Inflammation plays a pivotal role in the advancement of ischemic stroke, and Formononetin has been recognized for its potential benefits due to its anti-inflammatory effects. Although Formononetin shows promise for reducing cerebral ischemic injury, its precise effectiveness and the underlying molecular mechanisms still need to be thoroughly explored. The research aimed to investigate Formononetin’s impact and mechanisms on ischemic brain damage.
In this study, both the ischemia/reperfusion (I/R) mouse model and the oxygen-glucose deprivation/reperfusion (OGD/R) cell model were used. The I/R mouse model was prepared using the middle cerebral artery occlusion (MCAO) method, while the OGD/R SH-SY5Y cell model was established using the oxygen-glucose OGD/R method. Hematoxylin and Eosin (H&E) staining, Tunnel fluorescence staining, and Nissl staining were employed to observe the effects of Formononetin on neuronal damage, apoptosis, and survival in I/R mouse brain tissue. Additionally, the effects of Formononetin on the levels of pro-inflammatory factors in I/R mice and OGD/R cells were detected using Real-Time Quantitative Polymerase Chain Reaction (RT-qPCR) and Enzyme-Linked Immunosorbent Assay (ELISA) methods. The c-Fos/Interleukin-10 (IL-10)/Signal Transducer and Activator of Transcription 3 (STAT3) signaling pathway in I/R mice and OGD/R cells was examined using RT-qPCR and Western Blot (WB). Furthermore, rescue validation was performed using targeted interventions of IL-10 and c-Fos, confirming that the c-Fos/IL-10/STAT3 signaling pathway is a key target of Formononetin.
Our findings reveal that Formononetin notably decreased infarct size and neuronal damage in vivo (p < 0.001). Additionally, Formononetin decreased inflammation and lowered levels of pro-inflammatory cytokines (p < 0.05). In cell models, Formononetin effectively suppressed neuronal injury induced by OGD/R and the related inflammatory markers (p < 0.001). Mechanistic studies showed that Formononetin enhances IL-10 expression in both models of ischemic brain injury, a process crucial for its protective effects against inflammation (p < 0.05). This regulation is facilitated by increased nuclear translocation of c-Fos, highlighting the c-Fos/IL-10/STAT3 pathway as a crucial mechanism of Formononetin’s neuroprotective and anti-inflammatory effects in cerebral ischemia (p < 0.05).
We found Formononetin alleviates inflammation associated with I/R injury by activating the c-Fos/IL-10/STAT3 pathway, which highlights the potential of Formononetin as a promising therapeutic approach for I/R injury.
The medicinal phytochemical oleandrin (Ole) is obtained from the Nerium oleander plant. The exact relationship between Ole-induced apoptosis and autophagy in gastric cancer (GC) is unclear despite the fact that it has outstanding anti-tumor capabilities. This research aimed to demonstrate how autophagy and Ole-induced apoptosis interact in GC.
The Cell Counting Kit (CCK)-8 assay and colony formation assays were employed to evaluate cell proliferation. Cellular apoptosis was evaluated with Calcein/Propidium Iodide (PI) assays and flow cytometry. Confocal and electron microscopes were employed to examine the morphology of autophagy. Protein concentrations were assessed by western blotting. Luciferase-positive HGC-27 cells were administered subcutaneously to Balb/c nude mice to evaluate Ole’s anti-tumor activity. Immunohistochemistry assessed Ki67 expression and H&E staining in tumor tissue.
Ole causes GC cells to undergo intracellular apoptosis and autophagy at low nanomolar doses, halting the cell cycle at the G0/G1 phase. Whereas 3-methyladenine (3-MA), the inhibitor of autophagy, counteracts the apoptosis generated by Ole in vitro and in vivo.
Ole may trigger apoptosis through the activation of autophagy in GC. It offers a secure and efficacious candidate drug for the treatment of tumors in the digestive system.
Ischemic stroke triggers inflammatory responses that lead to neuronal damage, with microglial polarization significantly influencing post-stroke inflammation. This study explores the role of Fc gamma receptor Ia (FCGR1A) in microglial polarization and its regulatory mechanisms in ischemic stroke.
Differentially expressed genes (DEGs) associated with ischemic stroke were identified using the GSE58294 dataset. Hub genes were found by analyzing protein–protein interaction (PPI) networks. BV2 microglia were subjected to oxygen–glucose deprivation/reoxygenation (OGD/R) to mimic ischemic conditions in vitro, and FCGR1A and inflammatory marker levels were assessed. Besides, BV2 cells were stimulated with lipopolysaccharide (LPS) and interferon-gamma (IFN-γ) to induce M1 polarization, and the effects of FCGR1A overexpression and knockdown on cytokine production and microglial polarization were evaluated. The function of the AMP-activated protein kinase (AMPK)-mTOR pathway in regulating microglial polarization was further investigated using the mTOR inhibitor rapamycin (RAP).
From the 327 DEGs identified, FCGR1A was chosen as a hub gene. OGD/R treatment of BV2 cells produced a time-dependent rise in FCGR1A, induction of brown adipocytes 1 (Iba1), and interleukin 6 (IL-6) expression, indicating enhanced inflammation. FCGR1A overexpression induced a proinflammatory response and promoted M1 polarization, whereas FCGR1A knockdown reduced inflammation and shifted toward an anti-inflammatory M2 phenotype. Inhibition of the mTOR pathway using RAP, combined with FCGR1A knockdown, significantly enhanced AMPK activation and promoted a shift toward an anti-inflammatory M2 phenotype.
FCGR1A modulates microglial polarization by affecting the AMPK–mTOR signaling pathway in ischemic conditions. Targeting FCGR1A and related pathways could offer new therapeutic strategies to lessen inflammation and facilitate the healing process after an ischemic stroke.
The C-X-C chemokine receptors (CXCR) chemokine receptor family, consisting of seven primary members (CXCR1–CXCR7), is crucial in regulating immune cell recruitment, angiogenesis, cellular proliferation, and maintaining skin homeostasis and immune functions. This review evaluates the expression and roles of CXCR receptors across a range of skin cells, including keratinocytes, fibroblasts, endothelial cells, melanocytes, and various immune cells such as T cells, dendritic cells, and macrophages. Aberrations in CXCR signaling have been associated with a variety of skin disorders, such as psoriasis, atopic dermatitis, acne, and skin cancers. Despite significant advancements in the field, several critical questions persist. These include the differential effects of CXCR signaling in distinct skin pathologies and the intricate interactions between CXCR receptors and their ligands within diverse skin microenvironments. Moreover, the therapeutic targeting of the CXCR family remains unresolved, necessitating further research into its long-term efficacy and possible adverse effects. Future investigations should prioritize these critical issues to develop more effective therapeutic strategies for managing skin diseases, ultimately improving patient outcomes.
