The researchers extended a previously validated cellular Potts model into a coupled ordinary-differential-equation, partial-differential-equation and cellular framework. The molecular component used 16 equations to represent shear-induced AKT activation, endothelial nitric oxide synthase phosphorylation and nitric oxide production. Tissue-scale equations modeled the diffusion, production and decay of nitric oxide, reactive oxygen species and the chemokine MCP-1. At the cellular scale, a modified Monte Carlo algorithm updated endothelial states, cell movement, adhesion, proliferation, monocyte migration and macrophage differentiation. The simulations used a two-dimensional arterial slice measuring 0.8 by 0.2 millimeters, with one Monte Carlo step corresponding to one minute of biological time and a total duration of seven days.

Mechanical loading was prescribed rather than calculated from a resolved blood-flow field. The wall shear-stress amplitude varied spatially from 5 to 15 dyn/cm2 in a symmetric V-shaped profile, with its minimum at the center, and a cosine function introduced periodic reversal of flow direction. This configuration represented the low, oscillatory stresses associated with vulnerable regions such as arterial bends and bifurcations, but the model did not solve fluid velocity or direction explicitly. Each live endothelial cell contained the molecular signaling module, while its nitric oxide output entered the surrounding chemical field. Activated endothelial cells and pro-inflammatory M1 macrophages generated reactive oxygen species, and a non-linear Michaelis-Menten term represented direct chemical consumption of nitric oxide by reactive oxygen species, including formation of peroxynitrite.

In the untreated simulation, endothelial injury progressively worsened. Quiescent endothelial cells began disappearing around day 1, activated cells became the dominant endothelial population, and dead endothelial cells increased. By the end of the seven-day period, activated cells represented approximately 70% of the endothelial population, dead cells about 30%, and quiescent cells nearly none. Activated cells and M1 macrophages also secreted MCP-1, creating a chemical gradient that directed monocytes toward the injured region. Monocytes that contacted activated endothelium differentiated into M1 or M2 macrophages at a preset ratio of 2 to 1. At day 7, the approximate conversion proportions were 0.5 for M1 macrophages, 0.3 for M2 macrophages and 0.2 for dead monocytes, indicating a predominantly pro-inflammatory modeled environment.

The temporal redox results showed why total nitric oxide concentration alone was an inadequate measure of endothelial health. Nitric oxide increased during the simulation, but reactive oxygen species accumulated more rapidly at later stages. The contribution of nitric oxide from quiescent endothelial cells fell toward zero around day 1.5, because cells in the low-stress region crossed the model’s damage threshold and became activated. Their impaired production continued at roughly 0.6 times the quiescent-cell rate, allowing total nitric oxide to rise even as functional protection deteriorated. The model therefore attributes localized injury not to a universal shortage of nitric oxide, but to the mismatch between where nitric oxide is produced and where reactive oxygen species consume it. In the model, this chemical imbalance also reinforced inflammatory signaling and limited the return of activated cells to a quiescent state.

Validation focused on the molecular core rather than on direct testing of the complete repair model. Under static conditions, a 2.5-fold increase in simulated AKT activity predicted a 2.17-fold increase in endothelial nitric oxide synthase phosphorylation; published Western blot data reported a 2.27-fold increase after AKT overexpression. Simulations under different pulsatile shear stresses and after PI3K, AKT or endothelial nitric oxide synthase inhibition reproduced the trends and approximate magnitudes reported in experimental studies. Predicted nitric oxide concentrations at shear stresses of 8, 16 and 24 dyn/cm2 also closely matched a previously published reference model, including cases with enhanced AKT signaling. These checks support the parameterization of the AKT-to-nitric-oxide pathway, but they do not independently validate every cellular or biochemical rule in the larger multiscale framework.

Simulated intervention timing had a substantial effect on recovery. For this comparison, AKT activation was set at four times the baseline activation parameter. Starting treatment on day 1 produced the most favorable pattern: endothelial layers on either side of the central injury remained intact, activated cells began returning to a quiescent phenotype by day 5, reactive oxygen species accumulation was restrained and MCP-1 declined by day 7. Quiescent-cell recovery began around day 6, while the final proportion of quiescent cells exceeded that in the delayed-treatment groups. Starting on day 3 improved the outcome relative to no intervention, but nitric oxide production rose only after day 5, oxidative stress persisted and inflammation was stronger. Starting on day 5 produced little conversion back to quiescent cells, the lowest late nitric oxide peak, prominent central reactive oxygen species accumulation and the highest MCP-1 signal among the treatment groups.

The model also identified a non-linear response to activation strength when treatment began on day 3. Three simulated levels were compared: 3.4 times baseline as weak activation, 3.8 times as moderate activation and 4.2 times as strong activation. Under weak activation, the first activated-to-quiescent transition occurred at about 146 hours, nitric oxide recovery was slow and both reactive oxygen species and MCP-1 remained elevated. Moderate activation moved the first transition to 134 hours, produced a more uniform nitric oxide field and more closely balanced reductions in oxidative stress and inflammation. Strong activation advanced the transition to 124 hours, accelerated the fall in reactive oxygen species and drove the activated-cell proportion close to zero by the end of the simulation. However, the authors characterized the moderate condition as the most balanced combination of nitric oxide restoration, oxidative control and inflammatory resolution; the model did not explicitly simulate all downstream consequences of maximal AKT activation.

Immune effects in the simulations arose primarily through altered recruitment rather than a direct change in macrophage polarization. Because the M1-to-M2 differentiation ratio was fixed at 2 to 1, the model could not test whether AKT directly alters the choice between macrophage phenotypes. Instead, restoring endothelial function reduced the number of activated cells, which lowered their MCP-1 output and weakened the chemotactic signal drawing in monocytes. It also reduced cellular reactive oxygen species production, improving the local redox environment and indirectly changing the abundance of macrophages. This coupling created a feedback loop: better nitric oxide availability favored endothelial recovery, recovery reduced inflammatory recruitment, and lower inflammation reduced further damage. It also explains why delayed intervention performed poorly: newly generated nitric oxide entered an environment already loaded with reactive oxygen species that could rapidly neutralize it.

The findings remain limited by the model’s scope. It is a two-dimensional representation that does not capture the three-dimensional architecture of the vessel wall or diffusion barriers such as the internal elastic lamina. Its biochemical network concentrates on the AKT/eNOS/NO/ROS axis and omits crosstalk involving pathways such as Notch and TGF-β, as well as downstream effects of peroxynitrite, including possible eNOS uncoupling. Monocyte recruitment is regulated mainly by endothelial activation rather than directly by flow-dependent adhesion, and other immune cells and a wider cytokine network are absent. The results therefore provide a computational account of timing- and activation-dependent endothelial repair within the model, but they do not establish a clinical dosing schedule, demonstrate benefit in patients, or show that a particular AKT-targeted therapy can safely produce the modeled effects.

Subject of Research: Multiscale modeling of AKT-mediated endothelial repair under oscillatory shear stress

Article Title: Multiscale modeling reveals non-linear dynamics of AKT-mediated NO-ROS crosstalk in shear stress-induced endothelial phenotype

Article References: Zhang, Y.-Y., Wang, Y.-T., Qiu, Y., Li, Y.-J., Chen, D., & Qin, K.-R. (2026). Multiscale modeling reveals non-linear dynamics of AKT-mediated NO-ROS crosstalk in shear stress-induced endothelial phenotype. PLOS Computational Biology, 22(10), e1014860. https://doi.org/10.1371/journal.pcbi.1014860

Image Credits: AI Generated

DOI: 10.1371/journal.pcbi.1014860

Keywords: endothelial dysfunction, AKT signaling, nitric oxide, reactive oxygen species, oscillatory shear stress, atherosclerosis, multiscale modeling, inflammation, computational biology, vascular repair, Multiscale, modeling

The research team, working with specimens collected during Antarctic expeditions in 2014 and 2016, sequenced the fungus using both Illumina short-read and Oxford Nanopore long-read technologies, assembling a high-quality hybrid genome of approximately 37.25 million base pairs across 33 contigs. The assembly achieved over 96% completeness with an N50 of 1.91 megabases, indicating strong contiguity. Functional annotation identified 10,800 protein-coding genes, along with 48 rRNA and 199 tRNA genes, and nearly 10,000 proteins carrying conserved PFAM domains. Phylogenetic analysis based on concatenated ITS, RPB2, and Tef1 sequences confirmed the strain’s placement within the P. australis clade with 99% statistical support, clearly distinguishing it from related species such as P. griseus and P. pannorum.

What makes this genome particularly compelling is the density and diversity of its secondary metabolite machinery. Using antiSMASH version 8.0.1 with relaxed detection parameters, the researchers predicted 44 biosynthetic gene clusters, including 20 type I polyketide synthases, 8 nonribosomal peptide synthetases, 5 terpene clusters, and 4 NRPS-PKS hybrid systems. Only 13 of these clusters showed significant similarity to previously characterized entries in the MIBiG reference database, meaning the vast majority represent potentially novel biosynthetic pathways. Notable matches include clusters associated with geodin production (70% identity, 99.39% coverage), the F9775A/F9775B/orsellinic acid pathway (74% identity), and ustilaginoidin biosynthesis (66% identity), suggesting the fungus may produce compounds with structural parallels to known bioactive molecules.

