Differences In Ability Research Articles

Cannabidiol alleviates LPS-inhibited odonto/osteogenic differentiation in human dental pulp stem cells invitro.

Cannabidiol (CBD), derived from the Cannabis sativa plant, exhibits benefits in potentially alleviating a number of oral and dental pathoses, including pulpitis and periodontal diseases. This study aimed to explore the impact of CBD on several traits of human dental pulp stem cells (hDPSC), such as their proliferation, apoptosis, migration and odonto/osteogenic differentiation. hDPSCs were harvested from human dental pulp tissues. The cells were treated with CBD at concentrations of 1.25, 2.5, 5, 10, 25 and 50 μg/mL. Cell responses in terms of cell proliferation, colony-forming unit, cell cycle progression, cell migration, apoptosis and odonto/osteogenic differentiation of hDPSCs were assessed in the normal culture condition and P. gingivalis lipopolysaccharide (LPS)-induced 'inflammatory' milieus. RNA sequencing and proteomic analysis were performed to predict target pathways impacted by CBD. CBD minimally affects hDPSCs' behaviour under normal culture growth milieu in normal conditions. However, an optimal concentration of 1.25 μg/mL CBD significantly countered the harmful effects of LPS, indicated by the promoting cell proliferation and restoring the odonto/osteogenic differentiation potential of hDPSCs under LPS-treated conditions. The proteomic analysis demonstrated that several proteins involved in cell proliferation and differentiation were upregulated following CBD exposure, including CCL8, CDC42 and KFL5. RNA sequencing data indicated that CBD upregulated the Notch signalling pathway. In an inhibitory experiment, DAPT, a Notch inhibitor, reduced the effect of CBD-rescued LPS-attenuated mineralization in hDPSCs, suggesting that CBD potentially mediates Notch activation to exert its impact on odonto/osteogenic differentiation of hDPSCs. CBD recovers the proliferation and survival of hDPSCs following exposure to LPS. Additionally, we report that CBD-mediated Notch activation effectively restores the odonto/osteogenic differentiation ability of hDPSCs under inflamed conditions. These results underscore the potential role of CBD as a therapeutic option to enhance dentine regeneration.

Differentiation ability of hematopoietic stem cells and mesenchymal stem cells isolated from human peripheral blood

Human hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs) are the major stem cells of the bone marrow and are usually isolated from the peripheral blood. In the present study, we isolated these stem cells by an apheresis method from a donor who was administered granulocyte colony-stimulating factor (G-CSF). In vitro propagation of these stem cells showed a plastic-adherence property expressing CD73 and CD105 surface markers, which is a characteristic feature of MSCs. HSCs are non-adherent cells growing as a suspension culture, expressing CD150, CD133, CD34, and CD45 on their surface, which regulate the quiescence nature, and they derive energy from anaerobic glycolysis. The HSCs grow slowly compared to MSCs, are more viable, and survive for long periods under in vitro conditions, which are due to the expression of telomerase, BCL2, and Notch1 genes. The poor viability of MSCs in the culture due to the prominent expression of apoptotic genes BAX, caspase-3, and caspase-9 leads to rapid apoptosis. This was evident even in cells (astrocytes, osteocytes, and beta cells of the islets of Langerhans) differentiated from HSCs and MSCs, thus highlighting the importance of HSCs, the naive stem cells, in regeneration of tissues.

LL-37 regulates odontogenic differentiation of dental pulp stem cells in an inflammatory microenvironment

BackgroundInflammation often causes irreversible damage to dental pulp tissue. Dental pulp stem cells (DPSCs), which have multidirectional differentiation ability, play critical roles in the repair and regeneration of pulp tissue. However, the presence of proinflammatory factors can affect DPSCs proliferation, differentiation, migration, and other functions. LL-37 is a natural cationic polypeptide that inhibits lipopolysaccharide (LPS) activity, enhances cytokine production, and promotes the migration of stem cells. However, the potential of LL-37 in regenerative endodontics remains unknown. This study aimed to investigate the regulatory role of LL-37 in promoting the migration and odontogenic differentiation of DPSCs within an inflammatory microenvironment. These findings establish an experimental foundation for the regenerative treatment of pulpitis and provide a scientific basis for its clinical application.Materials and methodsDPSCs were isolated via enzyme digestion combined with the tissue block adhesion method and identified via flow cytometry. The impact of LL-37 on the proliferation of DPSCs was evaluated via a CCK-8 assay. The recruitment of DPSCs was assessed through a transwell assay. The mRNA expression levels of inflammatory and aging-related genes were assessed via reverse transcription‒polymerase chain reaction (RT‒PCR), western blotting, and enzyme‒linked immunosorbent assay (ELISA). The odontogenic differentiation of DPSCs was assessed through alkaline phosphatase (ALP) staining, alizarin red staining, and RT‒PCR analysis.ResultsLL-37 has the potential to enhance the migration of DPSCs. In an inflammatory microenvironment, LL-37 can suppress the expression of genes associated with inflammation and aging, such as TNF-α, IL-1β, IL-6, P21, P38 and P53. Moreover, it promotes odontogenic differentiation in DPSCs by increasing ALP activity, increasing calcium nodule formation, and increasing the expression of dentin-related genes such as DMP1, DSPP and BSP.ConclusionThese findings suggest that the polypeptide LL-37 facilitates the migration of DPSCs and plays a crucial role in resolving inflammation and promoting cell differentiation within an inflammatory microenvironment. Consequently, LL-37 has promising potential as an innovative therapeutic approach for managing inflammatory dental pulp conditions.

