Improvement of ACK1-targeted therapy efficacy in lung adenocarcinoma using chloroquine or bafilomycin A1.

BACKGROUND: Activated Cdc42-associated kinase 1 (ACK1) is a promising druggable target for cancer, but its inhibitors only showed moderate effects in clinical trials. The study aimed to investigate the underlying mechanisms and improve the antitumor efficacy of ACK1 inhibitors. METHODS: RNA-seq was performed to determine the downstream pathways of ACK. Using Lasso Cox regression analysis, we built a risk signature with ACK1-related autophagy genes in the lung adenocarcinoma (LUAD) patients from The Cancer Genome Atlas (TCGA) project. The performance of the signature in predicting the tumor immune environment and response to immunotherapy and chemotherapy were assessed in LUAD. CCK8, mRFP-GFP-LC3 assay, western blot, colony formation, wound healing, and transwell migration assays were conducted to evaluate the effects of the ACK1 inhibitor on lung cancer cells. A subcutaneous NSCLC xenograft model was used for in vivo study. RESULTS: RNA-seq revealed the regulatory role of ACK1 in autophagy. Furthermore, the risk signature separated LUAD patients into low- and high-risk groups with significantly different prognoses. The two groups displayed different tumor immune environments regarding 28 immune cell subsets. The low-risk groups showed high immune scores, high CTLA4 expression levels, high immunophenoscore, and low DNA mismatch repair capacity, suggesting a better response to immunotherapy. This signature also predicted sensitivity to commonly used chemotherapy and targeted drugs. In vitro, the ACK1 inhibitors (AIM-100 and Dasatinib) appeared to trigger adaptive autophagy-like response to protect lung cancer cells from apoptosis and activated the AMPK/mTOR signaling pathway, partially explaining its moderate antitumor efficacy. However, blocking lysosomal degradation with chloroquine/Bafilamycine A1 or inhibiting AMPK signaling with compound C/shPRKAA1 enhanced the ACK1 inhibitor's cytotoxic effects on lung cancer cells. The efficacy of the combined therapy was also verified using a mouse xenograft model. CONCLUSIONS: The resulting signature from ACK1-related autophagy genes robustly predicted survival and drug sensitivity in LUAD. The lysosomal degradation inhibition improved the therapeutic effects of the ACK1 inhibitor, suggesting a potential role for autophagy in therapy evasion.

Response to neoadjuvant immune checkpoint inhibitors and chemotherapy in Chinese patients with esophageal squamous cell carcinoma: the role of tumor immune microenvironment.

BACKGROUND: Immune checkpoint inhibitors (ICIs) through programmed cell death 1 blockade improve the survival outcomes of patients with advanced esophageal squamous cell carcinoma (ESCC). Recently, the use of neoadjuvant immunotherapy for the treatment of ESCC has been gradually increasing. We aimed to evaluate the efficacy of neoadjuvant treatment of ICIs with chemotherapy and explore tumor microenvironment (TME) immune profiles of ESCC samples during neoadjuvant therapy. METHODS: Patients with previously untreated, resectable, locally advanced ESCC (stage II or III) in Harbin Medical University Cancer Hospital were enrolled. Each patient received two to four cycles of neoadjuvant ICIs combined with chemotherapy before surgical resection. The TME immune profiles of formalin-fixed paraffin-embedded tumor samples at baseline and after surgery were evaluated by multiplex staining and multispectral imaging. RESULTS: In all, 18 patients were enrolled, and all patients received surgery with R0 resection. The postoperative pathological evaluation indicated that 7 (38.9%) patients had a pathological complete response (pCR) and 11 (61.1%) patients had a partial response. The neoadjuvant therapeutic regimens had acceptable side effect profiles. The TME immune profiles at baseline observed higher densities of stroma CD3 + , PD-1 + , and PD-1 + CD3 + cells in pCR patients than in non-pCR patients. Comparing TME immune profiles before and after neoadjuvant treatment, an increase in CD8 + T cells and a decrease in CD163 + CD68 + M2-like macrophage cells were observed after neoadjuvant treatment. CONCLUSIONS: Neoadjuvant ICIs combined with chemotherapy produced a satisfactory treatment response, demonstrating its anti-tumor efficacy in locally advanced ESCC. Further large-scale studies are required to understand the role of tumor immunities and ICIs underlying ESCC.

