Human versus Rat PRF on Collagen Membranes: A Pilot Study of Mineralization in Rat Calvaria Defect Model.

Platelet-rich fibrin, the coagulated plasma fraction of blood, is commonly used to support natural healing in clinical applications. The rat calvaria defect is a standardized model to study bone regeneration. It remains, however, unclear if the rat calvaria defect is appropriate to investigate the impact of human PRF (Platelet-Rich Fibrin) on bone regeneration. To this end, we soaked Bio-Gide® collagen membranes in human or rat liquid concentrated PRF before placing them onto 5 mm calvarial defects in Sprague Dawley rats. Three weeks later, histology and micro-computed tomography (µCT) were performed. We observed that the collagen membranes soaked with rat PRF show the characteristic features of new bone and areas of mineralized collagen matrix, indicated by a median mineralized volume of 1.5 mm3 (range: 0.9; 5.3 mm3). Histology revealed new bone growing underneath the membrane and hybrid bone where collagen fibers are embedded in the new bone. Moreover, areas of passive mineralization were observed. The collagen membranes soaked with human PRF, however, were devoid of histological features of new bone formation in the center of the defect; only occasionally, new bone formed at the defect margins. Human PRF (h-PRF) caused a median bone volume of 0.9 mm3 (range: 0.3-3.3 mm3), which was significantly lower than what was observed with rat PRF (r-PRF), with a BV median of 1.2 mm3 (range: 0.3-5.9 mm3). Our findings indicate that the rat calvaria defect model is suitable for assessing the effects of rat PRF on bone formation, but caution is warranted when extrapolating conclusions regarding the efficacy of human PRF.

The mylohyoid line is highly variable but does not affect the microarchitecture of the edentulous alveolar bone - an anatomical micro-CT study.

OBJECTIVES: To evaluate in the absence of teeth the variability of the mylohyoid line (ML), the microarchitecture of the adjacent bone, and whether the variable prominence/width of the ML is associated with the quality of the adjacent bone. METHODS: µCT scans of 28 human mandibles from anatomical specimens were analyzed. The following parameters were assessed in four edentulous areas (first and second premolar (PM), first, second, and third molar (M1/2/3)): ML width, cortical thickness (CtTh), average cortical- (Avg.Ct.BV/TV), and trabecular bone volume fraction (Avg.Tb.BV/TV). RESULTS: The ML width increased from the PM towards the M2 region, which also showed the highest variance (range: 0.4-10.2 mm). The CtTh showed a decrease in the M3 region, while Avg.Ct.BV/TV and Avg.Tb.BV/TV hardly differed among the regions. In the multivariable model on the effect of the various parameters on the ML width, only gender and tooth region were significant. Specifically, male specimens were associated with a wider ML width compared to female specimens and the M2 region was associated with a wider ML width compared to the other tooth regions. CONCLUSION: The ML width was not associated with the cortical and trabecular bone quality in the adjacent bone, while gender and tooth region had a significant effect. Specifically, the ML width was lower in female, but peaked in the M2 region with a median width of 3-4 mm. CLINICAL RELEVANCE: From a clinical point of view, it was confirmed that the ML is in general a highly variable structure, especially in the M2 region, but the ML width does not allow any conclusions on the bone quality. Altogether, this underlines the need for an individual and accurate diagnostic prior to any surgical intervention.

Synergistic effect of umbilical cord extracellular vesicles and rhBMP-2 to enhance the regeneration of a metaphyseal femoral defect in osteoporotic rats.

