Quantification of Total Ketone Bodies in Biological Matrices Using Stable Isotope-Labeled Internal Standards and RP-UHPLC-MS/MS.

Acetoacetate (AcAc) and D-beta-hydroxybutyrate (D-ßOHB), the two major ketone bodies found in circulation, are linked to multiple physiological and pathophysiological states. Therefore, analytical methodologies surrounding the quantification of total ketone body (TKB) concentrations in biological matrices are paramount. Traditional methods to quantify TKBs relied on indirect spectrophotometric assays with narrow dynamic ranges, which have been significantly improved upon by modern mass spectrometry (MS)-based approaches. However, the lack of stable isotope-labeled internal standards (ISs) for AcAc and the need to distinguish D-ßOHB from its closely related structural and enantiomeric isomers pose significant obstacles. Here, we provide a protocol to synthesize and quantify a [13C] stable isotope-labeled IS for AcAc, which, in conjunction with a commercially available [2H] stable isotope-labeled IS for ßOHB, allows TKBs to be measured across multiple biological matrices. This rapid (7 min) analysis employs reverse phase ultra-high performance liquid chromatography (RP-UHPLC) coupled to tandem MS (MS/MS) to distinguish ßOHB from three structural isomers using parallel reaction monitoring (PRM), providing excellent specificity and selectivity. Finally, a method is provided that distinguishes D-ßOHB from L-ßOHB using a simple one-step derivatization to produce the corresponding diastereomers, which can be chromatographically resolved using the same rapid RP-UHPLC separation with new PRM transitions. In summary, this method provides a rigorous analytical pipeline for the analysis of TKBs in biological matrices via leveraging two authentic stable isotope-labeled ISs and RP-UHPLC-MS/MS.

Liposomes-enabled cancer chemoimmunotherapy.

Chemoimmunotherapy is an emerging paradigm in the clinic for treating several malignant diseases, such as non-small cell lung cancer, breast cancer, and large B-cell lymphoma. However, the efficacy of this strategy is still restricted by serious adverse events and a high therapeutic termination rate, presumably due to the lack of tumor-targeted distribution of both chemotherapeutic and immunotherapeutic agents. Targeted drug delivery has the potential to address this issue. Among the most promising nanocarriers in clinical translation, liposomes have drawn great attention in cancer chemoimmunotherapy in recent years. Liposomes-enabled cancer chemoimmunotherapy has made significant progress in clinics, with impressive therapeutic outcomes. This review summarizes the latest preclinical and clinical progress in liposome-enabled cancer chemoimmunotherapy and discusses the challenges and future directions of this field.

Bioactive nanocomposite hydrogel enhances postoperative immunotherapy and bone reconstruction for osteosarcoma treatment.

Osteosarcoma, a malignant bone tumor often characterized by high hedgehog signaling activity, residual tumor cells, and substantial bone defects, poses significant challenges to both treatment response and postsurgical recovery. Here, we developed a nanocomposite hydrogel for the sustained co-delivery of bioactive magnesium ions, anti-PD-L1 antibody (αPD-L1), and hedgehog pathway antagonist vismodegib, to eradicate residual tumor cells while promoting bone regeneration post-surgery. In a mouse model of tibia osteosarcoma, this hydrogel-mediated combination therapy led to remarkable tumor growth inhibition and hence increased animal survival by enhancing the activity of tumor-suppressed CD8+ T cells. Meanwhile, the implanted hydrogel improved the microenvironment of osteogenesis through long-term sustained release of Mg2+, facilitating bone defect repair by upregulating the expression of osteogenic genes. After 21 days, the expression levels of ALP, COL1, RUNX2, and BGLAP in the Vis-αPD-L1-Gel group were approximately 4.1, 5.1, 5.5, and 3.4 times higher than those of the control, respectively. We believe that this hydrogel-based combination therapy offers a potentially valuable strategy for treating osteosarcoma and addressing the tumor-related complex bone diseases.

