Human Cells Research Articles

Unveiling the Neurodegenerative Alterations through Oral Stem Cells.

Parkinson's disease (PD) is a neurodegenerative condition characterized by the progressive and selective loss of dopaminergic (DAergic) neurons in the midbrain. The replacement of neuromelanin (NM)-containing DAergic neurons in the substantia nigra and the enhancement of NM concentration could offer a promising and safe approach to treating PD symptoms. The objective of this study was to investigate and compare the potential of human periapical-cysts mesenchymal stem cells (hPCy-MSCs) and dental pulp stem cells (DPSCs) to differentiate into DAergic NM-producing neurons and to generate functional 3-dimensional (3D) midbrain-like organoids in vitro. We assessed the changes in morphology and behavior of neuron-like cells (NLCs) as well as the expression of molecular markers characterizing the DAergic neurons. Furthermore, we observed electrically active and functionally mature DAergic neurons by means of electrophysiological assays, NM dosage assays, and the quantification of dopamine release by high-performance liquid chromatography. Our results demonstrate for the first time that both hPCy-MSCs and DPSCs are capable of differentiating into NLCs, further confirmed by the increase in lactate levels in the medium of cells exposed to neurogenic conditions. Importantly, we have induced such NLCs to further differentiate into functional DAergic NM-producing neurons. Finally, 3D midbrain-like organoids have been produced from oral stem cells: they appear as neurosphere-like structures diffusely expressing the neural marker β-III tubulin and containing NM-like granules. Our findings open up a novel and fascinating opportunity to rethink oral stem cells, and the derived 3D disease models, as a strategic and reliable tool for unveiling the neurodegenerative alterations.

A novel injectable boron doped-mesoporous nano bioactive glass loaded-alginate composite hydrogel as a pulpotomy filling biomaterial for dentin regeneration

BackgroundDifferent materials have been used as wound dressings after vital pulp therapies. Some of them have limitations such as delayed setting, difficult administration, slight degree of cytotoxicity, crown discoloration and high cost. Therefore, to overcome these disadvantages, composite scaffolds have been used in regenerative dentistry. This study aims to construct and characterize the physicochemical behavior of a novel injectable alginate hydrogel loaded with different bioactive glass nanoparticles in various concentrations as a regenerative pulpotomy filling material.MethodsAlginate hydrogels were prepared by dissolving alginate powder in alcoholic distilled water containing mesoporous bioactive glass nanoparticles (MBG NPs) or boron-doped MBG NPs (BMBG NPs) at 10 and 20 wt% concentrations. The mixture was stirred and incubated overnight in a water bath at 50 0 C to ensure complete solubility. A sterile dual-syringe system was used to mix the alginate solution with 20 wt% calcium chloride solution, forming the hydrogel upon extrusion. Then, constructed hydrogel specimens from all groups were characterized by FTIR, SEM, water uptake percentage (WA%), bioactivity and ion release, and cytotoxicity. Statistical analysis was done using One-Way ANOVA test for comparisons between groups, followed by multiple pairwise comparisons using Bonferroni adjusted significance level (p < 0.05).ResultsAlginate/BMBG loaded groups exhibited remarkable increase in porosity and pore size diameter [IIB1 (168), IIB2 (183) (µm)]. Similarly, WA% increased (~ 800%) which was statistically significant (p < 0.05). Alginate/BMBG loaded groups exhibited the strongest bioactive capability displaying prominent clusters of hydroxyapatite precipitates on hydrogel surfaces. Ca/P ratio of precipitates in IIA2 and IIB1 (1.6) were like Ca/P ratio for stoichiometric pure hydroxyapatite (1.67). MTT assay data revealed that the cell viability % of human gingival fibroblast cells have declined with increasing the concentration of both powders and hydrogel extracts in all groups after 24 and 48 h but still higher than the accepted cell viability % of (˃70%).ConclusionsThe outstanding laboratory performance of the injectable alginate/BMBGNPs (20 wt%) composite hydrogel suggested it as promising candidate for pulpotomy filling material potentially enhancing dentin regeneration in clinical applications.

Non-natural MUC1 Glycopeptide Homogeneous Cancer Vaccine with Enhanced Immunogenicity and Therapeutic Activity.

