Ultrasound-generated bubbles enhance osteogenic differentiation of mesenchymal stromal cells in composite collagen hydrogels.

Hydrogels can improve the delivery of mesenchymal stromal cells (MSCs) by providing crucial biophysical cues that mimic the extracellular matrix. The differentiation of MSCs is dependent on biophysical cues like stiffness and viscoelasticity, yet conventional hydrogels cannot be dynamically altered after fabrication and implantation to actively direct differentiation. We developed a composite hydrogel, consisting of type I collagen and phase-shift emulsion, where osteogenic differentiation of MSCs can be non-invasively modulated using ultrasound. When exposed to ultrasound, the emulsion within the hydrogel was non-thermally vaporized into bubbles, which locally compacted and stiffened the collagen matrix surrounding each bubble. Bubble growth and matrix compaction were correlated, with collagen regions proximal (i.e., ≤ ∼60 µm) to the bubble displaying a 2.5-fold increase in Young's modulus compared to distal regions (i.e., > ∼60 µm). The viability and proliferation of MSCs, which were encapsulated within the composite hydrogel, were not impacted by bubble formation. In vitro and in vivo studies revealed encapsulated MSCs exhibited significantly elevated levels of RUNX2 and osteocalcin, markers of osteogenic differentiation, in collagen regions proximal to the bubble compared to distal regions. Additionally, alkaline phosphatase activity and calcium deposition were enhanced adjacent to the bubble. An opposite trend was observed for CD90, a marker of MSC stemness. Following subcutaneous implantation, bubbles persisted in the hydrogels for two weeks, which led to localized collagen alignment and increases in nuclear asymmetry. These results are a significant step toward controlling the 3D differentiation of MSCs in a non-invasive and on-demand manner.

Wenshen Xiaozheng Tang alleviates fibrosis in endometriosis by regulating differentiation and paracrine signaling of endometrium-derived mesenchymal stem cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Wenshen Xiaozheng Tang (WXT), a traditional Chinese medicine (TCM) decoction, is effective for treating endometriosis. However, the effect of WXT on endometrium-derived mesenchymal stem cells (eMSCs) which play a key role in the fibrogenesis of endometriosis requires further elucidation. AIMS OF THE STUDY: The aim of this study was to clarify the potential mechanism of WXT in improving fibrosis in endometriosis by investigating the regulation of WXT on differentiation and paracrine of eMSCs. MATERIALS AND METHODS: The nude mice with endometriosis were randomly divided into model group, WXT group and mifepristone group. After 21 days of treatment, the lesion volume was calculated. Fibrosis in the lesions was evaluated by Masson staining and expression of fibrotic proteins. The differentiation of eMSCs in vivo was explored using a fate-tracking experiment. To further clarify the regulation of WXT on eMSCs, primary eMSCs from the ectopic lesions of endometriosis patients were isolated and characterized. The effect of WXT on the proliferation and differentiation of ectopic eMSCs was examined. To evaluate the role of WXT on the paracrine activity of ectopic eMSCs, the conditioned medium (CM) from ectopic eMSCs pretreated with WXT was collected and applied to treat ectopic endometrial stromal cells (ESCs), after which the expression of fibrotic proteins in ectopic ESCs was assessed. In addition, transcriptome sequencing was used to investigate the regulatory mechanism of WXT on ectopic eMSCs, and western blot and ELISA were employed to determine the key mediator. RESULTS: WXT impeded the growth of ectopic lesions in nude mice with endometriosis and reduced collagen deposition and the expression of fibrotic proteins fibronectin, collagen I, α-SMA and CTGF in the endometriotic lesions. The fate-tracking experiment showed that WXT prevented human eMSCs from differentiating into myofibroblasts in the nude mice. We successfully isolated eMSCs from the lesions of patients with endometriosis and demonstrated that WXT suppressed proliferation and myofibroblast differentiation of ectopic eMSCs. Moreover, the expression of α-SMA, collagen I, fibronectin and CTGF in ectopic ESCs was significantly down-regulated by the CM of ectopic MSCs pretreated with WXT. Combining the results of RNA sequencing, western blot and ELISA, we found that WXT not only reduced thrombospondin 4 expression in ectopic eMSCs, but also decreased thrombospondin 4 secretion from ectopic eMSCs. Thrombospondin 4 concentration-dependently upregulated the expression of collagen I, fibronectin, α-SMA and CTGF in ectopic ESCs, indicating that thrombospondin 4 was a key mediator of WXT in inhibiting the fibrotic process in endometriosis. CONCLUSION: WXT improved fibrosis in endometriosis by regulating differentiation and paracrine signaling of eMSCs. Thrombospondin 4, whose release from ectopic eMSCs is inhibited by WXT, may be a potential target for the treatment of endometriosis.

