Publications by authors named "Ilaria Bellantuono"

38 Publications

miR-24 and its target gene Prdx6 regulate viability and senescence of myogenic progenitors during aging.

Aging Cell 2021 Oct 24;20(10):e13475. Epub 2021 Sep 24.

Institute of Life Course and Medical Sciences, University of Liverpool, Liverpool, UK.

Satellite cell-dependent skeletal muscle regeneration declines during aging. Disruptions within the satellite cells and their niche, together with alterations in the myofibrillar environment, contribute to age-related dysfunction and defective muscle regeneration. In this study, we demonstrated an age-related decline in satellite cell viability and myogenic potential and an increase in ROS and cellular senescence. We detected a transient upregulation of miR-24 in regenerating muscle from adult mice and downregulation of miR-24 during muscle regeneration in old mice. FACS-sorted satellite cells were characterized by decreased levels of miR-24 and a concomitant increase in expression of its target: Prdx6. Using GFP reporter constructs, we demonstrated that miR-24 directly binds to its predicted site within Prdx6 mRNA. Subtle changes in Prdx6 levels following changes in miR-24 expression indicate miR-24 plays a role in fine-tuning Prdx6 expression. Changes in miR-24 and Prdx6 levels were associated with altered mitochondrial ROS generation, increase in the DNA damage marker: phosphorylated-H2Ax and changes in viability, senescence, and myogenic potential of myogenic progenitors from mice and humans. The effects of miR-24 were more pronounced in myogenic progenitors from old mice, suggesting a context-dependent role of miR-24 in these cells, with miR-24 downregulation likely a part of a compensatory response to declining satellite cell function during aging. We propose that downregulation of miR-24 and subsequent upregulation of Prdx6 in muscle of old mice following injury are an adaptive response to aging, to maintain satellite cell viability and myogenic potential through regulation of mitochondrial ROS and DNA damage pathways.
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http://dx.doi.org/10.1111/acel.13475DOI Listing
October 2021

Zoledronate extends healthspan and survival via the mevalonate pathway in a FOXO-dependent manner.

J Gerontol A Biol Sci Med Sci 2021 Jun 17. Epub 2021 Jun 17.

Healthy Lifespan Institute, Department of Oncology and Metabolism, The Medical School, University of Sheffield, Sheffield.

Over recent decades, increased longevity has not been paralleled by extended healthspan, resulting in more years spent with multiple diseases in older age. As such, interventions to improve healthspan are urgently required. Zoledronate is a nitrogen containing bisphosphonate, which inhibits the farnesyl pyrophosphate synthase (FPPS) enzyme, central to the mevalonate pathway. It is already used clinically to prevent fractures in osteoporotic patients, who have been reported to derive unexpected and unexplained survival benefits. Using Drosophila as a model we determined the effects of Zoledronate on lifespan, parameters of healthspan (climbing ability and intestinal dysplasia) and the ability to confer resistance to oxidative stress using a combination of genetically manipulated Drosophila strains and Western blotting. Our study shows that Zoledronate extended lifespan, improved climbing activity and reduced intestinal epithelial dysplasia and permeability with age. Mechanistic studies showed that Zoledronate conferred resistance to oxidative stress and reduced accumulation of X-ray-induced DNA damage via inhibition of FPPS. Moreover, Zoledronate was associated with inhibition of pAKT in the mTOR pathway downstream of the mevalonate pathway and required dFOXO for its action, both molecules associated with increased longevity. Taken together, our work indicates that Zoledronate, a drug already widely used to prevent osteoporosis and dosed only once a year, modulates important mechanisms of ageing. Its repurposing holds great promise as a treatment to improve healthspan.
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http://dx.doi.org/10.1093/gerona/glab172DOI Listing
June 2021

What researchers on ageing should know about multimorbidity, geroprotectors and artificial intelligence.

Mech Ageing Dev 2021 04 9;195:111453. Epub 2021 Feb 9.

Department of Clinical and Experimental Sciences, University of Brescia, Brescia, Italy. Electronic address:

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http://dx.doi.org/10.1016/j.mad.2021.111453DOI Listing
April 2021

Tackling immunosenescence to improve COVID-19 outcomes and vaccine response in older adults.

Lancet Healthy Longev 2020 Nov 9;1(2):e55-e57. Epub 2020 Nov 9.

AGE Research Group, NIHR Newcastle Biomedical Research Centre, Newcastle University, Newcastle upon Tyne Hospitals NHS Foundation Trust.

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http://dx.doi.org/10.1016/S2666-7568(20)30011-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7834195PMC
November 2020

The use of geroprotectors to prevent multimorbidity: Opportunities and challenges.

Mech Ageing Dev 2021 01 2;193:111391. Epub 2020 Nov 2.

