Philipp Herlyn

Vitamin D Increases Cellular Turnover and Functionally Restores the Skeletal Muscle after Crush Injury in Rats

Insufficient skeletal muscle regeneration after injury often impedes the healing process and is accompanied by functional deficiencies or pain. The aim of our study was to provide evidence that vitamin D improves muscle healing after muscle injury. Therefore, we used male rats and induced an injury of the soleus muscle. After crush injury, animals received either 8.3 mg/kg (332,000 IU/kg) body weight vitamin D or vehicle solution, s.c. After assessment of muscle force at days 1, 4, 14, and 42 after injury, sampling of muscle tissue served for analysis of proliferation, apoptosis, satellite cells, and prolyl-4-hydroxylase-β expression. Vitamin D application caused a significant increase in cell proliferation and a significant inhibition of apoptosis at day 4 after injury compared to control animals. The numbers of satellite cells were not influenced by the vitamin D application, but there was an increase in prolyl-4-hydroxylase-β expression, indicative of increased extracellular matrix proteins. This cellular turnover resulted in a faster recovery of contraction forces at day 42 in the vitamin D group. Current data support the hypothesis that vitamin D promotes the regenerative process in injured muscle. Thus, vitamin D treatment may represent a promising therapy to optimize recovery after injury.

Plyometric training improves voluntary activation and strength during isometric, concentric and eccentric contractions

OBJECTIVES This study investigated effects of plyometric training (6 weeks, 3 sessions/week) on maximum voluntary contraction (MVC) strength and neural activation of the knee extensors during isometric, concentric and eccentric contractions. DESIGN Twenty-seven participants were randomly assigned to the intervention or control group. METHODS Maximum voluntary torques (MVT) during the different types of contraction were measured at 110° knee flexion (180°=full extension). The interpolated twitch technique was applied at the same knee joint angle during isometric, concentric and eccentric contractions to measure voluntary activation. In addition, normalized root mean square of the EMG signal at MVT was calculated. The twitch torque signal induced by electrical nerve stimulation at rest was used to evaluate training-related changes at the muscle level. In addition, jump height in countermovement jump was measured. RESULTS After training, MVT increased by 20Nm (95% CI: 5-36Nm, P=0.012), 24Nm (95% CI: 9-40Nm, P=0.004) and 27Nm (95% CI: 7-48Nm, P=0.013) for isometric, concentric and eccentric MVCs compared to controls, respectively. The strength enhancements were associated with increases in voluntary activation during isometric, concentric and eccentric MVCs by 7.8% (95% CI: 1.8-13.9%, P=0.013), 7.0% (95% CI: 0.4-13.5%, P=0.039) and 8.6% (95% CI: 3.0-14.2%, P=0.005), respectively. Changes in the twitch torque signal of the resting muscle, induced by supramaximal electrical stimulation of the femoral nerve, were not observed, indicating no alterations at the muscle level, whereas jump height was increased. CONCLUSIONS Given the fact that the training exercises consisted of eccentric muscle actions followed by concentric contractions, it is in particular relevant that the plyometric training increased MVC strength and neural activation of the quadriceps muscle regardless of the contraction mode.

The standardized creation of a lumbar spine vertebral compression fracture in a sheep osteoporosis model induced by ovariectomy, corticosteroid therapy and calcium/phosphorus/vitamin D-deficient diet.

