Reactive oxygen species regulation by NCF1 governs ferroptosis susceptibility of Kupffer cells to MASH.

Impaired self-renewal of Kupffer cells (KCs) leads to inflammation in metabolic dysfunction-associated steatohepatitis (MASH). Here, we identify neutrophil cytosolic factor 1 (NCF1) as a critical regulator of iron homeostasis in KCs. NCF1 is upregulated in liver macrophages and dendritic cells in humans with metabolic dysfunction-associated steatotic liver disease and in MASH mice. Macrophage NCF1, but not dendritic cell NCF1, triggers KC iron overload, ferroptosis, and monocyte-derived macrophage infiltration, thus aggravating MASH progression. Mechanistically, elevated oxidized phospholipids induced by macrophage NCF1 promote Toll-like receptor (TLR4)-dependent hepatocyte hepcidin production, leading to increased KC iron deposition and subsequent KC ferroptosis. Importantly, the human low-functional polymorphic variant NCF190H alleviates KC ferroptosis and MASH in mice. In conclusion, macrophage NCF1 impairs iron homeostasis in KCs by oxidizing phospholipids, triggering hepatocyte hepcidin release and KC ferroptosis in MASH, highlighting NCF1 as a therapeutic target for improving KC fate and limiting MASH progression.

Autoantibodies against Endophilin A2 as a novel biomarker are beneficial to early diagnosis of breast cancer.

BACKGROUND: Due to the lack of early symptoms, breast cancer is frequently overlooked, leading to distant metastases and multi-organ lesions that directly threaten patients' lives. We have identified a novel tumor marker, antibodies to endophilin A2 (EA2), to improve early diagnosis of breast cancer. METHODS: Antibody levels of EA2 were analyzed in sera of patients with cancers of different origins and stages by indirect enzyme-linked immunosorbent assay (ELISA). Diagnostic accuracy and reference range were determined by the area under the receiver operating curve and distribution curve. The levels of EA2 antigen in sera were determined by sandwich ELISA. RESULTS: The levels of antibodies against EA2 were higher in sera of patients with breast cancer (P < 0.0001), liver cancer (P = 0.0005), gastric cancer (P = 0.0026), and colon cancer (P = 0.0349) than those in healthy controls, but not in patients with rectal cancer (P = 0.1151), leukemia (P = 0.7508), or lung cancer (P = 0.2247). The highest diagnostic value was for breast cancer, particularly in early cases (AUC = 0.8014) and those with distant metastases (AUC = 0.7885). The titers of EA2 antibodies in sera were correlated with levels of EA2 antigen in breast cancer patients. CONCLUSION: Antibodies to EA2 are novel blood biomarkers for early diagnosis of breast cancer that warrants further study in larger-scale cohort studies.

Pt3R5G inhibits colon cancer cell proliferation through inducing ferroptosis by down-regulating SLC7A11.

AIMS: Colon cancer (CC) is a prevalent malignancy worldwide and is one of the most easily altered cancers by dietary regulation. Petunidin 3-O-[rhamnopyranosyl-(trans-p-coumaroyl)]-5-O-(ß-D-glucopyranoside) (Pt3R5G) isolated and purified from Lycium ruthenicum Murray, which exhibits highly efficient antioxidant activity and specific anticancer effects, is the flavonoids compound. We aimed to study the effect of Pt3R5G on CC cells and elucidate the potential underlying mechanisms. MAIN METHODS: Cell proliferation was measured by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and colony formation assays. Cell cycle, cell apoptosis and reactive oxygen species (ROS) analysis were performed by flow cytometry. RNA-sequencing was performed to elucidate the potential underlying mechanisms. The lipid peroxidation level of cells was detected by malondialdehyde (MDA) assay. The mitochondrial morphology of cells was inspected using a transmission electron microscope. Additionally, we overexpressed SLC7A11 to perform rescue experiments. In vivo, xenograft mice assay was performed to verify the effect of Pt3R5G on the growth of colon cancer. KEY FINDINGS: Pt3R5G reduced the cell activity by blocking the cell cycle in G0/G1 phase, inducing the apoptosis and ferroptosis in RKO cells. The overexpressed of SLC7A11, a significantly down-regulated expression gene caused by Pt3R5G, rescued the cell proliferation inhibition and ferroptosis process. Furthermore, Pt3R5G inhibited tumor growth in nude mice. Our study suggests that Pt3R5G inhibits RKO cell proliferation through mainly reducing ferroptosis by down-regulated SLC7A11. SIGNIFICANCE: As a potential therapeutic drug, Pt3R5G showed efficient anticancer activity through a variety of pathways.

