Phenols and GABAA receptors: from structure and molecular mechanisms action to neuropsychiatric sequelae

γ-Aminobutyric acid type A receptors (GABAARs) are members of the pentameric ligand-gated ion channel (pLGIC) family, which are widespread throughout the invertebrate and vertebrate central nervous system. GABAARs are engaged in short-term changes of the neuronal concentrations of chloride (Cl−) and bicarbonate (HCO3−) ions by their passive permeability through the ion channel pore. GABAARs are regulated by various structurally diverse phenolic substances ranging from simple phenols to complex polyphenols. The wide chemical and structural variability of phenols suggest similar and different binding sites on GABAARs, allowing them to manifest themselves as activators, inhibitors, or allosteric ligands of GABAAR function. Interest in phenols is associated with their great potential for GABAAR modulation, but also with their subsequent negative or positive role in neurological and psychiatric disorders. This review focuses on the GABAergic deficit hypotheses during neurological and psychiatric disorders induced by various phenols. We summarize the structure–activity relationship of general phenol groups concerning their differential roles in the manifestation of neuropsychiatric symptoms. We describe and analyze the role of GABAAR subunits in manifesting various neuropathologies and the molecular mechanisms underlying their modulation by phenols. Finally, we discuss how phenol drugs can modulate GABAAR activity via desensitization and resensitization. We also demonstrate a novel pharmacological approach to treat neuropsychiatric disorders via regulation of receptor phosphorylation/dephosphorylation

Transcriptomic changes triggered by ouabain in rat cerebellum granule cells: Role of α3- and α1-Na+,K+-ATPase-mediated signaling

It was shown previously that inhibition of the ubiquitous α1 isoform of Na+,K+-ATPase by ouabain sharply affects gene expression profile via elevation of intracellular [Na+]i/[K+]i ratio. Unlike other cells, neurons are abundant in the α3 isoform of Na+,K+-ATPase, whose affinity in rodents to ouabain is 104-fold higher compared to the α1 isoform. With these sharp differences in mind, we compared transcriptomic changes in rat cerebellum granule cells triggered by inhibition of α1- and α3-Na+,K+-ATPase isoforms. Inhibition of α1- and α3-Na+,K+-ATPase isoforms by 1 mM ouabain resulted in dissipation of transmembrane Na+ and K+ gradients and differential expression of 994 transcripts, whereas selective inhibition of α3-Na+,K+-ATPase isoform by 100 nM ouabain affected expression of 144 transcripts without any impact on the [Na+]i/[K+]i ratio. The list of genes whose expression was affected by 1 mM ouabain by more than 2-fold was abundant in intermediates of intracellular signaling and transcription regulators, including augmented content of Npas4, Fos, Junb, Atf3, and Klf4 mRNAs, whose upregulated expression was demonstrated in neurons subjected to electrical and glutamatergic stimulation. The role [Na+]i/[K+]i-mediated signaling in transcriptomic changes involved in memory formation and storage should be examined further

Antifibrotic and regenerative effects of Treamid in pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive disease characterized by interstitial fibrosis and progressive respiratory failure. Pirfenidone and nintedanib slow down but do not stop the progression of IPF. Thus, new compounds with high antifibrotic activity and simultaneously regenerative activity are an unmet clinical need. Recently, we showed that Treamid can help restoring the pancreas and testicular tissue in mice with metabolic disorders. We hypothesized that Treamid may be effective in anti-fibrotic therapy and regeneration of damaged lung tissue in pulmonary fibrosis. In this study, experiments were performed on male C57BL/6 mice with bleomycin-induced pulmonary fibrosis. We applied histological and immunohistochemical methods, ELISA, and assessed the expression of markers of endothelial and epithelial cells in primary cultures of CD31+ and CD326+ lung cells. Finally, we evaluated esterase activity and apoptosis of lung cells in vitro. Our data indicate that Treamid exhibits antifibrotic activity in mice with pulmonary fibrosis and has a positive effect on capillaries of the lungs. Treamid also increases the number of endothelial progenitor cells in the lungs of animals with pulmonary fibrosis. Lastly, Treamid increases esterase activity and decreases apoptosis of CD31+ lung cells in vitro. Based on these findings, we suggest that Treamid may represent a promising compound for the development of new antifibrotic agents, which are capable of stimulating regeneration of lung endothelium in IPF patients

Cancer stem cells and somatic stem cells as potential new drug targets, prognosis markers, and therapy efficacy predictors in breast cancer treatment

