Considerations for cultivated crustacean meat: potential cell sources, potential differentiation and immortalization strategies, and lessons from crustacean and other animal models.

Cultivated crustacean meat (CCM) is a means to create highly valued shrimp, lobster, and crab products directly from stem cells, thus removing the need to farm or fish live animals. Conventional crustacean enterprises face increasing pressures in managing overfishing, pollution, and the warming climate, so CCM may provide a way to ensure sufficient supply as global demand for these products grows. To support the development of CCM, this review briefly details crustacean cell culture work to date, before addressing what is presently known about crustacean muscle development, particularly the molecular mechanisms involved, and how this might relate to recent work on cultivated meat production in vertebrate species. Recognizing the current lack of cell lines available to establish CCM cultures, we also consider primary stem cell sources that can be obtained non-lethally including tissues from limbs which are readily released and regrown, and putative stem cells in circulating hemolymph. Molecular approaches to inducing myogenic differentiation and immortalization of putative stem cells are also reviewed. Finally, we assess the current status of tools available to CCM researchers, particularly antibodies, and propose avenues to address existing shortfalls in order to see the field progress.

A novel role for Teneurin C-terminal Associated Peptide (TCAP) in the regulation of cardiac activity in the Sydney rock oyster, Saccostrea glomerata.

Teneurin C-terminal associated peptide (TCAP) is an ancient bioactive peptide that is highly conserved in metazoans. TCAP administration reduces cellular and behavioural stress in vertebrate and urochordate models, yet despite numerous studies in higher animals, there is limited knowledge of its role in invertebrates. In particular, there are no studies on TCAP's effects on the heart of any metazoan, which is a critical organ in the stress response. We used the Sydney rock oyster (SRO) as an invertebrate model to investigate a potential role for sroTCAP in regulating cardiac activity, including during stress. sroTCAP is localized to the neural innervation network of the SRO heart, and suggested binding with various heart proteins related to metabolism and stress, including SOD, GAPDH and metabotropic glutamate receptor. Intramuscular injection of sroTCAP (10 pmol) significantly altered the expression of heart genes that are known to regulate remodelling processes under different conditions, and modulated several gene families responsible for stress mitigation. sroTCAP (1 and 10 pmol) was shown to cause transient bradycardia (heart rate was reduced by up to 63% and for up to 40 min post-administration), indicative of an unstressed state. In summary, this study has established a role for a TCAP in the regulation of cardiac activity through modulation of physiological and molecular components associated with energy conservation, stress and adaptation. This represents a novel function for TCAP and may have implications for higher-order metazoans.

Secretion of IL-6 by fibroblasts exposed to Australian honeys involves lipopolysaccharide and is independent of floral source.

Honey stimulates cellular secretion of cytokines, which has been attributed to activation of lipopolysaccharide (LPS)-dependent and LPS-independent pathways. The objective of this study was to identify whether LPS is present in Australian honey samples at levels that can stimulate interleukin-6 (IL-6) secretion by fibroblasts and whether it can transduce cell signalling by activating toll-like receptor 4 (TLR4). IL-6 was measured in culture media of fibroblasts exposed to honey for 24 h. LPS was detected in a 0.125 mg/mL solution of grey ironbark honey (0.61 ± 0.05 ng/g honey). TLR4 signalling was observed in RAW264.7 macrophages that were exposed to honey and this was prevented by preincubating the honey with the LPS-neutralising agent, polymyxin B. Australian Eucalyptus, Leptospermum and Cyathode honeys stimulated IL-6 secretion in cultured human dermal fibroblasts. To examine whether the response was dependent on floral source, fibroblasts were exposed to four different samples of grey ironbark honey obtained from Queensland and New South Wales, Australia. The magnitude of the cytokine response to these honeys was highly varied. We conclude that Australian honeys contain endotoxin at levels that can stimulate IL-6 secretion by fibroblasts and that signalling in macrophages involves TLR4 activation. The IL-6 secretory response was independent of floral source.

Fingolimod does not prevent syndecan-4 shedding from the endothelial glycocalyx in a cultured human umbilical vein endothelial cell model of vascular injury.