BACKGROUND: The aim of this study was to evaluate potential synergistic effects of a single, local application of human umbilical cord MSC-derived sEVs in combination with a low dose of recombinant human rhBMP-2 to promote the regeneration of a metaphyseal femoral defect in an osteoporotic rat model. METHODS: 6 weeks after induction of osteoporosis by bilateral ventral ovariectomy and administration of a special diet, a total of 64 rats underwent a distal femoral metaphyseal osteotomy using a manual Gigli wire saw. Defects were stabilized with an adapted Y-shaped mini-locking plate and were subsequently treated with alginate only, or alginate loaded with hUC-MSC-sEVs (2 × 109), rhBMP-2 (1.5 µg), or a combination of sEVs and rhBMP-2 (n = 16 for each group). 6 weeks post-surgery, femora were evaluated by µCT, descriptive histology, and biomechanical testing. RESULTS: Native radiographs and µCT analysis confirmed superior bony union with callus formation after treatment with hUC-MSC-sEVs in combination with a low dose of rhBMP-2. This finding was further substantiated by histology, showing robust defect consolidation 6 weeks after treatment. Torsion testing of the explanted femora revealed increased stiffness after application of both, rhBMP-2 alone, or in combination with sEVs, whereas torque was only significantly increased after treatment with rhBMP-2 together with sEVs. CONCLUSION: The present study demonstrates that the co-application of hUC-MSC-sEVs can improve the efficacy of rhBMP-2 to promote the regeneration of osteoporotic bone defects.

Aged-vascular niche hinders osteogenesis of mesenchymal stem cells through paracrine repression of Wnt-axis.

Age-induced decline in osteogenic potential of bone marrow mesenchymal stem cells (BMSCs) potentiates osteoporosis and increases the risk for bone fractures. Despite epidemiology studies reporting concurrent development of vascular and bone diseases in the elderly, the underlying mechanisms for the vascular-bone cross-talk in aging are largely unknown. In this study, we show that accelerated endothelial aging deteriorates bone tissue through paracrine repression of Wnt-driven-axis in BMSCs. Here, we utilize physiologically aged mice in conjunction with our transgenic endothelial progeria mouse model (Hutchinson-Gilford progeria syndrome; HGPS) that displays hallmarks of an aged bone marrow vascular niche. We find bone defects associated with diminished BMSC osteogenic differentiation that implicate the existence of angiocrine factors with long-term inhibitory effects. microRNA-transcriptomics of HGPS patient plasma combined with aged-vascular niche analyses in progeria mice reveal abundant secretion of Wnt-repressive microRNA-31-5p. Moreover, we show that inhibition of microRNA-31-5p as well as selective Wnt-activator CHIR99021 boosts the osteogenic potential of BMSCs through de-repression and activation of the Wnt-signaling, respectively. Our results demonstrate that the vascular niche significantly contributes to osteogenesis defects in aging and pave the ground for microRNA-based therapies of bone loss in elderly.

In Vivo Engineering and Transplantation of Axially Vascularized and Epithelialized Flaps in Rats.

The arteriovenous loop (AVL) model allows the in vivo engineering of axially vascularized flaps, the so-called AVL flaps. Although AVL flaps can be transplanted microsurgically to cover tissue defects, they lack an epithelial layer on the surface. Therefore, the objective of this study was to engineer axially vascularized AVL flaps with an accompanying epithelial layer for local defect reconstruction. In this study, AVLs were established in 20 male Lewis rats. Minimally invasive injection of keratinocytes onto the surface of the AVL flaps was performed on postoperative day (POD) 21. AVL flaps were explanted from 12 rats on POD 24 or POD 30, then the epithelium formed by the keratinocytes on the surface of the flaps was evaluated using immunofluorescence staining. In six other rats, the AVL flap was locally transposed to cover a critical defect in the rats' leg on POD 30 and explanted for analysis on POD 40. In two control rats, sodium chloride was applied instead of keratinocytes. These control flaps were also transplanted on POD 30 and explanted on POD 40. Our results revealed that 3 days after keratinocyte application, a loose single-layered epithelium was observed histologically on the AVL flaps surface, whereas after 9 days, a multilayered and structured epithelium had grown. The epithelium on the transplanted AVL flaps showed its physiological differentiation when being exposed to an air-liquid interface. Histologically, a layered epithelium identical to the rats' regular skin was formed. In the sodium chloride control group, no epithelium had been grown. This study clearly demonstrates that axially vascularized AVL flaps can be processed in the subcutaneous chamber by minimally invasive injection of keratinocytes. Thus, AVL flaps with an intact epithelial layer were engineered and could be successfully transplanted for local defect coverage in a small animal model.

Active and Passive Mineralization of Bio-Gide® Membranes in Rat Calvaria Defects.