TMEM139 prevents NSCLC metastasis by inhibiting lysosomal degradation of E-cadherin.

Non-small-cell lung cancer (NSCLC) accounts for approximately 85% of all lung cancer cases and has the highest mortality rate among all solid tumors. It is characterized by early metastasis, and investigations of the molecular mechanisms underlying the progression and metastasis of NSCLC are urgently needed for the development of therapeutic targets. Here, we report that the transmembrane protein TMEM139 is significantly downregulated in NSCLC and that reduced expression of TMEM139 is correlated with a poor prognosis in NSCLC patients. Mechanistically, we found that TMEM139 directly interacts with E-cadherin at the plasma membrane and at focal adhesion sites. Moreover, TMEM139 can prevent the lysosomal degradation of E-cadherin, which inhibits epithelial-mesenchymal transition, migration and invasion of NSCLC cells both in vitro and in vivo. Our study not only expands our understanding of NSCLC metastasis but also provides a foundation to develop novel therapeutic strategies.

Pan-cancer analysis of UBE2T with a focus on prognostic and immunological roles in lung adenocarcinoma.

BACKGROUND: Ubiquitin-conjugating enzyme E2 T (UBE2T) is a potential oncogene. However, Pan-cancer analyses of the functional, prognostic and predictive implications of this gene are lacking. METHODS: We first analyzed UBE2T across 33 tumor types in The Cancer Genome Atlas (TCGA) project. We investigated the expression level of UBE2T and its effect on prognosis using the TCGA database. The correlation between UBE2T and cell cycle in pan-cancer was investigated using the single-cell sequencing data in Cancer Single-cell State Atlas (CancerSEA) database. The Weighted Gene Co-expression Network analysis (WGCNA), Univariate Cox and Least absolute shrinkage and selection operator (LASSO) Cox regression models, and receiver operating characteristic (ROC) were applied to assess the prognostic impact of UBE2T-related cell cycle genes (UrCCGs). Furthermore, the consensus clustering (CC) method was adopted to divide TCGA-lung adenocarcinoma (LUAD) patients into subgroups based on UrCCGs. Prognosis, molecular characteristics, and the immune panorama of subgroups were analyzed using Single-sample Gene Set Enrichment Analysis (ssGSEA). Results derived from TCGA-LUAD patients were validated in International Cancer Genome Consortium (ICGC)-LUAD data. RESULTS: UBE2T is highly expressed and is a prognostic risk factor in most tumors. CancerSEA database analysis revealed that UBE2T was positively associated with the cell cycle in various cancers(r > 0.60, p < 0.001). The risk signature of UrCCGs can reliably predict the prognosis of LUAD (AUC1 year = 0.720, AUC3 year = 0.700, AUC5 year = 0.630). The CC method classified the TCGA-LUAD cohort into 4 UrCCG subtypes (G1-G4). Kaplan-Meier survival analysis demonstrated that G2 and G4 subtypes had worse survival than G3 (Log-rank test PTCGA training set < 0.001, PICGC validation set < 0.001). A comprehensive analysis of immune infiltrates, immune checkpoints, and immunogenic cell death modulators unveiled different immune landscapes for the four subtypes. High immunophenoscore in G3 and G4 tumors suggested that these two subtypes were immunologically "hot," tending to respond to immunotherapy compared to G2 subtypes (p < 0.001). CONCLUSIONS: UBE2T is a critical oncogene in many cancers. Moreover, UrCCG classified the LUAD cohort into four subgroups with significantly different survival, molecular features, immune infiltrates, and immunotherapy responses. UBE2T may be a therapeutic target and predictor of prognosis and immunotherapy sensitivity.

