Valeriy G. Sergeyev

BACKGROUND: Olfactory dysfunction is a prodromal symptom of many neurodegenerative diseases. Neuroinflammatory and neurodegenerative processes in the olfactory bulbs may be initiated by infectious agents in the nasal cavity, particularly bacterial lipopolysaccharide (LPS). The molecular mechanisms mediating the effects of this endotoxin on the olfactory epithelium and the development of pathological processes in the olfactory bulbs remain insufficiently studied. AIM: This study aimed to investigate the effects of different doses of intranasally administered lipopolysaccharide on the intensity of immunopositive staining of the olfactory epithelium with antibodies against alpha-synuclein (α-syn) and the adaptor protein MyD88, as well as of olfactory bulbs with antibodies against α-syn and glial fibrillary acidic protein (GFAP). METHODS: The study included 18 male BALB/c mice weighing 20–33 g. Animals received unilateral intranasal injections of sterile saline 10 μL daily (control) or lipopolysaccharide at a high (0.1 μg/mL) or low (0.01 μg/mL) concentration. After 28 days, the brains and nasal structure complexes were excised and frozen using dry ice. Serial cryostat sections (14 μm) were prepared and stained with methylene blue and antibodies against α-syn, MyD88, and GFAP. The intensity of immunolabeling was quantitatively assessed using Image-Pro Insight 8.0 software. RESULTS: Unilateral intranasal administration of lipopolysaccharide solution to mice resulted in a dose-dependent increase in α-syn levels in receptor cells, olfactory nerve bundles, and olfactory bulb glomeruli, along with a dose-dependent increase in MyD88 in the receptor epithelium and GFAP in the glomerular layer of the olfactory bulb ipsilateral to endotoxin administration. CONCLUSION: The observed morphological and immunohistochemical changes in the olfactory bulbs following intranasal lipopolysaccharide administration indicate the development of dose-dependent neuroinflammation in the glomerular layer. Neuroinflammation in the olfactory bulb appears to be initiated by LPS-induced upregulation of α-syn expression in receptor neurons projecting to the bulbs, mediated by activation of MyD88, which participates in intracellular signaling downstream of Toll-like receptors and interleukin-1 receptor family members.

Malignant transformation of lymphopoiesis in lymph nodes (LN) is accompanied by structural rearrangement of the LN stroma and changes in the glycosylation of membrane and cytoplasmic proteins. For the histochemical detection of transforming lymphoid cells and remodeled LN stroma, we used the tomato lectin Lycopersicon esculentum, which is able to bind to surface and cytoplasmic glycoproteins of the majority of LN cells. The study aimed to investigate the characteristics of cell architectonics with a high level of protein glycosylation in the LN of patients with B-chronic lymphocytic leukаemia (B-CLL). The study material were biopsy specimens of supraclavicular and cervical LNs from patients of the First Republican Clinical Hospital of the Ministry of Health of the Udmurt Republic with a confirmed diagnosis of B-CLL (16 patients), aged 49-73 years, obtained prior to treatment with their informed voluntary consent. LN biopsies from the same body regions of 12 individuals aged 48-70 years with reactive LN hyperplasia served as control samples. Paraffin sections of 7 µm thick LN were stained with FITC-conjugated tomato lectin and fluorescent dye propidium iodide (IP) and examined using a Nikon Eclipse200 microscope equipped with a luminescence unit and digital camera. Analysis of LN preparations from patients with B-CLL revealed significant changes in the histotopography of cells and extracellular structures with a high degree of glycosylation. Follicles in the cortex were replaced by an array of small lymphocytes against a background of proliferating centers containing lymphocytes with dispersed packing of IP-labelled chromatin. In this area we also observed a uniform network of thin lectin-labelled reticular fibres and a large number of small blood vessels. Macrophage-like cells, clearly identifiable in the germinal centres of follicles in control, were absent in B-CLL. Their increased number and intensity of luminescence was observed in the subcapsular sinus area and in the paracortical area around collagen bundles formed by conduits, as well as around connective tissue trabeculae of the brain substance. The differences observed in the histological topography of highly glycosylated LN cells in B-CLL suggest that the proposed staining method is informative and facilitates the diagnosis of this disease in histological studies.

