain formation with dysplasia.

Engineered skin coverings have been adopted clinically to support extensive and deep wounds that result in fewer healthy skin remaining and therefore take longer to heal. Nonetheless, these biomaterials demand intensive labor and an expensive final cost. In comparison to conventional bandages, which do not meet all the requirements of wound care, electrospun fiber mats could potentially provide an excellent environment for healing. In this work, we developed two nanostructured scaffolds based on polyamide-6 (PA-6) to be tested as a wound covering in a rat model of full-thickness incisional wound healing. The central idea was to create a bioconstruct that is simple to implement and biologically safe, with a high survival rate, which provides physical support and biological recognition for new functional tissues. An unmodified PA-6 and a soybean-modified PA-6 were employed as nanofibrillar matrices in this study. The biomaterials showed a dimensional homology to natural extracellular matrix components and neither in vitro toxicity nor in vivo side effects. Both polymeric scaffolds were resistant to the sterilization process and could promote the attachment of 3T3 fibroblast cells, besides successfully incorporating the growth factor PDGF-BB, which had its bioactivity extended for up to 12 h under simulated conditions. The modification of PA-6 chains with a fatty acid derivative increased the scaffold's surface free energy, favoring cell proliferation, collagen formation, and ECM secretion. These results confirm the potential of these materials as a topical dermal covering for skin regeneration.

Summary Aims In previous studies, transplantation of bone marrow mononuclear cells (BMMCs) in epileptic animals has been found to be neuroprotective. However, the mechanism by which the BMMCs act remains unclear. We hypothesize that BMMCs may provide neuroprotection to the epileptic brain through trophic support. To test our hypothesis, we studied the temporal expression of neurotrophins after BMMC transplantation in the epileptic rat hippocampus. Methods Chronically epileptic rats were intravenously transplanted with 1 × 107 BMMCs isolated from GFP transgenic mice. Expression levels of BDNF, GDNF, NGF, VEGF, and TGF-β1, and their receptors, were evaluated by ELISA and/or qRT-PCR analysis. Results Our data revealed increased protein expression of BDNF, GDNF, NGF, and VEGF and reduced levels of TGF-β1 in the hippocampus of transplanted epileptic animals. Additionally, an increase in the mRNA expression of BDNF, GDNF, and VEGF, a reduction in TGF-β1, and a decrease in mRNA levels of the TrkA and TGFR-β1 receptors were also observed. Conclusion The gain provided by transplanted BMMCs in the epileptic brain may be related to the ability of these cells in modulating the network of neurotrophins and angiogenic signals.

Temporal lobe epilepsy (TLE) is a highly prevalent syndrome among people with epilepsy, and is usually refractory to drug treatment. Structural and physiological changes, such as hippocampal sclerosis, are often present in TLE patients. The objective of this study is to evaluate the feasibility and safety of intra-arterial infusion of autologous bone marrow mononuclear cells (BMMC) in adults with medically refractory mesial TLE (MTLE) and unilateral hippocampal sclerosis (HS). We enrolled 20 patients who had been diagnosed with MTLE-HS and were refractory to medical treatment. All patients underwent a neurological evaluation, magnetic resonance imaging with hippocampal volumetry, video-electroencephalography (EEG) with ictal recording, and a neuropsychological test battery focusing on verbal and nonverbal memory domains. After bone marrow aspiration and subsequent cell preparation, the BMMC were infused by selective posterior cerebral artery catheterization. Patients were followed for 6 months. Safety of the procedure, seizure frequency, neuropsychological evaluation, EEG variables, routine brain magnetic resonance imaging and hippocampal volumetry were considered measurements of outcome. Any serious intercurrent clinical event or adverse effects related to the procedure were reported. No additional lesions and no significant hippocampal volumetric changes were observed. EEG recordings showed a decrease in theta activity and spike density. At 6 months, eight patients (40%) were seizure free. A significant increase in the memory scores over time was observed. The BMMC autologous transplant for the treatment of temporal lobe epilepsy is feasible and safe. The seizure control achieved in this novel study supports the therapeutic potential of stem cell transplants in MTLE-HS patients. Copyright © 2016 John WileySons, Ltd.

