Autophagy In Cells Research Articles

RAB37 suppresses the EMT, migration and invasion of gastric cancer cells by mediating autophagic degradation of β-catenin.

Gastric cancer, characterized by its high morbidity and mortality rates, exhibits low levels of RAB37. The role and molecular mechanisms of RAB37, a small GTPase, in the pathogenesis of gastric cancer are still unclear. We assessed RAB37 expression in gastric cancer cells using quantitative Polymerase Chain Reaction (qPCR), Western blot, and immunohistochemical staining (IHC), and analyzed EMT marker proteins and autophagy changes via Western blot, immunofluorescence (IF), and transmission electron microscopy (TEM). Co-immunoprecipitation (co-IP) was used to identify protein-protein interactions. We studied the migration and invasion of gastric cancer cells using wound healing and transwell assays in vitro and a mouse pulmonary metastasis model in vivo. Overexpression of RAB37 suppressed EMT, invasion, and migration while enhancing autophagy in gastric cancer cells, which was dependent on its GTPase activity. However, all these effects could be reversed by the autophagy inhibitor chloroquine. Regarding the molecular mechanism, RAB37 strengthened the interaction between p62 and β-catenin, which consequently enhanced the p62-mediated autophagic degradation of β-catenin. Furthermore, RAB37 curbed the pulmonary metastasis of both general and cisplatin-resistant gastric cancer cells. The low level of RAB37 reduces interaction between p62 and β-catenin and then the autophagic degradation of β-catenin, thereby promoting the EMT, invasion, and migration in gastric cancer cells. The low expression of RAB37 in gastric cancer suggests a potential therapeutic target, especially for cisplatin-resistant gastric cancer.

Digoxigenin activates autophagy in hepatocellular carcinoma cells by regulating the PI3K/AKT/mTOR pathway

Hepatocellular carcinoma (HCC) is recognized as a highly malignant tumor. Targeted combination immunotherapy, the initially approved regimen, is compromised by adverse side effects and low response rates during clinical treatment. Traditional Chinese medicine and its derived natural compounds, known for their anticancer effects, offer advantages of low toxicity and cost. In this study, we performed high-throughput phenotypic screening in vitro to identify promising anti-HCC drugs. Among 1,444 bioactive compounds, digoxigenin (DIG) was found to significantly impede HCC cell progression. We validated DIG’s therapeutic effects through assays such as cell counting by CCK8, lactate dehydrogenase, and colony formation. Analyses including transmission electron microscopy, western blotting, and immunofluorescence demonstrated that DIG inhibits HCC cell proliferation via autophagy. Network pharmacology and molecular docking studies suggest that DIG targets the PI3K/AKT/mTOR signaling pathway. Comparative treatments of Hep3B and Huh7 cells with DIG or mTOR inhibitors revealed similar inhibitory impacts, indicating that DIG induces autophagy by inhibiting the PI3K/AKT/mTOR pathway. In vivo studies confirmed that DIG halts the growth of subcutaneous xenograft tumors. In conclusion, DIG represents a potential HCC treatment by modulating the PI3K/AKT/mTOR pathway to induce autophagy. This research, via phenotypic screening, accelerates drug discovery and the development of novel therapies targeting the underlying mechanisms of liver cancer.

Involvement of autophagy in germ cells in an experimental model of varicocele in rats before and after varicocelectomy.

Autophagy, a catabolic process enabling cellular organelles and proteins' reuse for energy, has been observed in varicocele models, but the effect of surgical treatment on this process remains unknown. This study aims to assess autophagy in varicocele models undergoing surgical correction. Twenty-one adolescent male rats were induced with varicocele and divided into three groups: sham, varicocele, and varicocele with varicocelectomy. After 21days, testicles were examined histologically for spermatogenesis (Jonhsen's score) and immunohistochemically for autophagy markers (LC3A, Beclin-1, Ambra-1, ULK-1, p62). Positive germ cells were quantitatively evaluated, and data were statistically analyzed (p < 0.05). Histological examination revealed significantly reduced Jonhsen's scores in varicocele compared to sham and varicocelectomy groups (p < 0.05). Expression of autophagy markers (LC3A, Beclin-1, Ambra-1, ULK-1, p62) was significantly higher in varicocele than sham and varicocelectomy groups (p < 0.05), and in varicocelectomy than sham (p < 0.05). Varicocele activates autophagy markers, with p62 potentially modulating autophagy despite being considered an inhibitor. While varicocelectomy improves histology, it doesn't fully inhibit autophagy, suggesting ongoing germ cell dysfunction despite treatment. This underscores varicocele's detrimental effects on germ cell functionality.

