-
Molecules (Basel, Switzerland) May 2023Metal-derived platinum complexes are widely used to treat solid tumors. However, systemic toxicity and tumor resistance to these drugs encourage further research into...
Metal-derived platinum complexes are widely used to treat solid tumors. However, systemic toxicity and tumor resistance to these drugs encourage further research into similarly effective compounds. Among others, organotin compounds have been shown to inhibit cell growth and induce cell death and autophagy. Nevertheless, the impact of the ligand structure and mechanisms involved in the toxicity of organotin compounds have not been clarified. In the present study, the biological activities of commercially available bis(tributyltin) oxide and tributyltin chloride, in comparison to those of specially synthesized tributyltin trifluoroacetate (TBT-OCOCF) and of cisplatin, were assessed using cells with different levels of tumorigenicity. The results show that tributyltins were more cytotoxic than cisplatin in all the tested cell lines. NMR revealed that this was not related to the interaction with DNA but to the inhibition of glucose uptake into the cells. Moreover, highly tumorigenic cells were less susceptible than nontumorigenic cells to the nonunique pattern of death induced by TBT-OCOCF. Nevertheless, tumorigenic cells became sensitive when cotreated with wortmannin and TBT-OCOCF, although no concomitant induction of autophagy by the compound was detected. Thus, TBT-OCOCF might be the prototype of a family of potential anticancer agents.
Topics: Cisplatin; Cell Line, Tumor; Trialkyltin Compounds; Antineoplastic Agents; Organotin Compounds; Coordination Complexes
PubMed: 37175265
DOI: 10.3390/molecules28093856 -
Biochemistry Jun 2022Inositol pyrophosphates are signaling molecules containing at least one phosphoanhydride bond that regulate a wide range of cellular processes in eukaryotes. With a...
Inositol pyrophosphates are signaling molecules containing at least one phosphoanhydride bond that regulate a wide range of cellular processes in eukaryotes. With a cyclic array of phosphate esters and diphosphate groups around -inositol, these molecular messengers possess the highest charge density found in nature. Recent work deciphering inositol pyrophosphate biosynthesis in revealed important functions of these messengers in nutrient sensing, hormone signaling, and plant immunity. However, despite the rapid hydrolysis of these molecules in plant extracts, very little is known about the molecular identity of the phosphohydrolases that convert these messengers back to their inositol polyphosphate precursors. Here, we investigate whether Plant and Fungi Atypical Dual Specificity Phosphatases (PFA-DSP1-5) catalyze inositol pyrophosphate phosphohydrolase activity. We find that recombinant proteins of all five PFA-DSP homologues display phosphohydrolase activity with a high specificity for the 5-β-phosphate of inositol pyrophosphates and only minor activity against the β-phosphates of 4-InsP and 6-InsP. We further show that heterologous expression of PFA-DSP1-5 rescues wortmannin sensitivity and deranged inositol pyrophosphate homeostasis caused by the deficiency of the PFA-DSP-type inositol pyrophosphate phosphohydrolase Siw14 in yeast. Heterologous expression in leaves provided evidence that PFA-DSP1 also displays 5-β-phosphate-specific inositol pyrophosphate phosphohydrolase activity . Our findings lay the biochemical basis and provide the genetic tools to uncover the roles of inositol pyrophosphates in plant physiology and plant development.
Topics: Arabidopsis; Diphosphates; Dual-Specificity Phosphatases; Inositol Phosphates; Saccharomyces cerevisiae
PubMed: 35640071
DOI: 10.1021/acs.biochem.2c00145 -
Annals of Hepatology 2022Type 2 diabetes mellitus (T2DM) increases the occurrence and mortality of liver cancer. Insulin growth factor (IGF) plays a crucial role in the development of diabetes...
INTRODUCTION AND OBJECTIVES
Type 2 diabetes mellitus (T2DM) increases the occurrence and mortality of liver cancer. Insulin growth factor (IGF) plays a crucial role in the development of diabetes and liver cancer, and specifically, IGF-1 may be involved in the development of liver cancer with preexisting T2DM. Autophagy contributes to cancer cell survival and apoptosis. However, the relationship between IGF-1 and autophagy has rarely been evaluated. The purpose of this study was to investigate whether IGF-1 promotes the development of liver cancer in T2DM patients by promoting autophagy.
