A simple method for the purification of human peripheral blood monocytes : A substitute for Sepracell-MN

Sepracell-MN has provided a simple, rapid means of isolating peripheral blood monocytes. However this product is no longer available. Consequentially we have developed a Percoll gradient wich matches Sepracell-MN in simplicity and yield of monocytes. Using this Percoll gradient, an average of 7 x 106 monocytes with a purity of 83% were obtained from 30-40 ml of blood. These monocytes were at least 97% viable and responded to chemotactic stimuli in comparable numbers to those prepared using Sepracell-MN.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/29026/1/0000056.pd

Effects of Greenshell™ mussel intervention on biomarkers of cartilage metabolism, inflammatory markers and joint symptoms in overweight/obese postmenopausal women: A randomized, double-blind, and placebo-controlled trial

ObjectiveTo investigate the effect of whole greenshell mussel (GSM) powder on biomarkers of cartilage metabolism, inflammatory cytokines, and joint symptoms in postmenopausal women with overweight/obesity and joint discomfort.DesignFifty-five postmenopausal women with overweight/obesity were randomly assigned to receive 3 g/day whole GSM powder or placebo for 12 weeks. Cartilage turnover biomarkers urinary C-telopeptide of type II collagen (CTX-II) and serum cartilage oligomeric matrix protein (COMP) were measured at baseline, week 6 and 12. Plasma cytokines were measured at baseline and week 12. Joint pain and knee-related problems were assessed at baseline and week 12 using a 100 mm Visual Analogue Scale (VAS) and the Knee injury and Osteoarthritis Outcome Score (KOOS) questionnaire, respectively.ResultsForty-nine participants completed the study (GSM n = 25, placebo n = 24). After 12 weeks, urinary CTX-II showed no significant change over time or between the groups (interaction effect P = 0.1). However, in women with symptomatic knees, a significant difference was noted between the group (treatment effect P = 0.04), as it was lower in the GSM group compared to placebo group at week 6 (P = 0.04) and week 12 (P = 0.03). Serum COMP and plasma cytokines were not affected. GSM supplementation showed greater reduction in the VAS pain score than placebo (−13.2 ± 20.3 vs. −2.9 ± 15.9; P = 0.04). No significant change in KOOS domains between the two groups was observed.ConclusionOral supplementation of whole GSM powder at 3 g/day may slow down the degradation of type II collagen in postmenopausal women with symptomatic knees. GSM treatment conferred clinical benefit on overall joint pain. No significant effect was noted for inflammatory cytokines, suggesting that GSM may act within the joint microenvironment rather than at the systemic level.Clinical trial registration[www.australianclinicaltrials.gov.au/clinical-trialregistries], identifier [ACTRN12620000413921p]

Hearing Ability with Age in Northern European Women: A New Web-Based Approach to Genetic Studies

Age-related hearing impairment (ARHI) affects 25–40% of individuals over the age of 65. Despite the high prevalence of this complex trait, ARHI is still poorly understood. We hypothesized that variance in hearing ability with age is largely determined by genetic factors. We collected audiologic data on females of Northern European ancestry and compared different audiogram representations. A web-based speech-to-noise ratio (SNR) hearing test was compared with pure-tone thresholds to see if we could determine accurately hearing ability on people at home and the genetic contribution to each trait compared. Volunteers were recruited from the TwinsUK cohort. Hearing ability was determined using pure-tone audiometry and a web-based hearing test. Different audiogram presentations were compared for age-correlation and reflection of audiogram shape. Using structural equation modelling based on the classical twin model the heritability of ARHI, as measured by the different phenotypes, was estimated and shared variance between the web-based SNR test and pure-tone audiometry determined using bivariate modelling. Pure-tone audiometric data was collected on 1033 older females (age: 41–86). 1970 volunteers (males and females, age: 18–85) participated in the SNR. In the comparison between different ARHI phenotypes the difference between the first two principle components (PC1–PC2) best represented ARHI. The SNR test showed a sensitivity and specificity of 89% and 80%, respectively, in comparison with pure-tone audiogram data. Univariate heritability estimates ranged from 0.70 (95% CI: 0.63–0.76) for (PC1–PC2) to 0.56 (95% CI: 0.48–0.63) for PC2. The genetic correlation of PC1–PC2 and SNR was −0.67 showing that the 2 traits share variances attributed to additive genetic factors. Hearing ability showed considerable heritability in our sample. We have shown that the SNR test provides a useful surrogate marker of hearing. This will enable a much larger sample to be collected at a fraction of the cost, facilitating future genetic association studies

Case Reports1. A Late Presentation of Loeys-Dietz Syndrome: Beware of TGFβ Receptor Mutations in Benign Joint Hypermobility