Identification and validation of an eight-lncRNA signature that predicts prognosis in patients with esophageal squamous cell carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is correlated with worse clinical prognosis and lacks available targeted therapy. Thus, identification of reliable biomarkers is required for the diagnosis and treatment of ESCC. METHODS: We downloaded the GSE53625 dataset as a training dataset to screen differentially expressed RNAs (DERs) with the criterion of false discovery rate (FDR) < 0.05 and |log2fold change (FC)| > 1. A support vector machine classifier was used to find the optimal feature gene set that could conclusively distinguish different samples. An eight-lncRNA signature was identified by random survival forest algorithm and multivariate Cox regression analysis. The RNA sequencing data from The Cancer Genome Atlas (TCGA) database were used for external validation. The predictive value of the signature was assessed using Kaplan-Meier test, time-dependent receiver operating characteristic (ROC) curves, and dynamic area under the curve (AUC). Furthermore, a nomogram to predict patients' 3-year and 5-year prognosis was constructed. CCK-8 assay, flow cytometry, and transwell assay were conducted in ESCC cells. RESULTS: A total of 1136 DERs, including 689 downregulated mRNAs, 318 upregulated mRNAs, 74 downregulated lncRNAs and 55 upregulated lncRNAs, were obtained in the GES53625 dataset. From the training dataset, we identified an eight-lncRNA signature, (ADAMTS9-AS1, DLX6-AS1, LINC00470, LINC00520, LINC01497, LINC01749, MAMDC2-AS1, and SSTR5-AS1). A nomogram based on the eight-lncRNA signature, age, and pathologic stage was developed and showed good accuracy for predicting 3-year and 5-year survival probability of patients with ESCC. Functionally, knockdown of LINC00470 significantly suppressed cell proliferation, G1/S transition, and migration in two ESCC cell lines (EC9706 and TE-9). Moreover, knockdown of LINC00470 downregulated the protein levels of PCNA, CDK4, and N-cadherin, while upregulating E-cadherin protein level in EC9706 and TE-9 cells. CONCLUSION: Our eight-lncRNA signature and nomogram can provide theoretical guidance for further research on the molecular mechanism of ESCC and the screening of molecular markers.

Hypoxia-induced oxidative stress and apoptosis in gills of scaleless carp (Gymnocypris przewalskii).

Scaleless carp (Gymnocypris przewalskii) are well adapted to low oxygen environment, but their specific adaptation mechanism to hypoxic condition remains unclear. The gill is an important respiratory organ that plays a crucial role in regulating hypoxic stress. Here, we established fish hypoxic stress model, as well as investigated oxidative stress, apoptotic responses, and relative enzyme activities in the gills of scaleless carp after exposure to various levels of hypoxic stress. The results demonstrated that gill lamellar height and basal length increased significantly under severe hypoxic stress, and interval lengths between lamellae increased significantly under hypoxic stress. Furthermore, lamellar epithelial cells underwent apoptosis, cytoplasmic contraction, and mitochondrial expansion, and the number of apoptotic cells increased significantly after exposure to severe hypoxic stress for 24 h. Subsequently, Bcl-2 and Caspase 3 mRNA levels, as well as Bcl-2/Bax expression ratio were significantly increased after exposure to severe hypoxic stress for 24 h, indicating upregulation of anti-apoptotic processes. Moreover, malondialdehyde and hydrogen peroxide levels were significantly increased after exposure to hypoxic stress for 24 h. Superoxide dismutase activity increased significantly after exposure to severe hypoxia for 8 h and then decreased, while glutathione peroxidase activity and total antioxidant capacity increased significantly under hypoxic stress. Taken together, the results indicated that scaleless carp gills respond to acute hypoxic conditions by undergoing lamellar morphology remodeling, enhanced apoptosis, and increased antioxidant enzymatic activity. The study findings provided new insight into the adaptation mechanisms of scaleless carp in response to hypoxic challenge.

Effects of Bombyx mori nuclear polyhedrosis virus on serpin and antibacterial peptide expression in B. mori.

The silkworm (Bombyx mori) is a typical and economically important lepidopteran species, and research has resulted in the development and accumulation of breeding lines. Studies of immune-related silkworm genes not only promote our understanding of silkworm immune response mechanisms, but they also inform insect immune molecular diversity research. Here, silkworm proteins were screened using proteomics after Bombyx mori nuclear polyhedrosis virus (BmNPV) infection, and 2368 silkworm proteins were identified, including six antimicrobial peptides and 12 serpins. The mRNA expression levels of these 18 proteins were examined at different times. The results indicated that attacin had the highest expression level, while serpin-5 and cecropin-D exhibited a negative regulatory correlation. These results provide a significant step toward a deeper understanding of B. mori immunoregulation.

A recombinant adenovirus targeting typical Aeromonas salmonicida induces an antibody-mediated adaptive immune response after immunization of rainbow trout.