Momordicine-I suppresses head and neck cancer growth by modulating key metabolic pathways

One of the hallmarks of cancer is metabolic reprogramming which controls cellular homeostasis and therapy resistance. Here, we investigated the effect of momordicine-I (M-I), a key bioactive compound from Momordica charantia (bitter melon), on metabolic pathways in human head and neck cancer (HNC) cells and a mouse HNC tumorigenicity model. We found that M-I treatment on HNC cells significantly reduced the expression of key glycolytic molecules, SLC2A1 (GLUT-1), HK1, PFKP, PDK3, PKM, and LDHA at the mRNA and protein levels. We further observed reduced lactate accumulation, suggesting glycolysis was perturbed in M-I treated HNC cells. Metabolomic analyses confirmed a marked reduction in glycolytic and TCA cycle metabolites in M-I-treated cells. M-I treatment significantly downregulated mRNA and protein expression of essential enzymes involved in de novo lipogenesis, including ACLY, ACC1, FASN, SREBP1, and SCD1. Using shotgun lipidomics, we found a significant increase in lysophosphatidylcholine and phosphatidylcholine loss in M-I treated cells. Subsequently, we observed dysregulation of mitochondrial membrane potential and significant reduction of mitochondrial oxygen consumption after M-I treatment. We further observed M-I treatment induced autophagy, activated AMPK and inhibited mTOR and Akt signaling pathways and leading to apoptosis. However, blocking autophagy did not rescue the M-I-mediated alterations in lipogenesis, suggesting an independent mechanism of action. M-I treated mouse HNC MOC2 cell tumors displayed reduced Hk1, Pdk3, Fasn, and Acly expression. In conclusion, our study revealed that M-I inhibits glycolysis, lipid metabolism, induces autophagy in HNC cells and reduces tumor volume in mice. Therefore, M-I-mediated metabolic reprogramming of HNC has the potential for important therapeutic implications.Graphical

Unveiling Key Genes Modulating Retinal Cell Survival and Autophagy in Glaucoma.

Glaucoma is a leading cause of irreversible blindness, with rising incidence globally. Effective treatment is challenging due to limited understanding of the disease mechanisms. Growth factor activity is crucial in glaucoma, with potential to reduce retinal ganglion cell (RGC) apoptosis and slow disease progression. This study aims to identify and analyze differentially expressed genes (DEGs) involved in growth factor activity to uncover new therapeutic targets. We analyzed the GSE9944 dataset from the Gene Expression Omnibus (GEO) to identify DEGs associated with glaucoma, resulting in 94 DEGs, including 29 down-regulated and 65 up-regulated genes. Functional enrichment and protein-protein interaction (PPI) network analyses were conducted using bioinformatics tools, highlighting the roles of Bone Morphogenetic Protein 1 (BMP1), Pleiotrophin (PTN), and f fibroblast Growth Factor 7 (FGF7). Aberrant expression vectors for these genes were transfected into RGCs derived from a glaucoma model to evaluate their impact on cell viability, apoptosis, and autophagy. Bioinformatics analysis of the GSE9944 dataset identified 94 DEGs, with 29 down-regulated and 65 up-regulated genes. Functional enrichment analysis revealed that these DEGs were involved in pathways related to growth factor activity, apoptosis, and autophagy, processes highly relevant to glaucoma pathogenesis. PPI network analysis identified BMP1, PTN, and FGF7 as central hub genes involved in extracellular matrix organization and growth factor signaling. In experimental validation using RGCs, we found that up-regulation of BMP1 significantly enhanced RGC viability and reduced apoptosis. Conversely, silencing PTN and FGF7 provided protective effects, enhancing RGC survival. Silencing BMP1 and upregulating PTN and FGF7 led to increased RGC apoptosis. Additionally, BMP1 was found to inhibit autophagy in RGCs, whereas PTN and FGF7 promoted autophagic activity, suggesting differential regulatory roles in glaucoma pathogenesis. Overall, BMP1, PTN, and FGF7 play critical roles in the regulation of RGC activity and autophagy in glaucoma, making them promising molecular targets for future therapeutic interventions.

