T-Lymphocytes Activated by Dendritic Cells Loaded by Tumor-Derived Vesicles Decrease Viability of Melanoma Cells In Vitro.

Immunotherapy represents an innovative approach to cancer treatment, based on activating the body's own immune system to combat tumor cells. Among various immunotherapy strategies, dendritic cell vaccines hold a special place due to their ability to activate T-lymphocytes, key players in cellular immunity, and direct them to tumor cells. In this study, the influence of dendritic cells processed with tumor-derived vesicles on the viability of melanoma cells in vitro was investigated. Dendritic cells were loaded with tumor-derived vesicles, after which they were used to activate T-cells. The study demonstrated that such modified T-cells exhibit high activity against melanoma cells, leading to a decrease in their viability. Our analysis highlights the potential efficacy of this approach in developing immunotherapy against melanoma. These results provide new prospects for further research and the development of antitumor strategies based on the mechanisms of T-lymphocyte activation using tumor-derived vesicles.

The Potential of Dendritic Cell Subsets in the Development of Personalized Immunotherapy for Cancer Treatment.

Since the discovery of dendritic cells (DCs) in 1973 by Ralph Steinman, a tremendous amount of knowledge regarding these innate immunity cells has been accumulating. Their role in regulating both innate and adaptive immune processes is gradually being uncovered. DCs are proficient antigen-presenting cells capable of activating naive T-lymphocytes to initiate and generate effective anti-tumor responses. Although DC-based immunotherapy has not yielded significant results, the substantial number of ongoing clinical trials underscores the relevance of DC vaccines, particularly as adjunctive therapy or in combination with other treatment options. This review presents an overview of current knowledge regarding human DCs, their classification, and the functions of distinct DC populations. The stepwise process of developing therapeutic DC vaccines to treat oncological diseases is discussed, along with speculation on the potential of combined therapy approaches and the role of DC vaccines in modern immunotherapy.

Serum Cytokine Profile, Beta-Hexosaminidase A Enzymatic Activity and GM2 Ganglioside Levels in the Plasma of a Tay-Sachs Disease Patient after Cord Blood Cell Transplantation and Curcumin Administration: A Case Report.

Tay-Sachs disease (TSD) is a progressive neurodegenerative disorder that occurs due to a deficiency of a ß hexosaminidase A (HexA) enzyme, resulting in the accumulation of GM2 gangliosides. In this work, we analyzed the effect of umbilical cord blood cell transplantation (UCBCT) and curcumin administration on the course of the disease in a patient with adult TSD. The patient's serum cytokine profile was determined using multiplex analysis. The level of GM2 gangliosides in plasma was determined using mass spectrometry. The enzymatic activity of HexA in the plasma of the patient was assessed using a fluorescent substrate assay. The HexA α-subunit (HexA) concentration was determined using ELISA. It was shown that both UCBCT and curcumin administration led to a change in the patient's cytokine profile. The UCBCT resulted in an increase in the concentration of HexA in the patient's serum and in an improvement in the patient's neurological status. However, neither UCBCT nor curcumin were able to alter HexA activity and the level of GM2 in patient's plasma. The data obtained indicate that UCBCT and curcumin administration can alter the immunity of a patient with TSD, reduce the level of inflammatory cytokines and thereby improve the patient's condition.

Proteomic profiling of the rat hippocampus from the kindling and pilocarpine models of epilepsy: potential targets in calcium regulatory network.

Herein proteomic profiling of the rat hippocampus from the kindling and pilocarpine models of epilepsy was performed to achieve new potential targets for treating epileptic seizures. A total of 144 differently expressed proteins in both left and right hippocampi by two-dimensional electrophoresis coupled to matrix-assisted laser desorption-mass spectrometry were identified across the rat models of epilepsy. Based on network analysis, the majority of differentially expressed proteins were associated with Ca2+ homeostasis. Changes in ADP-ribosyl cyclase (ADPRC), lysophosphatidic acid receptor 3 (LPAR3), calreticulin, ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1), synaptosomal nerve-associated protein 25 (SNAP 25) and transgelin 3 proteins were probed by Western blot analysis and validated using immunohistochemistry. Inhibition of calcium influx by 8-Bromo-cADP-Ribose (8-Br-cADPR) and 2-Aminoethyl diphenylborinate (2-APB) which act via the ADPRC and LPAR3, respectively, attenuated epileptic seizures. Considering a wide range of molecular events and effective role of calcium homeostasis in epilepsy, polypharmacy with multiple realistic targets should be further explored to reach the most effective treatments.