Type 2 cardiorenal syndrome (CRS) is a complex disease characterized by the interplay between the heart and kidneys. The pathophysiology of type 2 CRS involves multiple molecular signaling pathways. Transient receptor potential melastatin 2 (TRPM2) is a reactive oxygen species (ROS)-sensitive and non-selective calcium-permeable cation channel, which plays a regulatory role in intracellular Ca2+ homeostasis. Thus, this study aimed to explore the biological functions and mechanisms of the ROS–TRPM2 signaling axis in type 2 CRS.
Type 2 CRS model rats (a rat model of type 2 CRS induced through left anterior descending coronary artery ligation combined with 5/6 total nephrectomy) and lipopolysaccharide (LPS)-induced CRS cell lines, human kidney-2 (HK-2), were transfected with small interfering RNA (siRNA) to knock down TRPM2 or a calcium ion channel activator Yoda1 to evaluate the involvement of the ROS–TRPM2 signaling axis on type 2 CRS. Changes in kidney tissue morphology were observed using H&E staining; cell viability and apoptosis were monitored using CCK-8, Annexin V-FITC/PI, and TUNEL kits, alongside quantitative real-time polymerase chain reaction (qRT-PCR), Western blot, ELISA, and immunofluorescence assays to confirm the interaction between ROS, TRPM2, and Ca2+.
TRPM2 is highly expressed in HK-2 cells after LPS stimulation and renal tissues of type 2 CRS rats. Intervention via TRPM2 improves injured cell viability, mitigates apoptosis, inhibits the inflammatory cytokines interleukin 10 (IL-10) and tumor necrosis factor-α (TNF-α), as well as indices of oxidative stress—malondialdehyde (MDA) and ROS—promotes total antioxidant capacity (T-AOC) expression, and alleviates pathological changes in CRS; Yoda1 promoted a contrasting effect to the biological effect induced by TRPM2 deletion.
TRPM2 is abnormally highly expressed in damaged kidneys during the pathogenesis of type 2 CRS. Silencing TRPM2 can inhibit inflammatory and oxidative stress responses, reduce cell apoptosis, promote survival, and alleviate pathological loss; this may be related to the inhibition of Ca2+ influx. This suggests that the ROS–TRPM2 signaling pathway is significant for CRS development, and TRPM2 may be an effective therapeutic target for type 2 CRS.
Matrix metalloproteinases (MMPs) and the tissue inhibitors of metalloproteinases (TIMPs) play critical roles in the pathogenesis of liver diseases, particularly in conditions such as fibrosis, cirrhosis, and liver cancer. This review highlights the diagnostic and prognostic potential of MMPs, emphasizing their involvement in metastasis, de-differentiation, and hepatic cell proliferation. Utilizing advanced reporter mouse models has proven instrumental in assessing intraglandular MMP activity and predicting metastatic risks, paving the way for targeted therapeutic interventions. Current research indicates that specific MMPs and TIMPs can serve as valuable biomarkers for liver function and disease progression, although a clear consensus on their clinical utility remains elusive. Ongoing studies explore MMP-targeted therapies with potential applications in liver disease management, particularly in reducing fibrosis and enhancing liver regeneration. Future directions in this field involve elucidating the roles of MMPs in ischemia and transplantation, with the aim of improving clinical outcomes. Emerging therapeutic strategies focus on achieving a balance between MMP activity and TIMP expression to optimize liver function, highlighting the need for organ-specific targeting. Overall, this comprehensive overview underscores the importance of MMPs and TIMPs in liver function and liver disease, as well as the necessity for further research to harness their potential in clinical practice.
Inborn errors of immunity (IEIs) are a group of more than 485 disorders that impair immune development and function with variable reported incidence, severity, and clinical phenotypes. A subset of IEIs blend increased susceptibility to infection, autoimmunity, and malignancy and are known collectively as primary immune regulatory disorders (PIRDs). Programmed cell death, or apoptosis, is crucial for maintaining the balance of lymphocytes. Genetic-level identification of several human inherited diseases with impaired apoptosis has been achieved, such as autoimmune lymphoproliferative syndrome (ALPS), caspase-8 deficiency state (CEDS), X-linked lymphoproliferative syndrome (XLP), and Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway disorders. The consequences of this disease are manifested by abnormal lymphocyte accumulation, resulting in clinical features such as lymphadenopathy, hepatomegaly, splenomegaly, and an increased risk of lymphoma. Additionally, these disorders are often associated with autoimmune disease, particularly involving blood cells. Understanding the molecular pathogenesis of these conditions has provided critical insights into the signaling pathways that regulate apoptosis and lymphocyte activation, shedding light on mechanisms of immune dysregulation. This review focuses on the intersection between apoptosis, autoimmunity, and lymphoproliferation, discussing how dysregulation contributes to the development of these immune disorders. These conditions are characterized by excessive lymphocyte accumulation, autoimmunity, and/or immunodeficiency. Understanding their molecular pathogenesis has offered new insights into the signaling mechanisms that regulate apoptosis and lymphocyte activation.
Identifying novel biomarkers is a reliable approach to predict and diagnose human diseases as well as manage individual responses to therapeutic drugs. Heat shock proteins (HSPs) are molecular chaperones that play a major role in maintaining protein stability and folding. Many studies suggested their association with multiple types of diseases. HSPs from different categories play different roles; therefore, it is important to identify HSPs and their function to understand their biological functions clearly. This comprehensive review was performed to evaluate the role of HSPs as predictive biomarkers in cardiovascular diseases. The original publications related to HSPs from 2010 to 2024 were identified by using the keywords “heat-shock proteins”, “HSP in cardiovascular disorders” and “HSP in atherosclerosis”. The regulatory pathways involved in HSPs’ functioning are the important points of discussion in this review. HSPs play a critical role in key cellular processes, including apoptosis regulation, protein folding, immune responses, genomic stability, and DNA repair. Aberrant expression of HSPs causes dysregulation of these pathways resulting in the development and progression of diseases. A comprehensive understanding of HSPs in cardiovascular diseases and their associated regulatory pathways can have significant implications for disease intervention, diagnosis, and prognosis. In this review paper, we have highlighted the importance of HSPs as versatile biomarkers and their importance as targets for the therapeutic management of cardiovascular diseases.