Generating ESC-Derived RGCs for Cell Replacement Therapy.

In several ocular diseases, degeneration of retinal neurons can lead to permanent blindness. Transplantation of stem cell (SC)-derived RGCs has been proposed as a potential therapy for RGC loss. Although there are reports of successful cases of SC-derived RGC transplantation, achieving long-distance regeneration and functional connectivity remains a challenge. To address these hurdles, retinal organoids are being used to study the regulatory mechanism of stem cell transplantation. Here we present a modified protocol for differentiating human embryonic stem cells (ESCs) into retinal organoids and transplanting organoid-derived RGCs into the murine eyes.

Ecological linkages between top-down designed benzothiazole-degrading consortia and selection strength: From performance to community structure and functional genes.

The inefficient biodegradation and incomplete mineralization of nitrogenous heterocyclic compounds (NHCs) have emerged as a pressing environmental concern. The top-down design offers potential solutions to this issue by targeting improvements in community function, but the ecological linkages between selection strength and the structure and function of desired microbiomes remain elusive. Herein, the integration of metagenomics, culture-based approach, non-targeted metabolite screening and enzymatic verification experiments revealed the effect of enrichment concentration on the top-down designed benzothiazole (BTH, a typical NHC)-degrading consortia. Significant differences were observed for the degradation efficiency and community structure under varying BTH selections. Notably, the enriched consortia at high concentrations of BTH were dominated by genus Rhodococcus, possessing higher degradation rates. Moreover, the isolate Rhodococcus pyridinivorans Rho48 displayed excellent efficiencies in BTH removal (98 %) and mineralization (∼ 60 %) through the hydroxylation and cleavage of thiazole and benzene rings, where cytochrome P450 enzyme was firstly reported to participate in BTH conversion. The functional annotation of 460 recovered genomes from the enriched consortia revealed diverse interspecific cooperation patterns that accounted for the BTH mineralization, particularly Nakamurella and Micropruina under low selection strength, and Rhodococcus and Marmoricola under high selection strength. This study highlights the significance of selection strength in top-down design of synthetic microbiomes for degrading refractory organic pollutants, providing valuable guidance for designing functionally optimized microbiomes used in environmental engineering.

Effect of egg incubator temperature on sex differentiation in Korat chickens.