Glycopeptides derived from the glycoprotein mucin-1 (MUC1) have shown potential as tumor-associated antigens for cancer vaccine development. However, their low immunogenicity and non-selective conjugation to carriers present significant challenges for the clinical efficacy of MUC1-based vaccines. Here, we introduce a novel vaccine candidate based on a structure-guided design of an artificial antigen derived from MUC1 glycopeptide. This engineered antigen contains two non-natural amino acids and has an α-S-glycosidic bond, where sulfur replaces the conventional oxygen atom linking the peptide backbone to the sugar N-acetylgalactosamine. The glycopeptide is then specifically conjugated to the immunogenic protein carrier CRM197 (Cross-Reactive Material 197), a protein approved for human use. Conjugation involves selective reduction and re-bridging of a disulfide in CRM197, allowing the attachment of a single copy of MUC1. This strategy results in a chemically defined vaccine while maintaining both the structural integrity and immunogenicity of the protein carrier. The vaccine elicits a robust Th1-like immune response in mice and generates antibodies capable of recognizing human cancer cells expressing tumor-associated MUC1. When tested in mouse models of colon adenocarcinoma and pancreatic cancer, the vaccine is effective both as a prophylactic and therapeutic use, significantly delaying tumor growth. In therapeutic applications, improved outcomes were….

The impact of photobiomodulation on angiogenic differentiation of two different dental derived stem cells using two irradiation protocols: an in vitro investigation

The present study aimed to compare the effect of photobiomodulation with different energy densities on the angiogenic differentiation of human periodontal ligament stem cells (hPDLSCs) and stem cells from human exfoliated deciduous teeth (SHED). Photobiomodulation therapy with a 660 nm diode laser (2.4 J/cm2 and 3.9 J/cm2) on two consecutive days post-culture was applied to two types of stem cells (hPDLSCs and SHED). The Quantitative Real-time Polymerase Chain Reaction (RT-qPCR) test was undertaken to investigate Vascular Endothelial Growth Factor-A (VEGF-A) and Angiopoietin I (ANG-I) genes on days 1, 3, 5, 7, and 10 after the first session of laser application. The 4′,6-diamidino-2-phenylindole (DAPI) staining and Methyl Thiazolyl Tetrazolium (MTT) test were conducted on days 1, 3, and 5 after the first session of laser application, to assess the cell viability. The Two-way ANOVA with Tukey post hoc test was used to analyze the outcomes of the MTT and RT-qPCR tests. The results of the MTT and DAPI convergently illustrated that the groups receiving photobiomodulation with 2.4 J/cm2 had higher cell viability compared to 3.9 J/cm2. All experimental groups showed an upregulation of VEGF-A and ANG-I gene expression from day 1 to 5, followed by a downregulation from day 5 to 10. The groups with cultured hPDLSCs and SHED receiving photobiomodulation using 2.4 J/cm2 had the most amounts of VEGF-A and ANG-I gene expression on day 5, respectively. In conclusion, the 660 nm mediated photobiomodulation therapy of cultured SHED and hPDLSCs with 2.4 J/cm2 energy density may be associated with higher angiogenic differentiation (the expression of VEGF-A and ANG-I) as well as higher cell viability compared to the photobiomodulation therapy with 3.9 J/cm2.

The protective effects of apelin-13 in HIV-1 tat- induced macrophage infiltration and BBB impairment

ABSTRACT Impairment of the blood – brain barrier (BBB) and subsequent inflammatory responses contribute to the development of human immunodeficiency virus (HIV)-1-associated neurocognitive disorders (HAND). Apelin-13, the most abundant member of the apelin family, acts as the ligand of the angiotensin receptor-like 1 (APJ). However, its pharmacological function in HAND and its underlying mechanism are unknown. In the current study, we report that the presence of HIV-1 Tat reduced the levels of Apelin-13 and APJ in the cortex tissue of mice. Importantly, Apelin-13 preserved BBB integrity against HIV-1 Tat in mice by increasing the expression of the tight junction protein zonula occludens-1 (ZO-1) and occludin. Interestingly, increased macrophage infiltration, indicated by elevated CD68-positive staining was observed in the cortex after stimulation with HIV-1, which was mitigated by the administration of Apelin-13. Correspondingly, Apelin-13 reduced the expression of monocyte chemoattractant protein-1; (MCP-1). An in vitro two-chamber and two-cell trans-well assay demonstrated that HIV-1 Tat challenge significantly promoted macrophage migration, which was notably attenuated by the introduction of Apelin-13. Accordingly, treatment with Apelin-13 restored the HIV-1 Tat-induced reduction of occludin and ZO-1, while preventing the upregulation of MCP-1 in human brain microvascular endothelial cells (HBMVECs). Our results suggest that Apelin-13 may reduce macrophage infiltration into brain tissues and mitigate BBB dysfunction in patients with HAND.