Plant hydrogen sulfide (H2S) metabolism has garnered noteworthy attention due to its role in regulating many plant processes. The primary mechanism by which H2S exerts its signaling functions is through its reversible interaction with thiol groups on cysteine residues in proteins and peptides. This thiol-based oxidative post-translational modification (oxiPTM) is known as persulfidation. Transcription factors (TFs) are key proteins that control gene expression by interacting with distinguishing DNA sequences and other regulatory proteins. Their function is essential to almost all aspects of cellular biology, including development, differentiation, and responses to environmental biotic and abiotic cues. The persulfidation of TFs has emerged as an additional regulatory mechanism, linking H2S signaling with gene regulation. Although the available information on the crosstalk between the regulatory mechanisms of H2S metabolism and TF activity remains limited, existing data suggest that this connection influences not only H2S metabolism itself but also other metabolic pathways involved in various physiological and stress responses. This review provides an updated overview of an emerging research area, focusing on the mutual regulation between specific TFs and H2S metabolism, particularly in response to adverse environmental conditions.
Autoimmune hepatitis (AIH) is a chronic immune-mediated inflammatory liver disease characterized by recurring immune-triggered hepatic injury. While scientists have yet to fully elucidate the precise triggers of AIH, contemporary research indicates that both gut microbiota and their metabolic products significantly influence AIH progression. These factors contribute to multiple mechanisms, including compromised intestinal barrier function, altered microbial and metabolite trafficking, and disrupted immune balance, leading to inflammatory responses. This review begins by exploring the intestinal microbial populations and their byproducts linked to AIH. It highlights how disrupted gut flora compromises intestinal immune defenses, enables bacterial migration from the gut to hepatic tissue, and induces liver inflammatory responses. Research validates that metabolic products from microbes, such as short-chain fatty acids (SCFAs), bile acids (BAs), and specific amino acids (glutamine, cysteine, tryptophan, and branched-chain variants, among others), interact with immune cell populations. These interactions, coupled with immune cell modifications, contribute to AIH progression. Our review identifies promising treatment strategies, including the use of probiotic supplementation, engineered prebiotic compounds, microbiota transfer procedures, and specific medications targeting gut microorganisms and their byproducts. These approaches could potentially reduce immune-triggered hepatic damage, offering potential new avenues for AIH management.
Lysophosphatidic acid (LPA), a bioactive lipid molecule, has been identified as a critical regulator of several cellular processes in the central nervous system, with significant impacts on neuronal function, synaptic plasticity, and neuroinflammatory responses. While Alzheimer’s disease, Multiple Sclerosis, and Parkinson’s disease have garnered considerable attention due to their incidence and socioeconomic significance, many additional neurological illnesses remain unclear in terms of underlying pathophysiology and prospective treatment targets. This review synthesizes evidence linking LPA’s function in neurological diseases such as traumatic brain injury, spinal cord injury, cerebellar ataxia, cerebral ischemia, seizures, Huntington’s disease, amyotrophic lateral sclerosis, Hutchinson-Gilford progeria syndrome, autism, migraine, and human immunodeficiency virus (HIV)-associated complications Despite recent advances, the specific mechanisms underlying LPA’s actions in various neurological disorders remain unknown, and further research is needed to understand the distinct roles of LPA across multiple disease conditions, as well as to investigate the therapeutic potential of targeting LPA receptors in these pathologies. The purpose of this review is to highlight the multiple functions of LPA in the aforementioned neurological diseases, which frequently share the same poor prognosis due to a scarcity of truly effective therapies, while also evaluating the role of LPA, its receptors, and signaling as promising actors for the development of alternative therapeutic strategies to those proposed today.
Significant advancements have been achieved over recent decades in deciphering the molecular mechanisms driving malignant tumor development. Despite this progress, the precise roles of individual genes, their interactions, and the associated signaling pathways involved in tumor proliferation remain insufficiently characterized. Among these pathways, the mitogen-activated protein kinases (MAPKs) extracellular signal-regulated kinase (ERK)1/2 and p38, which regulate essential cellular functions such as growth, differentiation, and apoptosis, have garnered considerable research attention. Building on recent insights into MAPK signaling, we identified components closely linked to ERK1/2 and p38 activity and examined changes in their behavior during tumorigenesis. Furthermore, we developed quantifiable metrics to assess ERK1/2 and p38 activity, including the ERK/p38 ratio, a key indicator of tumor cell proliferative or quiescent states, along with activation levels of these signaling pathways. Our findings underscore the potential of ERK and p38-related gene expression and pathway dynamics as biomarkers for predicting clinical outcomes and informing tailored therapeutic approaches.
The Piezo2 transmembrane proteins were identified by Ardem Patapoutian and his team. They also found that Piezo2 is the principal mechanosensory ion channel responsible for proprioception. Even before the Nobel Prize was awarded to him, it was proposed that these Piezo2 channels could sustain acquired microdamage at the proprioceptive somatosensory terminals under allostatic stress. Moreover, the principality of Piezo2 is suggested to extend beyond its physiological function, highlighting its relevance in the context of microdamage as well. Hence, acquired Piezo2 channelopathy is proposed to constitute one principal gateway to pathophysiology underpinned by proton affinity, energy metabolism and a proprioceptive pathway switch. The differentiating incomparable hallmark of Piezo2 is theorized to be a low-frequency semiconductor Schottky barrier diode-like feature that provides proton handling for quantum tunnelling and ultrafast long-range signalling to the hippocampus. Accordingly, even the proposed acquired Piezo2 channelopathy is also enigmatic by causing the impairment of this Piezo2-initiated ultrafast proton-based long-range signalling and proper synchronization to the hippocampus. The revealing of this protonic word and the ultrafast long-range signalling within the nervous system and its microdamage brings an entirely new perspective in medicine with the interpretation of the quad-phasic non-contact injury model. This is why this Piezo2 microdamage has been coined as the primary damage or the root cause of ageing. Paired-associative electromagnetic stimulation appears to be a promising treatment method and heart rate variability detection could be used for diagnosing autonomic nervous system disbalance as one symptom of this proposed Piezo2 channelopathy.