Aeromonas salmonicida, the oldest known fish pathogen and currently endemic throughout most of the world in both fresh and marine waters, causes severe economic losses to the salmon farming industry. Although there have been many studies on the prevention of furunculosis over the past few decades, it is still prevalent in many fisheries. In this study, a recombinant adenovirus vaccine candidate harboring the highly immunogenic Vapa gene (pAd-easy-cmv-Vapa) was successfully constructed and tested. The immune protection rate and specific antibody levels in the peripheral blood were then determined after immunizing rainbow trout. In addition, relative levels of IgM and IgT in the head kidney and hindgut before and after immunization were measured by quantitative reverse transcription PCR. Western blotting results indicated that the recombinant adenovirus could infect HEK-293 cells and express the A layer protein (encoded by Vapa). Further, survival analysis of fish 28 days after challenge showed that immunization significantly lowered the mortality rate (40%) compared to that in the control group (76.6%) and empty vector group (73.6%). This also led to an increase in specific antibodies in peripheral serum. In addition, levels of IgM and IgT in the head kidney and hindgut were increased to varying degrees. In conclusion, our research provides a candidate vaccine for the prevention of Aeromonas salmonicida A450 infection in rainbow trout and lays the foundation for future research on adaptive immune mechanisms associated with rainbow trout antibodies.

Comparative transcriptomics and histopathological analysis of crucian carp infection by atypical Aeromonas salmonicida.

Aeromonas salmonicida is a ubiquitous fish pathogen known to cause furunculosis. With the emergence of new subtypes and the expansion of the host range, it has threatened the health of a variety of marine and freshwater fish, particularly the non-salmonids, manifesting differently from the classical furunculosis. Although there have been reports of infection by atypical strains on the crucian carp, the pathogenesis and tissue pathology remain unclear. In this study, transcriptomics and histopathology were used to analyze the immune response and lesions of crucian carp infected with A. salmonicida. Comparative analysis showed 6579 differentially expressed genes (DEGs) (3428 down-regulated and 3151 up-regulated) were identified on day 5 post-infection (5 dpi). Further annotation and analysis revealed that the DEGs were enriched in enzyme regulator activity, response to oxidative stress, iron ion homeostasis and other functions, and mitogen-activated protein kinase (MAPK), nuclear factor-κB (NF-κB), toll-like receptor (TLR), and nucleotide-binding oligomerization domain (NOD)-like receptor (NLR) etc., and immune-related signaling pathways. Meanwhile, the four C-type lysozyme genes found in all DEGs were significantly up-regulated after infection. In addition, there was severe bleeding on the body of the infected fish. Also, the intestine, liver, spleen, and kidney showed varying degrees of inflammatory damage, especially the goblet cell hyperplasia of intestinal mucosa epithelium and degeneration and necrosis of renal tubular epithelium cells. Additionally, with the increase in pathogen concentration, the cumulative mortality increased, the severity of lesions in the hindgut and head-kidney tissues increased. The relative expression levels of four immune-related genes (TNF-α, IL-1ß, IL-11, C-lysozyme) were also significantly upregulated, compared with the control (P < 0.05). In conclusion, this study provides a scientific basis for further study on the immune response, pathological diagnosis, and prevention of crucian carp infection caused by atypical A. salmonicida.

Baculoviral infection reduces the expression of four allergen proteins of silkworm pupa.

Silkworm (Bombyx mori) larvae are widely used to express exogenous proteins. Moreover, some silkworm pupal proteins can be used as drug-loading materials for selfexpressed oral tolerance drugs. However, several proteins expressed in silkworm pupae cause severe allergic reactions in humans and animals. Interestingly, some baculovirus vectors have been shown to alter the host gene and its expression in insect cells, but this has not been confirmed in silkworm. Here, we analyzed the effects of infection with an empty B. mori baculovirus (BmNPV) vector on silkworm pupal protein expression. Using a proteomics approach, the allergens thiol peroxiredoxin (Jafrac1), 27-kDa glycoprotein (p27k), arginine kinase, and paramyosin as well as 32 additional differentially expressed proteins were identified. Downregulation of the messenger RNA expression of the four known allergens was observed after BmNPV infection; subsequent changes in protein expression were confirmed by the western blot analysis using polyclonal antibodies prepared with recombinant proteins of the four allergens. Collectively, these data indicate that the four known allergens of silkworm pupae can be reduced by infection ith an empty BmNPV vector to increase the safety of silkworm pupa-based exogenous protein expression and drug delivery of oral pharmaceuticals. In addition, the four recombinant allergen proteins may contribute to the diagnosis of allergic diseases of silkworm pupa.