Healthy Lifespan Institute, Department of Oncology & Metabolism, University of Sheffield, UK. Electronic address:

Over 60 % of people over the age of 65 will suffer from multiple diseases concomitantly but the common approach is to treat each disease separately. As age-associated diseases have common underlying mechanisms there is potential to tackle many diseases with the same pharmacological intervention. These are known as geroprotectors and could overcome the problems related to polypharmacy seen with the use of the single disease model. With some geroprotectors now reaching the end stage of preclinical studies and early clinical trials, there is a need to review the evidence and assess how they can be translated practically and effectively into routine practice. Despite promising evidence, there are many gaps and challenges in our understanding that must be addressed to make geroprotective medicine effective in the treatment of age-associated multimorbidity. Here we highlight the key barriers to clinical translation and discuss whether geroprotectors such as metformin, rapamycin and senolytics can tackle all age-associated diseases at the same dose, or whether a more nuanced approach is required. The evidence suggests that geroprotectors' mode of action may differ in different tissues or in response to different inducers of accelerating ageing, suggesting that a blunt 'one drug for many diseases' approach may not work. We make the case for the use of artificial intelligence to better understand multimorbidity, allowing identification of clusters and networks of diseases that are significantly associated beyond chance and the underpinning molecular pathway of ageing causal to each cluster. This will allow us to better understand the development of multimorbidity, select a more homogenous group of patients for intervention, match them with the appropriate geroprotector and identify biomarkers specific to the cluster.
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http://dx.doi.org/10.1016/j.mad.2020.111391DOI Listing
January 2021

Senescence and Cancer: A Review of Clinical Implications of Senescence and Senotherapies.

Cancers (Basel) 2020 Jul 31;12(8). Epub 2020 Jul 31.

Departments of Internal Medicine, Geriatric Medicine and Gerontology, The Mayo Clinic, Rochester, MN 55905, USA.

Cellular senescence is a key component of human aging that can be induced by a range of stimuli, including DNA damage, cellular stress, telomere shortening, and the activation of oncogenes. Senescence is generally regarded as a tumour suppressive process, both by preventing cancer cell proliferation and suppressing malignant progression from pre-malignant to malignant disease. It may also be a key effector mechanism of many types of anticancer therapies, such as chemotherapy, radiotherapy, and endocrine therapies, both directly and via bioactive molecules released by senescent cells that may stimulate an immune response. However, senescence may contribute to reduced patient resilience to cancer therapies and may provide a pathway for disease recurrence after cancer therapy. A new group of drugs, senotherapies, (drugs which interact with senescent cells to interfere with their pro-aging impacts by either selectively destroying senescent cells (senolytic drugs) or inhibiting their function (senostatic drugs)) are under active investigation to determine whether they can enhance the efficacy of cancer therapies and improve resilience to cancer treatments. Senolytic drugs include quercetin, navitoclax, and fisetin and preclinical and early phase clinical data are emerging of their potential role in cancer treatments, although none are yet in routine use clinically. This article provides a review of these issues.
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http://dx.doi.org/10.3390/cancers12082134DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7464619PMC
July 2020

Modelling physical resilience in ageing mice.

Mech Ageing Dev 2019 01 2;177:91-102. Epub 2018 Oct 2.

MRC/Arthritis Research-UK Centre for Integrated Research into Musculoskeletal Ageing (CIMA), Department of Oncology and Metabolism, The Medical School, Beech Hill Road, Sheffield, S10 2RX, United Kingdom. Electronic address:

Geroprotectors, a class of drugs targeting multiple deficits occurring with age, necessitate the development of new animal models to test their efficacy. The COST Action MouseAGE is a European network whose aim is to reach consensus on the translational path required for geroprotectors, interventions targeting the biology of ageing. In our previous work we identified frailty and loss of resilience as a potential target for geroprotectors. Frailty is the result of an accumulation of deficits, which occurs with age and reduces the ability to respond to adverse events (physical resilience). Modelling frailty and physical resilience in mice is challenging for many reasons. There is no consensus on the precise definition of frailty and resilience in patients or on how best to measure it. This makes it difficult to evaluate available mouse models. In addition, the characterization of those models is poor. Here we review potential models of physical resilience, focusing on those where there is some evidence that the administration of acute stressors requires integrative responses involving multiple tissues and where aged mice showed a delayed recovery or a worse outcome then young mice in response to the stressor. These models include sepsis, trauma, drug- and radiation exposure, kidney and brain ischemia, exposure to noise, heat and cold shock.
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http://dx.doi.org/10.1016/j.mad.2018.10.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6445352PMC
January 2019

An -Ethyl--Nitrosourea (ENU) Mutagenized Mouse Model for Autosomal Dominant Nonsyndromic Kyphoscoliosis Due to Vertebral Fusion.

JBMR Plus 2018 May 8;2(3):154-163. Epub 2018 Mar 8.

Academic Endocrine Unit Radcliffe Department of Medicine University of Oxford Oxford Centre for Diabetes, Endocrinology and Metabolism Churchill Hospital Headington UK.

