Search Results (208)

Preterm infants often require oxygen supplementation, resulting in high risk for bronchopulmonary dysplasia (BPD) and neurodevelopmental deficits. Despite a growing number of studies, there is still little knowledge about brain injury in BPD models. Therefore, we exposed neonatal C57BL/6 mice to 85% oxygen from birth to postnatal day (P) 14. At P28, two weeks after recovery under normoxic conditions, right hemisphere was used for the analysis of mRNA and the left hemisphere for protein expression of neuronal cells, neuroinflammatory and vascularisation markers, analysed by real-time PCR and Western blot, respectively. Hyperoxia led to an altered expression of markers associated with neuronal and oligodendrocyte maturation and neuroinflammation such as Dcx, Nestin, Il-1β, Il-6, NG2, and YM1/2. These changes were accompanied by an increased expression of genes involved in angiogenesis and vascular remodelling, e.g., Vegf-a, Nrp-1, and Icam-1. Together, 14 days of hyperoxia triggered a phenotypic response, resembling signs of encephalopathy of prematurity (EoP). Full article

Background and Objectives: Alzheimer’s disease (AD) has become a serious health problem. Agomelatine (Ago) is a neuroprotective antidepressant. This study aimed to assess how Ago influences behavioral outcomes in AD-like rat model. Materials and Methods: Forty-eight Wistar albino rats were allocated into four groups: Control (C), Alzheimer’s disease-like model (AD), Alzheimer’s disease-like model treated with Ago (ADAgo), and Ago alone (Ago). Physiological saline was injected intrahippocampally in C and Ago animals, whereas Aβ peptide was delivered similarly in AD and ADAgo rats. On day 15, 0.9% NaCl was administered to the C and AD groups, and Agomelatine (1 mg/kg/day) was infused into ADAgo and Ago rats via osmotic pumps for 30 days. Behavioral functions were evaluated using Open Field (OF), Forced Swim (FST), and Morris Water Maze (MWM) tests. Brain tissues were examined histopathologically. Neuritin, Nestin, DCX, NeuN, BDNF, MASH1, MT1, and MT2 transcripts were quantified by real-time PCR. Statistical analyses were performed in R 4.3.1, with p < 0.05 deemed significant. Results: In the FST, swimming, climbing, immobility time, and mobility percentage differed significantly among groups (p < 0.05). In the MWM, AD rats exhibited impaired learning and memory that was ameliorated by Ago treatment (p < 0.05). DCX expression decreased in AD rats but was elevated by Ago (p < 0.05). Nestin levels differed significantly between control and AD animals; MT1 expression varied between control and AD cohorts; and MT2 transcript levels were significantly lower in AD, ADAgo, and Ago groups compared to C (all p < 0.05). Conclusions: Ago exhibits antidepressant-like activity in this experimental AD model and may enhance cognitive function via mechanisms beyond synaptic plasticity and neurogenesis. Full article

Astrocytes, which proliferate after brain injury, represent a promising target for cellular reprogramming due to their abundance and ability to support brain repair. In this study, we investigated the in vitro reprogramming of primary cortical astrocytes from neonatal rats into neuronal precursor cells (NPCs) using the transcription factors Oct4, Sox2, and Klf4 (OSK), delivered via lentiviral vectors. We designed a reporter system to trace the conversion of astrocytes to NPCs and neurons by using GFAP-driven iCre and Nestin- or Synapsin1-driven fluorescent reporters. After transduction, we observed morphological changes and the expression of neuronal markers in some cells, while many cells remained in a transitional state, expressing both astrocytic and neuronal features. Importantly, the study was not designed to quantify reprogramming efficiency or demonstrate full astrocyte-to-neuron conversion but rather to establish and evaluate a traceable reporter system. Our data suggest that OSK-mediated reprogramming in this in vitro model can initiate conversion of astrocytes to neuronal precursor-like cells, although the process is complex and incomplete within the one-week timeframe. We also highlight limitations in co-transduction efficiency and potential silencing of the reporter system during reprogramming. These findings provide an initial technical platform to explore astrocyte reprogramming in vitro and inform future studies aiming to refine these methods and apply them in vivo. Full article