An ultra-long-acting L-asparaginase synergizes with an immune checkpoint inhibitor in starvation-immunotherapy of metastatic solid tumors.

Metastasis stands as the primary contributor to mortality associated with tumors. Chemotherapy and immunotherapy are frequently utilized in the management of metastatic solid tumors. Nevertheless, these therapeutic modalities are linked to serious adverse effects and limited effectiveness in preventing metastasis. Here, we report a novel therapeutic strategy named starvation-immunotherapy, wherein an immune checkpoint inhibitor is combined with an ultra-long-acting L-asparaginase that is a fusion protein comprising L-asparaginase (ASNase) and an elastin-like polypeptide (ELP), termed ASNase-ELP. ASNase-ELP's thermosensitivity enables it to generate an in-situ depot following an intratumoral injection, yielding increased dose tolerance, improved pharmacokinetics, sustained release, optimized biodistribution, and augmented tumor retention compared to free ASNase. As a result, in murine models of oral cancer, melanoma, and cervical cancer, the antitumor efficacy of ASNase-ELP by selectively and sustainably depleting L-asparagine essential for tumor cell survival was substantially superior to that of ASNase or Cisplatin, a first-line anti-solid tumor medicine, without any observable adverse effects. Furthermore, the combination of ASNase-ELP and an immune checkpoint inhibitor was more effective than either therapy alone in impeding melanoma metastasis. Overall, the synergistic strategy of starvation-immunotherapy holds excellent promise in reshaping the therapeutic landscape of refractory metastatic tumors and offering a new alternative for next-generation oncology treatments.

Protective effects and molecular mechanisms of Litopenaeus vannamei treated with l-arginine/l-lysine against myofibrillar proteins oxidation and quality degradation during freeze-thaw cycles.

The storage and processing of Litopenaeus vannamei are often challenged by the freeze-thaw (F-T) cycle phenomenon. This study delved into the influence of pretreatment with l-arginine (Arg) and l-lysine (Lys) on the myofibrillar proteins oxidation and quality of shrimp subjected to F-T cycles. Arg and Lys pretreatment notably improved water-holding capacity (WHC), textural integrity as well as the myofibrillar structure of the shrimps. A lesser reduction in the amounts of immobile and bound water was found in the amino acid-treated groups, and the oxidation of lipids and proteins were both decelerated. Molecular simulation results indicated that Arg and Lys could form hydrogen and salt-bridge bonds with myosin, enhancing the stability of Litopenaeus vannamei. The study concludes that Arg and Lys are effective in alleviating the adverse effects of F-T cycles on the quality of Litopenaeus vannamei, and provides a new solution for the quality maintenance during storage and processing.

The Use of Multiplex Real-Time PCR for the Simultaneous Detection of Foodborne Bacterial Pathogens.

Foodborne pathogens continue to be a major health concern worldwide. Culture-dependent methodologies are still considered the gold standard to perform pathogen detection and quantification. These methods present several drawbacks, such as being time-consuming and labor intensive. The implementation of real-time PCR has allowed to overcome these limitations, and even reduce the cost associated with the analyses, due to the possibility of simultaneously and accurately detecting several pathogens in one single assay, with results comparable to those obtained by classical approaches. In this chapter, a protocol for the simultaneous detection of two of the most important foodborne pathogens, Salmonella spp. and Listeria monocytogenes, is described.

Application of MinION Long-Read Sequencer for Semi-targeted Detection of Foodborne Pathogens.

Foodborne pathogens remain a serious health issue in developed and developing countries. Safeness of food products has been assured for years with culture-based microbiological methods; however, these present several limitations such as turnaround time and extensive hands-on work, which have been typically address taking advantage of DNA-based methods such as real-time PCR (qPCR). These, and other similar techniques, are targeted assays, meaning that they are directed for the specific detection of one specific microbe. Even though reliable, this approach suffers from an important limitation that unless specific assays are design for every single pathogen potentially present, foods may be considered erroneously safe. To address this problem, next-generation sequencing (NGS) can be used as this is a nontargeted method; thus it has the capacity to detect every potential threat present. In this chapter, a protocol for the simultaneous detection and preliminary serotyping of Salmonella enterica serovar Enteritidis, Salmonella enterica serovar Typhimurium, Listeria monocytogenes, and Escherichia coli O157:H7 is described.

