Search Results (14,020)

Alpha-fetoprotein (AFP) is a glycoprotein primarily produced by the fetal liver and yolk sac during development. It is a multifaceted biomarker with significant applications in the prenatal screening of congenital abnormalities, cancer, and other disorders. The level of AFP in maternal blood may indicate several obstetric concerns and complications during pregnancy. Atypical AFP levels are commonly utilized as a biomarker for detecting fetal anomalies, placental complications, and other pregnancy-related issues. These findings raise concerns regarding the effectiveness of screening maternal serum alpha-fetoprotein (MS-AFP) as a primary indicator of pregnancy problems and underscore the need for further investigation into the functional role of AFP throughout pregnancy. The measurement of MS-AFP has been utilized for the past four decades. It is anticipated that MS-AFP measurement will continue to be utilized as a component of integrated or sequential tests for chromosomal abnormalities and may serve as a prognostic indicator for adverse obstetric outcomes. Critically, whether AFP functions solely as a passive marker or plays active biological roles in pregnancy physiology and pathology remains unresolved, necessitating additional mechanistic investigation and discourse. This review consolidates critical data from numerous studies on AFP, focusing specifically on its diagnostic and prognostic applications for congenital abnormalities and problems during pregnancy. This review also identifies key research gaps regarding the functional biology of AFP, particularly whether AFP functions as a passive biomarker or an active participant in the pathophysiology of adverse pregnancy outcomes. Full article

Blood disorders encompass a wide range of diseases including anemia, hemophilia, thrombotic disorders, platelet dysfunction, and hematological cancers, making blood disorders a major global health concern. These conditions can impair processes vital to human physiology including oxygenation, coagulation, and immune defense. Hematologic malignancies, both chronic and acute, require timely diagnosis and ongoing disease monitoring for effective clinical management. Microfluidic technologies have emerged as promising alternatives to benchtop techniques for diagnosing and monitoring hematological disorders. For example, microfluidic assays can be used for the isolation and characterization of liquid biopsy markers such as rare cells, extracellular vesicles, and cell-free molecules to support disease management in a minimally invasive manner while the process automation afforded by microfluidics decentralizes healthcare, making it more accessible. Advances in lab-on-a-chip technologies, including large-scale fabrication methods and novel design strategies, will provide tools for the clinical validation of biomarkers and the translation of these technologies from the laboratory bench to the patient bedside. In this review, we will show that microfluidic devices enable disease monitoring via high-throughput analysis of liquid biopsy samples for the detection of rare disease-specific biomarkers found in blood, plasma, urine, etc., providing an alternative to standard benchtop testing using specimens secured via invasive bone marrow procedures, typically used for managing blood-based diseases. A key advantage of microfluidics is their ability to manipulate blood components at scales that closely mimic the body’s microvascular environment. Not surprisingly, microfluidic vascular models have been developed to replicate physiological rheology enabling quantitative assessment of blood cell deformability, aggregation, or clot formation. We provide a critical perspective on the use of the microfluidic “organ-on-chip” designed for blood disorders’ modeling and employed to recapitulate the blood cancer microenvironment. A summary of advances in microfluidic strategies for detection, diagnosis, drug screening, and mechanistic investigations of blood disorders, and future directions for precision testing, will be presented. Full article