Cementless Total Hip Arthroplasty With a High Hip Center for Hartofilakidis Type B Developmental Dysplasia of the Hip: Results of Midterm Follow-Up.

BACKGROUND: Acetabular reconstruction in adults with Hartofilakidis type B developmental dysplasia of the hip is a major technical challenge. The purpose of this retrospective study was to evaluate hip function and radiographic outcomes regarding high hip center at midterm follow-up. METHODS: From January 1, 2007 to December 31, 2009, 37 patients who had Hartofilakidis type B developmental dysplasia of the hip underwent a primary total hip arthroplasty using a high hip center technique. Functional, radiographic, and survivorship outcomes were evaluated. RESULTS: Of the 37 patients, 31 patients (83.8%) were available for the mean follow-up of 6.1 years (range, 1.5-7.6 years). Thirty-one cementless cups were located at an average vertical distance of 38.1 ± 3.3 mm and at a mean horizontal distance of 35.5 ± 3.4 mm. The mean ratio of the height of the hip center was 2.4% (range, 2.0%-2.9%). The Harris Hip Scores were improved from 50.3 points (range, 38-63 points) preoperatively to 92.3 points (range, 85-100 points) at the final follow-up (P < .001). Four patients continued to present with Trendelenburg gait pattern at the last follow-up. With use of revision for any reason and aseptic loosening as the end point, the 5-year survival rates were 90.3% (95% CI, 79.9%-100%) and 93.3% (95% CI, 84.3%-100%), respectively. CONCLUSIONS: The high hip center technique in conjunction with a cementless acetabular component seems to be a valuable alternative to achieve satisfactory midterm outcomes for Hartofilakidis type B developmental dysplasia of the hip.

Single-cell RNA sequencing reveals distinct transcriptomic profiles and evolutionary patterns in lung cancer brain metastasis.

Background: Lung cancer metastasis to the brain presents significant clinical challenges. Therefore, elucidating its underlying mechanisms and characterizing its transcriptomic landscape is essential for developing therapeutic interventions. Methods: We analyzed two distinct single-cell RNA sequencing datasets of lung cancer metastasis to analyze the evolutionary trajectory of brain metastatic tumors. In addition, a systematic comparison of cell-cell interaction between tumor cells and lymphocytes was conducted within primary and brain metastatic tumors. Results: The brain metastatic tumors showed greater transcriptomic changes (reflected by a higher pseudotime) than tumors in the lymph nodes and primary tumors. Furthermore, our investigation has not only revealed specific shared ligand-receptor pairs in both mLN and mBrain, exemplified by the interaction between SPP1 and CD99 in T cells, but has also unveiled a diverse array of ligand-receptor pairs exclusive to the mBrain. Notably, this includes distinctive pairs such as APP and IL1 observed specifically in myeloid cells. Conclusion: The distinct microenvironment in the brain may influence the observed transcriptomic changes in tumors, emphasizing the significance of the specific environment in determining tumor behavior and therapeutic response.

Nucleoporin 93, a new substrate of the E3 ubiquitin protein ligase HECTD1, promotes esophageal squamous cell carcinoma progression.

Nucleoporin 93 (NUP93) is an important component of the nuclear pore complex, exhibiting pro-tumorigenic properties in some cancers. However, its function in esophageal squamous cell carcinoma (ESCC) has not been elucidated. This study aimed to investigate the effects of NUP93 in ESCC and the underlying mechanisms involved. Through analysis of public human cancer datasets, we observed higher expression of NUP93 in esophageal cancer tissues than in normal tissues. Stable ESCC cell lines with NUP93 overexpression or knockdown were established by lentiviral vector transduction and puromycin selection. NUP93 knockdown suppressed the proliferation, colony formation, cell cycle transition, migration, and invasion of ESCC cells, while the overexpression of NUP93 displayed opposite effects. NUP93 positively regulated epithelial-mesenchymal transition and AKT signaling transduction in ESCC cells. In addition, NUP93 increased the expression of programmed death ligand 1 (PD-L1) in ESCC cells and attenuated NK cell-mediated lysis of ESCC cells. In vivo experiments demonstrated that NUP93 promotes the growth of ESCC in nude mice, enhances Ki67 and PD-L1 expression, and promotes AKT signaling transduction in xenografts. Mechanistically, we demonstrated that the HECT domain E3 ubiquitin protein ligase 1 (HECTD1) contributes to the ubiquitination and degradation of NUP93 and acts as a tumor suppressor in ESCC. To conclude, this study has shown that NUP93 has pro-tumor properties in ESCC and that HECTD1 functions as an upstream regulator of NUP93 in ESCC. These findings may contribute to the investigation of potential therapeutic targets in ESCC.