MATERIALS AND METHODS
Thirty-three hepatocellular carcinoma (HCC) patients with T2DM and 33 age-matched patients with HCC without T2DM were included in this study. We analyzed the expression of IGF-1 and autophagy-related LC3 and p62 mRNA and the prognosis of two groups. In vitro, we stimulated HepG2 cells with IGF-1 and then detected changes in autophagy and cell proliferation, apoptosis, and migration in the presence/absence of wortmannin, an autophagy inhibitor.
RESULTS
IGF-1 promoted autophagy, resulting in inhibition of apoptosis and induction of growth and migration of HepG2 cells. Inhibition of autophagy by wortmannin impaired IGF-1 function. Higher expression of IGF-1 was detected in HCC patients with T2DM. IGF-1 expression was higher in liver cancer tissue compared to paracancerous tissue. Elevated IGF-1 was associated with a poor prognosis in patients with HCC.
CONCLUSIONS
IGF-1 participates in the development of liver cancer by inducing autophagy. Elevated IGF-1 was a prognostic factor for patients with HCC, especially when accompanied by T2DM.
Topics: Autophagy; Carcinoma, Hepatocellular; Cell Line, Tumor; Diabetes Mellitus, Type 2; Humans; Insulin; Insulin-Like Growth Factor I; Liver Neoplasms; Wortmannin
PubMed: 35297369
DOI: 10.1016/j.aohep.2022.100697 -
Journal of Leukocyte Biology Apr 2021Extracellular histones have been implicated as a cause of tissue inflammatory injury in a variety of disorders including sepsis, lung, and liver diseases. However,...
Extracellular histones have been implicated as a cause of tissue inflammatory injury in a variety of disorders including sepsis, lung, and liver diseases. However, little is known about their interactions with neutrophils and how this might contribute to injury. Here, it is shown that histone H4 acts as neutrophil activator by inducing hydrogen peroxide production, degranulation, cell adhesion, and IL-8 generation. Histone H4 caused permeabilization of the neutrophil membrane (a phenomenon described in other cell types) leading to accelerated cell death. H4 caused sustained rise in neutrophil intracellular calcium that is necessary for respiratory burst activation and degranulation. Convincing evidence was not found for TLRs or ATP receptors in H4 mediated activation. However, pertussis toxin and wortmannin (inhibitors of G protein and PI3K) inhibited H4-induced hydrogen peroxide production and degranulation. These studies suggest that release of histone H4 in sites of infection or inflammation may potentiate neutrophil activation and promote additional inflammatory responses. These studies may provide a better basis for developing novel therapeutic strategies to block neutrophil extracellular trap (NET) and H4-related pathology in sepsis and various forms of lung injury including that induced by viruses like influenza or SAR-CoV2.
Topics: Calcium; Cell Adhesion; Cell Degranulation; Cell Membrane; Cell Membrane Permeability; Histones; Humans; Hydrogen Peroxide; Integrins; Monocytes; Neutrophil Activation; Neutrophils; Peroxidase; Pertussis Toxin; Respiratory Burst; Signal Transduction; Wortmannin
PubMed: 32803840
DOI: 10.1002/JLB.3A0620-342R -
Cell Death & Disease Feb 2022Apoptotic exosome-like vesicles (ApoExos) are a novel type of extracellular vesicle that contribute to the propagation of inflammation at sites of vascular injury when...