Kyphosis and scoliosis are common spinal disorders that occur as part of complex syndromes or as nonsyndromic, idiopathic diseases. Familial and twin studies implicate genetic involvement, although the causative genes for idiopathic kyphoscoliosis remain to be identified. To facilitate these studies, we investigated progeny of mice treated with the chemical mutagen -ethyl--nitrosourea (ENU) and assessed them for morphological and radiographic abnormalities. This identified a mouse with kyphoscoliosis due to fused lumbar vertebrae, which was inherited as an autosomal dominant trait; the phenotype was designated as hereditary vertebral fusion (HVF) and the locus as . Micro-computed tomography (μCT) analysis confirmed the occurrence of nonsyndromic kyphoscoliosis due to fusion of lumbar vertebrae in HVF mice, consistent with a pattern of blocked vertebrae due to failure of segmentation. μCT scans also showed the lumbar vertebral column of HVF mice to have generalized disc narrowing, displacement with compression of the neural spine, and distorted transverse processes. Histology of lumbar vertebrae revealed HVF mice to have irregularly shaped vertebral bodies and displacement of intervertebral discs and ossification centers. Genetic mapping using a panel of single nucleotide polymorphic (SNP) loci arranged in chromosome sets and DNA samples from 23 HVF (eight males and 15 females) mice, localized to chromosome 4A3 and within a 5-megabase (Mb) region containing nine protein coding genes, two processed transcripts, three microRNAs, five small nuclear RNAs, three large intergenic noncoding RNAs, and 24 pseudogenes. However, genome sequence analysis in this interval did not identify any abnormalities in the coding exons, or exon-intron boundaries of any of these genes. Thus, our studies have established a mouse model for a monogenic form of nonsyndromic kyphoscoliosis due to fusion of lumbar vertebrae, and further identification of the underlying genetic defect will help elucidate the molecular mechanisms involved in kyphoscoliosis. © 2018 The Authors. is published by Wiley Periodicals, Inc. on behalf of the American Society for Bone and Mineral Research.
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http://dx.doi.org/10.1002/jbm4.10033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6124210PMC
May 2018

Find drugs that delay many diseases of old age.

Nature 2018 02;554(7692):293-295

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http://dx.doi.org/10.1038/d41586-018-01668-0DOI Listing
February 2018

Find drugs that delay many diseases of old age.

Nature 2018 Feb;554(7692):293-295

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http://dx.doi.org/10.1038/d41586-018-01668-0DOI Listing
February 2018

Longitudinal effects of Parathyroid Hormone treatment on morphological, densitometric and mechanical properties of mouse tibia.

J Mech Behav Biomed Mater 2017 11 25;75:244-251. Epub 2017 Jul 25.

Department of Mechanical Engineering, The University of Sheffield, Sheffield, UK; Insigneo Institute for in Silico Medicine, The University of Sheffield, Sheffield, UK.

The use of Parathyroid Hormone (PTH) as bone anabolic is limited due to cost-benefit assessments. Preclinical studies evaluating the effects of PTH on bone have reported variable and often contradictory results. Here, we have applied a new approach using a combination of in-vivo longitudinal µCT, image processing techniques and finite element models to monitor early local changes in the whole tibia (divided in 40 compartments) and mechanical properties of female C57BL/6J mice treated with PTH 1-34, compared to controls. Compared with standard 3D bone morphometric analysis, our new approach allowed detection of much smaller and localised changes in bone mineral content (BMC) at very early time points (1 week vs 3 weeks with standard methods) and showed that changes do not occur uniformly over time and across the anatomical space. Indeed, in the PTH treated mice, significant changes in BMC were observed in the medial and posterior sectors of the proximal tibia, a week after treatment, and in the medial sector of the tibia midshaft region a week later (p < 0.05). By the third week, two thirds of the regions showed significantly higher values of BMC (p < 0.05). The effect of PTH on bone regional volume is similar to that on BMC, but there is almost no effect of PTH on bone tissue mineral density. The differences in estimated mechanical properties became significant after three weeks of treatment (p < 0.05). These results provide the first evidence of an early and localised PTH effect on murine bone, and show that our novel partitioning approach, compared to the standard evaluation protocol, allows a more precise quantification of bone changes following treatment, which would facilitate preclinical testing of novel mono- and/or combination therapies throughout the bone.
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http://dx.doi.org/10.1016/j.jmbbm.2017.07.034DOI Listing
November 2017

Interventions for age-related diseases: Shifting the paradigm.

Mech Ageing Dev 2016 12 29;160:69-92. Epub 2016 Sep 29.

MRC Arthiritis Research UK Centre for Integrated Research into Musculoskeletal Ageing (CIMA), Department of Oncology and Metabolism, University of Sheffield, The Medical School, Sheffield, UK. Electronic address:

Over 60% of people aged over 65 are affected by multiple morbidities, which are more difficult to treat, generate increased healthcare costs and lead to poor quality of life compared to individual diseases. With the number of older people steadily increasing this presents a societal challenge. Age is the major risk factor for age-related diseases and recent research developments have led to the proposal that pharmacological interventions targeting common mechanisms of ageing may be able to delay the onset of multimorbidity. Here we review the state of the knowledge of multimorbidity, appraise the available evidence supporting the role of mechanisms of ageing in the development of the most common age-related diseases and assess potential molecules that may successfully target those key mechanisms.
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http://dx.doi.org/10.1016/j.mad.2016.09.009DOI Listing
December 2016

MouseAge: Leading the way in the development of geroprotectors in Europe.

Mech Ageing Dev 2016 12 22;160:32-33. Epub 2016 Jun 22.

National Helenic Research Foundation, Institute of Biology, Medicinal Chemistry and Biotechnology, Athens, Greece.

