Artificial selection on storage protein 1 possibly contributes to increase of hatchability during silkworm domestication.

Like other domesticates, the efficient utilization of nitrogen resources is also important for the only fully domesticated insect, the silkworm. Deciphering the way in which artificial selection acts on the silkworm genome to improve the utilization of nitrogen resources and to advance human-favored domestication traits, will provide clues from a unique insect model for understanding the general rules of Darwin's evolutionary theory on domestication. Storage proteins (SPs), which belong to a hemocyanin superfamily, basically serve as a source of amino acids and nitrogen during metamorphosis and reproduction in insects. In this study, through blast searching on the silkworm genome and further screening of the artificial selection signature on silkworm SPs, we discovered a candidate domestication gene, i.e., the methionine-rich storage protein 1 (SP1), which is clearly divergent from other storage proteins and exhibits increased expression in the ova of domestic silkworms. Knockout of SP1 via the CRISPR/Cas9 technique resulted in a dramatic decrease in egg hatchability, without obvious impact on egg production, which was similar to the effect in the wild silkworm compared with the domestic type. Larval development and metamorphosis were not affected by SP1 knockout. Comprehensive ova comparative transcriptomes indicated significant higher expression of genes encoding vitellogenin, chorions, and structural components in the extracellular matrix (ECM)-interaction pathway, enzymes in folate biosynthesis, and notably hormone synthesis in the domestic silkworm, compared to both the SP1 mutant and the wild silkworm. Moreover, compared with the wild silkworms, the domestic one also showed generally up-regulated expression of genes enriched in the structural constituent of ribosome and amide, as well as peptide biosynthesis. This study exemplified a novel case in which artificial selection could act directly on nitrogen resource proteins, further affecting egg nutrients and eggshell formation possibly through a hormone signaling mediated regulatory network and the activation of ribosomes, resulting in improved biosynthesis and increased hatchability during domestication. These findings shed new light on both the understanding of artificial selection and silkworm breeding from the perspective of nitrogen and amino acid resources.

New insight into the mechanism underlying the silk gland biological process by knocking out fibroin heavy chain in the silkworm.

BACKGROUND: Exploring whether and how mutation of silk protein contributes to subsequent re-allocation of nitrogen, and impacts on the timing of silk gland degradation, is important to understand silk gland biology. Rapid development and wide application of genome editing approach in the silkworm provide us an opportunity to address these issues. RESULTS: Using CRISPR/Cas9 system, we successfully performed genome editing of Bmfib-H. The loss-of-function mutations caused naked pupa and thin cocoon mutant phenotypes. Compared with the wild type, the posterior silk gland of mutant showed obviously degraded into fragments in advance of programmed cell death of silk gland cells. Comparative transcriptomic analyses of silk gland at the fourth day of the fifth instar larval stage(L5D4)identified 1456 differential expressed genes (DEGs) between posterior silk gland (PSG) and mid silk gland (MSG) and 1388 DEGs between the mutant and the wild type. Hierarchical clustering of all the DEGs indicated a remarkable down-regulated and an up-regulated gene clade in the mutant silk glands, respectively. Down-regulated genes were overrepresented in the pathways involved in cancer, DNA replication and cell proliferation. Intriguingly, up-regulated DEGs are significantly enriched in the proteasome. By further comparison on the transcriptome of MSG and PSG between the wild type and the mutant, we consistently observed that up-regulated DEGs in the mutant PSG were enriched in protein degrading activity and proteasome. Meantime, we observed a series of up-regulated genes involved in autophagy. Since these protein degradation processes would be normally occur after the spinning time, the results suggesting that these progresses were activated remarkably ahead of schedule in the mutant. CONCLUSIONS: Accumulation of abnormal fib-H protein might arouse the activation of proteasomes as well as autophagy process, to promote the rapid degradation of such abnormal proteins and the silk gland cells. Our study therefore proposes a subsequent process of protein and partial cellular degradation caused by mutation of silk protein, which might be helpful for understanding its impact of the silk gland biological process, and further exploration the re-allocation of nitrogen in the silkworm.

Anoikis-related genes as potential prognostic biomarkers in gastric cancer: A multilevel integrative analysis and predictive therapeutic value.