Bio-Gide® is a collagen membrane routinely used in guided bone regeneration. Recent studies have shown that this collagen membrane has osteoconductive properties, meaning that it can support the growth of new bone. However, it has also been observed that the collagen membrane has areas of mineralized fibers which can occur spontaneously and independently of osteoblasts. To better understand how this works, we established a model using minced collagen membranes to reduce the active mineralization of intact collagen membranes in favor of passive mineralization. We thus compared the original intact membrane with a minced collagen membrane in a 5 mm calvarial defect model in Sprague Dawley rats. After three weeks of healing, histology and microcomputed tomography (µCT) were performed. Histological analysis confirmed the osteoconductive properties, with new bone growing inside the intact collagen membrane. However, in minced collagen membranes, the osteoconductive properties were restricted to the defect margins. Interestingly, histology revealed large mineralized areas indicating passive mineralization with no signs of bone formation. In the µCT analysis, the intact collagen membranes caused a higher median mineralized volume (1.5 mm3) compared with the minced group (0.4 mm3), but this lacked significance (p = 0.09). The µCT analysis needs to be interpreted carefully, particularly in defects filled with minced membranes, considering that the mineralized tissue may not necessarily be bone but also the result of passive mineralization. Taken together, the findings suggest that Bio-Gide® collagen membranes support bone formation while also exhibiting potential for passive mineralization.

Comparison of Decellularized Human Dermal Scaffolds versus Bovine Collagen/Elastin Matrices for Engineering of Soft-Tissue Flaps.

BACKGROUND: Free flap-based soft-tissue reconstruction comes at the price of donor-site morbidity. The arteriovenous loop (AVL) technique can overcome this issue by allowing for the de novo generation of axially vascularized soft-tissue flaps from vein grafts embedded into different matrices. Application of the AVL technique has been limited by insufficient long-term volume retention and poor tissue stability. The authors investigated the suitability of a novel human dermal scaffold to improve volume retention and tissue stability. METHODS: AVLs were created in 28 immunocompetent rats and embedded in either decellularized human dermal scaffolds (experimental group, n = 14) (Epiflex) or bovine collagen/elastin matrices (control group, n = 14) (MatriDerm) in subcutaneous polytetrafluoroethylene chambers. The weight and volume of engineered tissues, the extent of angiogenesis, and the proportion of proliferating cells were compared between groups on postoperative days (PODs) 21 and 28 by means of immunohistochemistry and micro-computed tomography. RESULTS: On POD 28, both groups displayed homogeneous microvascular networks on histopathology and micro-computed tomography. Mean microvessel counts and surface areas and the percentage of proliferating cells did not differ between the groups. However, the experimental human scaffold group displayed significantly smaller volume loss and significantly less tissue degradation compared with bovine matrix controls (volume retention, 102% ± 5% versus 27% ± 7% on POD 21, and 79% ± 12% versus 12% ± 7% on POD 28, respectively; P < 0.0001). CONCLUSION: Compared with bovine matrices, decellularized human scaffolds allow for superior volume retention and tissue stability of de novo engineered soft-tissue AVL flaps in rats. CLINICAL RELEVANCE STATEMENT: AVLs allow for the de novo generation of vascularized soft-tissue flaps. However, insufficient long-term volume retention is still an issue. The authors' study shows that decellularized human matrices guarantee superior volume stability of de novo grown soft-tissue flaps in rats.

Lateral angle: A landmark-based method for the sex estimation in human cremated remains and application to an Austrian prehistoric sample.

OBJECTIVES: Estimating the sex of cremated human remains is difficult. The petrous bone frequently survives the cremation due to its density. Wahl observed the lateral angle to be sexually dimorphic in the 1980s. Previous studies showed various cut-off points to separate females from males, which are hardly replicable and difficult to apply. We want to test the Wahl method and compare it to a new landmark-based version. MATERIALS AND METHODS: In this study, we measured the lateral angle of 35 cremated petrous bones from late bronze age Austria using micro-CT scans. Technical errors of measurement were calculated for two different methods to intersect the internal acoustic meatus virtually in the midline (manual or landmark-based intersection). Furthermore, sex was estimated based on morphological features and metric measurements. This information was used in logistic regression modeling to define a cut-off point in our sample. RESULTS: The technical errors of measurement suggested that a landmark-based method was more precise in comparison to a manual intersection which was much more intuitive. Inter- and intra-observer errors were low which improved reliability. The logistic regression model produced good results in our sample (p = 0.02, R2 = 0.38, accuracy = 0.8). The mean lateral angle was similar to studies which focused on prehistoric cremated petrous bones. DISCUSSION: The proposed landmark-based method was precise, quick, and could be easily applied, even by unexperienced researchers. The size of the lateral angle seemed to be population-specific but also dependent on the method applied. We recommend to use the proposed landmark-based method which is more precise.