LC-MS coupled with diagnostic ion strategy facilitated the discovery of 5-methylcoumarin meroterpenoids from Gerbera piloselloides.

Plant-derived natural products remain crucial in drug development. However, the identification of undescribed natural products is becoming increasingly challenging. A comprehensive strategy combining LC-MS with diagnostic ions was proposed for the discovery of undescribed 5-methylcoumarin meroterpenoids. Thirteen undescribed 5-methylcoumarin meroterpenoids, including five pairs of enantiomers (1a/1b and 5a/5b-8a/8b), were isolated from the whole plant of Gerbera piloselloides. Their structures and absolute configurations were unambiguously determined based on their spectroscopic data, calculated and experimental ECD data and X-ray diffraction analysis. Bioassays conducted on scopolamine-induced injury PC12 cells revealed that compounds 5a/5b, 7a/7b and 8a/8b possessed mild protective effects. Additionally, compounds 2 and 8 showed notable IL-6 inhibition in lipopolysaccharide-induced BEAS-2B cells.

Effect of sodium L-lactate on bioactive properties of chitosan-hydroxyapatite/polycaprolactone conduits for peripheral nerve tissue engineering.

Biomaterials and synthetic polymers have been widely used to replicate the regenerative microenvironment of the peripheral nervous system. Chitosan-based conduits have shown promise in the regeneration of nerve injuries. However, to mimic the regenerative microenvironment, the scaffold structure should possess bioactive properties. This can be achieved by the incorporation of biomolecules (e.g., proteins, peptides) or trophic factors that should preferably be aligned and/or released with controlled kinetics to activate the process of positive axon chemotaxis. In this study, sodium L-lactate has been used to enhance the bioactive properties of chitosan-hydroxyapatite/polycaprolactone electrodeposits. Next, two methods have been developed to incorporate NGF-loaded microspheres - Method 1 involves entrapment and co-deposition of NGF-loaded microspheres, while Method 2 is based on absorption of NGF-loaded microspheres. The study shows that modification of chitosan-hydroxyapatite/polycaprolactone conduits by sodium L-lactate significantly improves their bioactive, biological, and physicochemical properties. The obtained implants are cytocompatible, enhancing the neurite regeneration process by stimulating its elongation. The absorption of NGF-loaded microspheres into the conduit structure may be considered more favorable for the stimulation of axonal elongation compared to entrapment, as it allows for trophic factor dose-dependent controlled release. The developed conduits possess properties essential for the successful treatment of peripheral nerve discontinuities.

Bioactivity-based Analysis and Chemical Characterization of Hypoglycemic Components from Helicteres angustifolia L.

Helicteres angustifolia L. (H. angustifolia), a well-known traditional Chinese medicine, has been demonstrated to have hypoglycemic activity. We found that the EtOAc extract of H. angustifolia (HAEF) showed stronger α-glucosidase inhibitory activity than that of positive control. Furthermore, the hypoglycemic activity of HAEF was evaluated in streptozotocin (STZ)-induced type 2 diabetes mellitus (T2DM) rats. The results demonstrated that HAEF reduced the drinking quantity, feeding quantity, and controlled weight loss in diabetic rats. Besides, the fasting blood glucose (FBG), viscera index, and the area under time-blood glucose curve (AUC) were significantly decreased, and the oral glucose tolerance was also improved after 5 weeks. Then, the high-performance liquid chromatography with quadrupole time of flight tandem mass spectrometry (HPLC-Q-TOF-MS/MS) method was performed for qualitative analysis of the chemical constituents in HAEF. Twenty-one compounds were identified from in HAEF. Four compounds were further isolated from HAEF and subjected to α-glucosidase inhibition experiments. At the end, molecular docking was empolyed simulate the interaction of three compounds with α-glucosidase. This is the first report on major hypoglycaemic components has been identified in the roots of H. angustifolia. These findings provide a material basis for the use of H. angustifolia in the treatment of diabetes.

