Myotube formation is affected by adipogenic lineage cells in a cell-to-cell contact-independent manner

Intramuscular adipose tissue (IMAT) formation is observed in some pathological conditions such as Duchenne muscular dystrophy (DMD) and sarcopenia. Several studies have suggested that IMAT formation is not only negatively correlated with skeletal muscle mass but also causes decreased muscle contraction in sarcopenia. In the present study, we examined w hether adipocytes affect myogenesis. For this purpose, skeletal muscle progenitor cells were transfected with siRNA of PPARγ (siPPARγ) in an attempt to inhibit adipogenesis. Myosin heavy chain (MHC)-positive myotube formation was promoted in cells transfected with siPPARγ compared to that of cells transfected with control siRNA. To determine whether direct cell-to-cell contact between adipocytes and myoblasts is a prerequisite for adipocytes to affect myogenesis, skeletal muscle progenitor cells were cocultured with pre- or mature adipocytes in a Transwell coculture system. MHC-positive myotube formation was inhibited when skeletal muscle progenitor cells were cocultured with mature adipocytes, but was promoted when they were cocultured with preadipocytes. Similar effects were observed when pre- or mature adipocyte-conditioned medium was used. These results indicate that preadipocytes play an important role in maintaining skeletal muscle mass by promoting myogenesis; once differentiated, the resulting mature adipocytes negatively affect myogenesis, leading to the muscle deterioration observed in skeletal muscle pathologies.     

Let-7 Represses Carcinogenesis and a Stem Cell Phenotype in the Intestine via Regulation of Hmga2

Let-7 miRNAs comprise one of the largest and most highly expressed family of miRNAs among vertebrates, and is critical for promoting differentiation, regulating metabolism, inhibiting cellular proliferation, and repressing carcinogenesis in a variety of tissues. The large size of the Let-7 family of miRNAs has complicated the development of mutant animal models. Here we describe the comprehensive repression of all Let-7 miRNAs in the intestinal epithelium via low-level tissue-specific expression of the Lin28b RNA-binding protein and a conditional knockout of the MirLet7c-2/Mirlet7b locus. This ablation of Let-7 triggers the development of intestinal adenocarcinomas concomitant with reduced survival. Analysis of both mouse and human intestinal cancer specimens reveals that stem cell markers were significantly associated with loss of Let-7 miRNA expression, and that a number of Let-7 targets were elevated, including Hmga1 and Hmga2. Functional studies in 3-D enteroids revealed that Hmga2 is necessary and sufficient to mediate many characteristics of Let-7 depletion, namely accelerating cell cycle progression and enhancing a stem cell phenotype. In addition, inactivation of a single Hmga2 allele in the mouse intestine epithelium significantly represses tumorigenesis driven by Lin28b. In aggregate, we conclude that Let-7 depletion drives a stem cell phenotype and the development of intestinal cancer, primarily via Hmga2.     

Skeletal Muscle Regeneration by the Exosomes of Adipose Tissue-Derived Mesenchymal Stem Cells

Profound skeletal muscle loss can lead to severe disability and cosmetic deformities. Mesenchymal stem cell (MSC)-derived exosomes have shown potential as an effective therapeutic tool for tissue regeneration. This study aimed to determine the regenerative capacity of MSC-derived exosomes for skeletal muscle regeneration. Exosomes were isolated from human adipose tissue-derived MSCs (AD-MSCs). The effects of MSC-derived exosomes on satellite cells were investigated using cell viability, relevant genes, and protein analyses. Moreover, NOD-SCID mice were used and randomly assigned to the healthy control (n = 4), muscle defect (n = 6), and muscle defect + exosome (n = 6) groups. Muscle defects were created using a biopsy punch on the quadriceps of the hind limb. Four weeks after the surgery, the quadriceps muscles were harvested, weighed, and histologically analyzed. MSC-derived exosome treatment increased the proliferation and expression of myocyte-related genes, and immunofluorescence analysis for myogenin revealed a similar trend. Histologically, MSC-derived exosome-treated mice showed relatively preserved shapes and sizes of the muscle bundles. Immunohistochemical staining revealed greater expression of myogenin and myoblast determination protein 1 in the MSC-derived exosome-treated group. These results indicate that exosomes extracted from AD-MSCs have the therapeutic potential for skeletal muscle regeneration.     

