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Background: Bone mineral density at the humeral head is reduced in patients with chronic rotator cuff tears. Bone loss in the humeral head is associated with repair failure after rotator cuff reconstruction. Bisphosphonates (eg, zoledronic acid) increase bone mineral density. Hypothesis: Zoledronic acid improves bone mineral density of the humeral head and biomechanical properties of the enthesis after reconstruction of chronic rotator cuff tears in rats. Study Design: Controlled laboratory study. Methods: A total of 32 male Sprague-Dawley rats underwent unilateral (left) supraspinatus tenotomy with delayed transosseous rotator cuff reconstruction after 3 weeks. All rats were sacrificed 8 weeks after rotator cuff repair. Animals were randomly assigned to 1 of 2 groups. At 1 day after rotator cuff reconstruction, the intervention group was treated with a single subcutaneous dose of zoledronic acid at 100 µg/kg bodyweight, and the control group received 1 mL of subcutaneous saline solution. In 12 animals of each group, micro–computed tomography scans of both shoulders were performed as well as biomechanical testing of the supraspinatus enthesis of both sides. In 4 animals of each group, histological analyses were conducted. Results: In the intervention group, bone volume fraction (bone volume/total volume [BV/TV]) of the operated side was higher at the lateral humeral head ( P = .005) and the medial humeral head ( P = .010) compared with the control group. Trabecular number on the operated side was higher at the lateral humeral head ( P = .004) and the medial humeral head ( P = .001) in the intervention group. Maximum load to failure rates on the operated side were higher in the intervention group ( P < .001). Cortical thickness positively correlated with higher maximum load to failure rates in the intervention group ( r = 0.69; P = .026). Histological assessment revealed increased bone formation in the intervention group. Conclusion: Single-dose therapy of zoledronic acid provided an improvement of bone microarchitecture at the humeral head as well as an increase of maximum load to failure rates after transosseous reconstruction of chronic rotator cuff lesions in rats. Clinical Relevance: Zoledronic acid improves bone microarchitecture as well as biomechanical properties after reconstruction of chronic rotator cuff tears in rodents. These results need to be verified in clinical investigations.

Background In spite of advances in the treatment of cartilage defects using cell and scaffold-based therapeutic strategies, the long-term outcome is still not satisfying since clinical scores decline years after treatment. Scaffold materials currently used in clinical settings have shown limitations in providing suitable biomechanical properties and an authentic and protective environment for regenerative cells. To tackle this problem, we developed a scaffold material based on decellularised human articular cartilage. Methods Human articular cartilage matrix was engraved using a CO2 laser and treated for decellularisation and glycosaminoglycan removal. Characterisation of the resulting scaffold was performed via mechanical testing, DNA and GAG quantification and in vitro cultivation with adipose-derived stromal cells (ASC). Cell vitality, adhesion and chondrogenic differentiation were assessed. An ectopic, unloaded mouse model was used for the assessment of the in vivo performance of the scaffold in combination with ASC and human as well as bovine chondrocytes. The novel scaffold was compared to a commercial collagen type I/III scaffold. Findings Crossed line engravings of the matrix allowed for a most regular and ubiquitous distribution of cells and chemical as well as enzymatic matrix treatment was performed to increase cell adhesion. The biomechanical characteristics of this novel scaffold that we term CartiScaff were found to be superior to those of commercially available materials. Neo-tissue was integrated excellently into the scaffold matrix and new collagen fibres were guided by the laser incisions towards a vertical alignment, a typical feature of native cartilage important for nutrition and biomechanics. In an ectopic, unloaded in vivo model, chondrocytes and mesenchymal stromal cells differentiated within the incisions despite the lack of growth factors and load, indicating a strong chondrogenic microenvironment within the scaffold incisions. Cells, most noticeably bone marrow-derived cells, were able to repopulate the empty chondrocyte lacunae inside the scaffold matrix. Interpretation Due to the better load-bearing, its chondrogenic effect and the ability to guide matrix-deposition, CartiScaff is a promising biomaterial to accelerate rehabilitation and to improve long term clinical success of cartilage defect treatment. Funding Austrian Research Promotion Agency FFG (“CartiScaff” #842455), Lorenz Böhler Fonds (16/13), City of Vienna Competence Team Project Signaltissue (MA23, #18-08), Graphical abstract Image, graphical abstract

