NMT1 sustains ICAM-1 to modulate adhesion and migration of tumor cells.

Protein modifications have significant effects on tumorigenesis. N-Myristoylation is one of the most important lipidation modifications, and N-myristoyltransferase 1 (NMT1) is the main enzyme required for this process. However, the mechanism underlying how NMT1 modulates tumorigenesis remains largely unclear. Here, we found that NMT1 sustains cell adhesion and suppresses tumor cell migration. Intracellular adhesion molecule 1 (ICAM-1) was a potential functional downstream effector of NMT1, and its N-terminus could be N-myristoylated. NMT1 prevented ubiquitination and proteasome degradation of ICAM-1 by inhibiting Ub E3 ligase F-box protein 4, which prolonged the half-life of ICAM-1 protein. Correlations between NMT1 and ICAM-1 were observed in liver and lung cancers, which were associated with metastasis and overall survival. Therefore, carefully designed strategies focusing on NMT1 and its downstream effectors might be helpful to treat tumors.

LPCAT3 Is Transcriptionally Regulated by YAP/ZEB/EP300 and Collaborates with ACSL4 and YAP to Determine Ferroptosis Sensitivity.

Aims: Lipid peroxidation occurring in lung adenocarcinoma (LUAD) cells leads to ferroptosis. Lysophosphatidylcholine acyl-transferase 3 (LPCAT3) plays a key role in providing raw materials for lipid peroxidation by promoting esterification of polyunsaturated fatty acids to phospholipids. Whether LPCAT3 determines ferroptosis sensitivity and the mechanism by which its expression is regulated in LUAD has not been reported. Results: LPCAT3 and acyl-coenzyme A (CoA) synthetase long-chain family member (ACSL)4 levels were positively associated with ferroptosis sensitivity in LUAD cell lines. Overexpression of LPCAT3 and ACSL4 sensitized LUAD cells to ferroptosis, while LPCAT3 and ACSL4 knockout showed the opposite effect. Zinc-finger E-box-binding (ZEB) was shown to directly bind the LPCAT3 promoter to stimulate its transcription in a Yes-associated protein (YAP)-dependent manner. An interaction between YAP and ZEB was also observed. E1A-binding protein p300 (EP300) simultaneously bound with YAP and ZEB, and induced H3K27Ac for LPCAT3 transcription. This mechanism was verified in primary LUAD cell and xenograft models. The ACSL4, LPCAT3, and YAP combination can jointly determine LUAD ferroptosis sensitivity. Innovation: The binding site of ZEB exists in the -1600 to -1401 nt region of LPCAT3 promoter, which promotes LPCAT3 transcription after ZEB binding. ZEB and YAP bind, and the ZEB zinc-finger cluster domain and YAP WW domain are crucial for their binding. EP300 may bind with YAP via its Bromo domain and with ZEB via its CBP/p300-HAT domain. In addition, the combination of ACSL4, LPCAT3, and YAP to determine ferroptosis sensitivity of LUAD cells is better than prostaglandin-endoperoxide synthase 2 (PTGS2), transferrin receptor (TFRC), or NADPH oxidase 1 (NOX1). Conclusion: LPCAT3 transcription is regulated by YAP, ZEB, and EP300. LUAD ferroptosis sensitivity can be determined by the combination of ACSL4, LPCAT3, and YAP. Antioxid. Redox Signal. 39, 491-511.

IGF2BP3 is an essential N6-methyladenosine biotarget for suppressing ferroptosis in lung adenocarcinoma cells.

A lack of promising targets leads to poor prognosis in patients with lung adenocarcinoma (LUAD). Therefore, it is urgent to identify novel therapeutic targets. The importance of the N6-methyladenosine (m6A) RNA modification has been demonstrated in various types of tumors; however, knowledge of m6A-related proteins in LUAD is still limited. Here, we found that insulin-like growth factor 2 mRNA binding protein 3 (IGF2BP3), an m6A reader protein, is highly expressed in LUAD and associated with poor prognosis. IGF2BP3 desensitizes ferroptosis (a new form of regulated cell death) in a manner dependent on its m6A reading domain and binding capacity to m6A-methylated mRNAs encoding anti-ferroptotic factors, including but not limited to glutathione peroxidase 4 (GPX4), solute carrier family 3 member 2 (SLC3A2), acyl-CoA synthetase long chain family member 3 (ACSL3), and ferritin heavy chain 1 (FTH1). After IGF2BP3 overexpression, expression levels and mRNA stabilities of these anti-ferroptotic factors were successfully sustained. Notably, significant correlations between SLC3A2, ACSL3, and IGF2BP3 were revealed in clinical LUAD specimens, further establishing the essential role of IGF2BP3 in desensitizing ferroptosis. Inducing ferroptosis has been gradually accepted as an alternative strategy to treat tumors. Thus, IGF2BP3 could be a potential target for the future development of new biomaterial-associated therapeutic anti-tumor drugs.