Skin aging is driven by cellular senescence, oxidative stress, and diminished regenerative capacity. In this study, we investigated the effects of PhytoCellTec™ Argan, an argan callus extract (PC), on primary human fibroblasts and adult stem cells. PC treatment (0.1% and 0.5%) significantly enhanced fibroblast proliferation, reduced senescence-associated β-galactosidase activity, and decreased the expression of p16, p21, and phosphorylated NFκB. PC treatment lowered intracellular ROS levels, increased ATP production, and promoted autophagy via LC3B-II accumulation and p62 reduction. In skin-derived precursor cells (SKPs), as well as mesenchymal stem cells (MSCs), PC treatment improved spheroid formation and growth while preserving the expression of key stemness markers, including Sox2, Oct4, and Nestin. Furthermore, PC exhibited antioxidant capacity (TEAC assay) and inhibited elastase, supporting its anti-aging potential. These findings suggest that PC is safe at concentrations below 1% and may serve as an effective natural compound to restore cellular homeostasis, reduce senescence and inflammation, and support stem cell health during aging. Full article

Background: Glioblastoma (GBM) remains almost uniformly fatal, owing in part to therapy-resistant cancer stem-like cells (CSCs) and to temozolomide (TMZ) resistance driven by O⁶-methylguanine-DNA methyltransferase (MGMT). Differentiation therapy with all-trans retinoic acid (ATRA) has the potential to attenuate stemness and sensitize GBM to TMZ. We therefore asked whether ATRA reduces expression of key CSC markers and MGMT in established GBM lines. Methods: Two established human GBM cell lines, U87-MG and A172, were cultured under neurosphere-promoting conditions to enrich for potential stem-like subpopulations. Cells were treated with either 1 µM ATRA or vehicle control (DMSO) for 5 days. Total RNA was extracted, and cDNA was synthesized. Quantitative Real-Time PCR (qPCR) assessed relative mRNA expression levels of key stemness transcription factors (SOX2, NES) and the DNA repair gene MGMT and corresponding protein levels were measured by an Enzyme-Linked Immunosorbent Assay (ELISA). Gene expression was normalized to the geometric mean of two validated housekeeping genes (GAPDH, ACTB). Relative quantification was calculated using the ΔΔCt method, and statistical significance was determined using Student’s t-tests. Results: ATRA markedly suppressed stemness and MGMT in both lines. In U87-MG, SOX2 mRNA fell 3.7-fold (p = 0.0008) and protein 2.99-fold (148.3 ± 6.0 → 49.7 ± 2.7 pg µg⁻¹; p = 0.0002); Nestin dropped 4.1-fold (p = 0.0005) and 3.51-fold (450.0 ± 17.3 → 128.3 ± 4.4 pg µg⁻¹; p = 0.00008). MGMT decreased 2.6-fold at transcript level (p = 0.0065) and 2.11-fold at protein level (81.7 ± 4.4 → 38.7 ± 1.8 pg µg⁻¹; p = 0.0005). In A172, SOX2 was reduced 2.9-fold (p = 0.0041) and 2.31-fold (p = 0.0007); Nestin 3.3-fold (p = 0.0028) and 2.79-fold (p = 0.00009). MGMT declined 2.2-fold (p = 0.0132) and 1.82-fold (p = 0.0015), respectively. Conclusions: Five-day exposure to ATRA diminishes SOX2, Nestin, and MGMT at both mRNA and protein levels in stem-enriched GBM cultures, supporting the premise that ATRA-induced differentiation can concurrently blunt CSC traits and TMZ-resistance mechanisms. These data provide a molecular rationale for testing ATRA in combination regimens aimed at improving GBM therapy. Full article

