Specific Role Research Articles

Essential Role of Cortactin in Myogenic Differentiation: Regulating Actin Dynamics and Myocardin-Related Transcription Factor A-Serum Response Factor (MRTFA-SRF) Signaling

Cortactin (CTTN) is an actin-binding protein regulating actin polymerization and stabilization, which are vital processes for maintaining skeletal muscle homeostasis. Despite the established function of CTTN in actin cytoskeletal dynamics, its role in the myogenic differentiation of progenitor cells remains largely unexplored. In this study, we investigated the role of CTTN in the myogenic differentiation of C2C12 myoblasts by analyzing its effects on actin cytoskeletal remodeling, myocardin-related transcription factor A (MRTFA) nuclear translocation, serum response factor (SRF) activation, expression of myogenic transcription factors, and myotube formation. CTTN expression declined during myogenic differentiation, paralleling the reduction in MyoD, suggesting a potential role in the early stages of myogenesis. We also found that CTTN knockdown in C2C12 myoblasts reduced filamentous actin, enhanced globular actin levels, and inhibited the nuclear translocation of MRTFA, resulting in suppressed SRF activity. This led to the subsequent downregulation of myogenic regulatory factors, such as MyoD and MyoG. Furthermore, CTTN knockdown reduced the nuclear localization of YAP1, a mechanosensitive transcription factor, further supporting its regulatory roles in cell cycle and proliferation. Consequently, CTTN depletion impeded proliferation, differentiation, and myotube formation in C2C12 myoblasts, highlighting its dual role in the coordination of cell cycle regulation and myogenic differentiation of progenitor cells during myogenesis. This study identifies CTTN as an essential regulator of myogenic differentiation via affecting the actin remodeling-MRTFA-SRF signaling axis and cell proliferation.

Machine learning-based prognostic model of lactylation-related genes for predicting prognosis and immune infiltration in patients with lung adenocarcinoma

BackgroundHistone lactylation is a novel epigenetic modification that is involved in a variety of critical biological regulations. However, the role of lactylation-related genes in lung adenocarcinoma has yet to be investigated.MethodsRNA-seq data and clinical information of LUAD were downloaded from TCGA and GEO datasets. Unsupervised consistent cluster analysis was performed to identify differentially expressed genes (DEGs) between the two clusters, and risk prediction models were constructed by Cox regression analysis and LASSO analysis. Kaplan–Meier (KM) survival analysis, ROC curves and nomograms were used to validate the accuracy of the models. We also explored the differences in risk scores in terms of immune cell infiltration, immune cell function, TMB, TIDE, and anticancer drug sensitivity. In addition, single-cell clustering and trajectory analysis were performed to further understand the significance of lactylation-related genes. We further analyzed lactate content and glucose uptake in lung adenocarcinoma cells and tissues. Changes in LUAD cell function after knockdown of lactate dehydrogenase (LDHA) by CCK-8, colony formation and transwell assays. Finally, we analyzed the expression of KRT81 in LUAD tissues and cell lines using qRT-PCR, WB, and IHC. Changes in KRT81 function in LUAD cells were detected by CCK-8, colony formation, wound healing, transwell, and flow cytometry. A nude mouse xenograft model and a KrasLSL-G12D in situ lung adenocarcinoma mouse model were used to elucidate the role of KRT81 in LUAD.ResultsAfter identifying 26 lactylation-associated DEGs, we constructed 10 lactylation-associated lung adenocarcinoma prognostic models with prognostic value for LUAD patients. A high score indicates a poor prognosis. There were significant differences between the high-risk and low-risk groups in the phenotypes of immune cell infiltration rate, immune cell function, gene mutation frequency, and anticancer drug sensitivity. TMB and TIDE scores were higher in high-risk score patients than in low-risk score patients. MS4A1 was predominantly expressed in B-cell clusters and was identified to play a key role in B-cell differentiation. We further found that lactate content was abnormally elevated in lung adenocarcinoma cells and cancer tissues, and glucose uptake by lung adenocarcinoma cells was significantly increased. Down-regulation of LDHA inhibits tumor cell proliferation, migration and invasion. Finally, we verified that the model gene KRT81 is highly expressed in LUAD tissues and cell lines. Knockdown of KRT81 inhibited cell proliferation, migration, and invasion, leading to cell cycle arrest in the G0/G1 phase and increased apoptosis. KRT81 may play a tumorigenic role in LUAD through the EMT and PI3K/AKT pathways. In vivo, KRT81 knockdown inhibited tumor growth.ConclusionWe successfully constructed a new prognostic model for lactylation-related genes. Lactate content and glucose uptake are significantly higher in lung adenocarcinoma cells and cancer tissues. In addition, KRT81 was validated at cellular and animal levels as a possible new target for the treatment of LUAD, and this study provides a new perspective for the individualized treatment of LUAD.

