Acta Pharmacologica Sinica最新文献

Selective serotonin reuptake inhibitors (SSRIs) are commonly used to treat depression, but their chronic use is associated with side effects and residual symptoms of depression. Both effects induced by SSRIs are mediated by serotonin receptor-dependent signaling pathways, yet the molecular mechanisms underlying these effects remain unclear. Here, we investigated the impact of chronic and acute activation of the 5-HT7 receptor (5-HT7R) using the selective agonist AGH-194 in male mice. Behavioral assessment revealed that chronic AGH-194 administration induced depressive-like effects in the novelty suppressed feeding test (NSFT), female urine sniffing test (FUST), and novel object location test (NOLT). After acute injection, depressive-like effects were observed only in NSFT. At the molecular level, AGH-194 administration activated matrix metalloproteinase 9 (MMP-9) through a 5-HT7R-Gαs signaling-dependent mechanism. Acute treatment induced transient activation, while chronic treatment led to prolonged enzymatic activity, accompanied by a reduction in the expression of the GluA1 subunit of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) in the hippocampus. At the cellular level, acute but not chronic AGH-194 treatment induced a shift toward more juvenile dendritic spine morphology in the CA1 and dentate gyrus (DG) regions of the hippocampus, along with an increase in dendritic spine density in DG. Electrophysiological recordings demonstrated that acute AGH-194 administration enhanced hippocampal excitability by increasing population spike amplitude in CA1. Chronic AGH-194 treatment further modulated short-term plasticity, increasing both population spike and extracellular field potential paired-pulse ratios (PS-PPR and EPSP-PPR) in CA1, while also enhancing the maximum EPSP slope amplitude. These findings provide novel evidence that chronic 5-HT7R activation can induce depressive-like behaviors in male mice, potentially through sustained MMP-9 activation and alterations in synaptic plasticity. Understanding the molecular and electrophysiological consequences of selective 5-HT7R stimulation may provide insights into receptor-specific mechanisms that could contribute to SSRI-induced side effects, thereby contributing to the development of improved antidepressant strategies.

Hypoxia-inducible factor 2-alpha (HIF-2α), a critical transcription factor, forms a heterodimer with aryl hydrocarbon receptor nuclear translocator (ARNT) to drive the transcription of erythropoietin (EPO), a key regulator of erythropoiesis. Activation of this pathway plays a pivotal role in the treatment of anemia. By discovered structure-based virtual screening and pharmacological assays, we herein discovered an amide thiazole AT-1 that bound to HIF-2α with a K_D of 2.63 μM, and enhanced the stability of the HIF-2α-ARNT heterodimer. Molecular docking and site-directed mutagenesis analysis revealed the critical roles of His293 and Tyr307 in the binding of AT-1 to HIF-2α. Pharmacological studies showed that AT-1 (10, 20, 40 μM) dose-dependently enhanced both the transcription and secretion of EPO in 786-O and Hep3B cells. In zebrafish (Danio rerio), AT-1 (10 or 50 μM) exhibited favorable safety profiles and, when combined with the prolyl hydroxylase (PHD) inhibitor Molidustat (10 μM), effectively mitigated doxorubicin-induced anemia. In adenine-induced chronic kidney disease (CKD) mouse model, combined administration of AT-1 (50 mg·kg^-1·d^-1, i.p.) and Molidustat (10 mg·kg^-1·d^-1, i.p.) for 15 days produced stronger effects on increasing EPO levels and alleviating anemia than Molidustat alone, further supporting the therapeutic potential of AT-1 in CKD-related anemia.