Endometriosis (EM) is a prevalent gynecological disorder in women. Although the underlying mechanisms have yet to be fully elucidated, EM may be related to oxidative stress. The current research aimed to identify possible pathways that control oxidative stress in EM, thereby providing a theoretical foundation for its clinical diagnosis and treatment.
High-throughput RNA sequencing (RNA-seq) data were integrated with GeneCards online data to screen for oxidative stress-related genes and potential targets in EM. The reverse transcription-quantitative polymerase chain reaction (RT-qPCR), Western blotting, and immunohistochemistry assays confirmed the expression of candidate genes. The in vivo and in vitro effects of CCAAT enhancer binding protein delta (CEBPD, C/EBP-delta) and DNA damage-inducible transcript 4 (DDIT4) on oxidative stress, cell proliferation, and angiogenesis in endometriotic cells were validated using loss- or gain-of-function approaches.
CEBPD was highly expressed in ectopic and eutopic endometrial tissue from patients with endometriosis. Loss- or gain-of-function experiments showed that CEBPD promoted oxidative stress, cell proliferation, and angiogenesis in vitro and in vivo. Integration of RNA-seq and online data revealed that CEBPD regulates DDIT4 expression, subsequently increasing oxidative stress, cell proliferation, and angiogenesis in endometriotic cells. Finally, CEBPD and DDIT4 were found to regulate the expression of extracellular signal-regulated kinase 1/2 (ERK1/2) proteins associated with the mitogen-activated protein kinase (MAPK) signaling pathway.
These results suggest that CEBPD may promote oxidative stress, cell proliferation, and angiogenesis in EM by activating MAPK via DDIT4. Hence, CEBPD may be a potential target for diagnosing and treating EM.
Radiotherapy is crucial for managing esophageal squamous cell carcinoma (ESCC). This research explored the potential and mechanism of enhancing ESCC radiosensitivity through targeting phosphoglycerate kinase 1 (PGK1).
After ESCC cells were exposed to X-rays and C-ions, hub genes were identified through proteomic analysis and bioinformatics. To elucidate PGK1’s function, small interfering RNAs and plasmids were used to silence and overexpress PGK1 in two human ESCC cell lines. Plate colony formation, cell counting kit 8, and 5-ethynyl-2′-deoxyuridine assays were conducted to detect cell proliferation after irradiation with different linear energy transfer rays (X-rays and carbon ions). Flow cytometry was used to assess radiation-induced perturbations in the cell cycle, apoptosis, reactive oxygen species (ROS), and mitochondrial membrane potential. Western blotting was performed to detect the protein expressions of protein kinase B (Akt), phosphorylated protein Kinase B (pAkt), mammalian target of rapamycin (mTOR), and phosphorylated mammalian target of rapamycin (pmTOR).
Proteomics and bioinformatics analyses revealed that PGK1 plays a key role in modulating ESCC radiosensitivity. Knockdown of PGK1 resulted in the suppression of cancer cell proliferation and viability, promoted apoptotic processes, and demonstrated a synergistic anti-tumor effect in conjunction with radiation. Conversely, overexpression of PGK1 promoted cancer cell growth and increased radiation resistance. This may be attributed to the accumulation of ROS and the inhibition of Akt/mTOR pathway following PGK1 inhibition.
Targeting PGK1 may be an effective strategy to increase ESCC radiation sensitivity, offering a promising strategy for improving treatment outcomes.
It is well known that the microenvironment in which an immune response develops, generally pro-inflammatory or immunosuppressive, along with other overproduced biomolecules recognized by pattern recognition receptors, may promote the stimulation and differentiation of monocytes into macrophages with effector functions. Low-density lipoprotein (LDL) plays a fundamental role in cholesterol transport. By contrast, its oxidized form (ox-LDL), which is overexpressed in conditions of obesity and chronic low-grade inflammation, has been associated with cardiovascular diseases. Depending on the microenvironmental context, prostaglandin E2 (PGE2) participates in various scenarios such as inflammation, anti-inflammation, and homeostasis. Therefore, obesity-derived biomolecules such as LDL, ox-LDL, and PGE2 could induce the differentiation of immune cells into effector populations with either pro-inflammatory or immunosuppressive profiles.
In the present work, we studied the effects of LDL, ox-LDL, and PGE2 on the differentiation of the human THP1 monocytic cell line into macrophages under two different protocols, analyzing several activation markers associated with either pro-inflammatory M1 or anti-inflammatory M2 profiles by flow cytometry and quantitative PCR (qPCR).
Our data suggest that native LDL induces the differentiation of human THP1 monocytes into M1 macrophages even more efficiently than classic phorbol 12-myristate 13-acetate (PMA) stimulation, whereas ox-LDL and PGE2 induce the expression of activation markers similarly to interferon gamma or interleukin 4 during PMA preactivation of macrophages.
The results of this study add evidence to the role of obesity-derived biomolecules as non-canonical differentiation stimuli in macrophages, which could be relevant in contexts where these biomolecules are chronically overproduced, such as obesity, low-grade inflammation, type 2 diabetes, and cancer.
Endometriosis (EMs) is a chronic gynecological disorder associated with ectopic endometrial tissue, inflammation, oxidative stress, and mitochondrial dysfunction. A promising strategy for treating EMs is to target ferroptosis, a programmed cell death mechanism regulated by reactive oxygen species (ROS) and glutamine metabolism. Solute carrier family 1 member 5 (SLC1A5), a glutamine transporter, and c-Myc play key roles in ferroptosis, forming a “ROS/c-Myc/SLC1A5” feedback loop. The aim of this study was to investigate the regulatory role of SLC1A5 in ferroptosis. In addition, we evaluated the ferroptosis inducer Erastin as a potential therapeutic agent for EMs.
