Molecular Medicine最新文献

Despite the established efficacy of hydroxyurea (HU) in increasing fetal hemoglobin (Hb F) levels in patients with intermedia beta-thalassemia (β-thal) and sickle cell anemia, the precise molecular mechanisms underlying these effects remain largely elusive. Understanding these mechanisms is paramount for identifying alternative therapeutic approaches to increase Hb F production while minimizing adverse effects. In this study, we employed weighted gene co-expression network analysis (WGCNA) to investigate the molecular underpinnings of γ-globin switching within GSE90878 dataset. Leveraging this information, we aimed to predict the transcriptome network and elucidate the mechanism of action of HU and Metformin (Met) on this network comprehensively. Through bioinformatic analysis, we identified IGF2BP1 and GCNT2 as key regulators of the γ-globin switching mechanism. To experimentally validate these findings, we utilized the K562 cell line as an erythroid model. Cells were treated with HU (50, 100, and 150 µM) and Met (50, 100, and 150 µM) for 24, 48, and 72 h. The expression levels of the GCNT2, γ-globin, IGF2BP1, miR-199a/b-5p, miR-451-5p and miR-144-3p were quantified using real-time polymerase chain reaction (qPCR). Our results revealed that treatment with HU (150 µM), Met (100 µM), and combination of HU-Met (150/100 µM) significantly increased IGF2BP1 expression by 6.2, 5.3, and 7.1-fold, respectively, after 24 h treatment. Furthermore, treatment with HU (50 µM), Met (50 µM) and HU/Met (50/50 µM) for 24 h led to a 3.3, 1.2, and 5-fold decrease in GCNT2 gene expression, respectively. Notably, the highest levels of γ-globin expression and Hb F production were observed with HU (100 µM), Met (50 µM), and HU/Met (100/50 µM). This study provides compelling evidence that HU and Met significantly enhance γ-globin expression and Hb F production in the K562 cell line. Our findings suggest that these drugs exert their effects by modulating the expression of IGF2BP1 and GCNT2, thus offering valuable insights into potential therapeutic strategies for disorders characterized by low Hb F levels.

Background: Temporomandibular joint osteoarthritis (TMJ-OA) is a disease characterized by cartilage degradation and synovial inflammation, with limited effective treatment currently. Synovial macrophage polarization is pivotal in TMJ-OA progression, making it a promising therapeutic aspect. This study investigated the effects of S-propargyl-cysteine (SPRC), an endogenous H2S donor, on macrophage polarization and its therapeutic potential in alleviating TMJ-OA.

Methods: A MIA-induced TMJ-OA rat model and LPS-stimulated RAW264.7 macrophages were employed to evaluate the effects of SPRC in vivo and in vitro. TMJ bone and cartilage were analyzed via micro-CT and histological methods, while macrophage polarization markers expression were assessed via RT-qPCR, western blot, and immunofluorescence. RNA sequencing was performed on macrophages, and the JAK2/STAT3 signaling pathway was validated using the JAK2-specific inhibitor AG490. The direct effects of SPRC on rat primary condylar chondrocytes were examined by evaluating ECM synthesis and degradation. Co-culture experiments further assessed macrophage-chondrocyte interactions.

Results: SPRC significantly alleviated cartilage and bone damage in the TMJ-OA rat model, as demonstrated by improved bone volume and cartilage structure. SPRC reduced pro-inflammatory M1 macrophage infiltration and enhanced anti-inflammatory M2 macrophage polarization. SPRC effectively inhibited the JAK2/STAT3, leading to reduction of inflammatory markers, including TNF-α, IL-6, and iNOS. Co-culture experiments revealed that SPRC-treated macrophage-conditioned medium improved chondrocyte metabolic activity and restored ECM integrity.

Conclusions: SPRC-modulated macrophage polarization alleviates TMJ-OA via JAK/STAT downregulation, thereby reducing synovial inflammation and cartilage degradation. These findings position SPRC as a promising therapeutic candidate for TMJ-OA and provide insights into novel strategies targeting macrophage polarization and synovium-cartilage crosstalk.

Osteoporosis (OP) is a common systemic metabolic bone disease characterized by the decrease in bone mass and hyperactivity of osteoclasts. ACT001 is approved as an orphan drug by FDA and has shown multiple protective effects against tissue injury. However, its role in prevention of osteoclast differentiation and the underlying mechanisms have not been elucidated. Herein, we show that ACT001 inhibited RANKL-induced osteoclast differentiation and F-actin ring formation through suppressing the expression of Nfatc1, TRAP, Ctsk, Dc-stamp without obvious cytotoxicity in vitro. ACT001 restrained the phosphorylation of NF-κB and the activation of NLRP3 inflammasome, thereby decreased the expression of pyroptosis-related protein. (GSDMD, caspase-1, IL-1β, IL-18). Consistent with ACT001, the NLRP3 inflammasome inhibitor MCC950 treatment also suppressed the osteoclastogenesis through inhibiting the transcriptional activation of Nfatc1. Furthermore, ACT001 protected ovariectomy-induced bone loss in mice, reduced the number of osteoclasts, downregulated the expression of NLRP3 and IL-1β. These data indicate that ACT001 can reduce RANKL-induced osteoclast differentiation through suppressing the NF-κB/NLRP3 pathway, and attenuate the bone loss induced by estrogen-deficiency, suggesting its therapeutic potential for bone homeostasis maintenance and osteoporosis treatment.