Neoadjuvant camrelizumab combined with paclitaxel and nedaplatin for locally advanced esophageal squamous cell carcinoma: a single-arm phase 2 study (cohort study).

BACKGROUND: Neoadjuvant administration of immune checkpoint inhibitors (ICIs) combined with chemotherapy demonstrated promising efficacy and manageable safety in locally advanced esophageal squamous cell carcinoma (ESCC). This prospective, single-arm, phase 2 study evaluated the efficacy and safety of neoadjuvant therapy with camrelizumab plus paclitaxel and nedaplatin for 2-4 cycles in ESCC. METHODS: Patients with locally advanced stage IIa-IIIb ESCC were enrolled in the study and received camrelizumab (200 mg), paclitaxel (155 mg/m 2 ), and nedaplatin (80 mg/m 2 ) intravenously on day one every 3 weeks. Patients underwent surgery after 2-4 cycles of treatment. The primary endpoint was the pathological complete response (pCR) rate. Secondary endpoints included the major pathological response (MPR) rate, R0 resection rate, tumor regression, objective response rate (ORR), and disease-free survival (DFS). Programmed cell death 1 ligand 1 (PD-L1) expression in tumor tissues was measured and quantified using immunohistochemistry staining and combined positive score (CPS), respectively. RESULTS: In total, 75 patients were enrolled and received neoadjuvant treatment. Of them, 45 (60%) received two cycles, 18 (24%) received three cycles, and 10 patients (13.3%) received four cycles of neoadjuvant therapy. Ultimately, 62 patients (82.7%) underwent surgery. The patients achieved a pCR of 27.4% (95% CI: 16.9-40.2), an MPR of 45.2% (95% CI: 33.1-59.2), and an ORR of 48.4% (95% CI: 35.5-61.4); all patients had an R0 resection. T and N downstaging occurred in 39 (62.9%) and 19 (30.6%) patients Moreover, patients with CPS ≥10 tended to have enhanced ORR, pCR, and MPR compared to those with CPS <10. Treatment-related adverse events (TRAEs) of grade 1-2 occurred in 59 (78.7%) patients, grade 3 TRAEs in four (5.3%), and one patient (1.3%) experienced a grade 4 TRAE. CONCLUSIONS: Neoadjuvant camrelizumab combined with chemotherapy showed promising efficacy in locally advanced ESCC, with a manageable safety profile, when administered flexibly in two to four cycles.

Expression characteristics and in vitro antibacterial properties of C-type lysozyme in crucian carp infected with Aeromonas salmonicida.

Aeromonas salmonicida is an ancient fish pathogen. Lysozymes are important molecules in the innate immune system that fight bacterial infections. The expression characteristics of C-type lysozyme in crucian carp infected with A. salmonicida and its antibacterial effect against A. salmonicida had not been investigated. Thus, we used bioinformatics to analyze the gene and protein characteristics of C-type lysozymes in crucian carp. Changes in C-type lysozyme expression before and after crucian carp infection with A. salmonicida were detected, and the in vitro antibacterial effect of recombinant carp C-type lysozyme on A. salmonicida was validated. The results showed that the coding DNA sequence region of the lysozyme gene sequence was 438 bp long, encoding 145 amino acids and containing two conserved catalytic sites: Glu53 and Asp69. Phylogenetic analysis revealed that crucian carp C-type lysozymes clustered with Cyprinus carpio lysozyme C. After crucian carp were infected with A. salmonicida, the gene and protein expression of C-type lysozymes in the liver, spleen, kidney, and hindgut were significantly upregulated, with the liver showing the highest upregulation that was 15 times higher than that in the uninfected group. In addition, recombinant C-type lysozyme exhibited significant antibacterial activity against A. salmonicida, with an average inhibition zone radius of 0.92 cm when using 40 µg recombinant lysozyme. In conclusion, this study reveals the important role of C-type lysozymes in the innate immune response of crucian carp and provides a theoretical basis for preventing crucian carp infection with A. salmonicida.