The human endometrial stromal cells (ESCs) line hEM15A was used in this study, together with a rat model of EMs. hEM15A cells and rats were treated with Erastin, with or without SLC1A5 modulation or ROS scavenging with N-acetylcysteine (NAC). Cell viability, ROS levels, glutamine metabolism, mitochondrial function, and ferroptosis markers (glutathione peroxidase 4 (GPX4)) were subsequently analyzed by Cell Counting Kit-8 (CCK-8) assay, reverse transcription quantitative polymerase chain reaction (RT-qPCR), Western blot, and fluorescent probes. Pathological changes, lesion volumes, and pelvic adhesions in the rat EM model were assessed using hematoxylin and eosin (HE) staining, ultrasound imaging, and Haber scoring.
Erastin treatment of ESCs induced ferroptosis by upregulating SLC1A5 and c-Myc expression, increasing ROS levels, and altering glutamine metabolism. Overexpression of SLC1A5 enhanced sensitivity to ferroptosis, whereas SLC1A5 knockdown and NAC treatment reversed these effects. Mechanistically, c-Myc bound to the SLC1A5 promoter, forming positive feedback with ROS. In the rat model of EMs, Erastin treatment reduced ectopic lesion volume, pelvic adhesions, and inflammatory markers (TNF-α, IL-6, IL-1β). These therapeutic effects were mitigated by NAC, highlighting the importance of the ROS/c-Myc/SLC1A5 pathway.
This study confirmed the involvement of the ROS/c-Myc/SLC1A5 pathway in regulating EMs sensitivity to ferroptosis and demonstrated the potential of Erastin as a therapeutic agent. Targeting this pathway offers a promising approach for the treatment of EMs.
Obesity induces chronic inflammation and hormonal imbalances that contribute to tumor growth. This study explores the less understood dynamics of tumor-related macrophages under a high-fat diet and its consequent impact on tumor growth, with a focus on elucidating the role of high-fat diets on macrophage behavior in liver cancer.
We established a mouse obesity model using a high-fat diet, combined with a liver cancer implantation approach. Tumor-infiltrating macrophages were isolated for analysis. We investigated the specific effects of a high-fat diet on macrophages through transcriptomic and metabolomic studies and further explored the influence of N6-methyladenosine (m6A) RNA modification on macrophage differentiation using in vitro and in vivo models.
Our findings reveal that a high-fat diet significantly accelerates in-situ liver cancer growth and fosters type II differentiation of tumor-associated macrophages. RNA sequencing indicated upregulation of Cpt1a and Mettl3 genes, which are crucial for m6A modification in macrophages. Using human and mouse macrophage cell lines with either elevated Mettl3 expression or Cpt1a gene knockout, we demonstrated that methyltransferase-like 3 (METTL3) enhances fatty acid metabolism in macrophages, a process reversible by Cpt1a gene knockout. These effects were corroborated in vivo. Further, macrophages infused with high Mettl3 expression, when combined with an in-situ implantation model and adoptive cell therapy, markedly promoted liver cancer growth and increased type II macrophage differentiation (p < 0.001). Knockout of the Cpt1a gene counteracted the METTL3 effect compared to the control group (p > 0.05). METTL3 and m6A RNA Immunoprecipitation (RIP) assays confirmed that METTL3 stabilizes Cpt1a mRNA. Additionally, multispectral staining of clinical specimens revealed a positive correlation between METTL3 protein levels in liver cancer tumor-associated macrophages and M2 macrophage prevalence, inversely correlating with M1 macrophages (p < 0.01). High Mettl3 expression in macrophages was associated with poor prognosis in liver cancer patients, correlating significantly with tumor size and tumor node metastasis (TNM) classification stage.
Our research identifies that a high-fat diet elevates METTL3-driven m6A modification of carnitine palmitoyltransferase 1A (CPT1A) in tumor macrophages, fostering type II macrophage differentiation, and exacerbating liver cancer growth and immune evasion.
Anxiety and depression-like behaviors are common in patients with type 2 diabetes mellitus (T2DM). This study explored the potential of swimming training (ST) to alleviate these symptoms by restoring mitochondrial function. While aerobic exercise is known to influence mitochondrial dysfunction and behavioral abnormalities, the mechanism by which ST achieves this remains unclear.
To investigate how ST improves T2DM and associated anxiety-like behaviors by regulating mitochondrial structure and function.
T2DM was induced in zebrafish with a high-sugar diet, followed by 20 days of ST. Behavioral analysis assessed anxiety-like behaviors, while ELISA and microscopic imaging techniques were used to evaluate changes in mitochondrial structure and function in liver tissue.
ST significantly alleviated anxiety-like behavior and mitigated mitochondrial damage. Furthermore, ST counteracted mitochondrial dysfunction induced by oxidative stress through regulation of reactive oxygen species levels (p < 0.01), stabilization of mitochondrial membrane potential (p < 0.0001), and increasing the production of adenosine triphosphate (p < 0.01). ST also improved T2DM markers, including blood glucose regulation (p < 0.001), insulin level (p < 0.05), and lipid metabolism (p < 0.01 for low-density lipoprotein cholesterol (LDL-C), p < 0.01 for high-density lipoprotein cholesterol (HDL-C), p < 0.01 for total cholesterol (T-CHO)).
This research provides insights into the intricate interplay between mitochondrial dysfunction in T2DM and behavioral outcomes while highlighting the potential of ST as a holistic therapeutic strategy for T2DM patients.
Fluorescent probes have become a powerful tool for monitoring biothiol concentrations, aiding in disease diagnosis and treatment while also facilitating the exploration of fundamental biological processes. However, the probes are limited by the short fluorescence emission wavelength and small Stokes shift, which makes them susceptible to background fluorescence interference and significant self-absorption. To overcome these limitations and achieve high-fidelity biothiols detection in complex biological systems, this study focuses on developing a near-infrared fluorescent probe with an extended Stokes shift.
(E)-4-(5-(2-(4-(dicyanomethylene)-4H-chromen-2-yl)vinyl)thiophen-2-yl)phenyl 2,4-dinitrobenzenesulfonate (DCMOS-N), a near-infrared (NIR) fluorescent probe featuring a large Stokes shift, was designed and synthesized for biothiols detection. The optical properties of DCMOS-N were evaluated using ultraviolet-visible (UV-Vis) and fluorescence spectroscopy. Additionally, its imaging capabilities for detecting biothiols in living cells were assessed through confocal fluorescence microscopy.
