Bright and stable monomeric green fluorescent protein derived from StayGold.

The high brightness and photostability of the green fluorescent protein StayGold make it a particularly attractive probe for long-term live-cell imaging; however, its dimeric nature precludes its application as a fluorescent tag for some proteins. Here, we report the development and crystal structures of a monomeric variant of StayGold, named mBaoJin, which preserves the beneficial properties of its precursor, while serving as a tag for structural proteins and membranes. Systematic benchmarking of mBaoJin against popular green fluorescent proteins and other recently introduced monomeric and pseudomonomeric derivatives of StayGold established mBaoJin as a bright and photostable fluorescent protein, exhibiting rapid maturation and high pH/chemical stability. mBaoJin was also demonstrated for super-resolution, long-term live-cell imaging and expansion microscopy. We further showed the applicability of mBaoJin for neuronal labeling in model organisms, including Caenorhabditis elegans and mice.

Quantitative assessment of near-infrared fluorescent proteins.

Recent progress in fluorescent protein development has generated a large diversity of near-infrared fluorescent proteins (NIR FPs), which are rapidly becoming popular probes for a variety of imaging applications. However, the diversity of NIR FPs poses a challenge for end-users in choosing the optimal one for a given application. Here we conducted a systematic and quantitative assessment of intracellular brightness, photostability, oligomeric state, chemical stability and cytotoxicity of 22 NIR FPs in cultured mammalian cells and primary mouse neurons and identified a set of top-performing FPs including emiRFP670, miRFP680, miRFP713 and miRFP720, which can cover a majority of imaging applications. The top-performing proteins were further validated for in vivo imaging of neurons in Caenorhabditis elegans, zebrafish, and mice as well as in mice liver. We also assessed the applicability of the selected NIR FPs for multicolor imaging of fusions, expansion microscopy and two-photon imaging.

Cpne1 deficiency preserves sperm motility under Ca2+ channel blockade.

Calcium ions (Ca2+) play crucial roles in sperm motility and fertilization. The copine (CPNE) family comprises several Ca2+-dependent phospholipid-binding proteins. Of these, CPNE1 is extensively expressed in mammalian tissues; however, its precise role in testicular development and spermatogenesis is yet to be fully characterized. In this study, we used proteomics to analyze testicular biopsies and found that levels of CPNE1 were significantly reduced in patients with non-obstructive azoospermia (defective spermatogenesis) compared to those in patients with obstructive azoospermia (physiological spermatogenesis). In mice, CPNE1 is expressed at various stages of germ cell development and is associated with the Golgi apparatus. Ultimately, CPNE1 is expressed in the flagella of mature sperms. To further examine the role of CPNE1, we developed a Cpne1 knockout mouse model. Analysis showed that the loss of Cpne1 did not impair testicular development, spermatogenesis, or sperm morphology and motility in physiological conditions. When treated with gadolinium (III) chloride or 2-aminoethoxydiphenyl borate, known inhibitors of store-operated Ca2+ entry, Ca2+ signals and sperm motility were significantly compromised in wild-type mice; however, both mechanisms were conserved in KO mice. These results suggested that CPNE1 is dispensable for testicular development, spermatogenesis or sperm motility in physiological conditions. In addition, CPNE1 may represent a target of Ca2+ channel inhibitors and may therefore be implicated in the regulation of Ca2+ signaling and sperm motility.

Construction and application of nasopharyngeal carcinoma-specific big data platform based on electronic health records.

OBJECTIVE: To establish a nasopharyngeal carcinoma-specific big data platform based on electronic health records (EHRs) to provide data support for real-world study of nasopharyngeal carcinoma. METHODS: A multidisciplinary expert team was established for this project. Based on industry standards and practical feasibility, the team designed the nasopharyngeal carcinoma data element standards including 14 modules and 640 fields. Data from patients diagnosed with nasopharyngeal carcinoma who visited Southern Hospital after 1999 were extracted from 15 EHRs systems and were cleaned, structured, and standardized using information technologies such as machine learning and natural language processing. In addition, a series of measures such as quality control and data encryption were taken to ensure data quality and patient privacy. At the platform application level, 10 functional modules were designed according to the needs of nasopharyngeal carcinoma research. RESULTS: As of 1 October 2022, the Big Data platform has included 11,617patients, of whom 8228 (70.83 %) were male and 3389 (29.17 %) were female, with a median age of 48 years (interquartile range, 40 years). The data in the platform were validated to have a high level of completeness and accuracy, especially for key variables such as social demographics, laboratory tests and vital signs. Currently, six projects involving risk factors, early diagnosis, treatment efficacy and prevention of treatment-related toxic reactions have been conducted on the platform. CONCLUSIONS: We have established a high-quality NPC-specific big data platform by integrating heterogeneous data from multiple sources in the EHR. The platform provides an effective tool and strong data support for real-world studies of nasopharyngeal carcinoma, which helps to improve research efficiency, reduce costs, and improve the quality of research results. We expect to promote multicenter nasopharyngeal carcinoma data sharing in the future to facilitate the generation of high-quality real-world evidence in nasopharyngeal carcinoma. This article may provide some reference value for other comprehensive hospitals to establish a big data platform for nasopharyngeal carcinoma.

