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Article ; Online: Histological Preparation and Evaluation of Cartilage Specimens.

Darwiche, Salim E / Tegelkamp, Milena / Nuss, Katja / von Rechenberg, Brigitte

Methods in molecular biology (Clifton, N.J.)

2022 Volume 2598, Page(s) 227–263

Abstract: In this chapter, an introduction is given into histological techniques to research related to hyaline cartilage and subchondral bone. Emphasis is placed on the importance to investigate cartilage and bone as a unit, which includes the transition zone of ... ...

Abstract	In this chapter, an introduction is given into histological techniques to research related to hyaline cartilage and subchondral bone. Emphasis is placed on the importance to investigate cartilage and bone as a unit, which includes the transition zone of the calcified cartilage and tidemark. Reasons for the appropriate selection of histological methods are presented such as when to use (decalcified) specimens for routine paraffin embedding including immunohistology, cryosections of cartilage alone, or non-decalcified specimens for embedding in polymethylmethacrylate with or without additional biomaterials. Appropriate staining methods are also outlined. Apart from detailed laboratory protocols for different embedding and staining methods including open communication about difficulties related to the various techniques, also practical instructions for state-of-the-art evaluation methods and their strengths and weaknesses are given. Sample figures for scoring methods are included.
MeSH term(s)	Cartilage, Articular/pathology ; Bone and Bones
Language	English
Publishing date	2022-11-10
Publishing country	United States
Document type	Journal Article
ISSN	1940-6029
ISSN (online)	1940-6029
DOI	10.1007/978-1-0716-2839-3_17
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Article ; Online: Studying Edema Formation After Release of the Infraspinatus Muscle as an Experimental Model of Rotator Cuff Lesions in Sheep: A Histological Analysis.

von Rechenberg, Brigitte / Gehrke, Rieke S / Klein, Karina / Kronen, Peter / Darwiche, Salim / Zbinden, Jeanni / Wieser, Karl / Lädermann, Alexandre

The American journal of sports medicine

2024 Volume 52, Issue 5, Page(s) 1319–1327

Abstract: Background: Muscle edema formation and inflammatory processes are early manifestations of acute rotator cuff lesions in sheep. Histological analysis of affected muscles revealed edema formation, inflammatory changes, and muscle tissue disruption in MRs.! ...

Abstract	Background: Muscle edema formation and inflammatory processes are early manifestations of acute rotator cuff lesions in sheep. Histological analysis of affected muscles revealed edema formation, inflammatory changes, and muscle tissue disruption in MRs. Hypothesis: Edema contributes to inflammatory reactions and early muscle fiber degeneration before the onset of fatty infiltration. Study design: Controlled laboratory study. Methods: Osteotomy of the greater tuberosity, including the insertion of the infraspinatus tendon, was performed on 14 sheep. These experimental animal models were divided into 2 groups: a nontrauma group with surgical muscle release alone (7 sheep) and a trauma group with standardized application of additional trauma to the musculotendinous unit (7 sheep). Excisional biopsy specimens of the infraspinatus muscle were taken at 0, 3, and 4 weeks. Results: Edema formation was histologically demonstrated in both groups and peaked at 3 weeks. At 3 weeks, signs of muscle fiber degeneration were observed. At 4 weeks, ingrowth of loose alveolar and fibrotic tissue between fibers was detected. Fatty tissue was absent. The diameter of muscle fibers increased in both groups, albeit to a lesser degree in the trauma group, and practically normalized at 4 weeks. Immunohistology revealed an increase in macrophage types 1 and 2, as well as inflammatory mediators such as prostaglandin E2 and nuclear factor kappa-light-chain-enhancer of activated B cells. Conclusion: Early muscle edema and concomitant inflammation precede muscle fiber degeneration and fibrosis. Edema formation results from tendon release alone and is only slightly intensified by additional trauma. Clinical relevance: This study illustrates that early edema formation and inflammation elicit muscle fiber degeneration that precedes fatty infiltration. Should this phenomenon be applicable to human traumatic rotator cuff tears, then surgery should be performed as soon as possible, ideally within the first 21 days after injury.
MeSH term(s)	Humans ; Animals ; Sheep ; Rotator Cuff/surgery ; Rotator Cuff Injuries/pathology ; Tendon Injuries/surgery ; Models, Theoretical ; Inflammation/pathology ; Adipose Tissue/pathology
Language	English
Publishing date	2024-03-08
Publishing country	United States
Document type	Journal Article
ZDB-ID	197482-8
ISSN	1552-3365 ; 0363-5465
ISSN (online)	1552-3365
ISSN	0363-5465
DOI	10.1177/03635465241226961
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Article ; Online: Biomechanical duality of fracture healing captured using virtual mechanical testing and validated in ovine bones.