BACKGROUND: Gastric cancer (GC) is a frequent malignancy of the gastrointestinal tract. Exploring the potential anoikis mechanisms and pathways might facilitate GC research. PURPOSE: The authors aim to determine the significance of anoikis-related genes (ARGs) in GC prognosis and explore the regulatory mechanisms in epigenetics. METHODS: After describing the genetic and transcriptional alterations of ARGs, we searched differentially expressed genes (DEGs) from the cancer genome atlas and gene expression omnibus databases to identify major cancer marker pathways. The non-negative matrix factorisation algorithm, Lasso, and Cox regression analysis were used to construct a risk model, and we validated and assessed the nomogram. Based on multiple levels and online platforms, this research evaluated the regulatory relationship of ARGs with GC. RESULTS: Overexpression of ARGs is associated with poor prognosis, which modulates immune signalling and promotes anti-anoikis. The consistency of the DEGs clustering with weighted gene co-expression network analysis results and the nomogram containing 10 variable genes improved the clinical applicability of ARGs. In anti-anoikis mode, cytology, histology, and epigenetics could facilitate the analysis of immunophenotypes, tumour immune microenvironment (TIME), and treatment prognosis. CONCLUSION: A novel anoikis-related prognostic model for GC is constructed, and the significance of anoikis-related prognostic genes in the TIME and the metabolic pathways of tumours is initially explored.

METTL3-mediated m6A methylation regulates ovarian cancer progression by recruiting myeloid-derived suppressor cells.

BACKGROUND: Ovarian cancer (OC) typically develops an immunosuppressive microenvironment by funtional changes of host immune cells. Dysregulated m6A level is associated with cancer progression via the intrinsic oncogenic pathways. However, the role of m6A in regulating host immune cell function during anti-tumor immunity needs comprehensive analysis. This study aimed to investigate the role of METTL3, a catalytic subunit of the methyltransferase complex, in regulating host immune cell response against OC. METHODS: In this study, myeloid-specific Mettl3 gene knockout (Mettl3-cKO) mice were bred using the Cre-LoxP system. Intraperitoneally injection of ID8 cells was used as a syngeneic OC model. Furthermore, the compositions of immune cell populations were analyzed by flow cytometry and single-cell sequencing. Moreover, chemokines and cytokines secretion were assessed using ELISA. Lastly, the role of METTL3 in regulating IL-1ß secretion and inflammasome activation in bone marrow-derived macrophages cocultured with ID8 cells was specified by ELISA and immunoblotting. RESULTS: It was revealed that OC cell growth was enhanced in Mettl3-cKO mice. Furthermore, a shift of decreased M1 to increased M2 macrophage polarization was observed during OC progression. Moreover, Mettl3 depletion in myeloid lineage cells increased secretion of CCL2 and CXCL2 in peritoneal lavage fluild. Interestingly, Mettl3 deficiency enhanced IL-1ß secretion induced by viable ID8 cells independent of inflammasome activation and cell death. Therefore, OC cells in tumor-bearing mice trigger a slight inflammatory response with a low-to-moderate secretion of pro-inflammatory cytokines and chemokines. CONCLUSION: This study provides new insights into METTL3-mediated m6A methylation, which regulates host immune response against OC.

In Situ Tyrosinase Monitoring by Wearable Microneedle Patch toward Clinical Melanoma Screening.

Despite the potential indicating role of tyrosinase (TYR) in cutaneous melanoma, how to capture the real changes of TYR in suspicious skin remains a major challenge. Unlike the traditional human serum test, this study reports a sensing platform that incorporates a wearable microneedle (MN) patch and trimetallic Au@Ag-Pt nanoparticles (NPs) for surface-enhanced Raman scattering (SERS) and colorimetric dual-mode detecting TYR in human skin in situ toward potential melanoma screening. In the presence of TYR, catechol immobilized on MN is preferentially oxidized to benzoquinone, which competitively impedes the interaction of MN and Au@Ag-Pt NPs, triggering the SERS-colorimetric signal reciprocal switch. Using a B16F10 mouse melanoma model, our platform is capable of noninvasively piercing the skin surface and detecting TYR levels before and during anti-PD-1 antibody treatment, which would be highly informative for prognostic judgment and illness monitoring of melanoma. Through in situ sensing for capturing the metabolic changes of TYR in advance, this platform was successfully applied to discriminate the melanoma subjects from skin moles and normal ones (p < 0.001), as well as screen potential melanoma from lactate dehydrogenase (LDH)-negative patients. Melanoma growth and prognosis can still be monitored through recording the continuous change of TYR levels. More importantly, the well-defined flexible and stretchable characteristics of the MN patch allow robustly adhering to the skin without inducing chemical or physical irritation. We believe this platform integrating MN-based in situ sensing, TYR responsiveness, and SERS/colorimetric dual-readout strategy will have high clinical importance in early diagnosis and monitoring of cutaneous melanoma.