Natural Mediterranean Spotted Fever Foci, Qingdao, China.

We sequenced DNA from spleens of rodents captured in rural areas of Qingdao, East China, during 2013-2015. We found 1 Apodemus agrarius mouse infected with Rickettsia conorii, indicating a natural Mediterranean spotted fever foci exists in East China and that the range of R. conorii could be expanding.

ISGylation of EMD promotes its interaction with PDHA to inhibit aerobic oxidation in lung adenocarcinoma.

Abnormal nuclear structure caused by dysregulation of skeletal proteins is a common phenomenon in tumour cells. However, how skeletal proteins promote tumorigenesis remains uncovered. Here, we revealed the mechanism by which skeletal protein Emerin (EMD) promoted glucose metabolism to induce lung adenocarcinoma (LUAD). Firstly, we identified that EMD was highly expressed and promoted the malignant phenotypes in LUAD. The high expression of EMD might be due to its low level of ubiquitination. Additionally, the ISGylation at lysine 37 of EMD inhibited lysine 36 ubiquitination and upregulated EMD stability. We further explored that EMD could inhibit aerobic oxidation and stimulate glycolysis. Mechanistically, via its ß-catenin interaction domain, EMD bound with PDHA, stimulated serine 293 and 300 phosphorylation and inhibited PDHA expression, facilitated glycolysis of glucose that should enter the aerobic oxidation pathway, and EMD ISGylation was essential for EMD-PDHA interaction. In clinical LUAD specimens, EMD was negatively associated with PDHA, while positively associated with EMD ISGylation, tumour stage and diameter. In LUAD with higher glucose level, EMD expression and ISGylation were higher. Collectively, EMD was a stimulator for LUAD by inhibiting aerobic oxidation via interacting with PDHA. Restricting cancer-promoting role of EMD might be helpful for LUAD treatment.

Essential roles of exosome and circRNA_101093 on ferroptosis desensitization in lung adenocarcinoma.

BACKGROUND: Resistance to ferroptosis, a regulated cell death caused by iron-dependent excessive accumulation of lipid peroxides, has recently been linked to lung adenocarcinoma (LUAD). Intracellular antioxidant systems are required for protection against ferroptosis. The purpose of the present study was to investigate whether and how extracellular system desensitizes LUAD cells to ferroptosis. METHODS: Established human lung fibroblasts MRC-5, WI38, and human LUAD H1650, PC9, H1975, H358, A549, and H1299 cell lines, tumor and matched normal adjacent tissues of LUAD, and plasma from healthy individuals and LUAD patients were used in this study. Immunohistochemistry and immunoblotting were used to analyze protein expression, and quantitative reverse transcription-PCR was used to analyze mRNA expression. Cell viability, cell death, and the lipid reactive oxygen species generation were measured to evaluate the responses to ferroptosis. Exosomes were observed using transmission electron microscope. The localization of arachidonic acid (AA) was detected using click chemistry labeling followed by confocal microscopy. Interactions between RNAs and proteins were detected using RNA pull-down, RNA immunoprecipitation and photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation methods. Proteomic analysis was used to investigate RNA-regulated proteins, and metabolomic analysis was performed to analyze metabolites. Cell-derived xenograft, patient-derived xenograft, cell-implanted intrapulmonary LUAD mouse models and plasma/tissue specimens from LUAD patients were used to validate the molecular mechanism. RESULTS: Plasma exosome from LUAD patients specifically reduced lipid peroxidation and desensitized LUAD cells to ferroptosis. A potential explanation is that exosomal circRNA_101093 (cir93) maintained an elevation in intracellular cir93 in LUAD to modulate AA, a poly-unsaturated fatty acid critical for ferroptosis-associated increased peroxidation in the plasma membrane. Mechanistically, cir93 interacted with and increased fatty acid-binding protein 3 (FABP3), which transported AA and facilitated its reaction with taurine. Thus, global AA was reduced, whereas N-arachidonoyl taurine (NAT, the product of AA and taurine) was induced. Notably, the role of NAT in suppressing AA incorporation into the plasma membrane was also revealed. In pre-clinical in vivo models, reducing exosome improved ferroptosis-based treatment. CONCLUSION: Exosome and cir93 are essential for desensitizing LUAD cells to ferroptosis, and blocking exosome may be helpful for future LUAD treatment.