Inglis, Brendan / Schwarzenberg, Peter / Klein, Karina / von Rechenberg, Brigitte / Darwiche, Salim / Dailey, Hannah L

Scientific reports

2022 Volume 12, Issue 1, Page(s) 2492

Abstract: Bone fractures commonly repair by forming a bridging structure called callus, which begins as soft tissue and gradually ossifies to restore rigidity to the bone. Virtual mechanical testing is a promising technique for image-based assessment of structural ...

Abstract	Bone fractures commonly repair by forming a bridging structure called callus, which begins as soft tissue and gradually ossifies to restore rigidity to the bone. Virtual mechanical testing is a promising technique for image-based assessment of structural bone healing in both preclinical and clinical settings, but its accuracy depends on the validity of the material model used to assign tissue mechanical properties. The goal of this study was to develop a constitutive model for callus that captures the heterogeneity and biomechanical duality of the callus, which contains both soft tissue and woven bone. To achieve this, a large-scale optimization analysis was performed on 2363 variations of 3D finite element models derived from computed tomography (CT) scans of 33 osteotomized sheep under normal and delayed healing conditions. A piecewise material model was identified that produced high absolute agreement between virtual and physical tests by differentiating between soft and hard callus based on radiodensity. The results showed that the structural integrity of a healing long bone is conferred by an internal architecture of mineralized hard callus that is supported by interstitial soft tissue. These findings suggest that with appropriate material modeling, virtual mechanical testing is a reliable surrogate for physical biomechanical testing.
MeSH term(s)	Animals ; Biomechanical Phenomena ; Bone and Bones/diagnostic imaging ; Bone and Bones/physiology ; Connective Tissue/diagnostic imaging ; Connective Tissue/physiology ; Finite Element Analysis ; Fracture Healing/physiology ; Fractures, Bone/physiopathology ; Mechanical Tests/methods ; Osteogenesis/physiology ; Sheep ; Tomography, X-Ray Computed/methods
Language	English
Publishing date	2022-02-15
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Research Support, U.S. Gov't, Non-P.H.S.
ZDB-ID	2615211-3
ISSN	2045-2322 ; 2045-2322
ISSN (online)	2045-2322
ISSN	2045-2322
DOI	10.1038/s41598-022-06267-8
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Article ; Online: Image-based radiodensity profilometry measures early remodeling at the bone-callus interface in sheep.

Ren, Tianyi / Klein, Karina / von Rechenberg, Brigitte / Darwiche, Salim / Dailey, Hannah L

Biomechanics and modeling in mechanobiology

2022 Volume 21, Issue 2, Page(s) 615–626

Abstract: Bone healing has been traditionally described as a four-phase process: inflammatory response, soft callus formation, hard callus development, and remodeling. The remodeling phase has been largely neglected in most numerical mechanoregulation models of ... ...

