Physiol. Res. 68 (Suppl. 3): S315-S323, 2019 https://doi.org/10.33549/physiolres.934350 Plasma Levels of Glial Cell Marker S100B in Children With Autism A. TOMOVA1, P. KEMÉNYOVÁ1, D. FILČÍKOVÁ1, Ž. SZAPUOVÁ1, A. KOVÁČ2, K. BABINSKÁ1, D. OSTATNÍKOVÁ1 1Institute of Physiology, Faculty of Medicine, Comenius University in Bratislava, Bratislava, Slovak Republic, 2Slovak Academy of Sciences, Institute of Neuroimmunology, Bratislava, Slovak Republic Received March 21, 2019 Accepted September 26, 2019 Summary interaction in the presence of restricted, repetitive Autism spectrum disorder (ASD) is a neurodevelopmental behaviors or interests. ASD is of expanding worldwide condition with increasing incidence. Recent evidences suggest medical and social concern because of its increasing glial cells involvement in autism pathophysiology. S100B is incidence (Onaolapo and Onaolapo 2017, Baio et al. a calcium binding protein, mainly found in astrocytes and 2018). Despite recent scientific interest, etiology and therefore used as a marker of their activity. In our study, children pathogenesis of ASD remain unclear what obstructs the with autism had higher plasma concentrations of S100B prevention, diagnostics and treatment of people with compared to non-autistic controls. No association of S100B these disorders. According to current hypotheses, plasma levels with behavioral symptoms (ADI-R and ADOS-2 etiology of ASD involves multiple mechanisms, scales) was found. Plasma S100B concentration significantly including genetic predisposition and environmental correlated with urine serotonin, suggesting their interconnection. factors. Their combination may lead to alterations in Correlation of plasma S100B levels with stool calprotectin epigenetic gene regulation, with consequences including concentrations was found, suggesting not only brain astrocytes, impairments in neuron-glia interactions (Zeidan-Chulia but also enteric glial cells may take part in autism pathogenesis. et al. 2014). Mounting evidence suggests the role of Based on our findings, S100B seems to have a potential to be neuroglial cells in autism, as well as in other used as a biomarker of human neurodevelopmental disorders, neurological, neurodevelopmental and psychiatric but more investigations are needed to clarify its exact role in disorders. Neuroglia described in 1858 by Virchow is pathomechanism of autism. represented by macroglia and microglia. The role of microglia is characterized in several reviews (Koyama Key words and Ikegaya 2015, Tay et al. 2017, Anderson and Vetter S100B • Autism • Serotonin 2019). Astrocytes, representatives of macroglia, are involved in neurogenesis, development and maintenance Corresponding author of homeostasis in the brain, neurotransmitter signaling, A. Tomova, Institute of Physiology, Faculty of Medicine, neuronal activity and mitochondrial functions in neural Comenius University in Bratislava, Bratislava, Slovakia, cells (Zeidan-Chulia et al. 2014). Since ASD is Sasinkova 2, 813 72 Bratislava, Slovak Republic. E-mail: characterized by deviations in brain development, [email protected] neurogenesis, synaptopathy and neuroinflammation (Gilbert and Man 2017), investigation of astrocytes and Introduction their products seems to be a reasonable and promising objective within surveys aimed at elucidating the autism ASD (Autism spectrum disorder) is brain-based pathogenesis. neurodevelopmental condition characterized by S100B is one of markers of astrocyte activation. impairments in social communication and social It is a Ca2+ (calcium)-binding protein, concentrated PHYSIOLOGICAL RESEARCH • ISSN 0862-8408 (print) • ISSN 1802-9973 (online) 2019 Institute of Physiology of the Czech Academy of Sciences, Prague, Czech Republic Fax +420 241 062 164, e-mail: [email protected], www.biomed.cas.cz/physiolres S316 Tomova et al. Vol. 68 mainly in intracellular compartment and found et al. 2008). This molecule is also actively investigated in extracellularly when actively released or in case of the connection to ASD. Evidence suggests that increased leakage from damaged cells. The intracellular role of or decreased serotonin level plays a critical role S100B includes participation in cell proliferation, in neurodevelopment (Garbarino et al. 2019). The differentiation, transcription, survival, enzyme activities. involvement of serotonin in ASD pathomechanisms is Its extracellular role is exerted mostly through receptors widely supported by findings that behavioral symptoms for advanced glycation end-products (RAGE) and of autism, including deficits in social behavior (Kiser triggering intracellular signaling cascade, that leads to et al. 2012) and restrictive repetitive behaviors processes such as neuroinflammation and neurode- (Hollander et al. 2005), are modulated by 5-HT signaling. generation (for review see Sorci et al. 2013, Michetti Some studies showed increased blood levels of serotonin et al. 2019). Interestingly, through this activation S100B in children with autism (Abdulamir et al. 2018), but other has been shown to regulate microglia activation and studies reported this might be a result of high level of migration, as well as to eventually cause neuronal death hyperserotonemia not in all, but in approx. 