Modulating the Adhesion of Haematopoietic Stem Cells with Chemokines to Enhance Their Recruitment to the Ischaemically Injured Murine Kidney

Introduction

Renal disease affects over 500 million people worldwide and is set to increase as treatment options are predominately supportive. Evidence suggests that exogenous haematopoietic stem cells (HSCs) can be of benefit but due to the rarity and poor homing of these cells, benefits are either minor or transitory. Mechanisms governing HSC recruitment to injured renal microcirculation are poorly understood; therefore this study determined (i) the adhesion molecules responsible for HSC recruitment to the injured kidney, (ii) if cytokine HSC pre-treatment can enhance their homing and (iii) the molecular mechanisms accountable for any enhancement.

Methods

Adherent and free-flowing HSCs were determined in an intravital murine model of renal ischaemia-reperfusion injury. Some HSCs and animals were pre-treated prior to HSC infusion with function blocking antibodies, hyaluronidase or cytokines. Changes in surface expression and clustering of HSC adhesion molecules were determined using flow cytometry and confocal microscopy. HSC adhesion to endothelial counter-ligands (VCAM-1, hyaluronan) was determined using static adhesion assays in vitro.

Results

CD49d, CD44, VCAM-1 and hyaluronan governed HSC adhesion to the IR-injured kidney. Both KC and SDF-1α pre-treatment strategies significantly increased HSC adhesion within injured kidney, whilst SDF-1α also increased numbers continuing to circulate. SDF-1α and KC did not increase CD49d or CD44 expression but increased HSC adhesion to VCAM-1 and hyaluronan respectively. SDF-1α increased CD49d surface clustering, as well as HSC deformability.

Conclusion

Increasing HSC adhesive capacity for its endothelial counter-ligands, potentially through surface clustering, may explain their enhanced renal retention in vivo. Furthermore, increasing HSC deformability through SDF-1α treatment could explain the prolonged systemic circulation; the HSC can therefore continue to survey the damaged tissue instead of becoming entrapped within non-injured sites. Therefore manipulating these mechanisms of HSC recruitment outlined may improve the clinical outcome of cellular therapies for kidney disease.     

New developments in the law for obesity discrimination protection

Background:

Obese individuals are frequent targets of weight‐based discrimination, particularly in the employment setting. Victims of weight discrimination have sought legal restitution like others who have suffered from different forms of discrimination. However, in the vast majority of the United States, body weight is not a protected class and weight‐based employment discrimination does not provide a basis for a legal claim. Some have attempted to seek legal recourse under the Rehabilitation Act of 1973 or the Americans with Disabilities Act of 1990 (collectively, the ADA), which protect against discrimination based on mental or physical disabilities in a variety of settings. Until recently, claims of weight discrimination under the ADA have also been largely unsuccessful. However, Congress recently passed the ADA Amendments Act, expanding the definition of what constitutes a disability and incorporating a broad view of ADA's coverage.

Objective:

This short communication provides an update of the law as it relates to employment based discrimination of obese people. The authors propose a legislative direction for future legal recourse.

Design and Methods:

The authors conducted legal research into the ADA Amendments Act, and synthesized this work relating to discrimination against weight in the employment context.

Results:

In light of the ADA Amendments Act, courts and the Equal Employment Opportunity Commission have provided protection for severely obese people from discrimination based on actual or perceived disability in the employment context.

Conclusion:

The authors discuss this positive legal development and additionally propose a targeted solution to address weight discrimination in the employment setting. National polling suggests there is considerable public support for such a measure. The authors thus recommend the implementation of a “Weight Discrimination in Employment Act” modeled after the Age Discrimination in Employment Act to adequately address this pervasive and damaging injustice toward individuals who are affected by obesity.     

Are our actions aligned With our evidence? The skinny on changing the landscape of obesity

Recent debate about the role of food deserts in the United States (i.e., places that lack access to healthy foods) has prompted discussion on policies being enacted, including efforts that encourage the placement of full‐service supermarkets into food deserts. Other initiatives to address obesogenic neighborhood features include land use zoning and parks renovations. Yet, there is little evidence to demonstrate that such policies effect change. While we suspect most researchers and policymakers would agree that effective neighborhood change could be a powerful tool in combating obesity, we desperately need strong and sound evidence to guide decisions about where and how to invest.     

The Bidirectional Association between Depressive Symptoms and Gait Speed: Evidence from the English Longitudinal Study of Ageing (ELSA)

Background

Depressive symptoms and physical performance are inversely associated, but it is unclear whether their association is bidirectional. We examined whether the association between depressive symptoms and physical performance measured using gait speed is bidirectional.

Methods

We used a national sample of 4,581 community-dwelling people aged 60 years and older from the English Longitudinal Study of Ageing (from 2002–03 to 2008-09). We fitted Generalized Estimating Equation (GEE) regression models to analyse repeated measurements of gait speed (m/sec) and elevated depressive symptoms (defined as a score of ≥4 on the eight-item Center for Epidemiological Studies-Depression scale).