Anti-inflammatory effects of proprotein convertase subtilisin/kexin 9 inhibitor therapy in the early phase of acute myocardial infarction.

This study examined the anti-inflammatory and endothelial function-enhancing effects of proprotein convertase subtilisin/kexin 9 (PCSK9) inhibitor therapy in the early phase after acute myocardial infarction (AMI) by assessing changes in tumor necrosis factor-α (TNF-α) levels and the L-arginine/asymmetric-dimethylarginine (ADMA) ratio. This retrospective, single-center cohort study included patients who underwent successful timely primary percutaneous coronary intervention (PCI) for first-onset AMI between September 2017 and March 2018. The PCSK9 inhibitor group comprised patients who received 75 mg alirocumab up to 7 days after AMI, while the standard therapy group comprised patients who did not. We evaluated the change in TNF-α levels and the L-arginine/ADMA ratio at the time of hospital admission and prior to discharge. PCSK9 inhibitor therapy in the early phase after AMI suppressed TNF-α levels (standard therapy group, 1.64 ± 2.14 pg/mL vs. PCSK9 inhibitor group, 0.26 ± 0.33 pg/mL; p = 0.033) and increased the L-arginine/ADMA ratio (standard therapy group, - 13.0 ± 39.7 vs. PCSK9 inhibitor group, 23.2 ± 39.7; p = 0.042). Upon multiple regression analysis adjusted for sex, age, and peak creatine kinase levels, PCSK9 inhibitor therapy was associated with TNF-α suppression (p = 0.025; ß = - 0.235, 95% confidence interval [CI], - 0.436 to - 0.033). The L-arginine/ADMA ratio was also analyzed using multiple regression, adjusted for sex, age, peak creatine kinase levels, and smoking, showing a significant improvement in the ratio (p = 0.018; ß = 41.913, 95% CI, 10.337-73.491). Moreover, a weak negative correlation was suggested between the change in TNF-α levels and the change in L-arginine/ADMA ratio (r = - 0.393, p = 0.058). PCSK9 inhibitor therapy in the early phase after AMI suppresses TNF-α levels and improves the L-arginine/ADMA ratio, potentially indicating anti-inflammatory and endothelial function-enhancing effects.

Pathogens transmitted by Ixodes ricinus.

Ixodes ricinus is the most important tick vector in central and western Europe and one of the most researched parasites. However, in the published literature on the tick and the pathogens it transmits, conjecture about specific transmission cycles and the clinical significance of certain microbes is not always clearly separated from confirmed facts. This article aims to present up-to-date, evidence-based information about the well-researched human pathogens tick-borne encephalitis virus, louping-ill virus, Anaplasma phagocytophilum, Borrelia burgdorferi sensu lato and several Babesia species, with a focus on their development in the tick, transmission dynamics and the competent reservoir hosts that support their circulation in the environment. Borrelia miyamotoi, Neoehrlichia mikurensis, Rickettsia helvetica and Rickettsia monacensis, which are much less common causes of disease but may affect immunocompromised patients, are also briefly discussed. Finally, the possible role of I. ricinus in the transmission of Coxiella burnetii, Francisella tularensis, Bartonella spp. and Spiroplasma ixodetis is reviewed.

Is YouTube a reliable source of education for the intravitreal injection procedure?