Sulforaphane and Benzyl Isothiocyanate Suppress Cell Proliferation and Trigger Cell Cycle Arrest, Autophagy, and Apoptosis in Human AML Cell Line

Isothiocyanates (ITCs) are naturally occurring sulfur-containing compounds with diverse biological effects. This study investigated the effects of sulforaphane (SFN, an aliphatic ITC) and benzyl isothiocyanate (BITC, an aromatic ITC) on human acute myeloid leukemia SKM-1 cells, focusing on cell proliferation, cell death, and drug resistance. Both drug-sensitive SKM-1 cells and their drug-resistant SKM/VCR variant, which overexpresses the drug transporter P-glycoprotein, were used. SFN and BITC reduced cell viability in a dose-dependent manner, with BITC showing greater potency. IC50 values ranged from 7.0–8.0 µM for SFN and 4.0–5.0 µM for BITC in both cell types, with only slight differences between the variants. Both ITCs induced autophagy as evidenced by increased LC3-II production and caused a significant increase in the sub-G0/G1 cell population, especially with BITC. Apoptosis was more pronounced after BITC treatment, whereas SFN had a weaker effect. These results suggest that autophagy may act as a defense mechanism in response to ITC-induced apoptosis in human AML cells.

Isothiocyanates induce autophagy and inhibit protein synthesis in primary cells via modulation of AMPK-mTORC1-S6K1 signaling pathway, and protect against mutant huntingtin aggregation

PurposeAutophagy is a degradation process whose activation underlies beneficial effects of caloric restriction. Isothiocyanates (ITCs) induce autophagy in cancer cells, however, their impact on primary cells remains insufficiently explored, particularly in non-epithelial cells. The aim of this study was to investigate whether ITCs induce autophagy in primary (non-immortalized) mesenchymal cells and if so, to determine the molecular mechanism underlying its activation and consequences on cell functioning.MethodsPrimary human dermal fibroblasts (HDFa) and prostate cancer cells (PC3) as well as two ITCs, sulforaphane and phenethyl isothiocyanate, were applied. Cell viability was measured by the MTT test, protein synthesis - by 3H-leucine incorporation, and protein level - by immunoblotting. A number of mutant huntingtin (mHtt) aggregates was assessed by fluorescence microscopy.ResultsBoth ITCs efficiently induced autophagy in fibroblasts which coincided with suppression of mTORC1 – a negative autophagy regulator - and protein synthesis arrest. A dephosphorylation of mTORC1 substrate, S6K1, and ribosomal S6 protein was preceded by activation of AMPK, an inhibitor of mTORC1 and autophagy activator. A similar response was observed in phenethyl isothiocyanate-treated prostate cancer cells. We also showed that ITCs-induced autophagy and/or translation block do not affect cells viability and can protect cells against an accumulation of mHtt aggregates – a main cause of Huntington’s disease.ConclusionOur study showed that ITCs induce autophagy and inhibit protein synthesis in both primary mesenchymal and cancer cells via modulation of the AMPK-mTORC1-S6K1 pathway. Moreover, it suggests that ITCs might have a potential in developing therapeutics for Huntington’s disease.

Enhancing the radiosensitivity of colorectal cancer cells by reducing spermine synthase through promoting autophagy and DNA damage

BACKGROUND Colorectal cancer (CRC), the third most common cancer worldwide, has increasingly detrimental effects on human health. Radiotherapy resistance diminishes treatment efficacy. Studies suggest that spermine synthase (SMS) may serve as a potential target to enhance the radiosensitivity. AIM To investigate the association between SMS and radiosensitivity in CRC cells, along with a detailed elucidation of the underlying mechanisms. METHODS Western blot was adopted to assess SMS expression in normal colonic epithelial cells and CRC cell lines. HCT116 cells were transfected with control/SMS-specific shRNA or control/pcDNA3.1-SMS plasmids. Assessments included cell viability, colony formation, and apoptosis via MTT assays, colony formation assays, and flow cytometry. Radiosensitivity was studied in SMS-specific shRNA-transfected HCT116 cells post-4 Gy radiation, evaluating cell viability, colony formation, apoptosis, DNA damage (comet assays), autophagy (immunofluorescence), and mammalian target of rapamycin (mTOR) pathway protein expression (western blot). RESULTS Significant up-regulation of SMS expression levels was observed in the CRC cell lines. Upon down-regulation of SMS expression, cellular viability and colony-forming ability were markedly suppressed, concomitant with a notable increase in apoptotic indices. Furthermore, attenuation of SMS expression significantly augmented the sensitivity of HCT116 cells to radiation therapy, evidenced by a pronounced elevation in levels of cellular DNA damage and autophagy. Importantly, down-regulation of SMS corresponded with a marked reduction in the expression levels of proteins associated with the mTOR signaling pathway. CONCLUSION Knocking down SMS attenuates the mTOR signaling pathway, thereby promoting cellular autophagy and DNA damage to enhance the radiosensitivity of CRC cells.