New drug targets, markers of disease prognosis, and more efficient treatment options are an unmet clinical need in breast cancer (BC). We have conducted a pilot study including patients with luminal B stage breast cancer IIA-IIIB. Presence and frequency of various populations of cancer stem cells (CSC) and somatic stem cells were assessed in the blood, breast tumor tissue, and normal breast tissue. Our results suggest that patients with BC can be divided into two distinct groups based on the frequency of aldehyde dehydrogenase positive cells (ALDH1+ cells) in the blood (ALDH1hi and ALDH1ow). In the ALDH1hi cells group, the tumor is dominated by epithelial tumor cells CD44+CD24low, CD326+CD44+CD24‒ and CD326‒CD49f+, while in the ALDH1low cells group, CSCs of mesenchymal origin and epithelial tumor cells (CD227+CD44+CD24‒ and CD44+CD24‒CD49f+) are predominant. In vitro CSCs of the ALDH1low cells group expressing CD326 showed high resistance to cytostatics, CD227+ CSCs of the ALDH1hi cells group are sensitive to cytostatics. Epithelial precursors of healthy mammary gland were revealed in normal breast tissue of patients with BC from both groups. The cells were associated with a positive effect of chemotherapy and remission in BC patients. Thus, dynamic control of their presence in blood and assessment of the sensitivity of CSCs to cytostatics in vitro can improve the effectiveness of chemotherapy in BC

Antidiabetic effects of Bisamide Derivative of Dicarboxylic Acid in metabolic disorders

In clinical practice, metabolic syndrome can lead to multiple complications, including diabetes. It remains unclear, which component of the metabolic syndrome (obesity, inflammation, hyperglycemia or insulin resistance) has the strongest inhibitory effect on stem cells involved in beta cell regeneration. This makes it challenging to develop effective treatment options for complications such as diabetes. In our study, experiments were performed on male C57BL/6 mice where metabolic disorders have been introduced experimentally by a combination of streptozotocin-treatment and high-fat diet. We evaluated the biological effects of Bisamide Derivative of Dicarboxylic Acid (BDDA) and its impact on pancreatic stem cells in vivo. To assess the impact of BDDA, we applied a combination of histological and biochemical methods along with a cytometric analysis of stem cell and progenitor cell markers. We show that in mice with metabolic disorders, BDDA has a positive effect on lipid and glucose metabolism. The pancreatic restoration was associated with a decrease of the inhibitory effects of inflammation and obesity factors on pancreatic stem cells. Our data show that BDDA increases the number of pancreatic stem cells. Thus, BDDA could be used as a new compound for treating complications of the metabolic syndrome such as diabetes

Analysis of circulating tumor and cancer stem cells provides new opportunities in diagnosis and treatment of small cell lung cancer

Current methods for diagnosis and treatment of small cell lung cancer (SCLC) have only a modest efficacy. In this pilot study, we analyzed circulating tumor cells (CTCs) and cancer stem cells (CSCs) in patients with SCLC to search for new diagnostic and prognostic markers and novel approaches to improve the treatment of the disease. In other forms of lung cancer, we showed a heterogeneity of blood CTCs and CSCs populations, as well as changes in other cell populations (ALDH+, CD87+CD276+, and EGF+Axl+) in smokers. A number of CTCs and CSCs in patients with SCLC have been shown to be resistant to chemotherapy (CT). High cytotoxic activity and resistance to apoptosis of reprogrammed CD3+CD8+ T-lymphocytes (rTcells) in relation to naive CD3+CD8+ T-lymphocytes was demonstrated in a smoking patient with SCLC (Patient G) in vitro. The target for rTcells was patient G’s blood CSCs. Reprogramming of CD3+CD8+ T-lymphocytes was carried out with the MEK1/2 inhibitor and PD-1/PD-L1 pathway blocker nivolumab. The training procedure was performed with a suspension of dead CTCs and CSCs obtained from patient’s G blood. The presented data show a new avenue for personalized SCLC diagnosis and targeted improvement of chemotherapy based on the use of both CTCs and CSCs

Intricacies of GABA_A Receptor Function: The Critical Role of the β3 Subunit in Norm and Pathology

Neuronal intracellular chloride ([Cl−]i) is a key determinant in γ-aminobutyric acid type A (GABA)ergic signaling. γ-Aminobutyric acid type A receptors (GABAARs) mediate both inhibitory and excitatory neurotransmission, as the passive fluxes of Cl− and HCO3− via pores can be reversed by changes in the transmembrane concentration gradient of Cl−. The cation–chloride co-transporters (CCCs) are the primary systems for maintaining [Cl−]i homeostasis. However, despite extensive electrophysiological data obtained in vitro that are supported by a wide range of molecular biological studies on the expression patterns and properties of CCCs, the presence of ontogenetic changes in [Cl−]i—along with the consequent shift in GABA reversal potential—remain a subject of debate. Recent studies showed that the β3 subunit possesses properties of the P-type ATPase that participates in the ATP-consuming movement of Cl− via the receptor. Moreover, row studies have demonstrated that the β3 subunit is a key player in GABAAR performance and in the appearance of serious neurological disorders. In this review, we discuss the properties and driving forces of CCCs and Cl−, HCO3−ATPase in the maintenance of [Cl−]i homeostasis after changes in upcoming GABAAR function. Moreover, we discuss the contribution of the β3 subunit in the manifestation of epilepsy, autism, and other syndromes

