Journal of advanced research最新文献

Introduction: Homologous recombination repair during meiosis is essential for the exchange of genetic information between sister chromosomes, underpinning spermatogenesis and, consequently, fertility. The disruption of this process can lead to infertility, highlighting the importance of identifying the molecular actors involved.

Objectives: This study aims to elucidate the role of the E3 ubiquitin ligase Rnf126 in spermatogenesis and its impact on fertility, particularly through its involvement in meiotic homologous recombination repair.

Methods: We used heterozygous and homozygous Rnf126 deletion models in mouse testes to examine the consequences on testicular health, sperm count, and the process of spermatogenesis. Additionally, we explored the association between RNF126 gene missense variants and nonobstructive male infertility in patients, with a focus on their functional impact on the protein's ubiquitin ligase activity.

Results: Rnf126 deletion led to testicular atrophy, disrupted seminiferous tubule structure, reduced sperm count, and spermatogenesis arrest at meiotic prophase I. Furthermore, male mice exhibited impaired homologous recombination repair and increased apoptosis within the seminiferous tubules. We identified four missense variants of the RNF126 (V68M, R241H, E261A, D253N) associated with male infertility. Specifically, the E261A and D253N variants, located in the RING domain, directly compromised the E3 ubiquitin ligase activity of RNF126.

Conclusion: Our findings demonstrate the pivotal role of RNF126 in maintaining spermatogenesis and fertility, offering insights into the molecular mechanisms underlying male infertility. The identified RNF126 variants present novel targets for diagnostic and therapeutic strategies in treating nonobstructive male infertility.

Introduction: Breast cancer, a heterogeneous disease, is influenced by multiple genetic and epigenetic factors. The majority of prognostic models for breast cancer focus merely on the main effects of predictors, disregarding the crucial impacts of gene-gene interactions on prognosis.

Objectives: Using DNA methylation data derived from nine independent breast cancer cohorts, we developed an independently validated prognostic prediction model of breast cancer incorporating epigenetic biomarkers with main effects and gene-gene interactions (ARTEMIS) with an innovative 3-D modeling strategy. ARTEMIS was evaluated for discrimination ability using area under the receiver operating characteristics curve (AUC), and calibration using expected and observed (E/O) ratio. Additionally, we conducted decision curve analysis to evaluate its clinical efficacy by net benefit (NB) and net reduction (NR). Furthermore, we conducted a systematic review to compare its performance with existing models.

Results: ARTEMIS exhibited excellent risk stratification ability in identifying patients at high risk of mortality. Compared to those below the 25th percentile of ARTEMIS scores, patients with above the 90th percentile had significantly lower overall survival time (HR = 15.43, 95% CI: 9.57-24.88, P = 3.06 × 10^-29). ARTEMIS demonstrated satisfactory discrimination ability across four independent populations, with pooled AUC_3-year = 0.844 (95% CI: 0.805-0.883), AUC_5-year = 0.816 (95% CI: 0.775-0.857), and C-index = 0.803 (95% CI: 0.776-0.830). Meanwhile, ARTEMIS had well calibration performance with pooled E/O ratio 1.060 (95% CI: 1.038-1.083) and 1.090 (95% CI: 1.057-1.122) for 3- and 5-year survival prediction, respectively. Additionally, ARTEMIS is a clinical instrument with acceptable cost-effectiveness for detecting breast cancer patients at high risk of mortality (P_t = 0.4: NB_3-year = 19‰, NB_5-year = 62‰; NR_3-year = 69.21%, NR_5-year = 56.01%). ARTEMIS has superior performance compared to existing models in terms of accuracy, extrapolation, and sample size, as indicated by the systematic review. ARTEMIS is implemented as an interactive online tool available at http://bigdata.njmu.edu.cn/ARTEMIS/.

Conclusion: ARTEMIS is an efficient and practical tool for breast cancer prognostic prediction.

Introduction: The efficiency of zinc oxide (ZnO) nanoparticles for environmental decontamination is limited by their reliance on ultraviolet (UV) light and rapid charge carrier recombination. Carbon doping has been proposed to address these challenges by potentially enhancing visible light absorption and charge separation.

Objectives: This study aims to introduce a novel, single-step synthesis method for carbon-doped ZnO (C-Z) nanoparticles, leveraging the decomposition of zinc nitrate hexahydrate and furfural under a nitrogen atmosphere to improve photocatalytic activity under visible light.

Methods: A series of C-Z variants (C-Z-1 to C-Z-5) and an undoped sample (ZnO-0) were synthesized. The influence of furfural on the synthesis process and doping mechanism was analyzed by X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD), and UV-visible diffuse reflectance spectroscopy (DRS).