The development of more effective disease-modifying treatments for Alzheimer’s disease (AD) is compromised by the lack of streamlined measures to detect and monitor the full spectrum of neurodegeneration, including white matter pathology, which begins early. This study utilized an established intracerebral streptozotocin (STZ) model of AD to examine the potential utility of a non-invasive serum extracellular vesicle (SEV)-based liquid biopsy approach for detecting a broad range of molecular pathologies related to neurodegeneration. The design enabled comparative analysis of immunoreactivity in frontal lobe tissue (FLTX), frontal lobe-derived EVs (FLEVs), and SEVs. Long Evans rats were administered i.c. STZ or saline (control) on postnatal day 3 (P3). Morris Water Maze testing was performed from P24 to P27. On P31–32, the rats were sacrificed to harvest FLTX and serum for EV characterization. STZ caused brain atrophy, with deficits in spatial learning and memory. STZ significantly impacted FLEV and SEV nanoparticle abundance and size distributions and concordantly increased AD (Tau, pTau, and Aβ) and oxidative stress (ubiquitin, 4-HNE) biomarkers, as well as immunoreactivity to immature oligodendrocyte (PLP), non-myelinating glial (PDGFRA, GALC) proteins, MAG, nestin, and GFAP in FLTX and FLEV. The SEVs also exhibited concordant STZ-related effects, but they were limited to increased levels of 4-HNE, PLP, PDGFRA, GALC, MAG, and GFAP. The findings suggest that non-invasive EV-based liquid biopsy approaches could potentially be used to detect and monitor some aspects of AD-type neurodegeneration. Targeting brain-specific EVs in serum will likely increase the sensitivity of this promising non-invasive approach for diagnostic and clinical management. Full article

Background/Objectives: Nestin is an intermediate filament protein and a marker of odontoblasts, but its function in tooth and bone formation is largely unknown. This study aimed to determine whether Nestin plays a role in postnatal tooth formation. Methods: 4-week-old Nestin knockout (KO) mice were analyzed with a range of techniques, including X-ray imaging, uCT, backscattered and acid-etched casted SEM, FITC-confocal microscopy, H&E and TRAP staining, and immunohistochemistry. Results: The KO mice had no apparent difference in crown formation compared to age-matched wild-type (WT) but showed delayed molar eruption with reduced TRAP⁺ osteoclasts. More importantly, KO mice developed expanded predentin and shorter, thinner roots with irregular and shortened dentin tubules. Additionally, the Nestin KO mice exhibited a reduced cellular cementum mass with sharp reductions in DMP1, OPN, and BSP. Conclusions: These findings suggest that Nestin plays a critical role in the postnatal development of root dentin and cellular cementum. Full article

Background: The intermediate filament nestin was first described in stem and progenitor cells of neural and mesenchymal origin. Additionally, it is expressed in endothelial cells during wound healing and tumorigenesis. Thus, nestin is widely regarded as a marker for proliferative endothelium. However, little is known about its role in lymphatic endothelium. Methods: Here, we analyzed the expression of nestin in the endothelium of ten human haemangiomas and ten lymphangiomas in situ by immunohistochemistry. This study aimed to investigate the expression of nestin in haemangiomas and lymphangiomas to determine its potential role as a vascular marker. Specifically, we aimed to assess whether nestin expression is restricted to proliferating endothelial cells or also present in non-proliferative blood vessels. Results: Immunohistochemically, haemangiomas were positive for CD31 but negative for D2-40. The endothelial cells within these lesions showed a homogeneous expression of nestin. In contrast, the endothelium of lymphangiomas reacted positively for D2-40 and CD31 but did not show any nestin expression. Additionally, only a few endothelial cells of capillary haemangiomas showed a Ki-67 positivity. Conclusions: The differential expression of nestin in haemangiomas and lymphangiomas indicates a specificity of nestin for the endothelium of blood vessels. The Ki-67 negativity in the majority of the endothelial cells reveals the proliferative quiescence of these cells. These findings indicate that nestin could be used as a marker to differentiate between blood and lymphatic vessels. Full article