SERPINA5 promotes tumour cell proliferation by modulating the PI3K/AKT/mTOR signalling pathway in gastric cancer.

SERPINA5 belongs to the serine protease inhibitor superfamily and has been reported to be lowly expressed in a variety of malignancies. However, few report of SERPINA5 in gastric cancer has been found. The purpose of this study was to determine the role of SERPINA5 in GC and to investigate potential tumorigenic mechanisms. We performed qPCR to determine the level of SERPINA5 expression in GC. We used public databases to evaluate whether SERPINA5 could be utilized to predict overall survival and disease-free survival in GC patients. We also knocked down the expression of SERPINA5 and evaluated its effect on cell proliferation and migration. Furthermore, we explored the signal pathways and regulatory mechanisms related to SERPINA5 functions. According to our findings, SERPINA5 was shown to exhibit high expression in GC. Notably, SERPINA5 was prognostic in GC with high expression being unfavourable. SERPINA5 was further observed to promote GC tumorigenesis by modulating GC cell proliferation ability. Mechanically, SERPINA5 could inhibit CBL to regulate the PI3K/AKT/mTOR signalling pathway, thereby promoting GC carcinogenesis progression. These results highlight the important role of SERPINA5 in GC cell proliferation and suggest that SERPINA5 could be a novel target for GC treatment and a predictor for GC prognosis.

Emerging Roles of T Helper Cells in Non-Infectious Neuroinflammation: Savior or Sinner.

CD4+ T cells, also known as T helper (Th) cells, contribute to the adaptive immunity both in the periphery and in the central nervous system (CNS). At least seven subsets of Th cells along with their signature cytokines have been identified nowadays. Neuroinflammation denotes the brain's immune response to inflammatory conditions. In recent years, various CNS disorders have been related to the dysregulation of adaptive immunity, especially the process concerning Th cells and their cytokines. However, as the functions of Th cells are being discovered, it's also found that their roles in different neuroinflammatory conditions, or even the participation of a specific Th subset in one CNS disorder may differ, and sometimes contrast. Based on those recent and contradictory evidence, the conflicting roles of Th cells in multiple sclerosis, Alzheimer's disease, Parkinson's disease, epilepsy, traumatic brain injury as well as some typical mental disorders will be reviewed herein. Research progress, limitations and novel approaches concerning different neuroinflammatory conditions will also be mentioned and compared.

LGR6 Acts as an Oncogene and Induces Proliferation and Migration of Gastric Cancer Cells.

Leucine rich repeat containing G protein-coupled receptor 6 (LGR6) belongs to the G protein-coupled receptor family, and it exhibits up-regulated expression in various types of human cancer. However, there are few reports of LGR6 contributing to gastric cancer (GC). Herein, we investigated the function of LGR6 and associated tumorigenic mechanisms in GC. LGR6 expression in GC was analyzed in the cancer genome atlas (TCGA) dataset and further confirmed in GC cell lines and fifteen paired tissue samples via quantitative real-time polymerase chain reaction (qRT-PCR). LGR6 expression was knocked down via small interfering RNA (siRNA), after which the impacts of silencing LGR6 on cell function were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT), cell colony formation, wound-healing, and cell cycle assays. Western blot was performed to explore signaling pathways and regulatory mechanisms associated with LGR6 function. In this study, we showed that LGR6 was at higher levels in GC cell lines and gastric adenocarcinoma tissues. We found that silencing LGR6 in MKN-45 and BGC-823 cells inhibited cell proliferation and migration ability, which accompanied with an obvious regulation of MMP-9, ß-catenin, CCNA2, CDK-2, and ERK1/2. In conclusion, this study demonstrated that LGR6 could act as an oncogene and may be a therapeutic target in GC.