Chronic Obstructive Pulmonary Disease (COPD) is a complex and diverse respiratory disorder, characterized by ongoing respiratory symptoms and restricted airflow. The major clinical manifestations typically encompass chronic cough, sputum production, and wheezing. The main pathological characteristics involve infiltration of inflammatory cells, overproduction of mucus, and damage to the alveolar walls. The underlying causes of COPD are complex and remain incompletely elucidated, thought to originate from the combined effect of various factors. Lipids, as hydrophobic molecules, fulfill three fundamental functions: energy storage, membrane biosynthesis, and signal transduction. Lipid metabolism is intricately intertwined with various metabolic pathways and plays a pivotal role in the complex pathogenesis of COPD. Delving into lipid metabolism, as well as the particular modifications and roles of lipid molecules in cells, is of paramount importance in the context of COPD. This review primarily aims to elucidate the role of fatty acid metabolism in the onset and progression of COPD. Additionally, it examines the potential of lipid metabolism reprogramming as a promising therapeutic approach, illuminating new paths for the management and treatment of this disabling respiratory condition.

Background: Cancer-induced bone pain (CIBP) is a complex chronic pain with poorly understood mechanisms. The anterior cingulate cortex (ACC) plays a critical role in processing and modulating chronic pain. This study investigates how the GluR2 receptors (calcium impermeable AMPA receptors) in ACC glutamatergic neurons regulate CIBP.

Methods: The CIBP models were established by injecting Walker 256 cells into the tibia of SD rats. Paw withdrawal threshold (PWT) and paw withdrawal latency (PWL) were used as indicators of hyperalgesia. The immunofluorescence staining was employed to detect the expression of c-Fos in ACC and identify the subtypes of co-labeled c-Fos⁺ neurons. Real-time monitoring of calcium activity in ACC glutamatergic neurons was achieved through the fiber photometry. The excitability of glutamatergic neurons in ACC was modulated using chemicalgenetics and optogenetics techniques. The expression of GluR2 at the mRNA and protein level in ACC were assessed using RT-qPCR and Western blotting.

Results: There were significant reductions in PWT and PWL of CIBP rats after Walker 256 cell injection. The ACC of CIBP rats showed increased c-Fos expression compared to sham rats, with mainly activated c-Fos co-localized with glutamatergic neurons. Optogenetic or chemogenetic activation of ACC glutamatergic neurons led to increased hyperalgesia in sham rats, while suppression of their activity alleviated hyperalgesia in CIBP rats. Calcium activity in ACC glutamatergic neurons of CIBP rats was increased with suprathreshold stimulation of von Frey filament. Notably, surface GluR2 protein and mRNA were reduced in ACC of CIBP rats. Furthermore, overexpression of GluR2 by AAV-CaMKII-GluR2 injection was decreased c-Fos expression in ACC and alleviated hyperalgesia in CIBP rats.

Conclusions: These findings suggest that decreased surface GluR2 receptors in ACC glutamatergic neurons contribute to calcium activity and excessive excitability, thereby inducing CIBP in rats. Conversely, GluR2 overexpression in ACC glutamatergic neurons alleviates CIBP in rats. This study provides a new potential therapeutic approach for targeting the GluR2 receptor to alleviate CIBP for cancer patients.

Pancreatic cancer (PC) has high lethality due to multiple reasons, and its limited response to conventional chemotherapy like gemcitabine (GEM) is a non-negligible one. Therefore, our study introduces Chlorophyllin (CHL) as an effective therapeutic candidate to enhance the therapeutic efficacy of GEM. Our results demonstrate that the combination of CHL and GEM exhibits a significant synergistic anti-tumor effect by targeting multiple oncogenic processes in PC, including inhibiting cell proliferation, invasion, and migration, as well as inducing cell apoptosis. Further investigations of mechanism have revealed that CHL induces cuproptosis in PC cells through a multifaceted process, involving depleting cellular intracellular glutathione (GSH), increasing reactive oxygen species (ROS) levels, and subsequently upregulating the HSP70 protein in response to heightened oxidative stress. Additionally, CHL releases free Cu²⁺, binds to the Ferredoxin 1 (FDX1) protein, and ultimately leads to the oligomerization of Dihydrolipoamide S-Acetyltransferase (DLAT) proteins to amplify the copper toxicity within PC cells. Moreover, in vivo experiments have demonstrated that the combination of CHL and GEM effectively inhibits the growth of subcutaneously transplanted tumors while maintaining a favorable biosafety profile. In conclusion, our study identifies CHL as a potent enhancer of GEM's anti-tumor effects in PC through the induction of cuproptosis, thus providing a novel therapeutic avenue for patients with PC.