Surgical sealants help achieve rapid haemostasis when applied as an adjunct to sutures in vascular surgery, but their use can lead to various side effects. This study compared the local inflammatory reaction to commercially available BioGlue and Coseal sealants in a rabbit aorta suture hole model. Twenty male New Zealand white rabbits were randomised to testing with either BioGlue or Coseal. Two weeks after sealant application to suture holes, sections of the aorta at the puncture site, and surrounding tissue, were processed for histopathological analysis. Inflammation was graded from 0 to 3 according to tissue alteration and presence of inflammatory cells. Material stiffness was measured in vitro using compression testing. From examination of the inflammatory response to the sealants, a less severe histopathological assessment score was assigned to the Coseal compared to the BioGlue group (mean ± SD: 1.56 ± 0.53 vs 2.67 ± 0.50; p = 0.002). While both materials triggered a typical foreign body reaction characterised by granulomatous inflammation, BioGlue additionally provoked eosinophilic cell infiltration. Lymphocytes, plasma cells and B cells were also more prevalent in the BioGlue compared to the Coseal specimens. Coseal residue was either absent or visible in only small quantities, while significant BioGlue deposits remained in the tissue 2 weeks after application. Coseal was much more elastic than BioGlue, with a compressive modulus an order of magnitude lower (mean ± SD: 91 ± 41 vs 1833 ± 297 kPa). Compared to BioGlue, Coseal elicited a less pronounced inflammatory response in the aortic and peri-aortic tissue in this model, and demonstrated greater elasticity.

Decellularization of blood vessels is a promising approach to generate native biomaterials for replacement of diseased vessels. The decellularization process affects the mechanical properties of the vascular graft and thus can have a negative impact for in vivo functionality. The aim of this study was to determine how detergents under different fluid dynamics affects decellularization efficacy and mechanical properties of the vascular graft. We applied a protocol utilizing 1% TritonX, 1% Tributyl phosphate (TnBP) and DNase on porcine vena cava. The detergents were applied to the vessels under different conditions; static, agitation and perfusion with 3 different perfusion rates (25, 100 and 400 mL/min). The decellularized grafts were analyzed with histological, immunohistochemical and mechanical tests. We found that decellularization efficacy was equal in all groups, however the luminal ultrastructure of the static group showed remnant cell debris and the 400 mL/min perfusion group showed local damage and tearing of the luminal surface. The mechanical stiffness and maximum tensile strength were not influenced by the detergent application method. In conclusion, our results indicate that agitation or low-velocity perfusion with detergents are preferable methods for blood vessel decellularization.

Background: Characteristics of chronic rotator cuff tears include continuous loss of tendon structure as well as tendon elasticity, followed by a high failure rate after surgical reconstruction. Several studies have already shown the beneficial effect of extracorporeal shockwave therapy (ESWT) on tissue regeneration in tendon pathologies. Hypothesis: ESWT improves biomechanical tendon properties as well as functional shoulder outcomes in chronic rotator cuff reconstruction in rodents. Study Design: Controlled laboratory study. Methods: After tendon detachment and 3 weeks of degeneration, a subsequent transosseous reattachment of the supraspinatus tendon was performed in 48 adult male Sprague-Dawley rats (n = 16 per group). Rodents were randomly assigned to 3 study groups: no ESWT/control group, intraoperative ESWT (IntraESWT), and intra- and postoperative ESWT (IntraPostESWT). Shoulder joint function, as determined by gait analysis, was assessed repeatedly during the observation period. Eight weeks after tendon reconstruction, the rats were euthanized, and biomechanical and gene expression analyses were performed. Results: Macroscopically, all repairs were intact at the time of euthanasia, with no ruptures detectable. Biomechanical analyses showed significantly improved load-to-failure testing results in both ESWT groups in comparison with the control group (control, 0.629; IntraESWT, 1.102; IntraPostESWT, 0.924; IntraESWT vs control, P≤ .001; IntraPostESWT vs control, P≤ .05). Furthermore, functional gait analyses showed a significant enhancement in intensity measurements for the IntraPostESWT group in comparison with the control group ( P≤ .05). Gene expression analysis revealed no significant differences among the 3 groups. Conclusion: Clearly improved biomechanical results were shown in the single-application and repetitive ESWT groups. Furthermore, functional evaluation showed significantly improved intensity measurements for the repetitive ESWT group. Clinical Relevance: This study underpins a new additional treatment possibility to prevent healing failure. Improved biomechanical stability and functionality may enable faster remobilization as well as an accelerated return to work and sports activities. Furthermore, as shockwave therapy is a noninvasive, easy-to-perform, cost-effective treatment tool with no undesired side effects, this study is of high clinical relevance in orthopaedic surgery. Based on these study results, a clinical study has already been initiated to clinically confirm the improved functionality by ESWT.

