GC-MS with Headspace Extraction for Non-Invasive Diagnostics of IBD Dynamics in a Model of DSS-Induced Colitis in Rats.

Inflammatory bowel diseases are extremely common throughout the world. However, in most cases, it is asymptomatic at the initial stage. Therefore, it is important to develop non-invasive diagnostic methods that allow identification of the IBD risks in a timely manner. It is well known that gastrointestinal microbiota secrete volatile compounds (VOCs) and their composition may change in IBD. We propose a non-invasive method to identify the dynamics of IBD development in the acute and remission stage at the level of VOCs in model of dextran sulfate sodium (DSS) with chemically induced colitis measured by headspace GC/MS (HS GC/MS). Methods: VOCs profile was identified using a headspace GC/MS (HS GC/MS). GC/MS data were processed using MetaboAnalyst 5.0 and GraphPad Prism 8.0.1 software. The disease activity index (DAI) and histological method were used to assess intestinal inflammation. The peak of intestinal inflammation activity was reached on day 7, according to the disease activity index. Histological examination data showed changes in the intestine due to different stages of inflammation. As the acute inflammation stage was reached, the metabolomic profile also underwent changes, especially at the short-fatty acids level. A higher relative amounts of acetic acid (p value < 0.025) and lower relative amounts of propanoic acid (p value < 0.0005), butanoic acid (p value < 0.005) and phenol 4-methyl- (p value = 0.053) were observed in DSS7 group on day 7 compared to the control group. In remission stage, disease activity indexes decreased, and the histological picture also improved. But metabolome changes continued despite the withdrawal of the DSS examination. A lower relative amounts of propanoic acid (p value < 0.025), butanoic acid (p value < 0.0005), pentanoic acid (p value < 0.0005), and a significant de-crease of hexanoic acid (p value < 0.0005) relative amounts were observed in the DSS14 group compared to the control group on day 14. A model of DSS-induced colitis in rats was successfully implemented for metabolomic assessment of different stages of inflammation. We demonstrated that the ratios of volatile compounds change in response to DSS before the appearance of standard signs of inflammation, determined by DAI and histological examination. Changes in the volatile metabolome persisted even after visual intestine repair and it confirms the high sensitivity of the microbiota to the damaging effects of DSS. The use of HS GC/MS may be an important addition to existing methods for assessing inflammation at early stages.

Alterations of the Composition and Neurometabolic Profile of Human Gut Microbiota in Major Depressive Disorder.

Major depressive disorder (MDD) is among the most prevalent mental disorders worldwide. Factors causing the pathogenesis of MDD include gut microbiota (GM), which interacts with the host through the gut-brain axis. In previous studies of GM in MDD patients, 16S rRNA sequencing was used, which provided information about composition but not about function. In our study, we analyzed whole metagenome sequencing data to assess changes in both the composition and functional profile of GM. We looked at the GM of 36 MDD patients, compared with that of 38 healthy volunteers. Comparative taxonomic analysis showed decreased abundances of Faecalibacterium prausnitzii, Roseburia hominis, and Roseburia intestinalis, and elevated abundances of Escherichia coli and Ruthenibacterium lactatiformans in the GM of MDD patients. We observed decreased levels of bacterial genes encoding key enzymes involved in the production of arginine, asparagine, glutamate, glutamine, melatonin, acetic, butyric and conjugated linoleic acids, and spermidine in MDD patients. These genes produced signature pairs with Faecalibacterium prausntizii and correlated with decreased levels of this species in the GM of MDD patients. These results show the potential impact of the identified biomarker bacteria and their metabolites on the pathogenesis of MDD, and should be confirmed in future metabolomic studies.

Peptide LCGA-17 Attenuates Behavioral and Neurochemical Deficits in Rodent Models of PTSD and Depression.