MARCH5 Promotes the Progression of Thyroid cancer by Regulating Mitochondrial Autophagy Protein FUNDC1-mediated Pyroptosis.

MARCH5 is a key regulatory factor in mitochondria. However, the expression and function of MARCH5 in thyroid cancer (TC) are not yet clear. The research explores the role and the potential mechanism of MARCH5 in the tumorigenesis of TC. MARCH5 expression were measured by qRT-PCR and Western blot. CCK-8 kits were used to measure the cell viability. Cell scratch assay and Tanswell assay were used to measure cell migration and invasion, respectively. The pyroptosis related proteins (NLRP3, caspase-1, GSDMD) and mitochondrial autophagy related proteins (LC3-II, p62, parkin, pink1) were detected. The mitochondrial ROS GSH, MDA, and SOD were detected using commercial kits. Finally, a TC mouse model was constructed to detect the role of MARCH5 in tumor growth in vivo. The results displayed that the expression of MARCH5 was increased in TC patients and cells, and was significantly correlated with prognosis. Functional studies have found that MARCH5 inhibits oxidative stress levels and mitochondrial autophagy in TPC-1 cells. Further research has found that MARCH5 promotes the progression of thyroid cancer by degrading FUNDC1 and inhibiting the mitochondrial autophagy mediated pyroptosis pathway, regulating cell proliferation, migration, and invasion in TPC-1 cells. More importantly, interference with MARCH5 inhibits tumor growth and further development of TC in vivo. In conclusion, MARCH5 promotes the progression of thyroid cancer by degrading FUNDC1 and inhibiting the mitochondrial autophagy mediated pyroptosis, regulating cell proliferation, migration, and invasion. This study provides new theoretical basis for the treatment and prevention of TC in clinical practice.

Canagliflozin differentially modulates carfilzomib-induced endoplasmic reticulum stress in multiple myeloma and endothelial cells.

Carfilzomib (CFZ), a second-generation proteasome inhibitor, is a key treatment for multiple myeloma (MM), but its use is associated with significant cardiovascular adverse events (CVAEs), including heart failure and hypertension. Endothelial dysfunction is believed to contribute to these CVAEs. Building on our previous findings that CFZ induces endothelial toxicity and that canagliflozin protects against CFZ-induced endothelial apoptosis, this study aimed to evaluate CFZ-induced endoplasmic reticulum (ER) stress and autophagy in endothelial and MM cells, as well as the impact of canagliflozin on these processes and its impact on the anticancer effects of CFZ in MM cells. Endothelial cells (HUVECs and EA.hy926) and multiple myeloma cells (RPMI8226) were treated with 0.5µM CFZ, either alone or in combination with canagliflozin (5-20µM), to assess the effects on ER stress and autophagy in both cell types. CFZ induced ER stress in endothelial and MM cells. In endothelial cells, canagliflozin mitigated CFZ-induced markers of ER stress, while unexpectedly upregulating CFZ-induced CHOP. Whereas, in MM cells, canagliflozin did not alter CFZ-induced ER stress, but instead further upregulated CFZ-induced ATF-4. In addition, CFZ induced autophagy in endothelial cells while inhibiting it in MM cells. Canagliflozin abrogated CFZ-induced autophagy in endothelial cells. In striking contrast to its effects in endothelial cells, canagliflozin enhanced the cytotoxic effects of CFZ in MM cells. Intriguingly, in an innovative co-culture system, canagliflozin enhanced CFZ-induced apoptosis in MM cells while protecting endothelial cells. These findings underscore the dual role of canagliflozin in reducing CFZ-induced endothelial toxicity, while enhancing its cytotoxic effect in MM.