Acetaminophen (APAP)-induced acute liver injury (AILI) is primarily driven by CYP3A4‒mediated overproduction of the toxic metabolite N-acetyl-p-benzoquinone imine (NAPQI), CYP3A4 activity serves as the rate-limiting determinant of NAPQI accumulation levels. Poly ADP-ribose polymerase 1 (PARP1)-driven ribosylation, a posttranslational modification, has been linked to drug-induced liver injury. PARP1 interacts with pregnane X receptor (PXR), a nuclear receptor that regulates drug-metabolizing enzymes including CYP3A4. In this study we investigated the specific sites of PARP1-mediated PXR ribosylation, particularly regarding their functional relevance to CYP3A4-driven NAPQI biosynthesis in AILI. To establish AILI models, mice were injected with APAP (300 mg·kg^-1, i.p.), liver tissues and serum were collected for analysis 24 h post-injection. In vitro study was conducted in primary hepatocytes isolated from AILI mice and in human hepatic L02 cells exposed to APAP (5, 10, 20 μM). We demonstrated that under AILI conditions, PARP1 catalyzed ribosylation of PXR at the residue E194, forming a PARP1-PXR‒CYP3A4 regulatory axis that amplified oxidative stress and NAPQI accumulation through a positive feedback loop. Specifically, PARP1 was significantly overexpressed in AILI models in vivo and in vitro, and its interaction with PXR was confirmed in immunoprecipitation and proximity biotinylation assays. Molecular dynamics (MD) simulations, mass spectrometry and E194A site-directed mutagenesis revealed that PARP1-mediated ribosylation of PXR E194 enhanced CYP3A4 transcription, ultimately leading to excessive NAPQI production. MD simulations also identified a natural compound schisandrin B (Sch B) that specifically bound to the ligand-binding domain of PXR, induced conformational changes and disrupted the PARP1-PXR interaction interface, thus suppressed the ribosylation. In AILI murine models, administration of Sch B (25, 50, and 100 mg·kg^-1·d^-1, i.g.) for 8 days significantly reduced serum ALT and AST levels, attenuated oxidative stress, and inhibited NAPQI generation by blocking complex formation. This study not only elucidates the mechanisms of PARP1-mediated PXR E194 ribosylation in AILI but also identifies Sch B as the first specific inhibitor of this pathway, providing a theoretical basis for precision detoxification strategies targeting posttranslational modifications.

Myeloid-derived suppressor cells (MDSCs) are a category of immature myeloid cells that have an important function in suppressing immune responses in a variety of pathological settings. Thus, MDSCs are the subject of intensive studies regarding their recruitment, expulsion, deactivation, and maturation promotion. Tumor necrosis factor superfamily member 15 (TNFSF15) is produced largely by vascular endothelial cells in mature blood vessels with expression also observed in tumor-associated macrophages (TAMs) and dendritic cells (DCs) within the tumor stroma. In addition to inhibiting the proliferation of vascular endothelial cells and the differentiation of bone marrow-derived endothelial cell progenitors, TNFSF15 is able to promote the maturation of DC, as well as to modulate the polarization of naive M2-macrophages into M1-macrophages capable of eliminating cancer cells, and activate T-cell. In this study, we investigated whether a recombinant TNFSF15 results in a substantial reduction of MDSC accumulation in Lewis lung cancer (LLC) tumor-bearing mice. LLC allograft model mice were administered recombinant TNFSF15 (5 mg·kg^-1·d^-1, i.p.) for 7 consecutive days. The tumor, bone marrow and spleen were retrieved on Day 8 and analyzed using flow cytometry or immunofluorescence staining. We showed that TNFSF15 treatment significantly inhibited the tumor growth, and caused a substantial reduction of MDSC accumulation in the tumors. The proportions of MDSC in the bone marrows and the spleens were also reduced. The diminished MDSC was mainly the monocyte-like MDSC (M-MDSC) subtype. Additionally, the reduction in M-MDSC population was accompanied by an increase of the proportions of macrophages and DCs in the tumors. We demonstrated that TNFSF15 promoted M-MDSC differentiation by activating the JAK1/STAT3 signaling pathway. Moreover, the treatment gave rise to a markedly escalated accumulation of cytotoxic T cells in the tumors, attributing to tumor growth inhibition. Our results support the view that TNFSF15-driven differentiation of M-MDSC into DCs and macrophages, and the subsequent activation of T cells, may contribute partially to reinstitution of immunity in the tumor microenvironment.