Abstract	Bone healing has been traditionally described as a four-phase process: inflammatory response, soft callus formation, hard callus development, and remodeling. The remodeling phase has been largely neglected in most numerical mechanoregulation models of fracture repair in favor of capturing early healing using a pre-defined callus domain. However, in vivo evidence suggests that remodeling occurs concurrently with repair and causes changes in cortical bone adjacent to callus that are typically neglected in numerical models of bone healing. The objective of this study was to use image processing techniques to quantify this early-stage remodeling in ovine osteotomies. To accomplish this, we developed a numerical method for radiodensity profilometry with optimization-based curve fitting to mathematically model the bone density gradients in the radial direction across the cortical wall and callus. After assessing data from 26 sheep, we defined a dimensionless density fitting function that revealed significant remodeling occurring in the cortical wall adjacent to callus during early healing, a 23% average reduction in density compared to intact. This fitting function is robust for modeling radial density gradients in both intact bone and fracture repair scenarios and can capture a wide variety of the healing responses. The fitting function can also be scaled easily for comparison to numerical model predictions and may be useful for validating future mechanoregulatory models of coupled fracture repair and remodeling.
MeSH term(s)	Animals ; Bony Callus/diagnostic imaging ; Fracture Healing/physiology ; Fractures, Bone ; Sheep
Language	English
Publishing date	2022-01-08
Publishing country	Germany
Document type	Journal Article
ZDB-ID	2093052-5
ISSN	1617-7940 ; 1617-7959
ISSN (online)	1617-7940
ISSN	1617-7959
DOI	10.1007/s10237-021-01553-2
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Article ; Online: Evaluation of the Potential of Umbilical Cord Mesenchymal Stromal Cell-Derived Small Extracellular Vesicles to Improve Rotator Cuff Healing: A Pilot Ovine Study.

Jenner, Florien / Wagner, Andrea / Gerner, Iris / Ludewig, Eberhard / Trujanovic, Robert / Rohde, Eva / von Rechenberg, Brigitte / Gimona, Mario / Traweger, Andreas

The American journal of sports medicine

2023 Volume 51, Issue 2, Page(s) 331–342

Abstract: Background: Despite significant advancements in surgical techniques to repair rotator cuff (RC) injuries, failure rates remain high and novel approaches to adequately overcome the natural biological limits of tendon and enthesis regeneration of the RC ... ...

Abstract	Background: Despite significant advancements in surgical techniques to repair rotator cuff (RC) injuries, failure rates remain high and novel approaches to adequately overcome the natural biological limits of tendon and enthesis regeneration of the RC are required. Small extracellular vesicles (sEVs) derived from the secretome of human multipotent mesenchymal stromal cells (MSCs) have been demonstrated to modulate inflammation and reduce fibrotic adhesions, and therefore their local application could improve outcomes after RC repair. Purpose: In this pilot study, we evaluated the efficacy of clinical-grade human umbilical cord (hUC) MSC-derived sEVs (hUC-MSC-sEVs) loaded onto a type 1 collagen scaffold in an ovine model of acute infraspinatus tendon injury to improve RC healing. Study design: Controlled laboratory study. Methods: sEVs were enriched from hUC-MSC culture media and were characterized by surface marker profiling. The immunomodulatory capacity was evaluated in vitro by T-cell proliferation assays, and particle count was determined by nanoparticle tracking analysis. Twelve skeletally mature sheep were subjected to partial infraspinatus tenotomy and enthesis debridement. The defects of 6 animals were treated with 2 × 10 Results: CD3/CD28-stimulated T-cell proliferation was significantly inhibited by hUC-MSC-sEVs ( Conclusion: A local, single-dose application of hUC-MSC-sEVs promoted tendon and enthesis healing in an ovine model of acute RC injury. Clinical relevance: Surgical repair of RC tears generally results in a clinical benefit for the patient; however, considerable rerupture rates have been reported. sEVs have potential as a cell-free biotherapeutic to improve healing outcomes after RC injury.
MeSH term(s)	Animals ; Sheep ; Humans ; Rotator Cuff/surgery ; Pilot Projects ; Collagen Type I/metabolism ; Rotator Cuff Injuries/surgery ; Mesenchymal Stem Cells/metabolism ; Extracellular Vesicles/pathology ; Umbilical Cord
Chemical Substances	Collagen Type I
Language	English
Publishing date	2023-01-16
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	197482-8
ISSN	1552-3365 ; 0363-5465
ISSN (online)	1552-3365
ISSN	0363-5465
DOI	10.1177/03635465221145958
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Article ; Online: Rotator Cuff Repair and Overlay Augmentation by Direct Interlocking of a Nonwoven Polyethylene Terephthalate Patch Into the Tendon: Evaluation in an Ovine Model.