Iridium(III) complex induces apoptosis in HeLa cells by regulating mitochondrial and PI3K/AKT signaling pathways: In vitro and in vivo experiments.

Cyclometalated iridium complexes with mitochondrial targeting show great potential as substitutes for platinum-based complexes because of their strong anti-cancer properties. Three novel cyclometalated iridium(III) compounds were synthesized and evaluated in five different cell lines as part of the ongoing systematic investigations of these compounds. The complexes were prepared using 4,7-dichloro-1,10-phenanthroline ligands. The cytotoxicity of complexes Ir1-Ir3 towards HeLa cells was shown to be high, with IC50 values of 0.83±0.06, 4.73±0.11, and 4.95±0.62 µM, respectively. Complex Ir1 could be ingested by HeLa cells in 3 h and has shown high selectivity toward mitochondria. Subsequent investigations demonstrated that Ir1 triggered apoptosis in HeLa cells by augmenting the generation of reactive oxygen species (ROS), reducing the mitochondrial membrane potential, and depleting ATP levels. Furthermore, the movement of cells was significantly suppressed and the progression of the cell cycle was arrested in the G0/G1 phase following the administration of Ir1. The Western blot analysis demonstrated that the induction of apoptosis in HeLa cells by Ir1 involves the activation of the mitochondria-dependent channel and the PI3K/AKT signaling pathway. No significant cytotoxicity was observed in zebrafish embryos at concentrations less than or equal to 16 µM, e.g., survival rate and developmental abnormalities. In vivo, antitumor assay demonstrated that Ir1 suppressed tumor growth in mice. Therefore, our work shows that complex Ir1 could be a promising candidate for developing novel antitumor drugs.

Loss of Raptor induces Sertoli cells into an undifferentiated state in mice.

In mammals, testis development is triggered by the expression of the sex-determining Y-chromosome gene SRY to commit the Sertoli cell (SC) fate at gonadal sex determination in the fetus. Several genes have been identified to be required to promote the testis pathway following SRY activation (i.e., SRY box 9 (SOX9)) in an embryo; however, it largely remains unknown about the genes and the mechanisms involved in stabilizing the testis pathway after birth and throughout adulthood. Herein, we report postnatal males with SC-specific deletion of Raptor demonstrated the absence of SC unique identity and adversely acquired granulosa cell-like characteristics, along with loss of tubular architecture and scattered distribution of SCs and germ cells. Subsequent genome-wide analysis by RNA sequencing revealed a profound decrease in the transcripts of testis genes (i.e., Sox9, Sox8, and anti-Mullerian hormone (Amh)) and, conversely, an increase in ovary genes (i.e., LIM/Homeobox gene 9 (Lhx9), Forkhead box L2 (Foxl2) and Follistatin (Fst)); these changes were further confirmed by immunofluorescence and quantitative reverse-transcription polymerase chain reaction. Importantly, co-immunofluorescence demonstrated that Raptor deficiency induced SCs dedifferentiation into a progenitor state; the Raptor-mutant gonads showed some ovarian somatic cell features, accompanied by enhanced female steroidogenesis and elevated estrogen levels, yet the zona pellucida 3 (ZP3)-positive terminally feminized oocytes were not observed. In vitro experiments with primary SCs suggested that Raptor is likely involved in the fibroblast growth factor 9 (FGF9)-induced formation of cell junctions among SCs. Our results established that Raptor is required to maintain SC identity, stabilize the male pathway, and promote testis development.