We have previously described the LCGA-17 peptide as a novel anxiolytic and antidepressant candidate that acts through the α2δ VGCC (voltage-gated calcium channel) subunit with putative synergism with GABA-A receptors. The current study tested the potential efficacy of acute and chronic intranasal (i.n.) LCGA-17 (0.05 mg/kg and 0.5 mg/kg) in rats on predator odor-induced conditioned place aversion (POCPA), a model of post-traumatic stress disorder (PTSD), and chronic unpredictable stress (CUS) that produce a range of behavioral and physiological changes that parallel symptoms of depression in humans. CUS and LCGA-17 treatment effects were tested in the sucrose preference (SPT) social interaction (SI), female urine sniffing (FUST), novelty-suppressed feeding (NSFT), and forced swim (FST) tests. Analysis of the catecholamines content in brain structures after CUS was carried out using HPLC. The efficacy of i.n. LCGA-17 was also assessed using the Elevated plus-maze (EPM) and FST. Acute LCGA-17 administration showed anxiolytic and antidepressant effects in EPM and FST, similar to diazepam and ketamine, respectively. In the POCPA study, LCGA-17 significantly reduced place aversion, with efficacy greater than doxazosin. After CUS, chronic LCGA-17 administration reversed stress-induced alterations in numerous behavioral tests (SI, FUST, SPT, and FST), producing significant anxiolytic and antidepressant effects. Finally, LCGA-17 restored the norepinephrine levels in the hippocampus following stress. Together, these results support the further development of the LCGA-17 peptide as a rapid-acting anxiolytic and antidepressant.

The Dynamics of ß-Amyloid Proteoforms Accumulation in the Brain of a 5xFAD Mouse Model of Alzheimer's Disease.

Alzheimer's disease (AD) is the leading cause of dementia among the elderly. Neuropathologically, AD is characterized by the deposition of a 39- to 42-amino acid long ß-amyloid (Aß) peptide in the form of senile plaques. Several post-translational modifications (PTMs) in the N-terminal domain have been shown to increase the aggregation and cytotoxicity of Aß, and specific Aß proteoforms (e.g., Aß with isomerized D7 (isoD7-Aß)) are abundant in the senile plaques of AD patients. Animal models are indispensable tools for the study of disease pathogenesis, as well as preclinical testing. In the presented work, the accumulation dynamics of Aß proteoforms in the brain of one of the most widely used amyloid-based mouse models (the 5xFAD line) was monitored. Mass spectrometry (MS) approaches, based on ion mobility separation and the characteristic fragment ion formation, were applied. The results indicated a gradual increase in the Aß fraction of isoD7-Aß, starting from approximately 8% at 7 months to approximately 30% by 23 months of age. Other specific PTMs, in particular, pyroglutamylation, deamidation, and oxidation, as well as phosphorylation, were also monitored. The results for mice of different ages demonstrated that the accumulation of Aß proteoforms correlate with the formation of Aß deposits. Although the mouse model cannot be a complete analogue of the processes occurring in the human brain in AD, and several of the observed parameters differ significantly from human values supposedly due to the limited lifespan of the model animals, this dynamic study provides evidence on at least one of the possible mechanisms that can trigger amyloidosis in AD, i.e., the hypothesis on the relationship between the accumulation of isoD7-Aß and the progression of AD-like pathology.

Plasma Neurotrophic Factor Levels are not Associated with the Severity of Depression: Prospective Pilot Study.

INTRODUCTION: Depression is one of the most common mental illnesses. Impaired neurogenesis is observed in depression. Biomarkers of impaired neurogenesis in depression can act as a useful objective and diagnostic and prognostic tool to determine the severity of depression. AIM: To study the concentration of biochemical indicators in the blood that may be involved in the pathogenesis of depression and their intercorrelations, and to determine any associations between the concentrations of biochemical indicators and severity of depressive symptoms. METHODS: We determined the plasma concentrations of serotonin, dopamine, and neurotrophic factors involved in neurogenesis (BDNF, CDNF and neuropeptide Y) using enzyme immunoassay and mass spectrometry in depressed patients (n=22) and healthy controls (n=16) matched by socio-demographic parameters. All participants were assessed using the Hamilton Depression Scale (HAMD), the Generalized Anxiety Disorder Questionnaire (GAD-7), and the Center for Epidemiologic Studies Depression Scale (CES-D) to enter the study. The standard cut-offs for the CES-D and GAD-7 scales were used to confirm the presence or absence of depression and anxiety. RESULTS: The concentrations of serotonin, dopamine, BDNF, CDNF, and neuropeptide Y in plasma did not differ between the groups and was not found to be associated with the scores on the scales. Positive correlations were found between the concentration of neuropeptide Y and serotonin, BDNF, and CDNF in blood plasma. CONCLUSIONS: Plasma concentrations of biomarkers related to the pathophysiology of depression did not correlate with the severity of its symptoms.