Tumour cells are sensitised to ferroptosis via RB1CC1-mediated transcriptional reprogramming.

BACKGROUND: Ferroptosis, a form of regulated cell death, is an important topic in the field of cancer research. However, the signalling pathways and factors that sensitise tumour cells to ferroptosis remain elusive. METHODS: We determined the level of ferroptosis in cells by measuring cell death and lipid reactive oxygen species (ROS) production. The expression of RB1-inducible coiled-coil 1 (RB1CC1) and related proteins was analyzed by immunoblotting and immunohistochemistry. Immunofluorescence was used to determine the subcellular localization of RB1CC1. We investigated the mechanism of RB1CC1 nuclear translocation by constructing a series of RB1CC1 variants. To examine the ferroptosis- and RB1CC1-dependent transcriptional program in tumour cells, chromatin immunoprecipitation sequencing was performed. To assess the effect of c-Jun N-terminal kinase (JNK) agonists on strenthening imidazole ketone erastin (IKE) therapy, we constructed cell-derived xenograft mouse models. Mouse models of hepatocellular carcinoma to elucidate the importance of Rb1cc1 in IKE-based therapy of liver tumourigenesis. RESULTS: RB1CC1 is upregulated by lipid ROS and that nuclear translocation of phosphorylation of RB1CC1 at Ser537 was essential for sensitising ferroptosis in tumour cells. Upon ferroptosis induction, nuclear RB1CC1 sharing forkhead box (FOX)-binding motifs recruits elongator acetyltransferase complex subunit 3 (ELP3) to strengthen H4K12Ac histone modifications within enhancers linked to ferroptosis. This also stimulated transcription of ferroptosis-associated genes, such as coiled-coil-helix-coiled-coil-helix domain containing 3 (CHCHD3), which enhanced mitochondrial function to elevate mitochondrial ROS early following induction of ferroptosis. FDA-approved JNK activators reinforced RB1CC1 nuclear translocation and sensitised cells to ferroptosis, which strongly suggested that JNK is upstream of RB1CC1. Nuclear localisation of RB1CC1 correlated with lipid peroxidation in clinical lung cancer specimens. Rb1cc1 was essential for ferroptosis agonists to suppress liver tumourigenesis in mice. CONCLUSIONS: Our findings indicate that RB1CC1-associated signalling sensitises tumour cells to ferroptosis and that targeting RB1CC1 may be beneficial for tumour treatment.

YAP ISGylation increases its stability and promotes its positive regulation on PPP by stimulating 6PGL transcription.

Yes-associated protein (YAP) activation is crucial for tumor formation and development, and its stability is regulated by ubiquitination. ISGylation is a type of ubiquitination like post-translational modification, whereas whether YAP is ISGylated and how ISGylation influences YAP ubiquitination-related function remains uncovered. In addition, YAP can activate glucose metabolism by activating the hexosamine biosynthesis pathway (HBP) and glycolysis, and generate a large number of intermediates to promote tumor proliferation. However, whether YAP stimulates the pentose phosphate pathway (PPP), another tumor-promoting glucose metabolism pathway, and the relationship between this stimulation and ISGylation needs further investigation. Here, we found that YAP was ISGylated and this ISGylation inhibited YAP ubiquitination, proteasome degradation, interaction with-beta-transducin repeat containing E3 ubiquitin-protein ligase (ßTrCP) to promote YAP stability. However, ISGylation-induced pro-YAP effects were abolished by YAP K497R (K, lysine; R, arginine) mutation, suggesting K497 could be the major YAP ISGylation site. In addition, YAP ISGylation promoted cell viability, cell-derived xenograft (CDX) and patient-derived xenograft (PDX) tumor formation. YAP ISGylation also increased downstream genes transcription, including one of the key enzymes of PPP, 6-phosphogluconolactonase (6PGL). Mechanistically, YAP promoted 6PGL transcription by simultaneously recruiting SMAD family member 2 (SMAD2) and TEA domain transcription factor 4 (TEAD4) binding to the 6PGL promoter to activate PPP. In clinical lung adenocarcinoma (LUAD) specimens, we found that YAP ISGylation degree was positively associated with 6PGL mRNA level, especially in high glucose LUAD tissues compared to low glucose LUAD tissues. Collectively, this study suggested that YAP ISGylation is critical for maintaining its stability and further activation of PPP. Targeting ISGylated YAP might be a new choice for hyperglycemia cancer treatment.