Unveiling Key Genes Modulating Retinal Cell Survival and Autophagy in Glaucoma.

Glaucoma is a leading cause of irreversible blindness, with rising incidence globally. Effective treatment is challenging due to limited understanding of the disease mechanisms. Growth factor activity is crucial in glaucoma, with potential to reduce retinal ganglion cell (RGC) apoptosis and slow disease progression. This study aims to identify and analyze differentially expressed genes (DEGs) involved in growth factor activity to uncover new therapeutic targets. We analyzed the GSE9944 dataset from the Gene Expression Omnibus (GEO) to identify DEGs associated with glaucoma, resulting in 94 DEGs, including 29 down-regulated and 65 up-regulated genes. Functional enrichment and protein-protein interaction (PPI) network analyses were conducted using bioinformatics tools, highlighting the roles of Bone Morphogenetic Protein 1 (BMP1), Pleiotrophin (PTN), and f fibroblast Growth Factor 7 (FGF7). Aberrant expression vectors for these genes were transfected into RGCs derived from a glaucoma model to evaluate their impact on cell viability, apoptosis, and autophagy. Bioinformatics analysis of the GSE9944 dataset identified 94 DEGs, with 29 down-regulated and 65 up-regulated genes. Functional enrichment analysis revealed that these DEGs were involved in pathways related to growth factor activity, apoptosis, and autophagy, processes highly relevant to glaucoma pathogenesis. PPI network analysis identified BMP1, PTN, and FGF7 as central hub genes involved in extracellular matrix organization and growth factor signaling. In experimental validation using RGCs, we found that up-regulation of BMP1 significantly enhanced RGC viability and reduced apoptosis. Conversely, silencing PTN and FGF7 provided protective effects, enhancing RGC survival. Silencing BMP1 and upregulating PTN and FGF7 led to increased RGC apoptosis. Additionally, BMP1 was found to inhibit autophagy in RGCs, whereas PTN and FGF7 promoted autophagic activity, suggesting differential regulatory roles in glaucoma pathogenesis. Overall, BMP1, PTN, and FGF7 play critical roles in the regulation of RGC activity and autophagy in glaucoma, making them promising molecular targets for future therapeutic interventions.

Self-assembled peptide-based nanofibers for cardiovascular tissue regeneration.

Cardiovascular diseases are the leading cause of death worldwide, claiming millions of lives every year. Cardiac tissue engineering has emerged as a versatile option for repairing cardiac tissue and helping its regeneration. The use of nanomaterials, particularly nanofiber-based scaffolds combined with biomolecular cues like peptides, has significantly improved the compatibility and efficacy of the scaffolds for cardiac tissue regeneration. By utilising the self-assembly properties of peptides to create nanofiber scaffolds, we can achieve stability that closely mimics the natural components of cardiac tissue, making them perfect for cardiac tissue regeneration. In this review, we highlighted the dynamic process of self-assembly into nanofibers and the use of various self-assembled nanofibers for cardiovascular tissue regeneration, focusing on their roles in antithrombotic, angiogenic, differentiation, proliferation, and anti-atherosclerotic interventions.