The effect of incubator temperature on sex differentiation in Korat chickens was investigated. The experiments were divided into two sets: temperature applied throughout the entire incubation period and temperature applied during certain periods (days 3-6 of incubation) by either increasing above the standard or decreasing below the standard temperature. In each experiment, 300 Korat chicken eggs were separated into three groups of 5 repetitions, with 20 eggs in each group. This was done using a completely randomized design for each experiment: a group using a temperature below the standard for incubation (36.0 °C), a group using the standard incubation temperature (37.7 °C), and a group using a temperature above the standard for incubation (38.0 °C). W chromosomes were detected at hatch; histology examined reproductive structures after 35 days. Increasing the temperature to 38.0 °C throughout the entire incubation period resulted in no significant difference in hatching rates compared to the standard temperature (P > 0.05). Raising the temperature to 38.0 °C throughout the entire incubation and during certain periods resulted in changes in the reproductive structure of chickens, leading to a mismatch between chromosomal and gonadal sex, observed at 9.7% and 5.9% of individuals with W chromosomes possessed testes, indicating a mismatch between chromosomal and gonadal sex. However, decreasing the temperature to 36.0 °C throughout the incubation period resulted in lower hatching rates compared to the standard temperature (P < 0.05). Incubating eggs at 36.0 °C for specific periods resulted in 19.4% of genetic males developing ovaries instead of testes. The presence of ovaries in individuals without W chromosomes indicated this mismatch. The results of this study provide evidence that temperature plays a role in sex differentiation in Korat chickens, as demonstrated by the detection of W chromosomes and histological studies of testes and ovaries. Moreover, this study presents the first evidence in broilers that temperature can affect sex differentiation.

MRI-based deep learning for differentiating between bipolar and major depressive disorders.

Mood disorders, particularly bipolar disorder (BD) and major depressive disorder (MDD), manifest changes in brain structure that can be detected using structural magnetic resonance imaging (MRI). Although structural MRI is a promising diagnostic tool, prevailing diagnostic criteria for BD and MDD are predominantly subjective, sometimes leading to misdiagnosis. This challenge is compounded by a limited understanding of the underlying causes of these disorders. In response, we present SE-ResNet, a Residual Network (ResNet)-based framework designed to discriminate between BD, MDD, and healthy controls (HC) using structural MRI data. Our approach extends the traditional Squeeze-and-Excitation (SE) layer by incorporating a dedicated branch for spatial attention map generation, equipped with soft-pooling, a 7 × 7 convolution, and a sigmoid function, intended to detect complex spatial patterns. The fusion of channel and spatial attention maps through element-wise addition aims to enhance the model's ability to discriminate features. Unlike conventional methods that use max-pooling for downsampling, our methodology employs soft-pooling, which aims to preserve a richer representation of input features and reduce data loss. When evaluated on a proprietary dataset comprising 303 subjects, the SE-ResNet achieved an accuracy of 85.8 %, a recall of 85.7 %, a precision of 85.9 %, and an F1 score of 85.8 %. These performance metrics suggest that the SE-ResNet framework has potential as a tool for detecting psychiatric disorders using structural MRI data.

Bone marrow mesenchymal stem cells-derived exosomal miR-381 alleviates lung ischemia-reperfusion injury by activating Treg differentiation through inhibiting YTHDF1 expression.

AIM: Our study aimed to investigate whether BMSCs-derived exosomal miR-381 promotes Treg cell differentiation in lung ischemia-reperfusion injury (LIRI), and the underlying mechanism. METHODS: The in vitro and in vivo models of LIRI were established by hypoxia/reoxygenation (H/R) treatment and lung ischemia/reperfusion (I/R) surgery, respectively. BMSCs-derived exosomes were isolated and identified by western blot, nanoparticle tracking analysis, and transmission electron microscopy. Cell viability, proliferation, and apoptosis were assessed by CCK-8, EdU, and flow cytometry assay, respectively. IL-18 secretion level in lung microvascular endothelial cells (LMECs) and lung tissue homogenate was examined by ELISA. Treg cell differentiation was determined using flow cytometry. The relationships between miR-381, YTHDF1, and IL-18 were investigated using dual-luciferase reporter gene, RIP, and/or RNA pull-down assays. MeRIP assay was employed to determine m6A modification of IL-18 mRNA in LMECs. The ubiquitination level of Foxp3 protein in CD4+ T cells was analyzed by Co-IP assay. RESULTS: BMSCs-derived exosomes reduced LMECs injury and increased Treg cell differentiation in LIRI, whereas miR-381 inhibition in BMSCs weakened these impacts. Mechanistically, miR-381 inhibited IL-18 translation in LMECs by inhibiting YTHDF1 expression via binding to its 3'-UTR. As expected, YTHDF1 overexpression in LMECs abolished the effects of miR-381-overexpressed exosomes on LMECs injury and Treg cell differentiation. Moreover, LMECs-secreted IL-18 inhibited Treg cell differentiation by promoting the ubiquitination degradation of Foxp3 protein. CONCLUSION: BMSCs-derived exosomal miR-381 suppressed IL-18 translation in LMECs through binding to YTHDF1 3'-UTR, thus suppressing the ubiquitination degradation of Foxp3 in CD4+ T cells, which promoted Treg cell differentiation and mitigated LIRI development.