Changes in the Surface Expression of Intercellular Adhesion Molecule 3, the Induction of Apoptosis, and the Inhibition of Cell-Cycle Progression of Human Multidrug-Resistant Jurkat/A4 Cells Exposed to a Random Positioning Machine

Experiments from flight- and ground-based model systems suggest that unexpected alterations of the human lymphoblastoid cell line Jurkat, as well as effects on cell growth, metabolism, and apoptosis, can occur in altered gravity conditions. Using a desktop random positioning machine (RPM), we investigated the effects of simulated microgravity on Jurkat cells and their multidrug-resistant subline, Jurkat/A4 cells. The viability of Jurkat/A4 cells decreased after simulated microgravity in contrast with the Jurkat cells. At the same time, the viability between the experimental Jurkat cells and control Jurkat cells was not significantly different. Of note, Jurkat cells appeared as less susceptible to apoptosis than their multidrug-resistant clone Jurkat/A4 cells, whereas cell-cycle analysis showed that the percentage of Jurkat/A4 cells in the S-phase was increased after 72 and 96 h of RPM-simulated microgravity relative to their static counterparts. The differences in Jurkat cells at all phases between static and simulated microgravity were not significant. The surface expression of the intercellular adhesion molecule 3 (ICAM-3)—also known as cluster of differentiation (CD)50—protein was changed for Jurkat/A4 cells following exposure to the RPM. Changes in cell morphology were observed in the Jurkat/A4 cells after 96 h of RPM-simulated microgravity. Thus, we concluded that Jurkat/A4 cells are more sensitive to RPM-simulated microgravity as compared with the parental Jurkat cell line. We also suggest that intercellular adhesion molecule 3 may be an important adhesion molecule involved in the induction of leukocyte apoptosis. The Jurkat/A4 cells with an acquired multidrug resistance phenotype could be a useful model for studying the effects of simulated microgravity and testing anticancer drugs

Sequential binding of calcium ions to the B-repeat domain of SdrD from Staphylococcus aureus

International audienceBiofilms of live bacteria forming on medical devices and implants contribute significantly to bacterial blood dissemination and to the spread of nosocomial infections. Cell surface SdrD protein plays a key role in the attachment of Staphylococcus aureus to the extracellular matrix (ECM) and in the formation of biofilm. SdrD binds calcium ions using its B1–B5 region bearing EF-hand Ca-binding sites, leading to conformational changes in the structure of SdrD. This alters the distance between the bacterial surface and the ECM-interacting domain of SdrD in a spring-like fashion, participating in bacterial attachment. In this study we investigated calcium binding to EF-hand sites of SdrD using isothermal titration calorimetry and determined the impact of this process on SdrD's thermodynamic stability. This allowed us to propose a model of B1–B5 reorganization upon binding of calcium and to get new insight into the molecular mechanism of SdrD's action

Zinc Inhibits the GABA_AR/ATPase during Postnatal Rat Development: The Role of Cysteine Residue

Zinc ions (Zn2+) are concentrated in various brain regions and can act as a neuromodulator, targeting a wide spectrum of postsynaptic receptors and enzymes. Zn2+ inhibits the GABAARs, and its potency is profoundly affected by the subunit composition and neuronal developmental stage. Although the extracellular amino acid residues of the receptor’s hetero-oligomeric structure are preferred for Zn2+ binding, there are intracellular sites that, in principle, could coordinate its potency. However, their role in modulating the receptor function during postembryonic development remains unclear. The GABAAR possesses an intracellular ATPase that enables the energy-dependent anion transport via a pore. Here, we propose a mechanistic and molecular basis for the inhibition of intracellular GABAAR/ATPase function by Zn2+ in neonatal and adult rats. The enzymes within the scope of GABAAR performance as Cl−ATPase and then as Cl−, HCO3−ATPase form during the first week of postnatal rat development. In addition, we have shown that the Cl−ATPase form belongs to the β1 subunit, whereas the β3 subunit preferably possesses the Cl−, HCO3−ATPase activity. We demonstrated that a Zn2+ with variable efficacy inhibits the GABAAR as well as the ATPase activities of immature or mature neurons. Using fluorescence recording in the cortical synaptoneurosomes (SNs), we showed a competitive association between Zn2+ and NEM in parallel changes both in the ATPase activity and the GABAAR-mediated Cl− and HCO3− fluxes. Finally, by site-directed mutagenesis, we identified in the M3 domain of β subunits the cysteine residue (C313) that is essential for the manifestation of Zn2+ potency