Background: Riboflavin kinase (RFK, also called flavokinase) is a catalytic enzyme that converts riboflavin to its active form in vivo. Dysfunction of the RFK gene has been associated with susceptibility to ischemic stroke. However, the protective role and mechanisms of RFK in ischemic stroke have not been elucidated.

Methods: Lentivirus-mediated RFK knock-up (RFK( +)) and knock-down (RFK(-)) were used to investigate the protective effect and mechanism of RFK in the rat middle cerebral artery occlusion (MCAO) model in vivo and in the oxygen and glucose deprivation (OGD) model of neurons in vitro; and the dependence of the protective effect of RFK on flavins was also investigated.

Results: We demonstrated that RFK was an endogenous protein against ischemia brain injury both in vivo and in vitro experiments. RFK inhibited cerebral infarction, cerebral edema and neuronal apoptosis after cerebral ischemia. Its mechanisms include inhibition of the protein expression of Caspase 12 and Caspase 3 induced by cerebral ischemia, and thus inhibiting endoplasmic reticulum stress (ERS) and neuronal apoptosis; the protective effect of RFK depends on the presence of the flavoprotein Ero1; exogenous riboflavin supplementation protected cortical neurons from ischemic injury and prolonged the lifespan in stroke-prone spontaneously hypertensive rats with low RFK gene function, but this protective effect is limited and cannot completely reverse the decreasing trend of neuronal tolerance to ischemic injury caused by RFK gene dysfunction; the protective effect of RFK against ischemic injury is independent of the presence of flavins and their concentrations.

Conclusions: The present study demonstrates that RFK is an important regulatory molecule against ischemia brain injury and its mechanism involves inhibition of ERS. The protective effect of RFK is independent of the presence of flavins and their concentrations. RFK deserves further investigation as a promising target gene for the detection of stroke susceptibility. Flavins may be used as a preventive or adjunctive treatments for ischemic brain injury.

Background: Colon anastomotic leakage (CAL) is a postoperative complication originating from disturbed colon anastomotic healing (CAH). Wound healing involves several well-coordinated stages, which have not been comprehensively studied for CAH or CAL. This study aims to provide transcriptional profiles of different intestinal layers of anastomotic tissues throughout distinct healing stages and to identify CAL-related genes.

Methods: Proximal colon anastomosis was constructed with 8 interrupted sutures in mice. Six hours, 24 h and 72 h after surgery, anastomotic complications were assessed. Transcriptional profiles of inner (mucosa and submucosa) and outer (muscularis externa) layer of the anastomotic and naive control tissues were analyzed with 3' bulk mRNA sequencing to identify the layer-specific healing and leakage pathways. Selective target genes differing between CAL and CAH were measured for their protein expression.

Results: Our data indicate that the mucosa/submucosa and muscularis externa enter inflammation stage at 6 h, proliferation stage at 24 h and tissue remodeling stage at 72 h during CAH. We observed that transcription profiles of the mucosa/submucosa, but not the muscularis externa, differ between CAH and CAL. Particularly, genes related to extracellular remodeling (including Col18a1 and Col16a1) and wound healing (Pdpn and Timp1) showed lower expression in the mucosa/submucosa of CAL tissue compared to CAH. Conformingly, protein levels for collagens as well IL-34 were decreased in CAL, while the TGF-β-pseudo-receptor BAMBI was increased in CAL compared to CAH tissues.

Conclusions: Mucosa/submucosa and muscularis externa are mostly in synchronization during the inflammation, proliferation, and extracellular remodeling stages during CAH. Transcriptional profiles within the anastomotic mucosa/submucosa differ between CAH and CAL in genes related to extracellular modelling and wound healing, indicating that genes of these pathways may contribute to CAL.

Background: Extrachromosomal circular DNA (eccDNA) has potential in tumor diagnosis, particularly for improving diagnostic accuracy and early cancer detection; however, many challenges remain in its application to clinical practice.

Methods: We conducted a Circle-Seq analysis on clinical samples at different stages of colorectal cancer progression to examine the dynamic changes of eccDNA during the progression of colorectal cancer. We used breakpoint-specific PCR to verify candidate eccDNAs identified by Circle-Seq. The results were further validated using the AOM/DSS-induced colorectal cancer model.

Results: There was an increase in the abundance of eccDNA with the progression of colorectal cancer. The genes associated with these eccDNA molecules were primarily related to signaling pathways involved in tumor development and metastasis. Our analysis also revealed that eccDNA abundance positively correlates with gene expression, and eccDNA derived from specific genes has potential value for the early diagnosis of tumors.

Conclusions: This study revealed a connection between eccDNA and colorectal cancer progression and highlights the clinical potential of eccDNA for the early diagnosis of colorectal cancer.