Antithrombotic revascularization strategy of bioengineered liver using a biomimetic polymer.

A bioengineered liver has the potential to save patients with end-stage liver disease, and a three-dimensional decellularized scaffold is a promising approach for practical use. The main challenge in bioengineered liver transplantation is thrombogenicity during blood perfusion. We aimed to apply a novel antithrombotic polymer to revascularize liver scaffolds and evaluate the thrombogenicity and biosafety of the polymer-treated scaffolds. A biomimetic polymer, 2-metacryloyloxyethyl phosphorylcholine (MPC) was prepared for modification of the extracellular matrix (ECM) in liver scaffolds. The polymer was injected into the rat liver scaffolds' portal vein (PV) and could extensively react to the vessel walls. In an ex-vivo blood perfusion experiment, we demonstrated significantly less platelet deposition in the polymer-treated scaffolds than non-treated or re-endothelialized scaffolds with human umbilical endothelial cells (HUVECs). In the heterotopic transplantation model, liver volume was better maintained in the polymer-treated groups and platelet deposition was suppressed in these groups. Additionally, the polymer-treated liver scaffolds maintained the metabolic function of the recellularized rat primary hepatocytes during perfusion culture. The MPC polymer treatment efficiently suppressed thrombus formation during blood perfusion in liver scaffolds and maintained the function of recellularized hepatocytes. Revascularizing liver scaffolds using this polymer is a promising approach for bioengineered liver transplantation.

Clusterin-carrying extracellular vesicles derived from human umbilical cord mesenchymal stem cells restore the ovarian function of premature ovarian failure mice through activating the PI3K/AKT pathway

BackgroundEmerging evidence has highlighted the therapeutic potential of human umbilical cord mesenchymal stem cells (UC-MSCs) in chemotherapy-induced premature ovarian failure (POF). This study was designed to investigate the appropriate timing and molecular mechanism of UC-MSCs treatment for chemotherapy-induced POF.MethodsOvarian structure and function of mice were assessed every 3 days after injections with cyclophosphamide (CTX) and busulfan (BUS). UC-MSCs and UC-MSCs-derived extracellular vesicles (EVs) were infused into mice via the tail vein, respectively. Ovarian function was analyzed by follicle counts, the serum levels of hormones and ovarian morphology. The apoptosis and proliferation of ovarian granulosa cells were analyzed in vitro and in vivo. Label-free quantitative proteomics was used to detect the differentially expressed proteins in UC-MSC-derived EVs.ResultsAfter CTX/BUS injection, we observed that the ovarian function of POF mice was significantly deteriorated on day 9 after CTX/BUS infusion. TUNEL assay indicated that the number of apoptotic cells in the ovaries of POF mice was significantly higher than that in normal mice on day 3 after CTX/BUS injection. Transplantation of UC-MSCs on day 6 after CTX/BUS injection significantly improved ovarian function, enhanced proliferation and inhibited apoptosis of ovarian granulosa cells, whereas the therapeutic effect of UC-MSCs transplantation decreased on day 9, or day 12 after CTX/BUS injection. Moreover, EVs derived from UC-MSCs exerted similar therapeutic effects on POF. UC-MSCs-derived EVs could activate the PI3K/AKT signaling pathway and reduce ovarian granulosa cell apoptosis. Quantitative proteomics analysis revealed that clusterin (CLU) was highly expressed in the EVs of UC-MSCs. The supplementation of CLU proteins prevented ovarian granulosa cells from chemotherapy-induced apoptosis. Further mechanistic analysis showed that CLU-knockdown blocked the PI3K/AKT signaling and reversed the protective effects of UC-MSCs-derived EVs.ConclusionsAdministration of UC-MSCs and UC-MSCs-derived EVs on day 6 of CTX/BUS injection could effectively improve the ovarian function of POF mice. UC-MSCs-derived EVs carrying CLU promoted proliferation and inhibited apoptosis of ovarian granulosa cells through activating the PI3K/AKT pathway. This study identifies a previously unrecognized molecular mechanism of UC-MSCs-mediated protective effects on POF, which pave the way for the use of cell-free therapeutic approach for POF.Graphical

Nucleolus and centromere Tyramide Signal Amplification-Seq reveals variable localization of heterochromatin in different cell types