The essential roles of m6A RNA modification to stimulate ENO1-dependent glycolysis and tumorigenesis in lung adenocarcinoma.

BACKGROUND: Lung adenocarcinoma (LUAD)  is the most common subtype of lung cancer. Patient prognosis is poor, and the existing therapeutic strategies for LUAD are far from satisfactory. Recently, targeting N6-methyladenosine (m6A) modification of RNA has been suggested as a potential strategy to impede tumor progression. However, the roles of m6A modification in LUAD tumorigenesis is unknown. METHODS: Global m6A levels and expressions of m6A writers, erasers and readers were evaluated by RNA methylation assay, dot blot, immunoblotting, immunohistochemistry and ELISA in human LUAD, mouse models and cell lines. Cell viability, 3D-spheroid generation, in vivo LUAD formation, experiments in cell- and patient-derived xenograft mice and survival analysis were conducted to explore the impact of m6A on LUAD. The RNA-protein interactions, translation, putative m6A sites and glycolysis were explored in the investigation of the mechanism underlying how m6A stimulates tumorigenesis. RESULTS: The elevation of global m6A level in most human LUAD specimens resulted from the combined upregulation of m6A writer methyltransferase 3 (METTL3) and downregulation of eraser alkB homolog 5 (ALKBH5). Elevated global m6A level was associated with a poor overall survival in LUAD patients. Reducing m6A levels by knocking out METTL3 and overexpressing ALKBH5 suppressed 3D-spheroid generation in LUAD cells and intra-pulmonary tumor formation in mice. Mechanistically, m6A-dependent stimulation of glycolysis and tumorigenesis occurred via enolase 1 (ENO1). ENO1 mRNA was m6A methylated at 359 A, which facilitated it's binding with the m6A reader YTH N6-methyladenosine RNA binding protein 1 (YTHDF1) and resulted in enhanced translation of ENO1. ENO1 positively correlated with METTL3 and global m6A levels, and negatively correlated with ALKBH5 in human LUAD. In addition, m6A-dependent elevation of ENO1 was associated with LUAD progression. In preclinical models, tumors with a higher global m6A level showed a more sensitive response to the inhibition of pan-methylation, glycolysis and ENO activity in LUAD. CONCLUSIONS: The m6A-dependent stimulation of glycolysis and tumorigenesis in LUAD is at least partially orchestrated by the upregulation of METTL3, downregulation of ALKBH5, and stimulation of YTHDF1-mediated ENO1 translation. Blocking this mechanism may represent a potential treatment strategy for m6A-dependent LUAD.

An Expectation Maximization Based Adaptive Group Testing Method for Improving Efficiency and Sensitivity of Large-Scale Screening of COVID-19.

The pathogen of the ongoing coronavirus disease 2019 (COVID-19) pandemic is a newly discovered virus called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Testing individuals for SARS-CoV-2 plays a critical role in containing COVID-19. For saving medical personnel and consumables, many countries are implementing group testing against SARS-CoV-2. However, existing group testing methods have the following limitations: (1) The group size is determined without theoretical analysis, and hence is usually not optimal. This adversely impacts the screening efficiency. (2) These methods neglect the fact that mixing samples together usually leads to substantial dilution of the SARS-CoV-2 virus, which seriously impacts the sensitivity of tests. In this paper, we aim to screen individuals infected with COVID-19 with as few tests as possible, under the premise that the sensitivity of tests is high enough. We propose an eXpectation Maximization based Adaptive Group Testing (XMAGT) method. The basic idea is to adaptively adjust its testing strategy between a group testing strategy and an individual testing strategy such that the expected number of samples identified by a single test is larger. During the screening process, the XMAGT method can estimate the ratio of positive samples. With this ratio, the XMAGT method can determine a group size under which the group testing strategy can achieve a maximal expected number of negative samples and the sensitivity of tests is higher than a user-specified threshold. Experimental results show that the XMAGT method outperforms existing methods in terms of both efficiency and sensitivity.