BiG-SCAPE comparative analysis grouped the 44 clusters into 41 gene cluster families, of which 39 were singletons, meaning they shared little to no similarity with other clusters in the same genome. This pattern of low genetic redundancy combined with high structural diversity is a hallmark of organisms with rich, underexplored chemical repertoires. Two multi-member families stood out: FAM_00002 comprises three NRPS-type clusters with the canonical A–T–C domain architecture, predicted to generate small cyclic dipeptides incorporating aromatic and aliphatic amino acids, closely resembling diketopiperazine synthases from Penicillium rubens. FAM_00001 encodes a highly conserved iterative type I PKS involved in synthesizing 1,3,6,8-tetrahydroxynaphthalene, a critical precursor in the DHN-melanin biosynthetic pathway, sharing 81% sequence identity with a known cluster from Glarea lozoyensis.

Having mapped the genetic blueprint, the team turned to the challenge of activating these silent pathways. In standard laboratory culture, many fungal biosynthetic gene clusters remain transcriptionally dormant, producing no detectable metabolites. To overcome this, the researchers employed two well-established elicitation strategies: sodium nitroprusside (SNP), which releases nitric oxide, and lipopolysaccharide (LPS) from Escherichia coli O55:B5, which mimics bacterial cell wall components that fungi perceive as threat signals. Nitric oxide functions as a signaling molecule in fungi, regulating developmental processes and neutralizing nitrosative stress while promoting the expression of cryptic biosynthetic genes. LPS, meanwhile, has been shown in prior studies to activate secondary metabolism in approximately 15% of tested fungal strains, including Penicillium and Aspergillus species.

The team cultivated P. australis UA-032-E in twelve different culture media using a micro-bioreactor system, then narrowed the screen to five media (M2, IMA, YES, ISP-4, and CG4) that best supported secondary metabolite production. Cultures were incubated at 15°C for seven days, mirroring the fungus’s natural psychrophilic growth conditions. Metabolites were extracted in situ with ethyl acetate and analyzed by LC–QTOF–MS/MS on an Agilent 6545 Q-TOF instrument equipped with a Zorbax C8 column. The resulting data were processed through MZmine for feature detection and then subjected to molecular networking via the GNPS Feature-Based Molecular Networking platform, complemented by in silico structure prediction using the SIRIUS, ZODIAC, CSI:FingerID, and CANOPUS computational tools.

The metabolomic analysis detected a core set of 75 mass spectral features consistently present across all four experimental treatments (LPS-negative, LPS-positive, SNP-negative, and SNP-positive). Among the annotated compounds were three diketopiperazine cyclic dipeptides: cyclo(L-Phe-L-Pro) at m/z 245.1280, cyclo(L-Val-L-Pro) at m/z 197.1281, and cyclo(L-Pro-L-Leu) at m/z 211.1437. These stable ring-structured metabolites, commonly produced by fungi, have been previously isolated from species including Penicillium chrysogenum, Aspergillus terreus, and marine-derived Pseudofusicoccum. The SIRIUS platform additionally identified maculosin [cyclo(L-Pro-L-Tyr)] at m/z 261.1230, a compound known from Alternaria alternata with documented antimicrobial properties, and methyl asterrate at m/z 385.0896, a polyketide-derived metabolite previously reported from Geomyces species with inhibitory activity against Aspergillus fumigatus.

Beyond small-molecule secondary metabolites, the analysis revealed the presence of DGTS 18:2, a betaine lipid producing a characteristic fragment ion at m/z 498.3800 corresponding to its monoacyl form. In other organisms such as the green alga Chlorella kessleri, DGTS species are known to replace phosphatidylcholine under phosphorus-deficient conditions as part of membrane remodeling strategies. The detection of this lipid in P. australis is consistent with a possible adaptive response to the nutrient-limited conditions of Antarctic sediments, although the researchers caution that a direct causal relationship in this specific context remains to be established through targeted lipidomic studies. The alkaloid harman, a β-carboline detected at m/z 183.0910, and N⁶,N⁶-dimethyladenine at m/z 164.0930, a methylated purine implicated in fungal cytokinin biosynthesis, further round out the chemical profile.

The YES medium supplemented with LPS elicitation yielded the highest metabolic diversity among all tested conditions, suggesting this combination effectively stimulates specialized metabolism in P. australis. The combined SNP-plus-LPS treatment also produced more features than either elicitor alone, consistent with previous findings in Antarctic bacteria where combined elicitation triggered dormant pathways and enhanced metabolite diversity. However, not all predicted biosynthetic gene clusters had corresponding metabolites detected in the analysis. The researchers note this could indicate a requirement for more specific activation signals, or that novel metabolites were produced but remained among the unannotated features in the metabolomic datasets. Certain clusters may also be epigenetically regulated, produce metabolites at very low concentrations, or generate unstable or non-ionizable compounds under the applied analytical conditions.

This study represents a significant step forward in the bioprospecting of polar fungi, demonstrating that the integration of genomic mining with chemical elicitation and metabolomic profiling can efficiently navigate the complex chemical space of psychrophilic organisms. With only approximately 10% of microbial biosynthetic gene clusters currently described in the literature, the findings underscore how much unexplored chemistry remains hidden in extreme environments. The annotated genome of P. australis UA-032-E, deposited in NCBI under accession JBLVTM000000000, now provides a foundational resource for future functional studies and targeted bioprospecting efforts aimed at discovering novel bioactive metabolites with potential applications in pharmaceuticals, agriculture, and industrial biotechnology.

Subject of Research: Genome mining and elicitation of silent biosynthetic pathways in the Antarctic fungus Pseudogymnoascus australis

Article Title: Exploration of inactive metabolic pathways in Antarctic Pseudogymnoascus australis through elicitation: a genomic and metabolomic approach to investigate its biotechnological potential

Article References: Leal, K., Machuca, J., Madariaga, D., Contreras, M. J., Barrientos, L., Nuñez-Montero, K., Chavarría, E., Bruna, P., & Iturrieta-González, I. (2026). Exploration of inactive metabolic pathways in Antarctic Pseudogymnoascus australis through elicitation: a genomic and metabolomic approach to investigate its biotechnological potential. IMA Fungus, 17, Article e156018. https://doi.org/10.3897/imafungus.17.156018

Image Credits: AI Generated

DOI: 10.3897/imafungus.17.156018

Keywords: Pseudogymnoascus australis, Antarctic fungi, biosynthetic gene clusters, secondary metabolites, elicitation, genomics, metabolomics, psychrophilic fungi, bioprospecting, natural products, nitric oxide, lipopolysaccharide

The idea rests on the gut-brain axis, the bidirectional communication network that links the digestive tract and the central nervous system through the vagus nerve, immune signaling, hormones, and a constant river of circulating metabolites. The gut microbiota can shape the integrity of the intestinal barrier, tune inflammatory signaling, and alter the availability of neurotransmitter precursors. In return, the brain influences the gut through the autonomic nervous system and the hypothalamic-pituitary-adrenal axis. When this dialogue goes wrong, the review suggests, the result may be amplified neuroinflammation, central sensitization, and a trigeminovascular system primed to fire. It is an appealing framework, and one that fits a long-observed clinical curiosity: people with migraine report gastrointestinal complaints at strikingly high rates.

The human evidence begins with patterns rather than mechanisms. Across pediatric cohorts, adults, and older women, researchers have reported differences in gut microbial composition between people with migraine and healthy controls. Children with migraine show shifts in taxa including Bacteroides, Parabacteroides, and Ruminococcaceae. In elderly women, migraine was associated with lower abundance of Faecalibacterium prausnitzii, a major butyrate-producing bacterium with anti-inflammatory and barrier-supporting functions. Patients with chronic migraine and medication-overuse headache show altered microbial profiles that track with circulating inflammatory markers. Even an early 2014 meta-analysis found a higher prevalence of Helicobacter pylori infection among migraine patients in some Asian populations, hinting that gastrointestinal microbes and headache have been entangled in the literature for over a decade.

But the review is refreshingly blunt about the limits of these findings. Most human microbiome studies are cross-sectional snapshots, which cannot tell us whether microbial changes precede migraine, result from migraine-related dietary restrictions and medication use, or simply reflect shared environmental factors. And here lies a crucial technical caveat that the authors hammer repeatedly: taxonomy is not chemistry. A drop in putative butyrate-producing bacteria suggests a reduced community capacity for short-chain fatty acid production, but it does not prove that actual fecal or blood levels of these molecules have fallen. Diet, age, sex, geography, sequencing platforms, and bioinformatic pipelines all inject noise. Without direct metabolomic measurements, the microbial signatures of migraine remain correlations in search of a mechanism.

The strongest mechanistic story centers on short-chain fatty acids, the acetate, propionate, and butyrate molecules that gut bacteria ferment from dietary fiber. These are not passive waste products. Acetate has been shown in non-migraine models to sustain the metabolic fitness of microglia, the brain’s resident immune cells. Propionate and butyrate bind the receptors FFAR2 and FFAR3 on epithelial and immune cells, shaping barrier function and inflammatory tone. In nitroglycerin-induced mouse models of migraine, the workhorse of the field, treatment with propionate and butyrate attenuated migraine-like responses, reduced inflammatory mediators in the trigeminal nucleus, and repaired intestinal mucosal injury. Most strikingly, in one recent experiment, antibiotic disruption of the microbiome prolonged migraine-like pain in mice, while supplementation restored intestinal acetate and propionate levels and FFAR2 signaling. In mice engineered to lack FFAR2, that protective effect vanished entirely, a rare causal demonstration inside an experimental model.