Background: Serum concentrations of 25-hydroxyvitamin D [25(OH)D] are associated with the risk of several chronic and acute diseases. However, updated data on vitamin D status in Mediterranean countries, including Italy, remain limited, hindering effective public health strategies. Objective: To assess serum 25(OH)D levels and their seasonal variation in healthy blood donors aged 18–65 years living in Northern Italy and not taking vitamin D supplements. Given the latitude and the high levels of environmental pollution, cutaneous vitamin D synthesis may be impaired in this population. Recent Italian guidelines on supplementation emphasize the need for updated data on the prevalence of hypovitaminosis D and seasonal variation in endogenous vitamin D synthesis. Methods: In this exploratory retrospective cross-sectional study, 534 blood donors (268 men and 266 women) attending the Transfusion Medicine Unit of the Verona University Hospital were enrolled between April 2016 and May 2018. Serum 25(OH)D concentrations were analyzed by season. Clinical, lifestyle, pharmacological and dietary characteristics were also collected. Results: Among healthy, normal-weight individuals, the prevalence of vitamin D insufficiency (25(OH)D < 50 nmol/L) was low and limited to one-two months per year. Overweight and obesity significantly reduced the likelihood of achieving adequate 25(OH)D levels through cutaneous synthesis for several months. Mean 25(OH)D concentrations were higher than those previously reported in the same area, while seasonal variation remained preserved. Conclusions: In a relatively small non-supplemented population of blood donors living in a high polluted urban area of Northern Italy, seasonal vitamin D synthesis seems to be preserved. These updated data show higher 25(OH)D levels compared to past findings. Although these data certainly warrant further validation through a national survey involving other regions of Italy and in not selected population, they appear to be in line with the SIOMMMS recommendations against indiscriminate serum 25(OH)D testing and against routine supplementation for healthy normal-weight individuals under 70 years. Full article

Tomatoes are the major dietary source of lycopene, a carotenoid that crosses the blood–brain barrier and exerts antioxidant and anti-inflammatory effects. However, the impact of tomato consumption on cognitive function in healthy adults remains unclear. This study assessed the effects of concentrated tomato paste on cognitive performance and explored potential mechanisms, including brain-derived neurotrophic factor (BDNF) and functional brain connectivity. A randomized, two-period crossover trial (ClinicalTrials.gov: NCT05891977) was conducted in 47 healthy adults aged 40–55 years assigned to two 3-month interventions separated by a 1-month washout: (a) daily consumption of concentrated tomato paste (0.5 g/kg body weight) and (b) a lycopene-restricted control diet. Cognitive performance was evaluated using validated neuropsychological tests (d2-R, Face-Name Associative Memory Exam, Modified Wisconsin Card Sorting Test), alongside plasma lycopene and BDNF, and resting-state functional magnetic resonance imaging (fMRI). Forty-two participants completed the study. Tomato intake improved selective attention (concentration performance: +7.2 points; processing speed: +8.3 points) and associative memory (face-name matching: +0.8 points). Plasma BDNF showed a borderline increase with tomato intake (mean difference 15.2 ng/mL). Resting-state fMRI revealed changes in brain networks, including reduced connectivity in frontoparietal and auditory networks, contrasting with reductions in the dorsal attention network during the control period. These findings provide evidence that tomato consumption may support cognitive function and modulate brain connectivity in healthy middle-aged adults. Full article

Background and Objectives: SGLT2 inhibitors increase hemoglobin and hematocrit in multiple clinical settings, an effect increasingly attributed to stimulation of erythropoiesis rather than hemoconcentration. However, most mechanistic evidence derives from diabetic populations, leaving uncertainty as to whether this response depends on diabetes-related metabolic changes. To evaluate whether dapagliflozin stimulates erythropoiesis in non-diabetic patients with heart failure and to determine whether hematologic changes correlate with renal, cardiac, inflammatory, hepatic, or iron-related parameters. Materials and Methods: In this retrospective observational study, each of 68 non-diabetic heart failure patients served as their own control. Hematologic, renal, cardiac, inflammatory, hepatic, and iron parameters were assessed at three time points: one year prior to dapagliflozin initiation, at baseline, and one year after initiation of therapy. Changes were analyzed using paired tests and correlation analyses. Results: Hemoglobin, hematocrit, and red blood cell count were significantly lower at the baseline compared with values recorded one year before dapagliflozin initiation and increased significantly during the year following treatment (all p < 0.001), while mean corpuscular indices remained stable. Serum iron decreased before treatment and increased significantly after dapagliflozin initiation (p < 0.05 vs. baseline); however, changes in serum iron did not correlate significantly with changes in hemoglobin after treatment. Inflammatory markers showed a modest reduction in C-reactive protein after treatment, while composite inflammatory indices remained largely stable. Liver enzymes showed no significant longitudinal changes. Correlation analyses demonstrated no association between changes in hemoglobin and changes in eGFR (ρ = 0.202, p = 0.098) or NT-proBNP (ρ = −0.003, p = 0.981) after treatment. Hematologic variables remained strongly intercorrelated, whereas cross-system correlations were minimal, indicating that erythropoietic stimulation occurred largely independently of renal or cardiac functional trajectories. Conclusions: Dapagliflozin robustly stimulates erythropoiesis in non-diabetic patients with heart failure, independent of improvements in kidney or cardiac function. Although serum iron levels improved after treatment, the absence of a direct correlation with hemoglobin suggests that iron mobilization may act as a permissive rather than a primary driver of erythropoietic response. These findings support the concept that erythropoiesis represents a diabetes-independent pharmacologic action of SGLT2 inhibitors and may involve renal, hepatic, inflammatory, and iron-regulatory pathways beyond those described in diabetic physiology. Dedicated mechanistic studies in non-diabetic populations are warranted. Full article