BACKGROUND: Shedding of the endothelial glycocalyx (EG) is associated with poor outcomes in a range of conditions including sepsis. Fresh frozen plasma (FFP) restores the damaged EG to baseline thickness, however the mechanism for this effect is unknown, and some components of FFP have adverse effects unrelated to the EG. There is some limited evidence that sphingosine-1-phosphate (S1P) within FFP restores the EG by activating the endothelial cell S1P receptor 1 (S1PR1). However, there are disadvantages to using S1P clinically as an EG restorative therapy. A potential alternative is the S1PR agonist fingolimod (FTY720). The aim of this study was to assess whether FTY720 prevents EG shedding in injured cultured human umbilical vein endothelial cells. METHODS: Shedding of the EG was induced in cultured human umbilical vein endothelial cells (HUVECs) by exposure to adrenaline, TNF-α and H2O2. The cells were then assigned to one of six conditions for 4 h: uninjured and untreated, injured and untreated, injured and treated with FTY720 with and without the S1PR1 inhibitor W146, and injured and treated with 25% FFP with and without W146. Syndecan-4, a component of the EG, was measured in cell supernatants, and syndecan-4 and thrombomodulin mRNA expression was quantitated in cell lysates. RESULTS: The injury resulted in a 2.1-fold increase in syndecan-4 (p < 0.001), consistent with EG shedding. Syndecan-4 and thrombomodulin mRNA expression was increased (p < 0.001) and decreased (p < 0.05), respectively, by the injury. Syndecan-4 shedding was not affected by treatment with FTY720, whereas FFP attenuated syndecan-4 shedding back to baseline levels in the injured cells and this was unaffected by W146. Neither treatment affected syndecan-4 or thrombomodulin mRNA expression. CONCLUSIONS: FTY720 did not prevent syndecan-4 shedding from the EG in the HUVEC model of endothelial injury, suggesting that activation of S1PR does not prevent EG damage. FFP prevented syndecan-4 shedding from the EG via a mechanism that was independent of S1PR1 and upregulation of SDC-4 production. Further studies to examine whether FTY720 or another S1PR agonist might have EG-protective effects under different conditions are warranted, as are investigations seeking the mechanism of EG protection conferred by FFP in this experimental model.

Anti-Fibrotic Potential of Tomentosenol A, a Constituent of Cerumen from the Australian Native Stingless Bee, Tetragonula carbonaria.

Bioactivity-guided fractionation was used to isolate two compounds, tomentosenol A (1) and torellianone A (2), from a cerumen extract from Tetragonula carbonaria. The anti-fibrotic activity of these compounds was examined using human cultured neonatal foreskin fibroblasts (NFF) and immortalised keratinocytes (HaCaTs). Tomentosenol A (1), inhibited NFF and HaCaT cell proliferation and prevented NFF and HaCaT scratch wound repopulation at 12.5-25 µM concentrations. These inhibitory effects were associated with reduced cell viability, determined by tetrazolium dye (MTT) and sulforhodamine B (SRB) assays. Compound 1 further inhibited transforming growth factor-ß1 (TGF-ß1)-stimulated, NFF-myofibroblast differentiation and soluble collagen production; and was an effective scavenger of the model oxidant, 2,2-diphenyl-1-picrylhydrazyl (DPPH·), with an EC50 value of 44.7 ± 3.1 µM. These findings reveal significant anti-fibrotic potential for cerumen-derived tomentosenol A (1).

The Association Between Contrast-Enhanced Ultrasound and Near-Infrared Spectroscopy-Derived Measures of Calf Muscle Microvascular Responsiveness in Older Adults.