Explicit Memory, Implicit Memory, and the Hippocampus: Insights From Early Neuroimaging Studies.

During the 1980s and 1990s, much memory research focused on the differential role of the hippocampus in various forms of memory. My work on the distinction between explicit and implicit memory led me to become involved in several early neuroimaging studies that made use of cognitive paradigms to investigate the conditions in which hippocampal activity does and does not occur, and to address the theoretical implications of these findings. Here, I summarize two such projects and some of the personal backstory associated with them.

Individual differences in the evolution of causal illusions.

In this research, we investigated individual differences in the formation and persistence of causal illusions. In a re-analysis of existing data, we identified two clusters of participants - persistent and adjusting - based on their trajectories in learning from repeated exposure to null contingencies. The persistent cluster maintained stable causal illusions, while the adjusting cluster demonstrated a reduction over time. This re-analysis provided a nuanced understanding of individual differences in causal learning, emphasizing the differential role of probability estimations in predicting causal judgements. These findings were replicated in a subsequent study, highlighting the robustness of the identified effects. In a pre-registered study, we extended the paradigm to include a second phase (active phase) to assess how individual differences in causal illusion trajectories in the passive phase would manifest when participants had agency in the information gathering process. The results were consistent with those of the two previous studies and confirmed our primary hypothesis that the adjusting cluster would exhibit a lower tendency to introduce the candidate cause on learning trials, and would, therefore, observe a higher frequency of cause-absent trials. Together, these studies provide comprehensive insights into the underpinnings of causal illusion development and persistence, potentially informing de-biasing interventions.

Iron deficiency negatively affects behavioral measures of learning, indirect neural measures of dopamine, and neural efficiency.

Iron deficiency (ID) is the most prevalent nutrient deficiency in the world, with a growing literature documenting the negative effects of ID on perception, attention, and memory. Animal models of ID suggest that dysregulation of dopamine is responsible for the deficits in memory. However, evidence that ID affects dopamine in humans is extremely limited. We report the results of a study involving college-aged women with and without ID learning two different category structures - a rule-based and an information-integration structure - selected based on the putative differential role of dopamine in learning these two structures. ID non-anemic (IDNA) and iron-sufficient (IS) women completed 1200 learning trials for each structure. EEG was collected to assess the effects of ID on features affected by dopaminergic state: error-related negativity (ERN) and positivity (Pe), feedback-related negativity (FRN), and task-related blink rate. In addition, we examined the EEG data for dynamics distinguishing IDNA from IS women, including a measure of neural efficiency. Both groups of women were able to learn both structures. However, IDNA women were initially slower and less accurate than IS women, specifically for the rule-based structure. There were large and persistent group differences in brain dynamics and neural efficiency measures. The results are discussed with respect to the selective impact of ID on initial rule-based learning and the persistent effect of ID on dopamine signaling and energetic efficiency.

Rarγ-Foxa1 signaling promotes luminal identity in prostate progenitors and is disrupted in prostate cancer.

Retinoic acid (RA) signaling is a master regulator of vertebrate development with crucial roles in body axis orientation and tissue differentiation, including in the reproductive system. However, a mechanistic understanding of how RA signaling governs cell lineage identity is often missing. Here, leveraging prostate organoid technology, we show that RA signaling orchestrates the commitment of adult mouse prostate progenitors to glandular identity, epithelial barrier integrity, and specification of prostatic lumen. RA-dependent RARγ activation promotes the expression of Foxa1, which synergizes with the androgen pathway for luminal expansion, cytoarchitecture and function. FOXA1 mutations are common in prostate and breast cancers, though their pathogenic mechanism is incompletely understood. Combining functional genetics with structural modeling of FOXA1 folding and chromatin binding analyses, we discover that FOXA1F254E255 is a loss-of-function mutation compromising its transcriptional function and luminal fate commitment of prostate progenitors. Overall, we define RA as an instructive signal for glandular identity in adult prostate progenitors. Importantly, we identify cancer-associated FOXA1 indels affecting residue F254 as loss-of-function mutations promoting dedifferentiation of adult prostate progenitors.