Genome differential positioning within interphase nuclei remains poorly explored. We extended and validated Tyramide Signal Amplification (TSA)-seq to map genomic regions near nucleoli and pericentric heterochromatin in four human cell lines. Our study confirmed that smaller chromosomes localize closer to nucleoli but further deconvolved this by revealing a preference for chromosome arms below 36-46 Mbp in length. We identified two lamina associated domain subsets through their differential nuclear lamina versus nucleolar positioning in different cell lines which showed distinctive patterns of DNA replication timing and gene expression across all cell lines. Unexpectedly, active, nuclear speckle-associated genomic regions were found near typically repressive nuclear compartments, which is attributable to the close proximity of nuclear speckles and nucleoli in some cell types, and association of centromeres with nuclear speckles in human embryonic stem cells (hESCs). Our study points to a more complex and variable nuclear genome organization than suggested by current models, as revealed by our TSA-seq methodology.

Association of Wild-Type TP53 with Downregulation of Lovastatin Sensitivity in Human Non-Small Cell Lung Cancer Cells

Association of Wild-Type TP53 with Downregulation of Lovastatin Sensitivity in Human Non-Small Cell Lung Cancer Cells

Establishment and characterization of a novel human gallbladder cancer cell line, GBC-X1

In this study, we successfully established a novel gallbladder cancer cell line, designated as GBC-X1, derived from a primary tumor of a gallbladder cancer patient. By comprehensively analyzing the cell line’s phenotype, molecular characteristics, biomarkers, and histological characteristics, we confirmed that GBC-X1 serves as a valuable model for investigating the pathogenesis of gallbladder cancer and developing therapeutic agents. GBC-X1 has been continuously cultured for one year, with over 60 stable passages. Morphologically, GBC-X1 exhibits typical features of epithelial tumors. The population doubling time of GBC-X1 is 32 h. STR analysis validated a high consistency between GBC-X1 and the patient’s primary tumor. Karyotype analysis revealed an abnormal hypertetraploid karyotype for GBC-X1, characterized by representative karyotypes of 98, XXXX del (4) p (12) del (5) p (21) der (10). Under suspension culture conditions, GBC-X1 efficiently forms tumor balls, while subcutaneous inoculation of GBC-X1 cells into NXG mice leads to xenograft formation with a rate of 80%. Drug sensitivity testing demonstrated that GBC-X1 is resistant to oxaliplatin and sensitive to 5-FU, gemcitabine, and paclitaxel. Immunohistochemistry revealed positive expression of CK7, CK19, E-cadherin, MMP-2, CD44, SOX2, and TP53 in GBC-X1 cells, weak positive expression of Vimentin, and a Ki67 positive rate of 35%. Our research highlights GBC-X1 as a novel gallbladder cancer cell line and emphasizes its potential as an effective experimental model for investigating the pathogenesis of gallbladder cancer and drug development.

Phosphoinositide 3-Kinase Inhibitors from Gladiolus segetum Ker-Gawl Corms Supported by Network Pharmacology.

Gladiolus segetum Ker-Gawl corms total extract exhibited remarkable in vitro anti-proliferative effects against panel of cancer cell lines; including human colon carcinoma (Caco-2), human breast cancer (MCF7) and hepatocellular carcinoma (HepG2) cell lines with IC50 values of 7.4, 9.1 and 11.2 µg/ml, respectively. The total ethanolic extract of G. segetum Ker-Gawl corms was subjected to untargeted metabolomics profiling using LC-HR-ESI-MS, which revealed the presence of various clusters of phytoconstituents as triterpenes, anthraquinones, flavonoids and phenolic derivatives. Network pharmacology study was performed for all identified compounds, the formed networks identified 73 intersected genes. The diagrammatic illustration of the top pathways revealed that phosphoinositide 3-kinase (PI3K) gene is the effective dominant gene in the top four KEGG pathways. Upon molecular docking and molecular dynamics investigation, kaempferol-3-O-glucopyranoside was suggested to be key anticancer metabolite. Interestingly, cytotoxic investigation of this compound revealed potential activity against the tested cancer cell lines (Caco-2, MCF7 and HepG2) with IC50 values of 6.2, 8.5 and 9.3 µg/ml, respectively. The present study highlighted the potential of G. segetum Ker-Gawl as a promising source of interesting anticancer scaffolds.