Fish have been shown to act as sentinel organisms in genetic toxicology studies and the micronucleus (MN) test using fish erythrocytes has been successfully used to evaluate the genotoxic impact of environmental pollutants on aquatic environments. This study used the MN test of peripheral blood of specimens of a native fish species (Hyphessobrycon luetkenii) to evaluate the genotoxicity of the water of two unconnected reservoirs of the Canela National Forest in southern Brazil. Fish were caught in the reservoirs and MN frequencies in blood smears were determined. Baseline MN frequency of H. luetkenii was determined in the uncontaminated reservoir (reference site) and compared with the results obtained at two different sampling periods (winter and spring) in the reservoir which receives discharges of industrial and municipal wastes. In the spring season, MN frequency in the contaminated reservoir was about five times higher than in the reference site (0.51‰ and 0.10‰, respectively). The results of this stu...

AIMS: To evaluate the profile of men with cancer who performed semen cryopreservation prior/during treatment and address the importance of this method for reproductive health.METHODS: This was a transversal and retrospective study which used a database from a Reproductive Medicine Center located in Brazil. A total of 150 male patients who performed semen cryopreservation due to cancer diagnosis, from January 2014 to December 2017, were included.RESULTS: The profile of men seeking fertility preservation prior/during treatment for cancer was young adults, single, childless, with higher education. Oncologists were the ones who reported more patients for semen cryopreservation followed by urologists and hematologists. With regards to tumor diagnosis frequency, testicular was the most diagnosed, followed by Hodgkin’s/non-Hodgkin’s lymphoma, leukemia, prostate and rectal tumor, along with retroperitoneal tumor.CONCLUSION: Data brought the reflection on the cultural and financial barriers involved for the accomplishment of cryopreservation. Health professionals attending cancer patients should consider the importance of educational and incentive activities to prevent male fertility. Future research on the subject should carried out.

OBJECTIVE: We studied transplants of bone marrow mononuclear cells (BMMC) by lumbar puncture (LP) in a severe model of spinal cord injury (SCI) using clip compression. METHODS: BMMCs or saline solution were transplanted by LP 48 hours and 9 days post injury. Motor function was evaluated by BBB scale, histological analysis by Nissl technique and the verification of cell migration by PCR analysis. RESULTS: The BBB had significantly improved in rats treated with BMMCs by LP compared with controls (p

The antioxidant effects of the hydro-alcoholic guaraná extract (Paullinia cupana var. sorbilis Mart.) on nitric oxide (NO) and other compounds generated from the degradation of sodium nitroprusside (SNP) in an embryonic fibroblast culture (NIH-3T3 cells) were evaluated. The guaraná bioactive compounds were initially determined by high-performance liquid chromatography: caffeine=12.240mg/g, theobromine=6.733mg/g and total catechins=4.336mg/g. Cells were exposed to 10μM SNP during a 6h period because the cells exhibited >90% mortality at this concentration. Guaraná was added to the cultures in five concentrations (0.5, 1, 5, 10 and 20mg/mL). The guaraná antioxidant effect was evaluated by viability assays, biochemical oxidation [lipid peroxidation, catalase and superoxide dismutase (SOD) activity] and genotoxicity (DNA Comet assay) analysis. Additionally, oxidative stress was evaluated by a 2,7-dihydrodichlorofluorescein diacetate fluorescence assay. Guaraná reverted the SNP toxicity mainly at lower concentrations (