tsRNA-5001a promotes proliferation of lung adenocarcinoma cells and is associated with postoperative recurrence in lung adenocarcinoma patients.

BACKGROUND: The biological role and clinical significance of transfer RNA-derived small RNAs (tsRNAs) remain largely unclear. The purpose of this study was to investigate the biological function, molecular mechanism, and clinical significance of tsRNA-5001a in lung adenocarcinoma. METHODS: The function of tsRNA-5001a on the growth of tumor cells was accessed by cell function experiments. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the expression of tsRNA-5001a in paired samples of lung adenocarcinoma. Cell localization of tsRNA-5001a was performed by nuclear-cytoplasmic separation assay. Transcriptome sequencing was used to screen the molecules involved in the regulatory network of tsRNA-5001a. Independent samples t-test was used to compare the two groups. Prism software (Prism 7.0) was used to analyze the statistical results. P<0.05 was considered statistically significant. RESULTS: tsRNA-5001a was significantly upregulated in lung adenocarcinoma tissues. Upregulation of tsRNA-5001a was found to increase the risk of postoperative recurrences in patients with lung adenocarcinoma and was associated with poor prognosis. Function assay showed that overexpression tsRNA-5001a could significantly promote cell proliferation. In contrast, knockdown of tsRNA-5001a significantly inhibited the proliferation of lung adenocarcinoma cells. In addition, nucleoplasmic isolation assay indicated that tsRNA-5001a was located mainly in the cytoplasm. According to the results of RNA sequencing and The Cancer Genome Atlas database (TCGA database) analysis, growth arrest and DNA damage 45G (GADD45G) was screened and may be the target gene of tsRNA-5001a. CONCLUSIONS: tsRNA-5001a promotes the proliferation of lung adenocarcinoma cells and increases the risk of postoperative recurrences in lung adenocarcinoma patients.

Transcriptional Repression of Ferritin Light Chain Increases Ferroptosis Sensitivity in Lung Adenocarcinoma.

Ferroptosis is an iron- and lipid peroxidation-dependent form of regulated cell death. The release of labile iron is one of the important factors affecting sensitivity to ferroptosis. Yes-associated protein (YAP) controls intracellular iron levels by affecting the transcription of ferritin heavy chain (FTH) and transferrin receptor (TFRC). However, whether YAP regulates iron metabolism through other target genes remains unknown. Here, we observed that the system Xc- inhibitor erastin inhibited the binding of the WW domain and PSY motif between YAP and transcription factor CP2 (TFCP2), and then suppressed the transcription of ferritin light chain (FTL) simultaneously mediated by YAP, TFCP2 and forkhead box A1 (FOXA1). Furthermore, inhibition of FTL expression abrogated ferroptosis-resistance in cells with sustained YAP expression. Unlike FTH, which exhibited first an increase and then a decrease in transcription, FTL transcription continued to decline after the addition of erastin, and a decrease in lysine acetyltransferase 5 (KAT5)-dependent acetylation of FTL was also observed. In lung adenocarcinoma (LUAD) tissues, lipid peroxidation and labile iron decreased, while YAP, TFCP2 and FTL increased compared to their adjacent normal tissues, and the lipid peroxidation marker 4-hydroxynonenal (4-HNE) was negatively correlated with the level of FTL or the degree of LUAD malignancy, but LUAD tissues with lower levels of 4-HNE showed a higher sensitivity to ferroptosis. In conclusion, the findings from this study indicated that the suppression of FTL transcription through the inhibition of the YAP-TFCP2-KAT5 complex could be another mechanism for elevating ferroptosis sensitivity and inducing cell death, and ferroptotic therapy is more likely to achieve better results in LUAD patients with a lower degree of lipid peroxidation.

