Journal of pharmacological sciences最新文献

Subretinal hemorrhages result in poor vision and visual field defects. During hemorrhage, several potentially toxic substances are released from iron-based hemoglobin and hemin, inducing cellular damage, the detailed mechanisms of which remain unknown. We examined the effects of excess intracellular iron on retinal pigment epithelial (RPE) cells. A Fe²⁺ probe, SiRhoNox-1 was used to investigate Fe²⁺ accumulation after treatment with hemoglobin or hemin in the human RPE cell line ARPE-19. We also evaluated the production of reactive oxygen species (ROS) and lipid peroxidation. Furthermore, the protective effect of-an iron chelator, 2,2′-bipyridyl (BP), and ferrostatin-1 (Fer-1) on the cell damage, was evaluated. Fe²⁺ accumulation increased in the hemoglobin- or hemin-treated groups, as well as intracellular ROS production and lipid peroxidation. In contrast, BP treatment suppressed RPE cell death, ROS production, and lipid peroxidation. Pretreatment with Fer-1 ameliorated cell death in a concentration-dependent manner and suppressed ROS production and lipid peroxidation. Taken together, these findings indicate that hemoglobin and hemin, as well as subretinal hemorrhage, may induce RPE cell damage and visual dysfunction via intracellular iron accumulation.

Imeglimin is a novel oral antidiabetic drug for treating type 2 diabetes. However, the effect of imeglimin on NLRP3 inflammasome activation has not been investigated yet. Here, we aimed to investigate whether imeglimin reduces LPS-induced NLRP3 inflammasome activation in THP-1 macrophages and examine the associated underlying mechanisms. We analyzed the mRNA and protein expression levels of NLRP3 inflammasome components and IL-1β secretion. Additionally, reactive oxygen species (ROS) generation, mitochondrial membrane potential, and mitochondrial permeability transition pore (mPTP) opening were measured by flow cytometry. Imeglimin inhibited NLRP3 inflammasome-mediated IL-1β production in LPS-stimulated THP-1-derived macrophages. In addition, imeglimin reduced LPS-induced mitochondrial ROS production and mitogen-activated protein kinase phosphorylation. Furthermore, imeglimin restored the mitochondrial function by modulating mitochondrial membrane depolarization and mPTP opening. We demonstrated for the first time that imeglimin reduces LPS-induced NLRP3 inflammasome activation by inhibiting mPTP opening in THP-1 macrophages. These results suggest that imeglimin could be a promising new anti-inflammatory agent for treating diabetic complications.

The ubiquitin-proteasome system (UPS) is a major proteolytic system that plays an important role in the regulation of various cell processes, such as cell cycle, stress response, and transcriptional regulation, especially in neurons, and dysfunction of UPS is considered to be a cause of neuronal cell death in neurodegenerative diseases. However, the mechanism of neuronal cell death caused by UPS dysfunction has not yet been fully elucidated.

In this study, we investigated the mechanism of neuronal cell death induced by proteasome inhibitors using human neuroblastoma SH-SY5Y cells. Z-Leu-D-Leu-Leu-al (MG132), a proteasome inhibitor, induced apoptosis in SH-SY5Y cells in a concentration- and time-dependent manner. Antioxidants N-acetylcysteine and EUK-8 attenuated MG132-induced apoptosis. Apocynin and diphenyleneiodonium, inhibitors of NADPH oxidase (NOX), an enzyme that produces superoxide anions, also attenuated MG132-induced apoptosis. It was also found that MG132 treatment increased the expression of NOX5, a NOX family member, and that siRNA-mediated silencing of NOX5 and BAPTA-AM, which inhibits NOX5 by chelating calcium, suppressed MG132-induced apoptosis and production of reactive oxygen species in SH-SY5Y cells.

These results suggest that MG132 induces apoptosis in SH-SY5Y cells through the production of superoxide anion by NOX5.