PURPOSE: This study aims to evaluate the quality of videos on YouTube that demonstrate the intravitreal injection (IVI) procedure as an educational tool. MATERIAL AND METHODS: A search on YouTube using the keywords "intravitreal injection", "intravitreal injection procedure", "eye injection", "eye injection procedure", "dexamethasone intraocular injection", and "anti-VEGF injection" was performed on January 10, 2023. Of the first 300 videos obtained, 70 met the inclusion criteria. The videos were evaluated for content and quality using the IVI procedure checklist score, DISCERN, modified Global Quality Score (GQS), Health on the Net Foundation (HON) code, and the Journal of American Medical Association (JAMA) scores. The quality of the videos was also compared with regard to the uploading source, such as a university or training hospital, educational channels, and individual medical doctors or healthcare professionals. RESULTS: The mean IVI procedure checklist score was 4.84±1.58, and 29 videos fulfilled more than 80% of the checklist items, indicating that only 41.4% of the videos conformed to the IVI procedure recommendations. The mean DISCERN, modified GQS, and JAMA benchmark scores were 34.75±10.46, 2.90±1.09, and 2.09±0.72, respectively, indicating poor overall video quality. The mean HON code score was 4.68±1.39, indicating moderate overall video quality. Videos uploaded by educational channels seem to be of better quality than those uploaded by others. CONCLUSIONS: The majority of evaluated YouTube videos on the IVI procedure appear to be of low quality as an educational tool. Videos uploaded by educational channels would be preferred to gain quality information about the IVI procedure.

Overexpression of SEZ6L2 and Immune Infiltration in Cancer Based on Gene Image Diagnosis.

BACKGROUND: With the rapid advancement of optical image diagnostic technology, researchers are delving into the potential applications in the field of cancer diagnosis and treatment. The exact link between the SEZ6L2 gene and cancer immune infiltration remains elusive. MATERIALS AND METHODS: This study aims to investigate the relationship between SEZ6L2 gene overexpression and cancer immune infiltration using optical image diagnostic technology, thereby presenting novel insights for enhancing cancer diagnosis and treatment strategies. Tissue samples obtained from cancer patients were meticulously analyzed to quantitatively assess the expression of the SEZ6L2 gene through light image diagnostic technology. Additionally, immunohistochemical techniques were employed to assess the nature and quantity of immune infiltrating cells within the cancerous tissues. RESULTS: The enrichment pathways were found to include complement activation, circulating immunoglobulin mediated humoral immune response, protein activation cascade, immunoglobulin complex, and immunoglobulin. In addition, the expression of SEZ6L2 is closely related to the infiltration level of tumor infiltrating immune cells (TIICs), and there is a potential relationship between the expression of SEZ6L2 and different marker genes of TIIC. CONCLUSION: Increased SEZ6L2 mRNA expression in breast invasive carcinoma was significantly associated with negative prognosis and immune invasion. SEZ6L2 may be a novel prognostic biomarker and a potential immunotherapeutic target in BRCA.

Mathematics and society reunited: The social aspects of Brouwer's intuitionism.

Brouwer's philosophy of mathematics is usually regarded as an intra-subjective, even solipsistic approach, an approach that also underlies his mathematical intuitionism, as he strived to create a mathematics that develops out of something inner and a-linguistic. Thus, points of connection between Brouwer's mathematical views and his views about and the social world seem improbable and are rarely mentioned in the literature. The current paper aims to challenge and change that. The paper employs a socially oriented prism to examine Brouwer's views on the construction, use, and practice of mathematics. It focuses on Brouwer's views on language, his social interactions, and the importance of group context as they appear in the significs dialogues. It does so by exploring the establishment and dissolution of the significs movement, focusing on Gerrit Mannoury's influence and relationship with Brouwer and analyzing several fragments from the significs dialogues while emphasizing the role Brouwer ascribed to groups in forming and sharing new ideas. The paper concludes by raising two questions that challenge common historical and philosophical readings of intuitionism.

Novel insights into the role of quercetin and kaempferol from Carthamus tinctorius L. in the management of nonalcoholic fatty liver disease via NR1H4-mediated pathways.