Meyer, Dominik C / Bachmann, Elias / Darwiche, Salim / Moehl, Andrea / von Rechenberg, Brigitte / Gerber, Christian / Snedeker, Jess G

The American journal of sports medicine

2023 Volume 51, Issue 12, Page(s) 3235–3242

Abstract: Background: Arthroscopic repair of large rotator cuff tendon tears is associated with high rates of retear. Construct failure often occurs at the suture-tendon interface. Patch augmentation can improve mechanical strength and healing at this interface.!# ...

Abstract	Background: Arthroscopic repair of large rotator cuff tendon tears is associated with high rates of retear. Construct failure often occurs at the suture-tendon interface. Patch augmentation can improve mechanical strength and healing at this interface. Purpose: To introduce a novel technique for suture-free attachment of an overlaid patch and evaluate its biomechanical strength and biological performance. Study design: Descriptive and controlled laboratory studies. Methods: An established ovine model of partial infraspinatus tendon resection and immediate repair was used. After a nonwoven polyethylene terephthalate patch was overlaid to the resected tendon, a barbed microblade was used to draw fibers of the patch directly into the underlying tissue. In vivo histological assessment of healing was performed at 6 and 13 weeks after implantation. Ex vivo models were used to characterize primary repair strength of the suture-free patch fixation to tendon. Additional ex vivo testing assessed the potential of the technique for patch overlay augmentation of suture-based repair. Results: The in vivo study revealed no macroscopic evidence of adverse tissue reactions to the interlocked patch fibers. Histological testing indicated a normal host healing response with minimal fibrosis. Uniform and aligned tissue ingrowth to the core of the patch was observed from both the tendon and the bone interfaces to the patch. There was no evident retraction of the infraspinatus muscle, lengthening of the tendon, or tendon gap formation over 13 weeks. Ex vivo testing revealed that direct patch interlocking yielded tendon purchase equivalent to a Mason-Allen suture (150 ± 58 vs 154 ± 49 N, respectively; Conclusion: Testing in an ovine model of rotator cuff tendon repair suggested that surgical interlocking of a nonwoven medical textile can provide effective biomechanical performance, support functional tissue ingrowth, and help avoid musculotendinous retraction after surgical tendon repair. Clinical relevance: The novel technique may facilitate patch augmentation of rotator cuff repairs.
MeSH term(s)	Sheep ; Animals ; Humans ; Rotator Cuff/pathology ; Polyethylene Terephthalates ; Rotator Cuff Injuries ; Tendons/surgery ; Orthopedic Procedures ; Suture Techniques ; Biomechanical Phenomena
Chemical Substances	Polyethylene Terephthalates
Language	English
Publishing date	2023-09-08
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	197482-8
ISSN	1552-3365 ; 0363-5465
ISSN (online)	1552-3365
ISSN	0363-5465
DOI	10.1177/03635465231189802
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Article: Poly(ADP-Ribose) Polymerases-Inhibitor Talazoparib Inhibits Muscle Atrophy and Fatty Infiltration in a Tendon Release Infraspinatus Sheep Model: A Pilot Study.

Olthof, Maurits G L / Hasler, Anita / Valdivieso, Paola / Flück, Martin / Gerber, Christian / Gehrke, Rieke / Klein, Karina / von Rechenberg, Brigitte / Snedeker, Jess G / Wieser, Karl

Metabolites

2024 Volume 14, Issue 4

Abstract: Structural muscle changes, including muscle atrophy and fatty infiltration, follow rotator cuff tendon tear and are associated with a high repair failure rate. Despite extensive research efforts, no pharmacological therapy is available to successfully ... ...