TRIB2 desensitizes ferroptosis via ßTrCP-mediated TFRC ubiquitiantion in liver cancer cells.

Tribbles homolog 2 (TRIB2) is known to boost liver tumorigenesis via regulating Ubiquitin (Ub) proteasome system (UPS). At least two ways are involved, i.e., acts as an adaptor protein to modulate ubiquitination functions of certain ubiquitin E3 ligases (E3s) and reduces global Ub levels via increasing the proteolysis activity of proteasome. Recently, we have identified the role of TRIB2 to relieve oxidative damage via reducing the availability of Ub that is essential for the ubiquitination and subsequent degradation of Glutathione peroxidase 4 (GPX4). Although GPX4 is a critical antioxidant factor to protect against ferroptosis, the exact evidence showing that TRIB2 desensitizes ferroptosis is lacking. Also, whether such function is via E3 remains unclear. Here, we demonstrated that deletion of TRIB2 sensitized ferroptosis via lifting labile iron in liver cancer cells. By contrast, overexpression of TRIB2 led to the opposite outcome. We further demonstrated that transferrin receptor (TFRC) was required for TRIB2 to desensitize the cells to ferroptosis. Without TFRC, the labile iron pool could not be reduced by overexpressing TRIB2. We also found that beta-transducin repeat containing E3 ubiqutin protein ligase (ßTrCP) was a genuine E3 for the ubiquitination of TFRC, and TRIB2 was unable to decline labile iron level once upon ßTrCP was knocked out. In addition, we confirmed that the opposite effects on ferroptosis and ferroptosis-associated lipid reactive oxygen species (ROS) generation resulted from knockout and overexpression of TRIB2 were all indispensible of TFRC and ßTrCP. Finally, we demonstrated that TRIB2 exclusively manipulated RSL3- and erastin-induced-ferroptosis independent of GPX4 and glutathione (GSH). In conclusion, we elucidated a novel role of TRIB2 to desensitize ferroptosis via E3 ßTrCP, by which facilitates TFRC ubiquitiation and finally decreases labile iron in liver cancer cells.

Targeting SLC3A2 subunit of system XC- is essential for m6A reader YTHDC2 to be an endogenous ferroptosis inducer in lung adenocarcinoma.

The m6A reader YT521-B homology containing 2 (YTHDC2) has been identified to inhibit lung adenocarcinoma (LUAD) tumorigenesis by suppressing solute carrier 7A11 (SLC7A11)-dependent antioxidant function. SLC7A11 is a major functional subunit of system XC-. Inhibition of system XC- can induce ferroptosis. However, whether suppressing SLC7A11 is sufficient for YTHDC2 to be an endogenous ferroptosis inducer in LUAD is unknown. Here, we found that induction of YTHDC2 to a high level can induce ferroptosis in LUAD cells but not in lung and bronchus epithelial cells. In addition to SLC7A11, solute carrier 3A2 (SLC3A2), another subunit of system XC- was equally important for YTHDC2-induced ferroptosis. YTHDC2 m6A-dependently destabilized Homeo box A13 (HOXA13) mRNA because a potential m6A recognition site was identified within its 3' untranslated region (3'UTR). Interestingly, HOXA13 acted as a transcription factor to stimulate SLC3A2 expression. Thereby, YTHDC2 suppressed SLC3A2 via inhibiting HOXA13 in an m6A-indirect manner. Mouse experiments further confirmed the associations among YTHDC2, SLC3A2 and HOXA13, and demonstrated that SLC3A2 and SLC7A11 were both important for YTHDC2-impaired tumor growth and -induced lipid peroxidation in vivo. Moreover, higher expression of SLC7A11, SLC3A2 and HOXA13 indicate poorer clinical outcome in YTHDC2-suppressed LUAD patients. In conclusion, YTHDC2 is believed to be a powerful endogenous ferroptosis inducer and targeting SLC3A2 subunit of system XC- is essential for this process. Increasing YTHDC2 is an alternative ferroptosis-based therapy to treat LUAD.

RRM2 protects against ferroptosis and is a tumor biomarker for liver cancer.