Microglia are the residential immune cells in the central nervous system. Their roles as innate immune cells and regulators of synaptic remodeling are critical to the development and the maintenance of the brain. Numerous studies have depleted microglia to elucidate their involvement in healthy and pathological conditions. PLX3397, a blocker of colony stimulating factor 1 receptor (CSF1R), is widely used to deplete mouse microglia due to its non-invasiveness and convenience. Recently, other small rodents, including Syrian hamsters (Mesocricetus auratus) and Mongolian gerbils (Meriones unguiculatus), have been recognized as valuable animal models for studying brain functions and diseases. However, whether microglia depletion via PLX3397 is feasible in these species remains unclear. Here, we administered PLX3397 orally via food pellets to hamsters and gerbils. PLX3397 successfully depleted gerbil microglia but had no effect on microglial density in hamsters. Comparative analysis of the CSF1R amino acid sequence in different species hints that amino acid substitutions in the juxtamembrane domain may potentially contribute to the inefficacy of PLX3397 in hamsters.

L-type amino acid transporter 1 (LAT1) is recognized as a promising target for cancer therapy; however, the cellular adaptive response to its pharmacological inhibition remains largely unexplored. This study examined the adaptive response to LAT1 inhibition using nanvuranlat, a high-affinity LAT1 inhibitor. Proteomic analysis revealed the activation of a stress-induced transcription factor ATF4 following LAT1 inhibition, aligning with the known cellular responses to amino acid deprivation. This activation was linked to the GCN2-eIF2α pathway which regulates translation initiation. Our results show that ATF4 upregulation counteracts the suppressive effect of nanvuranlat on cell proliferation in pancreatic ductal adenocarcinoma cell lines, suggesting a role for ATF4 in cellular adaptation to LAT1 inhibition. Importantly, dual targeting of LAT1 and ATF4 exhibited more substantial anti-proliferative effects in vitro than individual treatments. This study underscores the potential of combining LAT1 and ATF4 inhibition as a therapeutic strategy in cancer treatment.

Background

Gallic acid (GA) is an organic compound with phenolic properties that occurs naturally and can be found in Guizhi Fuling capsules, showcasing a wide range of biological functionalities.

Purpose

The objective of this study was to examine the influence of GA on endometrial hyperplasia (EH) and elucidate its underlying mechanism.

Methods

Initially, the induction of EH was achieved by administering estradiol to mice via continuous subcutaneous injection for a duration of 21 days. Concurrently, GA treatment was administered, and subsequently, the uterine tissue structure was assessed using hematoxylin and eosin (H&E) staining. Following this, the proliferation of human endometrial cells treated by GA was determined utilizing the CCK-8 method. Furthermore, network pharmacology and single-cell-RNA-seq data were employed to identify the target of GA action. In addition, we will employ immunofluorescence (IF), immunohistochemistry (IHC), flow cytometry, western blot and RT-qPCR methodologies to investigate the impact of GA on the expression level of cyclin D1, PI3K, p-PI3K, AKT, p-AKT.

Results

GA treatment ameliorated histopathological alterations in the uterus and suppress proliferation. Estradiol stimulation can activate the PI3K/AKT pathway, leading to up-regulation of cyclin D1 expression, whereas GA treatment results in down-regulation of its expression.

Conclusions

The expression of cyclin D1 is down-regulated by GA through the inhibition of the PI3K/AKT pathway, effectively mitigating estradiol-induced EH in mice.

Purpose

To determine whether combination of topical ripasudil and brimonidine has more effective neuroprotection on retinal ganglion cells (RGCs) following injury to axons composing the optic nerve.

Methods

Topical ripasudil, brimonidine, or mixture of both drugs were administered to adult mice after optic nerve injury (ONI). The influence of drug conditions on RGC health were evaluated by the quantifications of surviving RGCs, phosphorylated p38 mitogen-activated protein kinase (phospho-p38), and expressions of trophic factors and proinflammatory mediators in the retina.