Background: Pulmonary arterial hypertension is a rare but life-threatening condition in children, with hereditary forms often being linked to mutations in genes such as bone morphogenetic protein receptor type 2 (BMPR2), caveolin 1 (CAV1), and potassium channel subfamily K member 3 (KCNK3). Among these, CAV1 mutations are associated with severe disease phenotypes, though cases resulting from de novo heterozygous CAV1 mutations with multi-system involvement remain rarely reported. The CAV1 mutation (c.424C > T, p.Q142X) disrupts caveolin-1 function, leading to dysregulated pulmonary vascular remodeling and multi-system abnormalities. Methods: This was a retrospective case study of a pediatric patient with hereditary PAH. The patient was followed at our hospital from initial presentation until death. Clinical data were collected from medical records, including physical examinations, laboratory tests, echocardiography, chest X-ray, computed tomography pulmonary angiography (CTPA), and genetic analysis. The patient was treated sequentially with various PAH-targeted medications. This report also includes a review of the relevant literature on CAV1-associated PAH. Results: A female aged 3 years and 11 months was diagnosed with hereditary PAH associated with a de novo heterozygous CAV1 mutation (c.424C > T, p.Q142X). Both parents underwent genetic testing and were negative for the mutation, confirming its de novo origin. Clinical manifestations included special facial features, congenital telangiectasia, cutis marmorata (marbled skin), congenital cataract, hereditary lipodystrophy, and severe PAH. The patient presented with progressive exercise intolerance, syncope, and worsening dyspnea over nine years. Echocardiography revealed pulmonary hypertension with an estimated pulmonary artery systolic pressure of 69–105 mmHg, right heart enlargement, right ventricular hypertrophy, and moderate tricuspid regurgitation. Blood and urine metabolic screenings were normal. A chest X-ray showed progressive enlargement of the cardiac silhouette and bulging of the pulmonary artery segment. CTPA demonstrated pulmonary hypertension, secondary right heart dysfunction, decompensated right ventricular function, and mosaic perfusion in both lungs, suggestive of small arterial branch occlusion. Right heart catheterization was declined by the parents. Thus, the diagnosis of PAH was established based on clinical, echocardiographic, CTPA, and genetic findings. The patient was hospitalized four times and lost to follow-up from 2017 to 2023. She received sequential treatment with digoxin, hydrochlorothiazide, tadalafil, ambrisentan, selexipag, and treprostinil. Despite these therapies, pulmonary artery pressure continued to rise with progressive clinical deterioration. The patient ultimately died at 13 years of age due to a pulmonary hypertensive crisis and multiple organ failure following a severe episode of gastroenteritis. Conclusions: Despite aggressive treatment with multiple targeted reduced pulmonary artery pressure drug therapies, managing hereditary PAH caused by CAV1 mutations in children remains a significant challenge, with a high mortality rate. Early genetic diagnosis, regular follow-up, and individualized treatment are crucial. It requires the joint efforts of patients, parents, and healthcare providers. Full article