BACKGROUND AND AIM: Contrast-enhanced ultrasound (CEUS) measures of post-occlusion skeletal muscle microvascular responsiveness demonstrate the microvascular dysfunction associated with ageing and age-related disease. However, the accessibility of CEUS is limited by the need for intravenous administration of ultrasound contrast agents and sophisticated imaging analysis. Alternative methods are required for the broader assessment of microvascular dysfunction in research and clinical settings. Therefore, we aimed to evaluate the level of association and agreement between CEUS and near-infrared spectroscopy (NIRS)-derived measures of post-occlusion skeletal muscle microvascular responsiveness in older adults. METHODS: During supine rest, participants (n=15, 67±11 years) underwent 5 minutes of thigh cuff-occlusion (200 mmHg). Post-occlusion CEUS measures of calf muscle microvascular responsiveness were made, including time to 95% peak acoustic intensity (TTP95 AI) and the rate of rise (slope AI). Simultaneous measures, including time to 95% peak oxygenated haemoglobin (TTP95 O2Hb) and slope O2Hb, were made using continuous-wave NIRS in the same muscle region. RESULTS: There were strong correlations between TTP95 measures derived from CEUS and NIRS (r=0.834, p=<0.001) and the corresponding measures of slope (r=0.735, p=0.004). The limits of agreement demonstrated by Bland Altman plot analyses for CEUS and NIRS-derived measures of TTP95 (-9.67-1.98 s) and slope (-1.29-5.23%. s-1) were smaller than the minimum differences expected in people with microvascular dysfunction. CONCLUSIONS: The strong correlations and level of agreement in the present study support the use of NIRS as a non-invasive, portable and cost-effective method for assessing post-occlusion skeletal muscle microvascular responsiveness in older adults.

Measurement of the Concentration of Citrate in Human Biofluids in Patients Undergoing Continuous Renal Replacement Therapy Using Regional Citrate Anticoagulation: Application of a Two-Step Enzymatic Assay.

BACKGROUND: Continuous renal replacement therapy (CRRT) with regional citrate anticoagulation (RCA) is now commonly used to treat acute kidney injury in critically ill patients. The concentration of citrate is not routinely measured, with citrate accumulation and/or toxicity primarily assessed using surrogate measures. OBJECTIVES: The aim of this study was to measure the concentration of citrate in plasma and ultrafiltrate in patients receiving CRRT with RCA using a modified commercial enzymatic assay. METHODS: After meeting inclusion criteria, blood was sampled from 20 patients before, during, and after episodes of filtration. Using spectrophotometry, samples were tested for citrate concentration. Demographic and other clinical and biochemical data were also collected. Throughout, a 15 mmol/L solution of trisodium citrate was used as the prefilter anticoagulant. Results were analysed using STATA (v15.0) and presented as mean (SD), median (IQR), or simple proportion. Comparisons were made using either the Student t test or the Wilcoxon rank-sum test. Correlation was assessed using Pearson's r. RESULTS: Twenty patients (17 males) were enrolled in the study. Mean (SD) age was 63.7 years (9.9). Median (IQR) ICU length of stay was 281 h (199, 422) with 85% undergoing intermittent positive pressure ventilation. Median APACHE 3 score was 95 (87, 117) with an overall 30% mortality rate. Median filtration time was 85 h (46, 149). No difference was found between pre- and post-filtration plasma citrate concentrations (79 µmol/L [50] vs. 71 µmol/L [42], p = 0.65). Mean citrate concentration during filtration was 508 µmol/L (221) with a maximum of 1,070 µmol/L. This was significantly higher than the pre/post levels (p < 0.001). CONCLUSIONS: Plasma concentrations of citrate rose significantly during episodes of CRRT using RCA returning back to normal after cessation of treatment.

Use of kinetic data to model potential antioxidant activity: Radical scavenging capacity of Australian Eucalyptus honeys.

Antioxidant activity of honeys may be beneficial in wound healing processes by protecting cells against lipid oxidation. The DPPH assay assesses the efficacy of antioxidant molecules to reduce DPPH• to DPPHH. Studies determining EC50 are limited by single time-point determinations of antioxidant effect and can miss vital information about the rate of antioxidant response. Acquisition of kinetic data allows determination of the radical scavenging capacity (RSC) of honeys. The purpose of this study was to determine the RSC of 53 honeys from 16 species of Australian Eucalyptus trees and four samples of New Zealand manuka (Leptospermum scoparium) honey. Whereas honeys could not be differentiated based on EC50 values, significant differences were observed for RSC, supporting collection of kinetic data for honey analysis. The greatest RSC was observed for New Zealand manuka (4.6 ± 0.3 × 10-5 mg.mL-1.min-1), grey ironbark (E. paniculate; 3.4 ± 0.2 × 10-5 mg.mL-1.min-1) and river red gum honeys (E. camaldulensis; 3.2 ± 0.2 × 10-5 mg.mL-1.min-1).

