SIRT1 as a potential therapeutic target in pelvic organ prolapse due to protective effects against oxidative stress and cellular senescence in human uterosacral ligament fibroblasts.

INTRODUCTION: The pathogenesis of pelvic organ prolapse (POP), an age-related disease, has not been fully elucidated. Therapeutic targets of POP are limited. Silencing information regulator 2 related enzyme 1 (SIRT1), a gene considered capable of regulating oxidative stress and cellular senescence, has been widely demonstrated involved in aging and age-related diseases. The present study aimed to explore the role of SIRT1 in POP in vivo and in vitro. METHODS: Expression levels of SIRT1 in uterosacral ligament (USL) tissues from patients with or without POP were measured using immunohistochemical assays. SRT1720, a SIRT1 agonist, was used to upregulate SIRT1, and hydrogen peroxide (H2O2) was used to establish an oxidative stress model in human uterosacral ligament fibroblasts (hUSLFs). The effects of SIRT1 on cell viability, apoptosis, senescence, and reactive oxygen species (ROS) levels were detected, respectively. Western blot assays were used to examine expression levels of apoptosis- and senescence-associated biomarkers. Unpaired Student's t test, Mann-Whitney U test, χ2 test, and one-way ANOVA were performed for determining statistically significant differences. RESULTS: Compared to the control group, expression levels of SIRT1 were downregulated in USL tissues and hUSLFs from patients with POP, and associated with stage (p < 0.05). hUSLFs of patients with POP had lower growth rates (p < 0.0001) than those of the control group, which were improved by upregulating SIRT1 (p < 0.05). The senescent proportion was higher in the POP group than the control group (43.63 ± 10.62% vs. 4.84 ± 5.32%, p < 0.0001), which could be reduced by upregulating SIRT1 (p < 0.0001). High ROS levels in the POP group were also alleviated by SRT1720. H2O2 exposure increased ROS levels, inhibited proliferation, and triggered apoptosis and senescence in hUSLFs of patients without POP in a concentration-dependent manner. Further, these damages were alleviated by pretreatment with SRT1720. CONCLUSIONS: SIRT1 is downregulated in patients with POP, and the development of SIRT1 activators or agonists may have applications in the treatment and prevention of POP through antioxidative stress and antisenescence effects.

Effects of dextrose prolotherapy on tendinopathy, fasciopathy, and ligament injuries, fact or myth?: A systematic review and meta-analysis.

OBJECTIVES: Prolotherapy or proliferative therapy is a treatment option for damaged connective tissues involving the injection of a solution (proliferant) which theoretically causes an initial cell injury and a subsequent "proliferant" process of wound healing via modulation of the inflammatory process. Nonetheless, the benefits of dextrose prolotherapy have not been adequately evaluated. Therefore, the present study assesses the effectiveness and superiority of prolotherapy separately in treating dense fibrous connective tissue injuries. METHODS: PubMed, Scopus, and Embase were searched from the earliest record to February 18, 2019. This study included randomized controlled trials whichBoth analysis at individual studies level and pooled meta-analysis were performed. RESULTS: Ten trials involving 358 participants were included for review. At study level, the majority of comparisons did not reveal significant differences between dextrose prolotherapy and no treatment (or placebo) regarding pain control. The meta-analysis showed dextrose prolotherapy was effective in improving activity only at immediate follow-up (i.e., 0-1 month) (standardized mean difference [SMD]: 0.98; 95% confidence interval [CI]: 0.40-1.50; I = 0%); and superior to corticosteroid injections only in pain reduction at short-term follow-up (i.e., 1-3 month) (SMD: 0.70; 95% CI: 0.14-1.27; I = 51%). No other significant SMDs were found in this analysis. CONCLUSIONS: There is insufficient evidence to support the clinical benefits of dextrose prolotherapy in managing dense fibrous tissue injuries. More high-quality randomized controlled trials are warranted to establish the benefits of dextrose prolotherapy. REVIEW REGISTRATION: PROSPERO (CRD42019129044).

Effects of short-term moderate ZEN consumption on uterosacral ligament elasticity in pubertal gilts.