Abstract	Structural muscle changes, including muscle atrophy and fatty infiltration, follow rotator cuff tendon tear and are associated with a high repair failure rate. Despite extensive research efforts, no pharmacological therapy is available to successfully prevent both muscle atrophy and fatty infiltration after tenotomy of tendomuscular unit without surgical repair. Poly(ADP-ribose) polymerases (PARPs) are identified as a key transcription factors involved in the maintenance of cellular homeostasis. PARP inhibitors have been shown to influence muscle degeneration, including mitochondrial hemostasis, oxidative stress, inflammation and metabolic activity, and reduced degenerative changes in a knockout mouse model. Tenotomized infraspinatus were assessed for muscle degeneration for 16 weeks using a Swiss Alpine sheep model (n = 6). All sheep received daily oral administration of 0.5 mg Talazoparib. Due to animal ethics, the treatment group was compared with three different controls from prior studies of our institution. To mitigate potential batch heterogeneity, PARP-I was evaluated in comparison with three distinct control groups (n = 6 per control group) using the same protocol without treatment. The control sheep were treated with an identical study protocol without Talazoparib treatment. Muscle atrophy and fatty infiltration were evaluated at 0, 6 and 16 weeks post-tenotomy using DIXON-MRI. The controls and PARP-I showed a significant (control
Language	English
Publishing date	2024-03-26
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2662251-8
ISSN	2218-1989
ISSN	2218-1989
DOI	10.3390/metabo14040187
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Article ; Online: Combination of a Collagen Scaffold and an Adhesive Hyaluronan-Based Hydrogel for Cartilage Regeneration: A Proof of Concept in an Ovine Model.

Levinson, Clara / Cavalli, Emma / von Rechenberg, Brigitte / Zenobi-Wong, Marcy / Darwiche, Salim E

Cartilage

2021 Volume 13, Issue 2_suppl, Page(s) 636S–649S

Abstract: Objective: Hyaluronic acid-transglutaminase (HA-TG) is an enzymatically crosslinkable adhesive hydrogel with chondrogenic properties demonstrated : Design: Chondral defects of 6-mm diameter were created in the stifle joint of skeletally mature sheep. ...

Abstract	Objective: Hyaluronic acid-transglutaminase (HA-TG) is an enzymatically crosslinkable adhesive hydrogel with chondrogenic properties demonstrated Design: Chondral defects of 6-mm diameter were created in the stifle joint of skeletally mature sheep. In a 3-month study, 6 defects were treated with HA-TG in a collagen scaffold to test the stability and biocompatibility of the defect filling. In a 6-month study, 6 sheep had 12 defects treated with HA-TG and collagen and 2 sheep had 4 untreated defects. Histologically observed quality of repair tissue and adjacent cartilage was semiquantitatively assessed. Results: HA-TG adhered to the native tissue and did not cause any detectable negative reaction in the surrounding tissue. HA-TG in a collagen scaffold supported infiltration and chondrogenic differentiation of mesenchymal cells, which migrated from the subchondral bone through the calcified cartilage layer. Additionally, HA-TG and collagen treatment led to better adjacent cartilage preservation compared with empty defects ( Conclusions: This study demonstrates that the adhesive HA-TG hydrogel in a collagen scaffold shows good biocompatibility, supports
MeSH term(s)	Adhesives ; Animals ; Cartilage ; Collagen ; Hyaluronic Acid/pharmacology ; Hydrogels ; Mice ; Sheep
Chemical Substances	Adhesives ; Hydrogels ; Hyaluronic Acid (9004-61-9) ; Collagen (9007-34-5)
Language	English
Publishing date	2021-01-29
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2515870-3
ISSN	1947-6043 ; 1947-6035
ISSN (online)	1947-6043
ISSN	1947-6035
DOI	10.1177/1947603521989417
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Article ; Online: Thermal conditioning improves quality and speed of keratinocyte sheet production for burn wound treatment.

Frese, Laura / Darwiche, Salim E / von Rechenberg, Brigitte / Hoerstrup, Simon P / Giovanoli, Pietro / Calcagni, Maurizio

Cytotherapy

2021 Volume 23, Issue 6, Page(s) 536–547

Abstract: Background aims: Cultured patient-specific keratinocyte sheets have been used clinically since the 1970s for the treatment of large severe burns. However, despite significant developments in recent years, successful and sustainable treatment is still a ... ...