Titanium surfaces loaded with puerarin and exosomes derived from adipose stem cells promote the proliferation and differentiation of pre-osteoblasts.

The study is to evaluate the effects of collagen/hyaluronic acid coating with or without puerarin and exosomes (Exos) derived from adipose stem cells (ADSCs-Exos) on pre-osteoblast proliferation and differentiation on the surface of titanium materials. Titanium materials with different coatings were prepared by layer-by-layer technique, evaluating the surface characterization. Cell functions were assessed by cell biology experiments. Related genes and proteins were assessed by RT-qPCR and Western blot. Puerarin or ADSCs-Exos coating had better effects on promoting the adhesion, proliferation and differentiation of pre-osteoblasts, and the strongest effect was found after their co-coatings, manifesting as the up-regulations of alkaline phosphatase (ALP) activity, collagen type I alpha 1 (Col1a1), runt-related transcription factor 2 (Runx2), osterix and activating transcription factor-2 (ATF-2). Levels of phosphorylated-P38 (p-P38) and p-ATF-2 were up-regulated in pre-osteoblasts grown on puerarin and ADSCs-Exos-loaded titanium surfaces. Titanium surfaces loaded with puerarin and ADSCs-Exos promotes the proliferation and differentiation of pre-osteoblasts.

Favorable Response to Adjuvant Tofacitinib in a Case of Anti-Melanoma Differentiation-Associated Gene-5 Antibody Positive Clinically Amyopathic Dermatomyositis.

Anti-melanoma differentiation-associated gene-5 antibody (anti-MDA-5 Ab) associated with clinically amyopathic dermatomyositis (CADM) is characterized by vasculopathic ulcers, mechanic's hands, and progressive interstitial lung disease (ILD). We present a case of 38-year-old female who presented with all these classical clinical features. Her investigations revealed normal serum muscle enzyme levels and the presence of anti-Mi2 and anti-MDA-5 antibodies by immunoblot. Imaging study revealed changes suggestive of ILD. She was treated with rituximab along with oral glucocorticoid and other supportive treatment to which she didn't respond adequately. Recently, it has been postulated that plasmacytoid dendritic cells produce interferon which is responsible for tissue injury in dermatomyositis (DM). Tofacitinib, by inhibiting JAK-STAT pathway, inhibits downstream cytokines, mainly type 1 interferon. So, we added tofacitinib as adjuvant therapy in our patient. Post-six months of commencement of adjuvant tofacitinib, patient experienced remarkable improvement in cutaneous features as well as in pulmonary fibrosis.

Steroidogenic differentiation of human amniotic membrane-derived mesenchymal stem cells into a progesterone-/androgen-producing cell lineage by SF-1 and an estrogen-producing cell lineage by WT1-KTS.