BACKGROUND: Ferroptosis is the process of cell death triggered by lipid peroxides, and inhibition of glutathione (GSH) synthesis leads to ferroptosis. Liver cancer progression is closely linked to ferroptosis suppression. However, the mechanism by which inhibition of GSH synthesis suppresses potential ferroptosis of liver cancer cells and whether ferroptosis-related liver cancer biomarkers have a promising diagnostic value remain unknown. METHODS: Ribonucleotide reductase regulatory subunit M2 (RRM2) levels were measured using an enzyme linked immunosorbent assay (ELISA), quantitative RT-PCR (qPCR), immunoblotting (IB) and immunochemistry (IHC). Cell viability and cell death were measured by a CellTiter-Glo luminescent cell viability assay and staining with SYTOX Green followed by flow cytometry, respectively. Metabolites were measured using the indicated kits. The Interaction between glutathione synthetase (GSS) and RRM2 was measured using immunofluorescence (IF), co-immunoprecipitation (co-IP) and the proximal ligation assay (PLA). The diagnostic value was analyzed using the area under the receiver operating characteristic curve (AUC-ROC). Bioinformatics analysis was performed using the indicated database. RESULTS: RRM2 showed specifically elevated levels in liver cancer and inhibited ferroptosis by stimulating GSH synthesis via GSS. Mechanistically, phosphorylation of RRM2 at the Threonine 33 residue (T33) was maintained at normal levels to block the RRM2-GSS interaction and therefore protected RRM2 and GSS from further proteasome degradation. However, under ferroptotic stress, RRM2 was dephosphorylated at T33, thus the RRM2-GSS interaction was promoted. This resulted in the translocation of RRM2 and GSS to the proteasome for simultaneous degradation. Clinically, serum RRM2 was significantly associated with serum alpha-fetoprotein (AFP), carcinoembryonic antigen (CEA), alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), gamma glutamyl transpeptidase (γ-GT), albumin (ALB) and total bilirubin. The AUC-ROC for the combination of RRM2 with AFP was 0.947, with a sensitivity of 88.7% and a specificity of 97.0%, which indicates better diagnostic performance compared to either RRM2 or AFP alone. CONCLUSION: RRM2 exerts an anti-ferroptotic role in liver cancer cells by sustaining GSH synthesis. Serum RRM2 will be useful as a biomarker to evaluate the degree to which ferroptosis is suppressed and improve diagnostic efficiency for liver cancer.

Effect of TP53 rs1042522 on the susceptibility of patients to oral squamous cell carcinoma and oral leukoplakia: a meta-analysis.

BACKGROUND: There are different and inconsistent conclusions regarding the genetic relationship between the human tumor suppressor p53 (TP53) rs1042522 polymorphism and the risk of oral squamous cell carcinoma (OSCC) and oral leukoplakia (OL). Therefore, the aim of the study was to comprehensively reassess this association through the performance of an updated meta-analysis. METHODS: After searching the available databases, we systematically screened and included the eligible case-control studies, which contain the full genotype frequency data of the TP53 rs1042522 polymorphism for both OSCC/OL patients and the negative control groups. PA (P-value of the association test) and ORs (odd ratios) with their corresponding 95% CIs (confidence intervals) were calculated to quantitatively evaluate the influence of TP53 rs1042522 on the susceptibility of patients to OSCC or OL. RESULTS: In total, twenty eligible case-control articles were finally enrolled. Compared with the controls, no increased or decreased risk of OSCC was observed in the cases for six genetic models including allele C vs. G (PA = 0.741), carrier C vs. G (PA = 0.853), homozygote CC vs. GG (PA = 0.085), heterozygote GC vs. GG (PA = 0.882), dominant GC + CC vs. GG (PA = 0.969), and recessive CC vs. GG + GC (PA = 0.980). Furthermore, no statistically significant difference between the cases and controls was detected in most subgroup meta-analyses (PA > 0.05). For the risk of OL, we did not observe the difference between the cases and controls for most genetic models in the overall meta-analysis and subsequent subgroup analysis (PA > 0.05). Begg's test and Egger's test excluded the large risk of publication bias within the included studies in the meta-analysis of OSCC. The sensitivity analysis indicated the above relatively stable results. CONCLUSIONS: Our updated meta-analysis (based on the current evidence) shows that TP53 rs1042522 may not confer susceptibility to OSCC. In addition, for the first time, we provided evidence regarding the negative association between TP53 rs1042522 and OL risk.