Zearalenone (ZEN) is a potent estrogenic toxin in swine, contributing to economic losses in herds via reproductive consequences such as pelvic organ prolapse (POP). To better understand the relationship between ZEN-consumption and reproductive symptoms, an animal feeding study with pubertal gilts was designed. The gilts were exposed to three different treatments: solvent-only feed for 21 days (n = 10), ZEN-spiked feed for 7 days followed by solvent-only feed for 14 days (n = 10), and ZEN-spiked feed for 21 days (n = 10). The gilts did not display any ZEN-related symptoms throughout any of the treatments. At the end of the trial the elastic properties of the USLs from participating gilts were evaluated along two loading directions: main direction (MD) and perpendicular direction (PD). The elastic properties included average stresses at 2% and 4% strains, and secant moduli. Overall the elastic properties of the USLs did not vary across treatment groups or between loading directions. In the MD, average stress increased from 32.96 ± 4.43 kPa at 2% strain to 63.21 ± 9.69 kPa at 4% strain, with a secant modulus of 1.52 ± 0.27 MPa. In the PD, average stress increased from 40.82 ± 4.22 kPa at 2% strain to 83.38 ± 9.17 kPa at 4% strain, with a secant modulus of 2.13 ± 0.31 MPa. Continued research into the relationship between ZEN consumption and reproductive symptoms such as POP is necessary in order to mitigate their deleterious effects in herds.

Injectable hydrogels for tendon and ligament tissue engineering.

The problem of tendon and ligament (T/L) regeneration in musculoskeletal diseases has long constituted a major challenge. In situ injection of formable biodegradable hydrogels, however, has been demonstrated to treat T/L injury and reduce patient suffering in a minimally invasive manner. An injectable hydrogel is more suitable than other biological materials due to the special physiological structure of T/L. Most other materials utilized to repair T/L are cell-based, growth factor-based materials, with few material properties. In addition, the mechanical property of the gel cannot reach the normal T/L level. This review summarizes advances in natural and synthetic polymeric injectable hydrogels for tissue engineering in T/L and presents prospects for injectable and biodegradable hydrogels for its treatment. In future T/L applications, it is necessary develop an injectable hydrogel with mechanics, tissue damage-specific binding, and disease response. Simultaneously, the advantages of various biological materials must be combined in order to achieve personalized precision therapy.

Estrogen Enhances Osteogenic Differentiation of Human Periodontal Ligament Stem Cells by Activating the Wnt/ß-Catenin Signaling Pathway.

OBJECTIVES: This study was designed to investigate the role of the Wnt/ß-catenin signaling pathway in estrogen-enhanced osteogenic differentiation of human peridontal ligament stem cells (hPLSCs). METHODS: The limiting dilution technique was used for cloning and purification of hPLSCs. Flow cytometric analysis of STRO-1, CD146, and CD45 was conducted to identify hPLSCs. The P3 hPDLSCs were divided into 4 groups: Control, 10M E2, 10M E2+100 ng/mL Wnt3a, 10M E2+5 × 10M Xav939. After 7 days of osteogenic induction, qRT-PCR was used to detect the mRNA expression of ß-catenin, CyclinD1, alkaline phosphatase, Runx2, and OCN; Western blot was used to detect the protein expression of ß-catenin, GSK3ß, P-GSK3ß, CyclinD1, Runx2, and OCN; After 1, 3, 5, 7 days of osteogenic induction, the activity of alkaline phosphatase was detected. RESULTS: The authors' results showed that E2 was able to enhance the osteogenic differentiation of hPDLSCs and Wnt/ß-catenin signaling pathway was involved. Wnt3a activated the signaling pathway of Wnt/ß-catenin and further enhanced the osteogenesis of hPDLSCs. Xav939 inhibited the Wnt/ß-catenin signaling pathway in estrogen-mediated environment, but did not obviously inhibit the osteogenic differentiation of hPDLSCs. CONCLUSIONS: E2 enhanced osteogenic differentiation of hPDLSCs through the activation of the Wnt/ß-catenin signaling pathway.

Chemical Optimization for Functional Ligament Tissue Engineering.