Background: Sex steroid hormones, primarily synthesized by gonadal somatic cells, are pivotal for sexual development and reproduction. Mice studies have shown that two transcription factors, steroidogenic factor 1 (SF-1) and Wilms' tumor 1 (WT1), are involved in gonadal development. However, their role in human gonadal somatic differentiation remains unclear. We therefore aimed to investigate the roles of SF-1 and WT1 in human gonadal steroidogenic cell differentiation. Methods: Using a transient lentivirus-mediated gene expression system, we assessed the effects of SF-1 and WT1 expression on the steroidogenic potential of human amniotic membrane-derived mesenchymal stem cells (hAmMSCs). Results: SF-1 and WT1-KTS, a splice variant of WT1, played distinct roles in human steroidogenic differentiation of hAmMSCs. SF-1 induced hAmMSC differentiation into progesterone- and androgen-producing cell lineages, whereas WT1-KTS promoted hAmMSC differentiation into estrogen-producing cell lineages. Conclusion: Our findings revealed that SF-1 and WT1-KTS play important roles in human gonadal steroidogenic cell differentiation, especially during ovarian development. These findings may pave the way for future studies on human ovarian differentiation and development.

Clinical and genetic insights into ABCA12 variants in three Chinese families with ichthyosis: Genotype-phenotype correlation.

Autosomal recessive congenital ichthyosis (ARCI) comprises a series of non-syndromic ichthyoses. Pathogenic variants in several genes associated with ARCI have so far been identified. Notably, the variants in ABCA12 play a pivotal role in the pathology of ARCI. In this study, we report three Chinese families with compound heterozygous variants in the ABCA12 gene, including two novel variants and four reported variants. Clinical and genetic analyses were conducted to explore the genotype-phenotype correlation among the patients. Immunohistochemistry and transcriptome sequencing were utilized to assess the impact of pathogenic ABCA12 variants on skin homeostasis, revealing decreased levels of ABCA12 and claudin-1, alongside increased levels of involucrin and S100A8. In conclusion, our findings contribute to updating the genotype-phenotypic correlation and provide additional evidence for the long-term use of retinoic acid drugs in patients with causative ABCA12 variants.

Wnt3a-induced LRP6 phosphorylation enhances osteoblast differentiation to alleviate osteoporosis through activation of mTORC1/ß-catenin signaling.

OBJECTIVE: Osteoporosis (OP) is a common cause of morbidity and mortality in older individuals. The importance of Wnt3a in osteogenic activity and bone tissue homeostasis is well known. Here, we explored the possible molecular mechanism by which Wnt3a mediates the LRP6/mTORC1/ß-catenin axis to regulate osteoblast differentiation in OP. METHODS: OP-related key genes were identified through a bioinformatics analysis. A ROS17/2.8 cell differentiation system for rat osteogenic progenitors and a rat model of senile OP were constructed for in vitro and in vivo mechanism verification. RESULTS: Bioinformatics analysis revealed that LRP6 was poorly expressed in OP and may play a key role in the occurrence of OP by affecting osteoblast differentiation. LRP6 knockdown inhibited osteoblast differentiation in an in vitro model. In addition, Wnt3a promoted osteoblast differentiation by inducing LRP6 phosphorylation. Moreover, LRP6 promoted mTORC1 expression, which indirectly promoted ß-catenin expression, thus promoting osteoblast differentiation. Finally, an in vivo assay revealed that LRP6 inhibition improved OP. CONCLUSION: Our study provides evidence that Wnt3a induces phosphorylation of LRP6 to activate the mTORC1/ß-catenin axis, thus promoting osteoblast differentiation and ultimately improving OP in aged rats.

Standardization and optimization of the hiPSC-based PluriLum assay for detection of embryonic and developmental toxicants.