Objective: This study aimed to critically review the literature available regarding the Zika virus outbreak in Brazil and its possible association with microcephaly cases. Sources: Experts from Instituto do Cérebro do Rio Grande do Sul performed a critical (nonsystematic) literature review regarding different aspects of the Zika virus outbreak in Brazil, such as transmission, epidemiology, diagnostic criteria, and its possible association with the increase of microcephaly reports. The PubMed search using the key word “Zika virus” in February 2016 yielded 151 articles. The manuscripts were reviewed, as well as all publications/guidelines from the Brazilian Ministry of Health, World Health Organization and Centers for Disease Control and Prevention (CDC – United States). Summary of findings: Epidemiological data suggest a temporal association between the increased number of microcephaly notifications in Brazil and outbreak of Zika virus, primarily in the Brazil's Northeast. It has been previously documented that many different viruses might cause congenital acquired microcephaly. Still there is no consensus on the best curve to measure cephalic circumference, specifically in preterm neonates. Conflicting opinions regarding the diagnosis of microcephaly (below 2 or 3 standard deviations) that should be used for the notifications were also found in the literature. Conclusion: The development of diagnostic techniques that confirm a cause–effect association and studies regarding the physiopathology of the central nervous system impairment should be prioritized. It is also necessary to strictly define the criteria for the diagnosis of microcephaly to identify cases that should undergo an etiological investigation. Resumo: Objetivos: O objetivo deste estudo foi realizar uma revisão crítica da literatura sobre o surto de vírus Zika no Brasil e sua possível associação com casos de microcefalia. Fonte de dados: Especialistas em áreas afins do Instituto do Cérebro do Rio Grande do Sul realizaram uma revisão crítica (não sistemática) da literatura sobre o vírus Zika, suas formas de transmissão, a epidemia no Brasil, critérios diagnósticos e a possível associação com os casos crescentes de microcefalia. O uso da palavra chave “Zika virus” na base de dados do PubMed em fevereiro de 2016, retorna 151 publicações. Estes textos foram revisados assim como todas as publicações e recomendações do Ministério da Saúde, Organização Mundial da Saúde e Centro de Controle de Doenças (CDC – USA). Síntese dos dados: Os dados epidemiológicos sugerem uma relação temporal entre aumento da notificação de casos de microcefalia e o surto de vírus Zika, principalmente no Nordeste do Brasil. Agentes virais comprovadamente podem ser causadores de microcefalia congênita adquirida. Não existe um consenso sobre a melhor curva de perímetro cefálico a ser utilizada, principalmente nos prematuros. Assim como também existem divergências sobre a definição de microcefalia (abaixo de 2 ou 3 desvios padrões) a ser padronizada nas notificações. Conclusão: Deve-se priorizar o desenvolvimento de técnicas diagnósticas que confirmem uma relação causa–efeito e estudos sobre mecanismos da patogênese da infecção pelo Zika no sistema nervoso central. Também é necessário definir e universalizar os critérios diagnósticos para a identificação dos casos de microcefalia que devem ser submetidos à investigação etiológica. Keywords: Zika virus, Microcephaly, Pregnancy, Cortical development, Neuroimaging, Palavras-chave: Vírus Zika, Microcefalia, Gestação, Desenvolvimento cortical, Neuroimagem

In this study, we evaluated the cellular viability of various esthetic, metallic, and nickel-free orthodontic brackets.The sample was divided into 11 groups (n = 8): cellular control, negative control, positive control, metallic, polycarbonate, 2 types of monocrystalline ceramic, 3 types of nickel free, and polycrystalline ceramic brackets. Cell culture (NIH/3T3-mice fibroblasts) was added to the plates of 96 wells containing the specimens and incubated in 5% carbon dioxide at 37°C for 24 hours. Cytotoxicity was analyzed qualitatively and quantitatively. Cell growth was analyzed with an inverted light microscope, photomicrographs were obtained, and the results were recorded as response rates based on modifications of the parameters of Stanford according to the size of diffusion halo of toxic substances. Cell viability was analyzed (MTT assay); a microplate reader recorded the cell viability through the mitochondrial activity in a length of 570 nm. The values were statistically analyzed.All tested brackets had higher cytotoxicity values than did the negative control (P0.05), with the exception Rematitan and Equilibrium (both, Dentaurum, Ispringen, Germany) (P0.05), suggesting low toxicity effects. The values showed that only polycarbonate brackets were similar (P0.05) to the positive control, suggesting high toxicity.The brackets demonstrated different ranges of cytotoxicity; nickel-free brackets had better biocompatibility than the others. On the other hand, polycarbonate brackets were made of a highly cytotoxic material for the cells analyzed.