53 years old is a reasonable cut-off value to define young and old patients in clear cell renal cell carcinoma: a study based on TCGA and SEER database.

BACKGROUND: The objectives of this study were to screen out cut-off age value and age-related differentially expressed genes (DEGs) in clear cell renal cell carcinoma (CCRCC) from Surveillance Epidemiology and End Results (SEER) database and The Cancer Genome Atlas (TCGA) database. METHODS: We selected 45,974 CCRCC patients from SEER and 530 RNA-seq data from TCGA database. The age cut-off value was defined using the X-tile program. Propensity score matching (PSM) was used to balance the differences between young and old groups. Hazard ratio (HR) was applied to evaluate prognostic risk of age in different subgroups. Age-related DEGs were identified via RNA-seq data. Survival analysis was used to assess the relationship between DEGs and prognosis. RESULTS: In this study, we divided the patients into young (n = 14,276) and old (n = 31,698) subgroups according to cut-off value (age = 53). Age > 53 years was indicated as independent risk factor for overall survival (OS) and cancer specific survival (CSS) of CCRCC before and after PSM. The prognosis of old group was worse than that in young group. Eleven gene were differential expression between the younger and older groups in CCRCC. The expression levels of PLA2G2A and SIX2 were related to prognosis of the elderly. CONCLUSION: Fifty-three years old was cut-off value in CCRCC. The prognosis of the elderly was worse than young people. It remind clinicians that more attention and better treatment should be given to CCRCC patients who are over 53 years old. PLA2G2A and SIX2 were age-related differential genes which might play an important role in the poor prognosis of elderly CCRCC patients.

Difference in ambient-personal exposure to PM2.5 and its inflammatory effect in local residents in urban and peri-urban Beijing, China: results of the AIRLESS project.

Measurement of ambient fine particulate matter (PM2.5) is often used as a proxy of personal exposure in epidemiological studies. However, the difference between personal and ambient exposure, and whether it biases the estimates of health effects remain unknown. Based on an epidemiological study (AIRLESS) and simultaneously launched intensive monitoring campaigns (APHH), we quantified and compared the personal and ambient exposure to PM2.5 and the related health impact among residents in Beijing, China. In total, 123 urban and 128 peri-urban non-smoking participants were recruited from two well-established cohorts in Beijing. During winter 2016 and summer 2017, each participant was instructed to carry a validated personal air monitor (PAM) to measure PM2.5 concentration at high spatiotemporal resolution for seven consecutive days in each season. Multiple inflammatory biomarkers were measured, including exhaled NO, blood monocytes counts and C-reactive protein. Linear mixed-effect models were used for the associations between exposure and health outcomes with adjustment for confounders. The average level of daily personal exposure to PM2.5 was consistently lower than using corresponding ambient concentration, and the difference is greater during the winter. The personal to ambient (P/A) ratio of exposure to PM2.5 exhibited an exponentially declining trend, and showed larger variations when ambient PM2.5 levels < 25 µg m-3. Personal exposure to PM2.5 was significantly associated with the increase in respiratory and systemic inflammatory biomarkers; however, the associations were weaker or became insignificant when ambient concentrations were used. Exposure to ambient PM2.5 might not be a good proxy to estimate the health effect of exposure to personal PM2.5.

The I510V mutation in KLHL10 in a patient with oligoasthenoteratozoospermia.

Oligoasthenoteratozoospermia is a human infertility syndrome caused by defects in spermatogenesis, spermiogenesis, and sperm maturation, and its etiology remains unclear. Kelch-like 10 (KLHL10) is a component of ubiquitin ligase E3 10 (KLHL10) and plays an important role in male fertility. Deletion or mutation of the Klhl10 gene in Drosophila or mice results in defects in spermatogenesis or sperm maturation. However, the molecular mechanisms by which KLHL10 functions remain elusive. In this study, we identified a missense mutation (c.1528A→G, p.I510V) in exon 5 of KLHL10, which is associated with oligoasthenoteratozoospermia in humans. To investigate the effects of this mutation on KLHL10 function and spermatogenesis and/or spermiogenesis, we generated mutant mice duplicating the amino acid conversion using the clustered regularly interspaced palindromic repeat/caspase 9 (CRISPR/Cas9) system and designated them Klhl10I510V mice. However, the Klhl10I510V mice did not exhibit any defects in testis development, spermatogenesis, or sperm motility at ten-weeks-of-age, suggesting that this mutation does not disrupt the KLHL10 function, and may not be the cause of male infertility in the affected individual with oligoasthenoteratozoospermia.