Results: XPS confirmed the integration of carbon within the ZnO matrix, and XRD indicated increased lattice dimensions owing to doping. DRS revealed bandgap narrowing, suggesting enhanced charge separation. Among the variants, C-Z-3 significantly outperformed the others, showing a 12-fold increase in the photocatalytic degradation rate of Rhodamine B compared to undoped ZnO.

Conclusion: The developed single-step synthesis method for C-Z nanoparticles represents a major advancement in materials engineering for ecological applications. The enhanced photocatalytic activity under visible light, as demonstrated by C-Z-3, underscores the potential of these nanoparticles for environmental decontamination.

Introduction: Cetuximab (CTX) is an effective targeted drug for the treatment of metastatic colorectal cancer, but it is effective only in patients with wild-type KRAS genes. Even in this subset of patients, the sensitivity of CTX in patients with right hemi-colon cancer is much lower than that in patients with left hemi-colon cancer. This significantly limits its clinical application. Therefore, further elucidation of the underlying molecular mechanisms is needed. N-myc downstream-regulated gene 1 (NDRG1) plays an important role in solid tumor invasion and metastasis, but whether it can influence CTX sensitivity has not been thoroughly investigated.

Objective: Our study aimed to identify a novel mechanism by which NDRG1 affects CTX sensitivity.

Methods: Through mass spectrometry analysis of our previously constructed CTX-resistant RKO and HCT116 cells, we found that the signal transducer and activator of transcription-1 (Stat1) might be a potential target of NDRG1. By knocking out NDRG1 or/and Stat1 genes, we then applied the loss-of-function experiments to explore the regulatory relationship between NDRG1 and Stat1 and their roles in the cell cycle, epithelial-mesenchymal transition (EMT), and the sensitivity to CTX in these two colorectal cancer (CRC) cells. Finally, we used the nude-mouse transplanted tumor model and human CRC samples to verify the expression of NDRG1 and Stat1 and their impact on CTX sensitivity in vivo.

Results: Stat1 was upregulated in CTX-resistant cells, whereas NDRG1 was downregulated. Mechanically, NDRG1 was inversely correlated with Stat1 expression. It suppressed CRC cell proliferation, migration, and invasion, and promoted apoptosis and epithelial-mesenchymal transition (EMT) by inhibiting Stat1. In addition, NDRG1 directly interacted with Stat1 and promoted Smurf1-induced Stat1 ubiquitination. Importantly, this novel NDRG1-dependent regulatory loop also enhanced CTX sensitivity both in vitro and in vivo.

Conclusion: Our study revealed that NDRG1 enhanced the sensitivity to Cetuximab by inhibiting Stat1 expression and promoting its ubiquitination in colorectal cancer, elucidating NDRG1 might be a potential therapeutic target for refractory CTX-resistant CRC tumors. But its clinical value still needs to be validated in a larger sample size as well as a different genetic background.

Introduction: Angiogenesis plays a significant role in the development of tumor progression and inflammatory diseases. The role of IL-28A in angiogenesis and its precise regulatory mechanisms remain rarely elucidated.

Objectives: We report the novel regulatory role of IL-28A in physiological angiogenesis. The study aimed to elucidate the regulatory mechanisms involved in IL-28A-mediated angiogenesis and identify key genes associated with IL-28A-induced angiogenic responses.

Methods: To know the effect of IL-28A on angiogenesis, HUVECs were applied to perform proliferation, migration, invasion, tube formation, immunoblot, and EMSA. Gene expression changes in HUVECs following IL-28A treatment were analyzed by NGS. The functional role of HSP70-1 and IL-10Rβ in IL-28A-induced angiogenic responses was evaluated using PCR and siRNA knockdown. Animal studies were conducted by aortic ring ex vivo assays, Matrigel plug in vivo assays, and immunochemistry using HSP70-1 knockout and transgenic mice models. The efficacy of IL-28A in angiogenesis was confirmed in a hind-limb ischemia model.

Results: Autocrine/paracrine actions in HUVECs regulated IL-28A protein expression. Exogenous IL-28A increased the proliferation of HUVECs via eNOS/AKT and ERK1/2 signaling. IL-28A treatment promoted migration, invasion, and capillary tube formation of HUVECs through induction of the AP-1/NF-κB/MMP-2 network, which was associated with eNOS/AKT and ERK1/2 signaling. The efficacy of IL-28A-induced angiogenic potential was confirmed by aortic ring and Matrigel plug assay. HSP70-1 was identified as an IL-28A-mediated angiogenic effector gene using bioinformatics. Knockdown of HSP70-1 abolished angiogenic responses and eNOS/AKT signaling in IL-28A-treated HUVECs. IL-28A-induced microvessel sprouting formation was testified in HSP70-1-deficient and HSP70-1 transgenic mice. Flow recovery in hind-limb ischemia mice was accelerated by IL-28A injection. Finally, ablation of the IL-10Rβ gene impeded the angiogenic responses and eNOS/AKT signaling stimulated by IL-28A in HUVECs.

