Cellular regenerative therapy for acquired noncongenital musculoskeletal disorders

CITATION: Niesler, C. U. et al. 2019. Cellular regenerative therapy for acquired noncongenital musculoskeletal disorders. South African Medical Journal, 109(8b):58-63. doi:10.7196/SAMJ.2019.v109i8b.13860The original publication is available at http://www.samj.org.za/index.php/samj/indexENGLISH ABSTRACT: Stem cells have an inherent capacity to facilitate regeneration; this has led to unprecedented growth in their experimental use in various clinical settings, particularly in patients with diseases with few alternative treatment options. However, their approved clinical use has to date been restricted largely to haematological diseases and epidermal transplantation to treat severe burns. After thorough searching of two databases, this review illuminates the role of stem cell therapy for treatment of musculoskeletal diseases. Research suggests that successful application of stem cells as regenerative mediators is in all likelihood dependent on the ability of endogenous tissue-resident reparative mediators to respond to paracrine signals provided by the applied stem cells. Therefore, an understanding of how the pathological environment influences this process is crucial for the ultimate success of stem cell therapies. The current review presents both the progress and limitations of stem cells as regenerative mediators in the context of musculoskeletal disorders.DST/NRF South African Research Chair Initiativehttp://www.samj.org.za/index.php/samj/article/view/12714Publisher’s versio

Simple silicone chamber system for in vitro three-dimensonal skeletal muscle tissue formation

CITATION: Snyman, C., Goetsch, K. P., Myburgh, K. H. & Niesler, C. U. 2013. Simple silicone chamber system for in vitro three-dimensonal skeletal muscle tissue formation. Frontiers in Physiology, 4:1-6, doi:10.3389/fphys.2013.00349.The original publication is available at https://www.frontiersin.orgBioengineering skeletal muscle often requires customized equipment and intricate casting techniques. One of the major hurdles when initially trying to establish in vitro tissue engineered muscle constructs is the lack of consistency across published methodology. Although this diversity allows for specialization according to specific research goals, lack of standardization hampers comparative efforts. Differences in cell type, number and density, variability in matrix and scaffold usage as well as inconsistency in the distance between and type of adhesion posts complicates initial establishment of the technique with confidence. We describe an inexpensive, but readily adaptable silicone chamber system for the generation of skeletal muscle constructs that can readily be standardized and used to elucidate myoblast behavior in a three-dimensional space. Muscle generation, regeneration and adaptation can also be investigated in this model, which is more advanced than differentiated myotubes.https://www.frontiersin.org/articles/10.3389/fphys.2013.00349/fullPublisher's versio

Three-dimensional-printed gas dynamic virtual nozzles for x-ray laser sample delivery

Reliable sample delivery is essential to biological imaging using X-ray Free Electron Lasers (XFELs). Continuous injection using the Gas Dynamic Virtual Nozzle (GDVN) has proven valuable, particularly for time-resolved studies. However, many important aspects of GDVN functionality have yet to be thoroughly understood and/or refined due to fabrication limitations. We report the application of 2-photon polymerization as a form of high-resolution 3D printing to fabricate high-fidelity GDVNs with submicron resolution. This technique allows rapid prototyping of a wide range of different types of nozzles from standard CAD drawings and optimization of crucial dimensions for optimal performance. Three nozzles were tested with pure water to determine general nozzle performance and reproducibility, with nearly reproducible off-axis jetting being the result. X-ray tomography and index matching were successfully used to evaluate the interior nozzle structures and identify the cause of off-axis jetting. Subsequent refinements to fabrication resulted in straight jetting. A performance test of printed nozzles at an XFEL provided high quality femtosecond diffraction patterns. (C) 2016 Optical Society of Americ

Smoking Cessation Pharmacogenetics: Analysis of Varenicline and Bupropion in Placebo-Controlled Clinical Trials