The differential role of AI-operated WCF in L2 students’ noticing of errors and its impact on writing scores

Despite the plethora of studies on the role of noticing in second language learning, little is known about the role of AI-operated feedback in noticing errors and uptake during and after writing. To address this gap, this study primarily aimed to investigate the impact of feedback modes and timing on L2 students’ noticing of errors and writing scores. In pursuit of this objective, a quasi-experimental design was adopted in which 75 university undergraduate EFL participants were randomly distributed into four groups representing four feedback conditions. These included receiving feedback (a) from Grammarly while composing, (b) from E-rater after composing, (c) from the teacher after composing, and (d) a no feedback condition. Three argumentative prompts taken from IELTS task 2 were used as writing tasks and Noticing Sheets (NSs) were employed in recording and analyzing participants’ noticing. The findings generally demonstrate that participants exhibited increased noticing with teacher input compared to AI-operated feedback from Grammarly and E-rater, yet they still noticed errors when using the AI systems during and after writing. The results also show that noticing occurred in all feedback conditions, including the fourth one which involved no feedback from any source. This suggests that feedback is not an absolute prerequisite for noticing, but the source and timing of such corrective feedback can influence students’ noticing focus and intensity. The study provides further insights into the role of noticing in L2 learning and offers practical implications for teachers seeking to optimize conventional and automated feedback strategies for enhanced language learning.

Differential roles of problematic media use by mothers and toddlers in the relation between parenting stress and toddlers’ socioemotional development

The increasing prevalence of digital media devices in families with young children has raised concerns over problematic media use. However, the link between toddlers’ problematic media use, their socioemotional development, and the influence of parental factors remains unclear. The current study examined the roles of problematic media use by both mothers and toddlers in the association between maternal parenting stress and toddlers’ socioemotional development. Participants comprised 215 mothers and 73 lead child care classroom teachers of toddlers aged 24–36 months enrolled in child care centers in South Korea. Mothers reported parenting stress and problematic media use by themselves and their toddlers, and child care teachers reported toddlers’ socioemotional development. The results from structural equation modeling analysis revealed that maternal parenting stress was related to toddlers’ socioemotional development both directly and indirectly through toddlers’ problematic media use, both alone and in conjunction with mothers’ problematic media use. Although maternal parenting stress was related to mothers’ problematic media use, mothers’ problematic media use was not directly related to toddlers’ socioemotional development. These findings underscore the importance of toddlers’ problematic media use in their socioemotional development and highlight the need for comprehensive approaches that address family dynamics, such as maternal parenting stress and problematic media use.

Viscoelastic Alginate‐Based Hydrogels: Regulating Neural Stem Cell Behavior Through Viscosity

ABSTRACTThe mechanical properties of the extracellular matrix (ECM) play a crucial role in cell adhesion, proliferation, and differentiation. In this study, a series of viscoelastic alginate‐based blend hydrogels with tunable viscosity were prepared to investigate the effects of their viscosity on the spreading and viability of NE‐4C neural stem cells. The hydrogels with the same initial modulus but different viscosities were obtained by adjusting the degree of crosslinking through covalently and ionically crosslinked techniques. The study results indicated that at a low initial modulus, an increase in the viscosity of the viscoelastic substrate could lead to a rise in the spreading area of NE‐4C neural stem cells, along with the formation of synapses, suggesting that an increase in substrate viscosity is beneficial for cell adhesion and spreading. Furthermore, the survival rate of NE‐4C neural stem cells on a high‐viscosity matrix is significantly higher than on a low‐viscosity matrix, as the high‐viscosity matrix provides a more stable microenvironment for the cells. These results can not only enhance the understanding of the effect of the viscoelasticity of biomaterials on neural stem cell behavior but also provide experimental data and theoretical support for designing new biomaterials suitable for neural tissue engineering.