This study investigates the novel therapeutic potential of quercetin and kaempferol, two bioactive compounds derived from Carthamus tinctorius L., in treating nonalcoholic fatty liver disease (NAFLD) by modulating the bile acid receptor NR1H4 (Nuclear Receptor Subfamily 1 Group H Member 4) and its associated metabolic pathways. A rat model of NAFLD was established, and RNA sequencing and proteomics were carefully employed to identify differential gene expressions associated with the disease. The active components of Carthamus tinctorius L. were screened, followed by the construction of a comprehensive network that maps the interactions between these components, NR1H4 and NAFLD-related pathways. Both in vitro (using HepG2 cells) and in vivo experiments were conducted to evaluate the effects on NR1H4 expression levels through Western blot and RT-qPCR analyses. Our findings identify NR1H4 as a pivotal target in NAFLD. Network pharmacology analysis indicates that quercetin and kaempferol play crucial roles in combating NAFLD, with in vitro and in vivo experiments confirming their ability to mitigate hepatocyte steatosis by enhancing NR1H4 expression. Notably, the protective effects of these compounds were inhibited by the NR1H4 antagonist guggulsterone, highlighting the importance of NR1H4 upregulation. This study demonstrates the novel therapeutic efficacy of quercetin and kaempferol from Carthamus tinctorius L. in treating NAFLD through NR1H4 upregulation. This mechanism contributes to the regulation of lipid metabolism, improvement of liver function, reduction of inflammation, and alleviation of oxidative stress, offering a promising direction for future NAFLD treatment strategies.

International expert consensus statement on physiological interpretation of cardiotocograph (CTG): First revision (2024).

The first international consensus guideline on physiological interpretation of cardiotocograph (CTG) produced by 44 CTG experts from 14 countries was published in 2018. This guideline ensured a paradigm shift from classifying CTG by arbitrarily grouping certain features of the fetal heart rate into different "categories", and then, randomly combining them to arrive at an overall classification of CTG traces into "Normal, Suspicious and Pathological" (or Category I, II and III) to a classification which is based on the understanding of fetal pathophysiology. The guideline recommended the recognition of different types of fetal hypoxia, and the determination of features of fetal compensatory responses as well as decompensation to ongoing hypoxic stress on the CTG trace. Since its first publication in 2018, there have been several scientific publications relating physiological interpretation of CTG, especially relating to features indicative of autonomic instability due to hypoxic stress (i.e., the ZigZag pattern), and of fetal inflammation. Moreover, emerging evidence has suggested improvement in maternal and perinatal outcomes in maternity units which had implemented physiological interpretation of CTG. Therefore, the guideline on Physiological Interpretation of CTG has been revised to incorporate new scientific evidence, and the interpretation table has been expanded to include features of chorioamnionitis and relative utero-placental insufficiency of labour (RUPI-L).

Effects of sitagliptin and L-theanine combination therapy on testicular tissue in rats with experimental diabetes.

This study examines the impact of the combination of sitagliptin and L-theanine on the testis tissue of rats with experimental diabetes. Diabetes mellitus, a chronic metabolic illness, significantly reduces quality of life and can cause male infertility by decreasing sperm count, motility, and testosterone levels. Rats were allocated to five separate groups: control, diabetes, L-theanine, sitagliptin, and combination therapy. The measurements encompassed blood glucose levels, body weight, serum insulin levels, and the Homeostasis Model Assessment of Insulin Resistance (HOMA-IR). The histological examination of testicular tissue was conducted using H&E, PASH, caspase-12, and PCNA staining techniques, in addition to a TUNEL assay to detect apoptosis. Levels of oxidative stress indicators, including glutathione peroxidase (GPX), malondialdehyde (MDA), and catalase, were also evaluated. The results showed that the group of individuals with diabetes had significantly higher levels of blood glucose, apoptotic indices, GPX, catalase, and MDA levels and activities in comparison with the control group. Although both the L-theanine and sitagliptin groups exhibited some improvement, the combination therapy demonstrated the most significant decrease in histopathological damage and apoptotic markers. These results indicate that the combination of sitagliptin and L-theanine may significantly decrease testicular damage caused by diabetes, making it a promising therapeutic strategy.