Electrospun materials are widely used for functional tissue engineering for its robust production and biomimetic properties. Several issues persist, however, including heterogeneous cell distribution, insufficient matrix elaboration/accumulation, and limited construct size. We took three synergistic approaches to address these issues by modifying the chemical microenvironment for the seeded cells. Instead of the commonly used fibronectin, we demonstrated that type I collagen (COL) coating, facilitated by polydopamine treatment, promoted cell infiltration into the fibrous scaffold and resulted in homogeneous distribution in one week. Sequential treatment with fibroblast growth factor and transforming growth factor-ß after cell infiltration enhanced cell proliferation and matrix deposition, with increased lysyl oxidase and decreased matrix metalloproteinase-1 expressions. Finally, lamination of the fibrous sheets with fibrin gel not only increased construct size, but further stimulated COL deposition and improved construct mechanical functionalities in combination with sequential growth factor supplementation. These soluble and insoluble chemical optimizations encouraged rapid and robust construct development for a functional engineered ligament graft and can be adapted for the engineering of other tissues. Impact Statement Ligament and tendon injuries are some of the most common orthopedic injuries with long-term repercussions. Tissue engineered grafts provide a promising alternative to autograft and allografts. We present in this study robust and synergistic chemical optimization approaches for the functional engineering of ligament grafts. Moreover, these approaches can be adapted for a variety of other tissues to improve homogeneous construct development.

An overview of decellularisation techniques of native tissues and tissue engineered products for bone, ligament and tendon regeneration.

Decellularised tissues and organs have been successfully used in a variety of tissue engineering/regenerative medicine applications. Because of the complexity of each tissue (size, porosity, extracellular matrix (ECM) composition etc.), there is no standardised protocol and the decellularisation methods vary widely, thus leading to heterogeneous outcomes. Physical, chemical, and enzymatic methods have been developed and optimised for each specific application and this review describes the most common strategies utilised to achieve decellularisation of soft and hard tissues. While removal of the DNA is the primary goal of decellularisation, it is generally achieved at the expense of ECM preservation due to the harsh chemical or enzymatic processing conditions. As denaturation of the native ECM has been associated with undesired host responses, decellularisation conditions aimed at effectively achieving simultaneous DNA removal and minimal ECM damage will be highlighted. Additionally, the utilisation of decellularised matrices in regenerative medicine is explored, as are the most recent strategies implemented to circumvent challenges in this field. In summary, this review focusses on the latest advancements and future perspectives in the utilisation of natural ECM for the decoration of synthetic porous scaffolds.

Role of Biomaterials and Controlled Architecture on Tendon/Ligament Repair and Regeneration.

Engineering synthetic scaffolds to repair and regenerate ruptured native tendon and ligament (T/L) tissues is a significant engineering challenge due to the need to satisfy both the unique biological and biomechanical properties of these tissues. Long-term clinical outcomes of synthetic scaffolds relying solely on high uniaxial tensile strength are poor with high rates of implant rupture and synovitis. Ideal biomaterials for T/L repair and regeneration need to possess the appropriate biological and biomechanical properties necessary for the successful repair and regeneration of ruptured tendon and ligament tissues.

Effects of Human Placenta Extract (Laennec) on Ligament Healing in a Rodent Model.

Rich in bioactive substances such as amino acids and peptides, Laennec (human placenta hydrolysate) has been widely used to control various types of musculoskeletal pain. However, the effects of Laennec on tendon and ligament injuries are not clearly understood. In the present study, Laennec was tested to identify its in vivo effects on ligament injury in an animal model and its in vitro effects on tendon-derived fibrocytes. A total of 99 Sprague Dawley rats were divided into the negative control (normal) group (n = 11) and the ligament injury group (n = 88). The ligament injury group was subdivided into normal saline-treated group, Laennec-treated group, polydeoxyribonucleotide-treated group, and 20% dextrose-treated group. Ligaments were collected at 1 week and 4 weeks after treatment. Histologic and biomechanical properties were analyzed. In vitro effects of Laennec and polydeoxyribonucleotide on fibrocytes were also analyzed. Although all other treatment groups showed increased inflammatory cells, the Laennec-treated group maintained cell counts and activated macrophage levels that were similar to the normal group. Unlike the saline-treated group and dextrose-treated group, the Laennec-treated group had low levels of degenerative changes at 4 weeks after treatment. Supportively, in vitro results showed that the Laennec-treated group had increased collagen type I, scleraxis (Scx) and tenomodulin (Tnmd) expression (p < 0.05). Our study demonstrates that Laennec treatment enhances wound healing of damaged ligament by suppressing immune responses and reducing degenerative changes of damaged ligament. In addition, we found that Laennec induces the gene expression of type I collagen, Scx and Tnmd in fibrocytes, suggesting that Laennec may facilitate regeneration of damaged ligaments. Therefore, we expect that Laennec can be a useful drug to treat injured ligament.