Affective assemblages of kinship and single mothers’ labour migration from a ‘climate hotspot’

In coastal Bangladesh, ‘affective assemblages of kinship’ produce differential abilities for landless single mothers to migrate to brick kilns, the garment industry, and the Gulf. This group of women who return to their natal homes as a response to violence or abandonment is neglected by anthropologists of kinship and migration. Thinking of assemblages of kinship as open‐ended gatherings enables us to move away from fixed genealogical constructs of patrilineal and virilocal households and to theorize how different people may join or leave a kinship assemblage based on the fluctuations of emotional bonds over time. A rapidly changing socioeconomic landscape has contributed towards the unkinning of returning sisters from failed marriages, shifting filial duties, and matrifocal living. Such ‘divorced’ women make new kin through unofficial romantic partners, with whom they may choose to migrate together as a means to exert emotional agency. Thinking through ‘affective assemblages of kinship’ via single mothers reveals the gendered complexities of rural labour migration – of economic and affective‐sexual‐moral considerations – and how it is contingent on social reproductive support. Such an ethnographic corrective towards reductive explanations of climate‐induced migration and the vulnerability of ‘female‐headed households’ shows the importance of anthropology in understanding complex phenomena in a world undergoing rapid socioenvironmental change.

Taurine Protects Irinotecan-induced Muscle Dysfunction by Modulating Oxidative Stress and Endoplasmic Reticulum Stress in Human Skeletal Muscle Cells.

Irinotecan is a key component of standard first-line treatment for metastatic colorectal cancer. However, irinotecan-induced muscle dysfunction is a contributing factor to cancer cachexia. Here, we present the protective effect of taurine, a conditionally essential amino acid with great antioxidant properties, in attenuating muscle dysfunction induced by irinotecan. Irinotecan (20 μg/ml) was added to human skeletal muscle cells (HSkMCs) with or without pre-treatment of taurine (5 mM). The effects of taurine and irinotecan on the viability, cytotoxicity, and differentiation ability of HSkMC myoblasts were examined. The intracellular reactive oxygen species (ROS) and endoplasmic reticulum stress (ERS) were also monitored. Irinotecan caused cytotoxicity of HSkMCs, while taurine pretreatment increased cell viability and inhibited adenylate kinase release significantly in both myoblasts and myotubes. During differentiation, taurine increased ROS clearance and preserved the myotube differentiation ability impaired by irinotecan. Irinotecan exposure resulted in the up-regulation of CCAAT/enhancer-binding protein homologous protein (CHOP) and glucose-regulated protein 78 (GRP78). Taurine pretreatment could combat such irinotecan-induced ERS. The current in vitro study provides molecular evidence that taurine plays a beneficial role in protecting against irinotecan-induced muscle dysfunction by modulating oxidative stress and endoplasmic reticulum stress.

Negative Emotion Differentiation Promotes Cognitive Reappraisal: Evidence From Electroencephalogram Oscillations and Phase-Amplitude Coupling.

Cognitive reappraisal, an effective emotion regulation strategy, is influenced by various individual factors. Although previous studies have established a link between negative emotion differentiation (NED) and cognitive reappraisal, the underlying neural mechanisms remain largely unknown. Using electroencephalography, this study investigates the influence and neural basis of NED in cognitive reappraisal by integrating aspects of event-related potentials, neural oscillation rhythms, and cross-frequency coupling. The findings revealed that individuals with high NED demonstrated a significant decrease in parietal late positive potential amplitudes during cognitive reappraisal, suggesting enhanced cognitive reappraisal abilities. Moreover, high NED individuals displayed increased γ synchronization, parietal α-γ coupling, and frontal θ-γ coupling when reappraising negative emotions than those with low emotion differentiation ability. Machine learning analysis of these neural indicators highlighted the superior classification and predictive accuracy of multimodal indicators for NED as opposed to unimodal indicators. Overall, this multimodal evidence provides a comprehensive interpretation of the neurophysiological mechanisms through which NED influences cognitive reappraisal and provides preliminary empirical support for personalized cognitive reappraisal interventions to alleviate emotional problems.

Revitalizing Muscle Regeneration: Cocoa Polyphenols Shield Mitochondrial Integrity and Boost Myogenesis Under Oxidative Stress.