Results

Slower gait speed was associated with elevated depressive symptoms both concurrently and two years later. After adjustment for previous depressive symptoms and sociodemographic, clinical, lifestyle, psychosocial, and cognitive factors the concurrent association was partially explained (Odds Ratio [OR] 0.42, 95% confidence interval [CI], 0.30 to 0.59, per 1m/sec increase in gait speed) and the two-year lagged association fully (OR 0.75, 95% CI, 0.56 to 1.00). Elevated depressive symptoms were associated with slower gait speed. Full adjustment for covariates (including previous gait speed) partially explained both the concurrent (β regression coefficient [β] -0.038, 95% CI, -0.050 to -0.026, for participants with elevated depressive symptoms compared with those with no or one symptom) and the two-year lagged associations (β -0.017, 95% CI, -0.030 to -0.005). Subthreshold depressive symptoms (defined as a score of two or three on the eight-item Center for Epidemiological Studies-Depression scale) were also associated with slower gait speed. Full adjustment for covariates partially explained both the concurrent (β -0.029, 95% CI, -0.039 to -0.019, for participants with subthreshold symptoms compared with those with no or one symptom) and the two-year lagged associations (β -0.011, 95% CI, -0.021 to -0.001).

Conclusions

The inverse association between gait speed and depressive symptoms appears to be bidirectional.     

Akonni TruTip? and Qiagen? Methods for Extraction of Fetal Circulating DNA - Evaluation by Real-Time and Digital PCR

Due to the low percentage of fetal DNA present in maternal plasma (< 10%) during early gestation, efficient extraction processes are required for successful downstream detection applications in non-invasive prenatal diagnostic testing. In this study, two extraction methods using similar chemistries but different workflows were compared for isolation efficiency and percent fetal DNA recovery. The Akonni Biosystems TruTip technology uses a binding matrix embedded in a pipette tip; the Circulating Nucleic Acids Kit from Qiagen employs a spin column approach. The TruTip method adds an extra step to decrease the recovery of DNA fragments larger than 600 bp from the sample to yield an overall higher percentage of smaller molecular weight DNA, effectively enriching for fetal DNA. In this evaluation, three separate extraction comparison studies were performed - a dilution series of fragmented DNA in plasma, a set of clinical maternal samples, and a blood collection tube time point study of maternal samples. Both extraction methods were found to efficiently extract small fragment DNA from large volumes of plasma. In the amended samples, the TruTip extraction method was ~15% less efficient with overall DNA recovery, but yielded an 87% increase in % fetal DNA relative to the Qiagen method. The average percent increase of fetal DNA of TruTip extracted samples compared to the Qiagen method was 55% for all sets of blinded clinical samples. A study comparing extraction efficiencies from whole blood samples incubated up to 48 hours prior to processing into plasma resulted in more consistent % fetal DNA recoveries using TruTip. The extracted products were tested on two detection platforms, quantitative real-time PCR and droplet digital PCR, and yielded similar results for both extraction methods.     

Tissue-specific patterns of allelically-skewed DNA methylation

While DNA methylation is usually thought to be symmetrical across both alleles, there are some notable exceptions. Genomic imprinting and X chromosome inactivation are two well-studied sources of allele-specific methylation (ASM), but recent research has indicated a more complex pattern in which genotypic variation can be associated with allelically-skewed DNA methylation in cis. Given the known heterogeneity of DNA methylation across tissues and cell types we explored inter- and intra-individual variation in ASM across several regions of the human brain and whole blood from multiple individuals. Consistent with previous studies, we find widespread ASM with > 4% of the ～220,000 loci interrogated showing evidence of allelically-skewed DNA methylation. We identify ASM flanking known imprinted regions, and show that ASM sites are enriched in DNase I hypersensitivity sites and often located in an extended genomic context of intermediate DNA methylation. We also detect examples of genotype-driven ASM, some of which are tissue-specific. These findings contribute to our understanding of the nature of differential DNA methylation across tissues and have important implications for genetic studies of complex disease. As a resource to the community, ASM patterns across each of the tissues studied are available in a searchable online database: http://epigenetics.essex.ac.uk/ASMBrainBlood.     

Living on the edge: substrate competition explains loss of robustness in mitochondrial fatty-acid oxidation disorders

Background

Defects in genes involved in mitochondrial fatty-acid oxidation (mFAO) reduce the ability of patients to cope with metabolic challenges. mFAO enzymes accept multiple substrates of different chain length, leading to molecular competition among the substrates. Here, we combined computational modeling with quantitative mouse and patient data to investigate whether substrate competition affects pathway robustness in mFAO disorders.

Results

First, we used comprehensive biochemical analyses of wild-type mice and mice deficient for medium-chain acyl-CoA dehydrogenase (MCAD) to parameterize a detailed computational model of mFAO. Model simulations predicted that MCAD deficiency would have no effect on the pathway flux at low concentrations of the mFAO substrate palmitoyl-CoA. However, high concentrations of palmitoyl-CoA would induce a decline in flux and an accumulation of intermediate metabolites. We proved computationally that the predicted overload behavior was due to substrate competition in the pathway. Second, to study the clinical relevance of this mechanism, we used patients’ metabolite profiles and generated a humanized version of the computational model. While molecular competition did not affect the plasma metabolite profiles during MCAD deficiency, it was a key factor in explaining the characteristic acylcarnitine profiles of multiple acyl-CoA dehydrogenase deficient patients. The patient-specific computational models allowed us to predict the severity of the disease phenotype, providing a proof of principle for the systems medicine approach.

Conclusion

We conclude that substrate competition is at the basis of the physiology seen in patients with mFAO disorders, a finding that may explain why these patients run a risk of a life-threatening metabolic catastrophe.