Turkish coffee has an antitumor effect on breast cancer cells in vitro and in vivo

BackgroundBreast cancer is the most diagnosed cancer in women. Its pathogenesis includes several pathways in cancer proliferation, apoptosis, and metastasis. Some clinical data have indicated the association between coffee consumption and decreased cancer risk. However, little data is available on the effect of coffee on breast cancer cells in vitro and in vivo.MethodsIn our study, we assessed the effect of Turkish coffee and Fridamycin-H on different pathways in breast cancer, including apoptosis, proliferation, and oxidative stress. A human breast cancer cell line (MCF-7) was treated for 48 h with either coffee extract (5% or 10 v/v) or Fridamycin-H (10 ng/ml). Ehrlich solid tumors were induced in mice for in vivo modeling of breast cancer. Mice with Ehrlich solid tumors were treated orally with coffee extract in drinking water at a final concentration (v/v) of either 3%, 5%, or 10% daily for 21 days. Protein expression levels of Caspase-8 were determined in both in vitro and in vivo models using ELISA assay. Moreover, P-glycoprotein and peroxisome proliferator-activated receptor gamma (PPAR-γ) protein expression levels were analyzed in the in vitro model. β-catenin protein expression was analyzed in tumor sections using immunohistochemical analysis. In addition, malondialdehyde (MDA) serum levels were analyzed using colorimetry.ResultsBoth coffee extract and Fridamycin-H significantly increased Caspase-8, P-glycoprotein, and PPAR-γ protein levels in MCF-7 cells. Consistently, all doses of in vivo coffee treatment induced a significant increase in Caspase-8 and necrotic zones and a significant decrease in β- catenin, MDA, tumor volume, tumor weight, and viable tumor cell density.ConclusionThese findings suggest that coffee extract and Fridamycin-H warrant further exploration as potential therapies for breast cancer.

298 Botanicals reduce osteoarthritic-like inflammation influencing the endocannabinoid system in vitro

Abstract Osteoarthritis (OA) is a prevalent joint disease in dogs. Botanicals (BOT) interacting with the endocannabinoid system (ECS) can reduce pain, inflammation, and other OA-related processes, and could become potential treatments for pets. This study investigates various BOT in vitro for their potential to reduce OA-like inflammation, oxidative stress, and apoptosis, and influence the expression of ECS enzymes or receptors. Capsicum oleoresin (10% capsaicinoids) at 100 ppm, cinnamaldehyde at 5 ppm, turmeric extract (95% curcuminoids) at 10 ppm, and eugenol at 10 ppm were selected for their documented ability to interact with the ECS and used as treatments. The efficacy of these BOT was assessed by pre-treating human chondrosarcoma cells (SW1353 - ATCC HTB-94) with BOT for 2 h before exposure to interleukin (IL)1β and tumor necrosis factor (TNF) α for 24 h. Following the challenge, the cells and supernatants were collected. Then, qPCR on selected markers and IL8 quantification with ELISA were conducted. Additionally, to assess antioxidant and anti-apoptotic capabilities, BOT-treated cells were exposed to menadione for 1 h to measure reactive oxygen species (ROS) and for 6 h to induce apoptosis and evaluate chondrocyte viability. Data were analyzed using one-way ANOVA with Tukey’s multiple comparisons (n = 3; P &amp;lt; 0.05) and reported in this work compared against the positive control. Under inflammatory conditions, SW1353 cells exhibited a significant increase in IL6 (234-fold; P = 0.0333) and IL8 (4,500-fold; P = 0.0395) expression, and MMP1 (13-fold, P = 0.0052), MMP3 (40-fold; P = 0.0474), and MMP13 (26-fold; P = 0.0260) expression. Among the different BOTs, only cinnamaldehyde effectively countered IL6 and IL8 upregulation, reducing their expression to 0 for IL6 and by 4,400-fold for IL8 (P = 0.0086). Additionally, it reduced MMP3 by 40-fold (P = 0.0242) and MMP13 by 24-fold (P = 0.0275), while eugenol significantly reduced MMP3 by 36-fold (P = 0.0250). Only cinnamaldehyde significantly decreased IL8 secretion by 86% (P &amp;lt; 0.0001), and all tested botanicals reduced chondrocyte apoptosis (P &amp;lt; 0.0001). Eugenol and turmeric extract exhibited significant antioxidant effects, reducing ROS levels by 70% compared with the positive control (P &amp;lt; 0.05). Concerning ECS response to the inflammatory challenge, MAGL levels increased by 4.2-fold (P = 0.0253), with cinnamaldehyde significantly reducing its mRNA expression by 4-fold (P = 0.0078). PPAR-γ expression showed a numerical decrease of 0.7-fold under inflammatory conditions, but cinnamaldehyde and capsicum oleoresin upregulated its mRNA expression by 2.3-fold (P = 0.0162) and 1.3-fold (P = 0.0006), respectively. Cinnamaldehyde also reduced DAGLα expression by 0.3-fold (P = 0.0079), and upregulated CBR2 and NRF2 mRNA expression by 7-fold (P &amp;lt; 0.0001) and 1.6-fold (P = 0.0295) respectively, while other botanicals had no effects. In conclusion, the tested BOTs displayed promising properties to mitigate OA in vitro, with cinnamaldehyde proving to be the most promising one. These findings suggest the potential of BOT active on ECS as a novel treatment for canine OA, prompting further in vivo research for comprehensive characterization.