In this study, we describe the effect of cocoa polyphenol extract (CPE, from flavanols-rich cocoa) on myogenic differentiation in murine myoblasts (C2C12 cells) exposed to H2O2. The myogenic program was monitored using morphological, ultrastructural, and molecular approaches. Treatment with 100 μM of H2O2 for 1 h decreased cell viability. C2C12 (D1) exposed to H2O2 shows more apoptotic and necrotic cells, and mitochondria appear emptied, with cristae heavily damaged. To evaluate the effect of CPE on myoblast viability and myotube formation, 10 μg/mL of CPE were added 24 h prior to H2O2 treatment and cells were supplemented with fresh CPE every 24 h during differentiation. Supplementation with CPE protected C2C12 myoblasts from H2O2-induced oxidative damage both at early (D1) and late (D6) phases of differentiation, preventing cell death and mitochondrial damage. The number of mitochondria (per area of cell surface) increased 2-fold in both control and in CPE-supplemented and mitochondria in myotubes D6 showed a greater extension of mitochondrial cristae than mitochondria in D1. At D1 and D6 the monolayers showed surface and inner cell features relatively comparable to the untreated control suggesting that CPE supplementation significantly mitigated the effect of H2O2. Preliminary data obtained by the myogenic index (Giemsa staining) suggested that CPE-supplemented cells were partially protected from H2O2-induced myogenesis inhibition. The CPE supplementation seems to preserve the mitochondrial integrity and the myogenic differentiation ability of oxidatively injured C2C12 ensuing further nutraceutical perspectives.

Pathways to health: A longitudinal examination of protective factors in children with and without preschool anxiety.

Preschool anxiety is highly prevalent and well known to predict risk for future psychopathology. The present study explores whether a diagnosis of an anxiety disorder in preschool interacts with (a) social skills and (b) cognitive ability to longitudinally predict psychopathology, two well-known protective factors, among a sample of 207 children measured at preschool (Mage = 4.34 years) and early childhood (Mage = 6.61 years). To assess social skills and cognitive ability, we utilized the Social Skills Rating Scale and the Differential Abilities Scale, respectively. To assess psychopathology, we utilized the parent report of the Preschool Age Psychiatric Assessment. Hierarchical linear regression models revealed significant interactions between both social skills and cognitive ability with preschool anxiety. We observed that social skills protected against emergent psychopathology for both children with and without anxiety, although this association was stronger for children with preschool anxiety. Contrastingly, cognitive ability served as a protective factor against future psychopathology primarily among children without preschool anxiety. Results from this study identify targets for future intervention and inform our understanding of how preschool anxiety, a common disorder among young children, shapes future psychopathology risk in childhood.

Risk analysis and risk prediction of in-hospital heart failure in patients with acute myocardial infarction after emergency intervention surgery

BackgroundPercutaneous coronary intervention (PCI) can rapidly open the culprit vessels of acute myocardial infarction (AMI) and save ischemic myocardium, but it is often accompanied by a variety of complications, including heart failure (HF).Please check if the article title is presented correctlyYes, it is presented correctlyObjectiveWe aimed to (i) analyze the possible risk factors affecting the occurrence of in-hospital HF after emergency PCI in patients with AMI through clinical data and (ii) establish a personalized risk prediction model for the occurrence of HF after emergency PCI in patients with AMI.Please check if the author names and affiliations are captured correctlyYes, they are captured correctlyMethodsClinical data of 676 AMI patients who consecutively underwent emergency PCI between January 2020 and October 2023 at the First Affiliated Hospital of Guangxi University of Chinese Medicine were collected. Based on whether in-hospital HF occurred after PCI, the study subjects were divided into the HF group (91 cases) and the non-HF group (585 cases). Independent risk factors were screened using univariate and multivariate logistic regression. A nomogram model of the risk of HF was drawn using R, and the discriminative power was evaluated by calculating the area under the ROC curve and drawing the calibration curve and decision curve.ResultsIn this study, the incidence of in-hospital HF events in AMI patients after emergency PCI was 13.46%. The analysis showed that age, troponin levels, D-dimer levels, left ventricular ejection fraction (LVEF), and Gensini score were independent predictors of the occurrence of in-hospital HF in AMI patients after emergency PCI (P < 0.05). The AUC of the nomogram model were 0.87 (95% CI: 0. 82–0.91) and 0.85 (95% CI: 0. 76–0.93) in the training and validation sets, respectively. The Hosmer–Lemeshow goodness-of-fit test in the training set suggested that the difference between predicted and actual risks of the predictive model was not statistically significant (χ2 = 5.8185, P = 0.6676), and this was confirmed by the Hosmer–Lemeshow goodness-of-fit test in the validation set (χ2 = 9.4774, P = 0.3036).ConclusionsThe predictive model for the risk of in-hospital HF in AMI patients after emergency PCI includes age, troponin levels, D-dimer levels, LVEF, and Gensini score. It has a good differentiation ability and good accuracy, it can be used to intuitively and independently screen high-risk populations, and it has high predictive value for the occurrence of HF after PCI in AMI patients, so it can be used to assist clinicians in early screening, in identifying patients at high risk of postoperative HF, and in the implementation of targeted intervention therapy.Please check if "Strengths and limitations of this study" was captured and presented correctlyYes, it was captured and presented correctly