Tryptophan metabolism tells a subtler tale, because the three branches of this pathway differ in how much microbes actually own them. Gut bacteria directly convert tryptophan into indole derivatives such as indole-3-propionic acid and tryptamine, which activate the aryl hydrocarbon receptor and can modulate astrocyte activity, mucosal immunity, and barrier integrity. The kynurenine pathway, by contrast, is a host route that microbes can only nudge indirectly, yet its products are deeply relevant to headache biology: kynurenic acid dampens glutamatergic and NMDA receptor signaling, while 3-hydroxykynurenine is a redox-active compound tied to oxidative stress. In nitroglycerin models, a kynurenic acid analog attenuated hyperalgesia and altered CGRP-related signaling. Serotonin sits on the third branch, produced mainly by host enterochromaffin cells, and its centrality to migraine is beyond doubt, since triptans and lasmiditan both act on 5-HT receptors. But the authors caution that the clinical success of serotonergic drugs does not prove that microbiota-driven changes in serotonin metabolism cause migraine.

Perhaps the most provocative chapter involves nitric oxide, a molecule with a split identity. Host nitric oxide synthases are major NO producers, and the pharmacology is unambiguous: nitroglycerin, an NO donor, reliably provokes delayed migraine attacks in susceptible people, which is precisely why it became the field’s standard provocation paradigm. Independent labs have traced the downstream cascade through protein kinase C delta, CGRP antagonism, IL-17-driven neuroinflammation, and the interplay of all three NOS isoforms with peroxynitrite and soluble guanylate cyclase. Yet microbes hold a piece of this puzzle too. Oral bacteria reduce dietary nitrate to nitrite, feeding systemic NO production, and human studies show that periodontitis impairs this capacity while dietary nitrate reshapes the oral microbiome. In the American Gut Project cohort, people with migraine carried higher levels of nitrate- and nitrite-reducing oral microbes, and a 2025 case-control study linked several nitrate-reducing oral genera to headache day counts. Functional potential, the review stresses, is not metabolite flux, and no study has yet shown that microbe-derived NO reaches the trigeminovascular system in attack-relevant concentrations.

Beyond these three headline pathways, the review surveys a frontier of emerging suspects. Certain bacteria carry histidine decarboxylase and manufacture histamine, a molecule that can provoke headache in susceptible individuals and cross-talk with CGRP. Barrier markers like zonulin, which are host-derived rather than microbial, have been reported at altered levels in children with migraine alongside dysbiosis. The endocannabinoidome, the expanded family of lipid mediators involved in pain signaling, engages in bidirectional commerce with the microbiota. And the estrobolome, the collection of gut microbial beta-glucuronidases that deconjugate and reactivate estrogens, offers a plausible route by which microbial enzymes could influence menstrual and hormone-related migraine susceptibility, though direct evidence in migraine patients remains essentially absent.

What about treatment? The translational record is humbling. Randomized trials of probiotics and synbiotics have produced genuinely mixed results, with some reporting fewer attack days, reduced analgesic use, and lower inflammatory markers, and other placebo-controlled studies finding no significant benefit on primary outcomes. Meta-analyses describe a literature that is small, heterogeneous, and methodologically uneven. Fecal microbiota transplantation remains firmly experimental. Dietary interventions are attractive because food simultaneously shapes migraine triggers, microbial community structure, and the substrate supply for microbial metabolism, but observational associations between gut-friendly diets and headache severity cannot establish causation. Even the tantalizing question of whether microbiome profiles could predict response to CGRP monoclonal antibodies remains entirely open.

The verdict of the review is neither hype nor dismissal, but a disciplined call for better experiments. The microbiota-gut-brain axis offers a coherent framework in which microbial chemistry could plausibly modulate barrier integrity, neuroimmune signaling, and trigeminovascular nociception, and animal models now provide genuine causal footholds, particularly for the SCFA-FFAR2 axis. What is missing is the human bridge: longitudinal studies that pair standardized migraine phenotyping with metagenomics, direct measurement of fecal and circulating metabolites, dietary and medication data, and inflammatory biomarkers. Until such studies land, the authors conclude, microbiota-related metabolites should be treated as promising mechanistic and biomarker candidates rather than proven therapeutic targets. For a disorder that affects roughly one in seven people and resists many existing treatments, even a promising candidate is worth the follow-up. The next chapter of migraine research, it turns out, may be written partly in the language of bacterial chemistry, and partly in the fiber on our plates.

Subject of Research: The role of gut microbiota-derived metabolites in migraine pathogenesis via the gut-brain axis

Article Title: Roles of gut microbiota metabolites in the pathogenesis of migraine

Article References: Shi, Y., Cao, Y., Zhang, Z., Li, R., Zhou, G., & Xu, P. (2026). Roles of gut microbiota metabolites in the pathogenesis of migraine. Frontiers in Medicine, 13, Article 1949484. https://doi.org/10.3389/fmed.2026.1949484

Image Credits: AI Generated

DOI: 10.3389/fmed.2026.1949484

Keywords: migraine, gut microbiota, gut-brain axis, short-chain fatty acids, tryptophan metabolism, nitric oxide, CGRP, neuroinflammation, probiotics, intestinal barrier, kynurenine pathway, microbiome

The story of 6PPD-Q begins on the road. Every time a vehicle brakes, accelerates, or corners, its tires shed microscopic particles and a complex cocktail of chemicals into the surrounding environment. Among these chemicals, 6PPD-Q is generated by atmospheric oxidation of the tire antioxidant 6PPD and has attracted particular concern because of its extraordinary toxicity in sensitive aquatic species, most famously the acute mortality it induces in coho salmon. Environmental monitoring has since documented 6PPD-Q in air, dust, soil, road runoff, sediment, surface water, and even edible plants, creating multiple potential routes of human contact. Detection of the compound in human urine and cerebrospinal fluid indicates that environmental exposure translates into internal exposure, raising urgent questions about what such internal doses might be doing to human organs and tissues.

Previous research had already flagged the reproductive system as a potential target of 6PPD-Q. In ovarian granulosa cells, the compound has been reported to suppress PI3K/AKT/FOXO1 signaling and promote apoptosis. Chronic exposure in male mice disrupted endocrine balance, impaired sperm quality, increased germ-cell death, and altered sphingolipid signaling. An independent multi-omics study of non-obstructive azoospermia implicated oxidative injury, abnormal cell-cycle regulation, apoptosis, and loss of germ-cell homeostasis in 6PPD-Q-associated testicular damage. Growing evidence also links environmental pollution more broadly to male sexual dysfunction through oxidative stress, endocrine disruption, vascular endothelial injury, and impaired nitric oxide signaling, with occupational bisphenol A exposure, lead, arsenic, mixed heavy metals, and organophosphate metabolites all showing associations with erectile dysfunction. Yet whether 6PPD-Q directly impairs erectile function, and through what molecular mechanism, remained unclear until now.

To answer that question, the team first assembled the molecular landscape. They retrieved potential 6PPD-Q targets from SwissTargetPrediction, SEA, TargetNet, and the Comparative Toxicogenomics Database, and erectile dysfunction-associated genes from GeneCards. The intersection yielded 134 genes present in both collections, designated 6PPD-Q-ED-related genes. A protein-protein interaction network built in STRING and visualized in Cytoscape showed that these genes were mainly involved in responses to peptides and peptide hormones, responses to xenobiotic stimuli, and negative regulation of transport, while Gene Ontology and KEGG enrichment identified metabolism-, signaling-, and cancer-related pathways. The enrichment profile suggested that 6PPD-Q exposure might disrupt cellular homeostasis through mechanisms related to xenobiotic stress responses and altered transport regulation, providing the first hint of a biologically coherent link between the pollutant and cavernosal dysfunction.

The next step placed these candidate signals within the cellular architecture of the corpus cavernosum itself. Using single-cell RNA sequencing data from five human erectile tissue samples, the researchers resolved nineteen clusters and annotated six major cell types: fibroblasts, endothelial cells, smooth muscle cells, macrophages, Schwann cells, and T cells. Fibroblasts were the predominant population, accounting for 48.98 percent of all cells, followed by endothelial cells at 22.91 percent and smooth muscle cells at 22.49 percent. Gene set scoring with AUCell showed that the 6PPD-Q-ED-related gene set was relatively enriched in macrophages and moderately enriched in fibroblasts and endothelial cells, suggesting that 6PPD-Q exposure may disrupt the cavernosal microenvironment through inflammatory responses, extracellular matrix remodeling, and endothelial dysfunction, the very processes that govern penile vascular health.

To prioritize the leading candidates, the team integrated three rat corpus cavernosum transcriptomic datasets covering diabetes-associated, hypertension-associated, and neurogenic erectile dysfunction, batch-correcting the combined matrices with ComBat. Differential expression analysis identified 1,003 genes, seventeen of which overlapped with the 6PPD-Q-ED-related gene set. Two independent machine learning procedures, LASSO regression and support vector machine-recursive feature elimination, converged on four genes: ACO1, HIF1A, PDK4, and PTPN11. All four showed area under the receiver operating characteristic curve values greater than 0.70, with HIF1A reaching an AUC of 0.926. Critically, two-sample Mendelian randomization using cis-eQTL instruments from eQTLGen and erectile dysfunction summary statistics from FinnGen, comprising 1,154 cases and 94,024 controls, showed that genetically predicted HIF1A expression was significantly associated with erectile dysfunction risk, with an inverse-variance weighted odds ratio of 1.384 and a 95 percent confidence interval of 1.022 to 1.874. No significant associations were found for the other three genes, and sensitivity analyses including MR-Egger, weighted median, MR-PRESSO, and leave-one-out procedures supported the stability of the HIF1A result.