Morphologically bioinspired hierarchical nylon 6,6 electrospun assembly recreating the structure and performance of tendons and ligaments.

Reconstructions of ruptured tendons and ligaments currently have dissatisfactory failure rate. Failures are mainly due to the mechanical mismatch of commercial implants with respect to the host tissue. In fact, it is crucial to replicate the morphology (hierarchical in nature) and mechanical response (highly-nonlinear) of natural tendons and ligaments. The aim of this study was to develop morphologically bioinspired hierarchical Nylon 6,6 electrospun assemblies recreating the structure and performance of tendons and ligaments. First, we built different electrospun bundles to find the optimal orientation of the nanofibers. A 2nd-level hierarchical assembly was fabricated with a dedicated process that allowed tightly joining the bundles one next to the other with an electrospun sheath, so as to improve the mechanical performance. Finally, a further hierarchical 3rd-level assembly was constructed by grouping several 2nd-level assemblies. The morphology of the different structures was assessed with scanning electron microscopy and high-resolution X-ray tomography, which allowed measuring the directionality of the nanofibers in the bundles and in the sheaths. The mechanical properties of the single bundles and of the 2nd-level assemblies were measured with tensile tests. The single bundles and the hierarchical assemblies showed morphology and directionality of the nanofibers similar to the tendons and ligaments. The strength and stiffness were comparable to that of tendons and ligaments. In conclusion, this work showed an innovative electrospinning production process to build nanofibrous Nylon 6,6 hierarchical assemblies which are suitable as future implantable devices and able to mimic the multiscale morphology and the biomechanical properties of tendons and ligaments.

Encapsulation of a nanoporous simvastatin-chitosan composite to enhance osteointegration of hydroxyapatite-coated polyethylene terephthalate ligaments.

PURPOSE: This study was designed to evaluate the in vitro and in vivo biocompatibility and osteointegration of plasma-sprayed hydroxyapatite (HA)-coated polyethylene terephthalate (PET) ligaments encapsulated with a simvastatin (SV)-chitosan (CS) composite. METHODS: This study compared the in vitro and in vivo bone responses to three different PET ligaments: SV/CS/PET-HA, CS/PET-HA and PET-HA. A field emission scanning electron microscope was used to characterize the morphology, and the in vitro SV release profile was analyzed. MC3T3 cells were cocultured with SV/CS/PET-HA, CS/PET-HA and PET-HA to test their biocompatibility using CCK-8 tests. Osteogenic differentiation was investigated by the expression of marker genes using qPCR. Osteointegration was performed by implanting the PET ligaments into the proximal tibia bone tunnels of male Sprague-Dawley rats for 3 weeks and 6 weeks. The bone-implant interface was evaluated by micro-computed tomography (micro-CT) and histological analysis. RESULTS: The characteristic nanoporous structures mainly formed on the surface of the plasma-sprayed HA particles in the SV/CS/PET-HA and CS/PET-HA groups. The SV release test showed that the sustained release of simvastatin lasted for 25 days in the SV/CS/PET-HA group. The in vitro studies demonstrated that the SV/CS/PET-HA ligaments induced osteogenic differentiation in the MC3T3 cells, with higher mRNA expression levels of collagen-1, bone morphogenetic protein-2, osteocalcin and alkaline phosphatase than those in the CS/PET-HA and PET-HA ligament groups. The in vivo tests showed that both micro-CT analysis (bone mineral density and bone volume per total volume) and histological analysis (bone implant contact and interface area) revealed significantly higher peri-implant bone formation and less interface area in the SV/CS/PET-HA group than in the other groups. CONCLUSION: The SV-CS composite nanoporous structure was associated with the improved biocompatibility and osteogenic differentiation in vitro and enhanced osteointegration process in vivo of plasma-sprayed HA-coated PET ligaments.