Effect of WW-domain transcription regulator 1 on aging regulation of human dental pulp stem cells

Objective: Investigating the changes of phenotype and moleculars associated with aging with the increase of passage times of human dental pulp stem cells (hDPSC), to explore the role of WW-containing transcriptional regulator 1 (WWTR1) in the aging mechanism. Methods: hDPSCs were cultured by tissue block method, and were divided into 4 groups according to the age, algebra, cell knockdown and overexpression of WWTR1 in hDPSCs. Group I: hDPSCs from human teeth were further divided into youth group (15-25 years old) and group middle-aged group (40-50 years old)] according to different ages. Group Ⅱ: according to different passage, hDPSCs were divided into young cells group (hDPSCs were transmitted to P3 generation), and old cells group (hDPSCs were transmitted to P10 generation). Group Ⅲ: hDPSCs were knocked down of WWTR1, which were further divided into knockdown group (sh-WWTR1) and knockdown carrier group (sh-NC). Group Ⅳ: hDPSCs were overexpressed of WWTR1, which were further divided into overexpression group (OE-WWTR1) and overexpression carrier group (OE-NC). Real-time quantitative fluorescent PCR (RT-qPCR) was used to detect the changes of WWTR1 expression in groups Ⅰ and Ⅱ, and cell counting kit-8 (CCK-8) was used for groups Ⅱ, Ⅲ, and Ⅳ. Cell proliferation capacity was detected by CCK-8 assay. The ability of osteogenic differentiation was detected by alizarin red staining. Cell senescence positive rate was detected by age-related β-galactosidase staining. The expression levels of age-related genes p53 and p21 were detected by RT-qPCR. Results: The proportion of senescent cells increased gradually with continuous culture. The proliferation and osteogenic differentiation of hDPSCs in the old group were significantly lower than those in the young group (P<0.001). The expression levels of senescence related genes p53 (2.09±0.24) and p21 (4.91±0.54) in old cell group were higher than those in young cell group respectively [p53: (1.08±0.09) and p21: (1.09±0.08)] (P<0.01, P<0.001). The WWTR1 expression levels of hDPSCs in middle-aged group and old cells group were both decreased compared with those in young group and young cells group (P<0.01). The proportion of senescent cells in knockdown group (44.50±2.42) was higher than that in knockdown carrier group (22.27±0.56) (P<0.001). After knocking down WWTR1 in hDPSCs, the expression levels of age-related genes p53 and p21 were up-regulated (P<0.001), and the abilities of proliferation and osteogenic differentiation in the knockdown group were lower than those in the knockdown carrier group (P<0.001). The proportion of senescent cells in overexpression empty carrier group (20.4±0.79) was higher than that in overexpression group (10.07±0.61) (P<0.001). After WWTR1 overexpression ins hDPSCs, the expression levels of age-related genes p53 and p21 were down-regulated, and the proliferation and osteogenic differentiation ability in overexpression group were higher than those in OE-NC group (P<0.001). Conclusions: WWTR1 can inhibit the expression levels of age-related genes p53 and p21, thus delaying the aging process as well as promoting the proliferation and osteogenic differentiation of hDPSCs.

A study on the plasma proteomics of different types of depressive disorders based on label-free data-independent acquisition proteomic technology

BackgroundDue to the high incidence and high misdiagnosis rate of depressive disorder, biomarkers for the accurate diagnosis of depressive disorder are urgently needed to reduce the misdiagnosis rate and improve the cure rate. MethodsTo obtain original data, plasma samples were collected from patients suffering from various depressive disorders, including bipolar depression (BP), major depressive disorder (MDD), and persistent depressive disorder (PDD) prior to medication treatment, as well as from participants without psychiatric diagnoses (NP). Then these samples were analyzed using nano-LC–MS/MS. According to the screening criteria, differentially expressed proteins(DEPs) corresponding to different types of depressive disorders were identified, and validation proteins were identified via bioinformatics analysis and verified. ResultsNinety-nine DEPs were identified between BP and NP, 29 DEPs were identified between MDD and NP, and 14 DEPs were identified between PDD and NP. The plasma levels of PRDX2 in patients with depressive disorder were significantly increased. The plasma CRP level in BP patients was specifically increased, and the plasma SNCA level in MDD patients was specifically increased. CRP showed the best differential diagnostic ability in differentiating BP from NP, MDD and PDD, and SNCA showed the best differential diagnostic ability in differentiating MDD from NP, BP and PDD. ConclusionsThe mechanism of depressive disorder mainly involves biological processes and signaling pathways related to inflammation and lipid metabolism. The key biomarkers identified by proteomics and the signaling pathways involved are highly important for revealing the biological basis of depressive disorder and guiding its clinical diagnosis and treatment.