Apoptotic exosome-like vesicles (ApoExos) are a novel type of extracellular vesicle that contribute to the propagation of inflammation at sites of vascular injury when released by dying cells. ApoExos are characterized by the presence of the C-terminal perlecan LG3 fragment and 20S proteasome, and they are produced downstream of caspase-3 activation. In the present study, we assessed the relative roles of autophagy and caspase-3-mediated pathways in controlling the biogenesis and secretion of immunogenic ApoExos. Using electron microscopy and confocal immunofluorescence microscopy in serum-starved endothelial cells, we identified large autolysosomes resulting from the fusion of lysosomes, multivesicular bodies, and autophagosomes as a site of ApoExo biogenesis. Inhibition of autophagy with ATG7 siRNA or biochemical inhibitors (wortmannin and bafilomycin) coupled with proteomics analysis showed that autophagy regulated the processing of perlecan into LG3 and its loading onto ApoExos but was dispensable for ApoExo biogenesis. Caspase-3 activation was identified using caspase-3-deficient endothelial cells or caspase inhibitors as a pivotal regulator of fusion events between autolysosomes and the cell membrane, therefore regulating the release of immunogenic ApoExos. Collectively, these findings identified autolysosomes as a site of ApoExo biogenesis and caspase-3 as a crucial regulator of autolysosome cell membrane interactions involved in the secretion of immunogenic ApoExos.
Topics: Autophagosomes; Autophagy; Caspase 3; Endothelial Cells; Exosomes; Lysosomes
PubMed: 35149669
DOI: 10.1038/s41419-022-04591-5 -
Cell Death and Differentiation Apr 2023Regulated cell death (RCD) plays an important role in the progression of viral replication and particle release in cells infected by herpes simplex virus-1 (HSV-1)....
Regulated cell death (RCD) plays an important role in the progression of viral replication and particle release in cells infected by herpes simplex virus-1 (HSV-1). However, the kind of RCD (apoptosis, necroptosis, others) and the resulting cytopathic effect of HSV-1 depends on the cell type and the species. In this study, we further investigated the molecular mechanisms of apoptosis induced by HSV-1. Although a role of caspase-8 has previously been suggested, we now clearly show that caspase-8 is required for HSV-1-induced apoptosis in a FADD-/death receptor-independent manner in both mouse embryo fibroblasts (MEF) and human monocytes (U937). While wild-type (wt) MEFs and U937 cells exhibited increased caspase-8 and caspase-3 activation and apoptosis after HSV-1 infection, respective caspase-8-deficient (caspase-8-/-) cells were largely impeded in any of these effects. Unexpectedly, caspase-8-/- MEF and U937 cells also showed less virus particle release associated with increased autophagy as evidenced by higher Beclin-1 and lower p62/SQSTM1 levels and increased LC3-I to LC3-II conversion. Confocal and electron microscopy revealed that HSV-1 stimulated a strong perinuclear multivesicular body response, resembling increased autophagy in caspase-8-/- cells, entrapping virions in cellular endosomes. Pharmacological inhibition of autophagy by wortmannin restored the ability of caspase-8-/- cells to release viral particles in similar amounts as in wt cells. Altogether our results support a non-canonical role of caspase-8 in both HSV-1-induced apoptosis and viral particle release through autophagic regulation.
Topics: Animals; Mice; Humans; Herpesvirus 1, Human; Caspase 8; Apoptosis; Autophagy; Virion; Caspase 3
PubMed: 36418547
DOI: 10.1038/s41418-022-01084-y -
Calcium-Prolactin Secretion Coupling in Rat Pituitary Lactotrophs Is Controlled by PI4-Kinase Alpha.Frontiers in Endocrinology 2021The role of calcium, but not of other intracellular signaling molecules, in the release of pituitary hormones by exocytosis is well established. Here, we analyzed the...