The Effectiveness of Ultrasound-Guided Steroid Injection Combined with Miniscalpel-Needle Release in the Treatment of Carpal Tunnel Syndrome vs. Steroid Injection Alone: A Randomized Controlled Study.

OBJECTIVES: Carpal tunnel syndrome (CTS) is one of the most common nerve entrapment syndromes, which has a serious impact on patients' work and life. The most effective conservative treatment is steroid injection but its long-term efficacy is still not satisfactory. The aim of this study was to evaluate the effectiveness of steroid injection combined with miniscalpel-needle (MSN) release for treatment of CTS under ultrasound guidance versus steroid injection alone. We hypothesized that combined therapy could be more beneficial. METHODS: Fifty-one patients with CTS were randomly allocated into two groups, namely, steroid injection combined with MSN release group and steroid injection group. The therapeutic effectiveness was evaluated using Boston Carpal Tunnel Questionnaire (BCTQ), cross-sectional area (CSA) of the median nerve, and four electrophysiological parameters, including distal motor latency (DML), compound muscle action potential (CMAP), sensory nerve action potential (SNAP), and sensory nerve conduction velocity (SNCV) at baseline, 4 and 12 weeks after treatment. RESULTS: Compared with baseline, all the parameters in both groups showed statistically significant improvement at week 4 and week 12 follow-up, respectively (P<0.05). When compared with steroid injection group, the outcomes including BCTQ, DML, CMAP, SNCV, and CSA of the median nerve were significantly better in steroid injection combined with MSN release group at week 12 after treatment (P<0.05). CONCLUSIONS: The effectiveness of steroid injection combined with MSN release for CTS is superior to that of steroid injection alone, which may have important implications for future clinical practice. This Chinese clinical trial is registered with ChiCTR1800014530.

Gastric pentadecapeptide body protection compound BPC 157 and its role in accelerating musculoskeletal soft tissue healing.

There is a current need for a therapy that can alleviate the social and economic burden that presents itself with debilitating and recurring musculoskeletal soft tissue injuries and disorders. Currently, several therapies are emerging and undergoing trials in animal models; these focus on the manipulation and administration of several growth factors implicated with healing. However, limitations include in vivo instability, reliance on biocompatible and robust carriers and restricted application procedures (local and direct). The aim of this paper is therefore to critically review the current literature surrounding the use of BPC 157, as a feasible therapy for healing and functional restoration of soft tissue damage, with a focus on tendon, ligament and skeletal muscle healing. Currently, all studies investigating BPC 157 have demonstrated consistently positive and prompt healing effects for various injury types, both traumatic and systemic and for a plethora of soft tissues. However, to date, the majority of studies have been performed on small rodent models and the efficacy of BPC 157 is yet to be confirmed in humans. Further, over the past two decades, only a handful of research groups have performed in-depth studies regarding this peptide. Despite this, it is apparent that BPC 157 has huge potential and following further development has promise as a therapy to conservatively treat or aid recovery in hypovascular and hypocellular soft tissues such as tendon and ligaments. Moreover, skeletal muscle injury models have suggested a beneficial effect not only for disturbances that occur as a result of direct trauma but also for systemic insults including hyperkalamia and hypermagnesia. Promisingly, there are few studies reporting any adverse reactions to the administration of BPC 157, although there is still a need to understand the precise healing mechanisms for this therapy to achieve clinical realisation.

Enhanced biological properties of biomimetic apatite fabricated polycaprolactone/chitosan nanofibrous bio-composite for tendon and ligament regeneration.