New approach methodologies (NAMs) for predicting embryotoxicity and developmental toxicity are urgently needed for generating human relevant data, while reducing turnover time and costs, and alleviating ethical concerns related to the use of animal models. We have previously developed the PluriLum assay, a NKX2.5-reporter gene 3D model using human-induced pluripotent stem cells (hiPSCs) that are genetically modified to enable the assessment of adverse effects of chemicals on the early-stage embryo. Aiming at improving the predictive value of the PluriLum assay for future screening purposes, we sought to introduce standardization steps to the protocol, improving the overall robustness of the PluriLum assay, as well as a shortening of the assay protocol. First, we showed that the initial size of embryoid bodies (EBs) is crucial for a proper differentiation into cardiomyocytes and overall reproducibility of the assay. When the starting diameter of the EBs exceeds 500 µm, robust differentiation can be anticipated. In terms of reproducibility, exposure to the fungicide epoxiconazole at smaller initial diameters resulted in a larger variation of the derived data, compared to more reliable concentration-response curves obtained using spheroids with larger initial diameters. We further investigated the ideal length of the differentiation protocol, resulting in a shortening of the PluriLum assay by 24 h to 7 days. Following exposure to the teratogens all-trans and 13-cis retinoic acid, both cardiomyocyte contraction and measurement of NKX2.5-derived luminescence were recorded with a similar or increased sensitivity after 6 days of differentiation when compared to the original 7 days. Finally, we have introduced an efficient step for enzymatic dissociation of the EBs at assay termination. This allows for an even splitting of the individual EBs and testing of additional endpoints other than the NKX2.5-luciferase reporter, which was demonstrated in this work by the simultaneous assessment of ATP levels. In conclusion, we have introduced standardizations and streamlined the PluriLum assay protocol to improve its suitability as a NAM for screening of a large number of chemicals for developmental toxicity testing.

Geometric constraint of mechanosensing by modification of hydrogel thickness prevents stiffness-induced differentiation in bone marrow stromal cells.

Extracellular matrix (ECM) stiffness is fundamental in cell division, movement and differentiation. The stiffness that cells sense is determined not only by the elastic modulus of the ECM material but also by ECM geometry and cell density. We hypothesized that these factors would influence cell traction-induced matrix deformations and cellular differentiation in bone marrow stromal cells (BMSCs). To achieve this, we cultivated BMSCs on polyacrylamide hydrogels that varied in elastic modulus and geometry and measured cell spreading, cell-imparted matrix deformations and differentiation. At low cell density BMSCs spread to a greater extent on stiff compared with soft hydrogels, or on thin compared with thick hydrogels. Cell-imparted matrix deformations were greater on soft compared with stiff hydrogels or thick compared with thin hydrogels. There were no significant differences in osteogenic differentiation relative to hydrogel elastic modulus and thickness. However, increased cell density and/or prolonged culture significantly reduced matrix deformations on soft hydrogels to levels similar to those on stiff substrates. This suggests that at high cell densities cell traction-induced matrix displacements are reduced by both neighbouring cells and the constraint imposed by an underlying stiff support. This may explain observations of the lack of difference in osteogenic differentiation as a function of stiffness.

Altered Methylation Levels in LINE-1 in Dental Pulp Stem Cell-Derived Osteoblasts.

OBJECTIVES: Long interspersed nuclear element-1 (LINE-1) and Alu elements are major targets of methylation, an epigenetic mechanism that is associated with several biological processes. Alterations of methylation of LINE-1 and Alu have been reported in cancers, diseases, and ageing. However, these alterations have not been studied in osteogenic differentiation of dental pulp stem cells (DPSCs), which are a promising source of tissue regeneration. METHOD: This study was performed to investigate the methylation level of LINE-1 and Alu in dental pulp stem cell-derived osteoblasts (DPSC-DOs). By using the combined bisulfite restriction analysis, the levels of total methylation and 4 patterns of methylated cytosine-phosphate-guanine (CpG) dinucleotides of LINE-1 and Alu were compared between DPSC-DOs and DPSCs. RESULT: The levels of total methylation and hypermethylated CpG dinucleotides of LINE-1 were significantly lower (P = .015 and .021, respectively), whilst levels of one pattern of partial methylated CpG dinucleotides were significantly higher in DPSC-DOs than DPSCs (P = .021). The methylation of Alu was not significantly different between DPSCs and DPSC-DOs. CONCLUSIONS: Methylation alterations of LINE-1 but not Alu were found in osteogenic differentiation of DPSCs. The results of this study offer foundational insights into osteoblast differentiation from an epigenetic perspective and may contribute to advancements in bone regeneration therapy in the future.

