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The role of sarcopenia with and without fracture
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The role of sarcopenia with and without fracture Umberto Tarantinoa,* , Jacopo Baldia,b , Manuel Scimecac,d , Eleonora Piccirillia,b , Andrea Picciolie, Elena Bonannoc , Elena Gasbarraa a
Department of Orthopaedics and Traumatology, “Tor Vergata” University of Rome, “Policlinico Tor Vergata” Foundation, Viale Oxford 1, 00133 Rome, Italy School of Specialisation in Orthopaedics and Traumatology, “Tor Vergata” University of Rome, “Policlinico Tor Vergata” Foundation, Viale Oxford 1, 00133 Rome, Italy c Anatomic Pathology Section, Department of Biomedicine and Prevention, University of Rome “Tor Vergata”, Via Montpellier 1, 00133 Rome, Italy d “Multidisciplinary Study of the Effects of Microgravity on Bone Cells” Project, Italian Space Agency (ASI), Spatial Biomedicine Center, Via del Politecnico snc, 00133 Rome, Italy e Oncologic Centre, “Palazzo Baleani”, Azienda Policlinico Umberto I, Corso Vittorio Emanuele II 244, Rome, Italy b
A R T I C L E I N F O
Keywords: Sarcopenia Osteoporosis Osteoarthritis Hip fracture Aging Risk of falling Muscle fibre atrophy Satellite cells BMP2 Myostatin
A B S T R A C T
Introduction: Bone and muscle tissues are in a close relationship. They are linked from a biological and functional point of view and both are related to an increased fracture risk in the elderly. The aging process is involved in the loss of functionality of both bones and muscles. In particular, aging-induced decline in muscle size and quality accompanies catabolic alterations in bone tissue; furthermore, age-related changes in bone alter its response to muscle-derived stimulation. The increased fracture risk in individuals with sarcopenia and osteoporosis is due to the decline of muscle mass and strength, the decrease in bone mineral density (BMD) and limited mobility. In this study, we investigated the role of sarcopenia and the main age-related bone diseases, osteoporosis (OP) and osteoarthritis (OA). Methods: Muscular performance status was evaluated using the Physical Activity Scale for the Elderly (PASE) test in 27 female patients with OP who underwent total hip arthroplasty for hip fracture, and in 27 age-matched female patients with OA who underwent total hip arthroplasty. Dual-energy X-ray absorptiometry (DEXA) was performed and the T-score values were used to discriminate between OP and OA patients. Body Mass Index (BMI) was calculated. As part of a multiparametric model of evaluation, biopsies of vastus lateralis muscle were analysed by immunohistochemical reaction to find a correlation with the above mentioned functional index. Results: The PASE test showed that the OP patients had a low or moderate level of physical activity before fracture occurred, whereas the OA patients had more intensive pre-fracture physical performances. Histological analysis showed that osteoporosis is characterised by a preferential type II fibre atrophy; in particular, data correlation showed that lower PASE test scores were related to lower diameter of type II fibres. No correlation was found between bone mineral density (BMD) and PASE test results. Discussion and conclusion: Osteoporosis is closely related to sarcopenia before and after fracture. Bone remodelling is influenced by muscle morphological and functional impairment and sarcopenia is considered one of the major factors for functional limitation and motor dependency in elderly osteoporotic individuals. Therefore, physical activity should be strongly recommended for OP patients at diagnosis. ã 2016 Elsevier Ltd. All rights reserved.
Background Bone and muscle are in a close relationship: when the aging process affects one of these two tissues, the functionality of the other is compromised. Consequently, sarcopenia and osteoporosis
* Corresponding author. E-mail address: [email protected] (U. Tarantino).
represent two pathologies that are frequently associated in the elderly. The term sarcopenia was first used by Rosenberg in 1989 to describe the decline in muscle mass among older people [1,2]. More recently, sarcopenia was defined as a loss of skeletal muscle mass and strength that occurs with aging [3]. Nowadays, the term sarcopenia is used in the literature to describe several pathophysiological processes, such as denervation, mitochondrial dysfunction, inflammatory and hormonal changes that may lead to a
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decrease in muscle strength and mobility, a decrease in fatigue resistance and an increase in the risk of falls and fractures [4]. Moreover, studies in the literature show that muscles of patients with sarcopenia display severe alterations in cellular turnover, including an increase in oxidative stress, cellular vacuolisation, and mitochondrial alterations, which compromise the quality of oxidant-scavenging systems. The denervation of single muscle fibres is also known to lead to a substantial reduction in type II fibres, which are gradually replaced by type I fibres and fat tissue [5,6]. Mechanisms involved in the decline of muscle mass during sarcopenia converge on the failure of satellite cells to replace and repair damaged muscle fibres. The main cause of reduced satellite cell function may be alteration of systemic factors that regulate satellite cell activity and differentiation. Indeed, satellite cells are mostly quiescent and their activation is governed by multiple niche factors (e.g. injury or stress) and signalling pathways, such as transforming growth factor-beta (TGF-b) and myogenin. The TGFb superfamily plays a crucial role in normal physiology and pathogenesis of skeletal muscle. Numerous members of the TGF-b family have been shown to play important roles in regulating muscle growth and atrophy. The most extensively characterised ligands, in terms of the effects on skeletal muscle, are TGF-b, myostatin and bone morphogenetic proteins (BMPs). Myogenin is a transcription factor that induces myogenesis in a variety of cell types in tissue culture. It is a member of the helix-loop-helix (HLH) protein family, a large family of proteins related by sequence homology, and it is essential for the development of functional skeletal muscle [7]. Muscle and bone are proportionally matched in their functional capacity and geometric structure; however, this relationship appears to change significantly with age. For example, the capacity for muscle to generate force declines with age, and the anabolic response of bone to muscle-derived strains also appears to be altered with age. In addition, muscle is now recognised to have paracrine and endocrine effects that may also influence bone independently of a mechanical relationship [8,9]. In this study, we investigated the role of sarcopenia and the main age-related bone diseases, osteoporosis (OP) and osteoarthritis (OA). Patients, materials and methods A total of 54 patients who underwent hip arthroplasty for femoral fractures or for hip OA in the Orthopaedic Department