Identification of Ferroptosis-Associated Long Noncoding RNA Prognostic Model and Tumor Immune Microenvironment in Thyroid Cancer.

Background: Thyroid cancer (TC) is a rapidly increasing incidence of endocrine malignancies, occupying 3% of new cancer incidence, of which 10% has a heterogeneous prognosis. Ferroptosis is a form of cell death distinct from apoptosis, which involves antitumor drug-related research. Long noncoding RNAs (lncRNAs) could affect cancer prognosis by regulating the ferroptosis; thus, ferroptosis-associated lncRNAs are emerging as prospective biomarkers for cancer therapy and prognosis. However, the prognostic factors of ferroptosis-associated lncRNAs in this solid tumor and their mechanisms remain unknown. Methods: The TC lncRNA data were extracted from RNA sequencing files of The Cancer Genome Atlas (TCGA). Then, we performed a two-cluster analysis and grouped 502 patients with TC in a 7 : 3 ratio. Both the least absolute shrinkage and selection operator (LASSO) regression and Cox regression analysis were conducted to create and validate the ferroptosis-associated lncRNA prognostic model (Ferr-LPM). Based on the median Ferr-LPM-based risk score (LPM_score) of the training cohort, we categorized patients into high and low LPM_score groups, which were then subjected to prognostic correlation and difference analysis. We also created a nomogram and assessed its predictive ability. Furthermore, immune-related mechanisms were investigated by analyzing the tumor immune microenvironment (TIME) and applying algorithms such as CIBERSROT. Results: We built a highly accurate nomogram to promote the clinical applicability of Ferr-LPM. The area under the receiver operating characteristic curve (AUC-ROC) reached above 0.9. Survival analysis suggested that when the Ferr-LPM score was higher, the overall survival (OS) of patients within this group was shorter. Meanwhile, we found a strong association between Ferr-LPM and TIME. Interestingly, the LPM_score was inversely proportional to the tumor purity but positively related to immune checkpoint blockade (ICB) response. Conclusion: We constructed a novel ferroptosis-associated lncRNA nomogram that could highly predict the prognosis of TC patients. Ferroptosis-associated lncRNAs might possess potential functions in regulating TIME, and lncRNAs provide TC patients with new prognostic biomarkers and therapeutic targets.

MAIP1-Related Tumor Immune Infiltration: As a Potential Prognostic Biomarker for Esophageal Cancer.

Background: Esophageal cancer (EC), a common malignant tumor of digestive tract, is also one of the most deadly cancers. Accumulating studies have shown that the initiating and progressing multiple human diseases were closely related to the expression of MAIP. However, the specific roles and mechanisms of MAIP1 in EC remain incompletely defined. Purpose: This study aims to determine the clinical significance of MAIP1 in EC and explores its potential molecular mechanisms regulating tumor immune infiltration. Methods: We obtained RNA-seq datasets and corresponding clinical data for EC patients from the Cancer Genome Atlas (TCGA) database via the UCSC Xena browser to extract MAIP1 expression and plot survival curves to determine their prognosis. Based on the differential expression of MAIP1, EC patients were divided into high and low group to investigate the mechanism of MAIP1 in EC. In addition, the single sample gene set enrichment analysis (ssGSEA) quantified the expression of various immune cell signature marker genes and assessed the degree of immune infiltration in EC. Results: In the TCGA-EC cohort, the overexpression of MAIP1 was observed in tumor tissues compared to normal tissues (p = 0.0038). Overall survival analysis showed that EC patients with the overexpression of MAIP1 presented a lower overall survival and worse prognosis (p = 0.004). Enrichment analysis revealed that the differential genes (DEGs) between high and low group are involved in biological functions such as extracellular matrix and organization extracellular structure. The results of ssGSEA showed that DCs, iDCs, macrophages, mast cells, and NK cells were significantly different in MAIP1high and MAIP1low groups, and all showed high expression in the MAIP1low group. Conclusion: We proposed that MAIP1 overexpression was associated with poor prognosis and tumor immune infiltration in EC. At present, there are few MAIP1-related tumor immune infiltration studies in EC, and further investigation is needed.