Chronic stress inhibits testosterone synthesis in Leydig cells through mitochondrial damage via Atp5a1.

Stress is one of the leading causes of male infertility, but its exact function in testosterone synthesis has scarcely been reported. We found that adult male rats show a decrease in bodyweight, genital index and serum testosterone level after continual chronic stress for 21 days. Two-dimensional gel electrophoresis (2-DE) and MALDI-TOF-MS analysis identified 10 differentially expressed proteins in stressed rats compared with controls. A strong protein interaction network was found to be centred on Atp5a1 among these proteins. Atp5a1 expression significantly decreased in Leydig cells after chronic stress. Transfection of Atp5a1 siRNAs decreased StAR, CYP11A1, and 17ß-HSD expression by damaging the structure of mitochondria in TM3 cells. This study confirmed that chronic stress plays an important role in testosterone synthesis by regulating Atp5a1 expression in Leydig cells.

Effects of treadmill exercise on anxiety-like behavior in association with changes in estrogen receptors ERα, ERß and oxytocin of C57BL/6J female mice.

Exercise can reduce the incidence of stress-related mental diseases, such as depression and anxiety. Control group was neither exposed to CVMS nor TRE (noCVMS/noTRE). Females were tested and levels of serum17-beta-oestradiol (E2), estrogen receptors α immunoreactive neurons (ERα-IRs), estrogen receptors ß immunoreactive neurons (ERß-IRs) and oxytocin immunoreactive neurons (OT-IRs) were measured. The results showed there's increased anxiety-like behaviors for mice from CVMS/noTRE, CVMS/higher speed TRE (CVMS/HTRE) and noCVMS/HTRE groups when they were put in open field and elevated maze tests. They had lower serum E2 levels than mice from CVMS/low-moderate speed TRE (CVMS/LMTRE), noCVMS/LMTRE and noCVMS/noTRE groups. The three groups of CVMS/noTRE, CVMS/HTRE and noCVMS/HTRE mice had more ERα-IRs and less ERß-IRs in the medial preoptic area (mPOA), bed nucleus of the stria terminalis (BNST) and medial amygdala (MeA), hypothalamic paraventricular nucleus (PVN) and supraoptic nucleus (SON). The number of OT-IRs in PVN and SON of CVMS/noTRE, CVMS/HTRE and noCVMS/HTRE mice was also lower than that of mice from CVMS/LMTRE, noCVMS/LMTRE and noCVMS/noTRE groups. Interestingly, CVMS/LMTRE and noCVMS/LMTRE mice were similar to noCVMS/noTRE mice in that they did not show anxiety, while CVMS/HTRE and noCVMS/HTRE mice did not, which were similar to the mice in CVMS/noTRE. We propose that LMTRE instead of HTRE changes the serum concentration of E2. ERß/ERα ratio and OT level in the brain may be responsible for the decrease in anxiety-like behavior in female mice exposed to anxiety-inducing stress conditions.

Early maternal deprivation impairs learning and memory and alters hippocampal gene expression in adult male rats.

Maternal deprivation (MD) in early life severely disrupts hippocampal development, leading to persistent cognitive and behavior deficits. The current study uncovered that early MD (P1-P21) impaired spatial learning and memory capacity detected by Morris water maze (MWM) tests from juvenile (P31) to adult (P81) rats compared to age-matched controls. And the protein expression in hippocampus were detected by two-dimensional gel electrophoresis (2-DE) before MWM, respectively. Protein changes in hippocampal were examined to identify the molecular pathways underlying MD-induced hippocampal dysfunction. There were 11 differentially expressed proteins analyzed between adult MD and control male rats, while the 8 proteins were then identified by UPLC-ESI-Q-TOF-MS. Gene Ontology (GO) annotations of the identified proteins were related to neuronal and glial cytoskeletal dynamics, membrane signaling, stress responses, biosynthesis, and metabolism. The different expression proteins spectrin alpha chain, non-erythrocytic 1 (Sptan1), ATP-citrate synthase (Acly), and heat shock protein 90-alpha (Hsp90aa1) have been verified by western blot analysis, and their expression levels showed consistent with 2-DE analysis. In addition, glial fibrillary acidic protein (GFAP) was also found reduced in adult hippocampus of MD rats. This study identifies candidate proteins encompassing a range of functional categories that may contribute to persistent learning and memory deficits due to early life MD.