of “Tor Vergata” University from June 2014 to February 2015 were enrolled in this study. To evaluate bone mineral density (BMD), each patient with OA underwent dual-energy X-ray absorptiometry (DEXA) scan of the lumbar spine and femoral neck on the homolateral limb before surgery. Hip X-rays were performed to establish the grade of OA using the Kellgren-Lawrence Scale. To evaluate hip function, Harris Hip Score (HHS) was also calculated: the maximum score is 100 points, with the higher the HHS, the less the dysfunction. To estimate lumbar spine and non-fractured femur BMD, each OP patient underwent DEXA scan a few days after surgery; spinal Xrays were also performed in patients with femoral fracture or back pain to evaluate the presence of a vertebral compression fracture (VCF). A functional evaluation of performance status was conducted in each patient using the Physical Activity Scale for the Elderly (PASE) test. The questionnaire assesses two main groups of activities: recreational activities carried out during free time and domestic activities [10]. Each of these actions corresponds to a set value that is proportional to the “weight” that they could have in the life of each individual. The sum of the individual scores sets the patient
into one of four categories related to the degree of physical activity: inactivity (total score <42); lacking physical activity (43–105); moderate physical activity (106–145), and intense physical activity (>146). Body Mass Index (BMI) was also calculated. Exclusion criteria were: history of neoplastic diseases, myopathies or other neuromuscular diseases; taking anti-osteoporotic drugs or chronic administration (more than one month) of corticosteroid for autoimmune diseases; diabetes; alcohol abuse; cigarette smoking; and chronic viral infections (hepatitis B virus [HBV], hepatitis C virus [HCV], human immunodeficiency virus [HIV]). During surgery for total hip arthroplasty in both osteoarthritis (OA patients) and cervical femoral fragility fracture (OP patients), muscle biopsies were taken from the upper portion of the vastus lateralis muscle, which is normally involved in the surgical procedure, without damaging the tissue network. This muscle was chosen because it is hardly influenced by the fracture event and it is a good indicator of systemic muscle atrophy. All sampling and experiments were performed in agreement with the independent ethics committee, “Policlinico Tor Vergata”, approval reference number #85/12, and informed consent was obtained from all participants included in the study. Bone mineral density evaluation (DEXA) DEXA was performed with a Lunar DEXA apparatus (GE Healthcare, Madison, WI, USA). Lumbar spine (L1–L4) and femoral (neck and total) scans were performed, and BMD was analysed according to the manufacturer’s recommendations. DEXA measures BMD (in g/cm2) with a coefficient of variation of 0.7%. In the OA group, all measurements were performed on the non-dominant side; in the OP group, BMD was measured on the limb opposite to the fracture side. Results were expressed as absolute values and as T-scores [11,12]. Histology Muscle biopsies were fixed with 4% paraformaldehyde for 24 h and embedded in paraffin after alcoholic dehydration. Sections of 3 mm thick were stained with haematoxylin and eosin (H&E) and the pathological evaluation was performed by two pathologists who were blinded to the samples [13]. Atrophy assessment A minimum of 200 muscle fibres per biopsy were evaluated to assess fibre atrophy; minimum transverse diameter and crosssectional area of type I and type II fibres were compared for relative prevalence. A threshold diameter of less than 30 mm (minimum value of the normal range for women) characterised atrophic fibres [14,15]. To calculate muscle and bone areas, H&E slides were scanned at low-power field by Iscan Coreo (Ventana, Tucson, AZ, USA). Areas for each muscle and bone biopsy image were identified by a pathologist using Viewing software (Ventana, Tucson, AZ, USA). Immunohistochemistry Immunohistochemical characterisation was conducted to assess muscle fibre type (i.e. fast and slow) and the expression of myostatin, TGF-b, pax7 and myogenin. Briefly, 3 mm thick sections were pre-treated with EDTA citrate (pH 7.8) for 30 min at 95 C and then incubated, respectively, with mouse monoclonal anti-fast skeletal myosin for 60 min (1:100, clone MY-32, AbCam), mouse monoclonal anti-slow skeletal myosin for 60 min (1:100, clone NOQ7.5.4D, AbCam) and rabbit
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monoclonal anti-myostatin for 45 min (1:100, clone ab134682, AbCam). For myogenin, TGF-b and pax7, 3 mm thick sections were pre-treated with citrate (pH 6.0) for 30 min at 95 C and then incubated with mouse monoclonal anti-myogenin (1:100, clone F5D, AbCam), rabbit monoclonal anti-TGF-b for 60 min (1:250 clone 6B7, Novus Biologicals) and rabbit monoclonal anti-pax7 (1:500, NC, Novus Biologicals) [16]. A washing buffer composed of PBS 4% and Tween20 (pH 7.6) (UCS diagnostic, Rome, Italy) was used; reactions were revealed by HRP-DAB Detection Kit (UCS Diagnostic, Rome, Italy). Transmission electron microscopy (TEM) Small samples of muscle tissue from each patient were processed as previously described [17]. Briefly, muscle tissues were fixed in 4% paraformaldehyde, post-fixed in 2% osmium tetroxide and dehydrated by a series of incubations in 30%, 50% and 70%, ethanol. Samples were then embedded in EPON resin (Agar Scientific, Stansted, Essex, CM24 8GF, United Kingdom), for morphological ultrastructural analysis, and in LR-White resin (Agar Scientific, Stansted Essex, CM24 8GF, United Kingdom) for ultrastructural immunohistochemistry [18]. Tissues were cut and stained with heavy metal solutions, as described by Reynolds [17,18]. Statistical analysis Statistical analysis was performed using GraphPad Prism 5 Software (La Jolla, CA, USA). Morphometric data were expressed as percentage of atrophic fibres and percentage of tissue composition of muscle fasciculus and bone. The comparison of myostatin, TGF-b, BMP2, myogenin and pax7 expression between the two experimental groups was performed using the Mann– Whitney test. The difference between groups was considered statistically significant at P < 0.05. Immunohistochemistry data were expressed as mean value with standard deviation (n SD). Results Clinical evaluation Clinical evaluation was conducted and patients were divided into two groups: OP and OA. The OP group (femoral neck BMD range 0.454–0.645 g/cm2) included 27 females with fragility hip fracture, a T-score -2.5 SD and a negative radiographic framework for hip OA; the OA group (femoral neck BMD range 0.845–1.197 g/ cm2) included 27 females with a positive radiogram for hip OA with a Kellgren–Lawrence score of 3 or 4 and a T-score -2.5 SD (Table 1). The mean BMI of the OA group was statistically significantly higher than that of the OP group (26.88 0.85 kg/m2 vs 22.73 3.54 kg/m2, P < 0.001). These data confirm the frequently overweight condition of OA patients. The clinical results in this