Background:Given the unsatisfactory results and reported drawbacks of anterior cruciate ligament (ACL) reconstruction, such as donor site morbidity and the limited choice of grafts in revision surgery, new regenerative approaches based on tissue-engineering strategies are currently under investigation.Purposes:To determine (1) if a novel silk fiber–based ACL scaffold is able to initiate osteointegration in the femoral and tibial bone tunnels under in vivo conditions and (2) if the osteointegration process will be improved by intraoperatively seeding the scaffolds with the autologous stromal vascular fraction, an adipose-derived, stem cell–rich isolate from knee fat pads.Study Design:Controlled laboratory study.Methods:A total of 33 sheep underwent ACL resection and were then randomly assigned to 2 experimental groups: ACL reconstruction with a scaffold alone and ACL reconstruction with a cell-seeded scaffold. Half of the sheep in each group were randomly chosen and euthanized 6 months after surgery and the other half at 12 months. To analyze the integration of the silk-based scaffold in the femoral and tibial bone tunnels, hard tissue histology and micro–computed tomography measurements were performed.Results:Hard tissue histological workup showed that in all treatment groups, with or without the application of the autologous stromal vascular fraction, an interzone of collagen fibers had formed between bone and silk-based graft. This collagen-fiber continuity partly consisted of Sharpey fibers, comparable with tendon-bone healing known for autografts and allografts. Insertion sites were more broad based at 6 months and more concentrated on the slightly protruding, bony knoblike structures at 12 months. Histologically, no differences between the treatment groups were detectable. Analysis of micro–computed tomography measurements revealed a significantly higher tissue density for the cell-seeded scaffold group as compared with the scaffold-alone group in the tibial but not femoral bone tunnel after 12 months of implantation.Conclusion:The novel silk fiber–based scaffold for ACL regeneration demonstrated integration into the bone tunnels via the formation of a fibrous interzone similar to allografts and autografts. Histologically, additional cell seeding did not enhance osteointegration. No significant differences between 6 and 12 months could be detected. After 12 months, there was still a considerable amount of silk present, and a longer observation period is necessary to see if a true ligament-bone enthesis will be formed.Clinical Relevance:ACL regeneration with a silk fiber–based scaffold with and without additional cell seeding may provide an alternative treatment option to current techniques of surgical reconstruction.

Biomaterials currently in use for articular cartilage regeneration do not mimic the composition or architecture of hyaline cartilage, leading to the formation of repair tissue with inferior characteristics. In this study we demonstrate the use of “AuriScaff”, an enzymatically perforated bovine auricular cartilage scaffold, as a novel biomaterial for repopulation with regenerative cells and for the formation of high-quality hyaline cartilage. AuriScaff features a traversing channel network, generated by selective depletion of elastic fibers, enabling uniform repopulation with therapeutic cells. The complex collagen type II matrix is left intact, as observed by immunohistochemistry, SEM and TEM. The compressive modulus is diminished, but three times higher than in the clinically used collagen type I/III scaffold that served as control. Seeding tests with human articular chondrocytes (hAC) alone and in co-culture with human adipose-derived stromal/stem cells (ASC) confirmed that the network enabled cell migration throughout the scaffold. It also guides collagen alignment along the channels and, due to the generally traverse channel alignment, newly deposited cartilage matrix corresponds with the orientation of collagen within articular cartilage. In an osteochondral plug model, AuriScaff filled the complete defect with compact collagen type II matrix and enabled chondrogenic differentiation inside the channels. Using adult articular chondrocytes from bovine origin (bAC), filling of even deep defects with high-quality hyaline-like cartilage was achieved after 6 weeks in vivo. With its composition and spatial organization, AuriScaff provides an optimal chondrogenic environment for therapeutic cells to treat cartilage defects and is expected to improve long-term outcome by channel-guided repopulation followed by matrix deposition and alignment. Statement of Significance After two decades of tissue engineering for cartilage regeneration, there is still no optimal strategy available to overcome problems such as inconsistent clinical outcome, early and late graft failures. Especially large defects are dependent on biomaterials and their scaffolding, guiding and protective function. Considering the currently used biomaterials, structure and mechanical properties appear to be insufficient to fulfill this task. The novel scaffold developed within this study is the first approach enabling the use of dense cartilage matrix, repopulate it via channels and provide the cells with a compact collagen type II environment. Due to its density, it also provides better mechanical properties than materials currently used in clinics. We therefore think, that the auricular cartilage scaffold (AuriScaff) has a high potential to improve future cartilage regeneration approaches.