Fluorescence spectral analysis confirmed that the DCMOS-N probe exhibits high selectivity and strong anti-interference properties in biothiol detection. Moreover, its fluorescence intensity increases upon the addition of biothiols. Notably, a strong linear correlation was observed across the concentration range of 0 to 100 μmol/L (R2 = 0.9944 for glutathione (GSH), 0.9942 for cysteine (Cys), and 0.9946 for homocysteine (Hcy)), enabling the quantitative analysis of biothiol concentrations in biological systems. The detection limits for GSH, Cys, and Hcy were determined as 0.142 μmol/L, 0.129 μmol/L, and 0.143 μmol/L, respectively. Importantly, the practical application of DCMOS-N in living cells was validated, with confocal fluorescence imaging demonstrating its capability to detect both endogenous and exogenous biothiols in HeLa cells.
An NIR fluorescent probe, DCMOS-N, was developed and effectively utilized to monitor biothiols in living HeLa cells. The successful design of DCMOS-N presents significant potential and serves as an innovative platform for developing fluorescence probes targeted at biothiols.
Mitochondria are essential for cellular energy production and cell survival. Mitochondrial dysfunction has been implicated in various neurological disorders, prompting the development of novel therapeutic approaches targeting these organelles. Among these, mitochondrial transplantation (MT), which replaces dysfunctional mitochondria with healthy counterparts from donor tissues, has emerged as a promising strategy. While skeletal muscle is a rich source of mitochondria, the optimal muscle tissue for MT remains unidentified, and the potential functional differences among mitochondria from various muscle types are not fully understood. This study investigates the quantity, size, respiratory function, energy production, and anti-inflammatory effects of mitochondria isolated from red skeletal muscle (RSM), mixed skeletal muscle (MSM), and white skeletal muscle (WSM).
Mitochondria were extracted from the soleus muscle (RSM), pectoralis major and rectus abdominis (MSM), and biceps brachii and gastrocnemius (WSM) of healthy 8-week-old male Sprague Dawley rats. Nanoparticle tracking analysis was employed to determine mitochondrial quantity and size. The activities of mitochondrial complexes I, II, and IV and adenosine triphosphate (ATP) content were assessed. The protective effects of mitochondria (100 μg/mL) from each muscle type against lipopolysaccharide (LPS, 5 μg/mL)-induced cell death and mitochondrial membrane potential disruption were evaluated in PC-12 neuronal cells.
RSM-derived mitochondria exhibited a smaller average size and significantly higher mitochondrial content compared to those from MSM (mean size: p = 0.0056, vs. pectoralis major; p = 0.0056, vs. rectus abdominis; count of mitochondria: p < 0.0001, vs. pectoralis major; p < 0.0001, vs. rectus abdominis) and WSM (mean size: p = 0.0006, vs. biceps brachii; p < 0.0001, vs. gastrocnemius; count of mitochondria: p < 0.0001, vs. biceps brachii; p < 0.0001, vs. gastrocnemius). Additionally, RSM mitochondria demonstrated the highest activity of mitochondrial complex I among the three muscle types (p = 0.0001, vs. pectoralis major; p = 0.0095, vs. rectus abdominis; p < 0.0001, vs. biceps brachii; p < 0.0001, vs. gastrocnemius). WSM-derived mitochondria showed relatively lower complex II activity (p = 0.0006, biceps brachii vs. soleus; p = 0.0218, biceps brachii vs. rectus abdominis), while complex IV activity and ATP content were comparable across all groups. Supplementation with mitochondria isolated from RSM and WSM, but not MSM, effectively mitigated LPS-induced cell death (mitochondria isolated from soleus: p = 0.0031; biceps brachii: p = 0.0046; gastrocnemius: p = 0.0169) and preserved mitochondrial membrane potential (mitochondria isolated from soleus: p = 0.0204; biceps brachii: p = 0.0086; gastrocnemius: p = 0.0001) in PC-12 cells.
RSM emerges as the optimal source for mitochondrial extraction, demonstrating superior respiratory activity and significant protective effects against LPS-induced cell death and mitochondrial dysfunction. These findings provide critical insights into optimizing MT outcomes through the strategic selection of mitochondrial sources.
Frequent drug resistance seriously limits the therapeutic efficacy of sorafenib in advanced hepatocellular carcinoma (HCC). Strategies to increase the response to sorafenib are limited, and the underlying mechanism to facilitate such an increase is not entirely understood. Homeobox (HOX) transcript antisense intergenic RNA (HOTAIR) expression is high in HCC, promoting the occurrence and progression of HCC. In this study, we explored the mechanism through which HOTAIR knockdown affects the response of HCC cells to the chemotherapeutic sorafenib.
Cell viability and apoptosis were assessed using MTT assay, flow cytometry, and nuclear staining. Mitochondrial function and isolation were determined using flow cytometry and a mitochondrial isolation kit. Glycolysis was measured by glucose and lactic acid assay kits. The underlying mechanisms were explored through western blotting, quantitative reverse-transcription polymerase chain reaction (qRT-PCR), and chromatin immunoprecipitation (ChIP).
HOTAIR knockdown increased sorafenib-induced apoptosis in the HCC cells. HOTAIR and hexokinase 2 (HK2) expression levels were upregulated in human HCC tissues, demonstrating a significant correlation. The knockdown of HOTAIR or HK2 aggravated mitochondrial dysfunction and inhibited glycolysis. Further, HOTAIR knockdown promoted sorafenib-mediated HK2 mRNA downregulation, resulting in decreased HK2 protein levels in cells and mitochondria. This ultimately facilitated the mitochondrial apoptotic pathway. Moreover, it was demonstrated that HOTAIR regulated HK2 via polycomb repressive complex 2 (PRC2)-mediated epigenetic modification of miR-145-5p in HCC.
HOTAIR knockdown increased the sensitivity of HCC cells to sorafenib by disrupting the miR-145-5p/HK2 axis-mediated mitochondrial function and glycolysis, suggesting new strategies for HCC treatment.
The excessive formation of neutrophil extracellular traps (NETs) is involved in delayed wound healing under diabetic conditions. However, potential therapeutic agents remain underexplored. Our present study aimed to explore the effects of Ro 106-9920, a specific nuclear factor kappa B (NF-κB) inhibitor, on diabetic wound healing and to elucidate the underlying mechanisms.