Lipopolymer/siRNA Nanoparticles Targeting the Signal Transducer and Activator of Transcription 5A Disrupts Proliferation of Acute Lymphoblastic Leukemia.

The therapeutic potential of small interfering RNAs (siRNAs) in gene-targeted treatments is substantial, but their suboptimal delivery impedes widespread clinical applications. Critical among these is the inability of siRNAs to traverse the cell membranes due to their anionic nature and high molecular weight. This limitation is particularly pronounced in lymphocytes, which pose additional barriers due to their smaller size and scant cytoplasm. Addressing this, we introduce an innovative lipid-conjugated polyethylenimine lipopolymer platform, engineered for delivery of therapeutic siRNAs into lymphocytes. This system utilizes the cationic nature of the polyethylenimine for forming stable complexes with anionic siRNAs, while the lipid component facilitates cellular entry of siRNA. The resulting lipopolymer/siRNA complexes are termed lipopolymer nanoparticles (LPNPs). We comprehensively profiled the efficacy of this platform in human peripheral blood mononuclear cells (PBMCs) as well as in vitro and in vivo models of acute lymphoblastic leukemia (ALL), emphasizing the inhibition of the oncogenic signal transducer and activator of transcription 5A (STAT5A) gene. The lipopolymers demonstrated high efficiency in delivering siRNA to ALL cell lines (RS4;11 and SUP-B15) and primary patient cells, effectively silencing the STAT5A gene. The resultant gene silencing induced apoptosis and significantly reduced colony formation in vitro. Furthermore, in vivo studies showed a significant decrease in tumor volumes without causing substantial toxicity. The lipopolymers did not induce the secretion of proinflammatory cytokines (IL-6, TNF-α, and INF-γ) in PBMCs from healthy volunteers, underscoring their immune safety profile. Our observations indicate that LPNP-based siRNA delivery systems offer a promising therapeutic approach for ALL in terms of both safety and therapeutic efficacy.

Modeling Lewy body disease with SNCA triplication iPSC-derived cortical organoids and identifying therapeutic drugs.

Aggregated α-synuclein (α-SYN) proteins, encoded by the SNCA gene, are hallmarks of Lewy body disease (LBD), affecting multiple brain regions. However, the specific mechanisms underlying α-SYN pathology in cortical neurons, crucial for LBD-associated dementia, remain unclear. Here, we recapitulated α-SYN pathologies in human induced pluripotent stem cells (iPSCs)-derived cortical organoids generated from patients with LBD with SNCA gene triplication. Single-cell RNA sequencing, combined with functional and molecular validation, identified synaptic and mitochondrial dysfunction in excitatory neurons exhibiting high expression of the SNCA gene, aligning with observations in the cortex of autopsy-confirmed LBD human brains. Furthermore, we screened 1280 Food and Drug Administration-approved drugs and identified four candidates (entacapone, tolcapone, phenazopyridine hydrochloride, and zalcitabine) that inhibited α-SYN seeding activity in real-time quaking-induced conversion assays with human brains, reduced α-SYN aggregation, and alleviated mitochondrial dysfunction in SNCA triplication organoids and excitatory neurons. Our findings establish human cortical LBD models and suggest potential therapeutic drugs targeting α-SYN aggregation for LBD.

The alpha7 nicotinic acetylcholine receptor agonist PHA 568487 dampens inflammation in PBMCs from patients with newly discovered coronary artery disease.