Morphine-6-glucuronide (M6G), the active metabolite of morphine, is currently in clinical development due to its higher analgesic activity. In humans, intravenously administered M6G was predominantly eliminated unchanged through the kidney, whereas it was excreted into the urine as parent drug as well as its metabolites morphine and M3G in normal rats. In bile-duct-cannulated rats, however, bile excretion of the parent drug was the main route of clearance. In the study, we investigated the mechanisms underlying the species differences in vivo disposition of M6G. In hepatocyte uptake assay, we showed that M6G uptake in rat hepatocytes was 75-fold higher than that in human hepatocytes. Hepatic uptake transporter phenotyping study identified M6G as a substrate for rat rOatplal, rOatpla4, rOatp1b2, as well as for human hOATP1B1 and hOATP1B3. Among these, rOatps exhibited significantly stronger uptake of M6G compared to hOATPs. Furthermore, M6G was not a substrate for the canalicular efflux transporters MDR1, hBCRP/rBcrp, hBSEP/rBsep, and hMRP2, but it was recognized by rMrp2. These findings aligned with the observation that M6G exhibited significant biliary excretion in the rat sandwich cultured hepatocyte (SCH) model, but not in the human SCH. Additionally, no species differences were observed in renal uptake mediated by OAT3. Overall, M6G underwent renal clearance in humans via glomerular filtration and active secretion primarily mediated by hOAT3. Although a portion of M6G was also eliminated through the kidney in rats, the majority was subjected to enterohepatic circulation mediated primarily by rOatps and rMrp2, leading to the formation of morphine and M3G, which were subsequently excreted in the urine. The marked difference in the uptake activities of sinusoidal transporters hOATPs/rOatps and the substrate specificity of canalicular transporters hMRP2/rMrp2 were critical factors underlying the species differences in the hepatobiliary disposition of M6G.

Alcohol-associated liver disease (ALD) remains a predominant cause of chronic hepatic pathology, and effective therapeutic strategies are needed. Krüppel-like factor 15 (KLF15) is a member of the KLF family of zinc-finger transcription factors and is ubiquitously expressed in metabolically active tissues, with a particularly high abundance in the liver. KLF15 has been implicated in various hepatic disorders. In this study, we investigated the pathophysiological role of KLF15 in ALD. We established a National Institute on Alcohol Abuse and Alcoholism (NIAAA) model in mice by feeding them an ethanol Lieber-DeCarli liquid diet containing 5% (vol/vol) ethanol for 10 days. EtOH-fed mice were administered binge ethanol gavage (5 g/kg, body weight) on D11. We observed that the expression levels of KLF15 were significantly decreased in the livers of ALD patients and model mice. Overexpression of KLF15 conferred substantial protective effects in EtOH-fed mice, as evidenced by attenuated hepatic injury, apoptosis, steatosis and inflammation. In ethanol-treated AML-12 cells, overexpression of KLF15 reduced apoptosis and steatosis, whereas KLF15 knockdown exacerbated these pathological features. By performing RNA-seq and bioinformatics analyses, we observed that KLF15 regulated the AKT pathway by directly binding to the PFKFB3 promoter (-128 to -121). The physical interaction between PFKFB3 and AKT1 was further verified by Co-IP and molecular docking. These results suggest that KLF15 is a pivotal regulator of ALD pathogenesis through modulation of the PFKFB3/AKT axis, highlighting its potential as a novel therapeutic target for ALD intervention.

Myocardial hypertrophy is one of the most prominent features of heart failure. SET domain-containing protein 7 (Setd7), a catalytic enzyme responsible for histone H3K4 methylation, has been implicated in various cardiac diseases. In this study we investigated whether Setd7 contributed to the development of cardiac hypertrophy. Male mice were subjected to a hypobaric hypoxic environment for 8 weeks; neonatal rat cardiomyocytes (NRCMs) exposed to hypoxia for 6 h. We showed that hypoxic stimulation significantly upregulated the expression levels of Setd7 along with the expression of hypertrophic markers ANP and BNP in NRCMs. By conducting loss- and gain-of-function assays, we demonstrated that Setd7 modulated the hypertrophic and inflammatory markers in hypoxic cardiomyocytes. We further revealed that Setd7-mediated activation of E2F1 (E2 promoter binding factor 1) triggered the expression of E3 ubiquitin protein ligases WWP2, which catalyzed the ubiquitination and degradation of glutathione peroxidase 4 (GPx4), a critical lipid peroxide-reducing enzyme. This degradation drove extensive lipid peroxidation, thereby exacerbating pathological cardiac hypertrophy. Notably, GPx4 inhibition by ras-selective lethal small molecule 3 (RSL3) abolished the antihypertrophic effects of Setd7 knockdown in cardiomyocytes, underscoring the pivotal role of lipid peroxidation in Setd7-mediated hypertrophic responses. In summary, Setd7 promotes hypoxia-induced cardiac hypertrophy through the Setd7-E2F1-WWP2-GPx4 signaling pathway, suggesting that targeting Setd7 is a promising therapeutic strategy to alleviate hypoxia-induced myocardial hypertrophy.