Hypoxia-treated adipose mesenchymal stem cells derived exosomes enhance the therapeutic effects on unilateral ureteral obstruction mice.

The exosomes from adipose-derived mesenchymal stem cells (AMSCs) had therapeutic effects. However, whether the exosomes derived from hypoxia-treated AMSCs could improve renal functions in unilateral ureteral obstruction (UUO) mice remains unclear. The exosomes were characterized using a transmission electron microscope and Western blot. Its size distribution was determined using the Zetasizer Nano ZS analysis system. The differentiation ability was assessed by alkaline phosphatase and oil red staining. Consequently, the function of exosomes in inhibiting inflammatory factors was evaluated using an enzyme-linked immunosorbent assay, and apoptosis inhibition was evaluated by Western blot. Finally, the function of exosomes to ameliorate kidney fibrosis using qPCR, Western blot, hematoxylin-eosin staining and Masson staining. The cultured AMSCs could differentiate into osteoblast and adipocyte. Meanwhile, the cultured AMSCs could effectively secrete the exosomes, which were characterized with around 110 nm diameter and surface marker expression. Exosomes derived from hypoxia-treated AMSCs improved renal functions in UUO mice. The mechanism exploration revealed that exosomes could decrease the TNF-α and IL-6 and inhibit cell apoptosis. Finally, the fibrosis-associated protein was reversed, and the renal dysfunctions were ameliorated in UUO mice. The exosomes derived from the hypoxia-treated AMSCs have a better effect than those from normal AMSCs in ameliorating renal dysfunctions in UUO mice.

Exosomal miR-590-3p derived from bone marrow mesenchymal stem cells promotes osteoblast differentiation and osteogenesis by targeting TGFBR1.

Bone marrow mesenchymal stem cells (BMSCs) have been verified to be essential factors regulating osteogenic functions, which is mainly attributed to their secretion of extracellular vesicles. Exosomes derived from BMSCs (BMSCs-Exo) contribute to osteoblast functions that are critical for improving bone defect. Our current study aims to investigate the molecular mechanism dominated by BMSCs-Exo that affects osteoblast differentiation and osteogenesis. The first step this study validated that BMSCs co-culture enhanced the differentiation ability of osteoblast and promoted bone mineralization, while these tendencies were abolished after GW4869 treatment. Next, the BMSCs-Exo was isolated and identified by TEM observation, insight detection, and western blot analysis. Furthermore, BMSCs-Exo treatment could efficiently promote the differentiation ability and the bone mineralization of osteoblasts, decrease the mRNA levels of Collagen I and Collagen III, and increase the levels of osteogenic proteins, including alkaline phosphatase (ALP), Turning Bone Morphogenetic Protein 2 (BMP2), Bone sialoprofein (BSP), osteocalcin (OCN), and osterix (OSX). However, the abovementioned effects of BMSCs-Exo could be abolished by miR-590-3p silencing. Mechanistic analysis unmasked the negative regulation of miR-590-3p on its downstream target TGFBR1. Finally, the effects of miR-590-3p/TGFBR1 axis on the differentiation and osteogenesis of osteoblasts were validated by rescue assays. In conclusion, the present study demonstrates that exosomal miR-590-3p secreted by BMSCs can induce osteoblast differentiation and osteogenesis.

Mechanism of MiR-145a-3p/Runx2 pathway in dexamethasone impairment of MC3T3-E1 osteogenic capacity in mice.