Combination of epidural electrical stimulation with ex vivo triple gene therapy for spinal cord injury: a proof of principle study.

Despite emerging contemporary biotechnological methods such as gene- and stem cell-based therapy, there are no clinically established therapeutic strategies for neural regeneration after spinal cord injury. Our previous studies have demonstrated that transplantation of genetically engineered human umbilical cord blood mononuclear cells producing three recombinant therapeutic molecules, including vascular endothelial growth factor (VEGF), glial cell-line derived neurotrophic factor (GDNF), and neural cell adhesion molecule (NCAM) can improve morpho-functional recovery of injured spinal cord in rats and mini-pigs. To investigate the efficacy of human umbilical cord blood mononuclear cells-mediated triple-gene therapy combined with epidural electrical stimulation in the treatment of spinal cord injury, in this study, rats with moderate spinal cord contusion injury were intrathecally infused with human umbilical cord blood mononuclear cells expressing recombinant genes VEGF165, GDNF, NCAM1 at 4 hours after spinal cord injury. Three days after injury, epidural stimulations were given simultaneously above the lesion site at C5 (to stimulate the cervical network related to forelimb functions) and below the lesion site at L2 (to activate the central pattern generators) every other day for 4 weeks. Rats subjected to the combined treatment showed a limited functional improvement of the knee joint, high preservation of muscle fiber area in tibialis anterior muscle and increased H/M ratio in gastrocnemius muscle 30 days after spinal cord injury. However, beneficial cellular outcomes such as reduced apoptosis and increased sparing of the gray and white matters, and enhanced expression of heat shock and synaptic proteins were found in rats with spinal cord injury subjected to the combined epidural electrical stimulation with gene therapy. This study presents the first proof of principle study of combination of the multisite epidural electrical stimulation with ex vivo triple gene therapy (VEGF, GDNF and NCAM) for treatment of spinal cord injury in rat models. The animal protocols were approved by the Kazan State Medical University Animal Care and Use Committee (approval No. 2.20.02.18) on February 20, 2018.

Hippocampal asymmetry: differences in the left and right hippocampus proteome in the rat model of temporal lobe epilepsy.

The hippocampus is a complex brain structure and undergoes severe sclerosis and gliosis in temporal lobe epilepsy (TLE) as the most common type of epilepsy. The key features of the TLE may be reported in chronic animal models of epilepsy, such as pilocarpine model. Therefore, the current study was conducted in a rat pilocarpine model of acquired epilepsy. Two-dimensional gel electrophoresis based proteomic technique was used to compare the proteome map of the left and right hippocampus in both control and epileptic rats. Generally, 95 differentially expressed spots out of 1300 spots were identified in the hippocampus proteome using MALDI-TOF-TOF/MS. Within identified proteins, some showed asymmetric expression related to the mechanisms underlying TLE imposed by pilocarpine. Assessment of lateralization at the molecular level demonstrated that expression of proteins involved in dopamine synthesis was significantly more in the right hippocampus than the left one. In the epileptic model, reduction in dopamine pathway proteins was accompanied by an increase in the expression of proteins involved in polyamine synthesis, referring to a new regulating mechanism. Our results revealed changes in the laterality of protein expression due to pilocarpine-induced status epilepticus that could present some new proteins as potential candidates for antiepileptic drug design. BIOLOGICAL SIGNIFICANCE: In the current study, two-dimensional gel electrophoresis (2-DE) based proteomic technique was used to profile changes in the left and right hippocampus proteome after pilocarpine induced status epilepticus. Spots of proteome maps for two hemispheres were excised and identified with MALDI-TOF-TOF/MS. Analysis of proteome map of the left and right hippocampus revealed a lateralization at the molecular level, in which the expression of proteins involved in dopamine synthesis and release were significantly more in right hippocampi than the left ones in the normal rats. Also, the expression of proteins involved in polyamine synthesis significantly increased in epileptic hippocampus (considerably higher in right hippocampi), whilst the proteins which included in dopamine pathways were decreased. Our results revealed changes in the laterality of protein expression due to pilocarpine-induced status epilepticus that could present some new proteins as potential candidates for antiepileptic drug design.