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http://dx.doi.org/10.1016/j.mad.2016.06.007DOI Listing
December 2016

Does age matter? The impact of rodent age on study outcomes.

Lab Anim 2017 Apr 10;51(2):160-169. Epub 2016 Jul 10.

1 National Centre for the Replacement, Refinement and Reduction of Animals in Research, London, UK.

Rodent models produce data which underpin biomedical research and non-clinical drug trials, but translation from rodents into successful clinical outcomes is often lacking. There is a growing body of evidence showing that improving experimental design is key to improving the predictive nature of rodent studies and reducing the number of animals used in research. Age, one important factor in experimental design, is often poorly reported and can be overlooked. The authors conducted a survey to assess the age used for a range of models, and the reasoning for age choice. From 297 respondents providing 611 responses, researchers reported using rodents most often in the 6-20 week age range regardless of the biology being studied. The age referred to as 'adult' by respondents varied between six and 20 weeks. Practical reasons for the choice of rodent age were frequently given, with increased cost associated with using older animals and maintenance of historical data comparability being two important limiting factors. These results highlight that choice of age is inconsistent across the research community and often not based on the development or cellular ageing of the system being studied. This could potentially result in decreased scientific validity and increased experimental variability. In some cases the use of older animals may be beneficial. Increased scientific rigour in the choice of the age of rodent may increase the translation of rodent models to humans.
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http://dx.doi.org/10.1177/0023677216653984DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5367550PMC
April 2017

Development of a protocol to quantify local bone adaptation over space and time: Quantification of reproducibility.

J Biomech 2016 07 25;49(10):2095-2099. Epub 2016 May 25.

Department of Mechanical Engineering and INSIGNEO Institute for in Silico Medicine, the University of Sheffield, Sheffield, UK.

In vivo micro-computed tomography (µCT) scanning of small rodents is a powerful method for longitudinal monitoring of bone adaptation. However, the life-time bone growth in small rodents makes it a challenge to quantify local bone adaptation. Therefore, the aim of this study was to develop a protocol, which can take into account large bone growth, to quantify local bone adaptations over space and time. The entire right tibiae of eight 14-week-old C57BL/6J female mice were consecutively scanned four times in an in vivo µCT scanner using a nominal isotropic image voxel size of 10.4µm. The repeated scan image datasets were aligned to the corresponding baseline (first) scan image dataset using rigid registration. 80% of tibia length (starting from the endpoint of the proximal growth plate) was selected as the volume of interest and partitioned into 40 regions along the tibial long axis (10 divisions) and in the cross-section (4 sectors). The bone mineral content (BMC) was used to quantify bone adaptation and was calculated in each region. All local BMCs have precision errors (PE%CV) of less than 3.5% (24 out of 40 regions have PE%CV of less than 2%), least significant changes (LSCs) of less than 3.8%, and 38 out of 40 regions have intraclass correlation coefficients (ICCs) of over 0.8. The proposed protocol allows to quantify local bone adaptations over an entire tibia in longitudinal studies, with a high reproducibility, an essential requirement to reduce the number of animals to achieve the necessary statistical power.
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http://dx.doi.org/10.1016/j.jbiomech.2016.05.022DOI Listing
July 2016

Building for the future: essential infrastructure for rodent ageing studies.

Mamm Genome 2016 08 24;27(7-8):440-4. Epub 2016 May 24.

Department of Oncology and Metabolism, The Medical School, Beech Hill Road, Sheffield, S10 2RX, UK.

When planning ageing research using rodent models, the logistics of supply, long term housing and infrastructure provision are important factors to take into consideration. These issues need to be prioritised to ensure they meet the requirements of experiments which potentially will not be completed for several years. Although these issues are not unique to this discipline, the longevity of experiments and indeed the animals, requires a high level of consistency and sustainability to be maintained throughout lengthy periods of time. Moreover, the need to access aged stock or material for more immediate experiments poses many issues for the completion of pilot studies and/or short term intervention studies on older models. In this article, we highlight the increasing demand for ageing research, the resources and infrastructure involved, and the need for large-scale collaborative programmes to advance studies in both a timely and a cost-effective way.
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http://dx.doi.org/10.1007/s00335-016-9646-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4935732PMC
August 2016

Zoledronate Attenuates Accumulation of DNA Damage in Mesenchymal Stem Cells and Protects Their Function.

Stem Cells 2016 Mar 22;34(3):756-67. Epub 2015 Dec 22.

Mellanby Centre for Bone Research and MRC-Arthritis Research Centre for Integrated Research into Musculoskeletal Ageing (CIMA), Department of Human Metabolism, Medical School, University of Sheffield, Sheffield, United Kingdom.