Influence of cuff-occlusion duration on contrast-enhanced ultrasound assessments of calf muscle microvascular blood flow responsiveness in older adults.

NEW FINDINGS: What is the central question of the study? Cuff-occlusion duration may influence contrast-enhanced ultrasound (CEUS) assessments of skeletal muscle microvascular blood flow responsiveness: what are the effects of 1, 3 and 5 min cuff-occlusion on the magnitude and reliability of calf muscle microvascular blood flow responsiveness in older adults? What is the main finding and its importance? Calf muscle microvascular blood flow responsiveness was enhanced following 5 min cuff-occlusion compared with 1 min. The reliability of post-occlusion CEUS measurements was also improved following 5 min occlusion. The use of a standardized 5 min occlusion period should therefore be considered in future studies and clinical practice. ABSTRACT: Contrast-enhanced ultrasound (CEUS) is increasingly used in assessments of skeletal muscle microvascular blood flow responsiveness. In response to limb cuff-occlusion, some studies have reported significant impairments in CEUS measurements of microvascular blood flow in older adults with cardiovascular or metabolic disease, whereas others have failed to detect significant between-group differences, which has brought the reliability of the technique into question. In the absence of a standardized CEUS protocol, there is variance in the duration of cuff-occlusion used, which is likely to influence post-occlusion measurements of muscle microvascular blood flow. We aimed to determine the effect of cuff-occlusion duration by comparing the magnitude and reliability of CEUS measurements of calf muscle microvascular blood flow responsiveness in older adults (n = 15, 67 ± 11 years) following 1, 3 and 5 min occlusion periods. Microvascular blood flow (= microvascular volume × microvascular velocity) within the calf muscle was measured using real-time destruction-replenishment CEUS. Measurements were made following thigh cuff-occlusion (200 mmHg) periods of 1, 3 and 5 min in a random order. Microvascular blood flow was higher following 3 min (3.71 ± 1.46 aU s-1 ) and 5 min (3.47 ± 1.48 aU s-1 ) compared with 1 min (2.42 ± 1.27 aU s-1 , P = 0.002), which corresponded with higher microvascular volumes after 3 and 5 min compared with 1 min. Reliability was good following 5 min (intraclass correlation coefficient (ICC) 0.49) compared with poor following 1 min (ICC 0.34) and 3 min (ICC 0.35). This study demonstrates that the magnitude and reliability of calf muscle microvascular responsiveness is enhanced using a 5 min cuff-occlusion protocol compared with 1 min in older adults.

Endotoxin Tolerance in Abdominal Aortic Aneurysm Macrophages, In Vitro: A Case-Control Study.

Macrophages are implicated in the pathogenesis of abdominal aortic aneurysm (AAA). This study examined the environmentally conditioned responses of AAA macrophages to inflammatory stimuli. Plasma- and blood-derived monocytes were separated from the whole blood of patients with AAA (30-45 mm diameter; n = 33) and sex-matched control participants (n = 44). Increased concentrations of pro-inflammatory and pro-oxidant biomarkers were detected in the plasma of AAA patients, consistent with systemic inflammation and oxidative stress. However, in monocyte-derived macrophages, a suppressed cytokine response was observed in AAA compared to the control following stimulation with lipopolysaccharide (LPS) (tumor necrosis factor alpha (TNF-α) 26.9 ± 3.3 vs. 15.5 ± 3.2 ng/mL, p < 0.05; IL-6 3.2 ± 0.6 vs. 1.4 ± 0.3 ng/mL, p < 0.01). LPS-stimulated production of 8-isoprostane, a biomarker of oxidative stress, was also markedly lower in AAA compared to control participants. These findings are consistent with developed tolerance in human AAA macrophages. As Toll-like receptor 4 (TLR4) has been implicated in tolerance, macrophages were examined for changes in TLR4 expression and distribution. Although TLR4 mRNA and protein expression were unaltered in AAA, cytosolic internalization of receptors and lipid rafts was found. These findings suggest the inflamed, pro-oxidant AAA microenvironment favors macrophages with an endotoxin-tolerant-like phenotype characterized by a diminished capacity to produce pro-inflammatory mediators that enhance the immune response.