Table 1 Evaluated parameters in patients with osteoarthritis (OA) and osteoporosis (OP). Variables
OA Group
OP Group
Age (years) Body Mass Index (kg/m2) PASE test Bone Mineral Density L1-L4 (g/m2) T-score L1-L4 Femoral neck Bone Mineral Density (g/m2) Femoral neck T-score Kellgren-Lawrence grade
71.6 10.3 26.88 0.85 100 20.86 0.957 0.20 1.8 1.66 0.845–1.197 1.4 0.94 3–4
82.4 6.19 22.73 3.54 62.6 33 0.927 0.18 2.3 1.08 0.454–0.645 2.92 1.12 0–1
3
study showed an inverse correlation between BMI and HHS in the OA group. Indeed, a high body mass impairs hip joint mobility. The results of the PASE test showed that both OP and OA patients were characterised by poor physical activity; however, patients in the OP group had a lower PASE test score than those in the OA group. BMD, T-score and Kellgren–Lawrence grading scale were used for patients’ instrumental evaluation. OA patients were characterised by physiological age-related osteopenia limited to the lumbar region (T-score 1 <> 2.5 SD) and generally higher values of BMD compared to the OP group (Table 1). Conversely, all OP patients showed low levels of BMD and a T-score <-2.5 SD. Radiological analysis enabled classification of the OA patients, who were at least grade 3 on the Kellgren-Lawrence scale, whereas OP patients showed 0 or 1 Kellgren-Lawrence grade (Table 1). Morphometric analysis Muscle morphometric examination performed on digital scanned images showed a reduction in muscle tissues of about 20% in the OA and OP groups (Fig. 1a–b). In OA patients, muscle was mainly replaced by adipose tissue (17.93%), whereas in OP muscle fasciculus, atrophic muscle fibres were substituted by a mixture of adipose (8.01%) and connective tissue (6.31%) (Fig. 1a–b). Slow myosin antibody and fast myosin antibody stains enabled discrimination between type I and type II fibres, respectively. The morphometric analysis of muscle fibres in OP patients showed more than 50% of atrophic fibres with prevalence of type II fibres (19.13% type I and 35.93% type II) (Fig. 1a). In the OA group, there were 35% of atrophic fibres with a diameter of less than 30 mm (15.70% type I and 19.30% type II) (Fig. 1b). Immunohistochemistry study Myostatin and TGF-b expression were assessed by counting the number of positive fibres on 25 high-power field (HPF) (Fig. 2a–b). Remarkably, OP muscle biopsies showed a significantly higher number of myostatin-positive fibres (40.25 5.70) compared with the muscle of OA patients (15.36 2.45) (Figs. 2a and 3a–d). Conversely, there were no significant differences between the number of TGF-b-positive fibres among OA and OP groups (Figs. 2b and 3e–f). Nonetheless, TGF-b expression tends to increase in the elderly. The presence and activity of satellite cells were studied using pax7 and myogenin expression. Their expression was evaluated by counting the number of positive satellite cells on 25 HPF (Fig. 2c–d). Nuclear expression of pax7 was higher in muscle biopsy of OA patients compared with that of OP patients (OA 33.44 2.48 vs OP 18.74 1.84) (Figs. 2c and 3g–h). Notably, in OA muscle tissue, it was frequently observed that pax7-positive cells were aggregated to form syncytia. Finally, our results showed a significantly different rate of myogenin-positive cells in the OA group compared with the OP group (OA 250.30 89.65 vs OP 150.60 52.53) (Fig. 2d). In particular, in OA patients, many groups of myogenin-positive satellite cells were focally observed in the tissue (Fig. 3i–j). Transmission electron microscopy TEM analysis was performed to identify satellite cells according to their ultrastructural characteristics. In OA muscle biopsies, several satellite cells were found tightly associated or fused to form a syncytium. Notably, in the cytoplasm of these cells there was a de novo production of sarcomeric structures (Fig. 4a–c). In OP patients, the number of satellite cells was very low and their niches showed an obvious mark of degeneration (Fig. 4d–f).
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Fig. 1. Morphometric analysis. In OA patients, muscle was mainly replaced by adipose tissue (17.93%), whereas in OP muscle fasciculus, atrophic muscle fibres were substituted by a mixture of adipose (8.01%) and connective tissue (6.31%) (A–B). The morphometric analysis of muscle fibres in OP patients showed more than 50% of atrophic fibres with prevalence of type II fibres (19.13% type I and 35.93% type II) (Fig. 1a). In the OA group, 35% of atrophic fibres had a diameter of less than 30 mm (15.70% type I and 19.30% type II) (B–C).
Discussion This study was conducted to investigate the role of sarcopenia and the main age-related bone diseases, osteoporosis and osteoarthritis. To do this, a detailed clinical characterisation of patients was conducted by anamnesis, HHS, PASE test, BMI, DEXA and radiographic assessment by Kellgren–Lawrence. Although the protective role of fat tissues against osteoporosis is still controversial, the evidence that the OA patients in this study were frequently overweight supports this hypothesis [19,20]. As found in the literature, mean values for BMD and T-score were higher in OA patients compared with OP patients in this study; however, some patients showed different patterns of BMD and Tscore, which shows that OP and OA can co-exist in the same patient. According to our previous data, muscle morphometrical study demonstrated a higher percentage of atrophic fibres in OP patients compared with OA patients [21]. The OA patients were characterised particularly by a higher fat substitution of muscle fibres, which corresponded clinically to the
increased BMI in these patients. Recent studies have shown that in elderly patients, fat tissue induces sarcopenia through an inflammatory cytokine that inhibits muscle protein synthesis [22]. Moreover, the presence of adipose cells between muscle fibres could explain the molecular mechanisms of muscle metabolism (atrophy/regeneration) through the production of muscle growth factors (BMPs and myogenin) and muscle inhibitory molecules (TGF-b and myostatin). In this context, the immunohistochemical data from this study showed that muscle tissues of aged OP patients frequently express both myostatin and TGF-b next to infiltrating adipocyte. In detail, myostatin was shown to be more expressed in OP patients compared with OA patients. Conversely, the expression of TGF-b in muscle tissues seems to be linked to age rather than to bone diseases. The complexity of the TGF-b family signalling network is related to the different biological effects depending on the combination of ligands and receptors that are utilised [23]. Subsets of the TGF-b ligand family facilitate signal transduction via preferential recruitment of the Smad transcription regulators.
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Fig. 2. Immunohistochemical analysis. Immunostaining for myostatin (A) and TGF-b (B) was evaluated by counting the number of positive fibres on 25 high power field (HPF). Pax7 (C) and myogenin (D) were evaluated by counting the number of positive cells on 25 HPF.