The development of tailored nanofibrous scaffolds for tendon and ligament tissue engineering has been a goal of clinical research for current researchers. Here, we establish a formation of novel nanofibrous matrix with significant mechanical and biological properties by electro-spinning process. The fine fibrous morphology of the nanostructured hydroxyapatite (HAp) dispersed in the polycaprolactone/chitosan (HAp-PCL/CS) nanofibrous matrix was exhibited by microscopic (SEM and TEM) techniques. The favorable mechanical properties (load and modulus) were achieved. The load and modulus of the HAp-PCL/CS composite fibers was 250.1N and 215.5MPa, which is very similar to that of standard value of the human tendon and ligament tissues. The cellular responses and biocompatibility of HAp-PCL/CS nanofibrous scaffolds were investigated with human osteoblast (HOS) cells for tendon regeneration and examined the primary osteoblast mechanism by in vitro method. The morphological (FE-SEM and fluorescence) microscopic images clearly exhibited that HOS cells are well attached and flatted on the nanofibrous composites. The HAp dispersed PCL/CS nanofibrous scaffolds promoted higher adhesion and proliferation of HOS cells comparable to the nanofibrous scaffolds without HAp nanoparticles. The physic-chemical and biological properties of the synthesized nanofibrous scaffold were very close to that of normal ligament and tendon in human body. Over all, these studied results confirmed that the prepared nanofibrous scaffolds will be effective biomaterial of tendon ligament regeneration applications.

Nerve growth factor administration on cultured human ligamentocytes: an in vitro pilot study.

Nerve growth factor (NGF) is involved in several joint pathologies. It has been demonstrated that its concentration increases in synovial fluid and tissue from arthritis. However, its role in joint homeostasis and pathophysiology still remain to be clarified. This study analyzed the effect of 200 ng/ml on cultured human ligamentocytes by evaluating cell proliferation, cell phenotype and gene expression. The MTT test excluded an influence on cell viability at 7 and 14 days. Regarding cell phenotype, we observed that NGF might promote the synthesis of COL1A1. On the other hand, real time PCR showed that NGF did not influence gene expression of COL3A1, FGF-BETA, IGF1, MMP2, MMP3, MMP9 and MMP13. However, COL1A1 gene was significantly upregulated in treated cell at 14 days. Our results suggest that NGF may have an anabolic effect on ligament. Additional investigations are necessary to determine how NGF may influence ligamentocytes..

Use of Microcomputed Tomography to Measure the Relaxin-induced Expansion of Intrapubic Ligaments in Mice.

Relaxin is a 6-kDa peptide in the insulin superfamily of hormones. In addition to its effects on reproductive and musculoskeletal ligaments, relaxin has demonstrated beneficial effects on cardiac, renal, and vascular systems in preclinical models. The mouse intrapubic ligament ex vivo bioassay is the current standard for measuring in vivo relaxin bioactivity. However, this bioassay necessitates euthanasia and dissection of large cohorts to measure the intrapubic ligament at specified time points. We hypothesized that µCT imaging could be used to reduce the number of animals necessary for the intrapubic ligament bioassay by enabling a single animal to be followed longitudinally throughout the study rather than euthanizing different cohorts at established time points. Female CD1 mice were used to compare µCT imaging with the current standard. Both protocols revealed significant differences in intrapubic ligament length, with the µCT data having greater power when corrected for baseline imaging. From these data, we concluded that using µCT to measure the intrapubic ligament in mice primed with estrogen and dosed with relaxin is a viable refinement and will allow the use of fewer animals in longitudinal studies and provide more robust data, because animals can serve as their own controls.

The effect of estrogen on tendon and ligament metabolism and function.

Tendons and ligaments are crucial structures inside the musculoskeletal system. Still many issues in the treatment of tendon diseases and injuries have yet not been resolved sufficiently. In particular, the role of estrogen-like compound (ELC) in tendon biology has received until now little attention in modern research, despite ELC being a well-studied and important factor in the physiology of other parts of the musculoskeletal system. In this review we attempt to summarize the available information on this topic and to determine many open questions in this field.

* Fabrication and Characterization of Biphasic Silk Fibroin Scaffolds for Tendon/Ligament-to-Bone Tissue Engineering.