The role of calcium, but not of other intracellular signaling molecules, in the release of pituitary hormones by exocytosis is well established. Here, we analyzed the contribution of phosphatidylinositol kinases (PIKs) to calcium-driven prolactin (PRL) release in pituitary lactotrophs: PI4Ks - which control PI4P production, PIP5Ks - which synthesize PI(4, 5)P2 by phosphorylating the D-5 position of the inositol ring of PI4P, and PI3KCs - which phosphorylate PI(4, 5)P to generate PI(3, 4, 5)P. We used common and PIK-specific inhibitors to evaluate the strength of calcium-secretion coupling in rat lactotrophs. Gene expression was analyzed by single-cell RNA sequencing and qRT-PCR analysis; intracellular and released hormones were assessed by radioimmunoassay and ELISA; and single-cell calcium signaling was recorded by Fura 2 imaging. Single-cell RNA sequencing revealed the expression of and , as well as and , in lactotrophs. Wortmannin, a PI3K and PI4K inhibitor, but not LY294002, a PI3K inhibitor, blocked spontaneous action potential driven PRL release with a half-time of ~20 min when applied in 10 µM concentration, leading to accumulation of intracellular PRL content. Wortmannin also inhibited increase in PRL release by high potassium, the calcium channel agonist Bay K8644, and calcium mobilizing thyrotropin-releasing hormone without affecting accompanying calcium signaling. GSK-A1, a specific inhibitor of PI4KA, also inhibited calcium-driven PRL secretion without affecting calcium signaling and expression. In contrast, PIK93, a specific inhibitor of PI4KB, and ISA2011B and UNC3230, specific inhibitors of PIP5K1A and PIP5K1C, respectively, did not affect PRL release. These experiments revealed a key role of PI4KA in calcium-secretion coupling in pituitary lactotrophs downstream of voltage-gated and PI(4, 5)P2-dependent calcium signaling.
Topics: 3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; Animals; Calcium; Calcium Channel Agonists; Calcium Signaling; Exocytosis; Lactotrophs; Minor Histocompatibility Antigens; Phosphotransferases (Alcohol Group Acceptor); Prolactin; Protein Kinase Inhibitors; Rats; Sequence Analysis, RNA; Single-Cell Analysis; Wortmannin
PubMed: 35058881
DOI: 10.3389/fendo.2021.790441 -
Neural Regeneration Research Jul 2024Autism spectrum disorders are a group of neurodevelopmental disorders involving more than 1100 genes, including as a candidate gene. knockout mice, serving as an...
Autism spectrum disorders are a group of neurodevelopmental disorders involving more than 1100 genes, including as a candidate gene. knockout mice, serving as an animal model of autism, have been demonstrated to exhibit decreased density of dendritic spines. The role of melatonin, as a neurohormone capable of effectively alleviating social interaction deficits and regulating the development of dendritic spines, in deletion-induced nerve injury remains unclear. In the present study, we discovered that the deletion of exon 2 of the gene was linked to social interaction deficits, spine loss, impaired inhibitory neurons, and suppressed phosphatidylinositol-3-kinase (PI3K)/protein kinase B (Akt) signal pathway in the prefrontal cortex. Our findings demonstrated that the long-term oral administration of melatonin for 28 days effectively alleviated the aforementioned abnormalities in gene-knockout mice. Furthermore, the administration of melatonin in the prefrontal cortex was found to improve synaptic function and activate the PI3K/Akt signal pathway in this region. The pharmacological blockade of the PI3K/Akt signal pathway with a PI3K/Akt inhibitor, wortmannin, and melatonin receptor antagonists, luzindole and 4-phenyl-2-propionamidotetralin, prevented the melatonin-induced enhancement of GABAergic synaptic function. These findings suggest that melatonin treatment can ameliorate GABAergic synaptic function by activating the PI3K/Akt signal pathway, which may contribute to the improvement of dendritic spine abnormalities in autism spectrum disorders.
PubMed: 38051907
DOI: 10.4103/1673-5374.387973 -
Archives of Virology Mar 2022Japanese encephalitis (JE) is a zoonotic epidemic disease caused by Japanese encephalitis virus (JEV), and currently, no medicines are available to treat this disease....