In particular, myostatin and TGF-b proteins inhibit muscle growth by Smad2 and 3, whereas BMPs elicit muscle regeneration via Smad 1, 5 and 8 [23,24]. In this work, a correlation was demonstrated between BMP2 expression and muscle health. Indeed, BMP2 expression was noted in patients with low levels of fibre atrophy. With regard to the experimental groups, high levels of BMP2 were observed in muscle biopsies of OA patients. BMPs have recently been shown to induce adipocyte differentiation of mesenchymal stem cells [25]. In particular, high levels of BMPs induce the formation of brown adipose tissue, known as myogenic fat tissue, which secretes several factors (including myogenin) that trigger muscle regeneration. This phenomenon could initiate a circle in which BMPs are able to induce muscle regeneration by activating satellite cells and brown adipose tissue formation. To verify the possible relationship between BMPs, adipose tissues and muscle regeneration, the presence and activity of satellite muscle cells were investigated. Pax7 is a key transcriptional regulator expressed by quiescent satellite cells. This molecule is generally used to mark satellite cells in muscle tissues. In this study, OA patients were shown to preserve a higher spare of pax7-positive satellite cells compared with OP patients. Moreover, these cells appeared to spread
throughout the tissues. Data about satellite cells activity, obtained by myogenin expression analysis, further confirmed the potential ability of OA muscle to form new fibres. Indeed, there was a remarkable difference in myogenin expression between OP and OA groups. Notably, we did not observe myogenin-positive satellite cells associated to myofibres with a strong positivity for myostatin and vice versa. TEM was conducted to confirm this evidence. The ultrastructural analysis of OA tissues with a strong positivity for BMP2 and myogenin displayed the presence of a high rate of both single satellite cells and satellite cells forming syncytium. The finding of satellite cells syncytia is evidence of the muscle regeneration capability of OA muscle. Conversely, in muscle of OP patients with low BMP2, low myogenin expression and high myostatin expression, there were rare, sparse, single and not fused satellite cells. Moreover, these satellite cells showed marks of degeneration, such as nuclear anomalies and vacuolisations. These data clearly indicated that the spare of satellite cells and the presence of stimulating factor, such as BMP2 and myogenin, in OA patients slow the onset of age-related sarcopenia. There are also many other well-documented changes in muscle that occur with aging, including an overall decrease in myofibre size, and a decline in the number of excitable motor units within muscle [26]. There are also pronounced degradative changes in the
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Fig. 3. Satellite Stem Cells in muscle regeneration. A–B) Haematoxylin & eosin sections of muscle biopsies showed a significant increase in fat tissue in OA patients (A) compared to OP patients (B) (4). C) Image showing numerous myostatin-positive fibres (4). Myostatin expression was often not observed in OP patients (4) (D). The immunohistochemistry for TGF-b did not show a significant difference between OP and OA muscle tissue (40) (E–F). Immunohistochemical reaction displayed a significantly different rate of myogenin-positive cells in the OA group (G) compared to the OP group (H) (40) (arrows). Muscle biopsies of the OP group showed expression of TGF-b next to degenerative muscle tissue (asterisks) (40) (F) nuclear expression of pax7 was higher in muscle tissue of OA patients (I) compared with OP patients (J) (40).
neuromuscular junction with aging, which is known to contribute to synaptic loss and may be related to oxidative stress and mitochondrial dysfunction [27,37]. Aging is generally associated
with a preferential denervation of fast-twitch fibres along with reinnervation of some of those fibres by a-motorneurones from slow motor units [28].
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Fig. 4. Transmission electron microscopy analysis of muscle stem cells. TEM analysis was performed to identify satellite cells. In OA patients, fibres without degenerated markers were often observed (2.5000) (A). In these samples there were several satellite cells generally placed between normal and degenerated fibres (5.000 and 8000) (B–C). TEM analysis of muscle biopsies from OP patients showed numerous degeneration areas (asterisks) (2.500 and 5.000) (D–E). Rare satellite cells syncytia (arrow) with obvious mark of degeneration, such as nuclear anomalies and vacuolations, were found in OP patients (5.000) (F).
All of these changes will alter the contractile behaviour of aged muscle, which could be an important mechanistic link between muscle and bone formation (Fig. 4). One of the most exciting new developments in the area of muscle-bone interactions is the recognition that skeletal muscle secretes a variety of peptides, collectively termed myokines, and a number of these myokines are well recognised to have positive effects on bone formation, as well as inhibitory effects on osteoblast differentiation [8,9,29–31]. Importantly, muscle contraction stimulates the release of myokines in vivo and in vitro [32,33]. Insulin-like growth factor-1 (IGF1) expression in skeletal muscle tissue is elevated with muscle contraction, and circulating levels of IGF-1 are also increased with resistance exercise [34,35]. Aging has been shown to reduce the anabolic effects of resistance exercise on muscle protein synthesis and thus could potentially affect myokine synthesis and secretion [36]. Aged human myoblasts, for example, show increased levels of TGFb1 secretion in vitro compared with younger myoblasts, although it is not known how aging alters the secretion of other myokines with muscle contraction [37]. Moreover, it is well recognised that the bone marrow cavity accumulates fat with age, and it is certainly possible that increased bone marrow adipogenesis may attenuate the effects of circulating myokines on endocortical (endosteal) osteoprogenitor cells [38]. With increasing age, cortical thinning and trabecular loss may proceed at different rates and magnitude among various regions of the proximal femur; localised cortical thinning has been associated
with increased risk of hip fracture and trabecular loss has been found in hip fracture cases [39–43]. Conclusion Aging is associated with the development of osteoporosis in bone and sarcopenia in skeletal muscle; there are pronounced cellular and molecular changes in bone and muscle cells with age that likely underlie these observations. Such changes include the loss of osteocytes in bone matrix and a decline of osteoprogenitors in the periosteum, which attenuate the response of bone to muscle contraction and normal mechanical stimuli. Reduced fibre size, due to loss of motorneurones, decreased muscle protein synthesis and increased fatty infiltration in muscle with age also compromise the contractile function of aged muscle. Exercise could be a simple lifestyle approach that may produce localised adaptations in bone tissue, as seen in animal models [44,45]. Short bursts of regular hopping exercises have been shown to increase cortical mass surface density and endocortical trabecular density at the proximal femur, involving regions that may be important to structural integrity. According to the current study, physical activity should be strongly recommended in osteoporotic patients at diagnosis because it could be useful to delay muscle atrophy and bone loss as a result of the biomechanical stimuli the muscle exerts on bone. In particular, multi-task and balance exercises could improve fall-related self-efficacy, gait speed and balance performance, thereby reducing the risk of
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falling [46]. Exercise could then be a valid instrument to improve bone and muscle quality, and it could be a good strategy to reduce fracture risk and disability in the elderly population.