Background: The plant Griffonia simplicifolia is marketed as a dietary supplement; it is said to have antidepressant and sleep-promoting properties. Its main ingredient, 5-hydroxytryptophan (5-HTP), is the immediate precursor of serotonin and crosses the blood–brain barrier, thereby enhancing central serotonergic neurotransmission. Reduced serotonergic activity has been associated with affective disorders, sleep disturbances, and impaired central pain modulation. Despite this neurobiological rationale, evidence for analgesic efficacy remains limited. This study investigated the effects of Griffonia simplicifolia on peripheral and central sensitization and descending pain inhibition. Methods: In a randomized, double-blind, placebo-controlled crossover trial, 18 healthy volunteers underwent quantitative sensory testing (QST). Participants received 100 mg Griffonia simplicifolia orally once daily for 28 days or matching placebo. Sensory parameters were reassessed, followed by repetitive phasic heat application (RPHA) to induce short-term peripheral and central sensitization. After a 4-week washout period, participants crossed over to the alternate intervention. Results: A total of 17 participants completed the study. Griffonia simplicifolia showed no significant effect on acute pain perception after RPHA (β = −4.17; 95% CI −14.44 to 6.10; p = 0.401). The only significant difference was an increased distance of mechanical allodynia in the verum group (β = 0.82; 95% CI 0.05–1.59; p = 0.038). No differences were observed in thermal detection or pain thresholds, pressure pain thresholds, conditioned pain modulation, wind-up ratio, mechanical pain sensitivity, or flare area. Mild, transient adverse events occurred in two participants (11%) during Griffonia simplicifolia intake. Conclusions: Griffonia simplicifolia demonstrated limited effects on experimentally induced pain mechanisms compared with placebo and was well tolerated. Increased distance of allodynia may reflect serotonergic facilitation of pronociceptive pathways, suggesting an enhanced central and peripheral sensitization. Larger controlled trials are required to clarify its impact on pain perception. Full article

The scientific evidence regarding the use of plant-derived extracts to alleviate exercise-induced muscle damage in horses remains limited. Mulberry leaf flavonoids (MLFs) are the primary bioactive constituents of a traditional medicinal plant and are potent antioxidants. The aim of this study was to investigate the protective effects of MLFs against exercise-induced muscle damage. In this study, twelve Mongolian horses were used in a 3 × 3 Latin square design to investigate the protective effects of MLFs. Our results showed that high-intensity exercise negatively impacted the immune status, metabolic state, myofibrillar structure, and antioxidant capacity of the horses. Conversely, MLFs significantly reduced blood levels of white blood cells (WBC), monocytes (MON), aspartate aminotransferase (AST), creatine kinase (CK), and malondialdehyde (MDA) across various exercise distances and during recovery. Simultaneously, MLFs increased serum glutathione peroxidase (GPx), superoxide dismutase (SOD), and total antioxidant capacity (T-AOC). Mechanistically, transcriptomic analysis revealed that dietary MLFs upregulated genes associated with myofibrillar structural proteins (MYOZ2, MYOM3), the antioxidant defense system (GPX3, SOD3), and skeletal muscle satellite cell proliferation and differentiation (MYOD1, MRF6). Furthermore, quantitative proteomics indicated the enrichment of the PI3K-Akt and TGF-β signaling pathways, as well as ECM–receptor interactions, suggesting their potential involvement in regulating protein metabolism and facilitating myofibrillar restoration. Overall, MLFs effectively alleviated inflammation, metabolic disorder, and exercise-induced muscle damage. Under the tested conditions, a daily dosage of 10 g MLFs provided superior protective effects. Full article