In this experiment, we screened key miRNAs involved in the dexamethasone-induced decrease in osteogenic capacity of mouse precursor osteoblasts MC3T3-E1 over and investigated their specific regulatory mechanisms. In this experiment, cell counting kit assay was utilized to act on MC3T3-E1 cells at 0, 5μM, 10μM, 15μM concentrations of dexamethasone for 24h, 48h and 72h to observe the changes in cell viability in order to select the appropriate dexamethasone concentration. Apoptosis and reactive oxygen species were detected by flow cytometry. The transcription of osteogenesis-related genes (Runx2, ALP, OCN, OPN, OPG, COL1A1) and protein expression levels (Runx2, ALP, OCN, OPN) were detected by Western Blot and qRT-PCR to validate the changes in cellular osteogenesis. The differentially expressed miRNAs related to MC3T3-E1 osteogenic differentiation after dexamethasone action were screened out. The expression levels of selected target miRNAs were verified in the experimental group and the control group by qRT-PCR. The miRNA inhibitor was transfected to knock down miRNA in dexamethasone-induced MC3T3-E1 injury. Alkaline phosphatase staining and flow cytometry were performed to detect apoptosis and reactive oxygen species changes. transcript and protein expression levels of osteogenesis-related genes in mouse MC3T3-E1 were detected by qRT-PCR and Western blot experiments. By miRNA target gene prediction, luciferase reporter gene experiments, qRT-PCR and Western blot experiments were used to verify whether the selected target miRNAs targeted the target gene. First, it was determined that 10μM dexamethasone solution was effective in inducing a decrease in osteogenic function in mouse MC3T3-E1 by CCK8 experiments, which showed a significant decrease in alkaline phosphatase activity, a decrease in calcium nodules as shown by alizarin red staining, an increase in apoptosis and reactive oxygen species as detected by flow cytometry, as well as a decrease in the expression of osteogenesis-related genes and proteins. Five target miRNAs were identified: miR-706, miR-296-3p, miR-7011-5p, miR-145a-3p, and miR-149-3p. miR-145a-3p, which had the most pronounced and stable expression trend and was the most highly expressed miRNA, was chosen as the target of this experiment by qRT-PCR analysis. -145a-3p, as the subject of this experiment. Knockdown of miR-145a-3p in MC3T3-E1 cells after dexamethasone action significantly improved the expression of their impaired osteogenic indicators. It was shown that after knocking down the target miRNA, alkaline phosphatase staining was significantly increased compared with the dexamethasone-stimulated group and approached the level of the blank control group. Meanwhile, the expression of osteogenic function-related proteins and genes also increased in the dexamethasone-stimulated group after knocking down miR-145a-3p, and approached the level of the blank control group. A direct targeting relationship between miR-145a-3p and Runx2 was indeed confirmed by luciferase reporter gene assays, qRT-PCR and Western blot experiments. The results indicated that dexamethasone impaired the osteogenic differentiation ability of MC3T3-E1 cells by inducing the up-regulation of miR-145a-3p expression. MiR-145a-3p inhibited the osteogenic differentiation ability of MC3T3-E1 cells by targeting and suppressing the expression level of Runx2 protein. Inhibition of miR-145a-3p levels significantly improved the osteogenic differentiation ability of MC3T3-E1 cells.

Exploring mesenchymal stem cells homing mechanisms and improvement strategies.

Mesenchymal stem cells (MSCs) are multipotent cells with high self-renewal and multilineage differentiation abilities, playing an important role in tissue healing. Recent advancements in stem cell-based technologies have offered new and promising therapeutic options in regenerative medicine. Upon tissue damage, MSCs are immediately mobilized from the bone marrow and move to the injury site via blood circulation. Notably, allogenically transplanted MSCs can also home to the damaged tissue site. Therefore, MSCs hold great therapeutic potential for curing various diseases. However, one major obstacle to this approach is attracting MSCs specifically to the injury site following systemic administration. In this review, we describe the molecular pathways governing the homing mechanism of MSCs and various strategies for improving this process, including targeted stem cell administration, target tissue modification, in vitro priming, cell surface engineering, genetic modifications, and magnetic guidance. These strategies are crucial for directing MSCs precisely to the injury site and, consequently, enhancing their migration and local tissue repair properties. Specifically, our review provides a guide to improving the therapeutic efficacy of clinical applications of MSCs through optimized in vivo administration and homing capacities.

Cyclosporin A toxicity on endothelial cells differentiated from induced pluripotent stem cells: Assembling an adverse outcome pathway

Cyclosporin A (CSA) is a potent immunosuppressive agent in pharmacologic studies. However, there is evidence for side effects, specifically regarding vascular dysfunction. Its mode of action inducing endothelial cell toxicity is partially unclear, and a connection with an adverse outcome pathway (AOP) is not established yet. Therefore, we designed this study to get deeper insights into the mechanistic toxicology of CSA on angiogenesis. Stem cells, especially induced pluripotent stem cells (iPSCs) with the ability of differentiation to all organs of the body, are considered a promising in vitro model to reduce animal experimentation. In this study, we differentiated iPSCs to endothelial cells (ECs) as one cell type that in other studies would allow to generate multi-cell type organoids from single donors. Flow cytometry and immunostaining confirmed our scalable differentiation protocol. Then dose and time course experiments assessing CSA cytotoxicity on iPS derived endothelial cells were performed. Transcriptomic data suggested CSA dependent induction of reactive oxygen species (ROS), mitochondrial dysfunction, and impaired angiogenesis via ROS induction which was confirmed by in vitro experiments. In order to put these data into a potential adverse outcome pathway (AOP) context, we performed a literature review for CSA-mediated endothelial cell toxicity and combined our experimental data with the publicly available knowledge. Such an AOP will help to design in vitro test batteries and to model events observed in human toxicity studies, as well in predictive toxicology.