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The role of sarcopenia with and without fracture Umberto Tarantinoa,* , Jacopo Baldia,b , Manuel Scimecac,d , Eleonora Piccirillia,b , Andrea Picciolie, Elena Bonannoc , Elena Gasbarraa a
Department of Orthopaedics and Traumatology, “Tor Vergata” University of Rome, “Policlinico Tor Vergata” Foundation, Viale Oxford 1, 00133 Rome, Italy School of Specialisation in Orthopaedics and Traumatology, “Tor Vergata” University of Rome, “Policlinico Tor Vergata” Foundation, Viale Oxford 1, 00133 Rome, Italy c Anatomic Pathology Section, Department of Biomedicine and Prevention, University of Rome “Tor Vergata”, Via Montpellier 1, 00133 Rome, Italy d “Multidisciplinary Study of the Effects of Microgravity on Bone Cells” Project, Italian Space Agency (ASI), Spatial Biomedicine Center, Via del Politecnico snc, 00133 Rome, Italy e Oncologic Centre, “Palazzo Baleani”, Azienda Policlinico Umberto I, Corso Vittorio Emanuele II 244, Rome, Italy b
A R T I C L E I N F O
Keywords: Sarcopenia Osteoporosis Osteoarthritis Hip fracture Aging Risk of falling Muscle fibre atrophy Satellite cells BMP2 Myostatin
A B S T R A C T
Introduction: Bone and muscle tissues are in a close relationship. They are linked from a biological and functional point of view and both are related to an increased fracture risk in the elderly. The aging process is involved in the loss of functionality of both bones and muscles. In particular, aging-induced decline in muscle size and quality accompanies catabolic alterations in bone tissue; furthermore, age-related changes in bone alter its response to muscle-derived stimulation. The increased fracture risk in individuals with sarcopenia and osteoporosis is due to the decline of muscle mass and strength, the decrease in bone mineral density (BMD) and limited mobility. In this study, we investigated the role of sarcopenia and the main age-related bone diseases, osteoporosis (OP) and osteoarthritis (OA). Methods: Muscular performance status was evaluated using the Physical Activity Scale for the Elderly (PASE) test in 27 female patients with OP who underwent total hip arthroplasty for hip fracture, and in 27 age-matched female patients with OA who underwent total hip arthroplasty. Dual-energy X-ray absorptiometry (DEXA) was performed and the T-score values were used to discriminate between OP and OA patients. Body Mass Index (BMI) was calculated. As part of a multiparametric model of evaluation, biopsies of vastus lateralis muscle were analysed by immunohistochemical reaction to find a correlation with the above mentioned functional index. Results: The PASE test showed that the OP patients had a low or moderate level of physical activity before fracture occurred, whereas the OA patients had more intensive pre-fracture physical performances. Histological analysis showed that osteoporosis is characterised by a preferential type II fibre atrophy; in particular, data correlation showed that lower PASE test scores were related to lower diameter of type II fibres. No correlation was found between bone mineral density (BMD) and PASE test results. Discussion and conclusion: Osteoporosis is closely related to sarcopenia before and after fracture. Bone remodelling is influenced by muscle morphological and functional impairment and sarcopenia is considered one of the major factors for functional limitation and motor dependency in elderly osteoporotic individuals. Therefore, physical activity should be strongly recommended for OP patients at diagnosis. ã 2016 Elsevier Ltd. All rights reserved.
Background Bone and muscle are in a close relationship: when the aging process affects one of these two tissues, the functionality of the other is compromised. Consequently, sarcopenia and osteoporosis
* Corresponding author. E-mail address: [email protected] (U. Tarantino).
represent two pathologies that are frequently associated in the elderly. The term sarcopenia was first used by Rosenberg in 1989 to describe the decline in muscle mass among older people [1,2]. More recently, sarcopenia was defined as a loss of skeletal muscle mass and strength that occurs with aging [3]. Nowadays, the term sarcopenia is used in the literature to describe several pathophysiological processes, such as denervation, mitochondrial dysfunction, inflammatory and hormonal changes that may lead to a
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decrease in muscle strength and mobility, a decrease in fatigue resistance and an increase in the risk of falls and fractures [4]. Moreover, studies in the literature show that muscles of patients with sarcopenia display severe alterations in cellular turnover, including an increase in oxidative stress, cellular vacuolisation, and mitochondrial alterations, which compromise the quality of oxidant-scavenging systems. The denervation of single muscle fibres is also known to lead to a substantial reduction in type II fibres, which are gradually replaced by type I fibres and fat tissue [5,6]. Mechanisms involved in the decline of muscle mass during sarcopenia converge on the failure of satellite cells to replace and repair damaged muscle fibres. The main cause of reduced satellite cell function may be alteration of systemic factors that regulate satellite cell activity and differentiation. Indeed, satellite cells are mostly quiescent and their activation is governed by multiple niche factors (e.g. injury or stress) and signalling pathways, such as transforming growth factor-beta (TGF-b) and myogenin. The TGFb superfamily plays a crucial role in normal physiology and pathogenesis of skeletal muscle. Numerous members of the TGF-b family have been shown to play important roles in regulating muscle growth and atrophy. The most extensively characterised ligands, in terms of the effects on skeletal muscle, are TGF-b, myostatin and bone morphogenetic proteins (BMPs). Myogenin is a transcription factor that induces myogenesis in a variety of cell types in tissue culture. It is a member of the helix-loop-helix (HLH) protein family, a large family of proteins related by sequence homology, and it is essential for the development of functional skeletal muscle [7]. Muscle and bone are proportionally matched in their functional capacity and geometric structure; however, this relationship appears to change significantly with age. For example, the capacity for muscle to generate force declines with age, and the anabolic response of bone to muscle-derived strains also appears to be altered with age. In addition, muscle is now recognised to have paracrine and endocrine effects that may also influence bone independently of a mechanical relationship [8,9]. In this study, we investigated the role of sarcopenia and the main age-related bone diseases, osteoporosis (OP) and osteoarthritis (OA). Patients, materials and methods A total of 54 patients who underwent hip arthroplasty for femoral fractures or for hip OA in the Orthopaedic Department of “Tor Vergata” University from June 2014 to February 2015 were