Background: Thoracic aortic aneurysms (TAA) and dissections are not uncommon causes of sudden death in young adults. Loeys-Dietz syndrome (LDS) is a rare, recently described, autosomal dominant, connective tissue disease characterized by aggressive arterial aneurysms, resulting from mutations in the transforming growth factor beta (TGFβ) receptor genes TGFBR1 and TGFBR2. Mean age at death is 26.1 years, most often due to aortic dissection. We report an unusually late presentation of LDS, diagnosed following elective surgery in a female with a long history of joint hypermobility. Methods: A 51-year-old Caucasian lady complained of chest pain and headache following a dural leak from spinal anaesthesia for an elective ankle arthroscopy. CT scan and echocardiography demonstrated a dilated aortic root and significant aortic regurgitation. MRA demonstrated aortic tortuosity, an infrarenal aortic aneurysm and aneurysms in the left renal and right internal mammary arteries. She underwent aortic root repair and aortic valve replacement. She had a background of long-standing joint pains secondary to hypermobility, easy bruising, unusual fracture susceptibility and mild bronchiectasis. She had one healthy child age 32, after which she suffered a uterine prolapse. Examination revealed mild Marfanoid features. Uvula, skin and ophthalmological examination was normal. Results: Fibrillin-1 testing for Marfan syndrome (MFS) was negative. Detection of a c.1270G > C (p.Gly424Arg) TGFBR2 mutation confirmed the diagnosis of LDS. Losartan was started for vascular protection. Conclusions: LDS is a severe inherited vasculopathy that usually presents in childhood. It is characterized by aortic root dilatation and ascending aneurysms. There is a higher risk of aortic dissection compared with MFS. Clinical features overlap with MFS and Ehlers Danlos syndrome Type IV, but differentiating dysmorphogenic features include ocular hypertelorism, bifid uvula and cleft palate. Echocardiography and MRA or CT scanning from head to pelvis is recommended to establish the extent of vascular involvement. Management involves early surgical intervention, including early valve-sparing aortic root replacement, genetic counselling and close monitoring in pregnancy. Despite being caused by loss of function mutations in either TGFβ receptor, paradoxical activation of TGFβ signalling is seen, suggesting that TGFβ antagonism may confer disease modifying effects similar to those observed in MFS. TGFβ antagonism can be achieved with angiotensin antagonists, such as Losartan, which is able to delay aortic aneurysm development in preclinical models and in patients with MFS. Our case emphasizes the importance of timely recognition of vasculopathy syndromes in patients with hypermobility and the need for early surgical intervention. It also highlights their heterogeneity and the potential for late presentation. Disclosures: The authors have declared no conflicts of interes

Regulation of T-lymphocyte recruitment by endothelial selectins and alpha4 integrins in pulmonary inflammation.

The recruitment of leukocytes during inflammation is mediated by cell adhesion molecules expressed on both leukocytes and endothelium of blood vessels in inflamed tissue. This study defined the relationship between cell adhesion molecules on the pulmonary vasculature and on leukocytes recovered from the lung parenchyma and air space during a delayed-hypersensitivity response to intratracheal (IT) sheep red blood cells (SRBC) in sensitized C57BL/6J mice. Silver-enhanced immunogold staining and RT-PCR of lung tissues revealed low constitutive expression of VCAM-1, E-selectin and P-selectin in normal lung vasculature, and sustained induction of these molecules throughout the lung for up to 7 days following pulmonary challenge with SRBC. Selectin-chimera binding studies on lymphocytes following IT SRBC challenge revealed that the number of selectin ligand-positive T-cells in the peripheral blood decreased rapidly, at the same time as these cells rapidly increased in bronchoalveolar lavage fluid and the lung interstitium. Interdicting adhesion pathways mediated by $\alpha$4 integrins or P-selectin resulted in a partial blockage of mononuclear leukocyte accumulation in lung three days after pulmonary challenge. In contrast, blocking L-selectin or $\alpha4$ integrins, or genetically deleting expression of all selectin ligands, had no effect on neutrophil accumulation in the lung one day after challenge. Lymphocyte trafficking was examined by infusing and tracking fluorescently-labeled T-cells in vivo during the SRBC-induced pulmonary immune response. Labeled cells recovered from the lung displayed ligands for E- and P-selectin in proportions significantly higher than that seen in the original infused populations. Blocking $\alpha$4 integrins or genetically deleting selectin ligand expression on labeled T-lymphoblasts resulted in $\sim$50% decrease in the cells' ability to traffic into inflamed lung. Combining these two treatments nearly ablated recruitment of the labeled T-cells. These studies determined that both $\alpha$4 integrins and P-selectin may mediate lymphocyte and monocyte accumulation during pulmonary inflammation. However, selectin interactions are not required for neutrophil recruitment into inflamed lung in response to SRBC. Trafficking studies in vivo demonstrated that the $\alpha$4 integrins and endothelial selectins independently contribute to T-cell recruitment into inflamed lung.Ph.D.Biological SciencesCellular biologyHealth and Environmental SciencesImmunologyOncologyPathologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/130397/2/9722124.pd