The effect of osteocyte-derived RANKL on bone graft remodeling: An in vivo experimental study.

OBJECTIVES: Autologous bone is considered the gold standard for grafting, yet it suffers from a tendency to undergo resorption over time. While the exact mechanisms of this resorption remain elusive, osteocytes have been shown to play an important role in stimulating osteoclastic activity through their expression of receptor activator of NF-κB (RANK) ligand (RANKL). The aim of this study was to assess the function of osteocyte-derived RANKL in bone graft remodeling. MATERIALS AND METHODS: In Tnfsf11fl/fl ;Dmp1-Cre mice without osteocyte-specific RANKL as well as in Dmp1-Cre control mice, 2.6 mm calvarial bone disks were harvested and transplanted into mice with matching genetic backgrounds either subcutaneously or subperiosteally, creating 4 groups in total. Histology and micro-computed tomography of the grafts and the donor regions were performed 28 days after grafting. RESULTS: Histology revealed marked resorption of subcutaneous control Dmp1-Cre grafts and new bone formation around subperiosteal Dmp1-Cre grafts. In contrast, Tnfsf11fl/fl ;Dmp1-Cre grafts showed effectively neither signs of bone resorption nor formation. Quantitative micro-computed tomography revealed a significant difference in residual graft area between subcutaneous and subperiosteal Dmp1-Cre grafts (p < .01). This difference was not observed between subcutaneous and subperiosteal Tnfsf11fl/fl ;Dmp1-Cre grafts (p = .17). Residual graft volume (p = .08) and thickness (p = .13) did not differ significantly among the groups. Donor area regeneration was comparable between Tnfsf11fl/fl ;Dmp1-Cre and Dmp1-Cre mice and restricted to the defect margins. CONCLUSIONS: The results suggest an active function of osteocyte-derived RANKL in bone graft remodeling.

Corrigendum: Mapping the functional anatomy and topography of the cardiac autonomic innervation for selective cardiac neuromodulation using MicroCT.

[This corrects the article DOI: 10.3389/fcell.2022.968870.].

The p-rpS6-zone delineates wounding responses and the healing process.

The spatial boundaries of tissue response to wounding are unknown. Here, we show that in mammals, the ribosomal protein S6 (rpS6) is phosphorylated in response to skin injury, forming a zone of activation surrounding the region of the initial insult. This p-rpS6-zone forms within minutes after wounding and is present until healing is complete. The zone is a robust marker of healing as it encapsulates features of the healing process, including proliferation, growth, cellular senescence, and angiogenesis. A mouse model that is unable to phosphorylate rpS6 shows an initial acceleration of wound closure, but results in impaired healing, identifying p-rpS6 as a modulator but not a driver of healing. Finally, the p-rpS6-zone accurately reports on the status of dermal vasculature and the effectiveness of healing, visually dividing an otherwise homogeneous tissue into regions with distinct properties.

Acellular Human Placenta Small-Diameter Vessels as a Favorable Source of Super-Microsurgical Vascular Replacements: A Proof of Concept.