Abstract	Background aims: Cultured patient-specific keratinocyte sheets have been used clinically since the 1970s for the treatment of large severe burns. However, despite significant developments in recent years, successful and sustainable treatment is still a challenge. Reliable, high-quality grafts with faster availability and a flexible time window for transplantation are required to improve clinical outcomes. Methods: Keratinocytes are usually grown in vitro at 37°C. Given the large temperature differences in native skin tissue, the aim of the authors' study was to investigate thermal conditioning of keratinocyte sheet production. Therefore, the influence of 31°C, 33°C and 37°C on cell expansion and differentiation in terms of proliferation and sheet formation efficacy was investigated. In addition, the thermal effect on the biological status and thus the quality of the graft was assessed on the basis of the release of wound healing-related biofactors in various stages of graft development. Results: The authors demonstrated that temperature is a decisive factor in the production of human keratinocyte sheets. By using specific temperature ranges, the authors have succeeded in optimizing the individual manufacturing steps. During the cell expansion phase, cultivation at 37°C was most effective. After 6 days of culture at 37°C, three times and six times higher numbers of viable cells were obtained compared with 33°C and 31°C. During the cell differentiation and sheet formation phase, however, the cells benefited from a mildly hypothermic temperature of 33°C. Keratinocytes showed increased differentiation potential and formed better epidermal structures, which led to faster biomechanical sheet stability at day 18. In addition, a cultivation temperature of 33°C resulted in a longer lasting and higher secretion of the investigated immunomodulatory, anti-inflammatory, angiogenic and pro-inflammatory biofactors. Conclusions: These results show that by using specific temperature ranges, it is possible to accelerate the large-scale production of cultivated keratinocyte sheets while at the same time improving quality. Cultivated keratinocyte sheets are available as early as 18 days post-biopsy and at any time for 7 days thereafter, which increases the flexibility of the process for surgeons and patients alike. These findings will help to provide better clinical outcomes, with an increased take rate in severe burn patients.
MeSH term(s)	Burns/therapy ; Cell Differentiation ; Cells, Cultured ; Humans ; Keratinocytes ; Skin ; Skin Transplantation ; Wound Healing
Language	English
Publishing date	2021-03-06
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2039821-9
ISSN	1477-2566 ; 1465-3249
ISSN (online)	1477-2566
ISSN	1465-3249
DOI	10.1016/j.jcyt.2021.01.006
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Article ; Online: Domain-independent simulation of physiologically relevant callus shape in mechanoregulated models of fracture healing.

Schwarzenberg, Peter / Ren, Tianyi / Klein, Karina / von Rechenberg, Brigitte / Darwiche, Salim / Dailey, Hannah L

Journal of biomechanics

2021 Volume 118, Page(s) 110300

Abstract: Mechanoregulatory models have been used to predict the progression of bone fracture healing for more than two decades. However, many published studies share the same fundamental limitation: callus development proceeds within a pre-defined domain that ... ...

Abstract	Mechanoregulatory models have been used to predict the progression of bone fracture healing for more than two decades. However, many published studies share the same fundamental limitation: callus development proceeds within a pre-defined domain that both restricts and directs healing and leads to some non-physiologic healing patterns. To address this limitation, we added two spatial proximity functions to an existing mechanoregulatory model of fracture healing to control the localization of callus within the healing domain. We tested the performance of the new model in an idealized ovine tibial osteotomy with medial plate fixation using three sizes of healing domains and multiple variations of the spatial proximity functions. All model variations produced outward callus growth and bridging weighted toward the far cortex, which is consistent with in vivo healing. With and without the proximity functions, there were marked differences in the predicted callus volume and shape. With no proximity functions, the callus produced was strongly domain dependent, with a 15% difference in volume between the smallest and largest initialization domains. With proximity function control, callus growth was restricted to near the fracture line and there was only 2% difference in volume between domain sizes. Superimposing both proximity functions - one to control outward growth and one representing a decay in periosteal activity away from the fracture - produced a predicted callus size that was within the physiologic range for sheep and had a realistic morphology when compared with fluorescent dye co-localization with calcium deposition over time and histology.
MeSH term(s)	Animals ; Bone Plates ; Bony Callus ; Fracture Healing ; Fractures, Bone ; Osteotomy ; Sheep
Language	English
Publishing date	2021-02-06
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	218076-5
ISSN	1873-2380 ; 0021-9290
ISSN (online)	1873-2380
ISSN	0021-9290
DOI	10.1016/j.jbiomech.2021.110300
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