MS4A3 regulates hematopoietic myeloid differentiation through ROS/TGF-β/p38MAPK pathway

The hematopoietic homeostasis relies on the intricate regulation of hematopoietic stem cells during their proliferation and differentiation. Myeloid differentiation disorders can lead to chronic myeloid leukemia and acute myeloid leukemia. Previous studies have shown increased expression of MS4A3 in myeloid cells, suggesting that MS4A3 may play a critical role in hematopoietic myeloid differentiation. However, the underlying mechanism and its role in hematopoietic myeloid differentiation require further elucidation. In this study, using K562 cell lines with MS4A3 over-expression (oeMS4A3) and MS4A3 knockdown (shMS4A3), we demonstrated that the overexpression of MS4A3 resulted in an augmented skewing towards myeloid differentiation and cell cycle arrest at G0/G1. In addition, inhibition of ROS, TGF-β, and p38MAPK in oeMS4A3 K562 cells attenuated the skewing of myeloid differentiation. Furthermore, in vivo experiments revealed a slight myeloid differentiation suppression tendency in MS4A3 knockout mice. Taken together, we show that MS4A3 overexpression promote myeloid differentiation skewing through the activation of the ROS/p38MAPK/TGFβ pathway. This study underscored the role of MS4A3 in the hematopoietic myeloid differentiation.

The Multifaceted Roles of Hippo-YAP in Cardiovascular Diseases.

The Hippo-yes-associated protein (YAP) signaling pathway plays a crucial role in cell proliferation, differentiation, and death. It is known to have impact on the progression and development of cardiovascular diseases (CVDs) as well as in the regeneration of cardiomyocytes (CMs). However, further research is needed to understand the molecular mechanisms by which the Hippo-YAP pathway affects the pathological processes of CVDs in order to evaluate its potential clinical applications. In this review, we have summarized the recent findings on the role of the Hippo-YAP pathway in CVDs such as myocardial infarction, heart failure, and cardiomyopathy, as well as its in CM development. This review calls attention to the potential roles of the Hippo-YAP pathway as a relevant target for the future treatment of CVDs.

IRF2BP2 binds to a conserved RxSVI motif of protein partners and regulates megakaryocytic differentiation

IRF2BP2 is a transcriptional coregulator that plays diverse regulatory roles in various cellular processes in either IRF2-dependent or IRF2-independent manner through interactions with protein partners via its RING domain; however, the underlying molecular mechanisms remain unclear. In this study, we conduct a motif discovery search on the sequences of interacting proteins IRF2 and VGLL4 of IRF2BP2 and identify a conserved RxSVI motif. Biochemical and structural data reveal that the RING domain binds to the motif-containing peptides of IRF2 and VGLL4 with comparable affinities and in a similar manner. The motif-containing peptides tend to form a short loop along with a short β-strand, which facilitates effective recognition and tight binding by the RING domain. Further exploration of this motif in the human proteome identifies the transcription factor ZBTB16 as an interacting protein of IRF2BP2. Biochemical, structural, and cell biological data demonstrate that the RING domain binds to the motif-containing peptide of ZBTB16 in a manner similar to that of IRF2 and VGLL4. Moreover, IRF2BP2 plays a crucial regulatory role in megakaryocytic differentiation through interaction with ZBTB16. These findings elucidate the molecular basis for how IRF2BP2 can engage with different protein partners, thereby exerting diverse regulatory functions in many cellular processes.

Emerging functions of FMNL1 in myeloid neoplasms: insights from bioinformatics to biological and pharmacological landscapes.