Potential Therapeutic Application of Local Anesthetics in Cancer Treatment.

Local anesthetics are voltage-gated sodium channel blockers primarily administered locally or to the innervating nerves for anesthetic or analgesic purposes. In vitro studies have found direct effects of local anesthetics on cancer cells, such as impact on cancer cell proliferation, apoptosis, migration, invasion, and chemosensitivity, by multiple mechanisms. So far, in vivo evidence regarding the effect of local anesthetics on cancer cell lines is relatively lacking. Local and regional anesthesia administration has been reported to reduce postoperative pain and opioid use in cancer treatment. Additionally, regional anesthesia may reduce the perioperative stress response. However, the clinical therapeutic application of local anesthetics in cancer remains exploratory. In this review, we will discuss the direct and indirect effects of local anesthetics on cancer cells, and discuss the current evidence related to the use of local anesthetics in the treatment of cancer.

Prognostic Model and Immune Infiltration of Ferroptosis Subcluster-Related Modular Genes in Gastric Cancer.

Background: Gastric cancer (GC) is one of the gastrointestinal tumors with the highest mortality rate. The number of GC patients is still high. As a way of iron-dependent programmed cell death, ferroptosis activates lipid peroxidation and accumulates large reactive oxygen species. The role of ferroptosis in GC prognosis was underrepresented. The objective was to investigate the role of ferroptosis-related genes (FRGs) in the prognosis and development of GC. Methods: Datasets of GC patients were obtained from the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) database that include clinical information and RNA seq data. Through nonnegative matrix factorization (NMF) clustering, we identified and unsupervised cluster analysis of the expression matrix of FRGs. And we constructed the co-expression network between genes and clinical characteristics by consensus weighted gene co-expression network analysis (WGCNA). The prognostic model was constructed by univariate and multivariate regression analysis. The potential mechanisms of development and prognosis in GC were explored by Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, gene ontology (GO), tumor immune microenvironment (TIME), and tumor mutation burden (TMB). Results: Two molecular subclusters with different expression patterns of FRGs were identified, which have significantly different survival states. Ferroptosis subcluster-related modular genes were identified by WGCNA. Based on 8 ferroptosis subcluster-related modular genes (collagen triple helix repeat containing 1 (CTHRC1), podoplanin (PDPN), procollagen-lysine,2-oxoglutarate 5-dioxygenase 2 (PLOD2), glutamine-fructose-6-phosphate transaminase 2 (GFPT2), ATP-binding cassette subfamily A member 1 (ABCA1), G protein-coupled receptor 176 (GPR176), serpin family E member 1 (SERPINE1), dual specificity phosphatase 1 (DUSP1)) and clinicopathological features, a nomogram was constructed and validated for their predictive efficiency on GC prognosis. Through receiver operating characteristic (ROC) analysis, the results showed that the area under the curve (AUC) of 1-, 3-, and 5-year survival were 0.721, 0.747, and 0.803, respectively, indicating that the risk-scoring model we constructed had good prognosis efficacy in GC. The degree of immune infiltration in high-risk group was largely higher than low-risk group. It indicated that the immune cells have a good response in high-risk group of GC. The TMB of high-risk group was higher, which could generate more mutations and was more conducive to the body's resistance to the development of cancer. Conclusion: The risk-scoring model based on 8 ferroptosis subcluster-related modular genes has shown outstanding advantages in predicting patient prognosis. The interaction of ferroptosis in GC development may provide new insights into exploring molecular mechanisms and targeted therapies for GC patients.

Burden of atrial fibrillation and its attributable risk factors from 1990 to 2019: An analysis of the Global Burden of Disease study 2019.