[Short-chain fatty acid butyrate acid attenuates atherosclerotic plaque formation in apolipoprotein E-knockout mice and the underlying mechanism].

This study was designed to evaluate the role of short-chain fatty acid butyrate acid on intestinal morphology and function, and atherosclerotic plaque formation in apolipoprotein E-knockout (ApoE-/-) mice. ApoE-/- mice on high-fat, high-cholesterol diet were treated with butyrate acid (200 mmol/L) or NaCl (control) in the drinking water for 12 weeks, followed by histological evaluations of atherosclerotic lesion in aorta. Real-time PCR analysis and ELISA were used to measure the expression levels of proinflammatory cytokines. Butyrate acid significantly attenuated high-fat, high-cholesterol diet-induced atherosclerotic plaque formation in ApoE-/- mice. Butyrate acid prevented high-fat, high-cholesterol diet-induced inflammation in both the aorta and the circulation, as evidenced by reduced expression of proinflammatory cytokines. These changes were accompanied by a marked attenuation in metabolic endotoxemia lipopolysaccharide (LPS). Butyrate acid induced intestinal expression of the tight junction proteins (Occludin and zona occuldens protein-1), thereby preventing the gut permeability. Butyrate acid dose-dependently upregulated the expression of the tight junction proteins in Caco-2 cells in GPR41-dependent manner. In conclusion, butyrate acid attenuates atherosclerotic lesions by ameliorating metabolic endotoxemia-induced inflammation through restoration of the gut barrier.

Loss of Fbxw7 in Sertoli cells impairs testis development and causes infertility in mice†.

F-box and WD-40 domain protein 7 (Fbxw7) is a component of the Skp1-Cdc53/Cullin-F-box-protein complex (SCF/ß-TrCP), which is an E3 ubiquitin ligase that mediates protein degradation. This complex has recently been shown to negatively regulate spermatogonial stem cell self-renewal; however, its roles in Sertoli cell (SC) proliferation, differentiation, and function remain to be established. In this study, we generated conditional mutant mice with SC-specific deletion of Fbxw7 via the Cre-loxP system. Fbxw7 deficiency in SCs impaired testis development, which is characterized by age-dependent tubular atrophy, excessive germ cell loss, and spermatogenic arrest, and the mutant males were infertile at 7 months old. Fbxw7 ablation also compromised cytoskeletal organization and cell polarity of SCs, as well as integrity of the blood-testis barrier. In addition, the transcript levels of cell markers for germ cells, Leydig cells, and SCs were significantly decreased in Fbxw7 mutant mice. Importantly, protein levels of GATA-4, a transcription factor that plays a crucial role in SC maturation and testis development, were progressively decreased in control SCs after postnatal day 14, whereas levels were aberrantly elevated in Fbxw7-deleted SCs. Interestingly, the Gata-4 messenger RNA levels remained stable following Fbxw7 deletion. Fbxw7 silencing in SCs also induced progressive Leydig cell inefficiency and testosterone insufficiency. Collectively, these results demonstrate that Fbxw7 expression is required for SC maturation and function, potentially through degradation of GATA-4, to support pubertal testis development and spermatogenesis.

Adenylate kinase 1 deficiency disrupts mouse sperm motility under conditions of energy stress†.

Mammalian spermatozoa are highly polarized cells characterized by compartmentalized cellular structures and energy metabolism. Adenylate kinase (AK), which interconverts two ADP molecules into stoichiometric amounts of ATP and AMP, plays a critical role in buffering adenine nucleotides throughout the tail to support flagellar motility. Yet the role of the major AK isoform, AK1, is still not well characterized. Here, by using a proteomic analysis of testis biopsy samples, we found that AK1 levels were significantly decreased in nonobstructive azoospermia patients. This result was further verified by immunohistochemical staining of AK1 on a tissue microarray. AK1 was found to be expressed in post-meiotic round and elongated spermatids in mouse testis and subsequent mature sperm in the epididymis. We then generated Ak1 knockout mice, which showed that AK1 deficiency did not induce any defects in testis development, spermatogenesis, or sperm morphology and motility under physiological conditions. We further investigated detergent-modeled epididymal sperm and included individual or mixed adenine nucleotides to mimic energy stress. When only ADP was available, Ak1 disruption largely compromised sperm motility, manifested as a smaller beating amplitude and higher beating frequency, which resulted in less effective forward swimming. The energy restriction/recover experiments with intact sperm further addressed this finding. Besides, decreased AK activity was observed in sperm of a male fertility disorder mouse model induced by cadmium chloride. These results cumulatively demonstrate that AK1 was dispensable for testis development, spermatogenesis, or sperm motility under physiological conditions, but was required for sperm to maintain a constant adenylate energy charge to support sperm motility under conditions of energy stress.