Conclusion: HSP70-1 drives the progression of angiogenesis by the IL-28A/IL-10Rβ axis via eNOS/AKT signaling and the AP-1/NF-κB/MMP-2 network.

Introduction: Exosome-miR-146a is significantly increased in patients with Atherosclerosis (AS), but its mechanism and effect on AS have not been fully elucidated.

Objectives: To explore the change rule and mechanism of exosomes release, and the role and molecular mechanism of exosome-miR-146a in AS.

Methods: We isolated and identified exosomes from THP-1 macrophages after treating them with ox-LDL. Then used co-immunoprecipitation and silver staining to identify the proteins involved in regulating exosome release. PKH67 was used to label exosomes to confirm that cells can absorb them, and then co-culture with HVSMCs for cell proliferation and migration detection. The target genes of miR-146a were screened and identified through bioinformatics and luciferase activity assay, and the expression of miR-146a and related proteins was detected through qRT-PCR and Western blot in HUVECs. An AS model in LDLR^-/- mice induced by a high-fat diet was developed to investigate the impact of exosome-miR-146a on AS.

Results: The results showed that experimental foam cells from AS showed higher expression of miR-146a. It was observed that NMMHC IIA and HSP70 interacted to regulate the release of exosomes. And HUVECs can absorb exosomes derived from macrophages. In addition, we also found that miR-146a directly targeted the SMAD4 gene to modulate the p38 MAPK signaling pathway, thereby mediating HUVECs damage. Furthermore, exosome-miR-146a induced abnormal proliferation and migration of HVSMCs. The expression of miR-146a was significantly reduced in miR-146a-mimics mice and increased in miR-146a inhibitor mice whereas the inhibition of miR-146a effectively reduced while increasing miR-146a worsened AS in mice.

Conclusion: Our findings expressed the potential of miR-146a as a favorable therapeutic target for AS, however, further exploration is suggestive for deep understanding of the mechanisms regulating exosome-miR-146a release in vivo and to develop effective therapeutic strategies involving miR-146a.

Introduction: Immune checkpoint inhibitors (ICIs) are potent and precise therapies for various cancer types, significantly improving survival rates in patients who respond positively to them. However, only a minority of patients benefit from ICI treatments.

Objectives: Identifying ICI responders before treatment could greatly conserve medical resources, minimize potential drug side effects, and expedite the search for alternative therapies. Our goal is to introduce a novel deep-learning method to predict ICI treatment responses in cancer patients.

Methods: The proposed deep-learning framework leverages graph neural network and biological pathway knowledge. We trained and tested our method using ICI-treated patients' data from several clinical trials covering melanoma, gastric cancer, and bladder cancer.

Results: Our results demonstrate that this predictive model outperforms current state-of-the-art methods and tumor microenvironment-based predictors. Additionally, the model quantifies the importance of pathways, pathway interactions, and genes in its predictions. A web server for IRnet has been developed and deployed, providing broad accessibility to users at https://irnet.missouri.edu.

Conclusion: IRnet is a competitive tool for predicting patient responses to immunotherapy, specifically ICIs. Its interpretability also offers valuable insights into the mechanisms underlying ICI treatments.

Introduction: P. pastoris is a common host for effective biosynthesis of heterologous proteins as well as small molecules. Accurate regulation of gene transcription and protein synthesis is necessary to coordinate synthetic gene circuits and optimize cellular energy distribution. Traditional methanol or other inducible promoters, natural or engineered, have defects in either fermentation safety or expression capacity. The utilization of chemical inducers typically adds complexity to the product purification process, but there is no other well-controlled protein synthesis system than promoters yet.

Objective: The study aimed to address the aforementioned challenges by constructing light-regulated gene transcription and protein translation systems with excellent expression capacity and light sensitivity.

Methods: Trans-acting factors were designed by linking the N. crassa blue-light sensor WC-1 with the activation domain of endogenous transcription factors. Light inducible or repressive promoters were then constructed through chimeric design of cis-elements (light-responsive elements, LREs) and endogenous promoters. Various configurations of trans-acting factor/LRE pairs, along with different LRE positions and copy numbers were tested for optimal promoter performance. In addition to transcription, a light-repressive translation system was constructed through the "rare codon brake" design. Rare codons were deliberately utilized to serve as brakes during protein synthesis, which were switched on and off through the light-regulated changes in the expression of the corresponding pLRE-tRNA.