enrolled in this study. To evaluate bone mineral density (BMD), each patient with OA underwent dual-energy X-ray absorptiometry (DEXA) scan of the lumbar spine and femoral neck on the homolateral limb before surgery. Hip X-rays were performed to establish the grade of OA using the Kellgren-Lawrence Scale. To evaluate hip function, Harris Hip Score (HHS) was also calculated: the maximum score is 100 points, with the higher the HHS, the less the dysfunction. To estimate lumbar spine and non-fractured femur BMD, each OP patient underwent DEXA scan a few days after surgery; spinal Xrays were also performed in patients with femoral fracture or back pain to evaluate the presence of a vertebral compression fracture (VCF). A functional evaluation of performance status was conducted in each patient using the Physical Activity Scale for the Elderly (PASE) test. The questionnaire assesses two main groups of activities: recreational activities carried out during free time and domestic activities [10]. Each of these actions corresponds to a set value that is proportional to the “weight” that they could have in the life of each individual. The sum of the individual scores sets the patient
into one of four categories related to the degree of physical activity: inactivity (total score <42); lacking physical activity (43–105); moderate physical activity (106–145), and intense physical activity (>146). Body Mass Index (BMI) was also calculated. Exclusion criteria were: history of neoplastic diseases, myopathies or other neuromuscular diseases; taking anti-osteoporotic drugs or chronic administration (more than one month) of corticosteroid for autoimmune diseases; diabetes; alcohol abuse; cigarette smoking; and chronic viral infections (hepatitis B virus [HBV], hepatitis C virus [HCV], human immunodeficiency virus [HIV]). During surgery for total hip arthroplasty in both osteoarthritis (OA patients) and cervical femoral fragility fracture (OP patients), muscle biopsies were taken from the upper portion of the vastus lateralis muscle, which is normally involved in the surgical procedure, without damaging the tissue network. This muscle was chosen because it is hardly influenced by the fracture event and it is a good indicator of systemic muscle atrophy. All sampling and experiments were performed in agreement with the independent ethics committee, “Policlinico Tor Vergata”, approval reference number #85/12, and informed consent was obtained from all participants included in the study. Bone mineral density evaluation (DEXA) DEXA was performed with a Lunar DEXA apparatus (GE Healthcare, Madison, WI, USA). Lumbar spine (L1–L4) and femoral (neck and total) scans were performed, and BMD was analysed according to the manufacturer’s recommendations. DEXA measures BMD (in g/cm2) with a coefficient of variation of 0.7%. In the OA group, all measurements were performed on the non-dominant side; in the OP group, BMD was measured on the limb opposite to the fracture side. Results were expressed as absolute values and as T-scores [11,12]. Histology Muscle biopsies were fixed with 4% paraformaldehyde for 24 h and embedded in paraffin after alcoholic dehydration. Sections of 3 mm thick were stained with haematoxylin and eosin (H&E) and the pathological evaluation was performed by two pathologists who were blinded to the samples [13]. Atrophy assessment A minimum of 200 muscle fibres per biopsy were evaluated to assess fibre atrophy; minimum transverse diameter and crosssectional area of type I and type II fibres were compared for relative prevalence. A threshold diameter of less than 30 mm (minimum value of the normal range for women) characterised atrophic fibres [14,15]. To calculate muscle and bone areas, H&E slides were scanned at low-power field by Iscan Coreo (Ventana, Tucson, AZ, USA). Areas for each muscle and bone biopsy image were identified by a pathologist using Viewing software (Ventana, Tucson, AZ, USA). Immunohistochemistry Immunohistochemical characterisation was conducted to assess muscle fibre type (i.e. fast and slow) and the expression of myostatin, TGF-b, pax7 and myogenin. Briefly, 3 mm thick sections were pre-treated with EDTA citrate (pH 7.8) for 30 min at 95 C and then incubated, respectively, with mouse monoclonal anti-fast skeletal myosin for 60 min (1:100, clone MY-32, AbCam), mouse monoclonal anti-slow skeletal myosin for 60 min (1:100, clone NOQ7.5.4D, AbCam) and rabbit
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monoclonal anti-myostatin for 45 min (1:100, clone ab134682, AbCam). For myogenin, TGF-b and pax7, 3 mm thick sections were pre-treated with citrate (pH 6.0) for 30 min at 95 C and then incubated with mouse monoclonal anti-myogenin (1:100, clone F5D, AbCam), rabbit monoclonal anti-TGF-b for 60 min (1:250 clone 6B7, Novus Biologicals) and rabbit monoclonal anti-pax7 (1:500, NC, Novus Biologicals) [16]. A washing buffer composed of PBS 4% and Tween20 (pH 7.6) (UCS diagnostic, Rome, Italy) was used; reactions were revealed by HRP-DAB Detection Kit (UCS Diagnostic, Rome, Italy). Transmission electron microscopy (TEM) Small samples of muscle tissue from each patient were processed as previously described [17]. Briefly, muscle tissues were fixed in 4% paraformaldehyde, post-fixed in 2% osmium tetroxide and dehydrated by a series of incubations in 30%, 50% and 70%, ethanol. Samples were then embedded in EPON resin (Agar Scientific, Stansted, Essex, CM24 8GF, United Kingdom), for morphological ultrastructural analysis, and in LR-White resin (Agar Scientific, Stansted Essex, CM24 8GF, United Kingdom) for ultrastructural immunohistochemistry [18]. Tissues were cut and stained with heavy metal solutions, as described by Reynolds [17,18]. Statistical analysis Statistical analysis was performed using GraphPad Prism 5 Software (La Jolla, CA, USA). Morphometric data were expressed as percentage of atrophic fibres and percentage of tissue composition of muscle fasciculus and bone. The comparison of myostatin, TGF-b, BMP2, myogenin and pax7 expression between the two experimental groups was performed using the Mann– Whitney test. The difference between groups was considered statistically significant at P < 0.05. Immunohistochemistry data were expressed as mean value with standard deviation (n SD). Results Clinical evaluation Clinical evaluation was conducted and patients were divided into two groups: OP and OA. The OP group (femoral neck BMD range 0.454–0.645 g/cm2) included 27 females with fragility hip fracture, a T-score -2.5 SD and a negative radiographic framework for hip OA; the OA group (femoral neck BMD range 0.845–1.197 g/ cm2) included 27 females with a positive radiogram for hip OA with a Kellgren–Lawrence score of 3 or 4 and a T-score -2.5 SD (Table 1). The mean BMI of the OA group was statistically significantly higher than that of the OP group (26.88 0.85 kg/m2 vs 22.73 3.54 kg/m2, P < 0.001). These data confirm the frequently overweight condition of OA patients. The clinical results in this