Imperatorin promotes melanin degradation in keratinocytes through facilitating autophagy via the PI3K/Akt signaling pathway.

Melanin's pivotal role in skin protection and its overproduction leading to hyperpigmentation disorders highlight the necessity of regulating melanogenesis, with autophagy identified as a key degradation pathway. Imperatorin, a compound from Angelica dahurica, has been revealed to reduce melanin in epidermal keratinocytes, with the specific effects and mechanisms unknown. The purpose of this study was to investigate the mechanism by which imperatorin, reduces melanin production in HaCaT cells, with a focus on its potential role in promoting autophagy and regulating the PI3K/Akt signaling pathway. The study used HaCaT cells to investigate the effects of imperatorin on melanin production, autophagy, and PI3K/Akt signaling. Melanin content was measured using a spectrophotometric method. Protein levels of PMEL, ATG1, ATG5, and LC3B II were assessed by Western blotting. Autophagy was further visualized by GFP-LC3B puncta formation. The autophagy inhibitor 3-MA, the PI3K/Akt inhibitor LY294002 and PI3K/Akt activator 740 Y-P were used to assess the role of autophagy and PI3K/Akt signaling in imperatorin's effects. Cell viability was monitored to ensure that imperatorin's effects were not due to cytotoxicity. Imperatorin reduced melanin content in HaCaT cells in a dose-dependent manner without compromising cell viability. This reduction in melanin was accompanied by decreased levels of PMEL protein, a key player in melanosome formation. Additionally, imperatorin promoted autophagy in HaCaT cells, as evidenced by increased levels of autophagy-associated markers ATG1, ATG5, and LC3B II, as well as an increase in GFP-LC3B puncta. The autophagy inhibitor 3-MA partially reversed the effects of imperatorin on both autophagy markers and PMEL levels, indicating that autophagy plays a crucial role in imperatorin's depigmentation action. Furthermore, imperatorin inhibited Akt and mTOR phosphorylation, which are downstream targets of PI3K/Akt signaling, enhancing autophagy and further reducing melanin levels. The PI3K/Akt inhibitor LY294002 amplified imperatorin's effects on PI3K and Akt phosphorylation, autophagy, and melanin levels. While, PI3K/Akt activator 740 Y-P reversed imperatorin's effects on these factors. Imperatorin reduces melanin in HaCaT cells via promoting autophagy and melanin degradation, possibly via the PI3K/Akt signaling. Taken together, imperatorin has the therapeutic potential for the treatment of hyperpigmentation disorders.

Porphyromonas gingivalis inducing autophagy-related biological dysfunction in alveolar epithelial cells: an in vitro study

BackgroundChronic obstructive pulmonary disease (COPD) is a respiratory disease with high morbidity and mortality. Notably, the pathogenesis and progression of COPD are related to lung infection, inflammatory response, and biological dysfunction in alveolar epithelial cells. Studies also found that periodontitis is an independent risk factor for COPD. The inhalation of periodontal pathogens into the respiratory system is the most common method for periodontal pathogens to promote the development of COPD. Porphyromonas gingivalis (P. gingivalis), the keystone pathogen in periodontitis, has been found to migrate to the lungs, triggering inflammatory reactions and causing decreased lung function. However, the impact of P. gingivalis infection on the biological function of alveolar epithelial cells remains unclear. Therefore, this study aimed to investigate the effects of P. gingivalis infection on the biological functions of alveolar epithelial cells.MethodsMouse alveolar epithelial cells (MLE-12) were co-cultured with P. gingivalis and treated with autophagy inhibitor chloroquine (CQ) or LC3 siRNA in vitro. MTT assay and EdU staining were used to detect cell viability, and the TUNEL assay kit and Annexin V-FITC/PI method were used to detect cell apoptosis. Western blot was used to detect autophagic markers LC3 and P62, and mRFP-GFP-LC3 was used to observe autophagic flux.ResultsP. gingivalis inhibited the viability of alveolar epithelial cells in a dose- and time-dependent manner. P. gingivalis also promoted autophagy and apoptosis of alveolar epithelial cells in a dose-dependent manner. Interestingly, we found that inhibiting autophagy using CQ or silencing LC3 with siRNA significantly reduced cell apoptosis and viability damage induced by P. gingivalis. Thus, these data indicated the synergistic effect of autophagy in P. gingivalis-induced biological dysfunction of alveolar epithelial cells.ConclusionP. gingivalis infection can cause biological dysfunction of alveolar epithelial cells, manifested as decreased cell viability, increased autophagy and apoptosis. Notably, the up-regulation of autophagy induced by P. gingivalis plays a synergistic role in this dysfunction.