Role of R-spondin 2 on osteogenic differentiation of bone marrow mesenchymal stem cells and bone metabolism in ovariectomized mice

To investigate the effects of R-spondin 2 (Rspo2) on the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) and bone mineral content in ovariectomized mice. BMSCs were extracted from the bone marrow of the long bones of 7 4-week-old female C57BL/6 mice using whole bone marrow culture and passaged. After the cell phenotype was identified by flow cytometry, the 3rd generation cells were co-cultured with 10, 20, 40, 80, and 100 nmol/L Rspo2. Then, the cell activity and proliferative capacity were determined by cell counting kit 8 (CCK-8), and the intervention concentration of Rspo2 was screened for the subsequent experiments. The osteogenic differentiation ability of BMSCs was detected by alkaline phosphatase (ALP) staining, and the mRNA levels of osteogenesis-related genes [RUNX family transcription factor 2 (Runx2), collagen type Ⅰ alpha 1 (Col1), osteocalcin (OCN)] were detected by real-time fluorescence quantitative PCR (RT-qPCR). In addition, 18 10-week-old female C57BL/6 mice were randomly divided into sham operation group (sham group), ovariectomy group (OVX group), and OVX+Rspo2-intervention group (OVX+Rspo2 group), with 6 mice in each group. The sham group only underwent bilateral back incision and suturing, while the other two groups established osteoporosis mouse models by bilateral ovarian castration. Then, the mice were given a weekly intraperitoneal Rspo2 (1 mg/kg) treatment in OVX+Rspo2 group and saline at the same dosage in sham group and OVX group. After 12 weeks of treatment, the body mass and uterus mass of the mice were weighed in the 3 groups to assess whether the OVX model was successfully prepared; the tibia bones were stained with HE and immunohistochemistry staining to observe the changes in tibial bone mass and the expression level of Runx2 protein in the bone tissues. Blood was collected to detect the expressions of bone metabolism markers [ALP, OCN, type Ⅰ procollagen amino-terminal peptide (PINP)] and bone resorption marker [β-collagen degradation product (β-CTX)] by ELISA assay. Micro-CT was used to detect the bone microstructure changes in the tibia, and three-dimensional histomorphometric analyses were performed to analyze the trabeculae thickness (Tb.Th), trabeculae number (Tb.N), trabeculae separation (Tb.Sp), and bone volume fraction (BV/TV). CCK-8 assay showed that Rspo2 concentrations below 80 nmol/L were not cytotoxic ( P>0.05), and the cell viability of 20 nmol/L Rspo2 group was significantly higher than that of the control group ( P<0.05). Based on the above results, 10, 20, and 40 nmol/L Rspo2 were selected for subsequent experiments. ALP staining showed that the positive cell area of each concentration of Rspo2 group was significantly larger than that of the control group ( P<0.05), with the highest showed in the 20 nmol/L Rspo2 group. The expression levels of the osteogenesis-related genes (Runx2, Col1, OCN) significantly increased, and the differences were significant between Rspo2 groups and control group ( P<0.05) except for Runx2 in the 40 nmol/L Rspo2 group. In animal experiments, all groups of mice survived until the completion of the experiment, and the results of the body mass and uterus mass after 12 weeks of treatment showed that the OVX model was successfully prepared. Histological and immunohistochemical staining showed that the sparseness and connectivity of bone trabecula and the expression of Runx2 in the OVX group were lower than those in the sham group, whereas they were reversed in the OVX+Rspo2 group after treatment with Rspo2, and the differences were significant ( P<0.05). ELISA assay showed that compared with the sham group, the serum bone metabolism markers in OVX group had an increase in ALP and a decrease in PINP ( P<0.05). After Rspo2 intervention, PINP expression significantly reversed and increased, with significant differences compared to the sham group and OVX group ( P<0.05). The bone resorption marker (β-CTX) was significantly higher in the OVX group than in the sham group ( P<0.05), and it was significantly decreased in the OVX+Rspo2 group when compared with the OVX group ( P<0.05). Compared with the sham group, Tb.Th, Tb.N, and BV/TV significantly decreased in the OVX group, while Tb.Sp significantly increased ( P<0.05); after Rspo2 intervention, all of the above indexes significantly improved in the OVX+Rspo2 group ( P<0.05) except Tb.Th. Rspo2 promotes differentiation of BMSCs to osteoblasts, ameliorates osteoporosis due to estrogen deficiency, and promotes bone formation in mice.

BMSC Derived Exosomes Attenuate Apoptosis of Temporomandibular Joint Disc Chondrocytes in TMJOA via PI3K/AKT Pathway.