With HIF1A prioritized, the team turned to structural biology to ask whether 6PPD-Q could physically interact with HIF-1α, the protein it encodes. Molecular docking with AutoDock Vina predicted that 6PPD-Q could bind to a pocket in HIF-1α with a binding energy of −6.8 kcal/mol, below the −5 kcal/mol threshold conventionally interpreted as suggestive of potentially favorable binding. The docked complex also showed hydrogen-bond interactions between 6PPD-Q and nearby amino acid residues. A 100-nanosecond molecular dynamics simulation in GROMACS confirmed that the complex reached a relatively stable trajectory, with root-mean-square deviation rising during early structural accommodation and subsequently fluctuating around a plateau. Radius of gyration and solvent-accessible surface area decreased modestly before stabilizing, consistent with a more compact ensemble, and intermittent hydrogen bonds persisted throughout the trajectory. The free-energy landscape contained a dominant low-energy basin, providing a computational structural basis for the proposed interaction.

The decisive evidence came from the animal experiments. Twenty-four male Sprague-Dawley rats were allocated to four groups: control, 6PPD-Q, 6PPD-Q plus PX-478, and PX-478 alone. 6PPD-Q was suspended in corn oil and given by oral gavage at 4 mg/kg once daily for six weeks, while PX-478, a HIF-1α inhibitor, was administered orally at 10 mg/kg three times per week. After the exposure period, the researchers measured erectile hemodynamics by recording intracavernosal pressure in response to electrical stimulation of the pelvic ganglion, normalizing maximum intracavernosal pressure to mean arterial pressure as the functional endpoint. Rats exposed to 6PPD-Q showed a significantly lower ICPmax/MAP ratio than controls, indicating impaired erectile function. Co-treatment with PX-478 partially restored this ratio, providing pharmacological evidence that HIF-1α activation contributes to the erectile impairment.

The molecular changes in the corpus cavernosum told a coherent story. Immunohistochemistry showed increased HIF-1α staining in endothelial and smooth muscle cells of 6PPD-Q-exposed rats, which was reduced by PX-478 treatment. Western blot analysis revealed increased HIF-1α expression alongside decreased phosphorylated eNOS, decreased total eNOS, and a reduced p-eNOS/eNOS ratio in the 6PPD-Q group. Cavernosal nitric oxide levels, measured colorimetrically, were decreased following 6PPD-Q exposure and increased after PX-478 treatment. Notably, serum testosterone levels did not differ significantly among the four groups, suggesting that the erectile impairment in this model was attributable to local vascular dysfunction rather than altered androgen levels. The eNOS/NO pathway is the core mechanism responsible for maintaining normal erectile responses, as endothelial cell-derived nitric oxide promotes relaxation of corpus cavernosum smooth muscle and sinusoidal filling, and its disruption by 6PPD-Q points directly to endothelial injury as the site of toxic action.

The findings suggest that 6PPD-Q may contribute to erectile dysfunction by activating HIF1A-related stress responses and impairing eNOS/NO-mediated endothelial relaxation. HIF-1α is a central transcriptional regulator of cellular responses to hypoxia and stress, and excessive activation may enhance the release of inflammatory mediators and activate tissue remodeling signals, contributing to endothelial injury and smooth muscle dysfunction. Previous erectile dysfunction studies have shown that cavernosal nerve injury or hypoxic conditions are accompanied by increased HIF-1α expression, elevated fibrosis-related factors, and reduced NOS expression, while interventions that improve erectile function are often associated with HIF-1α downregulation and restoration of the NOS system. The authors acknowledge important limitations: only a single dose and exposure period were evaluated, precluding dose-response assessment; the transcriptomic datasets were derived from rat models; the Mendelian randomization analysis was constrained by available public data and tissue mismatch; downstream cGMP signaling was not evaluated; and the proposed 6PPD-Q-HIF-1α interaction lacked direct biochemical validation by surface plasmon resonance or co-immunoprecipitation. Nevertheless, this study extends the reproductive toxicological effects of 6PPD-Q to penile erectile function for the first time, adding a new dimension to concerns about traffic-derived contaminants and male sexual health, and suggesting that reducing environmental 6PPD-Q exposure could become a modifiable factor in preventing vascular erectile dysfunction.

Subject of Research: Mechanistic evidence linking the tire-derived environmental contaminant 6PPD-quinone to erectile dysfunction through dysregulation of the HIF-1α/eNOS/nitric oxide axis

Article Title: 6PPD-quinone exposure impairs erectile function through dysregulation of the HIF-1α/eNOS/NO axis: multi-omics and in vivo evidence

Article References: Meng, C., Li, Y., Zhu, X., Li, Y., Du, Y., & Xia, Z. (2026). 6PPD-quinone exposure impairs erectile function through dysregulation of the HIF-1α/eNOS/NO axis: multi-omics and in vivo evidence. Frontiers in Endocrinology, 17, Article 1912092. https://doi.org/10.3389/fendo.2026.1912092

Image Credits: AI Generated

DOI: 10.3389/fendo.2026.1912092

Keywords: 6PPD-quinone, erectile dysfunction, HIF-1α, eNOS, nitric oxide, environmental contaminant, tire rubber pollutant, corpus cavernosum, network toxicology, single-cell RNA sequencing, Mendelian randomization, molecular docking

The central idea of the review is metabolic reprogramming. Because plants are sessile organisms, they cannot flee environmental challenges; instead they must reorganise their internal chemistry in place. When stress strikes, cellular homeostasis collapses, photosynthesis falters and energy flows are diverted from growth toward defence. Melatonin, chemically N-acetyl-5-methoxytryptamine, intervenes at multiple points in this process. It shifts carbon fluxes through carbohydrate metabolism, alters amino acid biosynthesis, adjusts energy metabolism and stimulates the production of secondary metabolites such as phenolics, flavonoids and anthocyanins. The net effect is an accumulation of osmoprotectants, antioxidants and defence-related compounds that buffer the cell against damage and keep essential functions running under conditions that would otherwise be lethal.

One of the most striking aspects of the review is its emphasis on how modern analytical chemistry has transformed the field. High-throughput platforms such as liquid chromatography coupled to mass spectrometry, gas chromatography coupled to mass spectrometry and nuclear magnetic resonance spectroscopy now allow researchers to profile hundreds of stress-responsive metabolites simultaneously. These metabolomics approaches reveal, at a systems level, exactly which metabolic networks melatonin touches. When combined with transcriptomics, which measures changes in gene expression, and proteomics, which tracks shifts in protein abundance, metabolomics provides a multi-layered picture of how a single molecule can orchestrate such far-reaching changes. The authors highlight that this multi-omics integration is the key to understanding melatonin-mediated stress tolerance rather than viewing it as a collection of isolated physiological effects.

The mechanistic details are considerable. Melatonin biosynthesis in plants proceeds from tryptophan through a series of enzymatic steps, with serotonin N-acetyltransferases and related enzymes acting as key control points. Recent work cited in the review shows that manipulating these enzymes, for example by enhancing the expression of SNAT1 in kiwifruit, boosts melatonin biosynthesis, promotes growth and improves salt stress tolerance. Copy number variation in ASMT and COMT genes appears to drive differences in melatonin dosage between species, suggesting that breeders could select for naturally high-melatonin lines. This genetic dimension turns what might seem like an abstract biochemical story into a practical breeding target for climate-resilient agriculture.

Equally important is melatonin’s role as a signalling hub. The review describes extensive crosstalk with abscisic acid, auxin, cytokinin, gibberellin, ethylene, jasmonic acid, salicylic acid and strigolactones. In tomato, jasmonate and melatonin act synergistically to potentiate cold tolerance, while abscisic acid participates in melatonin-induced chilling tolerance in cucumber by regulating photosynthesis and the antioxidant system. In Arabidopsis, melatonin antagonises cytokinin responses to stimulate root growth, and cytokinin in turn modulates phytomelatonin responses under high light. Melatonin also represses auxin synthesis and polar auxin transport to regulate root meristem activity, and it can suppress ethylene biosynthesis during apple fruit ripening by inhibiting the transcription factor MdREM10. These interactions show that melatonin sits at a genuine crossroads of plant hormone networks, capable of both amplifying and damping specific hormonal signals depending on the stress context.

Reactive oxygen species and nitric oxide form another critical layer of this signalling web. Stress conditions cause bursts of reactive oxygen that damage membranes, proteins and DNA, but these molecules also serve as signals. Melatonin and its metabolites are exceptionally efficient radical scavengers, and the review documents how melatonin improves ROS metabolism and antioxidant defence responses in systems ranging from tomato seedlings exposed to copper to maize facing chromium toxicity. At the same time, melatonin engages nitric oxide in a stress-state-dependent signalling module, as recent work in Trends in Plant Science describes, allowing the plant to fine-tune its redox balance rather than simply eliminating all reactive species. This dual role, detoxifier and signal modulator, distinguishes melatonin from conventional antioxidants.

The review also covers melatonin’s contribution to defence against living attackers. In Arabidopsis, comparative metabolomic analysis has implicated sugars and glycerol in melatonin-mediated innate immunity against bacterial pathogens. Melatonin activates plant defence against root-knot nematodes through the jasmonate pathway, enhances resistance to Botrytis cinerea in tomato through a hierarchical cascade involving jasmonic acid and nitric oxide, and promotes jasmonic acid and phlorizin biosynthesis in pear against Botryosphaeria dothidea. Postharvest applications are particularly promising: melatonin treatment strengthens blueberry fruit defences by modulating jasmonic acid signalling and phenylpropanoid metabolites, and it improves chilling tolerance of peach fruit during cold storage by affecting jasmonate content and cell wall stability. There is even emerging evidence that melatonin may act as a natural antiviral compound in plant biocontrol.