Tissue engineering is an attractive strategy for tendon/ligament-to-bone interface repair. The structure and extracellular matrix composition of the interface are complex and allow for a gradual mechanical stress transfer between tendons/ligaments and bone. Thus, scaffolds mimicking the structural features of the native interface may be able to better support functional tissue regeneration. In this study, we fabricated biphasic silk fibroin scaffolds designed to mimic the gradient in collagen molecule alignment present at the interface. The scaffolds had two different pore alignments: anisotropic at the tendon/ligament side and isotropic at the bone side. Total porosity ranged from 50% to 80% and the majority of pores (80-90%) were <100-300 µm. Young's modulus varied from 689 to 1322 kPa depending on the type of construct. In addition, human adipose-derived mesenchymal stem cells were cultured on the scaffolds to evaluate the effect of pore morphology on cell proliferation and gene expression. Biphasic scaffolds supported cell attachment and influenced cytoskeleton organization depending on pore alignment. In addition, the gene expression of tendon/ligament, enthesis, and cartilage markers significantly changed depending on pore alignment in each region of the scaffolds. In conclusion, the biphasic scaffolds fabricated in this study show promising features for tendon/ligament-to-bone tissue engineering.

Vitamin C-enriched gelatin supplementation before intermittent activity augments collagen synthesis.

BACKGROUND: Musculoskeletal injuries are the most common complaint in active populations. More than 50% of all injuries in sports can be classified as sprains, strains, ruptures, or breaks of musculoskeletal tissues. Nutritional and/or exercise interventions that increase collagen synthesis and strengthen these tissues could have an important effect on injury rates. OBJECTIVE: This study was designed to determine whether gelatin supplementation could increase collagen synthesis. DESIGN: Eight healthy male subjects completed a randomized, double-blinded, crossover-design study in which they consumed either 5 or 15 g of vitamin C-enriched gelatin or a placebo control. After the initial drink, blood was taken every 30 min to determine amino acid content in the blood. A larger blood sample was taken before and 1 h after consumption of gelatin for treatment of engineered ligaments. One hour after the initial supplement, the subjects completed 6 min of rope-skipping to stimulate collagen synthesis. This pattern of supplementation was repeated 3 times/d with ≥6 h between exercise bouts for 3 d. Blood was drawn before and 4, 24, 48, and 72 h after the first exercise bout for determination of amino-terminal propeptide of collagen I content. RESULTS: Supplementation with increasing amounts of gelatin increased circulating glycine, proline, hydroxyproline, and hydroxylysine, peaking 1 h after the supplement was given. Engineered ligaments treated for 6 d with serum from samples collected before or 1 h after subjects consumed a placebo or 5 or 15 g gelatin showed increased collagen content and improved mechanics. Subjects who took 15 g gelatin 1 h before exercise showed double the amino-terminal propeptide of collagen I in their blood, indicating increased collagen synthesis. CONCLUSION: These data suggest that adding gelatin to an intermittent exercise program improves collagen synthesis and could play a beneficial role in injury prevention and tissue repair. This trial was registered at the Australian New Zealand Clinical Trials Registry as ACTRN12616001092482.

Annexin A2, up-regulated by IL-6, promotes the ossification of ligament fibroblasts from ankylosing spondylitis patients.

BACKGROUND: Annexin A2, a calcium-dependent phospholipid binding protein, is involved in osteogenesis. The objective of the present study was to explore the expression of Annexin A2 in spinal ligament tissues (LT) of ankylosing spondylitis (AS) patients and determine its pathological functions. METHODS: mRNA and protein expression of Annexin A2 was detected by real-time PCR and Western blotting, respectively. Interleukin-6 (IL-6) concentration in serum was assessed by enzyme linked immunosorbent assay. Alkaline phosphatase (ALP) activity was measured with ALP activity kit on a microplate reader. RESULTS: mRNA and protein expression of Annexin A2 in LT, and IL-6 concentration in serum were significantly increased in AS patients. Moreover, exogenous IL-6 treatment significantly up-regulated Annexin A2 expression and ALP activity. Silencing of Annexin A2 expression significantly ameliorated IL-6-induced ossification of fibroblasts from AS patients, as indicated by ALP activity, expression of proteins associated with osteogenic differentiation, including bone morphogenetic protein-2, osteocalcin and osterix, and the ratio of osteoprotegerin to receptor activator of NF-κB ligand. Further MEK inhibitor experiments suggested that Annexin A2 may exert its function through extracellular signal-related kinase pathway. CONCLUSIONS: Annexin A2, up-regulated by IL-6, may promote ligament ossification of AS patients.