Despite optimized guideline-directed medical therapy, patients with myocardial infarction (MI) often develop heart failure (HF) primarily because of excessive fibrosis. Bone morphogenetic protein 1 (BMP1) plays a critical role in the fibrotic process, yet its specific role in post-MI myocardial fibrosis remains unclear. In this study, we investigated the complex dynamics between BMP1 and fibrotic processes, offering critical insights for novel strategies to mitigate pathological fibrosis in cardiovascular diseases. An experimental MI model was established in mice by ligating the left anterior descending (LAD) coronary artery. We found that the expression levels of BMP1 were significantly elevated in both the serum of MI patients and the cardiac tissues of MI mice. Administration of the BMP1 inhibitor UK383367 (2 mg/kg, i.p., t.i.d., starting the day of myocardial infarction modeling and maintained for 7 days) in MI mice markedly improved cardiac function, reduced myocardial fibrosis, and attenuated the expression of proinflammatory cytokines, including TNF-α, IL-6 and MCP-1. Proteomic profiling revealed that BMP1 was associated with inflammation and oxidative phosphorylation pathways after MI. We demonstrated that UK383367 (250, 500, and 1000 nM) dose-dependently attenuated M1 macrophage polarization, protected mitochondrial function in lipopolysaccharide-stimulated primary macrophages, and inhibited collagen synthesis in Ang II-stimulated cardiac fibroblasts. Overall, these results reveal a pivotal yet detrimental role for BMP1 in driving myocardial fibrosis and amplifying inflammatory cascades after MI. This study highlights the therapeutic potential of the BMP1 inhibitor UK383367 as a promising alternative to conventional antifibrotic strategies, potentially curbing the progression toward HF.

The progression of colitis-associated cancer (CAC) is strongly associated with bone marrow-derived immunosuppressive cells (MDSCs). Although CAC could be suppressed by inducing MDSCs apoptosis, the immunosuppressive tumor microenvironment (TME) maintains immune homeostasis by upregulating M2-type tumor-associated macrophages (TAMs), thus leading to adaptive immune tolerance. Herein, we develop a dendritic cell (DC)-liposome conjugate to reverse immunosuppressive TME, showing remarkable efficiency against colorectal cancer. The DC-liposome conjugate is fabricated by conjugating resolvin E1-loaded liposomes with Fas ligand-transfected DCs, which eliminates tumor-infiltrated Fas⁺ MDSCs and enhances TAM phagocytosis in tumors. It shows significant therapeutic effects in preclinical CAC models and alleviates severe colitis when combined with immune checkpoint inhibitors. This study provides a feasible and customized cell-drug conjugate to overcome immunosuppressive TME for enhancing CAC immunotherapy.

Chemotherapeutic resistance is a significant issue in the treatment of breast cancer, which is related to pyroptosis inhibition. Increasing evidence suggests that long non-coding RNAs (lncRNAs) contribute to tumorigenesis and drug resistance. In this study we investigated the role of the lncRNA STMN1P2 in doxorubicin resistance in breast cancer, as well as its correlation with pyroptosis inhibition. Our results showed that the expression levels of lncRNA STMN1P2 were significantly elevated in doxorubicin-resistant breast cancer tissues and cells. We demonstrated that knockdown of STMN1P2 reduced doxorubicin resistance in breast cancer cells; overexpression of STMN1P2 inhibited doxorubicin-induced pyroptosis by reducing the expression of NLRP3, ASC, caspase-1 and GSDMD. Furthermore, STMN1P2 directly bound to and positively regulated heterogeneous nuclear ribonucleoprotein U (hnRNPU), and knockdown of hnRNPU reversed the inhibitory effect of STMN1P2 on pyroptosis and its ability to promote chemoresistance. In doxorubicin-resistant cells, hnRNPU directly bound to enhancer of zeste homologue 2 (EZH2), and STMN1P2 enhanced hnRNPU recruitment of EZH2 and increased EZH2 protein stability. EZH2 acted as a transcription factor to inactivate TNF receptor-associated factor 6 (TRAF6), thereby repressing the binding of TRAF6 with MALT1 and caspase-1, attenuating the canonical pathways of pyroptosis. In MCF7/DOX cells xenograft nude mouse model, we demonstrated that knockdown of STMN1P2 significantly enhanced the suppression of doxorubicin on the tumour growth. This study provides new clues and approaches for the prevention and treatment of breast cancer chemoresistance.