Analysis of multiple programmed cell death-related prognostic genes and functional validations of necroptosis-associated genes in oesophageal squamous cell carcinoma.

BACKGROUND: Oesophageal squamous cell carcinoma (ESCC) is a lethal malignancy. Immune checkpoint inhibitors (ICIs) showed great clinical benefits for patients with ESCC. We aimed to construct a model predicting prognosis and response to ICIs by integrating diverse programmed cell death (PCD) forms. METHODS: Genes related to 14 PCDs were collected to generate multi-gene signatures, including apoptosis, necroptosis, pyroptosis, ferroptosis, and cuproptosis. Bulk and single-cell RNA transcriptome datasets were used to develop and validate the model. We assessed the functions of two necroptosis-related genes in ESCC cells by Western blot, co-immunoprecipitation (Co-IP), LDH release assay, CCK-8, and migration assay, followed by immunohistochemistry (IHC) staining on samples of patients with ESCC (n = 67). FINDINGS: We built and validated a 16-gene prognostic combined cell death index (CCDI) by combining immunogenic cell death (ICD) and necroptosis signatures. The CCDI could also predict response to ICIs in cancer, as shown by Tumour Immune Dysfunction and Exclusion (TIDE) analysis, confirmed in four independent ICI clinical trials. Trajectory analysis revealed that HOOK1 and CUL4A might affect ESCC cell fate. We found that HOOK1 induced necroptosis and inhibited the proliferation and migration of ESCC cells, while CUL4A exhibited the opposite effects. Co-IP assay confirmed that HOOK1 and CUL4A promoted and reduced necrosome formation in ESCC cells. Data from patients with ESCC further supported that HOOK1 and CUL4A might be a tumour suppressor and oncogene, respectively. INTERPRETATION: We constructed a CCDI model with potential in predicting prognosis and response to ICIs in cancer. HOOK1 and CUL4A in the CCDI model are crucial prognostic biomarkers in ESCC. FUNDING: The Natural Science Foundation of China [82172786], The National Cancer Center Climbing Fund of China [NCC201908B06], The Natural Science Foundation of Heilongjiang Province [LH2021H077].

METTL3-mediated deficiency of lncRNA HAR1A drives non-small cell lung cancer growth and metastasis by promoting ANXA2 stabilization.

We previously reported lncRNA HAR1A as a tumor suppressor in non-small cell lung cancer (NSCLC). However, the delicate working mechanisms of this lncRNA remain obscure. Herein, we demonstrated that the ectopic expression of HAR1A inhibited the proliferation, epithelial-mesenchymal transition (EMT), migration, and invasion of NSCLC cells and enhanced paclitaxel (PTX) sensitivity in vitro and in vivo. We identified the oncogenic protein annexin 2 (ANXA2) as a potential interacting patterner of HAR1A. HAR1A overexpression enhanced ANXA2 ubiquitination and accelerated its degradation via the ubiquitin-proteasome pathway. We further uncovered that HAR1A promoted the interaction between E3 ubiquitin ligase TRIM65 and ANXA2. Moreover, the ANXA2 plasmid transfection could reverse HAR1A overexpression-induced decreases in proliferation, migration, and invasion of NSCLC cells and the activity of the NF-κB signaling pathway. Finally, we found that HAR1A loss in NSCLC might be attributed to the upregulated METTL3. The m6A modification levels of HAR1A were increased in cancer cells, while YTHDF2 was responsible for recognizing m6A modification in the HAR1A, leading to the disintegration of this lncRNA. In conclusion, we found that METTL3-mediated m6A modification decreased HAR1A in NSCLC. HAR1A deficiency, in turn, stimulated tumor growth and metastasis by activating the ANXA2/p65 axis.

Experimental and Numerical Study on Deflagration Characteristics of Large-Scale Propane-Air Mixture.