In this study, we aimed to evaluate the human placenta as a source of blood vessels that can be harvested for vascular graft fabrication in the submillimeter range. Our approach included graft modification to prevent thrombotic events. Submillimeter arterial grafts harvested from the human placenta were decellularized and chemically crosslinked to heparin. Graft performance was evaluated using a microsurgical arteriovenous loop (AVL) model in Lewis rats. Specimens were evaluated through hematoxylin-eosin and CD31 staining of histological sections to analyze host cell immigration and vascular remodeling. Graft patency was determined 3 weeks after implantation using a vascular patency test, histology, and micro-computed tomography. A total of 14 human placenta submillimeter vessel grafts were successfully decellularized and implanted into AVLs in rats. An appropriate inner diameter to graft length ratio of 0.81 ± 0.16 mm to 7.72 ± 3.20 mm was achieved in all animals. Grafts were left in situ for a mean of 24 ± 4 days. Decellularized human placental grafts had an overall patency rate of 71% and elicited no apparent immunological responses. Histological staining revealed host cell immigration into the graft and re-endothelialization of the vessel luminal surface. This study demonstrates that decellularized vascular grafts from the human placenta have the potential to serve as super-microsurgical vascular replacements.

Functionalizing Collagen Membranes with MSC-Conditioned Media Promotes Guided Bone Regeneration in Rat Calvarial Defects.

Functionalizing biomaterials with conditioned media (CM) from mesenchymal stromal cells (MSC) is a promising strategy for enhancing the outcomes of guided bone regeneration (GBR). This study aimed to evaluate the bone regenerative potential of collagen membranes (MEM) functionalized with CM from human bone marrow MSC (MEM-CM) in critical size rat calvarial defects. MEM-CM prepared via soaking (CM-SOAK) or soaking followed by lyophilization (CM-LYO) were applied to critical size rat calvarial defects. Control treatments included native MEM, MEM with rat MSC (CEL) and no treatment. New bone formation was analyzed via micro-CT (2 and 4 weeks) and histology (4 weeks). Greater radiographic new bone formation occurred at 2 weeks in the CM-LYO group vs. all other groups. After 4 weeks, only the CM-LYO group was superior to the untreated control group, whereas the CM-SOAK, CEL and native MEM groups were similar. Histologically, the regenerated tissues showed a combination of regular new bone and hybrid new bone, which formed within the membrane compartment and was characterized by the incorporation of mineralized MEM fibers. Areas of new bone formation and MEM mineralization were greatest in the CM-LYO group. Proteomic analysis of lyophilized CM revealed the enrichment of several proteins and biological processes related to bone formation. In summary, lyophilized MEM-CM enhanced new bone formation in rat calvarial defects, thus representing a novel 'off-the-shelf' strategy for GBR.

Impact of a Static Magnetic Field on Early Osseointegration: A Pilot Study in Canines.

A static magnetic field generated by neodymium-iron-boron (NdFeB) magnets placed in the inner cavity of dental implants can enhance bone regeneration in rabbits. It is, however, unknown whether static magnetic fields support osseointegration in a canine model. We therefore determined the potential osteogenic effect of implants carrying NdFeB magnets inserted in the tibia of six adult canines in the early stages of osseointegration. Here, we report that after 15 days of healing, magnetic and regular implants showed a high variation with a median new bone-to-implant contact (nBIC) in the cortical (41.3% and 7.3%) and the medullary (28.6% and 44.8%) region, respectively. Consistently, the median new bone volume/tissue volume (nBV/TV) in the cortical (14.9% and 5.4%) and the medullary (22.2% and 22.4%) region were not significantly different. One week of healing only resulted in negligible bone formation. These findings suggest that considering the large variation and the pilot nature of this study, magnetic implants failed to support peri-implant bone formation in a canine model.

FasL is a catabolic factor in alveolar bone homeostasis.

AIM: Fas ligand (FasL) belongs to the tumour necrosis factor superfamily regulating bone turnover, inflammation, and apoptosis. The appendicular and axial skeleton phenotype of mature Faslgld mice has been reported. The impact of FasL on the alveolar bone providing support for the teeth at mature stages under healthy and induced inflammatory conditions remains unknown. MATERIALS AND METHODS: We performed a phenotypical analysis of mice carrying the homozygous Faslgld mutation and wild-type (WT) mice (C57BL/6) under healthy conditions and upon ligature-induced periodontitis. After 12 days, micro-computed tomography analysis revealed the distance between the cement enamel junction and the alveolar bone crest. Additional structural parameters, such as the bone volume fraction (BV/TV) and the periodontal ligament space volume, were measured. Histological analyses were performed to visualize the catabolic changes at the defect site. RESULTS: Healthy Faslgld mice were found to have more periodontal bone than their WT littermates. Faslgld had no significant effect on inflammatory osteolysis compared to WT controls with ligatures. Histology revealed eroded surfaces at the root and in the inter-proximal bone in both strains. CONCLUSIONS: Our findings suggest that FasL is a catabolic factor in alveolar bone homeostasis but it does not affect the inflammatory osteolysis.