Role of cardiac progenitor cell‐derived exosome‐mediated microRNA‐210 in cardiovascular disease

Cardiac progenitor cells are considered to be one of the most promising stem cells for heart regeneration and repair. The cardiac protective effect of CPCs is mainly achieved by reducing tissue damage and/or promoting tissue repair through a paracrine mechanism. Exosome is a factor that plays a major role in the paracrine effect of CPCs. By delivering microRNAs to target cells and regulating their functions, exosomes have shown significant beneficial effects in slowing down cardiac injury and promoting cardiac repair. Among them, miRNA‐210 is an important anoxic‐related miRNA derived from CPCs exosomes, which has great cardiac protective effect of inhibiting myocardial cell apoptosis, promoting angiogenesis and improving cardiac function. In addition, circulating miR‐210 may be a useful biomarker for the prediction or diagnosis of related cardiovascular diseases. In this review, we briefly reviewed the mechanism of miR‐210 derived from CPCs exosomes in cardiac protection in recent years.     

Neuronal Differentiation of Human Mesenchymal Stem Cells Using Exosomes Derived from Differentiating Neuronal Cells

Exosomes deliver functional proteins and genetic materials to neighboring cells, and have potential applications for tissue regeneration. One possible mechanism of exosome-promoted tissue regeneration is through the delivery of microRNA (miRNA). In this study, we hypothesized that exosomes derived from neuronal progenitor cells contain miRNAs that promote neuronal differentiation. We treated mesenchymal stem cells (MSCs) daily with exosomes derived from PC12 cells, a neuronal cell line, for 1 week. After the treatment with PC12-derived exosomes, MSCs developed neuron-like morphology, and gene and protein expressions of neuronal markers were upregulated. Microarray analysis showed that the expression of miR-125b, which is known to play a role in neuronal differentiation of stem cells, was much higher in PC12-derived exosomes than in exosomes from B16-F10 melanoma cells. These results suggest that the delivery of miRNAs contained in PC12-derived exosomes is a possible mechanism explaining the neuronal differentiation of MSC.     

Therapeutic Potential of Human Adipose-Derived Stem Cells (ADSCs) from Cancer Patients: A Pilot Study

Mesenchymal stem cells from adipose tissue (ADSCs) are an important source of cells for regenerative medicine. The therapeutic effect of culture-expanded adipose derived stem cells has been shown; however, optimal xeno-free culture conditions remain to be determined. Cancer patients, specifically those undergoing invasive surgery, constitute a subgroup of patients who could benefit from autologous stem cell transplantation. Although regenerative potential of their ADSCs could be affected by the disease and/or treatment, we are not aware of any study that has evaluated the therapeutic potential of ADSCs isolated from cancer patients in reference to that of ADSCs derived from healthy subjects. Here we report that ADSCs isolated from subabdominal adipose tissue of patients with urological neoplasms yielded similar growth kinetics, presented equivalent mesenchymal surface markers and showed similar differentiation potential into distinct mesodermal cell lineages: adipocytes, chondroblasts and osteoblasts than ADSCs isolated from adipose tissue of age-matched non-oncogenic participants, all under xeno-free growth culture conditions. Molecular karyotyping of patient expanded ADSCs genomes showed no disease-related alterations indicating their safety. In addition, vesicles <100 nm identified as exosomes (EXOs) which may be at least partly responsible for the attributed therapeutic paracrine effects of the ADSCs were effectively isolated from ADSCs and showed equivalent miRNA content regardless they were derived from cancer patients or non-oncogenic participants indicating that the repair capabilities of xeno-free expanded ADSCs are not compromised by patient condition and therefore their xeno-free culture expanded ADSCs should be suitable for autologous stem cell transplantation in a clinical setting.     