Results: As demonstrated with GFP, the light-inducible promoter 4pLRE-cP_AOX1 was 70 % stronger than the constitutive promoter P_GAP, with L/D ratio = 77. The light-repressive promoter P_GAP-pLRE was strictly suppressed by light, with expression capacity comparable with P_GAP in darkness. As for the light-repressive translation system, the "triple brake" design successfully eliminated leakage and achieved light repression on protein synthesis without any impact on mRNA expression.

Conclusion: The newly designed light-regulated transcription and translation systems offer innovative tools that optimize the application of P. pastoris in biotechnology and synthetic biology.

Introduction: Heterosis has revolutionized crop breeding, enhancing global agricultural production. However, the mechanisms underlying heterosis remain obscure. Xiangzamian 2# (XZM2), a super hybrid upland cotton (Gossypium hirsutum L.) characterized by high-yield heterosis, has been developed and extensively planted in China.

Objectives: We conducted a systematic analysis of CRI12 and J8891, two parents of XZM2. We aimed to reveal the precise genetic information and the role of non-syntenic divergence in shaping heterosis, laying a foundation for advancing understanding of heterosis.

Methods: We de novo assembled high-quality genomes of CRI12 and J8891, and further uncovered abundant genetic variations and non-syntenic regions between the parents. Whole-genome comparison, association analysis, transcriptomic analysis and relative identity-by-descent (rIBD) estimation were conducted to identify structural variations (SVs) and introgressions within non-syntenic blocks and to analyze their impacts on promoting heterosis.

Results: Parental genetic divergence increased in non-syntenic regions. Furthermore, these regions, accounting for only 16.71% of the total genome, contained more loci with significantly higher heterotic effects, far exceeding the syntenic background. SVs covered 97.26% of non-syntenic sequences and caused widespread gene expression differences in these regions, driving dynamic complementation of gene expression in the hybrid. A set of SVs were responsible for trait improvement and had positive effects on heterosis, contributing larger heritability than short variations. We characterized numerous parental-specific introgressions from G. barbadense. Specifically, a functional introgression segment within non-syntenic blocks introduced an elite haplotype, which significantly increased lint yield and enhanced heterosis.

Conclusion: Our study clarified non-syntenic regions to harbor more loci with higher heterotic effects, revealed their importance in promoting heterosis and supported the crucial role of genetic complementation in heterosis. SVs and introgressions were identified as key factors responsible for non-syntenic divergence between the parents. They had important effects on gene expression and trait improvement, positively contributing to heterosis.

Introduction: The accumulation of microbiota-derived trimethylamine N-oxide (TMAO) in the atrium is linked to the development and progression of atrial arrhythmia. Butyrate, a major short-chain fatty acid, plays a crucial role in sustaining intestinal homeostasis and alleviating systemic inflammation, which may reduce atrial arrhythmogenesis.

Objectives: This study explored the roles of butyrate in regulating TMAO-mediated atrial remodeling and arrhythmia.

Methods: Whole-cell patch clamp experiments, Western blotting, and immunocytochemistry were used to analyze electrical activity and signaling, respectively, in TMAO-treated HL-1 atrial myocytes with or without sodium butyrate (SB) administration. Telemetry electrocardiographic recording and echocardiography and Masson's trichrome staining and immunohistochemistry were employed to examine atrial function and histopathology, respectively, in mice treated with TMAO with and without SB administration.

Results: Compared with control cells, TMAO-treated HL-1 myocytes exhibited reduced action potential duration (APD), elevated sarcoplasmic reticulum (SR) calcium content, larger L-type calcium current (I_Ca-L), increased Na⁺/Ca²⁺ exchanger (NCX) current, and increased potassium current. However, the combination of SB and TMAO resulted in similar APD, SR calcium content, I_Ca-L, transient outward potassium current (I_to), and ultrarapid delayed rectifier potassium current (I_Kur) compared with controls. Additionally, TMAO-treated HL-1 myocytes exhibited increased activation of endoplasmic reticulum (ER) stress signaling, along with increased PKR-like ER stress kinase (PERK)/IRE1α axis activation and expression of phospho-IP3R, NCX, and Kv1.5, compared with controls or HL-1 cells treated with the combination of TMAO and SB. TMAO-treated mice exhibited atrial ectopic beats, impaired atrial function, increased atrial fibrosis, and greater activation of ER stress signaling with PERK/IRE1α axis activation compared with controls and mice treated with TMAO combined with SB.

Conclusion: TMAO administration led to PERK/IRE1α axis activation, which may increase atrial remodeling and arrhythmogenesis. SB treatment mitigated TMAO-elicited ER stress. This finding suggests that SB administration is a valuable strategy for treating TMAO-induced atrial arrhythmia.