Seven deflagration tests of a propane-air mixture were carried out in a 22.5 m3 large-scale chamber. The effects of initial volume, gas concentration, and initial turbulence intensity on deflagration characteristics were analyzed. The main frequency of the explosion wave was quantitatively determined by the combination of the wavelet transform and energy spectrum analysis. The results show that the explosive overpressure is formed by the discharge of combustion products and secondary combustion, and the effects of turbulence and gas concentration on the explosive overpressure are higher than the initial volume. Under the condition of weak initial turbulence, the main frequency of gas explosion wave is between 32.13 and 48.33 Hz. Under strong initial turbulence conditions, the main frequency of the gas explosion wave increases with the increase of overpressure, and the empirical formula of the relationship between the main frequency and overpressure is summarized, which could provide theoretical support for the design of mechanical metamaterials for oil and gas explosion. Finally, the flame acceleration simulator numerical model was calibrated through tests, and the overpressure simulation values were in good agreement with the experimental data. The leakage, diffusion, and explosion of a liquefied hydrocarbon loading station in a petrochemical enterprise were simulated. The lethal distance and explosion overpressure at key buildings are predicted for different wind speed conditions. The simulation results can provide a technical basis for evaluating personnel injury and building damage.

Development of a cuproptosis-related signature for prognosis prediction in lung adenocarcinoma based on WGCNA.

Background: Cuproptosis is a novel mitochondrial respiration-dependent cell death mechanism induced by copper that can kill cancer cells via copper carriers in cancer therapy. However, the clinical significance and prognostic value of cuproptosis in lung adenocarcinoma (LUAD) remains unclear. Methods: We performed a comprehensive bioinformatics analysis of the cuproptosis gene set, including copy number aberration, single-nucleotide variation, clinical characteristics, survival analysis, etc. Cuproptosis-related gene set enrichment scores (cuproptosis Z-scores) were calculated in The Cancer Genome Atlas (TCGA)-LUAD cohort using single-sample gene set enrichment analysis (ssGSEA). Modules significantly associated with cuproptosis Z-scores were screened by weighted gene co-expression network analysis (WGCNA). The hub genes of the module were then further screened by survival analysis and least absolute shrinkage and selection operator (LASSO) analysis, in which TCGA-LUAD (497 samples) and GSE72094 (442 samples) were used as the training and validation cohorts, respectively. Finally, we analyzed the tumor characteristics, immune cell infiltration levels, and potential therapeutic agents. Results: Missense mutation and copy number variant (CNV) events were general in the cuproptosis gene set. We identified 32 modules, of which the MEpurple (107 genes) and MEpink (131 genes) modules significantly positively and negatively correlated with cuproptosis Z-scores, respectively. We identified 35 hub genes significantly related to overall survival and constructed a prognostic model consisting of 7 cuproptosis-related genes in patients with LUAD. Compared with the low-risk group, patients in the high-risk group had a worse overall survival and gene mutation frequency, as well as significantly higher tumor purity. In addition, infiltration of immune cells was also significantly different between the 2 groups. Furthermore, the correlation between the risk scores and half-maximum inhibitory concentration (IC50) of antitumor drugs in the Genomics of Drug Sensitivity in Cancer (GDSC) v. 2 database was explored, revealing differences in drug sensitivity across the 2 risk groups. Conclusions: Our study provided a valid prognostic risk model for LUAD and improved understanding of its heterogeneity, which may aid in the development of personalized treatment strategies.

Thermal runaway features of large-format power lithium-ion cells under various thermal abuse patterns and capacities.

Herein, a comprehensive investigation is performed to research the thermal runaway features of large-format power lithium-ion cells under various heating patterns (2 kW electric heating oven and 600 W electric heating plate) and capacities (60, 150, and 180 Ah). Although the electric heating plate induces the cell to encounter thermal runaway earlier in comparison with the electric heating oven, the combustion does not appear for the former case since the compact stacking of the electric heating plate restrains the heat release of the heater such that the surrounding temperature is too low to induce the ignition of the thermal runaway combustibles. Besides that, it is interesting to find that the color of the ejected products under the electric heating plate condition becomes shallower as the thermal runaway proceeds, which implies that the ejecta in the initial of thermal runaway is mixed with quantities of solid particles and the proportion would gradually decrease. With the increase of the cell capacity, thermal runaway emerges later as a result of the greater cell height which delays the cell temperature rise, when exposed to an electric heating oven. In addition, the cell with a larger capacity demonstrates a lower peak temperature, a lower maximum temperature rise rate, a shorter combustion, a lower flame temperature, and a weaker radiation heat strength during thermal runaway; that is, less heat is released due to its violent thermal runaway behaviour. Finally, the severe explosion risk for the larger-capacity cell should be especially noted considering the larger amount of explosive gases released.