Mesenchymal stem cells (MSCs) undergo a decline in function following ex vivo expansion and exposure to irradiation. This has been associated with accumulation of DNA damage and has important implications for tissue engineering approaches or in patients receiving radiotherapy. Therefore, interventions, which limit accumulation of DNA damage in MSC, are of clinical significance. We were intrigued by findings showing that zoledronate (ZOL), an anti-resorptive nitrogen containing bisphosphonate, significantly extended survival in patients affected by osteoporosis. The effect was too large to be simply due to the prevention of fractures. Moreover, in combination with statins, it extended the lifespan in a mouse model of Hutchinson Gilford Progeria Syndrome. Therefore, we asked whether ZOL was able to extend the lifespan of human MSC and whether this was due to reduced accumulation of DNA damage, one of the important mechanisms of aging. Here, we show that this was the case both following expansion and irradiation, preserving their ability to proliferate and differentiate in vitro. In addition, administration of ZOL before irradiation protected the survival of mesenchymal progenitors in mice. Through mechanistic studies, we were able to show that inhibition of mTOR signaling, a pathway involved in longevity and cancer, was responsible for these effects. Our data open up new opportunities to protect MSC from the side effects of radiotherapy in cancer patients and during ex vivo expansion for regenerative medicine approaches. Given that ZOL is already in clinical use with a good safety profile, these opportunities can be readily translated for patient benefit.
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http://dx.doi.org/10.1002/stem.2255DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4832316PMC
March 2016

The chondrocyte clock gene Bmal1 controls cartilage homeostasis and integrity.

J Clin Invest 2016 Jan 14;126(1):365-76. Epub 2015 Dec 14.

Osteoarthritis (OA) is the most prevalent and debilitating joint disease, and there are currently no effective disease-modifying treatments available. Multiple risk factors for OA, such as aging, result in progressive damage and loss of articular cartilage. Autonomous circadian clocks have been identified in mouse cartilage, and environmental disruption of circadian rhythms in mice predisposes animals to OA-like damage. However, the contribution of the cartilage clock mechanisms to the maintenance of tissue homeostasis is still unclear. Here, we have shown that expression of the core clock transcription factor BMAL1 is disrupted in human OA cartilage and in aged mouse cartilage. Furthermore, targeted Bmal1 ablation in mouse chondrocytes abolished their circadian rhythm and caused progressive degeneration of articular cartilage. We determined that BMAL1 directs the circadian expression of many genes implicated in cartilage homeostasis, including those involved in catabolic, anabolic, and apoptotic pathways. Loss of BMAL1 reduced the levels of phosphorylated SMAD2/3 (p-SMAD2/3) and NFATC2 and decreased expression of the major matrix-related genes Sox9, Acan, and Col2a1, but increased p-SMAD1/5 levels. Together, these results define a regulatory mechanism that links chondrocyte BMAL1 to the maintenance and repair of cartilage and suggest that circadian rhythm disruption is a risk factor for joint diseases such as OA.
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http://dx.doi.org/10.1172/JCI82755DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4701559PMC
January 2016

Evaluation of in-vivo measurement errors associated with micro-computed tomography scans by means of the bone surface distance approach.

Med Eng Phys 2015 Nov 1;37(11):1091-7. Epub 2015 Oct 1.

Department of Mechanical Engineering and INSIGNEO Institute for in silico Medicine, The University of Sheffield, Sheffield, UK.

In vivo micro-computed tomography (µCT) scanning is an important tool for longitudinal monitoring of the bone adaptation process in animal models. However, the errors associated with the usage of in vivo µCT measurements for the evaluation of bone adaptations remain unclear. The aim of this study was to evaluate the measurement errors using the bone surface distance approach. The right tibiae of eight 14-week-old C57BL/6 J female mice were consecutively scanned four times in an in vivo µCT scanner using a nominal isotropic image voxel size (10.4 µm) and the tibiae were repositioned between each scan. The repeated scan image datasets were aligned to the corresponding baseline (first) scan image dataset using rigid registration and a region of interest was selected in the proximal tibia metaphysis for analysis. The bone surface distances between the repeated and the baseline scan datasets were evaluated. It was found that the average (±standard deviation) median and 95th percentile bone surface distances were 3.10 ± 0.76 µm and 9.58 ± 1.70 µm, respectively. This study indicated that there were inevitable errors associated with the in vivo µCT measurements of bone microarchitecture and these errors should be taken into account for a better interpretation of bone adaptations measured with in vivo µCT.
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http://dx.doi.org/10.1016/j.medengphy.2015.08.017DOI Listing
November 2015

Shared Ageing Research Models (ShARM): a new facility to support ageing research.

Biogerontology 2013 Dec 2;14(6):789-94. Epub 2013 Oct 2.

Department of Human Metabolism, Medical School, Mellanby Centre for Bone Research, University of Sheffield, Beech Hill Road, Sheffield, S10 2RX, UK,

In order to manage the rise in life expectancy and the concomitant increased occurrence of age-related diseases, research into ageing has become a strategic priority. Mouse models are commonly utilised as they share high homology with humans and show many similar signs and diseases of ageing. However, the time and cost needed to rear aged cohorts can limit research opportunities. Sharing of resources can provide an ethically and economically superior framework to overcome some of these issues but requires dedicated infrastructure. Shared Ageing Research Models (ShARM) ( www.ShARMUK.org ) is a new, not-for-profit organisation funded by Wellcome Trust, open to all investigators. It collects, stores and distributes flash frozen tissues from aged murine models through its biorepository and provides a database of live ageing mouse colonies available in the UK and abroad. It also has an online environment (MICEspace) for collation and analysis of data from communal models and discussion boards on subjects such as the welfare of ageing animals and common endpoints for intervention studies. Since launching in July 2012, thanks to the generosity of researchers in UK and Europe, ShARM has collected more than 2,500 tissues and has in excess of 2,000 mice registered in live ageing colonies. By providing the appropriate support, ShARM has been able to bring together the knowledge and experience of investigators in the UK and Europe to maximise research outputs with little additional cost and minimising animal use in order to facilitate progress in ageing research.
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http://dx.doi.org/10.1007/s10522-013-9457-0DOI Listing
December 2013

Intra-femoral injection of human mesenchymal stem cells.