Tandem Delivery of Multiple Therapeutic Genes Using Umbilical Cord Blood Cells Improves Symptomatic Outcomes in ALS.

Current treatment options of chronic, progressive degenerative neuropsychiatric conditions offer only marginal efficacy, and there is no therapy which arrests or even reverses these diseases. Interest in genetic engineering and cell-based approaches have constantly been increasing, although most of them so far proved to be fruitless or at best provided very slight clinical benefit. In the light of the highly complex patho-mechanisms of these maladies, the failure of drugs aimed at targeting single molecules is not surprising. In order to improve their effectiveness, the role of a unique triple-combination gene therapy was investigated in this study. Intravenous injection of human umbilical cord blood mononuclear cell (hUCBMC) cotransduced with adenoviral vectors expressing vascular endothelial growth factor (VEGF), glial cell-derived neurotrophic factor (GDNF), and neural cell adhesion molecule (NCAM) resulted in prominent increase of life span and performance in behavioral tests in amyotrophic lateral sclerosis (ALS). Expression of the recombinant genes in hUCBMCs was confirmed as soon as 5 days after transduction by RT-PCR, and cells were detectable for as long as 1 month after grafting in lumbar spinal cord by immunofluorescent staining. Xenotransplantation of cells into mice blood without any immunosuppression demonstrated a high level of hUCBMCs homing and survivability in the central nervous system (CNS), most conspicuously in the spinal cord, but not in the spleen or liver. This study confirms an increased addressed homing and notable survivability of triple-transfected cells in lumbar spinal cord, yielding a remarkably enhanced therapeutic potential of hUCBMCs overexpressing neurotrophic factors.

Symptomatic improvement, increased life-span and sustained cell homing in amyotrophic lateral sclerosis after transplantation of human umbilical cord blood cells genetically modified with adeno-viral vectors expressing a neuro-protective factor and a neural cell adhesion molecule.

Amyotrophic lateral sclerosis (ALS) is an incurable, chronic, fatal neuro-degenerative disease characterized by progressive loss of moto-neurons and paralysis of skeletal muscles. Reactivating dysfunctional areas is under earnest investigation utilizing various approaches. Here we present an innovative gene-cell construct aimed at reviving inert structure and function. Human umbilical cord blood cells (hUCBCs) transduced with adeno-viral vectors encoding human VEGF, GDNF and/or NCAM genes were transplanted into transgenic ALS mice models. Significant improvement in behavioral performance (open-field and grip-strength tests), as well as increased life-span was observed in rodents treated with NCAM-VEGF or NCAM-GDNF co-transfected cells. Active trans-gene expression was found in the spinal cord of ALS mice 10 weeks after delivering genetically modified hUCBCs, and cells were detectable even 5 months following transplantation. Our gene-cell therapy model yielded prominent symptomatic control and prolonged life-time in ALS. Incredible survivability of xeno-transpanted cells was also observed without any immune-suppression. These results suggest that engineered hUCBCs may offer effective gene-cell therapy in ALS.

Bacterial enzymes effectively digest Alzheimer's ß-amyloid peptide.

Aggregated ß-amyloid peptides play key roles in the development of Alzheimer's disease, and recent evidence suggests that microbial particles, among others, can facilitate their polymerization. Bacterial enzymes, however, have been proved to be beneficial in degrading pathological fibrillar structures in clinical settings, such as strepto-kinases in resolving blood-clots. The purpose of this study was to investigate the ability of bacterial substances to effectively hydrolyze ß-amyloid peptides. Degrading products of several proteinases from Bacillus pumilus were evaluated using MALDI-TOF mass-spectrometry, and their toxicity was assessed in vitro using cell-culture assays and morphological studies. These enzymes have proved to be non-toxic and were demonstrated to cleave through the functional domains of ß-amyloid peptide. By yielding inactive fragments, proteinases of Bacillus pumilus may be used as candidate anti-amyloid agents.