25 % of (Bianchi et al. 2011). people with ASD (Gabriele et al. 2014, Marler et al. Since S100B is released from damaged 2016). At the same time, serotonin is a neurotransmitter astrocytes, its increased extracellular concentration might that is produced in large quantities in the gastrointestinal be used as a biomarker of acute brain injury, neural tract (Jameson and Hsiao 2018). We choose to study distress, and, possibly, neurodegenerative disorders. serotonin levels in urine not only because it is a non- Moreover, it is described that S100B biological activity is invasive procedure, but 5-HT levels in plasma and urine associated with its concentration (Michetti et al. 2019). are shown to correlate (Audhya et al. 2012). There is However, for interpretation of S100B role in even a study, reporting that hyperserotonemia increases pathophysiology it is important to remember, that in 5-HT level in urine in individuals autism (Mulder et al. neural cultures it demonstrated neurotrophic effects at 2010). nanomolar levels, while apoptotic effects at micromolar In the presented study, we investigated serotonin levels (Sorci et al. 2013, Michetti et al. 2019). For levels in connection to S100B, since according to example, in nanomolar concentrations S100B has been scientific evidence their actions seem to be shown to exert neurotrophic and neuroprotective effects interconnected. Most of serotonin effects are mediated on serotonergic neurons (Eriksen and Druse 2001). For through the release of S100B (Shapiro et al. 2010). last decades, S100B has been shown to be a reliable The level of S100B in the periphery comprises biomarker of neural injury (Astrand and Unden 2019, its release from glial cells in the brain and in the intestinal Michetti et al. 2019). It is actively investigated as tract. Enteric glial cells (EGCs) are responding to gut a possible biomarker in such disorders as Alzheimer’s inflammation (Langness et al. 2017), which can be disease, Parkinson’s disease, Schizophrenia, or mood approximated by measuring fecal calprotectin levels. disorders (for review see Michetti et al. (2019)) and Calprotectin is routinely used as a validated non-invasive others. However, if is it possible to use it as a marker in marker of intestinal inflammation (Alibrahim et al. 2015). patients with neurodevelopmental disorders, including In inflammatory bowel disease it is used to evaluate the autism, remains unclear. The main goal of our study was severity of disease and the response to treatment to investigate the plasma concentrations of Ca2+ binding (Erbayrak et al. 2009, Alibrahim et al. 2015). It can be protein S100B, as a biomarker of the status of astrocytes used also as a marker of gastrointestinal status in children in central nervous system and glial cells in enteric with autism (Babinska et al. 2017). nervous system in children with and without autism, and its possible correlation with their behavior. Methods Additionally, in attempt to elucidate the mechanism of autism, in a randomly selected fraction of Subjects children with autism (n=30) urine was analyzed for The study included 93 children diagnosed with serotonin levels. Serotonin (5-hydroxytryptamin, 5-HT) autism and 35 non-autistic children. The age of children is a neurotransmitter, well known for its pleiotropic with autism was 6.22±0.30 years (mean ± SEM), ranging effects, including role in the central nervous system and from 2 to 16 years, and matching control group of behavior (Frazer and Hensler 1999, Mohammad-Zadeh children aged 6.99±0.59 years (mean ± SEM), ranging 2019 Plasma S100B in Children With Autism in Slovakia S317 from 2 to 12 years. All children were boys. Children were laboratory within 3 h, where frozen at -20 °C until recruited from the Academic Research Center for Autism analysis. Serotonin levels were measured by (ARCA), based at the Institute of Physiology, Faculty of UHPLC/MS/MS method using the AbsoluteIDQ® p180 Medicine Comenius University in Bratislava. The kit (Biocrates Life Sciences AG, Innsbruck, Austria), diagnosis of ASD was performed meeting criteria for UPLC-Xevo TQS. Sample preparation and measurements DSM-V (The Diagnostic and Statistical Manual of were carried out according to the manufacturer’s protocol Mental Disorders, Fifth Edition), using