Background: Blast traumatic brain injury (bTBI) can result in depression-like behaviors in the acute and chronic phases. SSRIs have been shown to significantly alleviate depression-like behaviors in animal models of traumatic brain injury (TBI) by increasing serotonin (5-HT) and brain-derived neurotrophic factor (BDNF) in the hippocampus. However, the therapeutic effects of SSRIs on depression caused by bTBI remain unclear. Objective: Therefore, this study was aimed at investigating the therapeutic effects of SSRIs on depression-like behaviors in bTBI models. Methods: We created a rat model to study mild TBI by subjecting rats to increased blast overpressures (BOP) and injecting fluoxetine and escitalopram SSRIs intraperitoneally for 28 days. Results: On day 14 post-BOP exposure, rats treated with SSRIs showed decreased depression-like behaviors. This finding was accompanied by higher 5-HT levels in the hippocampus and increased numbers of Nestin-positive cells in the dentate gyrus. Furthermore, rats treated with SSRIs exhibited increased pCREB and BDNF protein expression in the hippocampus on days 7, 14, and 28 after bTBI. Conclusions: Overall, our findings indicate that SSRI-induced recovery from depression-like behaviors after mild bTBI is associated with the upregulation of 5-HT levels, pCREB and BDNF expression, and neurogenesis in the hippocampus. Full article

Retinoic acid (RA) is commonly used to differentiate SH-SY5Y neuroblastoma cells. This effect is sustained by a specific modulation of gene transcription, leading to marked changes in cellular proteins. In this scenario, autophagy may be pivotal in balancing protein synthesis and degradation. The present study analyzes whether some autophagy-related proteins and organelles are modified during RA-induced differentiation of SH-SY5Y cells. RA-induced effects were compared to those induced by starvation. SH-SY5Y cells were treated with a single dose of 10 µM RA or grown in starvation, for 3 days or 7 days. After treatments, cells were analyzed at light microscopy and transmission electron microscopy to assess cell morphology and immunostaining for specific markers (nestin, βIII-tubulin, NeuN) and some autophagy-related proteins (Beclin 1, LC3). We found that both RA and starvation differentiate SH-SY5Y cells. Specifically, cell differentiation was concomitant with an increase in autophagy proteins and autophagy-related organelles. However, the effects of a single dose of 10 μM RA persist for at least 7 days, while prolonged starvation produces cell degeneration and cell loss. Remarkably, the effects of RA are modulated in the presence of autophagy inhibitors or stimulators. The present data indicate that RA-induced differentiation is concomitant with an increased autophagy. Full article

Many endodontic procedures within the pediatric population are performed with patients aged 12 years and older, using intracanal irrigants to complement mechanical debridement for the removal of debris and to disinfect the root canal system. The use of antimicrobial irrigants that limit damage to the dental pulp are the goals of endodontic biomaterials research. Using an existing biorepository of dental pulp stem cells (DPSCs), Endocyn was evaluated in varying concentrations in proliferation and viability assays, and compared with positive (sodium hypochlorite or bleach) and negative (phosphate-buffered saline) controls. The DPSC viability was reduced in the range of −8.3% to −15.8%, p = 0.22 to p = 0.042, while the growth inhibition varied between −29.7% and −63%, p = 0.041 to p = 0.022. However, the RNA analysis revealed that no significant changes in biomarker mRNA expression (Nestin, NANOG, Sox2, Oct4, CD73, CD90, and CD105) were observed. These data demonstrated that all of the concentrations of Endocyn inhibited the DPSC viability and growth, although only high concentrations were statistically significant. Moreover, the administration of Endocyn did not alter the DPSC biomarker expression, which are novel and important findings not previously observed or reported that may assist with the development of clinical decision protocols and methods for the treatment of vital pulp tissue. Full article

Nestin is a type VI intermediate filament protein and a well-known neural stem cell marker. It is also expressed in high-grade cancer cells, forming copolymerized filaments with vimentin. We previously showed that nestin inhibits the binding of vimentin’s tail domain to actin filaments (AFs) by steric hindrance through its large nestin tail domain (NTD), thereby increasing three-dimensional cytoskeleton network mobility, enhancing cell flexibility, and promoting cancer progression. Further, we found that nestin itself stably binds to AFs via the NTD. We therefore hypothesized that the NTD may form a flexible cytoskeletal structure by extending with weak force. In vitro tensile tests using atomic force microscopy were performed to assess the mechanical properties of NTDs. The C-terminus of the NTD bound AFs by bringing the AFM tip modified with the NTD into contact with the AFs on the substrate. NTDs were elongated to approximately 80% of their maximum length at weak forces < 150 pN. Repeated tensile tests revealed that the NTD refolded quickly and behaved like a soft elastic material. We speculate that nestin stably binds AFs, and the NTD extends with weak force, contracting quickly upon load release. Thereby, nestin would absorb mechanical load and maintain cytoskeletal integrity. Full article