Sarcopenia Is Independently Associated with Cardiovascular Disease in Older Korean Adults: The Korea National Health and Nutrition Examination Survey (KNHANES) from 2009

Background

The association between sarcopenia and cardiovascular disease (CVD) in elderly people has not been adequately assessed. The aim of this study was to investigate whether CVD is more prevalent in subjects with sarcopenia independent of other well-established cardiovascular risk factors in older Korean adults.

Method

This study utilized the representative Korean population data from the Korea National Health and Nutrition Examination Survey (KNHANES) which was conducted in 2009. Subjects older than 65 years of age with appendicular skeletal muscle mass (ASM) determined by dual energy X-ray absorptiometry were selected. The prevalence of sarcopenia in the older Korean adults was investigated, and it was determined whether sarcopenia is associated with CVD independent of other well-known risk factors.

Results

1,578 subjects aged 65 years and older with the data for ASM were selected, and the overall prevalence of sarcopenia was 30.3% in men and 29.3% in women. Most of the risk factors for CVD such as age, waist circumference, body mass index, fasting plasma glucose and total cholesterol showed significant negative correlations with the ratio between appendicular skeletal muscle mass and body weight. Multiple logistic regression analysis demonstrated that sarcopenia was associated with CVD independent of other well-documented risk factors, renal function and medications (OR, 1.768; 95% CI, 1.075–2.909, P = 0.025).

Conclusions

Sarcopenia was associated with the presence of CVD independent of other cardiovascular risk factors after adjusting renal function and medications.     

Aging differentially alters the expression of angiogenic genes in a tissue-dependent manner

Organ functions are altered and impaired during aging, thereby resulting in increased morbidity of age-related diseases such as Alzheimer’s disease, diabetes, and heart failure in the elderly. Angiogenesis plays a crucial role in the maintenance of tissue homeostasis, and aging is known to reduce the angiogenic capacity in many tissues. Here, we report the differential effects of aging on the expression of angiogenic factors in different tissues, representing a potentially causes for age-related metabolic disorders. PCR-array analysis revealed that many of angiogenic genes were down-regulated in the white adipose tissue (WAT) of aged mice, whereas they were largely up-regulated in the skeletal muscle (SM) of aged mice compared to that in young mice. Consistently, blood vessel density was substantially reduced and hypoxia was exacerbated in WAT of aged mice compared to that in young mice. In contrast, blood vessel density in SM of aged mice was well preserved and was not different from that in young mice. Moreover, we identified that endoplasmic reticulum (ER) stress was strongly induced in both WAT and SM during aging in vivo. We also found that ER stress significantly reduced the expression of angiogenic genes in 3T3-L1 adipocytes, whereas it increased their expression in C2C12 myotubes in vitro. These results collectively indicate that aging differentially affects the expression of angiogenic genes in different tissues, and that aging-associated down-regulation of angiogenic genes in WAT, at least in part through ER stress, is potentially involved in the age-related adipose tissue dysfunction.     

Metabolic Syndrome,Sarcopenia and Role of Sex and Age: Cross-Sectional Analysis of Kashiwa Cohort Study