Long Chain Omega-3 Polyunsaturated Fatty Acids Improve Vascular Stiffness in Abdominal Aortic Aneurysm: A Randomized Controlled Trial.

Abdominal aortic aneurysm (AAA) is a vascular disease involving permanent focal dilation of the abdominal aorta (≥30 mm) that can lead to catastrophic rupture. Destructive remodeling of aortic connective tissue in AAA contributes to wall stiffening, a mechanical parameter of the arterial system linked to a heightened risk of cardiovascular morbidity and mortality. Since aortic stiffening is associated with AAA progression, treatment options that target vascular inflammation would appear prudent. Given this, and growing evidence indicating robust anti-inflammatory and vasoprotective properties for long chain omega-3 polyunsaturated fatty acids (LC n-3 PUFAs), this study evaluated the impact of these nutrients (1.8 g/day for 12 weeks) on indices of vascular stiffness in patients with AAA. At baseline, pulse wave velocity (PWV) and augmentation index normalized to a heart rate of 75 bpm (AIx75) were significantly higher in patients with AAA compared to control participants (PWV: 14.2 ± 0.4 m.s-1 vs. 12.6 ± 0.4 m.s-1, p = 0.014; AIx75: 26.4 ± 1.7% vs. 17.3 ± 2.7%, p = 0.005). Twelve-week LC n-3 PUFA supplementation significantly decreased PWV (baseline: 14.2 ± 0.6 m.s-1, week 12: 12.8 ± 0.7 m.s-1, p = 0.014) and heart rate (baseline: 63 ± 3 bpm, week 12: 58 ± 3 bpm, p = 0.009) in patients with AAA. No change was observed for patients receiving placebo capsules. While this raises the possibility that LC n-3 PUFAs provide improvements in aortic stiffness in patients with AAA, the clinical implications remain to be fully elucidated.

Aortic and Systemic Arterial Stiffness Responses to Acute Exercise in Patients With Small Abdominal Aortic Aneurysms.

OBJECTIVE/BACKGROUND: Elevated arterial stiffness is a characteristic of abdominal aortic aneurysm (AAA), and is associated with AAA growth and cardiovascular mortality. A bout of exercise transiently reduces aortic and systemic arterial stiffness in healthy adults. Whether the same response occurs in patients with AAA is unknown. The effect of moderate- and higher intensity exercise on arterial stiffness was assessed in patients with AAA and healthy adults. METHODS: Twenty-two men with small diameter AAAs (36 ± 5 mm; mean age 74 ± 6 years) and 22 healthy adults (mean age 72 ± 5 years) were included. Aortic stiffness was measured using carotid to femoral pulse wave velocity (PWV), and systemic arterial stiffness was estimated from the wave reflection magnitude (RM) and augmentation index (Alx75). Measurements were performed at rest and during 90 min of recovery following three separate test sessions in a randomised order: (i) moderate intensity continuous exercise; (ii) higher intensity interval exercise; or (iii) seated rest. RESULTS: At rest, PWV was higher in patients with AAA than in healthy adults (p < .001), while AIx75 and RM were similar between groups. No differences were observed between AAA patients and healthy adults in post-exercise aortic and systemic arterial stiffness after either exercise protocol. When assessed as the change from baseline (delta, Δ), post-exercise ΔAIx75 was not different to the seated rest protocol. Conversely, post-exercise ΔPWV and ΔRM were both lower at all time points than seated rest (p < .001). ΔPWV was lower immediately after higher intensity than after moderate intensity exercise (p = .015). CONCLUSION: High resting aortic stiffness in patients with AAA is not exacerbated after exercise. There was a similar post-exercise attenuation in arterial stiffness between patients with AAA and healthy adults compared with seated rest. This effect was most pronounced following higher intensity interval exercise, suggesting that this form of exercise may be a safe and effective adjunctive therapy for patients with small AAAs.