Myeloid neoplasms encompass disorders characterized by abnormal myeloid cell proliferation and differentiation, including myelodysplastic syndromes (MDS), myeloproliferative neoplasms, acute myeloid leukemia (AML), and chronic myeloid leukemia (CML). Formin-like protein 1 (FMNL1) is involved in the regulation of the actin cytoskeleton and is predominantly expressed in hematopoietic cells. Given its role in leukemia cell proliferation, survival, migration, and invasion, this study investigates FMNL1 expression in normal hematopoiesis and myeloid neoplasms and explores associations with clinical-laboratory characteristics, mutational status, and survival outcomes in AML. Transcript levels of FMNL1 from several blood-forming cell populations and myeloid neoplasms were extracted from publicly available databases. Myeloid neoplasm cell lines were used for gene/protein expression and cell differentiation studies. Functional genomics analysis was performed using RNA-seq data from The Cancer Genome Atlas (TCGA) AML study, and drug sensitivity predictions were investigated using Beat AML and Genomics of Drug Sensitivity in Cancer (GDSC) datasets. Statistical analyses assessed the impact of FMNL1 expression on clinical outcomes. FMNL1 was highly expressed in metamyelocytes, neutrophils, and monocytes compared to hematopoietic stem cells, and its expression increased with granulocytic differentiation. FMNL1 expression was elevated in AML and CML patients compared to healthy donors. FMNL1 expression was not significantly associated with clinical-laboratory characteristics or survival outcomes but showed a higher frequency of WT1 transcription factor (WT1) mutations with low FMNL1 expression in AML patients. High FMNL1 expression in AML correlated with immune response and inflammatory activity pathways. FMNL1 mRNA levels influenced drug sensitivity in AML models, with correlations observed for specific antineoplastic agents. FMNL1 plays a potential role in granulocyte differentiation and function, and its differential expression is linked to critical signaling pathways in leukemogenesis and inflammation. These findings highlight FMNL1's potential therapeutic implications in myeloid neoplasia, warranting further investigation.

The effect of osteoclasts on epigenetic regulation by long non-coding RNAs in osteoblasts grown on titanium with nanotopography

Titanium (Ti) implant osseointegration is regulated by the crosstalk among bone cells that are affected by epigenetic machinery, including the regulation of long non-coding RNAs (lncRNAs). Nanotopography Ti (Ti Nano) induces the differentiation of osteoblasts that are inhibited by osteoclasts through epigenetic mechanisms. Thus, we hypothesize that osteoclasts affect lncRNA expression in Ti Nano-cultivated osteoblasts. Osteoblasts were grown on Ti Nano and Ti Control that were then co-cultured with osteoclasts for 48 h. Using RNAseq, we identified 252 modulated lncRNAs in osteoblasts regulated by both surfaces of Ti, but mainly in Ti Nano-cultivated osteoblasts. A negative correlation was observed between Kcnq1ot1 and the mRNAs of Alpl, Bglap, Bmp8a, Col1a1, and Vim in Ti Nano-cultivated osteoblasts with osteoclasts. The pull-down indicated that Bglap mRNA is a direct target of Kcnq1ot1, with enhanced physical interaction in Ti Nano-cultivated osteoblasts, and greater osteoclast inhibition than the Ti Control. The bone marker expression at the levels of mRNA and protein were downregulated by the Kcnq1ot1 silencing, indicating its pivotal role in osteoblast differentiation. These results showed that nanostructured Ti surface modulates the osteoblast–osteoclast crosstalk, at least in part, through the regulation of lncRNA expression in osteoblasts. We demonstrate that the lncRNA Kcnq1ot1 directly interacts with Bglap mRNA, and this interaction is enhanced by nanotopography and reduced by osteoclasts with greater intensity in Ti Nano-cultivated osteoblasts. These findings confirm the molecular mechanisms associated with the high osteogenic potential of nanotopography and can potentially support osteointegration of dental and skeletal prostheses.