Two self-adhering hemostatic patches, based on either PEG-coated collagen (PCC) or PEG-coated oxidized cellulose (PCOC), are compared regarding to maximum burst pressure, mechanical stability, and swelling. In addition, the induction of tissue adhesions by the materials was assessed in a rabbit liver abrasion model. Both materials showed comparable sealing efficacy in a burst pressure test (37 ± 16 vs. 35 ± 8 mmHg, P = 0.730). After incubation in human plasma, PCC retained its mechanical properties over the test period of 8 h, while PCOC showed faster degradation after the 2 h time-point. The degradation led to a significantly decreased force at break (minimum force at break 0.55 N during 8 h for PCC, 0.27 N for PCOC; p

Purpose: Topical hemostatic agents are an important means of controlling or preventing bleeding. This study was performed to compare gelatin–thrombin matrix with smooth particles (SmGM) versus gela...

Biomaterials based on decellularized tissues are increasingly attracting attention as functional alternatives to other natural or synthetic materials. However, a source of non-cadaver human allograft material would be favorable. Here we establish a decellularization method of vascular tissue from cryopreserved human placenta chorionic plate starting with an initial freeze–thaw step followed by a series of chemical treatments applied with a custom-made perfusion system. This novel pulsatile perfusion set-up enabled us to successfully decellularize the vascular tissue with lower concentrations of chemicals and shorter exposure times compared to a non-perfusion process. The decellularization procedure described here lead to the preservation of the native extracellular matrix architecture and the removal of cells. Quantitative analysis revealed no significant changes in collagen content and a retained glycosaminoglycan content of approximately 29%. In strain-to-failure tests, the decellularized grafts showed similar mechanical behavior compared to native controls. In addition, the mechanical values for ultimate tensile strength and stiffness were in an acceptable range for in vivo applications. Furthermore, biocompatibility of the decellularized tissue and its recellularizationability to serve as an adequate substratum for upcoming recellularization strategies using primary human umbilical vein endothelial cells (HUVECs) was demonstrated. HUVECs cultured on the decellularized placenta vessel matrix performed endothelialization and maintained phenotypical characteristics and cell specific expression patterns. Overall, the decellularized human placenta vessels can be a versatile tool for experimental studies on vascularization and as potent graft material for future in vivo applications. Statement of significance In the US alone more than 1 million vascular grafts are needed in clinical practice every year. Despite severe disadvantages, such as donor site morbidity, autologous grafting from the patient’s own arteries or veins is regarded as the gold standard for vascular tissue repair. Besides, strategies based on synthetic or natural materials have shown limited success. Tissue engineering approaches based on decellularized tissues are regarded as a promising alternative to clinically used treatments to overcome the observed limitations. However, a source for supply of non-cadaver human allograft material would be favorable. Here, we established a decellularization method of vascular tissue from the human placenta chorionic plate, a suitable human tissue source of consistent quality. The decellularized human placenta vessels can be a potent graft material for future in vivo applications and furthermore might be a versatile tool for experimental studies on vascularization.