Potential of Dental Pulp Stem Cell Exosomes: Unveiling miRNA-Driven Regenerative Mechanisms.

Human dental pulp stem cells (hDPSCs) have emerged as a promising resource in regenerative medicine due to their unique ability to secrete exosomes containing a diverse array of bioactive molecules, particularly microRNAs (miRNAs). These exosomes appear to be essential for stimulating regenerative mechanisms, especially those associated with stem cell pluripotency and tissue repair. However, several challenges such as cargo specificity and delivery efficiency need to be addressed to maximise the therapeutic potential of hDPSC-derived exosomes and miRNA-based therapies. This narrative review explores hDPSCs' potential in regenerative medicine by examining their role in tissue engineering, secretome composition, exosome function, exosomal miRNA in diverse models, and miRNA profiling. Therefore, it is imperative to sustain ongoing research on miRNA to advance clinical applications in the field of regenerative medicine and dentistry. A comprehensive understanding of the specific miRNA composition within hDPSC-derived exosomes is essential to elucidate their mechanistic roles in diverse disease states and to inform the development of innovative therapeutic strategies. These findings hold significant potential for the development of innovative regenerative therapies and emphasises the importance of establishing a strong connection between translational research discoveries and clinical applications. hDPSC-derived exosomes and miRNA-based therapies play a crucial role in immune modulation, regenerative dentistry, and tissue repair.

The transcription regulators ZNF750 and LSD1/KDM1A dampen inflammation on the skin's surface by silencing pattern recognition receptors.

The surface of the skin is continually exposed to pro-inflammatory stimuli; however, it is unclear why it is not constantly inflamed due to this exposure. Here, we showed undifferentiated keratinocytes residing in the deep epidermis could trigger a strong inflammatory response due to their high expression of pattern recognition receptors (PRRs) that detect damage or pathogens. As keratinocytes differentiated, they migrated outward toward the surface of the skin and decreased their PRR expression, which led to dampened immune responses. ZNF750, a transcription factor expressed only in differentiated keratinocytes, recruited the histone demethylase KDM1A/LSD1 to silence genes coding for PRRs (TLR3, IFIH1/MDA5, and DDX58/RIG1). Loss of ZNF750 or KDM1A in human keratinocytes or mice resulted in sustained and excessive inflammation resembling psoriatic skin, which could be restored to homeostatic conditions upon silencing of TLR3. Our findings explain how the skin's surface prevents excessive inflammation through ZNF750- and KDM1A-mediated suppression of PRRs.

AEBP1 restores osteoblastic differentiation under dexamethasone treatment by activating PI3K/AKT signalling.

Adipocyte enhancer-binding protein 1 (AEBP1) is closely implicated in osteoblastic differentiation and bone fracture; this research aimed to investigate the effect of AEBP1 on restoring osteoblastic differentiation under dexamethasone (Dex) treatment, and its interaction with the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway. Pre-osteoblastic MC3T3-E1 cells were cultured in osteogenic medium and treated by Dex to mimic steroid-induced osteonecrosis cellular model. They were then further transfected with control or AEBP1-overexpressed lentiviral vectors. Finally, cells were treated with the PI3K inhibitor LY294002, with or without AEBP1-overexpressed lentiviral vectors. AEBP1 expression showed a downward trend in MC3T3-E1 cells under Dex treatment in a dose-dependent manner. AEBP1-overexpressed lentiviral vectors increased relative cell viability, alkaline phosphatase (ALP) staining, Alizarin red staining and osteoblastic differentiation markers including osteocalcin (OCN), osteopontin (OPN), collagen type I alpha 1 (COL1A1), runt-related transcription factor 2 (RUNX2) and bone morphogenetic protein 2 (BMP2), but decreased cell apoptosis rate in MC3T3-E1 cells under Dex treatment; besides, AEBP1-overexpressed lentiviral vectors positively regulated p-PI3K and p-AKT expressions. Furthermore, LY294002 treatment decreased relative cell viability, Alizarin red staining, osteoblastic differentiation markers including OCN, OPN, RUNX2 and BMP, increased cell apoptosis rate and did not affect ALP staining in MC3T3-E1 cells under Dex treatment; meanwhile, LY294002 treatment weakened the effect of AEBP1 overexpression vectors on the above cell functions. AEBP1 restores osteoblastic differentiation under Dex treatment by activating the PI3K/AKT pathway.