BACKGROUND: Neuroinflammation is a key pathophysiological mechanism in age-related neurodegenerative diseases such as Parkinson's disease. Dihydroquercetin's water-soluble form (DHQ-WF) is considered a promising agent capable of inhibiting the neuroinflammatory process. Nevertheless, uncertainties persist regarding the cellular and molecular mechanisms governing its effects, taking into account nervous tissue's gliovascular organization. AIM: To study structural changes of microcirculatory vessels and functional responses of micro- and astroglial cells in the substantia nigra of young and old rats in response to intranigral injection of lipopolysaccharide (LPS) and subsequent oral administration of DHQ-WF. MATERIALS AND METHODS: Young (250–320 g) and old (390–450 g) Wistar rats were injected into the substantia nigra using a stereotaxic device with 2 μL of LPS solution at a concentration of 0.01 μL/mL (experimental groups; n=24) or 2 μL of sterile saline (control groups; n=12). Half of the animals in the experimental groups (6 animals of each age group) received 2 ml of a solution containing DHQ-WF ("Taxifolin aqua"; Advanced Technologies Ltd., Russia) at a concentration of 3 mg/mL by gavage daily for 8 weeks. At the end of the experiment, the animals were transcardially perfused with 4% paraformaldehyde, the brain was extracted and frozen on dry ice. Cryostat sections obtained on the cryotome were stained with FITC-labelled tomato lectin for the detection of vascular endothelium and antibodies against GFAP and CD-11β for the immunohistochemical detection of astrocytes and microglia, respectively. The length and number of vessels and their branches were counted using AngioTool software. The areas of glial cell bodies and their processes were measured using the morphometric software ImagePro Inside 8.0. RESULTS: 8 weeks after LPS administration into the substantia nigra (SN) of old rats, a significant excess of areas occupied by cell bodies and processes of microglial and astroglial cells, as well as the number of vessels on the standard plot, was found both in young animals that had experienced similar effects and in old control animals. Oral administration of DHQ-WF to rats significantly reduced LPS-induced glial activation in young and old animals. In addition, administration of DHQ-WF to old animals reduced the intensity of microvascular SN remodeling induced by LPS administration. CONCLUSIONS: Administration of LPS to the SN of rats of different ages causes neuroinflammation, which is maximally expressed in aged animals. In addition, LPS-induced microvessel angiogenesis is observed in aged animals. Administration of DHQ-WF for 8 weeks significantly reduces these LPS-induced changes, which allows us to consider it as a promising anti-neuroinflammatory agent.

Histotopography of lymphocytes expressing bitter taste receptors of T2R family and α-gustducin (receptor-associated subunit of the G-protein complex) in mouse spleen in the norm and in 48 h after intraperitoneal administration of LPS was studied by the immunohistochemical method. Two populations of immunopositive lymphocytes expressing the above proteins were detected in the spleen; they were located in the marginal zones of lymphoid follicles and periarteriolar lymphoid sheaths. In most α-gustducin-positive lymphocytes, co-expression of α-gustducin and CD19 (B-cell marker) was found. Intraperitoneal administration of LPS significantly increased the number of gustducinergic lymphocytes in periarteriolar lymphoid sheaths. We hypothesize that spleen lymphocytes with gustducin signaling are involved in T-dependent immune response to the blood bacterial polysaccharides.

Hormone peptide tyrosine–tyrosine (PYY) is secreted into circulation from the gut L-endocrine cells in response to food intake, thus inducing satiation during interaction with its preferred receptor, Y₂R. Clinical applications of systemically administered PYY for the purpose of reducing body weight were compromised as a result of the common side effect of visceral sickness. We describe here a novel approach of elevating PYY in saliva in mice, which, although reliably inducing strong anorexic responses, does not cause aversive reactions. The augmentation of salivary PYY activated forebrain areas known to mediate feeding, hunger, and satiation while minimally affecting brainstem chemoreceptor zones triggering nausea. By comparing neuronal pathways activated by systemic versus salivary PYY, we identified a metabolic circuit associated with Y₂R-positive cells in the oral cavity and extending through brainstem nuclei into hypothalamic satiety centers. The discovery of this alternative circuit that regulates ingestive behavior without inducing taste aversion may open the possibility of a therapeutic application of PYY for the treatment of obesity via direct oral application.

Administration of autologous macrophages isolated from the abdominal cavity and stimulated in vitro with bacterial LPS to rats increased blood level of antibodies against α-synuclein. Antibody titer reached its maximum during week 5 of the experiment and exceeded the levels of anti-α-synuclein antibodies appeared in response to transplantation of non-stimulated macrophages. Brain immunohistochemistry showed that additional administration of LPS (250 μg/kg) to animals during week 4 after injection of LPS-stimulated macrophages led to α-synuclein accumulation in 9.4±3.2% dopaminergic neurons of the substantia nigra. These findings attest to induction of humoral immune response to α-synuclein in rats after administration of autologous LPS-stimulated macrophages, which can affect α-synuclein metabolism in dopaminergic neurons of the brain.