Background: Understanding the pattern and trend of the atrial fibrillation (AF) burden are essential for developing effective preventive strategies. The purpose of this study was to estimate AF burdens and risk factors in 204 countries and territories between 1990 and 2019. Materials and methods: Data were extracted from the Global Burden of Disease 2019, including incidence, death, disability-adjusted life-years (DALYs), and the attributable risk factors. In order to quantify changes in the age-standardized incidence rate (ASIR), age-standardized death rate (ASDR), and age-standardized DALY rate between 1990 and 2019, the estimated annual percentage change (EAPC) was used. Also, AF burden was assessed in relation to the Socio-demographic Index (SDI). Results: Globally, there were 4,720,324 incident cases, 117,038 deaths and 8,393,635 DALYs in 2019. There were no significant changes in ASIR, ASDR, or age-standardized DALY rates from 1990 to 2019. Although the burden and trend of AF varied in different regions and countries, the ASIR, ASDR and age-standardized DALY rate were positively correlated with SDI. Furthermore, the burden of AF was higher in males and elderly. The age-standardized DALY rate worldwide was primarily attributable to high systolic blood pressure, followed by high body-mass index, alcohol use, smoking, diet high in sodium and lead exposure. Conclusion: AF remained a major public health challenge worldwide, with substantial variation at regional and national levels. There is an urgent need to increase public awareness about AF risk factors and to bring about cost-effective interventions for AF in order to reduce its future burden.

Multifunctional Au nano-bridged nanogap probes as ICP-MS/SERS dual-signal tags and signal amplifiers for bacteria discriminating, quantitative detecting and photothermal bactericidal activity.

Ultra-sensitive detection of pathogenic bacteria is of great significance in the early stage of bacterial infections and treatment. In this work, we report a novel strategy using multifunctional Au nano-bridged nanogap nanoparticles (Au NNPs)-based sandwich nanocomposites, that made of Concanavalin A-conjugated Fe3O4@SiO2 NPs (ConA-Fe3O4@SiO2 NPs)/bacteria/aptamer-modified Au NNPs (apt-Au NNPs), for bacteria discrimination and quantitative detection by surface-enhanced Raman scattering (SERS) and inductively coupled plasma mass spectrometry (ICP-MS), and subsequently photothermal antibacterial assay. The sandwich nanocomposite consists of ConA-Fe3O4@SiO2 NPs to magnetically enrich and photothermal killing bacteria, and dual-signal tags of apt-Au NNPs for both SERS sensing and ICP-MS quantification. This strategy can specifically distinguish different kinds of pathogenic bacteria, and provided a good linear relationship of Staphylococcus aureus (S. aureus) in the range from 50 to 104 CFU/mL with a detection limit of 11 CFU/mL, as well as realized ultralow amounts of bacterial detection in serum sample with high accuracy. Based on the quantitative detection, high antibacterial efficiency was monitored by ICP-MS. Overall, the established method combines bacteria discrimination, quantitative detection, and photothermal elimination with a simple and rapid process, which provides a novel way for the early diagnosis and treatment of bacterial infection.

Tumor targeting and penetrating biomimetic mesoporous polydopamine nanoparticles facilitate photothermal killing and autophagy blocking for synergistic tumor ablation.