Effects of Endochondral and Intramembranous Ossification Pathways on Bone Tissue Formation and Vascularization in Human Tissue-Engineered Grafts.

Bone grafts can be engineered by differentiating human mesenchymal stromal cells (MSCs) via the endochondral and intramembranous ossification pathways. We evaluated the effects of each pathway on the properties of engineered bone grafts and their capacity to drive bone regeneration. Bone-marrow-derived MSCs were differentiated on silk scaffolds into either hypertrophic chondrocytes (hyper) or osteoblasts (osteo) over 5 weeks of in vitro cultivation, and were implanted subcutaneously for 12 weeks. The pathways' constructs were evaluated over time with respect to gene expression, composition, histomorphology, microstructure, vascularization and biomechanics. Hypertrophic chondrocytes expressed higher levels of osteogenic genes and deposited significantly more bone mineral and proteins than the osteoblasts. Before implantation, the mineral in the hyper group was less mature than that in the osteo group. Following 12 weeks of implantation, the hyper group had increased mineral density but a similar overall mineral composition compared with the osteo group. The hyper group also displayed significantly more blood vessel infiltration than the osteo group. Both groups contained M2 macrophages, indicating bone regeneration. These data suggest that, similar to the body's repair processes, endochondral pathway might be more advantageous when regenerating large defects, whereas intramembranous ossification could be utilized to guide the tissue formation pattern with a scaffold architecture.

Bone Graft Packing and Its Association with Bone Regeneration in Maxillary Sinus Floor Augmentations: Histomorphometric Analysis of Human Biopsies.

Research in maxillary sinus floor augmentation (MSFA) focussed on the optimisation of microstructural parameters such as microporosity and particle size of bone substitute particles (BS). However, little is known about the impact of BS packing and the corresponding (void) interparticular space on bone regeneration. The aim of this study was to characterise the spatial distribution of BS and its association with BS integration 6 ± 1 months after MSFA. Histological thin-ground sections of 70 human sinus biopsies were histomorphometrically analysed: In serial zones of 100 µm proceeding from the sinus floor (SF) up to the apical end of the biopsy, we measured the distribution of BS particles within these zones in terms of volume (BSV/TV), number and size of BS particles, interparticle spacing (BS.Sp) and bone-to-BS contact. BS particles were not homogeneously distributed over the length of biopsies: The first 200 µm directly adjacent to the SF represented a zone poor in BS particles but with high osteogenic potential. Graft packing density increased from the SF towards the apical part of the AA. Integration of BS particles was inversely associated with the distance to the SF and the graft packing density. A high packing density through excessive compaction of BS particles should be avoided to optimise the macrostructural environment for bone regeneration.

Mapping the functional anatomy and topography of the cardiac autonomic innervation for selective cardiac neuromodulation using MicroCT.

Background: Vagus nerve stimulation (VNS) has gained great importance as a promising therapy for a myriad of diseases. Of particular interest is the therapy of cardiovascular diseases, such as heart failure or atrial fibrillation using selective cardiac VNS. However, there is still a lack of organ-specific anatomical knowledge about the fascicular anatomy and topography of the cardiac branch (CB), which diminishes the therapeutic possibilities for selective cardiac neuromodulation. Here, we established a topographical and anatomical map of the superior cardiac VN in two animal species to dissect cervical and cardiac VN morphology. Methods: Autonomic nerves including superior CBs were harvested from domestic pigs and New Zeeland rabbits followed by imaging with microcomputed tomography (µCT) and 3D rendering. The data were analyzed in terms of relevant topographical and anatomical parameters. Results: Our data showed that cardiac vagal fascicles remained separated from other VN fascicles up to 22.19 mm (IQR 14.02-41.30 mm) in pigs and 7.68 mm (IQR 4.06-12.77 mm) in rabbits from the CB point and then started merging with other fascicles. Exchanges of nerve fascicles between sympathetic trunk (ST) and VN were observed in 3 out of 11 nerves, which might cause additional unwanted effects in unselective VNS. Our 3D rendered digital model of the cardiac fascicles was generated showing that CB first remained on the medial side where it branched off the VN, as also shown in the µCT data of 11 pig nerves, and then migrated towards the ventromedial site the further it was traced cranially. Conclusion: Our data provided an anatomical map of the cardiac vagal branches including cervical VN and ST for future approaches of selective cardiac neurostimulation, indicating the best position of selective cardiac VNS just above the CB point.