Japanese encephalitis (JE) is a zoonotic epidemic disease caused by Japanese encephalitis virus (JEV), and currently, no medicines are available to treat this disease. Autophagy modulators play an important role in the treatment of tumors, heart disease, and some viral diseases. The aim of this study was to investigate the effects of autophagy modulators on JEV infection and the host response in mice. The experimental mice were grouped as follows: DMEM (control), JEV, JEV+rapamycin (JEV+Rapa), JEV+wortmannin (JEV+Wort), JEV+chloroquine (JEV+CQ), Rapa, Wort, and CQ. The control group was treated with DMEM. The mice in other groups were infected with 10 PFU of JEV, and Rapa, Wort, and CQ were administered 2 h prior to JEV challenge and then administered daily for 10 consecutive days. All mice were monitored for neurological signs and survival. The damage of subcellular structures in the mouse brain was evaluated by transmission electron microscopy. The distribution of virus in the mouse brain was determined by RNAScope staining and immunohistochemical staining. The neuroinflammatory responses in the brain were examined via quantitative real-time PCR, and the signal pathways involved in neuroinflammation were identified by Western blot. The mice in the JEV+Wort and JEV+CQ groups showed milder neurological symptoms, less damage to the mitochondria in the brain tissue, and a higher survival rate than those in the JEV+Rapa and JEV groups. Compared with the JEV+Rapa and JEV groups, the distribution of JEV in the brain of mice in the JEV+Wort and JEV+CQ groups was lower, and the inflammatory response was weaker. No significant difference was observed in the expression of the PI3K/AKT/NF-κB pathway in mouse brain among the different groups. Our study suggests that the autophagy inhibitors Wort and CQ reduce JEV infection and weaken the inflammatory response, which does not depend on the PI3K/AKT/NF-κB pathway in mouse brain.
Topics: Animals; Autophagy; Encephalitis Virus, Japanese; Encephalitis, Japanese; Inflammation; Mice; Phosphatidylinositol 3-Kinases
PubMed: 35119507
DOI: 10.1007/s00705-021-05283-9 -
Frontiers in Oncology 2021Development of new therapeutic strategies for breast cancer is urgently needed due to the sustained emergence of drug resistance, tumor recurrence and metastasis. To...
Development of new therapeutic strategies for breast cancer is urgently needed due to the sustained emergence of drug resistance, tumor recurrence and metastasis. To gain a novel insight into therapeutic approaches to fight against breast cancer, the cytocidal effects of hellebrigenin (Helle) and arenobufagin (Areno) were investigated in human estrogen receptor (ER)-positive breast cancer cell line MCF-7 and triple-negative breast cancer cell line MDA-MB-231. Helle exhibited more potent cytotoxicity than Areno in both cancer cells, and MCF-7 cells were more susceptible to both drugs in comparison with MDA-MB-231 cells. Apoptotic-like morphological characteristics, along with the downregulation of the expression level of Bcl-2 and Bcl-xL and the upregulation of the expression level of Bad, were observed in Helle-treated MCF-7 cells. Helle also caused the activation of caspase-8, caspase-9, along with the cleavage of poly(ADP-ribose) polymerase in MCF-7 cells. Helle-mediated necrosis-like phenotype, as evidenced by the increased propidium iodide (PI)-positive cells was further observed. G/M cell cycle arrest was also induced by Helle in the cells. Upregulation of the expression level of p21 and downregulation of the expression level of cyclin D1, cyclin E1, cdc25C and survivin were observed in MCF-7 cells treated with Helle and occurred in parallel with G/M arrest. Autophagy was triggered in MCF-7 cells and the addition of wortmannin or 3-MA, two well-known autophagy inhibitors, slightly but significantly rescued the cells. Furthermore, similar alterations of some key molecules associated with the aforementioned biological phenomena were observed in MDA-MB-231 cells. Intriguingly, the numbers of PI-positive cells in Helle-treated MCF-7 cells were significantly reduced by wortmannin and 3-MA, respectively. In addition, Helle-triggered G/M arrest was significantly corrected by wortmannin, suggesting autophagy induction contributed to Helle-induced cytotoxicity of breast cancer cells by modulating necrosis and cell cycle arrest. Collectively, our results suggested potential usefulness of both Helle and Areno in developing therapeutic strategies to treat patients with different types of breast cancer, especially ER-positive breast cancer.
PubMed: 34513690
DOI: 10.3389/fonc.2021.711220