A diabetic mouse model was established, and full-thickness wounds were created. Ro 106-9920 was administered, and wound healing was monitored. Protein levels of NET markers and nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) inflammasome components were assessed via western blotting and histological analysis. Functional assays were conducted to evaluate the effect of NETs on fibroblasts, endothelial cells, and keratinocytes using conditioned media (CM) from a phorbol 12-myristate 13-acetate (PMA)-treated neutrophil-macrophage coculture model (NET-CM). CM collected from the coculture model after Ro 106-9920 treatment ((NET+Ro 106-9920)-CM) was used to determine its therapeutic potential.
NET formation and NLRP3 inflammasome activation were significantly elevated in wound tissues of diabetic mice from day 7 (p < 0.001). Similar results were observed in PMA-treated neutrophils and macrophages (p < 0.001). The viability and migration of endothelial cells, fibroblasts, and keratinocytes, as well as the angiogenic potential of endothelial cells, were significantly impaired by NET-CM treatment (all p < 0.001), whereas Ro 106-9920 effectively attenuated these alterations (NET-CM vs. (NET+Ro 106-9920)-CM, cell viability, p < 0.001; cell migration, p < 0.01; tube formation, p < 0.001). In vivo, Ro 106-9920 treatment inhibited NET formation, as evidenced by the decreased citrullinated histone H3 (CitH3) and peptidyl arginine deiminase 4 (PAD4) expression (p < 0.05). This was followed by a decrease in NLRP3 inflammasome activation (p < 0.05), an increase in angiogenesis in wound tissues (p < 0.001), and improved wound healing (p < 0.001) compared with those in Ro 106-9920-untreated mice.
Ro 106-9920 enhances diabetic wound healing by suppressing NET formation and inhibiting NLRP3 inflammasome activation, providing a novel therapeutic choice for improving chronic wound healing in patients with diabetes.
The chick embryo chorioallantoic membrane (CAM) model is gaining increasing attention from cancer researchers worldwide. Its affordability, short experimental duration, robustness, and ease of tumor xenograft visualization make it a valuable tool in cancer research. This review explores recent advancements and potential applications of the avian CAM model, including the following: (1) studying tumor growth and metastasis, (2) investigating mechanisms of tumor chemoresistance, (3) optimizing drug delivery methods, (4) improving bioimaging techniques, (5) evaluating immuno-oncology drug efficacy, (6) examining tumor-extracellular matrix interactions, (7) analyzing tumor angiogenesis, and (8) exploring the roles of microRNAs in cancer. Additionally, we compare the in ovo CAM model with other in vivo animal models and in vitro cell culture systems. Positioned between in vitro and in vivo models in terms of cost-effectiveness and accuracy in cancer recapitulation, the CAM model enhances both preclinical and translational research. Its expanding use in cancer studies and therapy development is expected to continue growing.
Multiple sclerosis (MS) is characterized as a chronic inflammatory autoimmune disorder affecting the central nervous system (CNS). Prior research has explored the involvement of pyroptosis and high mobility group box 1 (HMGB1) in the pathophysiology of MS. Nevertheless, the underlying pathogenic mechanisms and their interactions have yet to be fully elucidated.
Myelin oligodendrocyte glycoprotein (MOG)35-55-treated mice and BV-2 microglial cells were utilized as a model for MS. Subsequently, these subjects were transfected with lentiviral vectors that express short hairpin RNA targeting HMGB1. HT-22 cells and Ma-c cells were exposed to conditioned medium (CM) derived from BV-2 cells following treatment. The levels of HMGB1, tumor necrosis factor (TNF)-α, and interleukin-1β (IL-1β) were quantified using enzyme-linked immunosorbent assay (ELISA). Additionally, western blot (WB) analysis was performed to further elucidate the mechanisms involved.
Mice treated with MOG35-55 (experimental autoimmune encephalomyelitis, EAE) exhibited reduced body weights and significant nerve function impairment (p < 0.001), accompanied by increased activation of microglia within the CNS (p < 0.05). Additionally, the secretion of HMGB1 was found to be upregulated in the MS cell model (p < 0.05), and CM from these cells induced the release of pro-inflammatory cytokines in HT-22 and Ma-c cell lines (p < 0.001). Notably, the modulation of HMGB1 and NOD-like receptor family pyrin domain containing 3 (NLRP3) expression was shown to mitigate the release of pro-inflammatory cytokines (p < 0.01), TUNEL-positive cells (p < 0.01) in both HT-22 cells and Ma-c cells, which were induced by CM from BV-2 cells treated with MOG35-55. Furthermore, WB analysis indicated that the suppression of HMGB1 expression can inhibit the activation of the toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB) signaling pathway, as well as pyroptosis in EAE mice and HT-22/ Ma-c cells exposed to CM from BV-2 cells (p < 0.05).
HMGB1 has the potential to act as a promoter of MS through the activation of TLR4/NF-κB signaling pathway and the induction of pyroptosis in microglial and other cells. Consequently, the modulation of HMGB1 may represent a novel therapeutic strategy for the management of MS.
Polycystic ovary syndrome (PCOS) has increasingly emerged as a significant cause of impaired reproductive outcomes, primarily characterized by a combination of ovulatory dysfunction and decreased oocyte quality. However, the molecular mechanisms underlying the decreased oocyte quality caused by PCOS and preventative strategies still require further investigation.
All procedures were approved by the Animal Ethics Committee of Chongqing Medical University. We established a mice model of PCOS using dehydroepiandrosterone (DHEA) treatment. The estrous cycle was recorded, and plasma sex hormone and trimethylamine N-oxide (TMAO) levels were measured. Ovarian indices and follicular formation were compared. Time-lapse imaging was used to observe in vitro maturation and blastocyst formation. Reactive oxygen species (ROS), MitoSOX level, and mitochondrial membrane potential were measured to analyze the mitochondrial function of oocytes. Confocal laser scanning microscopy was used to detect spindle function and chromosomes.
Our study found that DHEA-induced PCOS mice exhibited significantly lower plasma TMAO levels compared to normal mice. Consequently, we supplemented TMAO in PCOS mice and found that the abnormal estrous cycle and reduced ovarian function induced by PCOS could be restored. Additionally, TMAO rescued PCOS-induced defects in oocyte maturation, spindle and chromosome morphology, and embryonic developmental potential. Mechanically, we found that TMAO effectively reduced ROS levels by improving mitochondrial function in PCOS oocytes.