Heavy metal and contaminant stress receives detailed treatment as well. Melatonin mitigates nickel toxicity in tomato seedlings by improving nutrient uptake, root architecture and antioxidant potential; it reduces heavy metal accumulation in pepper; it alleviates mercury toxicity in moss by modulating redox homeostasis and the urea cycle; and it helps rice cope with arsenic by reinforcing antioxidant defences and secondary metabolism. In alfalfa, melatonin alleviates aluminium toxicity, and in tomato, anthocyanin synthesis proves critical for melatonin-induced chromium tolerance. Combined applications are a recurring theme: melatonin with strigolactone mitigates salinity in maize through antioxidant defence, osmotic adjustment and ion homeostasis; melatonin with hydrogen sulfide modulates secondary metabolites and metal sequestration in arsenic-stressed tomato; and melatonin with salicylic acid improves drought performance in Brassica napus.

For agriculture, the practical implications are substantial. Exogenous melatonin application, whether as foliar spray, seed treatment or postharvest dip, has repeatedly improved crop performance under stress. In cotton, combined abscisic acid and melatonin application protects carbohydrate metabolism and yield under drought. In sugar beet, melatonin with jasmonic acid protects against heat stress by modulating enzymatic antioxidants and organic osmolytes. In sorghum, exogenous melatonin improves drought responses through defined physiological mechanisms. The review frames these findings within the broader push toward climate-smart agriculture, noting that melatonin-enabled omics approaches are helping researchers understand plant responses to single and combined stresses, which is precisely the kind of multifactorial challenge that field crops increasingly face as weather patterns become more erratic.

The authors conclude that integrating metabolomics with transcriptomics and proteomics offers the clearest path forward for decoding melatonin’s mechanisms and translating them into strategies for enhancing crop resilience and productivity. What was once considered merely an animal sleep hormone has turned out to be a versatile green signal, one that plants deploy to rewire their metabolism in the face of adversity. As sequencing costs fall and mass spectrometry becomes ever more sensitive, the coming years should reveal the precise regulatory circuits through which melatonin operates, opening the door to crops engineered or bred to harness this molecule’s remarkable protective chemistry. For a world where heatwaves, droughts and saline soils threaten harvests with growing frequency, melatonin’s journey from night hormone to agricultural ally may prove to be one of the more consequential stories in plant biology.

Subject of Research: Melatonin-mediated metabolic reprogramming and stress adaptation in plants

Article Title: Melatonin-mediated metabolic reprogramming and stress adaptation in plants

Article References: Hussain, B., Colgecen, H., Karahan, H., & Fariduddin, Q. (2026). Melatonin-mediated metabolic reprogramming and stress adaptation in plants. Plant Cell Reports, 45(11), Article 335. https://doi.org/10.1007/s00299-026-04016-z

Image Credits: AI Generated

DOI: 10.1007/s00299-026-04016-z

Keywords: melatonin, phytomelatonin, metabolic reprogramming, metabolomics, multi-omics, plant stress tolerance, reactive oxygen species, nitric oxide, phytohormone crosstalk, secondary metabolites, crop resilience, Plant Cell Reports

The trial enrolled 66 adults aged 60 to 75 with type 2 diabetes who had not been meeting recommended physical activity levels. Participants were randomly assigned in equal numbers to a morning exercise group, an evening exercise group, or a control group. Both exercise groups completed 12 weeks of moderate-intensity combined aerobic and resistance training delivered online three times per week, with the morning group training from 9:00 to 10:10 and the evening group from 19:00 to 20:10. The control group maintained usual lifestyle habits and attended weekly health education sessions without structured exercise. Fifty-eight participants completed the trial and were included in the final analysis.

The researchers measured four complementary indicators of vascular health: flow-mediated vasodilation, or FMD, which gauges how well the brachial artery expands in response to increased blood flow; brachial-ankle pulse wave velocity, or baPWV, the standard clinical marker of arterial stiffness; brachial blood pressure; and circulating plasma nitric oxide, the key vasodilator molecule produced by endothelial cells. Assessments were performed at baseline and after the intervention, with post-intervention testing conducted 48 to 72 hours after the final session to eliminate acute exercise effects, and always at the same time of day to control for circadian confounding.

The results showed that both exercise groups improved. FMD increased significantly from baseline in the morning group, rising from about 5.0 to 6.3 percent, and in the evening group, rising from about 5.5 to 7.2 percent, while systolic blood pressure and mean arterial pressure fell in both. Arterial stiffness also declined in both groups, but the magnitude differed strikingly: baPWV dropped by an average of 89.2 centimetres per second in the morning group and by 221.1 centimetres per second in the evening group. An exploratory analysis directly comparing the two exercise groups found a statistically significant interaction for baPWV, indicating a greater reduction with evening exercise.

Nitric oxide told a similar story. Plasma NO levels, measured indirectly through its metabolites nitrite and nitrate, increased significantly only in the evening group, from 31.75 to 46.30 micromoles per litre, whereas the morning group showed essentially no change. Post-intervention, the evening group exhibited higher FMD and NO levels and lower mean arterial pressure than the control group. The exploratory comparison of the two exercise groups for NO showed a numerically greater increase with evening exercise, although the interaction just missed statistical significance at p = 0.051.

Why might the clock matter? The answer lies in circadian physiology. In the early morning, sympathetic nervous system activity surges, accompanied by elevated cortisol, catecholamines, angiotensin II, platelet aggregates, and plasminogen activator inhibitor-1. Heart rate and blood pressure rise, serum nitric oxide falls, and brachial artery FMD is attenuated compared with other times of day. These alterations are thought to underlie the well-documented morning peak of cardiovascular events. By contrast, in the early evening most of these patterns reverse, creating a vascular environment with lower sympathetic tone and greater vasodilatory capacity, which may allow exercise-induced shear stress on endothelial cells to translate more effectively into nitric oxide synthesis and arterial adaptations.

The nitric oxide pathway is central to the trial’s mechanistic logic. Exercise increases blood flow and shear stress on the endothelium, which promotes endothelial nitric oxide synthase phosphorylation and NO production while reducing oxidative stress and proinflammatory signalling. NO, in turn, modulates vascular tone and arterial elasticity, so a larger training-induced NO response could plausibly contribute to the greater reduction in arterial stiffness observed after evening sessions. The authors note that plasma NO and FMD provide complementary biochemical and functional perspectives on vascular adaptation, and that considerable inter-individual variability in NO responses may reflect heterogeneity in vascular status within this population.

The trial has notable strengths. It is, to the researchers’ knowledge, the first randomised controlled trial to compare the long-term effects of morning versus evening combined aerobic and resistance exercise on endothelial function, arterial stiffness, and blood pressure in older adults with type 2 diabetes. Attendance was high in both exercise groups, at 95.3 percent in the morning and 97.5 percent in the evening, no exercise-related adverse events occurred, and outcome assessors and data analysts were blinded to group allocation. The consistent timing of vascular assessments before and after the intervention minimised the confounding influence of circadian variation on the outcome measures, a limitation that plagues many acute time-of-day studies.

Limitations temper the conclusions. The study did not include an afternoon exercise group, so whether midday sessions confer different vascular benefits remains unknown. Baseline use of lipid-lowering medication differed significantly among the three groups, an imbalance that likely arose by chance but may have influenced between-group vascular responses. Dietary intake was not strictly controlled, and sleep, physical activity outside prescribed sessions, and medication timing were not monitored during the intervention. Shear rate was not assessed during FMD measurement because the ultrasound system could not record brachial artery diameter and blood velocity simultaneously after cuff deflation. The authors call for future studies to monitor these variables and to include longer follow-up to determine whether the observed adaptations are sustained.

Even so, the findings carry practical weight. FMD is an independent predictor of future cardiovascular events, and evidence suggests that a 1 percent increase in FMD is associated with roughly a 13 percent reduction in cardiovascular risk. In this trial, FMD increased by an average of 1.45 percent across the two exercise groups, indicating potentially clinically relevant improvements in endothelial function. Current exercise guidelines for people with type 2 diabetes specify the type, intensity, frequency, duration, and progression of exercise but offer no clear guidance on timing within the day. This trial suggests that evening exercise may provide additional vascular benefits, particularly for arterial stiffness, and that exercise timing deserves consideration in exercise planning for this population. The study was registered in the Chinese Clinical Trial Registry under identifier ChiCTR2300073737.

Subject of Research: Exercise timing and vascular health in type 2 diabetes

Article Title: Effects of morning versus evening exercise on vascular function in older adults with type 2 diabetes: a randomised controlled trial

Article References: Ma, Z., Lin, Y., Zhu, Z., & Cao, Z.-B. (2026). Effects of morning versus evening exercise on vascular function in older adults with type 2 diabetes: a randomised controlled trial. Frontiers in Endocrinology, 17, Article 1976870. https://doi.org/10.3389/fendo.2026.1976870

Image Credits: AI Generated

DOI: 10.3389/fendo.2026.1976870

Keywords: type 2 diabetes, exercise timing, vascular function, arterial stiffness, nitric oxide, randomised controlled trial, circadian rhythm, cardiovascular risk, Effects, morning, versus, evening

Skeletal muscle dysfunction is a hallmark consequence of diabetes mellitus. Patients experience reduced strength, impaired perfusion, lipid accumulation within muscle fibers and chronic inflammation, all of which erode mobility and quality of life. The Chilean researchers hypothesized that a specific class of membrane proteins — large-pore channels, including connexin-formed hemichannels and P2X7 receptors — drives much of this damage. When these channels open, they dramatically increase membrane permeability, allowing ions and signaling molecules to flood across the sarcolemma and triggering inflammatory cascades. Boldine, known for its antioxidant and anti-inflammatory properties, is a recognized inhibitor of these channels, making it an ideal candidate to test.