Bacterial Metabolites of Human Gut Microbiota Correlating with Depression.

Depression is a global threat to mental health that affects around 264 million people worldwide. Despite the considerable evolution in our understanding of the pathophysiology of depression, no reliable biomarkers that have contributed to objective diagnoses and clinical therapy currently exist. The discovery of the microbiota-gut-brain axis induced scientists to study the role of gut microbiota (GM) in the pathogenesis of depression. Over the last decade, many of studies were conducted in this field. The productions of metabolites and compounds with neuroactive and immunomodulatory properties among mechanisms such as the mediating effects of the GM on the brain, have been identified. This comprehensive review was focused on low molecular weight compounds implicated in depression as potential products of the GM. The other possible mechanisms of GM involvement in depression were presented, as well as changes in the composition of the microbiota of patients with depression. In conclusion, the therapeutic potential of functional foods and psychobiotics in relieving depression were considered. The described biomarkers associated with GM could potentially enhance the diagnostic criteria for depressive disorders in clinical practice and represent a potential future diagnostic tool based on metagenomic technologies for assessing the development of depressive disorders.

Copy Number Variation of Satellite III (1q12) in Patients With Schizophrenia.

Introduction: It was shown that copy number variations (CNVs) of human satellite III (1q12) fragment (f-SatIII) reflects the human cells response to stress of different nature and intensity. Patients with schizophrenia (SZ) experience chronic stress. The major research question: What is the f-SatIII CNVs in human leukocyte as a function of SZ? Materials and Methods: Biotinylated pUC1.77 probe was used for f-SatIII quantitation in leukocyte DNA by the non-radioactive quantitative hybridization for SZ patients (N = 840) and healthy control (HC, N = 401). SZ-sample included four groups. Two groups: first-episode drug-naïve patients [SZ (M-)] and medicated patients [SZ (M+)]. The medical history of these patients did not contain reliable confirmed information about fetal hypoxia and obstetric complications (H/OCs). Two other groups: medicated patients with documented H/OCs [hypoxia group (H-SZ (M+)] and medicated patients with documented absence of H/OCs [non-hypoxia group (NH-SZ (M+)]. The content of f-SatIII was also determined in eight post-mortem brain tissues of one SZ patient. Results: f-SatIII in human leukocyte varies between 5.7 to 44 pg/ng DNA. f-SatIII CNVs in SZ patients depends on the patient's history of H/OCs. f-SatIII CN in NH-SZ (M+)-group was significantly reduced compared to H-SZ (M+)-group and HC-group (p < 10-30). f-SatIII CN in SZ patients negatively correlated with the index reflecting the seriousness of the disease (Positive and Negative Syndrome Scale). Antipsychotic therapy increases f-SatIII CN in the untreated SZ patients with a low content of the repeat and reduces the f-SatIII CN in SZ patients with high content of the repeat. In general, the SZ (M+) and SZ (M-) groups do not differ in the content of f-SatIII, but significantly differ from the HC-group by lower values of the repeat content. f-SatIII CN in the eight regions of the brain of the SZ patient varies significantly. Conclusion: The content of f-SatIII repeat in leukocytes of the most patients with SZ is significantly reduced compared to the HC. Two hypotheses were put forward: (1) the low content of the repeat is a genetic feature of SZ; and/or (2) the genomes of the SZ patients respond to chronic oxidative stress reducing the repeats copies number.

Diffusion Tensor Imaging in Diagnosis of Post-Traumatic Syringomyelia in Spinal Cord Injury in Rats.