Methods Mol Biol 2013 ;976:131-41

Department of Human Metabolism, Mellanby Centre for Bone Research, University of Sheffield, Sheffield, UK.

In vivo transplantation of putative populations of hematopoietic stem cells (HSC) and assessment of their engraftment is considered the golden standard to assess their quality and degree of stemness. Transplantation is usually carried out by intravenous injection in murine models and assessment of engraftment is performed by monitoring the number and type of mature blood cells produced by the donor cells in time. In contrast intravenous injection of mesenchymal stem cells (MSC), the multipotent stem cells present in bone marrow and capable of differentiating to osteoblasts, chondrocytes and adipocytes, has not been successful. This is due to limited or absent engraftment levels. Here, we describe the use of intra-femoral injection as an improved method to assess MSC engraftment to bone and bone marrow and their quality.
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http://dx.doi.org/10.1007/978-1-62703-317-6_10DOI Listing
July 2013

Autosomal dominant hypercalciuria in a mouse model due to a mutation of the epithelial calcium channel, TRPV5.

PLoS One 2013 30;8(1):e55412. Epub 2013 Jan 30.

Academic Endocrine Unit, Nuffield Department of Medicine, University of Oxford, Oxford Centre for Diabetes, Endocrinology and Metabolism-OCDEM, Churchill Hospital, Headington, Oxford, United Kingdom.

Hypercalciuria is a major cause of nephrolithiasis, and is a common and complex disorder involving genetic and environmental factors. Identification of genetic factors for monogenic forms of hypercalciuria is hampered by the limited availability of large families, and to facilitate such studies, we screened for hypercalciuria in mice from an N-ethyl-N-nitrosourea mutagenesis programme. We identified a mouse with autosomal dominant hypercalciuria (HCALC1). Linkage studies mapped the Hcalc1 locus to a 11.94 Mb region on chromosome 6 containing the transient receptor potential cation channel, subfamily V, members 5 (Trpv5) and 6 (Trpv6) genes. DNA sequence analysis of coding regions, intron-exon boundaries and promoters of Trpv5 and Trpv6 identified a novel T to C transition in codon 682 of TRPV5, mutating a conserved serine to a proline (S682P). Compared to wild-type littermates, heterozygous (Trpv5(682P/+)) and homozygous (Trpv5(682P/682P)) mutant mice had hypercalciuria, polyuria, hyperphosphaturia and a more acidic urine, and ∼10% of males developed tubulointerstitial nephritis. Trpv5(682P/682P) mice also had normal plasma parathyroid hormone but increased 1,25-dihydroxyvitamin D(3) concentrations without increased bone resorption, consistent with a renal defect for the hypercalciuria. Expression of the S682P mutation in human embryonic kidney cells revealed that TRPV5-S682P-expressing cells had a lower baseline intracellular calcium concentration than wild-type TRPV5-expressing cells, suggesting an altered calcium permeability. Immunohistological studies revealed a selective decrease in TRPV5-expression from the renal distal convoluted tubules of Trpv5(682P/+) and Trpv5(682P/682P) mice consistent with a trafficking defect. In addition, Trpv5(682P/682P) mice had a reduction in renal expression of the intracellular calcium-binding protein, calbindin-D(28K), consistent with a specific defect in TRPV5-mediated renal calcium reabsorption. Thus, our findings indicate that the TRPV5 S682P mutant is functionally significant and study of HCALC1, a novel model for autosomal dominant hypercalciuria, may help further our understanding of renal calcium reabsorption and hypercalciuria.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0055412PLOS
July 2013

A small molecule modulator of prion protein increases human mesenchymal stem cell lifespan, ex vivo expansion, and engraftment to bone marrow in NOD/SCID mice.

Stem Cells 2012 Jun;30(6):1134-43

Mellanby Centre for Bone Research, Department of Human Metabolism, University of Sheffield, Sheffield, United Kingdom.

Human mesenchymal stem cells (hMSCs) have been shown to have potential in regenerative approaches in bone and blood. Most protocols rely on their in vitro expansion prior to clinical use. However, several groups including our own have shown that hMSCs lose proliferation and differentiation ability with serial passage in culture, limiting their clinical applications. Cellular prion protein (PrP) has been shown to enhance proliferation and promote self-renewal of hematopoietic, mammary gland, and neural stem cells. Here we show, for the first time, that expression of PrP decreased in hMSC following ex vivo expansion. When PrP expression was knocked down, hMSC showed significant reduction in proliferation and differentiation. In contrast, hMSC expanded in the presence of small molecule 3/689, a modulator of PrP expression, showed retention of PrP expression with ex vivo expansion and extended lifespan up to 10 population doublings. Moreover, cultures produced a 300-fold increase in the number of cells generated. These cells showed a 10-fold increase in engraftment levels in bone marrow 5 weeks post-transplant. hMSC treated with 3/689 showed enhanced protection from DNA damage and enhanced cell cycle progression, in line with data obtained by gene expression profiling. Moreover, upregulation of superoxide dismutase-2 (SOD2) was also observed in hMSC expanded in the presence of 3/689. The increase in SOD2 was dependent on PrP expression and suggests increased scavenging of reactive oxygen species as mechanism of action. These data point to PrP as a good target for chemical intervention in stem cell regenerative medicine.
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http://dx.doi.org/10.1002/stem.1065DOI Listing
June 2012