Recent epidemiological evidence suggests that effects of cardiovascular risk factors may vary depending on sex and age. In this study, we assessed the associations of metabolic syndrome (MetS) with sarcopenia and its components in older adults, and examined whether the associations vary by sex and age. We also tested if any one of the MetS components could explain the associations. We conducted a cross-sectional analysis of the baseline data from the cohort study conducted in Kashiwa city, Chiba, Japan in 2012 which included 1971 functionally-independent, community-dwelling Japanese adults aged 65 years or older (977 men, 994 women). Sarcopenia was defined based on appendicular skeletal muscle mass, grip strength and usual gait speed. MetS was defined based on the National Cholesterol Education Program’s Adult Treatment Panel-III criteria. The prevalence of sarcopenia was 14.2% in men and 22.1% in women, while the prevalence of MetS was 43.6% in men and 28.9% in women. After adjustment for potential confounders, MetS was positively associated with sarcopenia in men aged 65 to 74 years (odds ratio 5.5; 95% confidence interval 1.9–15.9) but not in older men or women. Among the sarcopenia components, MetS was associated with lower muscle mass and grip strength, particularly in men aged 65 to 74 years. The associations of MetS with sarcopenia and its components were mainly driven by abdominal obesity regardless of sex or age. In conclusion, MetS is positively associated with sarcopenia in older men. The association is modified by sex and age, but abdominal obesity is the main contributor to the association across sex and age.     

Deep Sequencing of RNA from Three Different Extracellular Vesicle (EV) Subtypes Released from the Human LIM1863 Colon Cancer Cell Line Uncovers Distinct Mirna-Enrichment Signatures

Secreted microRNAs (miRNAs) enclosed within extracellular vesicles (EVs) play a pivotal role in intercellular communication by regulating recipient cell gene expression and affecting target cell function. Here, we report the isolation of three distinct EV subtypes from the human colon carcinoma cell line LIM1863 – shed microvesicles (sMVs) and two exosome populations (immunoaffinity isolated A33-exosomes and EpCAM-exosomes). Deep sequencing of miRNA libraries prepared from parental LIM1863 cells/derived EV subtype RNA yielded 254 miRNA identifications, of which 63 are selectively enriched in the EVs - miR-19a/b-3p, miR-378a/c/d, and miR-577 and members of the let-7 and miR-8 families being the most prominent. Let-7a-3p*, let-7f-1-3p*, miR-451a, miR-574-5p*, miR-4454 and miR-7641 are common to all EV subtypes, and 6 miRNAs (miR-320a/b/c/d, miR-221-3p, and miR-200c-3p) discern LIM1863 exosomes from sMVs; miR-98-5p was selectively represented only in sMVs. Notably, A33-Exos contained the largest number (32) of exclusively-enriched miRNAs; 14 of these miRNAs have not been reported in the context of CRC tissue/biofluid analyses and warrant further examination as potential diagnostic markers of CRC. Surprisingly, miRNA passenger strands (star miRNAs) for miR-3613-3p*, -362-3p*, -625-3p*, -6842-3p* were the dominant strand in A33-Exos, the converse to that observed in parental cells. This finding suggests miRNA biogenesis may be interlinked with endosomal/exosomal processing.     

Sarcopenia is not due to lack of regenerative drive in senescent skeletal muscle

Sarcopenia, loss of skeletal muscle mass, is a hallmark of aging commonly attributed to a decreased capacity to maintain muscle tissue in senescence, yet the mechanism behind the muscle wasting remains unresolved. To address these issues we have explored a rodent model of sarcopenia and age-related sensorimotor impairment, allowing us to discriminate between successfully and unsuccessfully aged cohort members. Immunohistochemistry and staining of cell nuclei revealed that senescent muscle has an increased density of cell nuclei, occurrence of aberrant fibers and fibers expressing embryonic myosin. Using real-time PCR we extend the findings of increased myogenic regulatory factor mRNA to show that very high levels are found in unsuccessfully aged cohort members. This pattern is also reflected in the number of embryonic myosin-positive fibers, which increase with the degree of sarcopenia. In addition, we confirm that there is no local down-regulation of IGF-I and IGF-IR mRNA in aged muscle tissue; on the contrary, the most sarcopenic individuals showed significantly higher local expression of IGF-I mRNA. Combined, our results show that the initial drive to regenerate myofibers is most marked in cases with the most advanced loss of muscle mass, a pattern that may have its origin in differences in the rate of tissue deterioration and/or that regenerating myofibers in these cases fail to mature into functional fibers. Importantly, the genetic background is a determinant of the pace of progression of sarcopenia.     