Omega-3 fatty acids decrease oxidative stress and inflammation in macrophages from patients with small abdominal aortic aneurysm.

Abdominal aortic aneurysm (AAA) is associated with inflammation and oxidative stress, the latter of which contributes to activation of macrophages, a prominent cell type in AAA. Omega-3 polyunsaturated fatty acids (n-3 PUFAs) have been reported to limit oxidative stress in animal models of AAA. The aim of this study was to evaluate the effect of the n-3 PUFA docosahexaenoic acid (DHA) on antioxidant defence in macrophages from patients with AAA. Cells were obtained from men with small AAA (diameter 3.0-4.5 cm, 75 ± 6 yr, n = 19) and age- matched male controls (72 ± 5 yr, n = 41) and incubated with DHA for 1 h before exposure to 0.1 µg/mL lipopolysaccharide (LPS) for 24 h. DHA supplementation decreased the concentration of tumour necrosis factor-α (TNF-α; control, 42.1 ± 13.6 to 5.1 ± 2.1 pg/ml, p < 0.01; AAA, 25.2 ± 9.8 to 1.9 ± 0.9 pg/ml, p < 0.01) and interleukin-6 (IL-6; control, 44.9 ± 7.7 to 5.9 ± 2.0 pg/ml, p < 0.001; AAA, 24.3 ± 5.2 to 0.5 ± 0.3 pg/ml, p < 0.001) in macrophage supernatants. DHA increased glutathione peroxidase activity (control, 3.2 ± 0.3 to 4.1 ± 0.2 nmol/min/ml/µg protein, p = 0.004; AAA, 2.3 ± 0.5 to 3.4 ± 0.5 nmol/min/ml/µg protein, p = 0.008) and heme oxygenase-1 mRNA expression (control, 1.5-fold increase, p < 0.001). The improvements in macrophage oxidative stress status serve as a stimulus for further investigation of DHA in patients with AAA.

A simple and effective method for the isolation and culture of human monocytes from small volumes of peripheral blood.

Innate immune cell defects contribute to severe autoimmunity and the pathogenesis of inflammatory disease. Monocyte-derived macrophages typically retain disease related signatures and represent an excellent in vitro model to uncover and validate mechanisms contributing to specific pathological states. Monocyte isolation procedures vary widely in terms of purity, yield, cost, degree of technical difficulty and volume of peripheral blood needed. This paper outlines a novel isolation method that yields monocytes through density gradient centrifugation (Ficoll® and hyperosmotic Percoll®). The protocol has been optimised for small volumes of blood (42 ml) and is simple, reproducible and inexpensive compared to other methods. Monocyte recovery is 70% (relative to monocyte numbers within the buffy coat) and the highly functional macrophages produced are characterised by excellent purity (98.6 ±â€¯0.6%) and intact activation and phagocytic capacities. The method is well suited to investigations involving patient populations where a particular subset of immune cells is known to contribute to the pathogenesis of a specific disease or is aberrant as a consequence of that disease.

n-3 PUFAs improve erythrocyte fatty acid profile in patients with small AAA: a randomized controlled trial.