Background/Objectives: Sacral giant cell tumor of bone (GCTB) is a rare, mostly benign but locally aggressive neoplasm that carries significant diagnostic and treatment challenges due to its anatomic complexity, proximity to sacral nerve roots as well as the blood vessels, and potential impact on bowel, bladder, sexual, and lumbopelvic function and stability. This narrative review aimed to synthesize current evidence on the epidemiology, clinical presentation, diagnostic evaluation, classification, management strategies, outcomes, and surveillance of sacral GCTB. Methods: A focused literature search of PubMed/MEDLINE and Google Scholar was conducted for studies published between January 2000 and January 2026, with additional manual review of reference lists. Given the rarity of the tumor and the observed heterogeneity in study designs, treatment strategies, and outcomes, the evidence was synthesized narratively. Results: Sacral GCTB mainly affects young adults with an indolent nature, often presenting late with progressive low back or buttock pain, radiculopathy, or neurological deficits. Magnetic resonance imaging is the preferred modality for determining local extent, whereas histopathologic biopsy and molecular testing remain essential for definitive diagnosis. Conventional grading systems, such as Enneking and Campanacci, have limited value in sacral disease, as anatomical extent and anticipated neurological morbidity are more relevant for treatment planning. Surgery remains the cornerstone for resectable disease, yet management plans should balance local tumor control against preservation of sacral nerve roots and mechanical stability. Denosumab, selective arterial embolization, and radiotherapy may play important roles in selected unresectable or high-morbidity cases. Local recurrence remains a major concern, and long-term surveillance is recommended because tumor relapse, treatment-related morbidity, and distant metastasis may occur late. Conclusions: Current evidence supports a multidisciplinary, individualized approach to sacral GCTB, guided by tumor extent, expected neurological morbidity, and patient-centered functional outcomes. Prospective multicenter studies are needed to refine treatment algorithms and improve risk stratification. Full article

DNA must be efficiently extracted from samples to accurately test the authenticity of food, particularly from processed matrices in which DNA integrity may be compromised. We systematically evaluated the efficiency of extracting DNA from dairy and blood products by four methods, namely SDS-CTAB, SDS-isopropanol precipitation, guanidine isothiocyanate magnetic beads, and a commercial kit. The guanidine isothiocyanate-magnetic bead method yields high quantities and purity of DNA; for example, the yield obtained from chicken blood samples was 318.34 ± 4.77 ng/µL, with an A260/A280 ratio ranging from 1.8 to 2.0. The processing time of this method was compared with the DNA Extraction Kit shorter by 40% and unlike methods such as the SDS-CTAB protocol, does not require the use of toxic reagents such as phenol or chloroform, meeting green chemistry requirements. Among the dairy and blood samples tested, it enables the extraction of DNA in quantities comparable to those obtained using commercial kits; moreover, the DNA yield achieved is 20–30% higher than that of these kits. Furthermore, this method is free from the limitations associated with protein contamination and amplification instability often encountered in protocols such as the CTAB-SDS and SDS-isopropanol methods. The magnetic bead approach was adaptable for complex matrices and demonstrated strong tolerance to coexisting contaminants, thereby improving extraction performance in challenging food samples. The magnetic bead surface functionalization and buffer systems could be improved to further increase their versatility. This method enables reliable DNA extraction and advanced technical support for DNA analysis. Full article

Malaria in a Greek citizen with prior malaria history residing and working in Cameroon returning in his home country is a medical emergency warranting prompt and accurate diagnosis and effective treatment. We describe a mixed malaria case of a febrile patient, a professional returning to Greece from a malaria-endemic country whose initial diagnosis was a false-negative malaria rapid diagnostic test. Subsequent alternative rapid diagnostic test, malaria thin-film blood examination and molecular diagnosis revealed mixed malaria infection from Plasmodium ovale and Plasmodium malariae. The patient was successfully treated and achieved complete clinical recovery. The case described here highlights important points regarding prompt and accurate malaria diagnosis in returning travelers in non-endemic countries, emphasizing the importance of revealing cryptic mixed malaria cases and providing molecular approaches to malaria diagnosis in combination with the gold-standard microscopy. Full article