Despite effective therapies for smoking cessation, most smokers find quitting difficult and most successful quitters relapse. Considerable evidence supports a genetic risk for nicotine dependence; however, less is known about the pharmacogenetics of smoking cessation. In the first pharmacogenetic investigation of the efficacy of varenicline and bupropion, we examined whether genes important in the pharmacodynamics and pharmacokinetics of these drugs and nicotine predict medication efficacy and adverse events. Subjects participated in randomized, double-blind, placebo-controlled smoking cessation clinical trials, comparing varenicline, a nicotinic acetylcholine receptor (nAChR) partial agonist, with bupropion, a norepinephrine/dopamine reuptake inhibitor, and placebo. Primary analysis included 1175 smokers of European ancestry, and 785 single nucleotide polymorphisms from 24 genes, representing 254 linkage disequilibrium (LD) bins (genes included nAChR subunits, additional varenicline-specific genes, and genes involved in nicotine or bupropion metabolism). For varenicline, continuous abstinence (weeks 9–12) was associated with multiple nAChR subunit genes (including CHRNB2, CHRNA5, and CHRNA4) (OR=1.76; 95% CI: 1.23–2.52) (p<0.005); for bupropion, abstinence was associated with CYP2B6 (OR=1.78; 95% CI: 1.27–2.50) (p<0.001). Incidence of nausea was associated with several nAChR subunit genes (OR=0.50; 95% CI: 0.36–0.70) (p<0.0001) and time to relapse after quitting was associated with HTR3B (HR=1.97; 95% CI: 1.45–2.68) (p<0.0001). These data provide evidence for multiple genetic loci contributing to smoking cessation and therapeutic response. Different loci are associated with varenicline vs bupropion response, suggesting that additional research may identify clinically useful markers to guide treatment decisions

The changing AMPK expression profile in differentiating mouse skeletal muscle myoblast cells helps confer increasing resistance to apoptosis.

AMP-activated protein kinase (AMPK) functions as a alpha/beta/gamma heterotrimer to preserve ATP levels and so cell viability during stressful conditions. However, its role in aiding survival of adult skeletal muscle precursor cells is unclear. Using the differentiating mouse C2C12 postnatal skeletal muscle myoblast cell line, we have determined that proteins for the AMPK subunit isoforms alpha2 and gamma2 are constitutively expressed, while those for alpha1, beta1 and beta2 are undetectable in undifferentiated myoblasts but increasingly expressed with differentiation to myotubes. Although the gamma3 subunit is expressed at a low level in myoblasts, it too is expressed increasingly with differentiation to myotubes. The p50 but not the p72 isoform of the embryonic alpha subunit homologue MELK is expressed only in proliferating myoblasts, while the ARK5 alpha subunit homologue is increasingly expressed with differentiation. Myotubes displayed higher basal and stimulated alpha1/alpha2 AMPK activation than myoblasts. Furthermore, serum starvation resulted in less apoptosis of differentiated myotubes than of undifferentiated myoblasts. This reflects, in part, the increased expression of functional AMPK in the myotubes, since specific inhibition of AMPK activity with 6-[4-(2-piperidin-1-ylethoxy)-phenyl]-3-pyridin-4-ylpyrazolo[1,5-alpha] pyrimidine (Compound C) exacerbated the apoptosis resulting from serum withdrawal. If these in vitro events can also occur in vivo, they could have implications for pathologies such as muscle wasting, in which undifferentiated satellite stem cells may be easier apoptotic targets than their differentiated counterparts. Furthermore, these results suggest that when interpreting results from in vitro or in vivo experiments on AMPK, the subunit expression profile should be taken into account

Satellite cell pool expansion is affected by skeletal muscle characteristics

INTRODUCTION We investigated changes in satellite cell (SC) pool size after an acute bout of strenuous exercise and evaluated the influence of baseline SC count and fiber type. METHODS Participants completed a downhill running (DHR) intervention (5 &times; 8 min, 2-min rest; 80% VO2max ; -10% gradient). Muscle biopsies were taken 7 days before VO₂max and 7-9 days after the DHR intervention. Delayed-onset muscle soreness (DOMS) and creatine kinase activity (CK) were measured on days 1, 2, 7, and 9 post-DHR. SCs were identified by Pax7 and laminin staining. Relative distribution of MHC isoforms was determined by electrophoresis. RESULTS DOMS and CK peaked on day 1 post-DHR (P &lt; 0.01). The SC pool increased (26%) after DHR (P = 0.005). SCs/total myonuclei after recovery correlated with baseline SCs (r = 0.979, P = 0.003) and VO₂max (r = 0.956, P = 0.011), whereas change in SC pool (Pax7(+) cells/total myonuclei: recovery minus baseline) tended to correlate with percent MHC II (r = 0.848; P = 0.06). CONCLUSION Interindividual physiological characteristics affect SC pool expansion after a single bout of DHR and are influenced by VO₂max