Activation of mTORC1 in fibroblasts accelerates wound healing and induces fibrosis in mice.

Wound healing is a multicellular process that involves the coordinated efforts of several cell types, including keratinocytes, fibroblasts, and endothelial cells. This process is also regulated by an equally complex signaling network involving numerous growth factors, cytokines, and chemokines. The mechanistic target of rapamycin complex 1 (mTORC1) is a central regulator of cell growth, proliferation, and differentiation. A recent study showed that mTORC1 activation in epithelial cells dramatically enhanced epithelial cell proliferation, migration, and cutaneous wound healing; however, the roles of mTORC1 in fibroblasts during wound healing remain unknown. Here, we generated genetically mutated mice with activated mTORC1 in fibroblasts by conditionally deleting the mTORC1 inhibitor, TSC1. Activation of mTORC1 in fibroblasts significantly increased fibroblastic cell proliferation and contractile α-smooth muscle actin expression, thus promoting wound closure. Elevated mTORC1 activity also adversely induced excessive collagen production, leading to excessive scaring and fibrosis. Importantly, both accelerated wound healing and fibrotic phenotypes were largely reversed by the mTORC1 inhibitor, rapamycin. These observations were also replicated in primary human dermal fibroblasts. These results collectively demonstrated that mTORC1 activity in skin fibroblasts was a critical orchestrator in cutaneous wound healing and scarring.

TSC1 deletion in fibroblasts alleviates lipopolysaccharide-induced acute kidney injury.

Mechanistic target of rapamycin complex 1 (mTORC1) signaling is active in inflammation, but its involvement in septic acute kidney injury (AKI) has not been shown. mTORC1 activation (p-S6) in renal fibroblasts was increased in a mouse AKI model induced by 1.5 mg/kg lipopolysaccharide (LPS). Deletion of tuberous sclerosis complex 1 (TSC1), an mTORC1 negative regulator, in fibroblasts (Fibro-TSC1-/-) inhibited the elevation of serum creatinine and blood urea nitrogen in AKI compared with that in TSC1fl/fl control mice. Endothelin-1 (EDN1) and phospho-Jun-amino-terminal kinase (p-JNK) were up-regulated in Fibro-TSC1-/- renal fibroblasts after LPS challenge. Rapamycin, an mTORC1 inhibitor, and bosentan, an EDN1 antagonist, eliminated the difference in renal function between TSC1fl/fl and Fibro-TSC1-/- mice after LPS injection. Rapamycin restored LPS-induced up-regulation of EDN1, endothelin converting enzyme-1 (ECE1), and p-JNK in TSC1-knockdown mouse embryonic fibroblasts (MEFs). SP600125, a Jun-amino-terminal kinase (JNK) inhibitor, attenuated LPS-induced enhancement of EDN1 and ECE1 in TSC1-knockdown MEFs without a change in phospho-S6 ribosomal protein (p-S6) level. The results indicate that mTORC1-JNK-dependent up-regulation of ECE1 elevated EDN1 in TSC1-knockout renal fibroblasts and contributed to improvement of renal function in Fibro-TSC1-/- mice with LPS-induced AKI. Renal fibroblast mTORC1 plays an important role in septic AKI.

Cyclin D1 (CCND1) G870A polymorphisms and cervical cancer susceptibility: a meta-analysis based on ten case-control studies.