The alpha7 nicotinic acetylcholine receptor (α7nAChR) regulates inflammation in experimental models and is expressed in human peripheral blood mononuclear cells (PBMCs) and in human atherosclerotic plaques. However, its role in regulating inflammation in patients with cardiovascular disease is unknown. This study aims to investigate whether α7nAChR stimulation can reduce the inflammatory response in PBMCs from patients with newly diagnosed coronary artery disease (CAD). Human PBMCs, extracted from patients with verified CAD (n=38) and control participants with healthy vessels (n=38), were challenged in vitro with lipopolysaccharide (LPS) in combination with the α7nAChR agonist PHA 568487. Cytokine levels of the supernatants were analyzed using a multiplex immunoassay. Patients in the CAD group were re-examined after 6 months. The immune response to LPS did not differ between PBMCs from control group and CAD patients. α7nAChR stimulation decreased TNFα in both control participants and CAD patients. The most pronounced effect of α7nAChR stimulation was observed in CAD patients at their first visit, where 15 of 17 cytokines were decreased (IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-10, IL-12 (p70), IL-17A, G-CSF, GM-CSF, IFN-γ, MCP-1, MIP-1β and TNFα). In conclusion, stimulation with α7nAChR agonist PHA 568487 dampens the inflammatory response in human PBMCs. This finding suggests that the anti-inflammatory properties of the α7nAChR may have a role in treating CAD.

Metformin Attenuates Partial Epithelial-Mesenchymal Transition in Salivary Gland Inflammation via PI3K/Akt/GSK3β/Snail Signaling Axis.

Chronic inflammation in the salivary glands (SG) often triggers epithelial-mesenchymal transition (EMT), leading to the loss of acinar function and promoting fibrosis. This study explores the role of Metformin in mitigating partial EMT in SG inflammation. In vitro, human salivary gland epithelial cells (hSGECs) were treated with lipopolysaccharide (LPS) and Metformin. EMT markers and the PI3K/Akt/GSK3β/Snail signaling axis were assessed using RNA-seq and Western blot analysis. In vivo, a Wharton's duct ligation rat model was employed to mimic chronic sialadenitis (CS). Nine Wistar rats were randomly divided into three groups: Control, Ligation and Ligation + Metformin groups, with three rats per group. After ductal ligation, the Ligation + Metformin group received 100mg/kg of Metformin via intragastric administration, while the Control and Ligation groups received an equivalent saline every 24h. Histological analysis, immunohistochemical and immunofluorescence staining were conducted to evaluate acinar morphology, EMT, and the PI3K/Akt/GSK3β/Snail signaling axis. The results showed that in CS tissues, atrophied acinar cells underwent partial EMT. In vitro, Metformin reversed LPS-induced EMT in hSGECs. RNA-seq and Western blot revealed that Metformin achieved this effect by targeting the PI3K/Akt/GSK3β/Snail signaling axis (P < 0.01). In ductal ligation models, Metformin treatment restored ligation-induced acinar damage and functional loss (P < 0.01). Further histological evidence supported that Metformin mitigated EMT by inhibiting inflammatory activation of PI3K/Akt/GSK3β/Snail signaling axis (P < 0.01). In conclusion, Metformin alleviates partial EMT in SG inflammation by targeting the PI3K/Akt/GSK3β/Snail signaling axis, highlighting its potential as a therapeutic strategy for SG inflammation.

Roles and action mechanisms of NRIP1 in pre-eclampsia.

Pre-eclampsia (PE) is characterized by the onset of hypertension and proteinuria during pregnancy. Here, we aimed to explore the functions of nuclear receptor-interacting protein 1 (NRIP1) in PE mice and human placental JEG-3 cells. We evaluated its effects on JEG-3 cell proliferation, apoptosis, invasion, and inflammatory response and regulation of Wnt/β-catenin pathway. NRIP1 levels in human serum and placental tissues, JEG-3 cells, and mouse models were assessed via quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and western blotting. JEG-3 cell growth, apoptosis, migration, and invasion were evaluated via 5-ethynyl-2'-deoxyuridine, flow cytometry, and transwell assays. Levels of the inflammatory factors, matrix metalloproteinase (MMP)-2, tumor necrosis factor (TNF)-α, and interleukin (IL)-6, were determined via enzyme-linked immunosorbent assay. Wnt/β-catenin pathway was assessed via western blotting and qRT-PCR. Systolic blood pressure and proteinuria were measured using the non-invasive tail cuff method and Coomassie brilliant blue assay, respectively. TdT-mediated dUTP nick-end labeling assay was used to assess cell apoptosis in the placental tissues of PE mice. NRIP1 levels were upregulated in the serum and placental tissues of patients with PE. In vitro experiments revealed that NRIP1-small interfering RNA (siRNA) increased the cell viability, migration, and invasion and reduced the cell apoptosis compared to the control siRNA. Moreover, NRIP1-siRNA activated the Wnt/β-catenin signaling pathway, as indicated by the increased Wnt3a, β-catenin, p-glycogen synthase kinase-3β, c-Myc, and cyclin D1 levels. Levels of the inflammatory factors, IL-6, TNF-α, and MMP-2, were decreased in the NRIP1-siRNA-treated group. Notably, NRIP1 downregulation improved the PE-like symptoms, inhibited the inflammatory responses, and reduced apoptosis in PE mice. This study revealed the crucial roles of NRIP1 in PE. Our findings revealed that NRIP1 downregulation relieved PE symptoms by inhibiting cell proliferation, migration, and invasion via the Wnt/β-catenin pathway, thus providing a novel candidate for PE treatment.