The synergistic manipulation of autophagy blocking with tumor targeting and penetration effects to enhance cancer cell killing during photothermal therapy (PTT) remains a substantial challenge. Herein, we fabricated a biomimetic nanoplatform by precisely coating homologous prostate cancer cell membranes (CMs) onto the surface of mesoporous polydopamine nanoparticles (mPDA NPs) encapsulating the autophagy inhibitor chloroquine (CQ) for synergistically manipulating PTT and autophagy for anticancer treatment. The resulting biomimetic mPDA@CMs NPs-CQ system could escape macrophage phagocytosis, overcome the vascular barrier, and home in the homologous prostate tumor xenograft with high tumor targeting and penetrating efficiency. The mPDA NPs core endowed the mPDA@CMs NPs-CQ with good photothermal capability to mediate PTT killing of prostate cancer cells, while NIR-triggered CQ release from the nanosystem further arrested PTT-induced protective autophagy of cancer cells, thus weakening the resistance of prostate cancer cells to PTT. This combined PTT killing and autophagy blocking anticancer strategy could induce significant autophagosome accumulation, ROS generation, mitochondrial damage, endoplasmic reticulum stress, and apoptotic signal transduction, which finally results in synergistic prostate tumor ablation in vivo. This prostate cancer biomimetic nanosystem with synergistically enhanced anticancer efficiency achieved by manipulating PTT killing and autophagy blocking is expected to serve as a more effective anticancer strategy against prostate cancer. STATEMENT OF SIGNIFICANCE: Autophagy is considered as one of the most efficient rescuer and reinforcement mechanisms of cancer cells against photothermal therapy (PTT)-induced cancer cell eradication. How to synergistically manipulate autophagy blocking with significant tumor targeting and penetration to enhance PTT-mediated cancer cell killing remains a substantial challenge. Herein, we fabricated a biomimetic nanoplatform by precisely coating homologous cancer cell membranes onto the surface of mesoporous polydopamine nanoparticles with encapsulation of the autophagy inhibitor chloroquine for synergistic antitumor treatment with high tumor targeting and penetrating efficiency both in vitro and in vivo. This biomimetic nanosystem with synergistically enhanced anticancer efficiency by manipulating PTT killing and autophagy blocking is expected to serve as a more effective anticancer strategy.

Role of juvenile hormone receptor Methoprene-tolerant 1 in silkworm larval brain development and domestication.

The insect brain is the central part of the neurosecretory system, which controls morphology, physiology, and behavior during the insect's lifecycle. Lepidoptera are holometabolous insects, and their brains develop during the larval period and metamorphosis into the adult form. As the only fully domesticated insect, the Lepidoptera silkworm Bombyx mori experienced changes in larval brain morphology and certain behaviors during the domestication process. Hormonal regulation in insects is a key factor in multiple processes. However, how juvenile hormone (JH) signals regulate brain development in Lepidoptera species, especially in the larval stage, remains elusive. We recently identified the JH receptor Methoprene tolerant 1 ( Met1) as a putative domestication gene. How artificial selection on Met1 impacts brain and behavioral domestication is another important issue addressing Darwin's theory on domestication. Here, CRISPR/Cas9-mediated knockout of Bombyx Met1 caused developmental retardation in the brain, unlike precocious pupation of the cuticle. At the whole transcriptome level, the ecdysteroid (20-hydroxyecdysone, 20E) signaling and downstream pathways were overactivated in the mutant cuticle but not in the brain. Pathways related to cell proliferation and specialization processes, such as extracellular matrix (ECM)-receptor interaction and tyrosine metabolism pathways, were suppressed in the brain. Molecular evolutionary analysis and in vitro assay identified an amino acid replacement located in a novel motif under positive selection in B. mori, which decreased transcriptional binding activity. The B. mori MET1 protein showed a changed structure and dynamic features, as well as a weakened co-expression gene network, compared with B. mandarina. Based on comparative transcriptomic analyses, we proposed a pathway downstream of JH signaling (i.e., tyrosine metabolism pathway) that likely contributed to silkworm larval brain development and domestication and highlighted the importance of the biogenic amine system in larval evolution during silkworm domestication.

Genetic Mapping of Climbing and Mimicry: Two Behavioral Traits Degraded During Silkworm Domestication.

Behavioral changes caused by domestication in animals are an important issue in evolutionary biology. The silkworm, Bombyx mori, is an ideal fully domesticated insect model for studying both convergent domestication and behavior evolution. We explored the genetic basis of climbing for foraging and mimicry, two degraded behaviors during silkworm domestication, in combination of bulked segregant analysis (BSA) and selection sweep screening. One candidate gene, ASNA1, located in the 3-5 Mb on chromosome 19, harboring a specific non-synonymous mutation in domestic silkworm, might be involved in climbing ability. This mutation was under positive selection in Lepidoptera, strongly suggesting its potential function in silkworm domestication. Nine candidate domesticated genes related to mimicry were identified on chromosomes 13, 21, and 27. Most of the candidate domesticated genes were generally expressed at higher levels in the brain of the wild silkworm. This study provides valuable information for deciphering the molecular basis of behavioral changes associated with silkworm domestication.