Our findings indicate that the reduction in TMAO levels induced by PCOS may be a key factor influencing reproductive outcomes. TMAO supplementation in vivo can effectively enhance mitochondrial function and oocyte quality in PCOS, holding significant clinical importance for improving assisted reproductive outcomes in patients with PCOS.
This study aimed to evaluate the effects of selected phosphodiesterase-4 inhibitors (PDE-4 inhibitors)—roflumilast, ibudilast, and crisaborole—on the activity of blood coagulation factor XII (FXII). In the intrinsic coagulation pathway, FXII is known to initiate the kallikrein–kinin system (KKS), causing an increase in the system expression, which ultimately leads to inflammation and coagulation states. Additionally, the activation of KKS downstream effectors leads to inflammation. Inflammation signaling was found to be initiated when the bradykinin (BK) protein binds to its B2 receptor because of the FXII-dependent pathway activation. BK abnormalities can cause a critical condition, hereditary angioedema (HAE), which is characterized by recurring serious swelling. While it is considered unnecessary for hemostasis, FXII is an important enzyme for pathogenic thrombosis. Because of this special characteristic, FXII is a desirable therapeutic target. Our hypothesis is to identify the inhibitory effects of roflumilast, ibudilast, and crisaborole on the activated FXII and to reveal their beneficial impacts in the reduction of the pathogenesis of FXII-related conditions, HAE, and thrombosis. In a current study, we presented the inhibitory effect of tested drugs on the main target activated factor XII (FXIIa) as well as two other plasma protease enzymes included in the target pathway, plasma kallikrein and FXIa.
To achieve our aim, in vitro chromogenic enzymatic assays were utilized to assess the inhibitory effects of these drugs by monitoring the amount of para-nitroaniline (pNA) chromophore released from the substrate of FXIIa, FXIa, or plasma kallikrein.
Our study findings exhibited that among assessed PDE-4 inhibitor drugs, roflumilast at micromolar concentrations significantly inhibited FXIIa in a dose-dependent manner. The FXIIa was clearly suppressed by roflumilast, but not the other related KKS members, plasma kallikrein, or the activated factor XI. On the other hand, ibudilast and crisaborole showed no inhibitory effects on the activities of all enzymes.
Overall, roflumilast could be used as a lead compound for developing a novel multifunctional therapeutic drug used for the prevention of HAE or thrombotic disorders.
Interferons (IFNs) are ototoxic drugs leading to vestibular and auditory disorders. This study investigated the effect of pro-inflammatory cytokine IFN-α2b on the afferent glutamatergic synaptic transmission of the vestibular end organ, focusing on ionotropic glutamate receptors (iGluRs).
In order to characterize the role of IFN-α2b in the glutamatergic synaptic transmission in vestibular epithelium, we investigated its influence on responses evoked by D,L-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), N-methyl-D-aspartate (NMDA) and kainic acid (kainate). This was carried out using external perfusion of the vestibular apparatus and multiunit recording of afferent firing activity of semicircular canal ampullary nerve fibers. The change in the ratio of the maximum frequency of pulse activity to the preceding background was chosen as a criterion for evaluating the evoked responses of glutamate receptor (GluR) agonists.
Acute perfusion of the vestibular apparatus with IFN-α2b and AMPA did not alter the AMPA-evoked response. However, a significant increase in the response was observed 15 min after cessation of drug application and washing with normal solution (paired-samples t-test p = 0.018; n = 20). IFN-α2b significantly increased the kainate-evoked response during cytokine application (Wilcoxon signed-rank test p = 0.016; n = 11), and further potentiates the response 15 min after rinsing with normal solution, compared to the test value (Wilcoxon signed-rank test p = 0.05; n = 11). IFN had no effect on NMDA-induced responses. AMPA receptors (AMPARs) potentiated by IFN-α2b increase NMDA-evoked responses (Repeated measures analysis of variance [ANOVA RM], p = 0.028; n = 10).
IFN-α2b stimulates AMPARs and kainate receptors (KARs) through various mechanisms but has no direct effect on NMDA receptors (NMDARs). Interferon-activated AMPARs can stimulate NMDARs activity, thereby altering synaptic plasticity of the glutamatergic afferent synapse in vestibular epithelium.
Lung cancer is the primary cause of cancer-related mortality, but the molecular mechanisms behind this malignancy remain unclear.
The Cancer Genome Atlas (TCGA) online database and tissue chips were used to analyze the expression levels of tumor necrosis factor receptor-associated factor 2 (TRAF2)- and non-catalytic region of tyrosine kinase adaptor protein (NCK)- interacting kinase (TNIK) protein in lung cancer. A549 and PC-9 lung adenocarcinoma (LUAD) cells with stable TNIK knockdown were generated by lentivirus infection. The tumor phenotypes were subsequently examined both in vitro and in vivo. The TCGA online database and RNA-sequencing of TNIK-knockdown cells were used to study the molecular mechanism underlying the TNIK-mediated phenotype of LUAD cells. The effects of TNIK knockdown on focal adhesion dynamics and mitosis were examined by indirect immunofluorescence and Western blot, on the sensitivity to chemotherapy drugs by cell counting kit-8 (CCK-8) assay, on apoptosis by flow cytometry, and on cell proliferation by 5-ethynyl-2′-deoxyuridine (EDU).
TNIK was highly expressed in LUAD (p < 0.0001), predominantly in the cytosol. Phenotype assays revealed that TNIK knockdown in LUAD cells led to a significant increase in cell spreading (p < 0.0001), but also inhibition of cell growth and movement (p < 0.01). Mechanistically, TNIK was found to regulate F-actin and microtubule organization, as well as the Ras homolog gene family (RHO)/RHO-associated kinase 2 (ROCK2)/LIM motif-containing protein kinase 1 (LIMK1) signaling pathway, thereby playing a crucial role in the control of focal adhesion turnover and mitosis. Additionally, the silencing of TNIK enhanced the sensitivity of LUAD cells to chemotherapeutic drugs.
Our findings suggest that TNIK regulates focal adhesion turnover and mitosis to promote tumor malignancy via the RHO/ROCK2/LIMK1 pathway. The combination of TNIK targeting with chemotherapeutic drugs could be an effective strategy to overcome resistance in LUAD.