Disease-specific expression of the serotonin-receptor 5-HT(2C) in natural killer cells in Alzheimer's dementia.

Alzheimer's dementia (AD) is a degenerative brain disorder characterized mainly by cholinergic failure, but other neuro-transmitters are also deficient especially at late stages of the disease. Misfolded ß-amyloid peptide has been identified as a causative agent, however inflammatory changes also play a pivotal role. Even though the most prominent pathology is seen in the cognitive functions, specific abnormalities of the central nervous system (CNS) are also reflected in the periphery, particularly in the immune responses of the body. The aim of this study was to characterize the dopaminergic and serotonergic systems in AD, which are also markedly disrupted along with the hallmark acetyl-choline dysfunction. Peripheral blood mono-nuclear cells (PBMCs) from demented patients were judged against comparison groups including individuals with late-onset depression (LOD), as well as non-demented and non-depressed subjects. Cellular sub-populations were evaluated by mono-clonal antibodies against various cell surface receptors: CD4/CD8 (T-lymphocytes), CD19 (B-lymphocytes), CD14 (monocytes), and CD56 (natural-killer (NK)-cells). The expressions of dopamine D(3) and D(4), as well as serotonin 5-HT(1A), 5-HT(2A), 5-HT(2B) and 5-HT(2C) were also assessed. There were no significant differences among the study groups with respect to the frequency of the cellular sub-types, however a unique profound increase in 5-HT(2C) receptor exclusively in NK-cells was observed in AD. The disease-specific expression of 5-HT(2C), as well as the NK-cell cyto-toxicity, has been linked with cognitive derangement in dementia. These changes not only corroborate the existence of bi-directional communication between the immune system and the CNS, but also elucidate the role of inflammatory activity in AD pathology, and may serve as potential biomarkers for less invasive and early diagnostic purposes as well.

Peripheral blood mono-nuclear cells derived from Alzheimer's disease patients show elevated baseline levels of secreted cytokines but resist stimulation with ß-amyloid peptide.

OBJECTIVES: Among several other factors, the neuro-toxic ß-amyloid peptide (ßAP)-induced inflammatory mechanisms have also been implicated in the pathogenesis of Alzheimer's dementia (AD). Cytokines have recently emerged as prime candidates underlying this immune reaction. The purpose of this study was to evaluate the inflammatory response of peripheral blood mono-nuclear cells (PBMC) in AD. DESIGN: Cross-sectional (observational) study. SETTING: Behavioral and cognitive neurology clinic of the Universidade Federal de Minas Gerais in Belo Horizonte, Brazil. PARTICIPANTS: AD patients (n=19), healthy elderly (n=19) and young (n=14) individuals. MEASUREMENTS: Cytokine levels were assessed by enzyme-linked immuno-sorbent assay (ELISA) after exposing cells to a broad range of ßAP concentrations (10(-4)-10(-10)M) as a stimulus. AD samples were weighed against leukocytes harvested from non-demented young and elderly subjects. RESULTS: Cytokine production of PBMCs in the youth was characterized by low baseline levels when compared to cells from the older generation. In the aging population, AD cells were distinguished from the healthy elderly sub-group by an even higher basal cytokine secretion. The low resting concentration in young individuals was markedly increased after treatment with ßAP, however cells from the elderly, irrespective of their disease status, showed unchanged cytokine release following ßAP administration. Non-specific activation of PBMCs with anti-CD3/CD28 antibodies resulted in elevated interleukin (IL)-1ß concentrations in AD. CONCLUSIONS: These results demonstrate a general over-production of cytokines and resistance to ßAP in the old comparison group, with a more pronounced disruption/boosted pattern in AD. Our findings are in line with the hypothesis of "inflammaging", i.e. an enhanced inflammatory profile with normal aging and a further perturbed environment in AD. The observed cytokine profiles may serve as diagnostic biomarkers in dementia.