INTRODUCTION Vertebral compression fractures (VCFs) are one of the most common injuries in the aging population presenting with an annual incidence of 1.4 million new cases in Europe. Current treatment strategies focus on cement-associated solutions (kyphoplasty/vertebroplasty techniques). Specific cement-associated problems as leakage, embolism and the adjacent fracture disease are reported adding to open questions like general fracture healing properties of the osteoporotic spine. In order to analyze those queries animal models are of great interest; however, both technical difficulties in the induction of experimental osteoporosis in animal as well as the lack of a standardized fracture model impede current and future in vivo studies. This study introduces a standardized animal model of an osteoporotic VCF type A3.1 that may enable further in-depth analysis of the afore mentioned topics. MATERIAL AND METHODS Twenty-four 5-year-old female Merino sheep (mean body weight: 67 kg; range 57-79) were ovariectomized (OP1) and underwent 5.5 months of weekly corticosteroid injections (dexamethasone and dexamethasone-sodium-phosphate), adding to a calcium/phosphorus/vitamin D-deficient diet. Osteoporosis induction was documented by pQCT and micro-CT BMD (bone mineral density) as well as 3D histomorphometric analysis postoperatively of the sheep distal radius and spine. Non osteoporotic sheep served as controls. Induction of a VCF of the second lumbar vertebra was performed via a mini-lumbotomy surgical approach with a standardized manual compression mode (OP2). RESULTS PQCT analysis revealed osteoporosis of the distal radius with significantly reduced BMD values (0.19 g/cm(3), range 0.13-0.22 vs. 0.27 g/cm(3), range 0.23-0.32). Micro-CT documented significant lowering of BMD values for the second lumbar vertebrae (0.11 g/cm(3), range 0.10-0.12) in comparison to the control group (0.14 g/cm(3), range 0.12-0.17). An incomplete burst fracture type A3.1 was achieved in all cases and resulted in a significant decrease in body angle and vertebral height (KA 4.9°, range: 2-12; SI 4.5%, range: 2-12). With OP1, one minor complication (lesion of small bowel) occurred, while no complications occurred with OP2. CONCLUSIONS A suitable spinal fracture model for creation of VCFs in osteoporotic sheep was developed. The technique may promote the development of improved surgical solutions for VCF treatment in the experimental and clinical setting.

Dietary Restriction-Induced Alterations in Bone Phenotype: Effects of Lifelong Versus Short-Term Caloric Restriction on Femoral and Vertebral Bone in C57BL/6 Mice

Caloric restriction (CR) is a well‐described dietary intervention that delays the onset of aging‐associated biochemical and physiological changes, thereby extending the life span of rodents. The influence of CR on metabolism, strength, and morphology of bone has been controversially discussed in literature. Thus, the present study evaluated whether lifelong CR versus short‐term late‐onset dietary intervention differentially affects the development of senile osteoporosis in C57BL/6 mice. Two different dietary regimens with 40% food restriction were performed: lifelong CR starting in 4‐week‐old mice was maintained for 4, 20, or 74 weeks. In contrast, short‐term late‐onset CR lasting a period of 12 weeks was commenced at 48 or 68 weeks of age. Control mice were fed ad libitum (AL). Bone specimens were assessed using microcomputed tomography (μCT, femur and lumbar vertebral body) and biomechanical testing (femur). Adverse effects of CR, including reduced cortical bone mineral density (Ct.BMD) and thickness (Ct.Th), were detected to some extent in senile mice (68+12w) but in particular in cortical bone of young growing mice (4+4w), associated with reduced femoral failure force (F). However, we observed a profound capacity of bone to compensate these deleterious changes of minor nutrition with increasing age presumably via reorganization of trabecular bone. Especially in lumbar vertebrae, lifelong CR lasting 20 or 74 weeks had beneficial effects on trabecular bone mineral density (Tb.BMD), bone volume fraction (BV/TV), and trabecular number (Tb.N). In parallel, lifelong CR groups showed reduced structure model index values compared to age‐matched controls indicating a transformation of vertebral trabecular bone microarchitecture toward a platelike geometry. This effect was not visible in senile mice after short‐term 12‐week CR. In summary, CR has differential effects on cortical and trabecular bone dependent on bone localization and starting age. Our study underlines that bone compartments possess a lifelong capability to cope with changing nutritional influences.

Cementless Titanium Mesh Fixation of Osteoporotic Burst Fractures of the Lumbar Spine Leads to Bony Healing: Results of an Experimental Sheep Model