Bone marrow mesenchymal stem cell-derived exosomes (BMSC-Exos) are crucial means of intercellular communication and can regulate a range of biological processes by reducing inflammation, decreasing apoptosis and promoting tissue repair. We treated temporomandibular joint (TMJ) disc chondrocytes with TNF-α and performed local injection of sodium iodoacetate (MIA) in the TMJ of rats to establish in vitro and in vivo models of TMJ osteoarthritis (TMJOA). BMSC-Exos were isolated and extracted to evaluate their proliferation and trilineage differentiation abilities, and their antiapoptotic and chondroprotective effects were assessed. This study revealed that BMSC-Exos can be endocytosed by TMJ disc chondrocytes in vitro and that BMSC-Exos pretreatment strongly attenuated the inhibitory effect of TNF-α on the proliferative and chondrogenic potential of TMJ disc chondrocytes. The administration of BMSC-Exos significantly suppressed TNF-α-induced apoptosis in TMJ disc chondrocytes by increasing the phosphorylation level of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB/AKT) pathway-related proteins, whereas the PI3K inhibitor LY294002 neutralized this antiapoptotic effect. Intradiscal injection of BMSC-Exos alleviated the degeneration and inflammation of TMJ discs in a rat model of TMJOA. Our study revealed that BMSC-Exos can attenuate the apoptosis of TMJ disc chondrocytes and destruction of TMJ discs partially by inhibiting the apoptotic pathway and activating the PI3K/AKT pathway, thereby providing a promising treatment strategy for the regeneration of damaged TMJ discs.

Specific human gene expression in response to infection is an effective marker for diagnosis of latent and active tuberculosis

RNA sequencing and microarray analysis revealed transcriptional markers expressed in whole blood can differentiate active pulmonary TB (ATB) from other respiratory diseases (ORDs), and latent TB infection (LTBI) from healthy controls (HC). Here we describe a streamlined reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) assay that could be applied at near point-of-care for diagnosing and distinguishing ATB from ORDs and LTBI from HC. A literature review was undertaken to identify the most plausible host-gene markers (HGM) of TB infection. Primers, and dual labelled hydrolysis probes were designed and analytically evaluated for accuracy in an in-vitro model of infection using a lung fibroblast cell-line. Best performing genes were multiplexed into panels of 2–4 targets and taken forward for clinical evaluation. Mycobacteria Growth Indicator Tube and QuantiFERON-TB Gold Plus were used as reference tests for ATB and LTBI respectively. A total of 16 HGM were selected and incorporated into five multiplex RT-qPCR panels. PCR assay efficiency of all evaluated targets was ≥ 90% with a median analytical sensitivity of 292 copies/µl [IQR: 215.0-358.3 copies/µl], and a median limit of quantification of 61.7 copies/µl [IQR: 29.4-176.3 copies/µl]. Clinically, ATB was characterised by higher gene expression than ORDs, while LTBI was associated with lower gene expression than HC, Kruskal-Wallis p < 0.0001. Crucially, BATF2, CD64, GBP5, C1QB, GBP6, DUSP3, and GAS6 exhibited high differentiative ability for ATB from ORDs, LTBI or HC while KLF2, PTPRC, NEMF, ASUN, and ZNF296 independently discriminated LTBI from HC. Our results show that different HGM maybe required for ATB and LTBI differentiation from ORDs or HC respectively and demonstrate the feasibility of host gene-based RT-qPCR to diagnose ATB and LTBI at near point-of-care.

Metformin improves HPRT1-targeted purine metabolism and repairs NR4A1-mediated autophagic flux by modulating FoxO1 nucleocytoplasmic shuttling to treat postmenopausal osteoporosis.

Osteoporosis is a major degenerative metabolic bone disease that threatens the life and health of postmenopausal women. Owing to limitations in detection methods and prevention strategy awareness, the purpose of osteoporosis treatment is more to delay further deterioration rather than to fundamentally correct bone mass. We aimed to clarify the pathogenesis of postmenopausal osteoporosis and optimize treatment plans. Our experiments were based on previous findings that oxidative stress mediates bone metabolism imbalance after oestrogen deficiency. Through energy metabolism-targeted metabolomics, we revealed that purine metabolism disorder is the main mechanism involved in inducing oxidative damage in bone tissue, which was verified via the use of machine-learning data from human databases. Xanthine and xanthine oxidase were used to treat osteoblasts to construct a purine metabolism disorder model. The activity and differentiation ability of osteoblasts decreased after X/XO treatment. Transcriptomic sequencing indicated that autophagic flux damage was involved in purine metabolism-induced oxidative stress in osteoblasts. Additionally, we performed serum metabolomics combined with network pharmacology to determine the pharmacological mechanism of metformin in the treatment of postmenopausal osteoporosis. HPRT1 was the potential target filtered from the hub genes, and FoxO1 signalling was the key pathway mediating the effect of metformin in osteoblasts. We also revealed that SIRT3-mediated deacetylation promoted the nuclear localization of FoxO1 to increase the expression of HPRT1. HPRT1 upregulation promoted purine anabolism and prevented the accumulation of ROS caused by purine catabolism to reverse oxidative damage in osteoblasts. We propose that purine metabolism disorder-induced oxidative stress is important for the pathogenesis of postmenopausal osteoporosis. The therapeutic mechanism of metformin should be confirmed through subsequent drug optimization and development studies to improve bone health in postmenopausal women.