Nuclear Factor I family members are key transcription factors regulating gene expression

The Nuclear Factor I (NFI) family of transcription factors (TFs) plays key roles in cellular differentiation, proliferation, and homeostasis. As such, NFI family members engage in large number of interactions with other proteins and the chromatin. However, despite their well-established significance, the NFIs interactomes, their dynamics, and their functions have not been comprehensively examined. Here, we employed complementary omics-level techniques, i.e. interactomics (affinity purification mass spectrometry (AP-MS) and proximity-dependent biotinylation (BioID)), and chromatin immunoprecipitation sequencing (ChIP-Seq), to obtain a comprehensive view of the NFI proteins and their interactions in different cell lines. Our analyses included all four NFI family members, and a less studied short isoform of NFIB (NFIB4), which lacks the DNA binding domain. We observed that, despite exhibiting redundancy, each family member had unique high-confidence interactors and target genes, suggesting distinct roles within the transcriptional regulatory networks. The study revealed that NFIs interact with other TFs to co-regulate a broad range of regulatory networks and cellular processes. Notably, time-dependent proximity-labeling unveiled a highly dynamic nature of NFI protein-protein interaction networks and hinted at temporal modulation of NFI interactions. Furthermore, gene ontology (GO) enrichment analysis of NFI interactome and targetome revealed the involvement of NFIs in transcriptional regulation, chromatin organization, and cellular signaling pathways, and pathways related with cancer. Additionally, we observed that NFIB4 engages with proteins associated with mRNA regulation, which suggests that NFIs have roles beyond traditional DNA binding and transcriptional modulation. We propose that NFIs may function as potential pioneering TFs, given their role in regulating the DNA binding ability of other TFs and their interactions with key chromatin remodeling complexes, thereby influencing a wide range of cellular processes. These insights into NFI protein-protein interactions and their dynamic, context-dependent nature provide a deeper understanding of gene regulation mechanisms and hint at the role of NFIs as master regulators.

Inhibiting autophagy further promotes Ginkgolide B's anti-osteoclastogenesis ability

Excessive osteoclast activity could cause skeletal diseases including osteoporosis. Additionally, autophagy plays an initial role in osteoclast differentiation and function. Ginkgolide B (GB), a key compound in Ginkgo biloba, improves bone mass and suppresses mature osteoclast formation in vitro. This study examines the role of GB role in regulating osteoclast formation via autophagy. Using murine bone marrow-derived macrophages in vitro, we explored GB's effects on autophagy and osteoclast formation. We also assessed bone loss prevention in an ovariectomized (OVX) mouse model using GB combined with an autophagy inhibitor. Tartrate-resistant acid phosphatase staining was used to observe osteoclast formation. Autophagy-related proteins and intracellular microtubule associated protein 1 light chain 3 beta puncta were observed using western blotting and immunofluorescence. The impact of GB on OVX mice was evaluated using micro-computed tomography and hematoxylin and eosin staining. GB directly promoted autophagy in osteoclast precursors (OCPs), but inhibited osteoclast differentiation by reducing receptor activator of nuclear factor kappa B ligand (RANKL)-induced autophagy. GB inhibits the phosphorylation of RANKL downstream signaling pathways, such as Jun N-terminal kinase (JNK), p38, and extracellular signal-regulated kinase (ERK). Anisomycin (ANI), a JNK activator, reversed GB's inhibitory effects on RANKL-induced autophagy and osteoclastogenesis. Inhibiting autophagy using 3-Methyladenine (3-MA) or small interfering RNA significantly suppressed osteoclast differentiation both in vitro and in vivo. Our findings suggested that GB inhibits osteoclast formation by decreasing JNK phosphorylation and autophagy under RANKL stimulation. Interestingly, GB also directly promotes autophagy in OCPs. Thus, GB markedly reduces osteoclast differentiation and prevents bone loss, with its anti-osteoclastogenesis effect being enhanced by 3-MA. Accordingly, inhibiting GB-induced direct autophagy could further increase its therapeutic effect on bone disease resulting from excessive osteoclast activity.