OCTN2 expression and function in the Sertoli cells of testes from patients with non-obstructive azoospermia.

Among couples, male factors account for approximately 50% of infertility cases, with nonobstructive azoospermia (NOA) representing one of the most clinically common and severe categories of male infertility, affecting approximately 10-15% of patients. Currently, L-carnitine is clinically used to improve spermatogenesis by regulating Sertoli cell function. Multiple clinical trials have described the efficacy of L-carnitine in treating NOA. Notably, Sertoli cells rely on organic carnitine transporter 2 (OCTN2) for carnitine transport. However, it remains unknown whether OCTN2 expression is involved in the pathological process of NOA. To investigate the expression and function of OCTN2 in Sertoli cells from patients with NOA. Ten testicular tissue samples were collected, including five from a healthy group and five from a group of patients with NOA. Immunohistochemistry and immunofluorescence were used to detect the expression of OCTN2 in testicular tissue. Additionally, an Octn2-KO TM4 cell line (a mouse testicular Sertoli cell line) was constructed to explore the function of OCTN2 expression in Sertoli cells through transcriptomic sequencing, cell proliferation experiments, metabolomic analysis, and Western blot analysis. Compared with those of the healthy group, the immunohistochemistry results revealed a significant decrease in OCTN2 expression in the Sertoli cells of the NOA group. Further investigation through cell proliferation experiments revealed a reduction in the proliferative capacity of the Octn2-KO TM4 cell line. Transcriptomic sequencing and metabolomic data analysis revealed a decrease in autophagy in the Octn2-KO TM4 cell line. Western blot analysis subsequently verified the expression levels of autophagy-related proteins. In the Sertoli cells of NOA patients, decreased OCTN2 protein expression leads to decreased cell proliferation and autophagy abnormalities, which may play a crucial role in the spermatogenic dysfunction observed in NOA patients.

TMEM16A Activation Inhibits Autophagy in Dorsal Root Ganglion Cells, Which is Associated with the p38 MAPK/mTOR Pathway

Transmembrane member 16A (TMEM16A) exhibits a negative correlation with autophagy, though the underlying mechanism remains elusive. This study investigates the mechanism between TMEM16A and autophagy by inducing autophagy in DRG neuronal cells using Rapamycin. Results indicated that TMEM16A interference augmented cell viability and reduced Rapamycin-induced apoptosis. Autophagosome formation increased with TMEM16A interference but decreased upon overexpression. A similar increase in autophagosomes was observed with SB203580 treatment. Furthermore, TMEM16A interference suppressed Rapamycin-induced gene and protein expression of p38 MAPK and mTOR, whereas overexpression had the opposite effect. These findings suggest that TMEM16A activation inhibits autophagy in DRG cells, which is associated with the p38 MAPK/mTOR pathway, offering a potential target for mitigating neuropathic pain (NP).Graphical abstract

Metabolism: a potential regulator of neutrophil fate.

Neutrophils are essential components of the innate immune system that defend against the invading pathogens, such as bacteria, viruses, and fungi, as well as having regulatory roles in various conditions, including tissue repair, cancer immunity, and inflammation modulation. The function of neutrophils is strongly related to their mode of cell death, as different types of cell death involve various cellular and molecular alterations. Apoptosis, a non-inflammatory and programmed type of cell death, is the most common in neutrophils, while other modes of cell death, including NETOsis, necrosis, necroptosis, autophagy, pyroptosis, and ferroptosis, have specific roles in neutrophil function regulation. Immunometabolism refers to energy and substance metabolism in immune cells, and profoundly influences immune cell fate and immune system function. Intercellular and intracellular signal transduction modulate neutrophil metabolism, which can, in turn, alter their activities by influencing various cell signaling pathways. In this review, we compile an extensive body of evidence demonstrating the role of neutrophil metabolism in their various forms of cell death. The review highlights the intricate metabolic characteristics of neutrophils and their interplay with various types of cell death.