Usutu virus (USUV) is a mosquito-borne flavivirus, widely distributed in Central Europe, where it causes avian outbreaks with large-scale mortality. Although most human infections are asymptomatic or mild, the reports of USUV neurologic infections are increasing, especially among immunocompromised patients. Cross-reactivity in serological and molecular assays is often seen between USUV and the genetically and antigenically related West Nile virus (WNV). Here, we report the first USUV infection in Greece in an asymptomatic blood donor who tested positive in the automated nucleic acid test during screening for WNV, which is endemic in the country. Following the blood donation surveillance plan, a serum sample taken two weeks post-donation was tested for WNV IgM and IgG antibodies. The borderline index of the IgM antibodies, combined with negative result for IgG antibodies, raised the suspicion of molecular cross-reactivity with USUV. Although the USUV-specific PCR in donor’s plasma was negative, its result was positive following amplification of the virus in cell culture, as USUV RNA was detected in the culture supernatant confirming the USUV infection. Whole genome sequence was taken using an Ion Torrent next-generation sequencing platform. Phylogenetic analysis showed that the Greek strain clusters within the USUV Europe 2A genetic lineage. The detection of USUV human infection in Greece prompts for surveillance studies to elucidate its epidemiology and ecology in the country. Full article

Background/Objectives: Hypoxia and hypoxia-inducible factors (HIFs) have emerged as pivotal factors in the pathophysiology of chronic rhinosinusitis (CRS). Obstructive sleep apnea (OSA) may exacerbate hypoxia-driven sinonasal inflammation. This study evaluated the association between OSA risk and sinonasal HIF expression in CRS patients, focusing on the distinct profiles of eosinophilic (ECRS) and non-eosinophilic (NECRS) endotypes. Methods: Ethmoid mucosal tissues were collected from 64 CRS patients undergoing surgery. Patients were classified into ECRS or NECRS groups based on blood eosinophil counts, and into high- or low-risk OSA groups based on apnea-hypopnea index on polysomnography or sleep domain score from the 22-item sinonasal outcome test. Protein levels of HIF-1α, HIF-2α, and various inflammatory mediators were measured via multiplex immunoassay. Results: HIF-2α expression was significantly higher in the high-risk OSA group (116.80 ± 131.48 vs. 47.37 ± 42.14, p < 0.05), whereas HIF-1α levels were independent of OSA status. Following stratification by endotype, HIF-1α expression was significantly higher in NECRS than in ECRS (0.042 ± 0.020 vs. 0.034 ± 0.024, p < 0.05). Notably, high-risk OSA was associated with markedly increased HIF-2α only within the NECRS subgroup (115.52 ± 61.07 vs. 47.97 ± 31.03, p < 0.05). Correlation analyses demonstrated endotype-specific inflammatory coupling, showing that HIF-1α and HIF-2α were selectively linked to MMP-9 (r = 0.689, p < 0.05) and neutrophil-related markers (r = 0.925, p < 0.05) in NECRS while exhibiting broader cytokine correlations in ECRS (p < 0.05). Conclusions: Sinonasal HIF expression in CRS varies according to OSA risk and CRS endotype. OSA-associated hypoxic stress preferentially influences HIF-2α expression in NECRS, whereas this effect is attenuated in ECRS, likely due to the dominance of local type 2 inflammatory signaling. Full article