Many studies have evaluated the association between cyclin D1 (CCND1) G870A polymorphism and cervical cancer susceptibility. However, these studies showed inconsistent results. The aim of this study was to derive a more precise estimation of this association. We searched PubMed and Embase for related studies that had been published in English, and ten case-control studies with a total of 2,864 cases and 3,898 controls were finally identified to be eligible studies in the meta-analysis. The association was assessed by summarizing the odds ratios (ORs) with the corresponding 95 % confidence intervals (CIs). Overall, there was no significant association between cyclin D1 (CCND1) G870A polymorphism and cervical cancer risk (for the allele model A vs. G: OR = 1.02, 95 % CI 0.88-1.19, p = 0.76; for the co-dominant model AA vs. GG: OR = 1.03, 95 % CI 0.75-1.41, p = 0.85; for the dominant model AA + GA vs. GG: OR = 1.00, 95 % CI 0.78-1.28, p = 0.99; for the recessive comparison AA vs. GA + GG: OR = 1.06, 95 % CI 0.85-1.32, p = 0.62). In subgroup analysis by ethnicity, no significant difference was found in both Asians and Caucasians. In summary, the present meta-analysis provides evidence that genotypes for the cyclin D1 (CCND1) G870A polymorphism may be not associated with genetic susceptibility of cervical cancer.

Smooth muscle progenitor cells involved in the development of airway remodeling in a murine model of asthma.

BACKGROUND: The mechanisms regulating airway remodeling changes remain poorly understood. Recently, a smooth muscle progenitor cell was identified in the peripheral circulatory system that plays an important role in the reconstruction of injured blood vessels. However, to the best of our knowledge, there is no report in the medical literature regarding the role of smooth muscle progenitor cells (SPCs) in asthma. OBJECTIVE: The aim of this study was to investigate the relationship between SPCs and the development of airway remodelling in a murine model of asthma. METHODS: Chronic asthma with airway remodeling was generated by sensitizing and stimulating BALB/c mice with atomized ovalbumin (OVA). Bronchoalveolar lavage fluid (BALF) was collected for eosinophils (EOS) counting and histological analysis. The Ficoll method was used to isolate mononuclear cells from peripheral blood. Smooth muscle myosin heavy chain (SM-MHC) and highly glycosylated type I transmembrane protein (CD34⁺) were selected as two markers to detect the expression of SPCs by Flow Cytometry. RESULTS: Long-term inhalation of OVA produced thickening of the epithelial and smooth muscle layer, goblet cell hyperplasia, collagen deposition around smooth muscle, luminal exudates and inflammatory cell infiltration. The number of SPCs in the asthma group was significantly higher than in the control group. CONCLUSION: Long-term inhalation of OVA results in airway remodeling and the smooth progenitor muscle cell are involved in the development of airway remodeling.

Effects of four polyphenols on mouse wound healing and the gene expression profile of resveratrol action.

Studies have demonstrated the potent effects of polyphenols on cutaneous wound healing. However, the molecular mechanisms underlying polyphenol activity are incompletely understood. Herein, mice were experimentally wounded, intragastrically treated with four polyphenols, resveratrol, tea polyphenols, genistein, and quercetin; and monitored for 14 days. Resveratrol was the most effective compound, promoting wound healing starting at day 7 after wounding, by enhancing cell proliferation and reducing apoptosis and subsequently promoting epidermal and dermal repair, collagen synthesis and scar maturation. RNA sequencing was performed in control and resveratrol-treated tissues on day 7 after wounding. Resveratrol treatment upregulated 362 genes and downregulated 334 genes. Gene Ontology enrichment analysis showed that differentially expressed genes (DEGs) were associated with different biological processes (keratinization, immunity, and inflammation), molecular functions (cytokine and chemokine activities), and cellular components (extracellular region and matrix). Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that DEGs were predominantly enriched in inflammatory and immunological pathways, including cytokine-cytokine receptor interaction, chemokine signaling, and tumor necrosis factor (TNF) signaling. These results show that resveratrol accelerates wound healing by promoting keratinization and dermal repair and attenuating immune and inflammatory responses.

Plasma Metabolic Profiles-Based Prediction of Induction Chemotherapy Efficacy in Nasopharyngeal Carcinoma: Results of a Bidirectional Clinical Trial.