Predicting the progression of ophthalmic disease based on slit-lamp images using a deep temporal sequence network.

Ocular images play an essential role in ophthalmology. Current research mainly focuses on computer-aided diagnosis using slit-lamp images, however few studies have been done to predict the progression of ophthalmic disease. Therefore exploring an effective approach of prediction can help to plan treatment strategies and to provide early warning for the patients. In this study, we present an end-to-end temporal sequence network (TempSeq-Net) to automatically predict the progression of ophthalmic disease, which includes employing convolutional neural network (CNN) to extract high-level features from consecutive slit-lamp images and applying long short term memory (LSTM) method to mine the temporal relationship of features. First, we comprehensively compare six potential combinations of CNNs and LSTM (or recurrent neural network) in terms of effectiveness and efficiency, to obtain the optimal TempSeq-Net model. Second, we analyze the impacts of sequence lengths on model's performance which help to evaluate their stability and validity and to determine the appropriate range of sequence lengths. The quantitative results demonstrated that our proposed model offers exceptional performance with mean accuracy (92.22), sensitivity (88.55), specificity (94.31) and AUC (97.18). Moreover, the model achieves real-time prediction with only 27.6ms for single sequence, and simultaneously predicts sequence data with lengths of 3-5. Our study provides a promising strategy for the progression of ophthalmic disease, and has the potential to be applied in other medical fields.

Automatic diagnosis of imbalanced ophthalmic images using a cost-sensitive deep convolutional neural network.

BACKGROUND: Ocular images play an essential role in ophthalmological diagnoses. Having an imbalanced dataset is an inevitable issue in automated ocular diseases diagnosis; the scarcity of positive samples always tends to result in the misdiagnosis of severe patients during the classification task. Exploring an effective computer-aided diagnostic method to deal with imbalanced ophthalmological dataset is crucial. METHODS: In this paper, we develop an effective cost-sensitive deep residual convolutional neural network (CS-ResCNN) classifier to diagnose ophthalmic diseases using retro-illumination images. First, the regions of interest (crystalline lens) are automatically identified via twice-applied Canny detection and Hough transformation. Then, the localized zones are fed into the CS-ResCNN to extract high-level features for subsequent use in automatic diagnosis. Second, the impacts of cost factors on the CS-ResCNN are further analyzed using a grid-search procedure to verify that our proposed system is robust and efficient. RESULTS: Qualitative analyses and quantitative experimental results demonstrate that our proposed method outperforms other conventional approaches and offers exceptional mean accuracy (92.24%), specificity (93.19%), sensitivity (89.66%) and AUC (97.11%) results. Moreover, the sensitivity of the CS-ResCNN is enhanced by over 13.6% compared to the native CNN method. CONCLUSION: Our study provides a practical strategy for addressing imbalanced ophthalmological datasets and has the potential to be applied to other medical images. The developed and deployed CS-ResCNN could serve as computer-aided diagnosis software for ophthalmologists in clinical application.

Localization and diagnosis framework for pediatric cataracts based on slit-lamp images using deep features of a convolutional neural network.

Slit-lamp images play an essential role for diagnosis of pediatric cataracts. We present a computer vision-based framework for the automatic localization and diagnosis of slit-lamp images by identifying the lens region of interest (ROI) and employing a deep learning convolutional neural network (CNN). First, three grading degrees for slit-lamp images are proposed in conjunction with three leading ophthalmologists. The lens ROI is located in an automated manner in the original image using two successive applications of Candy detection and the Hough transform, which are cropped, resized to a fixed size and used to form pediatric cataract datasets. These datasets are fed into the CNN to extract high-level features and implement automatic classification and grading. To demonstrate the performance and effectiveness of the deep features extracted in the CNN, we investigate the features combined with support vector machine (SVM) and softmax classifier and compare these with the traditional representative methods. The qualitative and quantitative experimental results demonstrate that our proposed method offers exceptional mean accuracy, sensitivity and specificity: classification (97.07%, 97.28%, and 96.83%) and a three-degree grading area (89.02%, 86.63%, and 90.75%), density (92.68%, 91.05%, and 93.94%) and location (89.28%, 82.70%, and 93.08%). Finally, we developed and deployed a potential automatic diagnostic software for ophthalmologists and patients in clinical applications to implement the validated model.