Genetic Structure of Monochamus alternatus (Hope) in Qinling-Daba Mountains and Expansion Trend: Implications for Pest Prevention and Management.

Pine wilt disease (PWD), caused by Bursaphelenchus xylophilus, severely threatens global pine forests. Monochamus alternatus is the primary vector of B. xylophilus in East Asia. Understanding the population structure and evolutionary forces of vector insects is critical for establishing effective PWD management strategies. The present work explores the genetic structure and phylogenetic relationships of 20 populations of M. alternatus from the Qinling-Daba Mountains (QDM) in China using the mitochondria DNA dataset, supplemented by ecological niche modeling (ENM). All M. alternatus populations were categorized into three phylogeographic clusters (Clade A, Clade B, and Clade C), with Clade A and Clade B corresponding to the western and eastern QDM, respectively. The results of divergence time estimation concur with environmental changes induced by Quaternary glacial climate oscillations in QDM of China. M. alternatus populations exhibited significant genetic differentiation, with expansion in their population size. Ecological niche modeling (ENM) demonstrated that precipitation and temperature significantly influence the distribution of M. alternatus and the species is anticipated to grow into higher latitude and higher altitude regions in the future. In a nutshell, exploring the genetic structure and evolutionary dynamics of M. alternatus can provide valuable insights into the prevention and occurrence of B. xylophilus. These findings also serve as a reference for research on population differentiation and phylogeography of other species in QDM and adjacent areas.

13-cis Retinoic Acid-Mediated Modulation of Human Meibomian Gland Epithelial Cells Development: Implications for In Vitro Modeling of Meibomian Gland Dysfunction.

Purpose: This study aimed to investigate the effect of 13-cis retinoic acid (13-cis RA) on human meibomian gland epithelial cells (HMGECs) and explore the potential of using this experimental model as an in vitro approach for studying meibomian gland dysfunction (MGD). Methods: First, HMGECs were cultured with 13-cis RA at different doses and times, and cell viability and proliferation rates were assessed to determine the appropriate stimulation concentration and time. Subsequently, during the proliferation stage, the expression of proliferation, inflammation, and oxidative stress genes and their products were evaluated. The meibum synthesis capacity was determined during the differentiation stage. Additionally, the peroxisome proliferator-activated receptor gamma (PPARγ) antagonist GW9662 was used as a control to assess the impact of 13-cis RA on PPARγ. Results: 13-cis RA significantly inhibited cell viability and proliferation in a time-dose response manner. Under the stimulation of 2 and 5 µM for 48 h during the proliferation stage, a significant decrease was observed in the expression of cell proliferation markers Ki67, antioxidant SOD-2, and Nrf-2. However, the expression of the pro-inflammatory factors IL-1ß, IL-8, MMP9, and oxidative stress markers NOX-4 and reactive oxygen species increased. During the differentiation stage, it suppressed meibum synthesis and the expression of meibocyte differentiation-related proteins adipose differentiation-associated protein 4 (ADFP4), elongation of very long chain fatty acid protein 4 (ELOVL4), sterol regulatory element-binding protein 2 (SREBP-2), and PPARγ. Conclusion: 13-cis RA inhibited cell viability, promoted inflammation and oxidative stress, and suppressed meibum synthesis through the PPARγ pathway. Our study shed light on the effect of 13-cis RA on HMGECs and provided a promising direction for studying MGD in vitro.