Resveratrol inhibits white adipose deposition by the ESR1-mediated PI3K/AKT signaling pathway.

Excessive adipose accumulation is the primary cause of obesity. Resveratrol (RES), a natural polyphenolic compound, has garnered significant attention for its anti-obesity properties. However, the precise mechanisms by which RES influences fat deposition have not yet been explored. In this study, the aim was to identify the target proteins and associated pathways of RES in order to elucidate the mechanisms by which RES reduces fat deposition. In this study, mice were administered 400 mg/kg of RES via gavage for 12 weeks. We found that while 400 mg/kg RES had no impact on the growth of the mice, it significantly reduced the weight of various white adipose tissues, as well as the serum and liver concentrations of total cholesterol and triglycerides. Network pharmacology identified 15 potential targets of RES and highlighted the PI3K/AKT signaling pathway as a key pathway. Molecular docking and dynamic simulations suggested that ESR1 might be the target protein through which RES exerts its anti-fat deposition effects. In vitro experiments revealed that ESR1 promotes the proliferation and inhibits the differentiation of 3 T3-L1 adipocytes, and suppresses the PI3K/AKT signaling pathway. Silencing the ESR1 gene altered the ability of RES to inhibit cell differentiation via the PI3K/AKT pathway. Gene expression results in subcutaneous adipose tissue, epididymal fat tissue, and liver tissue of mice were consistent with observations in cells. In summary, RES reduces white fat deposition by directly targeting the ESR1 protein and inhibiting the PI3K/AKT signaling pathway. Our findings provide new insights into the potential use of RES in the prevention and treatment of obesity.

HNRNPD/MAD2L2 axis facilitates lung adenocarcinoma progression and is a potential prognostic biomarker.

BACKGROUND: Although progress has been made in the treatment of LAUD, the survival rate for patients remains poor. An in-depth grasp of the molecular pathways implicated in LUAD progression is vital for improving diagnosis and treatment strategies. This study aims to explore novel molecular mechanisms driving LUAD progression and identify new potential prognostic biomarkers for LAUD patients. METHODS: Based on mass spectrometry analysis of human LUAD tissues, HNRNPD and MAD2L2 were identified as potential key proteins involved in LUAD progression. Subsequently, the interplay between HNRNPD and MAD2L2 was examined through dual-luciferase reporter assays, RNA-seq analysis, and various molecular biology techniques. Ultimately, the role of the HNRNPD/MAD2L2 axis in LUAD advancement and its potential as a prognostic indicator were investigated utilizing LUAD specimens, cell lines, and xenograft mouse models. RESULTS: In human LAUD tissues and cell lines, elevated levels of HNRNPD and MAD2L2 proteins were discovered. It was determined that HNRNPD binds to the MAD2L2 promoter, forming a regulatory axis at the transcriptional level. Subsequently, both in vitro and in vivo data demonstrated that the downregulation of the HNRNPD/MAD2L2 axis inhibited LUAD progression, while this effect could be rescued by MAD2L2 upregulation. Conversely, the upregulation of the HNRNPD/MAD2L2 axis facilitated LUAD progression, and this outcome could be reversed by MAD2L2 knockdown. Mechanistically, the downregulation of HNRNPD suppressed the promoter activity and transcription of MAD2L2, thus inhibiting the PI3K/HIF1α/ANGPTL4 pathway and tumor angiogenesis. Finally, it was confirmed that LUAD patients with high levels of both HNRNPD and MAD2L2 exhibited the poorest prognosis. Therefore, the HNRNPD/MAD2L2 axis has been identified as a potential predictive indicator for LUAD patients. CONCLUSIONS: The HNRNPD/MAD2L2 axis facilitates LUAD progression and serves as a potential prognostic biomarker.