To model diabetes, the team injected male C57BL/6J mice with streptozotocin at 40 mg/kg/day for five days, a standard protocol that destroys insulin-producing beta cells and produces sustained hyperglycemia. Diabetic animals then received boldine for four weeks. The results were striking. Untreated diabetic mice showed markedly reduced muscle strength and depolarization of the resting membrane potential of myofibers, meaning the electrical charge across the muscle cell membrane shifted toward a less negative value, impairing excitability. Basal gastrocnemius perfusion dropped by approximately 20 percent, reflecting compromised vascular supply to the muscle. Boldine treatment prevented all three deficits, preserving strength, membrane potential and blood flow at near-normal levels.

Metabolic disruption was equally dramatic. Lipid accumulation in the muscle of diabetic mice reached 52.4 ± 3.6 percent of the measured area, compared with just 3.1 ± 1.3 percent in healthy controls. This ectopic fat deposition, a well-known feature of diabetic muscle, was reduced to 15.2 ± 4.1 percent with boldine treatment, a statistically significant rescue (p < 0.05). The alkaloid also tamed the inflammatory arm of the disease: mRNA levels of NLRP3, a core component of the inflammasome, surged 17.7-fold in diabetic muscle, and boldine cut this increase roughly in half. NLRP3 inflammasome activation is a key driver of interleukin-1β maturation and sterile inflammation, so its suppression represents a meaningful anti-inflammatory effect at the tissue level.

To dissect the cellular mechanism, the researchers turned to human myoblasts of the AB1167 line, exposed to low or high glucose with or without boldine. Under high glucose, the cells showed increased ethidium uptake — a standard readout of large-pore channel opening — along with elevated intracellular calcium, increased nitric oxide production, upregulated inflammasome-related gene expression and enhanced nuclear localization of PPARγ, a nuclear receptor that promotes lipid storage. Boldine attenuated every one of these changes, suggesting a coherent causal chain linking channel opening to metabolic and inflammatory damage.

The proposed mechanism, illustrated in the study’s graphical abstract, works as follows. In myoblasts, high glucose increases large-pore channel activity, raising cytoplasmic calcium concentration and nitric oxide generation. These signals feed back positively, further increasing the activity of connexin-formed hemichannels and P2X7 receptors, elevating levels of inflammasome components and promoting lipid accumulation. In streptozotocin-diabetic mice, de novo expression of large-pore channels in skeletal muscle contributes to reducing the resting membrane potential of myofibers, reducing muscle blood perfusion, causing weakness, activating PPARγ and driving intramuscular fat deposition. Boldine interrupts this vicious cycle at its entry point by inhibiting channel activity.

Calcium is the linchpin of this cascade. Sustained calcium influx through open large-pore channels activates calcium-dependent enzymes, including nitric oxide synthases, whose product nitric oxide then potentiates channel opening in a self-amplifying loop. The resulting inflammatory signaling upregulates NLRP3 and related genes, while PPARγ activation diverts metabolic programs toward lipid storage. By clamping the channel gate shut, boldine simultaneously protects membrane excitability, vascular reactivity and metabolic homeostasis — three domains of muscle biology that diabetes attacks in parallel.

The vascular findings deserve particular attention. A 20 percent reduction in basal muscle perfusion implies that large-pore channels are active not only in myofibers but also in the endothelial and mural cells that regulate vessel tone. Boldine’s ability to restore perfusion suggests the compound preserves vascular reactivity in the diabetic muscle microcirculation, ensuring that oxygen and nutrients continue to reach working fibers. This integrated muscle-vascular protection distinguishes the approach from strategies that target only metabolic or only inflammatory pathways.

The work, conducted across Chilean institutions including Pontificia Universidad Católica de Chile, Universidad Andrés Bello, Universidad del Bío-Bío, Universidad de Talca, Universidad Autónoma de Chile and Universidad de Valparaíso, was funded by ANID grants and published open access under a Creative Commons licence. Animal procedures were approved by the Bioethics Committee of the Universidad de Valparaíso. The authors emphasize that their effects involve inhibition of large-pore channels, reducing membrane permeability and calcium-dependent inflammatory signaling, and they highlight these channels as therapeutic targets for diabetes-induced muscle dysfunction.

Translational caveats remain. The study used a streptozotocin model of type 1-like diabetes in male mice, and human diabetic myopathy — most often associated with type 2 diabetes — involves additional metabolic complexity. Dosing, pharmacokinetics and long-term safety of boldine in humans are unestablished. Nevertheless, the convergence of in vivo and in vitro evidence, the magnitude of the lipid and inflammasome rescue, and the identification of a mechanistically coherent, druggable target make this one of the more compelling recent entries in the search for therapies against diabetic muscle disease. If future studies confirm these effects in models of type 2 diabetes and ultimately in patients, a compound derived from a traditional Chilean medicinal tree could become a genuine candidate for preserving muscle function in millions of people living with diabetes.

Subject of Research: Study showing boldine prevents diabetes-induced skeletal muscle dysfunction by inhibiting large-pore channels

Article Title: Boldine attenuates diabetes-induced skeletal muscle dysfunction

Article References: Vásquez, W., Lira, A., Troncoso, F., Sandoval, H., Cea, L. A., Figueroa, X. F., Escudero, C., & Sáez, J. C. (2026). Boldine attenuates diabetes-induced skeletal muscle dysfunction. Cellular and Molecular Life Sciences. https://doi.org/10.1007/s00018-026-06404-w

Image Credits: AI Generated

DOI: 10.1007/s00018-026-06404-w

Keywords: boldine, skeletal muscle, diabetes, large-pore channels, connexin hemichannels, P2X7 receptor, NLRP3 inflammasome, PPARγ, lipid accumulation, nitric oxide, calcium signaling, Peumus boldus

The central finding is striking: the parts of the parsnip that most people throw away are precisely the parts richest in bioactive compounds. Among the edible tissues, the skin of the ‘Warrior’ cultivar recorded the highest flavonoid content, at 15.27 milligrams of catechin equivalents per 100 grams of fresh weight. Among the inedible tissues, ‘Warrior’ leaves topped the chart for phenolic content, reaching 525.17 milligrams of gallic acid equivalents per 100 grams of fresh weight. These numbers matter because phenolic compounds and flavonoids are among the most studied classes of plant secondary metabolites, valued for their capacity to neutralize reactive oxygen species and modulate inflammatory signaling pathways in human cells.

Antioxidant activity in the study was assessed using two widely accepted colorimetric assays, DPPH and ABTS, each of which measures the ability of plant extracts to scavenge synthetic free radicals in solution. The DPPH assay relies on the reduction of a stable nitrogen-centered radical, which changes color from deep purple to pale yellow as antioxidants donate electrons or hydrogen atoms. The ABTS assay works on a similar principle, generating a green-blue radical cation that is decolorized by antioxidant molecules. When the researchers ran these assays on extracts from different parsnip tissues, the results tracked closely with the distribution of phenolic compounds, reinforcing a long-standing observation in food chemistry that total phenolic content is often a reliable predictor of radical-scavenging capacity.

Cultivar differences emerged as a consistent theme throughout the data. The ‘Warrior’ cultivar generally exhibited higher total flavonoid content, higher total phenolic content, and stronger antioxidant activities than ‘Albion’ across the tissues examined. This kind of genotype-dependent variation is well documented in other vegetables; comparable studies of onions, blueberries, and aronia berries have all shown that polyphenol profiles can differ substantially between cultivars grown under identical conditions. For breeders and food scientists, the implication is that cultivar selection is not merely a matter of yield, texture, or flavor, but also a lever for enhancing the nutritional and functional value of a crop.

The anti-inflammatory experiments took the investigation a step further, moving from chemical assays into living cells. The team used RAW 264.7 macrophages, a mouse cell line that has become a standard model for studying inflammatory responses in vitro. These immune cells were stimulated with lipopolysaccharide, or LPS, a molecule derived from the outer membrane of Gram-negative bacteria that triggers a potent inflammatory response, including the production of nitric oxide and a burst of intracellular reactive oxygen species. Nitric oxide, while serving important signaling functions at low concentrations, contributes to tissue damage and inflammatory pathology when overproduced by activated macrophages, making its inhibition a key benchmark for anti-inflammatory activity.

Against this backdrop, ‘Warrior’ parsnip skin extract delivered the study’s most compelling result. The skin extract showed the strongest anti-inflammatory activity of all the tissues tested, suppressing both nitric oxide production and the generation of intracellular reactive oxygen species in the LPS-stimulated macrophages. This dual action is significant because oxidative stress and inflammation are tightly intertwined processes; reactive oxygen species can activate inflammatory signaling cascades, and inflammatory enzymes in turn generate more free radicals. Compounds that interrupt this feedback loop at both points are of particular interest to researchers exploring dietary approaches to chronic inflammatory conditions, which underlie diseases ranging from atherosclerosis to arthritis.