Effects of Extracorporeal Shockwave Therapy on Functional Recovery and Circulating miR-375 and miR-382-5p after Subacute and Chronic Spinal Cord Contusion Injury in Rats.

Extracorporeal shockwave therapy (ESWT) can stimulate processes to promote regeneration, including cell proliferation and modulation of inflammation. Specific miRNA expression panels have been established to define correlations with regulatory targets within these pathways. This study aims to investigate the influence of low-energy ESWT-applied within the subacute and chronic phase of SCI (spinal cord injury) on recovery in a rat spinal cord contusion model. Outcomes were evaluated by gait analysis, µCT and histological analysis of spinal cords. A panel of serum-derived miRNAs after SCI and after ESWT was investigated to identify injury-, regeneration- and treatment-associated expression patterns. Rats receiving ESWT showed significant improvement in motor function in both a subacute and a chronic experimental setting. This effect was not reflected in changes in morphology, µCT-parameters or histological markers after ESWT. Expression analysis of various miRNAs, however, revealed changes after SCI and ESWT, with increased miR-375, indicating a neuroprotective effect, and decreased miR-382-5p potentially improving neuroplasticity via its regulatory involvement with BDNF. We were able to demonstrate a functional improvement of ESWT-treated animals after SCI in a subacute and chronic setting. Furthermore, the identification of miR-375 and miR-382-5p could potentially provide new targets for therapeutic intervention in future studies.

Improved biomechanics in experimental chronic rotator cuff repair after shockwaves is not reflected by bone microarchitecture.

PURPOSE: The aim of this study was to investigate the effect of extracorporeal shockwave therapy (ESWT) on bone microstructure as well as the bone-tendon-interface and the musculo-tendinous transition zone to explain the previously shown improved biomechanics in a degenerative rotator cuff tear animal model. This study hypothesized that biomechanical improvements related to ESWT are a result of improved bone microstructure and muscle tendon properties. METHODS: In this controlled laboratory study unilateral supraspinatus (SSP) tendon detachment was performed in 48 male Sprague-Dawley rats. After a degeneration period of three weeks, SSP tendon was reconstructed transosseously. Rats were randomly assigned into three groups (n = 16 per group): control (noSW); intraoperative shockwave treatment (IntraSW); intra- and postoperative shockwave treatment (IntraPostSW). Eight weeks after SSP repair, all rats were sacrificed and underwent bone microstructure analysis as well as histological and immunohistochemical analyses. RESULTS: With exception of cortical porosity at the tendon area, bone microstructure analyses revealed no significant differences between the three study groups regarding cortical and trabecular bone parameters. Cortical Porosity at the Tendon Area was lowest in the IntraPostSW (p≤0.05) group. Histological analyses showed well-regenerated muscle and tendon structures in all groups. Immunohistochemistry detected augmented angiogenesis at the musculo-tendinous transition zone in both shockwave groups indicated by CD31 positive stained blood vessels. CONCLUSION: In conclusion, bone microarchitecture changes are not responsible for previously described improved biomechanical results after shockwave treatment in rotator cuff repair in rodents. Immunohistochemical analysis showed neovascularization at the musculo-tendinous transition zone within ESWT-treated animals. Further studies focusing on neovascularization at the musculo-tendinous transition zone are necessary to explain the enhanced biomechanical and functional properties observed previously. CLINICAL RELEVANCE: In patients treated with a double-row SSP tendon repair, an improvement in healing through ESWT, especially in this area, could prevent a failure of the medial row, which is considered a constantly observed tear pattern.