BACKGROUND Post-traumatic syringomyelia (PTS) is a common disease after spinal cord injury (SCI). The present study was performed to evaluate the advantages of diffusion tensor imaging (DTI) in estimating SCI and prognosing PTS in SCI rats. MATERIAL AND METHODS Forty rats were divided into 3 groups based on the extent of the individual SCI and PTS: a control group (n=10), a PTS group (n=8), and an SCI group (n=22). BBB tests were performed preoperatively and postoperatively at (1 d, 3 d, 5 d, 1 w, 2 w, 1 w, 2 w, 3 w, 4 w, 5 w, and 6 w). MRI T2 scanning was conducted postoperatively at (1 w, 2 w, 3 w, 4 w, 5 w, 6 w). DTI and diffusion tensor tractography were used for analyzing neuro-fiber changes after SCI. RESULTS BBB scoring showed no differences between the PTS group and SCI group (P<0.05). PTS was found in 8 rats after SCI. MRI showed PTS formation in 3 rats at 2 w after SCI, and 5 rats showed PTS formation at postoperative 3w after SCI. Compared with the control group, ADC showed significant increase in both the PTS group (P<0.05) and the SCI group (P<0.05), FA showed significant decreases in the PTS (P<0.05) and SCI (P<0.05) groups. Compared with the SCI group, the PTS group showed an increase in ADC, but no statistical difference was found in ADC (P>0.05). The PTS group showed a significant increase in FA (P<0.05). CONCLUSIONS The combination of diffusion tensor imaging and diffusion tensor tractography has characteristics of high-sensitivity and quantitation for PTS prognosis. FA is predictive in the prognosis of PTS formation after SCI.

Prenatal exposure to brain-specific anion transporter-1-specific monoclonal antibodies impairs cognitive function in post-natal life.

Subclinical hypothyroidism is caused by thyroid hormone deficit and can lead to impairments in mood and cognition. In brain, supply with thyroxine (T4) is mediated by thyroid hormone transporters including the brain-specific anion transporter-1 (BSAT-1). In humans and rodents, BSAT-1 is expressed in brain microvessels and astrocytes. In this study, we tested whether exposure in utero with BSAT-1-specific monoclonal antibodies (MabBSAT) will affect the cognitive function of the progeny. On gestation day 16th, females were intravenously treated with MabBSAT, non-specific antibodies (control 1), and saline (control 2). 72h after injection, MabBSAT were still detectable in the rat brain while non-specific antibodies were found. Immunocytochemistry showed that MabBSAT can bind to cultured primary cerebrovascular rat cells. At the age of 1month, the progeny was subjected to the Y-maze test, novel object recognition test, passive avoidance test, and Morris water maze, which revealed significant impairments in the cognitive function in the MabBSAT-exposed progeny compared to both control progeny groups. Therefore, prenatal exposure to MabBSAT blocks brain BSAT-1 and limits T4 influx to the brain. This impairs the cognitive function in exposed progeny in the post-natal life.

Precise Delivery Into Chronic Spinal Cord Injury Syringomyelic Cysts with Magnetic Nanoparticles MRI Visualization.

BACKGROUND: Traumatic spinal cord injury (SCI) often results in the deficiency of glia and neurons in cystic cavities. These syringomyelic cysts can prevent axonal regeneration and sprouting. Details of the mechanism of syringomyelic cyst formation are unknown and an effective treatment for overcoming syringomyelic cysts is not available. MATERIAL AND METHODS: Ten adult female Wistar rats underwent contusion SCI modeling resulting in syringomyelic cyst formation. A novel method for locating the cysts was developed and employed. MRI safe silver needles were inserted through the erector spinae of anesthetized rats to create a stable reference point. MRI images of the rodent spine were taken with the needles in situ. This information was used to accurately locate the cyst and determine the 3-dimensional entry point coordinates for nanoparticle delivery. Nanoparticles were injected into the cyst during a primary injection of 8 ul and a secondary injection of 8 ul, to prove the procedure can be accurately repeated. RESULTS: None of the rats died intra- or post-operatively. The syringomyelic cysts were accurately located with the 3-dimensional entry point coordinates. After nanoparticle delivery twice into each rat, the visualized syringomyelic cyst volume significantly decreased from 5.71±0.21 mm3 to 3.23±0.364 mm3 and to 1.48±0.722 mm3. CONCLUSIONS: The present study describes a novel strategy for precise nanoparticle delivery into a syringomyelic cyst, using measurements obtained from MRI images. This strategy may aid in developing a new method for studying chronic spinal cord injury and a novel treatment for syringomyelic cysts.