The concentration of bioactive compounds in outer tissues is not unique to parsnip, but the study adds a valuable quantitative dimension to a pattern seen across the plant kingdom. In apples, grapes, and onions, the peel routinely outperforms the flesh in phenolic content, a consequence of the plant’s own defensive strategy. The outer layers of a root or fruit are the first line of contact with the environment, exposed to ultraviolet radiation, pathogens, and physical damage, and plants respond by accumulating protective secondary metabolites precisely where they are needed most. Leaves, similarly, face intense oxidative pressure from photosynthesis and sunlight, which helps explain why ‘Warrior’ parsnip leaves, normally discarded as agricultural waste, turned out to be the single richest source of phenolics in the entire analysis.

That last point carries real economic and environmental weight. Parsnip is grown primarily for its taproot, and the foliage is typically left in the field or composted after harvest. If the leaves contain more than 500 milligrams of phenolic compounds per 100 grams of fresh weight, they represent a largely untapped reservoir of functional ingredients that could be recovered for use in nutraceuticals, functional foods, or natural preservatives. The same logic applies to peeling waste from food processing operations, where parsnip skins are removed on an industrial scale. Converting these by-products into value-added extracts would align with a broader movement in food science toward circular economy approaches, in which agricultural residues are reimagined as raw materials rather than refuse.

The study also situates parsnip within a growing body of literature on root vegetables as sources of health-promoting compounds. Previous work by the same research group had already characterized the physicochemical properties, organic acids, and sugar profiles of parsnip cultivars harvested in Korea, and other teams have documented the antioxidant activity of essential oils from wild parsnip subspecies. Historical texts and traditional medicine systems have long attributed therapeutic properties to Pastinaca sativa, and modern pharmacological reviews have catalogued its chemical constituents. What the new research adds is a systematic, tissue-by-tissue, cultivar-by-cultivar comparison that connects specific chemical measurements to measurable biological effects in cell-based models, bridging the gap between compositional analysis and functional validation.

For consumers, the practical takeaways are straightforward, if somewhat counterintuitive. Keeping the skin on parsnips when cooking preserves the tissue shown to have the highest flavonoid content and the strongest anti-inflammatory activity in laboratory tests, and choosing cultivars such as ‘Warrior’ may offer a modest nutritional edge over alternatives. The researchers caution, as all in vitro studies must, that cellular assays do not directly translate into clinical outcomes; compounds must survive digestion, reach target tissues, and act in the complex environment of the human body before any health benefit can be claimed. Nevertheless, the work provides a clear roadmap for future research, from identifying the individual polyphenols responsible for the observed effects to testing bioavailability and efficacy in more advanced models. In the meantime, it offers a fresh reason to reconsider one of the supermarket’s most overlooked vegetables, and a reminder that in the chemistry of plants, the parts we discard often hold the greatest treasures.

Subject of Research: Antioxidant and anti-inflammatory bioactive compound distribution in parsnip cultivars and plant parts

Article Title: Distribution of antioxidant compounds, antioxidant activities, and anti-inflammatory properties in different cultivars and plant parts of parsnip (Pastinaca sativa)

Article References: Shim, H., Jung, S. K., & Shin, Y. (2026). Distribution of antioxidant compounds, antioxidant activities, and anti-inflammatory properties in different cultivars and plant parts of parsnip (Pastinaca sativa). Food Science and Biotechnology. https://doi.org/10.1007/s10068-026-02344-0

Image Credits: AI Generated

DOI: 10.1007/s10068-026-02344-0

Keywords: parsnip, Pastinaca sativa, antioxidant, anti-inflammatory, polyphenols, flavonoids, phenolic compounds, cultivar, RAW 264.7 macrophages, nitric oxide, DPPH assay, food by-products

Blastocystis is an anaerobic single-celled protozoan that colonizes the intestines of an enormous share of the global population, and it has been increasingly linked to gastrointestinal disorders, including irritable bowel syndrome and other functional complaints. Because the organism thrives without oxygen, its survival depends heavily on glycolysis, the ancient metabolic pathway that breaks down glucose without the need for mitochondrial respiration. GAPDH sits at a critical junction in that pathway, catalyzing the conversion of glyceraldehyde-3-phosphate into 1,3-bisphosphoglycerate, a step that conserves energy in a form the cell can spend. In an anaerobic parasite, interfering with this enzyme strikes at the heart of its energy economy. At the same time, Blastocystis lives in a gut microenvironment saturated with nitric oxide, produced both by host immune defenses and by neighboring bacteria, and the interplay between the parasite’s metabolism and this nitrosative pressure has remained poorly understood.

The motivation for the study is a practical one: metronidazole, the drug most commonly deployed against Blastocystis and other anaerobic pathogens, is losing ground. Resistance to the nitroimidazole class has been documented with growing frequency, and clinicians have few well-validated alternatives. That therapeutic bottleneck has pushed researchers toward two strategies reflected in this experiment—repurposing existing drugs such as metformin, whose antiparasitic potential has attracted attention in recent years, and screening plant-derived compounds, in this case an extract of Allium cepa, the common onion, whose sulfur-containing constituents have long been suspected of antimicrobial activity. The Egyptian team, led by Gehad A. Basuony and Magda S. A. Abdeltawab of the Department of Medical Parasitology at Cairo University’s Faculty of Medicine, together with colleagues from the Egyptian Drug Authority, Umm Al Qura University in Saudi Arabia, and Cairo University’s plant physiology department, set out to test all three agents side by side under identical conditions.

The experimental design was straightforward but rigorous. The researchers obtained twenty-one stool samples that had tested positive for Blastocystis and cultured the parasites in Jones’ medium under xenic conditions, meaning the cultures retained the accompanying bacteria that normally support the parasite’s growth in the gut. That choice matters: many laboratory studies use axenic cultures stripped of other microbes, which can mask the ecological interactions that determine whether a drug works in a real biological setting. Each culture was then treated with graded concentrations of metronidazole, metformin, or onion extract. Parasite numbers were assessed by direct microscopic counting, while the biochemical readouts—GAPDH activity and nitric oxide levels—were quantified using gas chromatography–mass spectrometry, a sensitive analytical technique that allowed the team to measure metabolic changes within the complex culture medium rather than in isolated enzyme preparations.

The results were unambiguous at the population level. All three agents significantly reduced Blastocystis counts compared with untreated controls, with metronidazole demonstrating the strongest antiparasitic effect overall. More intriguing were the metabolic findings. Nitric oxide levels and GAPDH activity both dropped significantly across every treatment group, with the reductions in each case reaching a high level of statistical significance. The two readouts, however, did not respond identically to the different drugs. Metformin, at a concentration of 1000 micrograms per milliliter, proved the most powerful inhibitor of GAPDH activity, suggesting that the diabetes drug interferes directly or indirectly with the parasite’s glycolytic machinery. Metronidazole, at 500 micrograms per milliliter, produced the lowest nitric oxide levels in the cultures, indicating a distinct effect on the nitrosative chemistry of the microbial milieu.

The correlation analysis added a layer of biochemical nuance that the authors consider central to the study’s significance. In cultures treated with metronidazole, nitric oxide and GAPDH activity showed a significant negative association—as one fell, the other rose—while in metformin-treated cultures the two variables moved together in a positive correlation. These divergent patterns suggest that the two drugs perturb the parasite’s biology through different mechanisms. Metronidazole, which requires chemical reduction inside anaerobic organisms to become active, appears to reshape the relationship between nitrosative stress and glycolytic output, whereas metformin’s effect seems more tightly coupled to the energy pathway itself. Onion extract, meanwhile, achieved significant suppression of both parasite counts and the metabolic markers, lending quantitative support to the idea that plant-derived compounds can act on the same biochemical axes as conventional pharmaceuticals.

Placing these findings in context requires appreciating how unusual the gut environment is as a therapeutic battleground. Blastocystis does not live in isolation; it coexists with a dense bacterial community, and the nitric oxide measured in the cultures comes from that entire ecosystem, not solely from the parasite. A drug that lowers nitric oxide levels, as metronidazole did most dramatically, may be altering the chemistry of the whole microbial milieu, potentially with consequences for both the parasite and its bacterial partners. Conversely, an agent that cripples GAPDH, as metformin did most strongly, attacks a metabolic vulnerability that is shared, in some form, by many anaerobic organisms. The authors argue that targeting GAPDH and nitric oxide–related mechanisms represents a promising therapeutic direction precisely because these pathways sit at the intersection of parasite metabolism and host–microbiota interaction.

The repurposing angle deserves particular emphasis. Metformin is among the most prescribed medications on earth, with a decades-long safety record in millions of patients with type 2 diabetes, and its anti-inflammatory and metabolic effects have been studied in contexts far removed from blood sugar control. If its capacity to inhibit GAPDH-dependent glycolysis in anaerobic parasites can be translated into a clinically useful antiparasitic regimen, the path from bench to bedside could be considerably shorter than for a novel compound. The onion extract result, for its part, speaks to a growing interest in food-derived antimicrobials. Allium species are rich in organosulfur compounds and flavonoids, and while the present study did not isolate a single active constituent, the demonstration that a crude extract can measurably suppress both parasite proliferation and specific metabolic markers provides a quantitative foundation for further fractionation and characterization work.

Important caveats temper the enthusiasm. The experiments were conducted in vitro, in culture tubes, at concentrations—up to 1000 micrograms per milliliter—that may or may not be achievable in the human intestine through oral dosing. Xenic cultures, while more realistic than axenic ones, still cannot reproduce the full complexity of a living gut, with its immune system, mucus layer, variable oxygen gradients, and diverse microbiota. The study also measured short-term effects on parasite counts and biochemical markers rather than long-term eradication or clinical outcomes. The authors are appropriately careful in their conclusions, framing the work as evidence that these agents exhibit significant anti-Blastocystis activity accompanied by modulation of key metabolic and nitrosative pathways, rather than as proof of therapeutic efficacy in patients.