PURPOSE: The efficacy of induction chemotherapy (IC) as a primary treatment for advanced nasopharyngeal carcinoma (NPC) remains a topic of debate, with a lack of dependable biomarkers for predicting its efficacy. This study seeks to establish a predictive classifier using plasma metabolomics profiles. PATIENTS AND METHODS: A total of 166 NPC patients enrolled in the clinical trial NCT05682703 who were undergoing IC were included in the study. Plasma lipoprotein profiles were obtained using 1H-nuclear magnetic resonance before and after IC treatment. An artificial intelligence-assisted radiomics method was developed to effectively evaluate its efficacy. Metabolic biomarkers were identified through a machine learning approach based on a discovery cohort and subsequently validated in a validation cohort that mimicked the most unfavorable real-world scenario. RESULTS: Our research findings indicate that the effectiveness of IC varies among individual patients, with a correlation observed between efficacy and changes in metabolite profiles. Using machine learning techniques, it was determined that the extreme gradient boosting model exhibited notable efficacy, attaining an area under the curve (AUC) value of 0.792 (95% CI, 0.668-0.913). In the validation cohort, the model exhibited strong stability and generalizability, with an AUC of 0.786 (95% CI, 0.533-0.922). CONCLUSIONS: In this study, we found that dysregulation of plasma lipoprotein may result in resistance to IC in NPC patients. The prediction model constructed based on the plasma metabolites' profile has good predictive capabilities and potential for real-world generalization. This discovery has implications for the development of treatment strategies and may offer insight into potential targets for enhancing the effectiveness of IC.

VLDL and LDL Subfractions Enhance the Risk Stratification of Individuals Who Underwent Epstein-Barr Virus-Based Screening for Nasopharyngeal Carcinoma: A Multicenter Cohort Study.

Serological tests for Epstein-Barr virus (EBV) antibodies have been widely conducted for the screening of nasopharyngeal carcinoma (NPC) in endemic areas. Further risk stratification of NPC can be achieved through plasma lipoprotein and metabolic profiles. A total of 297 NPC patients and 149 EBV-positive participants are enrolled from the NCT03919552 and NCT05682703 cohorts for plasma nuclear magnetic resonance (NMR) metabolomic analysis. Small, dense very low density lipoprotein particles (VLDL-5) and large, buoyant low density lipoprotein particles (LDL-1) are found to be closely associated with nasopharyngeal carcinogenesis. Herein, an NMR-based risk score (NRS), which combines lipoprotein subfractions and metabolic biomarkers relevant to NPC, is developed and well validated within a multicenter cohort. Combining the median cutoff value of the NRS (N50) with that of the serological test for EBV antibodies, the risk stratification model achieves a satisfactory performance in which the area under the curve (AUC) is 0.841 (95% confidence interval: 0.811-0.871), and the positive predictive value (PPV) reaches 70.08% in the combined cohort. These findings not only suggest that VLDL-5 and LDL-1 particles can serve as novel risk factors for NPC but also indicate that the NRS has significant potential in personalized risk prediction for NPC.

Nitric oxide-induced lipophagic defects contribute to testosterone deficiency in rats with spinal cord injury.

Introduction: Males with acute spinal cord injury (SCI) frequently exhibit testosterone deficiency and reproductive dysfunction. While such incidence rates are high in chronic patients, the underlying mechanisms remain elusive. Methods and results: Herein, we generated a rat SCI model, which recapitulated complications in human males, including low testosterone levels and spermatogenic disorders. Proteomics analyses showed that the differentially expressed proteins were mostly enriched in lipid metabolism and steroid metabolism and biosynthesis. In SCI rats, we observed that testicular nitric oxide (NO) levels were elevated and lipid droplet-autophagosome co-localization in testicular interstitial cells was decreased. We hypothesized that NO impaired lipophagy in Leydig cells (LCs) to disrupt testosterone biosynthesis and spermatogenesis. As postulated, exogenous NO donor (S-nitroso-N-acetylpenicillamine (SNAP)) treatment markedly raised NO levels and disturbed lipophagy via the AMPK/mTOR/ULK1 pathway, and ultimately impaired testosterone production in mouse LCs. However, such alterations were not fully observed when cells were treated with an endogenous NO donor (L-arginine), suggesting that mouse LCs were devoid of an endogenous NO-production system. Alternatively, activated (M1) macrophages were predominant NO sources, as inducible NO synthase inhibition attenuated lipophagic defects and testosterone insufficiency in LCs in a macrophage-LC co-culture system. In scavenging NO (2-4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (CPTIO)) we effectively restored lipophagy and testosterone levels both in vitro and in vivo, and importantly, spermatogenesis in vivo. Autophagy activation by LYN-1604 also promoted lipid degradation and testosterone synthesis. Discussion: In summary, we showed that NO-disrupted-lipophagy caused testosterone deficiency following SCI, and NO clearance or autophagy activation could be effective in preventing reproductive dysfunction in males with SCI.