Differential TLR2 and TLR4 mediated inflammatory and apoptotic responses in asymptomatic and symptomatic Leptospira interrogans infections in canine uterine tissue

Leptospirosis is major zoonotic disease with global implications, affecting both domestic animals and humans. It is caused by Leptospira interrogans (L. interrogans), which can damage multiple organs, including the kidneys, liver, testes, and uterus. Despite this, L. interrogans can also persist asymptomatically in tissues, akin to nonpathogenic strains. The mechanisms driving asymptomatic infections remain poorly understood. This study investigated the role of L. interrogans in asymptomatic infection within the uterine tissue of canines, focusing on the differential expression of Toll-like receptors (TLRs)2 and 4 and their roles in inflammatory and apoptotic pathways. We hypothesized that TLR2 and TLR4 coexpression is crucial for eliciting inflammation and apoptosis, whereas TLR4 alone might be insufficient. Our findings revealed that in symptomatic infections, both TLR2 and TLR4 are coexpressed, leading to markedly elevated levels of the proinflammatory cytokines IL-10, IL-1β, TNF-α, and IL-6. This enhanced inflammatory response is further evidenced by increased CD4 expression, indicating robust T helper cell activation. In contrast, asymptomatic infections are characterized by exclusive TLR4 expression, with inflammatory markers remaining at baseline levels. Additionally, we observed that L. interrogans induces apoptosis in symptomatic animals through TLR2 and TLR4 mediated activation of Caspase 8 and Caspase 3. These findings illustrate that L. interrogans drives both inflammation and apoptosis via the combination of TLR2 and TLR4 actions. When only TLR4 is activated, the immune response is insufficient, resulting in an asymptomatic disease course. This study provides novel insights into the differential roles of TLR receptors in leptospirosis, offering potential directions for targeted therapeutic strategies.

The incidence and significance of iliac vein stenosis in patients with deep vein thrombosis.

ObjectiveThe role of iliac vein stenosis (IVS) in the development of DVT is poorly understood. This study determines the incidence of IVS in patients diagnosed with DVT and assesses its impact on presentation and outcomes. Our hypothesis is that the presence of IVS increases the risk of long-term ipsilateral DVT recurrence. MethodsA retrospective study of the electronic medical records of consecutive adult patients treated for lower extremity DVT was performed. Only patients with cross-sectional imaging (CT or MR with intravenous contrast) were included. Patient and DVT characteristics were recorded. Cross-sectional imaging was reviewed for the presence or absence of ipsilateral IVS (≥50%). Patients were divided into 2 groups based on the presence or absence of IVS to compare characteristics and outcomes. Subgroup analyses on patients with provoked DVT, Cancer-related DVT, and unprovoked DVT were performed. ResultsThere were 548 patients with DVT and 32% had evidence of ipsilateral IVS on cross-sectional imaging. There were no significant differences in baseline characteristics or treatment methods between the 2 groups. There was a trend toward patients with IVS having less incidence of PE on presentation (22.9% vs 29.7%, P=0.1) but that difference did not reach statistical significance in the overall comparison. Subgroup analysis in patients with Cancer-related DVT (n=227) showed that patients with IVS were significantly more likely to develop ipsilateral recurrent DVT compared to patients with no IVS (12.9% vs 4.5%, P=0.045). Patients with unprovoked DVT with IVS had significantly lower PE on presentation compared to patients with unprovoked DVT without IVS (24.2% vs 39.8%, P<0.03). ConclusionIpsilateral ≥50% IVS is present in approximately a third of patients presenting with DVT. The presence of IVS seems to play a differential role in ipsilateral DVT recurrence and prevention of pulmonary embolization in different groups of patients presenting with DVT.