Type 2 diabetes mellitus (T2DM) significantly impairs fracture healing, with neutrophils playing a crucial role in this process. In T2DM, these immune cells are over-activated, leading to the excessive release of neutrophil extracellular traps (NETs), increasing inflammation and hindering recovery. Thus, a need for markers to assess patients in the risk group arises. This study demonstrates that circulating cell-free DNA (cfDNA) can be efficiently quantified from serum samples by a single-step qPCR and be used as a marker for NETosis. Our results revealed that trauma patients with T2DM have the highest cfDNA levels, followed by trauma patients, and the healthy group has the lowest. The method shows strong correlations between cfDNA and neutrophil-specific markers such as MPO, citH3, AZU1, and α-defensin, highlighting its potential as a rapid indicator of NETosis. This approach could allow the timely interference for high-risk patients, ultimately improving healing outcomes and reducing complications such as chronic inflammation, non-union fractures, and diabetic foot ulcers.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a systemic condition associated with compromised bone integrity. Emerging evidence suggests that disturbances in hepatic vitamin D metabolism may contribute to these skeletal impairments. However, the hepatic mechanisms driving bone deterioration remain poorly defined. This study aimed to establish a human 3D in vitro model of MASLD and demonstrate that hepatic vitamin D dysregulation adversely affects bone homeostasis. Liver spheroids composed of HepaRG cells, LX-2 stellate cells, and HUVECs were stimulated with 600 µM free fatty acids (2:1 oleic: palmitic acid) to induce MASLD-like features, validated by BODIPY staining and gene expression. MASLD model induction led to downregulation of hepatic genes regulating lipid and vitamin D metabolism. ELISA confirmed significantly reduced 25-hydroxyvitamin D levels, aligning with downregulation of CYP2R1 and CYP27A1 . Transcriptomic profiling of human MASLD liver biopsies validated these molecular changes. To evaluate MASLD’s systemic impact on bone, THP-1-derived macrophages and SCP-1 mesenchymal stem cells were seeded onto bone scaffolds and co-cultured with spheroids. Bone scaffolds co-cultured with MASLD spheroids showed impaired mineralization and elevated expression of bone resorption markers. These findings mirror skeletal dysfunction observed in MASLD patients and suggest a mechanistic link between hepatic vitamin D dysregulation and bone pathology. This study introduces a pioneering 3D human liver-bone co-culture model that reveals MASLD-driven disruption of hepatic vitamin D metabolism as a direct contributor to bone deterioration. This 3D model develops a powerful translational platform for decoding systemic disease mechanisms and targeting the liver-bone axis therapeutically.

Undiagnosed diabetes and prediabetes are common among trauma patients and increase perioperative complication rates, yet routine glycemic screening at hospital admission is rarely performed. We compared 10 supervised machine learning (ML) algorithms for early detection of impaired glucose metabolism in 1789 trauma surgery patients (1095 normal; 694 prediabetes/diabetes) using questionnaire data and preoperative laboratory values within a nested cross-validation framework with hyperparameter optimization. On the combined feature set, XGBoost achieved the highest area under the receiver operating characteristic curve (ROC-AUC; 0.815), followed by multilayer perceptron (0.814) and Gradient Boosting (0.813); adding blood parameters significantly improved performance for nine of ten algorithms (DeLong’s test, p < 0.05). SHapley Additive exPlanations (SHAP) and XGBoost feature gains identified six overlapping predictors, yielding a simplified model with comparable mean discrimination. Inter-model differences were small, and a fully interpretable logistic regression model remained within one percentage point of XGBoost. The best-performing model demonstrated strong calibration and a higher net benefit than either universal testing or no testing across the evaluated threshold probabilities. In routine trauma practice, such a model could identify patients who would benefit from confirmatory glycated hemoglobin (HbA1c) testing, enabling earlier detection of dysglycemia, risk-stratified glucose management, and timely referral for lifestyle intervention in patients with prediabetes.

Chronic wounds represent an increasing global health burden for both patients and healthcare systems. These non-healing lesions frequently result in amputation and are particularly prevalent among individuals with diabetes. Hallmarks of chronic wounds include a prolonged inflammatory phase characterized by sustained neutrophil infiltration, excessive formation of neutrophil extracellular traps (NETs), and an elevated alkaline tissue pH. Emerging evidence suggests a link between pH and wound healing, with acidic conditions being more favorable for tissue repair. Here, we investigated the pH-dependent modulation of primary neutrophil functions, including viability, reactive oxygen species (ROS) production, NET formation, and exocytosis. Neutrophils cultured in media with pH values ranging from 6.1 to 8.4 exhibited no significant differences in viability. In contrast, acidic conditions suppressed ROS production and NET formation in a pH-dependent manner, whereas alkaline conditions caused responses comparable to those observed at physiological pH (7.4). Furthermore, levels of antioxidant enzymes were markedly increased under acidic conditions following phorbol 12‑myristate 13‑acetate (PMA) stimulation. Notably, alkaline pH promoted granule polarization in neutrophils, thereby impairing exocytosis. In contrast, acidic conditions (pH < 7.0) reduced granule polarization and enhanced the release of granule-associated proteins upon PMA stimulation. Pharmacological inhibition of degranulation confirmed that these effects were mediated by neutrophil exocytosis. In summary, extracellular pH critically modulates neutrophil effector functions and may contribute to the distinct healing dynamics observed in acute versus chronic wounds.