Background: Peripheral artery disease (PAD) impacts more than 200 million individuals globally. Despite its prevalence, management remains suboptimal, partly due to the lack of reliable blood-based biomarkers. The ankle–brachial index (ABI), the current gold-standard test for PAD, is limited by inter-operator variability, misinterpretation, and reduced accuracy in patients with diabetes. Fatty acid binding protein 3 (FABP3) has emerged as a potential biomarker for PAD; however, its prognostic performance relative to ABI remains unclear. This study compared FABP3 and ABI for predicting PAD outcomes using statistical and machine learning approaches. Methods: A total of 1001 participants were prospectively recruited, including 644 patients with PAD and 357 without PAD. The primary outcome was 2-year major adverse limb event (MALE), defined as a composite of vascular intervention, major amputation, or acute limb ischemia. At enrollment, plasma FABP3 was quantified using a validated multiplex immunoassay. Kaplan–Meier analysis of MALE-free survival was performed across pre-specified FABP3 tertiles (high [>3.55 ng/mL], moderate [1.55–3.55 ng/mL], and low [<1.55 ng/mL]) and ABI tertiles (severe [<0.40], moderate [0.40–<0.70], and mild [0.70–0.90]), with curve separation assessed using log-rank tests. Multivariable Cox proportional hazards modelling was used to evaluate the independent relationships of FABP3 and ABI with 2-year MALE after adjustment for baseline demographic and clinical covariates. To assess predictive performance for 2-year MALE, an extreme gradient boosting (XGBoost) classification model incorporating 10-fold cross-validation was trained using a combination of clinical covariates, plasma FABP3 levels, and ABI. Discriminatory performance was assessed using the area under the receiver operating characteristic curve (AUC). Results: The average participant age was 68 years (SD 12), and 34% (n = 340) were women. Mean ABI was 0.75 ± 0.25 and mean FABP3 concentration was 2.97 ± 2.06 ng/mL. Among the 644 participants with PAD, 558 (86.6%) had complete time-to-event data for MALE status, FABP3, and ABI. Over the median follow-up period of 2 years, 140 (25.1%) participants with PAD experienced MALE. Kaplan–Meier analyses demonstrated significant separation in MALE-free survival across FABP3 tertiles (log-rank p < 0.001). At 24 months, MALE-free survival was 100.0% in the FABP3 < 1.55 group, compared with 71.1% in the FABP3 1.55–3.55 group and 67.7% in the FABP3 > 3.55 group. In contrast, ABI severity groups showed less pronounced separation, with 24-month MALE-free survival rates of 80.3% for mild ABI, 73.2% for moderate ABI, and 71.3% for severe ABI, without a statistically significant overall difference (p = 0.170). In adjusted Cox proportional hazards models, FABP3 demonstrated strong prognostic performance for 2-year MALE. A 1 SD increase in log-transformed FABP3 was independently associated with a higher risk of 2-year MALE (HR 1.90, 95% CI 1.60–2.25; p < 0.001), with minimal change after additional adjustment for ABI (HR 1.90, 95% CI 1.60–2.24; p < 0.001). Machine learning analyses similarly favored FABP3 over ABI, with the FABP3-based model achieving an AUC of 0.773 compared to 0.686 for the ABI-based model. Adding ABI to the FABP3 model did not improve discrimination. Conclusions: Circulating plasma levels of FABP3 are strongly associated with PAD outcomes. Specifically, FABP3 demonstrated a stronger and more robust association with 2-year MALE compared to ABI. This study validates the prognostic value of FABP3 for PAD outcomes in comparison to ABI. Full article

Objectives: Seasonal influenza leads to substantial global morbidity and mortality, especially in adults aged 65 years and older, who present poor immune responses to standard-dose influenza vaccines. This study aimed to systematically evaluate the preclinical safety and immunogenicity of a high-dose quadrivalent seasonal influenza split vaccine (HD-QIV), providing preclinical evidence for its clinical application in the elderly. Methods: Following GLP guidelines, we performed single-dose and repeated-dose toxicity tests in Sprague–Dawley rats, active systemic anaphylaxis assays in guinea pigs, and immunogenicity assessments in young and aged BALB/c mice. Safety indicators included general clinical signs, hematology, blood biochemistry, histopathology and allergic reactions; immunogenicity was evaluated via hemagglutination inhibition (HI) antibody titers and antigen-specific cellular immune responses. Results: HD-QIV only caused mild and reversible local irritation in rats without obvious systemic toxicity, and no dose-related systemic anaphylaxis was observed in guinea pigs. HD-QIV induced robust and dose-dependent humoral immune responses, and showed significantly higher HI antibody titers, earlier seroconversion and longer antibody persistence than standard quadrivalent influenza vaccine in aged mice. Cellular immunity was slightly enhanced but not the dominant protective response. Conclusions: The HD-QIV demonstrates favorable preclinical safety and superior immunogenicity, supporting its further clinical development and use as a priority vaccine for the elderly population. Full article