Transcriptomic Analysis and Finding of Potential Key mRNA Expression Profile in Human Cumulus Cells During in Vitro Culture and Different Passages Based on Integrated Bioinformatics Analysis.

This study leveraged microarray datasets to investigate differentially expressed genes (DEGs) in cumulus cells and their relevance in predicting the successful implantation of embryos in human in-vitro fertilization procedures. The microarray data were obtained from the GEO database, encompassing samples of cumulus cells during in vitro culture and different passages. To ensure data consistency, inter-batch normalization was performed, and Principal Component Analysis (PCA) was applied to assess the impact of normalization on sample group clustering. The integrated dataset included samples from cumulus cells during in vitro culture, comprising 17,662 genes. Utilizing the "limma" software package, 1906 DEGs were identified, with 437 genes downregulated and 589 genes upregulated in the cumulus cells of infertility cases, while 748 genes were upregulated, and 1317 genes were downregulated in cumulus cells of successful implantation cases. Functional enrichment analysis utilized Gene Ontology, Metascape, and Kyoto Encyclopedia of Genes and Genomes pathway enrichment tools. Biological processes and molecular functions were enriched, including protein targeting, mRNA processing, and molecular binding among the identified DEGs. Furthermore, target prediction and functional enrichment analysis of microRNAs (miRNAs) revealed 25 key genes and 13 relevant miRNAs were identified. Notably, hsa-miR-149, hsa-miR23b, hsa-miR-877, hsa-miR593, hsa-miR-18a, hsa-miR25, hsa-miR185, mmu-miR-207, hsa-miR425, hsa-miR214, hsa-miR-129, hsa-miR-629, and hsa-miR-194 emerged as the most prominent miRNAs with potential regulatory roles in successful embryo implantation. This comprehensive analysis provides valuable insights into the molecular mechanisms underlying embryo implantation, offering potential targets for further research and therapeutic interventions in assisted reproductive technologies.

Telomerase reverse transcriptase protects against diabetic kidney disease by promoting AMPK/PGC-1a-regulated mitochondrial energy homeostasis

Disordered glucose and lipid metabolism, coupled with disturbed mitochondrial bioenergetics, are pivotal in the initiation and development of diabetic kidney disease (DKD). While the essential role of telomerase reverse transcriptase (TERT) in regulating mitochondrial function in the cardiovascular system has been recognized, its specific function in maintaining mitochondrial homeostasis in DKD remains unclear. This study aimed to explore how TERT regulates mitochondrial function and the underlying mechanisms. In vitro, human renal proximal tubular HK-2 cells exposed to high glucose/high fat (HG/HF) presented significant downregulation of TERT and AMPK dephosphorylation. This led to decreased ATP production, altered NAD+/NADH ratios, reduced mitochondrial complex activities, increased mitochondrial dysfunction, lipid accumulation, and reactive oxygen species (ROS) production. Knockdown of TERT (si-TERT) further exacerbated mitochondrial dysfunction, decreased mitochondrial membrane potential, and lowered levels of cellular oxidative phosphorylation and glycolysis, as determined via a Seahorse X24 flux analyzer. Conversely, mitochondrial dysfunction was significantly alleviated after pcDNA-TERT plasmid transfection and adeno-associated virus (AAV) 9-TERT gene therapy in vivo. Notably, treatment with an AMPK inhibitor, activator, and si-PGC-1a (peroxisome proliferator-activated receptor γ coactivator-1α), resulted in mitochondrial dysfunction and decreased expression of genes related to energy metabolism and mitochondrial biogenesis. Our findings reveal that TERT protects mitochondrial function and homeostasis by partially activating the AMPK/PGC-1a signaling pathway. These results establish a crucial foundation for understanding TERT's critical role inmitochondrial regulation and its protective effect on DKD.