Platelet microparticle delivered microRNA-Let-7a promotes the angiogenic switch

Platelet microparticle (PMP)-induced angiogenesis plays a key role in tumour metastasis and has been proposed to contribute towards cardiovascular disease by enhancing atherosclerotic plaque vulnerability. However, the mechanisms underlying PMP induced angiogenesis are ill defined. Recent reports demonstrate that PMPs deliver micro-RNAs (miRNAs) to recipient cells, controlling gene expression. We therefore evaluated whether miRNA transfer was a key regulator of PMP-induced angiogenesis. Co-culturing PMPs with human umbilical vein endothelial cells (HUVEC) on extracellular matrix gel induced robust capillary like structure formation. PMP treatment altered the release of angiogenesis modulators from HUVEC, including significantly reducing production of anti-angiogenic thrombospondin-1 (THBS-1). Both functional responses were abrogated by treating PMPs with RNase, suggesting the transfer of PMP-derived RNA was a critical event. PMPs were an abundant source of miRNA Let-7a, which was transferred to HUVEC following co-incubation. Using luciferase reporter assays we have shown that Let-7a directly targets the 3’UTR of the THBS-1 mRNA. HUVEC transfection with a Let-7a anti-sense oligonucleotide reduced the ability of PMPs to inhibit THBS-1 release, and significantly decreased PMP induced in vitro angiogenesis. Antibody neutralisation of THBS-1 reversed the anti-angiogenic effect of let-7a inhibition in PMP treated HUVEC, highlighting Let-7a dependent translational repression of THBS-1 drives angiogenesis. Importantly, plasmid overexpression of Let-7a in HUVEC alone induced robust tubule formation on extracellular matrix gel. These data reveal a new role for Let-7a in promoting angiogenesis and show for the first time PMPs induced angiogenic responses occur through miRNA regulation of HUVEC.     

Prevalence of Sarcopenia and Its Relationship with Sites of Fragility Fractures in Elderly Chinese Men and Women

Objective

Sarcopenia might be associated with bone fragility in elderly individuals. This study aimed to investigate the prevalence of sarcopenia and its association with fragility fracture sites in elderly Chinese patients.

Methods

Patients (322 men and 435 women) aged 65–94 years and with a history of fragility fractures in the ankle, wrist, vertebrae or hip, and healthy men (n = 1263) and women (n = 1057) aged 65–92 years without a history of fractures were enrolled. Whole-body dual energy X-ray absorptiometry was used to analyze skeletal muscle mass index (SMI), fat mass and bone mineral density. Sarcopenia was defined as SMI less than two standard deviations below the mean of a young reference group.

Results

Sarcopenia occurrence varied with fracture location. Sarcopenia was more common in females with vertebral and hip fractures and in men with hip and ankle fractures than in the non-fracture group). Sarcopenia was significantly more prevalent in men with wrist, hip and ankle fractures than in women. SMI was correlated with BMD in different fracture groups. Logistic regression analyses revealed that lower SMI was associated with an increased risk of hip fracture both in men and women and ankle fracture in men.

Discussion

Sarcopenia may be an independent risk factor for hip and ankle fractures in men, and for hip fractures in women.     

Is Iron Accumulation a Possible Risk Factor for Sarcopenia?

Sarcopenia has a high incidence among the elderly, with significant negative effects on the quality of life. The pathogenesis of sarcopenia is complex, and many factors are involved in its development and progression. Sarcopenia might be associated with iron accumulation given that (1) age-related iron accumulation was found in the skeletal muscle, (2) excess iron could cause skeletal muscle damage or atrophy, and (3) patients with sarcopenia showed higher levels of serum ferritin. Understanding the etiology and pathogenesis of sarcopenia would help to develop new treatment and preventive methods, thereby improving the quality of life of the elderly patients.