Introduction. Current treatment strategies for osteoporotic vertebral compression fractures (VCFs) focus on cement-associated solutions. Complications associated with cement application are leakage, embolism, adjacent fractures, and compromise in bony healing. This study comprises a validated VCF model in osteoporotic sheep in order to (1) evaluate a new cementless fracture fixation technique using titanium mesh implants (TMIs) and (2) demonstrate the healing capabilities in osteoporotic VCFs. Methods. Twelve 5-year-old Merino sheep received ovariectomy, corticosteroid injections, and a calcium/phosphorus/vitamin D-deficient diet for osteoporosis induction. Standardized VCFs (type AO A3.1) were created, reduced, and fixed using intravertebral TMIs. Randomly additional autologous spongiosa grafting (G1) or no augmentation was performed (G2, n = 6 each). Two months postoperatively, macroscopic, micro-CT and biomechanical evaluation assessed bony consolidation. Results. Fracture reduction succeeded in all cases without intraoperative complications. Bony consolidation was proven for all cases with increased amounts of callus development for G2 (58.3%). Micro-CT revealed cage integration. Neither group showed improved results with biomechanical testing. Conclusions. Fracture reduction/fixation using TMIs without cement in osteoporotic sheep lumbar VCF resulted in bony fracture healing. Intravertebral application of autologous spongiosa showed no beneficial effects. The technique is now available for clinical use; thus, it offers an opportunity to abandon cement-associated complications.

Cementless Fixation of Osteoporotic VCFs Using Titanium Mesh Implants (OsseoFix): Preliminary Results

Introduction. Vertebral compression fractures (VCFs) affect 20% of people over the age of 70 with increasing incidence. Kypho-/vertebroplasty as standard operative procedures are associated with limitations like cement leakage, limited reduction capabilities, and risk for adjacent fractures. To address these shortcomings, we introduce a new minimal invasive cementless VCF fixation technique. Methods. Four patients (72.3 years, range 70–76) with VCFs type AO/Müller A1.3 and concomitant osteoporosis were treated by minimal invasive transpedicular placement of two intervertebral mesh cages for fracture reduction and maintenance. Follow-up included functional/radiological assessment and clinical scores and averaged 27.7 months (24–28). Results. Endplate reduction was achieved in all cases (mean surgery time: 28.5 minutes). Kyphotic (KA) and Cobb angle revealed considerable improvements postoperatively (KA 14.5° to 10.7°/Cobb 10.1° to 8.3°). Slight loss of vertebral reduction (KA: 12.6°) and segment rekyphosis (Cobb: 10.7°) were observed for final follow-up. Pain improved from 8.8 to 2.8 (visual analogue scale). All cases showed signs of bony healing. No perioperative complications and no adjacent fractures occurred. Conclusion. Preliminary results in a small, selected patient collective indicate the ability of bony healing for osteoporotic VCFs. Cementless fixation using intravertebral titanium mesh cages revealed substantial pain relief, adequate reduction, and reduction maintenance without complications. Trial registration number is DRKS00005657, German Clinical Trials Register (DKRS).

A pan-caspase inhibitor reduces myocyte apoptosis and neuropathic pain in rats with chronic constriction injury of the sciatic nerve.

BACKGROUND:Chronic constriction injury is a widely used model for neuropathic pain in rats. It presents with symptoms resembling human neuropathic pain, such as spontaneous pain, hyperalgesia, and allodynia. Recently, myocyte apoptosis was found in neuropathic rats as a possible promoter of pain and motor dysfunction. Our aim in this study was to demonstrate whether muscle cell apoptosis contributes to neuropathic pain in this animal model. METHODS:To clarify this issue, we examined pain, nutritive perfusion, and inflammation in muscle tissue as well as myocyte apoptosis in rats with neuropathic pain established by chronic constriction injury of the sciatic nerve. Animals received either the pan-caspase inhibitor zVAD (OMe)-fmk (n = 5) or equivalent volumes of vehicle (n = 6). Sham-operated rats served as controls (n = 6). RESULTS:At day 4 after nerve ligation, there were no signs of perfusion failure or muscle tissue inflammation in all experimental groups. However, animals treated with the vehicle had marked myocyte apoptosis, which was found almost completely blocked in zVA-Dtreated animals. The zVA-Dtreated animals presented with a significant reduction of pain upon heat, cold, and mechanical stimulation comparable with values found in sham controls. CONCLUSIONS:Myocyte apoptosis possibly contributes to thermal and mechanical allodynia in this experimental model for neuropathic pain. The development of neuropathic pain symptoms did not depend on disturbances in microcirculation or muscle tissue inflammation.