Despite the beneficial effects of cell-based therapies on brain repair shown in most studies, there has not been a consensus regarding the optimal dose of human umbilical cord blood cells (HUCBC) for neonatal hypoxia-ischemia (HI). In this study, we compared the long-term effects of intravenous administration of HUCBC at three different doses on spatial memory and brain morphological changes after HI in newborn Wistar rats. In addition, we tested whether the transplanted HUCBC migrate to the injured brain after transplantation. Seven-day-old animals underwent right carotid artery occlusion and were exposed to 8% O(2) inhalation for 2 h. After 24 h, randomly selected animals were assigned to four different experimental groups: HI rats administered with vehicle (HI+vehicle), HI rats treated with 1×10(6) (HI+low-dose), 1×10(7) (HI+medium-dose), and 1×10(8) (HI+high-dose) HUCBC into the jugular vein. A control group (sham-operated) was also included in this study. After 8 weeks of transplantation, spatial memory performance was assessed using the Morris water maze (MWM), and subsequently, the animals were euthanized for brain morphological analysis using stereological methods. In addition, we performed immunofluorescence and polymerase chain reaction (PCR) analyses to identify HUCBC in the rat brain 7 days after transplantation. The MWM test showed a significant spatial memory recovery at the highest HUCBC dose compared with HI+vehicle rats (P

article i nfo Article history: Received 25 January 2011 Accepted 3 June 2011 Aims: Epilepsy affects 0.5-1% of the world's population, and approximately a third of these patients are refractory to current medication. Given their ability to proliferate, differentiate and regenerate tissues, stem cells could restore neural circuits lost during the course of the disease and reestablish the physiological excitability of neurons. This study verified the therapeutic potential of bone marrow mononuclear cells (BMMCs) on seizure control and cognitive impairment caused by experimentally induced epilepsy. Main methods: Status epilepticus (SE) was induced by lithium-pilocarpine injection and controlled with diazepam 90 min after SE onset. Lithium-pilocarpine-treated rats were intravenously transplanted 22 days after SE with BMMCs obtained from enhanced green fluorescent protein (eGFP) transgenic C57BL/6 mice. Control epileptic animals were given an equivalent volume of saline or fibroblast injections. Animals were video-monitored for the presence of spontaneous recurrent seizures prior to and following the cell administration procedure. In addition, rats underwent cognitive evaluation using a Morris water maze. Key findings: Our data show that BMMCs reduced the frequency of seizures and improved the learning and long-term spatial memory impairments of epileptic rats. EGFP-positive cells were detected in the brains of transplanted animals by PCR analysis. Significance: The positive behavioral effects observed in our study indicate that BMMCs could represent a promising therapeutic option in the management of chronic temporal lobe epilepsy.

Human umbilical cord blood is an attractive source of stem cells; however, it has a heterogeneous cell population with few mesenchymal stem cells. Cell reprogramming induced by different methodologies can confer pluripotency to differentiated adult cells. The objective of this study was to evaluate the reprogramming of fibroblasts and their subsequent neural differentiation after co-culture with umbilical cord blood mononuclear cells. Cells were obtained from four human umbilical cords. The mononuclear cells were cultured for 7 d and subsequently co-cultured with mouse fibroblast NIH-3T3 cells for 6 d. The pluripotency of the cells was evaluated by RT-PCR using primers specific for pluripotency marker genes. The pluripotency was also confirmed by adipogenic and osteogenic differentiation. Neural differentiation of the reprogrammed cells was evaluated by immunofluorescence. All co-cultured cells showed adipogenic and osteogenic differentiation capacity. After co-cultivation, cells expressed the pluripotency gene KLF4. Statistically significant differences in cell area, diameter, optical density, and fractal dimension were observed by confocal microscopy in the neurally differentiated cells. Contact in the form of co-cultivation of fibroblasts with umbilical cord blood mononuclear fraction for 6 d promoted the reprogramming of these cells, allowing the later induction of neural differentiation.