Table 1 Evaluated parameters in patients with osteoarthritis (OA) and osteoporosis (OP). Variables
OA Group
OP Group
Age (years) Body Mass Index (kg/m2) PASE test Bone Mineral Density L1-L4 (g/m2) T-score L1-L4 Femoral neck Bone Mineral Density (g/m2) Femoral neck T-score Kellgren-Lawrence grade
71.6 10.3 26.88 0.85 100 20.86 0.957 0.20 1.8 1.66 0.845–1.197 1.4 0.94 3–4
82.4 6.19 22.73 3.54 62.6 33 0.927 0.18 2.3 1.08 0.454–0.645 2.92 1.12 0–1
3
study showed an inverse correlation between BMI and HHS in the OA group. Indeed, a high body mass impairs hip joint mobility. The results of the PASE test showed that both OP and OA patients were characterised by poor physical activity; however, patients in the OP group had a lower PASE test score than those in the OA group. BMD, T-score and Kellgren–Lawrence grading scale were used for patients’ instrumental evaluation. OA patients were characterised by physiological age-related osteopenia limited to the lumbar region (T-score 1 <> 2.5 SD) and generally higher values of BMD compared to the OP group (Table 1). Conversely, all OP patients showed low levels of BMD and a T-score <-2.5 SD. Radiological analysis enabled classification of the OA patients, who were at least grade 3 on the Kellgren-Lawrence scale, whereas OP patients showed 0 or 1 Kellgren-Lawrence grade (Table 1). Morphometric analysis Muscle morphometric examination performed on digital scanned images showed a reduction in muscle tissues of about 20% in the OA and OP groups (Fig. 1a–b). In OA patients, muscle was mainly replaced by adipose tissue (17.93%), whereas in OP muscle fasciculus, atrophic muscle fibres were substituted by a mixture of adipose (8.01%) and connective tissue (6.31%) (Fig. 1a–b). Slow myosin antibody and fast myosin antibody stains enabled discrimination between type I and type II fibres, respectively. The morphometric analysis of muscle fibres in OP patients showed more than 50% of atrophic fibres with prevalence of type II fibres (19.13% type I and 35.93% type II) (Fig. 1a). In the OA group, there were 35% of atrophic fibres with a diameter of less than 30 mm (15.70% type I and 19.30% type II) (Fig. 1b). Immunohistochemistry study Myostatin and TGF-b expression were assessed by counting the number of positive fibres on 25 high-power field (HPF) (Fig. 2a–b). Remarkably, OP muscle biopsies showed a significantly higher number of myostatin-positive fibres (40.25 5.70) compared with the muscle of OA patients (15.36 2.45) (Figs. 2a and 3a–d). Conversely, there were no significant differences between the number of TGF-b-positive fibres among OA and OP groups (Figs. 2b and 3e–f). Nonetheless, TGF-b expression tends to increase in the elderly. The presence and activity of satellite cells were studied using pax7 and myogenin expression. Their expression was evaluated by counting the number of positive satellite cells on 25 HPF (Fig. 2c–d). Nuclear expression of pax7 was higher in muscle biopsy of OA patients compared with that of OP patients (OA 33.44 2.48 vs OP 18.74 1.84) (Figs. 2c and 3g–h). Notably, in OA muscle tissue, it was frequently observed that pax7-positive cells were aggregated to form syncytia. Finally, our results showed a significantly different rate of myogenin-positive cells in the OA group compared with the OP group (OA 250.30 89.65 vs OP 150.60 52.53) (Fig. 2d). In particular, in OA patients, many groups of myogenin-positive satellite cells were focally observed in the tissue (Fig. 3i–j). Transmission electron microscopy TEM analysis was performed to identify satellite cells according to their ultrastructural characteristics. In OA muscle biopsies, several satellite cells were found tightly associated or fused to form a syncytium. Notably, in the cytoplasm of these cells there was a de novo production of sarcomeric structures (Fig. 4a–c). In OP patients, the number of satellite cells was very low and their niches showed an obvious mark of degeneration (Fig. 4d–f).
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Fig. 1. Morphometric analysis. In OA patients, muscle was mainly replaced by adipose tissue (17.93%), whereas in OP muscle fasciculus, atrophic muscle fibres were substituted by a mixture of adipose (8.01%) and connective tissue (6.31%) (A–B). The morphometric analysis of muscle fibres in OP patients showed more than 50% of atrophic fibres with prevalence of type II fibres (19.13% type I and 35.93% type II) (Fig. 1a). In the OA group, 35% of atrophic fibres had a diameter of less than 30 mm (15.70% type I and 19.30% type II) (B–C).
Discussion This study was conducted to investigate the role of sarcopenia and the main age-related bone diseases, osteoporosis and osteoarthritis. To do this, a detailed clinical characterisation of patients was conducted by anamnesis, HHS, PASE test, BMI, DEXA and radiographic assessment by Kellgren–Lawrence. Although the protective role of fat tissues against osteoporosis is still controversial, the evidence that the OA patients in this study were frequently overweight supports this hypothesis [19,20]. As found in the literature, mean values for BMD and T-score were higher in OA patients compared with OP patients in this study; however, some patients showed different patterns of BMD and Tscore, which shows that OP and OA can co-exist in the same patient. According to our previous data, muscle morphometrical study demonstrated a higher percentage of atrophic fibres in OP patients compared with OA patients [21]. The OA patients were characterised particularly by a higher fat substitution of muscle fibres, which corresponded clinically to the
increased BMI in these patients. Recent studies have shown that in elderly patients, fat tissue induces sarcopenia through an inflammatory cytokine that inhibits muscle protein synthesis [22]. Moreover, the presence of adipose cells between muscle fibres could explain the molecular mechanisms of muscle metabolism (atrophy/regeneration) through the production of muscle growth factors (BMPs and myogenin) and muscle inhibitory molecules (TGF-b and myostatin). In this context, the immunohistochemical data from this study showed that muscle tissues of aged OP patients frequently express both myostatin and TGF-b next to infiltrating adipocyte. In detail, myostatin was shown to be more expressed in OP patients compared with OA patients. Conversely, the expression of TGF-b in muscle tissues seems to be linked to age rather than to bone diseases. The complexity of the TGF-b family signalling network is related to the different biological effects depending on the combination of ligands and receptors that are utilised [23]. Subsets of the TGF-b ligand family facilitate signal transduction via preferential recruitment of the Smad transcription regulators.
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Fig. 2. Immunohistochemical analysis. Immunostaining for myostatin (A) and TGF-b (B) was evaluated by counting the number of positive fibres on 25 high power field (HPF). Pax7 (C) and myogenin (D) were evaluated by counting the number of positive cells on 25 HPF.