Intrabody application of eptotermin alpha enhances bone formation in osteoporotic fractures of the lumbar spine; however, fails to increase biomechanical stability - results of an experimental sheep model.

Abstract This study analyses the effect of eptotermin α application into fractured vertebrae. It is hypothesized that eptotermin α is capable to enhance bony healing of the osteoporotic spine. In 10 Merino sheep osteoporosis induction was performed by ovariectomy, corticosteroid therapy and calcium/phosphorus/vitamin D-deficient diet; followed by standardized creation of lumbar vertebral compression fractures (VCFs) type A3.1 and consecutive fracture reduction/fixation using expandable mesh cages. Randomly, intravertebral eptotermin α (G1) or no augmentation was added (G2). Macroscopic, micro-CT, and biomechanical evaluation assessed bony consolidation two months postoperatively: Micro-CT data revealed bony consolidation for all cases with significant increased callus development for G2 (60%) and BV/TV (bone volume/total volume 73.45%, osteoporotic vertebrae 35.76%). Neither group showed improved biomechanical stability. Eptotermin α enhanced mineralisation in VCFs in an experimental setup with use of cementless augmentation via an expandable cage. However, higher bone mineral density did not lead to superior biomechanical properties.

The Bindex(®) ultrasound device: reliability of cortical bone thickness measures and their relationship to regional bone mineral density.

The Bindex(®) quantitative ultrasound (QUS) device is currently available and this study analyzed (I) its relative and absolute intra- and inter-session reliability and (II) the relationship between the data provided by Bindex(®)-QUS and the bone mineral density (BMD) measured by dual-energy x-ray absorptiometry at corresponding skeletal sites in young and healthy subjects (age: 25.0  ±  3.6 years). Bindex(®)-QUS calculates a density index on the basis of the thickness of cortical bone measured at the distal radius and the distal plus proximal tibia. The data show a very good relative and absolute intra- (ICC  =  0.977, CV  =  1.5%) and inter-session reliability (ICC  =  0.978, CV  =  1.4%) for the density index. The highest positive correlations were found between cortical thickness and BMD for the distal radius and distal tibia (r  ⩾  0.71, p  <  0.001). The data indicate that the Bindex(®)-QUS parameters are repeatable within and between measurement sessions. Furthermore, the measurements reflect the BMD at specific skeletal sites. Bindex(®)-QUS might be a useful tool for the measurement of skeletal adaptations.

Fibroblast Growth Factor-23, Sclerostin, and Bone Microarchitecture in Patients With Osteoporotic Fractures of the Proximal Femur: A Cross-sectional Study

This cross-sectional observational cohort study was designed to simultaneously investigate bone microarchitecture and serum markers of bone metabolism in elderly osteoporotic patients experiencing a trochanteric or femoral neck fracture. Special emphasis was put on renal function, sclerostin and fibroblast growth factor-23 (FGF-23). Eighty-two patients (median age: 84 years; 49 trochanteric fractures) scheduled for emergency surgery due to an osteoporotic fracture participated. Bone specimens for ex vivo microcomputed X-ray tomography were sampled during surgery. Blood samples for laboratory workup were collected before surgery (t0) and 1 day afterward (t1). Fifty-eight patients consented to dual-energy X-ray absorptiometry scanning of the lumbar spine and/or contralateral femoral neck after recovery during the in-patient stay. Samples were grouped according to the site of fracture. Regression coefficients were controlled for age and/or estimated glomerular filtration rate (eGFR), if appropriate. Patients experiencing a femoral neck fracture presented with better preserved renal function (eGFR) and lower C-terminal fragment of fibroblast growth factor-23 (cFGF-23) concentrations compared to those with trochanteric fractures. By contrast, serum sclerostin was similar at both time points and did not differ between groups. Age-adjusted correlation analysis revealed negative associations between eGFR and cFGF-23 determined at t1 (R=-0.34; p<0.05) as well as between eGFR and sclerostin levels at t0 (R=-0.45; p<0.05) in patients with trochanteric and femoral neck fractures, respectively. Our study provides evidence that not only an age-related decline of renal function but also the type of skeletal injury may contribute to the circulating concentrations of cFGF-23.