Environmental pollution by pesticide residues is a major environmental concern due to the extensive use of these substances in agriculture. The insecticide beta-cyfluthrin is a synthetic pyrethroid widely used in agricultural and other domestic activities. The aim of the present study was to assess the genotoxic effects of a sublethal exposure of the fish Bryconamericus iheringii (Characidae) to a commercial formulation of beta-cyfluthrin using the comet assay. Fish were exposed to sublethal concentrations (4.2 and 5.6 microg/L) of beta-cyfluthrin under static conditions during 24- and 48-h exposure periods. Fish in tap water were used as negative controls. Results obtained by the comet assay revealed genotoxic effects of the pyrethroid in the higher concentration and at the longer exposure period. The mean DNA damage index of fish exposed to 5.6 microg/L beta-cyfluthrin for 48 h was significantly higher (145.9 +/- 51.8) than in the control group (69.3 +/- 39.5). These findings indicate that native fish species might be at risk for genotoxic damage in waters contaminated with beta-cyfluthrin.

As celulas mononucleares da medula ossea tem sido utilizadas em diversas afeccoes na tentativa de regeneracao tecidual. (O objetivo deste estudo foi o de avaliar a adesao das celulas mononucleares sobre defeito critico, com a adicao do plasma rico em plaquetas (PRP) e / ou TGF-β1 (Fator de crescimento transformador Beta 1), e, por fim, verificar o reparo osseo dos sitios defeituosos dos femures dos ratos. Foi criado um defeito bilateral critico nos femures de 33 Wistar-Kyoto ratos. Celulas mononucleares de medula ossea, TGF-β e PRP foram adicionadas no lado tratado da lesao e soro fisiologico no lado contralateral. Determinou-se a adesao de celulas mononucleares sobre o defeito critico e reparo osseo. A presenca e consequente adesao das celulas mononucleares administradas nos animais tratados nao foi evidenciada atraves da tecnica de PCR. As analises radiologicas evidenciam fechamento da lesao, porem, nos nao podemos afirmar que foi pelos tratamentos administrados ou pela propria regeneracao ossea, quando analisadas em seis e 10 semanas pos-operatorias, haja visto nao apresentarem diferencas significantes entre os grupos. Conclusoes: a) As celulas mononucleares nao aderiram ao defeito critico criado no femur do rato; b) Nao foi possivel avaliar a eficiencia dos tratamentos propostos para o reparo osseo, por nao apresentarem diferencas significativas entre os grupos.

Introduction Spinal cord injury occurs mainly in young healthy individuals, causing motor impairment and temporary or sensory loss and leading to chronic disability. An attractive possibility of treatment for TRM is the use of stem cells for restoration of neural cells in the spinal cord. Support Matrices for use in cell therapy must be able to provide structural and microenvironmental support to ensure the viability of the transplanted cells and to facilitate integration in the target tissue. Materials and Methods Biocompatible polymeric films composed of polylactic acid-co-acid (PLGA) polymer nanofiber polypyrrole (PPy) were used polypyrrole nanofibers and films made containing 10% (W/W) PLGA in PPy by solvent evaporation method were synthesized. The PPy nanofibers showed electrical conductivity of semiconductor and the dispersion of the nanofibers in PLGA resulted in PPy films with fibrous topographical morphology with suitable for use as scaffolds for cell growth driven by PPy nanofiber voids. Cell toxicity test method of extraction was performed. Samples of the polymers were maintained in DMEM culture medium obeying the proportions established by ISO 10993 (2013) by 24, 48, 72 hours, 7 and 14 days. After each time period, NIH3T3 fibroblasts were cultured for 24 hours with the film exposed to the environment, and then was held cellular toxicity test by MTT assay. Wistar rats were anesthetized and underwent laminectomy of the eighth thoracic vertebra. On the local display of the spinal cord was placed a cut polymer approximately 3 mm wide and 8 mm long. The motor recovery of the animals 24 hours and 7, 14, and 28 days after injury was assessed. To assess the degree of inflammation generated by the polymer in tissues other than the spinal cord, a 1 cm incision in the flank and placement polymer was performed. In the region against lateral incision was performed only. Histological evaluation of the spinal cord and the fabric flank was held. Results The electrical conductivity of PPy fibers showed values of 8.10 to 4 S/cm, lying within the range of semiconductors. The molecular structure of PPy was evaluated in the range of 400 to 4,000/cm and all spectra were similar. There was no change in cell viability after exposure of the cells to the conditioned medium with the polymer in both seasons. Cell viability was maintained at 100% when compared with the control. The motor functional assessment of mice showed no alteration in any evaluation period are because of the surgical procedure or a possible effect of inflammatory irritation of the polymer on the spinal nerve tissue. Histological evaluation showed no changes suggestive of inflammation or infection of the marrow tissue adjacent to the polymer or the flank of the animal tissues. Conclusion PLGA/PPy polymer has conductivity expected for semiconductors and structural heterogeneity, without any type of cell toxicity in periods. The polymer is safe to be used as a scaffold in the TRM model because it does not cause any functional, structural, or inflammatory change.