Glycogen synthase kinase-3α/β inhibition promotes in vivo amplification of endogenous mesenchymal progenitors with osteogenic and adipogenic potential and their differentiation to the osteogenic lineage.

J Bone Miner Res 2011 Apr;26(4):811-21

Mellanby Centre for Bone Research, Department of Human Metabolism, School of Medicine, Dentistry and Health, University of Sheffield, Sheffield, United Kingdom.

Small molecules are attractive therapeutics to amplify and direct differentiation of stem cells. They also can be used to understand the regulation of their fate by interfering with specific signaling pathways. Mesenchymal stem cells (MSCs) have the potential to proliferate and differentiate into several cell types, including osteoblasts. Activation of canonical Wnt signaling by inhibition of glycogen synthase kinase 3 (GSK-3) has been shown to enhance bone mass, possibly by involving a number of mechanisms ranging from amplification of the mesenchymal stem cell pool to the commitment and differentiation of osteoblasts. Here we have used a highly specific novel inhibitor of GSK-3, AR28, capable of inducing β-catenin nuclear translocation and enhanced bone mass after 14 days of treatment in BALB/c mice. We have shown a temporally regulated increase in the number of colony-forming units-osteoblast (CFU-O) and -adipocyte (CFU-A) but not colony-forming units-fibroblast (CFU-F) in mice treated for 3 days. However, the number of CFU-O and CFU-A returned to normal levels after 14 days of treatment, and the number of CFU-F was decreased significantly. In contrast, the number of osteoblasts increased significantly only after 14 days of treatment, and this was seen together with a significant decrease in bone marrow adiposity. These data suggest that the increased bone mass is the result of an early temporal wave of amplification of a subpopulation of MSCs with both osteogenic and adipogenic potential, which is driven to osteoblast differentiation at the expense of adipogenesis.
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http://dx.doi.org/10.1002/jbmr.266DOI Listing
April 2011

Alterations in the self-renewal and differentiation ability of bone marrow mesenchymal stem cells in a mouse model of rheumatoid arthritis.

Arthritis Res Ther 2010 22;12(4):R149. Epub 2010 Jul 22.

Department of Human Metabolism, University of Sheffield, Sheffield S10 2RX, UK.

Introduction: Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease primarily involving the synovium. Evidence in recent years has suggested that the bone marrow (BM) may be involved, and may even be the initiating site of the disease. Abnormalities in haemopoietic stem cells' (HSC) survival, proliferation and aging have been described in patients affected by RA and ascribed to abnormal support by the BM microenvironment. Mesenchymal stem cells (MSC) and their progeny constitute important components of the BM niche. In this study we test the hypothesis that the onset of inflammatory arthritis is associated with altered self-renewal and differentiation of bone marrow MSC, which alters the composition of the BM microenvironment.

Methods: We have used Balb/C Interleukin-1 receptor antagonist knock-out mice, which spontaneously develop RA-like disease in 100% of mice by 20 weeks of age to determine the number of mesenchymal progenitors and their differentiated progeny before, at the start and with progression of the disease.

Results: We showed a decrease in the number of mesenchymal progenitors with adipogenic potential and decreased bone marrow adipogenesis before disease onset. This is associated with a decrease in osteoclastogenesis. Moreover, at the onset of disease a significant increase in all mesenchymal progenitors is observed together with a block in their differentiation to osteoblasts. This is associated with accelerated bone loss.

Conclusions: Significant changes occur in the BM niche with the establishment and progression of RA-like disease. Those changes may be responsible for aspects of the disease, including the advance of osteoporosis. An understanding of the molecular mechanisms leading to those changes may lead to new strategies for therapeutic intervention.
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http://dx.doi.org/10.1186/ar3098DOI Listing
January 2011

Aging of marrow stromal (skeletal) stem cells and their contribution to age-related bone loss.

Biochim Biophys Acta 2009 Apr 27;1792(4):364-70. Epub 2009 Jan 27.

University of Sheffield Medical School, Sheffield S10 2RX, UK.

Marrow stromal cells (MSC) are thought to be stem cells with osteogenic potential and therefore responsible for the repair and maintenance of the skeleton. Age related bone loss is one of the most prevalent diseases in the elder population. It is controversial whether MSC undergo a process of aging in vivo, leading to decreased ability to form and maintain bone homeostasis with age. In this review we summarize evidence of MSC involvement in age related bone loss and suggest new emerging targets for intervention.
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http://dx.doi.org/10.1016/j.bbadis.2009.01.008DOI Listing
April 2009

Telomere length dynamics differ in foetal and early post-natal human leukocytes in a longitudinal study.