Abamectin is an insecticide, miticide and nematicide that has been extensively used in agriculture for many years. The excessive use of abamectin inevitably pollutes water and soil and might even cause adverse effects on aquatic biota. However, it is currently unclear how abamectin exposure causes neurotoxicity in aquatic organisms. Herein, the early neural system development was assessed in zebrafish embryos following abamectin exposure. After treatment with a concentration gradient of abamectin (0.055, 0.0825, 0.11 mg/L), the survival rate, average heart rate, pericardial edema area and yolk sac edema were all documented in zebrafish embryos (96 hpf). It was found that after abamectin exposure, embryonic brain development was impaired, and motor behaviors were also affected. The fluorescence intensity was reduced in the transgenic embryos (Eno2: GFP). The activities of acetylcholinesterase (AChE) and ATPase were decreased, and the expression of neurodevelopment-related genes, such as sox10, gap43, grin1b, abat, gad1b, grin2b, nestin and glsa, were all inhibited in zebrafish embryo treatment with abamectin. Furthermore, the reactive oxygen species (ROS) were triggered upon exposure to abamectin in zebrafish embryos along with the accumulation of ROS, eventually resulting in neuroapoptosis in the developing embryonic brain. In conclusion, neurodevelopmental toxicity was caused by oxidative stress-induced apoptosis in zebrafish embryos following abamectin exposure. Full article

This study aimed to evaluate the effect of leukocyte and platelet-rich plasma (L-PRP) on the pulpal healing process following immediate and intentionally delayed tooth replantation in mice. After the maxillary first molars of 3-week-old mice were extracted, the teeth were immersed for 1 min [immediate reimplantation (IR)] or 30 min [intentionally delayed reimplantation (IDR)] in phosphate-buffered saline (PBS) solution. The alveolar socket was filled with or without 1.5 μL of L-PRP [experimental or control groups (EG or CG)] followed by tooth replantation. Samples were collected from day 1 to week 4 after the operation, processed for histology, and evaluated by immunohistochemistry for Nestin and Ki-67 expression. Quantitative analysis revealed positive Nestin staining during pulpal healing in the EG at week 1 following IR and week 2 following IDR. Hard tissue deposition was significantly increased in the EG after IR at week 2. Cell proliferation was higher in the EG compared with that in the CG at week 1 and significantly decreased in the coronal pulp of the EG after the IDR at week 2. Our data suggest that treatment with L-PRP may have a positive effect on pulpal healing, even in teeth replanted after an extended extra-oral period. Full article

Direct pro-neural reprogramming is a conversion of differentiated somatic cells to neural cells without an intermediate pluripotency stage. It is usually achieved via ectopic expression (EE) of certain transcription factors (TFs) or other reprogramming factors (RFs). Determining the transcriptional changes (TCs) caused by particular RFs in a given cell line enables an informed approach to reprogramming initiation. Here, we characterized TCs in the human fibroblast cell line LF1 on the 5th day after EE of the single well-known pro-neural RFs NGN2, ASCL1, SOX2, and MSI1. As assessed by expression analysis of the bona fide neuronal markers nestin and beta-III tubulin, all four RFs initiated pro-neuronal phenotype conversion; analysis by RNA-seq revealed striking differences in the resulting TCs, although some pathways were overlapping. ASCL1 and SOX2 were not sufficient to induce significant pro-neural phenotype switches using our EE system. NGN2 induced TCs indicative of cell phenotype changes towards neural crest cells, neural stem cells, mature neurons, as well as radial glia, astrocytes, and oligodendrocyte precursors and their mature forms. MSI1 mainly induced a switch towards early stem-like cells, such as radial glia. Full article