Results

Topical ripasudil and brimonidine suppressed ONI-induced RGC death respectively, and mixture of both drugs further stimulated RGC survival. Topical ripasudil and brimonidine suppressed ONI-induced phospho-p38 in the whole retina. In addition, topical ripasudil suppressed expression levels of TNFα, IL-1β and monocyte chemotactic protein-1 (MCP-1), whereas topical brimonidine increased the expression level of basic fibroblast growth factor (bFGF).

Conclusions

Combination of topical ripasudil and brimonidine may enhance RGC protection by modulating multiple signaling pathways in the retina.

We previously identified a spinal astrocyte population that expresses hairy and enhancer of split 5 (Hes5) and is selectively present in superficial laminae in mice. However, it was unclear whether such astrocyte heterogeneity is commonly observed across species. Using adeno-associated viral (AAV) vectors incorporating a rat Hes5 promotor (AAV-Hes5P), we found that AAV-Hes5P-captured astrocytes were selectively located in the superficial laminae in rats. Furthermore, activation of AAV-Hes5P⁺ astrocytes elicited allodynia-like behavior and increased c-FOS⁺ cells in the superficial laminae. Thus, laminar-selective Hes5⁺ astrocytes are conserved beyond species and have the capability to convert tactile information to nociceptive.

Diabetic nephropathy is a common complication of diabetes, accumulating evidence underscores the pivotal role of tubulointerstitial fibrosis in the progression of diabetic nephropathy. However, the underlying mechanisms remain incompletely understood. Although the mechanisms in diabetic nephropathy fibrosis have been the focus of many studies, only limited information is currently available concerning microRNA regulation in tubulointerstitial fibrosis. In this study, we aimed to investigate the roles of miR-320a-3p and bone morphogenetic protein-6 (BMP6) in tubulointerstitial fibrosis. After inducing fibrosis with high glucose in HK-2 cells, we found that miR-320a-3p is significantly up-regulated, whereas BMP6 is markedly down-regulated. These changes suggest close link between miR-320a-3p and BMP6 in tubulointerstitial fibrosis. To elucidate this phenomenon, miR-320a-3p mimic, inhibitor and siBMP6 were employed. We observed in miR-320a-3p mimic group the fibrosis marker include alpha smooth muscle actin and type I collagen was significantly up-regulated, whereas BMP6 exhibited the opposite trend. Additionally, we found icariin could alleviate tubulointerstitial fibrosis by downregulation the miR-320a-3p expression. In conclusion, miR-320a-3p promotes tubulointerstitial fibrosis during the development of DN by suppressing BMP signal pathway activity via inhibiting BMP6 expression. Suggesting that miR-320a-3p represents a potential therapeutic target for tubulointerstitial fibrosis induced by diabetic nephropathy.

Amino acid transporter LAT1 is highly upregulated in various cancer types, including cholangiocarcinoma (CHOL), and contributes to the rapid proliferation of cancer cells and disease progression. However, the molecular mechanisms underlying the pathological upregulation of LAT1 remain largely unknown. This study pursued the possibility of miRNA-mediated regulation of the LAT1 expression in CHOL cells. Using online target prediction methods, we extracted five candidate miRNAs commonly predicted to regulate the LAT1 expression. Three of them, miR-194-5p, miR-122-5p, and miR-126-3p, were significantly downregulated in CHOL cancer compared to normal tissues. Correlation analysis revealed weak-to-moderate negative correlations between the expression of these miRNAs and LAT1 mRNA in CHOL cancer tissues. We selected miR-194-5p and miR-122-5p for further analyses and found that both miRNAs functionally target 3′UTR of LAT1 mRNA by a luciferase-based reporter assay. Transfection of the miRNA mimics significantly suppressed the LAT1 expression at mRNA and protein levels and inhibited the proliferation of CHOL cells, with a trend of affecting intracellular amino acids and amino acid-related signaling pathways. This study indicates that the decreased expression of these LAT1-targeting tumor-suppressive miRNAs contributes to the upregulation of LAT1 and the proliferation of CHOL cells, highlighting their potential for developing novel cancer therapeutics and diagnostics.