Nanoformula Design for Inducing Non-Apoptotic Cell Death Regulation: A Powerful Booster for Cancer Immunotherapy.

Cancer treatment has witnessed revolutionary advancements marked by the emergence of immunotherapy, specifically immune checkpoint blockade (ICB). However, the inherent low immunogenicity of tumor cells and the intricate immunosuppressive network within the tumor microenvironment (TME) pose significant challenges to the further development of immunotherapy. Nanotechnology has ushered in unprecedented opportunities and vast prospects for tumor immunotherapy. Nevertheless, traditional nano-formulations often rely on inducing apoptosis to kill cancer cells, which encounters the issue of immune silencing, hindering effective tumor immune activation. The non-apoptotic modes of regulated cell death (RCD), including pyroptosis, ferroptosis, autophagy, necroptosis, and cuproptosis, have gradually garnered attention. These non-apoptotic cell death pathways can induce effective immunogenic cell death (ICD), enhancing cancer immunotherapy. This review comprehensively explores advanced nano-formulation design strategies and their applications in enhancing cancer immunotherapy by promoting non-apoptotic RCD in recent years. It also discusses the potential advantages of these strategies in inducing tumor-specific non-apoptotic RCD. By deeply understanding the significance of non-apoptotic RCD in synergistic cancer immunotherapy, this article provides valuable insights for developing more advanced nano-delivery systems that can robustly induce highly immunogenic non-apoptotic modes, offering novel research and development avenues to address the clinical challenges encountered by immunotherapy represented by ICB.

ULK1 methylation promotes TGF-β1-induced endometrial fibrosis via the FOXP1/DNMT1 axis.

Intrauterine adhesion (IUA) is the second most common cause of secondary infertility in women and can also lead to menstrual abnormalities and multiple adverse pregnancy outcomes. Therefore, elucidating the mechanism of its development is crucial for the prevention and treatment of IUA. This study will investigate the function and mechanism of forkhead box P1 (FOXP1)/DNA methyltransferase 1 (DNMT1)/unc-51-like autophagy activating kinase 1 (ULK1) in IUA. Expression levels of key genes were detected using western blot and quantitative - real time reverse transcription polymerase chain reaction. Cell proliferation was detected by CCK-8 and EdU staining. Transcriptional regulation relationships were detected by dual luciferase reporter gene and chromatin immunoprecipitation (ChIP) assay. Methylation station of ULK1 was detected by methylmion specific PCR (MSP). Fibrosis and pathological changes in the uterine cavity tissues were detected by Masson and hematoxylin and eosin staining. It was observed that the expression of FOXP1 and DNMT1 increased in transforming growth factor (TGF)-β1-induced cells, while ULK1 expression decreased. Downregulation of FOXP1 could inhibit human endometrial stromal cells proliferation and autophagy, as well as decrease the expression of fibrogenic factors (collagen type I alpha 1 chain [COL1A1], fibronectin [FN], and alpha-smooth muscle actin [α-SMA]). The results of MSP and ChIP experiments showed that DNMT1 promotes methylation of the ULK1 promoter region and inhibits its transcription. In an animal model, knockdown of FOXP1 alleviated pathological fibrosis and uterine adhesions. Knockdown of FOXP1 can inhibit endometrial fibrosis in IUA rats; FOXP1 could be a potential target for the treatment of IUA.

Human adenovirus type 7 (HAdV-7) infection induces pulmonary vascular endothelial injury through the activation of endothelial autophagy