Biogerontology 2009 Jun 7;10(3):279-84. Epub 2008 Nov 7.

Royal Manchester Children's Hospital, Manchester, M27 4HA, UK.

Haemopoietic stem cells (HSC) undergo a process of self renewal to constantly maintain blood cell turnover. However, it has become apparent that adult HSC lose their self-renewal ability with age. Telomere shortening in peripheral blood leukocytes has been seen to occur with age and it has been associated with loss of HSC proliferative capacity and cellular ageing. In contrast foetal HSC are known to have greater proliferative capacity than post-natal stem cells. However it is unknown whether they undergo a similar process of telomere shortening. In this study we show a more accentuated rate of telomere loss in leukocytes from pre term infants compared to human foetuses of comparable age followed longitudinally for 8-12 weeks in a longitudinal study. Our results point to a difference in HSC behaviour between foetal and early postnatal life which is independent of age but may be influenced by events at birth itself.
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http://dx.doi.org/10.1007/s10522-008-9194-yDOI Listing
June 2009

Direct muscle delivery of GDNF with human mesenchymal stem cells improves motor neuron survival and function in a rat model of familial ALS.

Mol Ther 2008 Dec 16;16(12):2002-10. Epub 2008 Sep 16.

The Waisman Center, University of Wisconsin-Madison, Madison, Wisconsin 53705-2280, USA.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease in which there is a progressive loss of motor neurons and their connections to muscle, leading to paralysis. In order to maintain muscle connections in a rat model of familial ALS (FALS), we performed intramuscular transplantation with human mesenchymal stem cells (hMSCs) used as "Trojan horses" to deliver growth factors to the terminals of motor neurons and to the skeletal muscles. hMSCs engineered to secrete glial cell line-derived neurotrophic factor (hMSC-GDNF) were transplanted bilaterally into three muscle groups. The cells survived within the muscle, released GDNF, and significantly increased the number of neuromuscular connections and motor neuron cell bodies in the spinal cord at mid-stages of the disease. Further, intramuscular transplantation with hMSC-GDNF was found to ameliorate motor neuron loss within the spinal cord where it connects with the limb muscles receiving transplants. While disease onset was similar in all the animals, hMSC-GDNF significantly delayed disease progression, increasing overall lifespan by up to 28 days, which is one of the largest effects on survival noted for this rat model of FALS. This preclinical data provides a novel and practical approach toward ex vivo gene therapy for ALS.
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http://dx.doi.org/10.1038/mt.2008.197DOI Listing
December 2008

Stem cell ageing: does it happen and can we intervene?

Expert Rev Mol Med 2007 Nov 19;9(31):1-20. Epub 2007 Nov 19.

Academic Unit of Bone Biology, University of Sheffield Medical School, Sheffield, S10 2RX, UK.

Adult stem cells have become the focus of intense research in recent years as a result of their role in the maintenance and repair of tissues. They exert this function through their extensive expansion (self-renewal) and multipotent differentiation capacity. Understanding whether adult stem cells retain this capacity throughout the lifespan of the individual, or undergo a process of ageing resulting in a decreased stem cell pool, is an important area of investigation. Progress in this area has been hampered by lack of suitable models and of appropriate markers and assays to identify stem cells. However, recent data suggest that an understanding of the mechanisms governing stem cell ageing can give insight into the mechanism of tissue ageing and, most importantly, advance our ability to use stem cells in cell and gene therapy strategies.
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http://dx.doi.org/10.1017/S146239940700049XDOI Listing
November 2007

Hematopoietic progenitor cell deficiency in fetuses and children affected by Down's syndrome.

Exp Hematol 2006 Dec;34(12):1611-5

Stem Cell Research Group, Royal Manchester Children's Hospital, Manchester, UK.

Objectives: There is an increased risk of myeloid malignancy in individuals with Down's syndrome (DS), which is associated with a mutation in exon 2 of the transcription factor GATA-1. It is recognized that there is accelerated telomere shortening in blood cells of children with DS similar to that in conditions such as Fanconi anemia and dyskeratosis congenita. The latter conditions are associated with stem cell deficiency and clonal change, including acute myeloid leukemia. In this study we address the questions 1) whether the accelerated telomere shortening is associated with progenitor/stem cell deficiency in individuals with DS, predisposing to clonal change and 2) whether the occurrence of reduced numbers of stem/progenitor cells precede the incidence of mutations in exon 2 of GATA-1.

Material And Methods: Peripheral blood from fetuses (23-35 weeks gestation) and/or bone marrow from children affected by DS and age-matched hematologically healthy controls were analyzed for telomere length, content of stem/progenitor cells, and mutations in exon 2 of GATA-1.

Results: We found that hematopoietic stem/progenitor cell deficiency and telomere shortening occurs in individuals with DS in fetal life. Moreover, the presence of a low number of progenitor cells was not associated with mutations in exon 2 of GATA-1.

Conclusions: We propose that stem cell deficiency may be a primary predisposing event to DS leukemia development.
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http://dx.doi.org/10.1016/j.exphem.2006.10.013DOI Listing
December 2006
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