Abdominal aortic aneurysm (AAA) is an important cause of death in older adults, which has no current drug therapy. Inflammation and abnormal redox status are believed to be key pathogenic mechanisms for AAA. In light of evidence correlating inflammation with aberrant fatty acid profiles, this study compared erythrocyte fatty acid content in 43 AAA patients (diameter 3.0-4.5 cm) and 52 healthy controls. In addition, the effect of omega-3 PUFA (n-3 PUFA) supplementation on erythrocyte fatty acid content was examined in a cohort of 30 AAA patients as part of a 12 week randomized placebo-controlled clinical trial. Blood analyses identified associations between AAA and decreased linoleic acid (LA), and AAA and increased Δ6-desaturase activity and biosynthesis of arachidonic acid (AA) from LA. Omega-3 PUFA supplementation (1.5 g DHA + 0.3 g EPA/day) decreased red blood cell distribution width (14.8 ± 0.4% to 13.8 ± 0.2%; P = 0.003) and levels of pro-inflammatory n-6 PUFAs (AA, 12.46 ± 0.23% to 10.14 ± 0.3%, P < 0.001; adrenic acid, 2.12 ± 0.13% to 1.23 ± 0.09%; P < 0.001). In addition, Δ-4 desaturase activity increased (DHA/docosapentaenoic acid ratio, 1.85 ± 0.14 to 3.93 ± 0.17; P < 0.001) and elongase 2/5 activity decreased (adrenic acid/AA ratio, 0.17 ± 0.01 to 0.12 ± 0.01; P < 0.01) following supplementation. The findings suggest that n-3 PUFAs improve fatty acid profiles and ameliorate factors associated with inflammation in AAA patients.

Cytokine Responses to Acute Exercise in Healthy Older Adults: The Effect of Cardiorespiratory Fitness.

Markers of chronic inflammation increase with aging, and are associated with cardiovascular disease prevalence and mortality. Increases in fitness with exercise training have been associated with lower circulating concentrations of cytokines known to have pro-inflammatory actions (such as interleukin-6 [IL-6]) and higher circulating concentrations of anti-inflammatory cytokines (interleukin-10 [IL-10]). However, the effect of cardiorespiratory fitness on acute cytokine responses to a single bout of exercise in healthy older individuals is unknown. We compared the response of plasma cytokines IL-6, tumor necrosis factor-alpha (TNF-α) and IL-10 to a bout of moderate-intensity continuous and higher-intensity interval exercise between older individuals with higher and lower levels of cardiorespiratory fitness. Sixteen lower-fit (VO2peak: 22.6±2.8 mL.kg-1.min-1) and fourteen higher-fit participants (VO2peak: 37.4±5.9 mL.kg-1.min-1) completed three 24 min experimental protocols in a randomized order: (1) moderate-intensity continuous exercise (40% of peak power output [PPO]); (2) higher-intensity interval exercise (12 × 1 min intervals at 70% PPO separated by 1 min periods at 10% PPO); or (3) non-exercise control. Plasma cytokines were measured at rest, immediately after, and during 90 min of recovery following exercise or control. Plasma IL-6 concentrations at baseline were greater in the higher-fit compared to the lower-fit group (P = 0.02), with no difference in plasma IL-10 or TNF-α concentrations at baseline between groups. Plasma IL-6 and IL-10 concentrations in both groups increased immediately after all protocols (IL-6: P = 0.02, IL-10: P < 0.01). However, there was no difference in the IL-6 and IL-10 response between the exercise and non-exercise (control) protocols. After all protocols, no changes in plasma TNF-α concentrations were observed in either the higher- or lower-fit groups. In this study, basal concentrations of circulating IL-6 were elevated in older individuals with higher levels of cardiorespiratory fitness. However, changes in plasma cytokine concentrations after exercise were not different to changes after non-exercise control in both the lower- and higher-fit groups.

Acute Inflammatory Responses to Exercise in Patients with Abdominal Aortic Aneurysm.