BackgroundHAdV-7 is a prevalent pathogen that can cause severe pneumonia in children. Previous studies have shown a significant increase in serum levels of vascular permeability factor (VPF/VEGF) and viral load in pediatric patients with fatal HAdV-7 infection, suggesting potential damage to the pulmonary vascular endothelium. Further research is necessary to elucidate the underlying mechanism.MethodsThe human lung microvascular endothelial cell line-5a and human CD46 mice were used for in vitro and in vivo experiments, respectively. RNA-seq was employed for correlative omics analysis. Viral infection and copy status were examined using transmission electron microscopy to observe virus particles, immunofluorescence to detect the viral protein Hexon, and qPCR to assess HAdV-7 fiber gene copies. Various methods, including ELISAs for VEGF and other injury markers, the CCK8 assay for cell viability, and flow cytometry for endothelium numbers, were employed to evaluate endothelial damage. Acute lung injury severity was evaluated by scoring pathological inflammation and measuring pulmonary vascular permeability. Autophagy activation was assessed by observing autophagosomes and validating marker proteins.ResultsGSEA analysis showed significant enrichment of gene sets related to endothelial functions (barrier, defense, and regeneration) and ALI in the HAdV-7-infected group. GO analysis indicated an enrichment of autophagy-related pathways linked to cell death. Subsequently, successful signs of HAdV-7 infection and replication were observed in the endothelium, including cytopathic effects, intracellular virions, and increased HAdV-7 fiber gene copies. Endothelial injury, including mitochondrial damage, decreased endothelium, and elevated levels of endothelial injury markers such as VEGF, sICAM-1, sVCAM-1, E-selectin, ESM1, MCP1, and IL1β were observed after HAdV-7 infection. Additionally, evidence of leaky lung blood vessels and ALI was observed, including progressive weight loss, elevated pulmonary vascular permeability, and severe lung consolidation. Furthermore, HAdV-7 infection induced autophagosome formation in the endothelium and triggered complete cell autophagy. Importantly, inhibiting autophagic flux reduced VEGF levels and other endothelial injury markers, decreased viral load, improved cell survival rate, alleviated pulmonary vessel leakage, and mitigated lung inflammation.ConclusionsHAdV-7 successfully infects pulmonary vascular endothelium and replicates effectively, causing injury to the endothelium, high VEGF expression and viral load in the serum, as well as ALI/ARDS. Autophagy inhibitors can alleviate endothelial injury, inhibit viral replication, relieve leakage from the vasculature, and reduce lung inflammation.

Effects of autophagy on the selective death of human breast cancer cells exposed to plasma-activated Ringer’s lactate solution

Plasma-activated Ringer’s lactate (PAL) solution prepared by irradiating an intravenous solution with a non-equilibrium atmospheric pressure plasma is a potential new cancer therapy having no side effects. However, the induction of autophagy to avoid cell death has been confirmed to occur following exposure to PAL solution. It is thought that the antitumor effect of PAL solution could be weakened by this process, which is meant to maintain homeostasis in cells and assists tumorigenesis. Thus, it would be helpful to devise PAL-based cancer therapies that inhibit autophagy. Unfortunately, it is not yet clear which substances in PAL solution promote autophagy. The present work examined the mechanism by which PAL solution induces autophagy when treating MCF-7 human breast cancer cells. Autophagy was found to be temporarily induced upon exposure to PAL solution, suggesting that this effect contributes to cell proliferation. Although autophagy is associated with reactive oxygen and nitrogen species and/or acidic environments, in this study, significant autophagy was observed using a PAL solution diluted 1/256x without these stressors. Acetate, glyoxylate and 2,3-dimethyltartrate in the PAL solution were determined to promote autophagy. Interestingly, 2,3-dimethyltartrate was found to either induce cell death or autophagy depending on the concentration.

A novel antifungal peptide, SP1.2, from Rhodopseudomonas palustris against the rice blast pathogen.

Rice blast has a significant detrimental impact on rice yields, so developing efficient biological control technologies is an effective means for rice blast prevention and control. The GroEL protein has proven to be effective at preventing and managing the pathogenicity of rice blast. Here, we analyzed the amino acid sequence of the GroEL protein and synthesized the '60 kDa chaperonin signature' (350-373 amino acids) peptide SP1.2, which has potent antifungal activity. Notably, the SP1.2 peptide exhibited potent fungicidal activity against Magnaporthe oryzae, effectively inhibiting appressorium germination. Electron microscopy revealed that SP1.2 disrupted the fungal plasma membrane and bound to multiple bioactive phosphoinositides in vitro, triggering the production of reactive oxygen species. Furthermore, it also caused an increase in the acetylation of M. oryzae and induced autophagy in cells. The spray application of SP1.2 significantly reduced the number of disease spots caused by the fungal pathogen M. oryzae in rice, enhancing the defense response of rice plants. Field trials showed that the control effect was 64.59% after spraying SP1.2. Our study illustrates the antifungal activity of the structurally unique SP1.2 peptide against plant fungal pathogens and paves the way for the future development of this class of peptides as antifungal agents. © 2024 Society of Chemical Industry.