Even with those limitations, the study contributes something the field has lacked: a side-by-side biochemical comparison of a standard drug, a repurposed medication, and a plant extract, all measured against the same molecular yardsticks in the same culture system. It suggests that the fight against drug-resistant Blastocystis may not require entirely new chemical entities, but rather a smarter exploitation of the metabolic seams that anaerobic parasites cannot easily patch. As metronidazole resistance continues to spread, and as interest in microbiome-aware therapeutics accelerates, the humble GAPDH enzyme and the nitric oxide swirling through the gut may prove to be the targets on which the next generation of antiparasitic strategies is built. The Cairo team’s work, conducted with anonymous leftover diagnostic specimens under institutional ethical approval, shows how a relatively simple culture experiment, paired with modern analytical chemistry, can illuminate those seams with unusual clarity.

Subject of Research: In vitro effects of metronidazole, metformin, and onion extract on GAPDH activity and nitric oxide production in Blastocystis cultures

Article Title: Evaluating the effects of metronidazole, metformin, and onion extract on GAPDH activity and nitric oxide production in Blastocystis sp. cultures: a biochemical and therapeutic approach

Article References: Basuony, G. A., Hamed, A. M. R., Mahfoz, A. M., Mahmoud, A. W. M., Ramadan, H., & Abdeltawab, M. S. A. (2026). Evaluating the effects of metronidazole, metformin, and onion extract on GAPDH activity and nitric oxide production in Blastocystis sp. cultures: a biochemical and therapeutic approach. BMC Complementary Medicine and Therapies, 26(1), Article 295. https://doi.org/10.1186/s12906-026-05605-x

Image Credits: AI Generated

DOI: 10.1186/s12906-026-05605-x

Keywords: Blastocystis, metronidazole, metformin, onion extract, GAPDH, nitric oxide, glycolysis, drug resistance, intestinal parasite, in vitro culture, GC-MS, gut microbiota

The endothelium, the delicate single layer of cells lining the interior of every blood vessel, is far more than passive plumbing. It actively regulates vascular tone by releasing vasodilating and vasoconstricting compounds, controls oxidative processes, modulates cell adhesion, and influences arterial stiffness. When the endothelium falters, vessels stiffen, their dilatory responses weaken, and the earliest silent stages of cardiovascular disease take hold. Flow-mediated dilation of the brachial artery, measured with high-resolution ultrasound after a temporary ischemic stimulus, captures this dysfunction early and tracks closely with coronary artery function, which is why the research team led by Simon Iskra and Armin Paravlić of the University of Ljubljana centered their analysis on it.

To assemble the evidence, the researchers searched six major databases from inception through multiple updated searches in 2023, 2025 and 2026, screening nearly 1,900 records against strict PICOS criteria. Only randomized controlled trials in adults that reported brachial artery flow-mediated dilation alongside at least one secondary outcome made the cut. The final pool spanned healthy volunteers and patients with cardiovascular disease, metabolic syndrome, diabetes, kidney disease, cancer, and even mental and neurological disorders. Participants ranged from about 20 to 78 years old, 43 percent were women, and interventions lasted from four to 26 weeks, with roughly two-thirds of study arms using continuous training and one-third using interval formats.

The headline result survived every robustness check the team threw at it. A random-effects model yielded a mean difference of 2.35 percent in flow-mediated dilation, a fixed-effects model produced 1.90 percent, and a trim-and-fill adjustment for publication bias still left a significant adjusted estimate of 1.74 percent. Sensitivity analysis removing the single largest study did not change the conclusion. Heterogeneity was high, as expected when pooling trials across such diverse populations and protocols, but the direction and clinical relevance of the effect were consistent. By the classification scheme the authors applied, the pooled gain qualifies as small but clinically meaningful, sitting above the 1 percent threshold considered the minimal clinically important difference.

The most novel contribution, however, lies in the meta-regression, which asked which physiological changes actually drive the vascular improvements. Four clusters emerged as significant moderators. Greater reductions in body weight and body fat were associated with larger gains in endothelial function, with regression coefficients of -4.021 and -1.721 respectively. Drops in systolic and diastolic blood pressure also tracked with better flow-mediated dilation, at coefficients of -0.525 and -0.484. On the positive side, improvements in maximal oxygen uptake, the classic index of cardiorespiratory fitness, showed a strong association with endothelial gains at a coefficient of 0.656, and changes in insulin levels moderated the effect as well. Notably, blood lipids, fasting glucose, HbA1c, and insulin resistance measured by HOMA-IR did not significantly moderate the vascular response, suggesting lipid and glucose improvements run on partly independent tracks.

Beneath these statistical associations lies a coherent mechanistic story. During aerobic exercise, blood flow to working muscles surges, subjecting the vessel walls to elevated shear stress. This mechanical signal activates endothelial nitric oxide synthase, boosting production of nitric oxide, the principal vasodilator of healthy arteries. The meta-analysis confirmed this biochemistry in blood: nitric oxide and its metabolites rose substantially, with standardized mean differences of 1.23 for both, while endothelin-1, the vessels’ chief constricting agent, fell moderately. Enhanced nitric oxide bioavailability combined with reduced endothelin-1 shifts vascular tone toward dilation, plausibly explaining much of the measured improvement in flow-mediated dilation, even though the biomarkers themselves did not reach statistical significance as moderators, likely because relatively few studies measured them.

The hemodynamic findings add another layer. Aerobic training produced small but significant reductions in systolic pressure, diastolic pressure, and mean arterial pressure, and the meta-regression tied these blood pressure drops directly to endothelial gains. The authors interpret this as evidence of a feedback loop: healthier endothelia dilate more readily, lowering vascular resistance and pressure, which in turn protects the vessel wall. Arterial stiffness told a subtler tale. Peripheral stiffness, measured at the brachial-ankle segment, decreased significantly, but central stiffness, measured at the carotid-femoral segment, and the augmentation index did not change. The researchers attribute this to the time course of vascular remodeling, since peripheral arteries adapt faster than the stiffer central vessels, and to the small number of trials that measured central outcomes alongside flow-mediated dilation.

Cardiorespiratory fitness emerged as perhaps the most intriguing partner in the adaptation. Maximal oxygen uptake rose moderately across trials, and the stronger the fitness gains, the stronger the endothelial improvements. The relationship is likely bidirectional. Repeated shear stress from training enhances nitric oxide signaling and vascular homeostasis, while preserved endothelial function optimizes skeletal muscle perfusion and oxygen delivery, supporting further fitness gains. The authors caution that because the meta-regression works at the study level, causality cannot be established; the pattern reflects interconnected adaptations rather than a simple cause-and-effect chain. Still, the convergence of fitness, body composition, blood pressure, and insulin changes as moderators paints exercise as a multi-front intervention rather than a single-target drug.

Body composition results carried their own surprise. Even though the analysis excluded trials that combined exercise with dieting, aerobic training alone significantly reduced body weight, body mass index, and body fat percentage, with no change in skeletal muscle mass. The regression confirmed that fat loss amplified the vascular benefit, and the authors suggest that shrinking adipose tissue dampens chronic low-grade inflammation, a process tightly linked to endothelial health. Inflammation markers moved in the expected direction, with a small significant fall in C-reactive protein and a moderate, near-significant decrease in TNF-alpha. Lipid changes were real but modest, with small rises in HDL cholesterol and small falls in LDL and triglycerides, while glucose metabolism improved through fasting glucose, HbA1c, and insulin resistance, even without significant changes in insulin itself.

The authors are candid about limitations. The restriction to trials reporting brachial artery flow-mediated dilation means conclusions may not generalize to other endothelial assessments. Flow-mediated dilation tends to normalize over longer interventions as structural remodeling proceeds, so pooled changes may understate peak functional adaptation. Pooling healthy and clinical populations yields general tendencies rather than disease-specific prescriptions, and the use of standardized mean differences for secondary outcomes, while necessary given heterogeneous measurement methods, obscures absolute changes. Publication bias was detected for the primary outcome and several secondary markers, though corrections preserved the core finding. Even so, the message is hard to dismiss: aerobic exercise acts simultaneously on fat mass, fitness, blood pressure, insulin regulation, nitric oxide signaling, and inflammation, and these converging pathways collectively rebuild endothelial function. For clinicians and the public alike, the analysis reinforces aerobic training as one of the most powerful non-pharmacological tools available for protecting the vascular system, with benefits that compound across nearly every measurable dimension of cardiovascular health.

Subject of Research: Effects of aerobic exercise on endothelial function and cardiovascular disease markers

Article Title: Impact of Aerobic Exercise on Endothelial Function and Other Cardiovascular Disease Markers in Health and Disease: A Systematic Review and Meta-analysis with Meta-regression of Randomized Controlled Trials

Article References: Iskra, S., Turnšek, G., Drole, K., & Paravlić, A. (2026). Impact of Aerobic Exercise on Endothelial Function and Other Cardiovascular Disease Markers in Health and Disease: A Systematic Review and Meta-analysis with Meta-regression of Randomized Controlled Trials. Sports Medicine – Open, 12(1), Article 150. https://doi.org/10.1186/s40798-026-01110-0

Image Credits: AI Generated

DOI: 10.1186/s40798-026-01110-0

Keywords: aerobic exercise, endothelial function, flow-mediated dilation, cardiovascular disease, meta-analysis, nitric oxide, blood pressure, cardiorespiratory fitness, body composition, insulin, arterial stiffness, vascular health