PURPOSE: Inflammation and extracellular matrix degeneration contribute to abdominal aortic aneurysm (AAA) development. We aimed to assess the effect of exercise intensity on circulating biomarkers of inflammation and extracellular matrix degeneration in patients with AAA and healthy older adults. METHODS: Twenty patients with AAA (74 ± 6 yr) and 20 healthy males (72 ± 5 yr) completed moderate-intensity cycling at 40% peak power output, higher-intensity intervals at 70% peak power output, and control (rest) on separate days. Circulating matrix metalloproteinase-9 (MMP-9), transforming growth factor beta 1, interleukin-6 (IL-6), IL-10, and tumor necrosis factor alpha (TNF-α) were analyzed at rest and 0 to 90 min postexercise. RESULTS: Biomarkers at baseline were similar between groups. IL-6 responses to exercise were similar between groups, with a greater increase in ΔIL-6 after moderate-intensity compared with higher-intensity exercise (P < 0.001). Delta MMP-9 showed a 118-ng·mL (95% confidence interval = 23 to 214, P = 0.02) greater increase immediately after higher-intensity exercise compared with changes in control in both groups. Delta MMP-9 then decreased by 114 ng·mL (18 to 211, P = 0.02) 90 min after higher-intensity exercise compared with the changes in control. Delta TNF-α was not different between protocols in healthy adults. In patients with AAA, delta TNF-α showed a greater decrease after higher-intensity compared with moderate-intensity exercise (-6.1 pg·mL, -8.5 to -3.6, P < 0.001) and control (-4.9 pg·mL, -7.4 to -2.4, P < 0.001). IL-10 and transforming growth factor beta 1 did not change in either group. CONCLUSIONS: These findings suggest that a bout of higher-intensity exercise elicits a greater anti-inflammatory response compared with moderate-intensity exercise, which may be further augmented in patients with AAA. Exercise-induced reductions in biomarkers associated with AAA progression may represent a protective effect of exercise in patients with AAA.

Natural products isolated from Tetragonula carbonaria cerumen modulate free radical-scavenging and 5-lipoxygenase activities in vitro.

BACKGROUND: Propolis and cerumen are plant-derived products found in honeybees and stingless bees, respectively. Although propolis is an ancient folk medicine, the bioactivities of cerumen obtained from Australian native stingless bees (Tetragonula carbonaria) have not been widely studied. Therefore, we investigated selected anti-oxidant and anti-inflammatory properties of T. carbonaria cerumen. METHODS: A methanolic extract was prepared from the combined cerumen of 40 T. carbonaria hives, and HPLC was used to screen for chemical constituents that scavenged 2,2-azobis(2-methylpropionamidine) dihydrochloride (AAPH). The ability of cerumen extracts to scavenge 1,1-diphenyl-2-picrylhydrazyl (DPPH) and to interfere with leukotriene B4 (LTB4) production in ionomycin-stimulated human neutrophils was also examined. RESULTS: The extract dose-dependently scavenged DPPH (EC50 = 27.0 ± 2.3 µg/mL); and inhibited the 5-lipoxygenase (5-LOX)-mediated oxidation of linoleic acid (IC50 = 67.1 ± 9.6 µg/mL). Pre-treatment of isolated human neutrophils with the methanolic cerumen extract additionally inhibited the ionomycin-stimulated production of LTB4 from these cells (IC50 = 13.3 ± 5.3 µg/mL). Following multi-solvent extraction, the free radical-scavenging and 5-LOX-inhibiting activities of the initial cerumen extract were retained in a polar, methanol-water extract, which contained gallic acid and a range of flavonone and phenolic natural products. CONCLUSIONS: The findings identify free radical scavenging activity, and interference by extracts of T. carbonaria cerumen in 5-LOX-LTB4 signaling. Further investigation is needed to determine whether the extracts will provide therapeutic benefits for medical conditions in which oxidative stress and inflammation are implicated, including cardiovascular disease and impaired wound healing.

Abdominal aortic aneurysm and omega-3 polyunsaturated fatty acids: Mechanisms, animal models, and potential treatment.

Abdominal aortic aneurysm (AAA) is an inflammatory disease associated with macrophage accumulation in the adventitia, oxidative stress, medial elastin degradation and aortic dilation. Progression of AAA is linked to increased risk of rupture, which carries a high mortality rate. Drug therapies trialled to date lack efficacy and although aneurysm repair is available for patients with large aneurysm, peri-surgical morbidity and mortality have been widely reported. Recent studies using rodent models of AAA suggest that long chain omega-3 polyunsaturated fatty acids (LC n-3 PUFAs) and their metabolites can moderate inflammation and oxidative stress perpetuated by infiltrating macrophages and intervene in the destruction of medial elastin. This review examines evidence from these animal studies and related reports of inhibition of inflammation and arrest of aneurysm development following prophylactic supplementation with LC n-3 PUFAs. The efficacy of LC n-3 PUFAs for management of existing aneurysm is unclear and further investigations involving human clinical trials are warranted.