Osteoporosis: Modern Paradigms for Last Century’s Bones

The skeleton is a metabolically active organ undergoing continuously remodelling. With ageing and menopause the balance shifts to increased resorption, leading to a reduction in bone mineral density and disruption of bone microarchitecture. Bone mass accretion and bone metabolism are influenced by systemic hormones as well as genetic and lifestyle factors. The classic paradigm has described osteoporosis as being a “brittle bone” disease that occurs in post-menopausal, thin, Caucasian women with low calcium intakes and/or vitamin D insufficiency. However, a study of black women in Africa demonstrated that higher proportions of body fat did not protect bone health. Isoflavone interventions in Asian postmenopausal women have produced inconsistent bone health benefits, due in part to population heterogeneity in enteric bacterial metabolism of daidzein. A comparison of women and men in several Asian countries identified significant differences between countries in the rate of bone health decline, and a high incidence rate of osteoporosis in both sexes. These studies have revealed significant differences in genetic phenotypes, debunking long-held beliefs and leading to new paradigms in study design. Current studies are now being specifically designed to assess genotype differences between Caucasian, Asian, African, and other phenotypes, and exploring alternative methodology to measure bone architecture

Digestion behaviour of capsaicinoid-loaded emulsion gels and bioaccessibility of capsaicinoids: Effect of emulsifier type

In this study, the effect of emulsifier type, i.e. whey protein versus Tween 80, on the digestion behaviour of emulsion gels containing capsaicinoids (CAPs) was examined. The results indicate that the CAP-loaded Tween 80 emulsion gel was emptied out significantly faster during gastric digestion than the CAP-loaded whey protein emulsion gel. The Tween-80-coated oil droplets appeared to be in a flocculated state in the emulsion gel, had no interactions with the protein matrix and were easily released from the protein matrix during gastric digestion. The whey-protein-coated oil droplets showed strong interactions with the protein matrix, and the presence of thick protein layer around the oil droplets protected their liberation during gastric digestion. During intestinal digestion, the CAP-loaded Tween 80 emulsion gel had a lower extent of lipolysis than the CAP-loaded whey protein emulsion gel, probably because the interfacial layer formed by Tween 80 was resistance to displacement by bile salts, and/or because Tween 80 formed interfacial complexes with bile salts/lipolytic enzymes. Because of the softer structure of the CAP-loaded Tween 80 emulsion gel, the gel particles were broken down much faster and the oil droplets were liberated from the protein matrix more readily than for the CAP-loaded whey protein emulsion gel during intestinal digestion; this promoted the release of CAP molecules from the gel. In addition, the Tween 80 molecules displaced from the interface would participate in the formation of mixed micelles and would help to solubilize the released CAP molecules, leading to improved bioaccessibility of CAP. Information obtained from this study could be useful in designing functional foods for the delivery of lipophilic bioactive compounds

Effects of Greenshell Mussel (Perna canaliculus) Intake on Pathological Markers of Multiple Phenotypes of Osteoarthritis in Rats

The prevalence of metabolic osteoarthritis has been increasing worldwide, particularly among women. The aim of this study was to investigate the effects of the New Zealand greenshell mussel (Perna canaliculus; GSM) on osteoarthritis (OA) prevention in a rat model. One-hundred-and-eight female rats aged 12 weeks were divided into four test groups, containing 24 rats each, plus an additional control group. Each test group received one of the four experimental diets: normal control diet (ND), normal control diet supplemented with GSM (ND + GSM), high fat/high sugar diet (HFHS), or high fat/high sugar diet supplemented GSM (HFHS + GSM), for 36 weeks (end of the study). After 8 weeks on experimental diets, half of each group was subjected to ovariectomy (OVX) and the remaining half received a sham operation (ovaries left intact). The study evaluated body composition, bone mass, plasma cytokines, adipokines, HbA1c, CTX-II, and knee joint’s histopathology. HFHS diet and OVX significantly induced body weight gain and leptin production. OVX rats lost bone mineral density but increased adiponectin, HbA1C, and MCP-1. The OVX rats fed HFHS showed the highest Mankin scores. Importantly, inclusion of GSM reduced these pathological features. In conclusion, GSM might be beneficial in halting the progression of OA