Dependence of SARS-CoV-2 infection on cholesterol-rich lipid raft and endosomal acidification.

Coronavirus disease 2019 is a kind of viral pneumonia caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, the mechanism whereby SARS-CoV-2 invades host cells remains poorly understood. Here we used SARS-CoV-2 pseudoviruses to infect human angiotensin-converting enzyme 2 (ACE2) expressing HEK293T cells and evaluated virus infection. We confirmed that SARS-CoV-2 entry was dependent on ACE2 and sensitive to pH of endosome/lysosome in HEK293T cells. The infection of SARS-CoV-2 pseudoviruses is independent of dynamin, clathrin, caveolin and endophilin A2, as well as macropinocytosis. Instead, we found that the infection of SARS-CoV-2 pseudoviruses was cholesterol-rich lipid raft dependent. Cholesterol depletion of cell membranes with methyl-ß-cyclodextrin resulted in reduction of pseudovirus infection. The infection of SARS-CoV-2 pseudoviruses resumed with cholesterol supplementation. Together, cholesterol-rich lipid rafts, and endosomal acidification, are key steps of SARS-CoV-2 required for infection of host cells. Therefore, our finding expands the understanding of SARS-CoV-2 entry mechanism and provides a new anti-SARS-CoV-2 strategy.

ß-Endorphin Induction by Psychological Stress Promotes Leydig Cell Apoptosis through p38 MAPK Pathway in Male Rats.

: Psychological stress (PS) disturbs the reproductive endocrine system and promotes male infertility, but the underlying pathogenic mechanisms have not been extensively studied. This study aimed to uncover the mechanisms of PS-induced male reproductive related abnormalities subjected to a 'terrified sound' exposure. Male rats subjected to PS displayed slow growth, decreased sperm quality, abnormal levels of the reproductive endocrine hormones, decreased expression of the reproductive-related proteins androgen-binding protein (ABP) and bromodomain-containing protein (BRDT), increased apoptosis in the testis, and accompanied by elevated levels of ß-endorphin (ß-EP). These effects were reversed by naloxone. Furthermore, PS-induced ß-EP could promote mu opioid receptor (MOR) activation and ensure intracellular p38 MAPK phosphorylation and then lead to Leydig cells (LCs) apoptosis. The current result showed that ß-EP was a key factor to PS-induced male infertility.

RNA-Binding Motif Protein 6 is a Candidate Serum Biomarker for Pancreatic Cancer.

PURPOSE: Early diagnosis is crucial to improve outcomes for pancreatic cancer patients (PC). The present study is designed to identify differently expressed peptides involved in PC as potential biomarkers. EXPERIMENTAL DESIGN: The serum proteome of 22 PC patients, 12 pancreatitis patients (PP), and 45 healthy controls (HC) are analyzed using magnetic bead-based weak cation exchange (MB-WCX) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Next, a supervised neural network (SNN) algorithm model is established by ClinProTools and the candidate biomarker identified using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Finally, the candidate biomarker is validated in tissue samples. RESULTS: The SNN algorithm model discriminates PC from HC with 92.97% sensitivity and 94.55% specificity. Seventy-six differentially expressed peptides are identified, seven of which are significantly different among PC, PP, and HC (p < 0.05). Only one peak (m/z: 1466.99) tends to be upregulated in samples from HC, PP, and PC, which is identified as region of RNA-binding motif protein 6 (RBM6). In subsequent tissue analysis, it is verified that RBM6 expression is significantly higher in PC tissues than paracancerous tissue. CONCLUSIONS AND CLINICAL RELEVANCE: The results indicate that RBM6 might serve as a candidate diagnostic biomarker for PC. CLINICAL RELEVANCE: Methods used in this study could generate serum peptidome profiles of PC, PP, and HC, and present an approach to identify potential biomarkers for diagnosis of this malignancy.