Introduction Experimental studies of the spinal cord before and after injury caused by trauma are necessary to evaluate the new therapeutic options, such as stem cell therapy. In those experiments, it is mandatory to measure motor and sensitive function adequately to the bias and increase true results. The objective is to describe the technique used to obtain the compound and extracellular action potentials in the spinal cord. Materials and Methods To obtain the spinal cord compound action potentials (CAPs), the spinal cord of the rat spine was immediately placed in a Perspex chamber covered with Sylgard and superfused at (2 mL/min) with Krebs solution and glucose. The solution was maintained at pH 7.2 and 5% CO2/95% O2. The Ag/AgCl needle electrodes fixed the tissue in the chamber. The needle electrodes were connected to an operational amplifier and to a programmable signal conditioner model. The electrical stimulation was done by suction bipolar electrode placed in the inferior segment of the spinal cord. The electrode was connected to an analogical stimulator, which delivered stimulus at 01 Hz for a 0.2 milliseconds duration. The data always began to be recorded after 15 minutes of stimulation to ensure the stability of the preparation. The recordings of the CAPs were made during 30 minutes directly to an AxoScope software. The events were analyzed by the Clampfit software. To acquire extracellular action potential, the spinal cord was cooled in Krebs solution, previously gassed with a 95% O2 and 5% CO2 mixture to obtain a pH value of 7.2 to 7.4. A 400 µm thick coronal section was made. The spinal cord slice was recovered during at least 30 minutes at 32°C in Krebs solution and transferred to a submersion-type recording chamber with Krebs solution, 95% O2 and 5% CO2. Extracellular recordings were made using 5 to 10 MΩ resistance glass pipettes. An iridium stimulation electrode was connected to an analogical stimulator and positioned at the proximal side of the tissue to deliver electrical stimuli (0.1 Hz, 0.2 ms). The recording electrode was placed in the opposite or distal side. The recordings were performed using an amplifier connected to a programmable signal conditioner. The experiments were recorded during 1 hour. The data were retrieved by an AxoScope software and analyzed by the Clampfit software. Results The compound and extracellular action potential could be generated in both experiments indicating that the functional analysis of spinal cord neurons can be assessed by in vitro stimulation of the spinal cord tissue or its slice. The control values for the amplitude of CAPs was 4.4 ± 0.3 mV ( n = 3). The rise time of the CAPs was 1.73 ± 03 ms; the decay time was 4.2 ± 2 ms, and the duration was 4.2 ± 0.8 ms. The analysis of the extracellular action potentials showed that the mean amplitude, rise time, decay time, and duration were 2.17 ± 0.3 mV, 0.4 ± 0.2, 4 ± 1, and 2.1 ± 0.2 ms, respectively. Conclusion The analysis of compound and extracellular action potential can be used as outcome measurement of spinal cord functionality. This can measure more precisely, for example, the neuronal repopulation and motor and sensitive recovery of spinal cord injury.

In this study, we analysed the genotoxic and cytotoxic effects of treated tannery wastewater using the micronucleus (MN) test in fish erythrocytes (Bryconamericus iheringii) and the Allium cepa test system. Fish were exposed to three different concentrations of tannery effluent (2, 5 and 10%) for 3 and 6 days. Onion bulbs were exposed to four dilutions of the effluent (2, 5, 10 and 25%) for 60 hours. The results indicated absence of genotoxicity in the treated tannery effluent; however, toxic compounds were identified, threatening the survival of fish species at the effluent discharge site.