In particular, myostatin and TGF-b proteins inhibit muscle growth by Smad2 and 3, whereas BMPs elicit muscle regeneration via Smad 1, 5 and 8 [23,24]. In this work, a correlation was demonstrated between BMP2 expression and muscle health. Indeed, BMP2 expression was noted in patients with low levels of fibre atrophy. With regard to the experimental groups, high levels of BMP2 were observed in muscle biopsies of OA patients. BMPs have recently been shown to induce adipocyte differentiation of mesenchymal stem cells [25]. In particular, high levels of BMPs induce the formation of brown adipose tissue, known as myogenic fat tissue, which secretes several factors (including myogenin) that trigger muscle regeneration. This phenomenon could initiate a circle in which BMPs are able to induce muscle regeneration by activating satellite cells and brown adipose tissue formation. To verify the possible relationship between BMPs, adipose tissues and muscle regeneration, the presence and activity of satellite muscle cells were investigated. Pax7 is a key transcriptional regulator expressed by quiescent satellite cells. This molecule is generally used to mark satellite cells in muscle tissues. In this study, OA patients were shown to preserve a higher spare of pax7-positive satellite cells compared with OP patients. Moreover, these cells appeared to spread
throughout the tissues. Data about satellite cells activity, obtained by myogenin expression analysis, further confirmed the potential ability of OA muscle to form new fibres. Indeed, there was a remarkable difference in myogenin expression between OP and OA groups. Notably, we did not observe myogenin-positive satellite cells associated to myofibres with a strong positivity for myostatin and vice versa. TEM was conducted to confirm this evidence. The ultrastructural analysis of OA tissues with a strong positivity for BMP2 and myogenin displayed the presence of a high rate of both single satellite cells and satellite cells forming syncytium. The finding of satellite cells syncytia is evidence of the muscle regeneration capability of OA muscle. Conversely, in muscle of OP patients with low BMP2, low myogenin expression and high myostatin expression, there were rare, sparse, single and not fused satellite cells. Moreover, these satellite cells showed marks of degeneration, such as nuclear anomalies and vacuolisations. These data clearly indicated that the spare of satellite cells and the presence of stimulating factor, such as BMP2 and myogenin, in OA patients slow the onset of age-related sarcopenia. There are also many other well-documented changes in muscle that occur with aging, including an overall decrease in myofibre size, and a decline in the number of excitable motor units within muscle [26]. There are also pronounced degradative changes in the
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Fig. 3. Satellite Stem Cells in muscle regeneration. A–B) Haematoxylin & eosin sections of muscle biopsies showed a significant increase in fat tissue in OA patients (A) compared to OP patients (B) (4). C) Image showing numerous myostatin-positive fibres (4). Myostatin expression was often not observed in OP patients (4) (D). The immunohistochemistry for TGF-b did not show a significant difference between OP and OA muscle tissue (40) (E–F). Immunohistochemical reaction displayed a significantly different rate of myogenin-positive cells in the OA group (G) compared to the OP group (H) (40) (arrows). Muscle biopsies of the OP group showed expression of TGF-b next to degenerative muscle tissue (asterisks) (40) (F) nuclear expression of pax7 was higher in muscle tissue of OA patients (I) compared with OP patients (J) (40).
neuromuscular junction with aging, which is known to contribute to synaptic loss and may be related to oxidative stress and mitochondrial dysfunction [27,37]. Aging is generally associated
with a preferential denervation of fast-twitch fibres along with reinnervation of some of those fibres by a-motorneurones from slow motor units [28].
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Fig. 4. Transmission electron microscopy analysis of muscle stem cells. TEM analysis was performed to identify satellite cells. In OA patients, fibres without degenerated markers were often observed (2.5000) (A). In these samples there were several satellite cells generally placed between normal and degenerated fibres (5.000 and 8000) (B–C). TEM analysis of muscle biopsies from OP patients showed numerous degeneration areas (asterisks) (2.500 and 5.000) (D–E). Rare satellite cells syncytia (arrow) with obvious mark of degeneration, such as nuclear anomalies and vacuolations, were found in OP patients (5.000) (F).
All of these changes will alter the contractile behaviour of aged muscle, which could be an important mechanistic link between muscle and bone formation (Fig. 4). One of the most exciting new developments in the area of muscle-bone interactions is the recognition that skeletal muscle secretes a variety of peptides, collectively termed myokines, and a number of these myokines are well recognised to have positive effects on bone formation, as well as inhibitory effects on osteoblast differentiation [8,9,29–31]. Importantly, muscle contraction stimulates the release of myokines in vivo and in vitro [32,33]. Insulin-like growth factor-1 (IGF1) expression in skeletal muscle tissue is elevated with muscle contraction, and circulating levels of IGF-1 are also increased with resistance exercise [34,35]. Aging has been shown to reduce the anabolic effects of resistance exercise on muscle protein synthesis and thus could potentially affect myokine synthesis and secretion [36]. Aged human myoblasts, for example, show increased levels of TGFb1 secretion in vitro compared with younger myoblasts, although it is not known how aging alters the secretion of other myokines with muscle contraction [37]. Moreover, it is well recognised that the bone marrow cavity accumulates fat with age, and it is certainly possible that increased bone marrow adipogenesis may attenuate the effects of circulating myokines on endocortical (endosteal) osteoprogenitor cells [38]. With increasing age, cortical thinning and trabecular loss may proceed at different rates and magnitude among various regions of the proximal femur; localised cortical thinning has been associated
with increased risk of hip fracture and trabecular loss has been found in hip fracture cases [39–43]. Conclusion Aging is associated with the development of osteoporosis in bone and sarcopenia in skeletal muscle; there are pronounced cellular and molecular changes in bone and muscle cells with age that likely underlie these observations. Such changes include the loss of osteocytes in bone matrix and a decline of osteoprogenitors in the periosteum, which attenuate the response of bone to muscle contraction and normal mechanical stimuli. Reduced fibre size, due to loss of motorneurones, decreased muscle protein synthesis and increased fatty infiltration in muscle with age also compromise the contractile function of aged muscle. Exercise could be a simple lifestyle approach that may produce localised adaptations in bone tissue, as seen in animal models [44,45]. Short bursts of regular hopping exercises have been shown to increase cortical mass surface density and endocortical trabecular density at the proximal femur, involving regions that may be important to structural integrity. According to the current study, physical activity should be strongly recommended in osteoporotic patients at diagnosis because it could be useful to delay muscle atrophy and bone loss as a result of the biomechanical stimuli the muscle exerts on bone. In particular, multi-task and balance exercises could improve fall-related self-efficacy, gait speed and balance performance, thereby reducing the risk of
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falling [46]. Exercise could then be a valid instrument to improve bone and muscle quality, and it could be a good strategy to reduce fracture risk and disability in the elderly population.
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Please cite this article in press as: U. Tarantino, et al., The role of sarcopenia with and without fracture, Injury (2016), http://dx.doi.org/10.1016/j. injury.2016.07.057