Natural extracellular matrix-derived biomaterials from decellularized allogenic tissues are of increasing interest for tissue engineering because their structure and composition provide a complexity that is not achievable with current manufacturing techniques. The prerequisite to bring allogenic tissue from bench to bedside as a functional biomaterial is the full removal of cells while preserving most of its native characteristics such as structure and composition. The exceptionally dense structure of articular cartilage, however, poses a special challenge for decellularization, scaffold preparation, and reseeding. Therefore, we tested 24 different protocols aiming to remove cells and glycosaminoglycans (GAG) while preserving the collagen backbone and ultrastructure. The resulting matrices were analyzed for cell removal (DNA quantification, haematoxylin and eosin staining), GAG content (dimethyl methylene blue assay, Alcian blue staining and micro-computed tomography), collagen integrity (immunohistochemistry and ultrastructure), and biomechanics (compression test). Furthermore, seeding tests were conducted to evaluate cell viability and attachment to the scaffolds. Sodium dodecyl sulfate-based protocols yielded satisfactory reduction of DNA content, yet had negative effects on cell viability and attachment. Hydrochloric acid efficiently decellularized the scaffold and pepsin emerged as best option for GAG depletion. Combining these two reagents led to our final protocol, most efficient in DNA and GAG depletion while preserving the collagen architecture. The compressive modulus decreased in the absence of GAG to ∼1/3 of native cartilage, which is significantly higher than that by commercially available scaffolds tested as a reference (ranging from 1/25 to 1/100 of native cartilage). Cytocompatibility tests showed that human adipose-derived stromal cells readily adhered to the scaffold. In this study, we established a protocol combining freeze-thaw cycles, osmotic shock, and treatment with hydrochloric acid followed by a pepsin digestion step, achieving successful decellularization and GAG depletion within 1 week, resulting in a cytocompatible material with intact collagen structure. The protocol provides a basis for the generation of allogeneic scaffolds, potentially substituting manufactured scaffolds currently used in clinical articular cartilage treatment.

Various tissue engineering (TE) approaches are based on silk fibroin (SF) as scaffold material because of its superior mechanical and biological properties compared to other materials. The translation of one-step TE approaches to clinical application has generally failed so far due to the requirement of a prolonged cell seeding step before implantation. Here, we propose that the plant lectin WGA (wheat germ agglutinin), covalently bound to SF, will mediate cell adhesion in a time frame acceptable to be part of a one-step surgical intervention. After the establishment of a modification protocol utilizing carbodiimide chemistry, we examined the attachment of cells, with a special focus on adipose-derived stromal cells (ASC), on WGA-SF compared to pure native SF. After a limited time frame of 20min the attachment of ASCs to WGA-SF showed an increase of about 17-fold, as compared to pure native SF. The lectin-mediated cell adhesion further showed an enhanced resistance to trypsin (as a protease model) and to applied fluid shear stress (mechanical stability). Moreover, we could demonstrate that the adhesion of ASCs on the WGA-SF does not negatively influence proliferation or differentiation potential into the osteogenic lineage. To test for in vitro immune response, the proliferation of peripheral blood mononuclear cells in contact with the WGA-SF was determined, showing no alterations compared to plain SF. All these findings suggest that the WGA modification of SF offers important benefits for translation of SF scaffolds into clinical applications.

Schwann cells play a key role in peripheral nerve regeneration. Failure in sufficient formation of Bungner bands due to impaired Schwann cell proliferation has significant effects on the functional outcome after regeneration. Therefore, the growth substrate for Schwann cells should be considered with highest priority in any peripheral nerve tissue engineering approach. Due to its excellent biocompatibility silk fibroin has most recently attracted considerable interest as a biomaterial for use as conduit material in peripheral nerve regeneration. In this study we established a protocol to covalently bind collagen and laminin, which have been isolated from human placenta, to silk fibroin utilizing carbodiimide chemistry. Altered adhesion, viability and proliferation of Schwann cells were evaluated. A cell adhesion assay revealed that the functionalization with both, laminin or collagen, significantly improved Schwann cell adhesion to silk fibroin. Moreover laminin drastically accelerated adhesion. Schwann cell proliferation and viability assessed with BrdU and MTT assay, respectively, were significantly increased in the laminin-functionalized groups. The results suggest beneficial effects of laminin on both, cell adhesion as well as proliferative behaviour of Schwann cells. To conclude, the covalent tailoring of silk fibroin drastically enhances its properties as a cell substratum for Schwann cells, which might help to overcome current hurdles bridging long distance gaps in peripheral nerve injuries with the use of silk-based nerve guidance conduits.

Recent studies have demonstrated that combining cells with meshes prior to implantation successfully enhanced hernia repair. The idea is to create a biologic coating surrounding the mesh with autologous cells, before transplantation into the patient. However, due to the lack of a prompt and robust cell adhesion to the meshes, extensive in vitro cultivation is required to obtain a homogenous cell layer covering the mesh. In this context, the objective of this publication is to manufacture meshes made of silk fibres and to enhance the cytoadhesion and cytocompatibility of the biomaterial by surface immobilization of a pro-adhesive wheat germ agglutinin (lectin WGA). We first investigated the affinity between the glycoprotein WGA and cells, in solution and then after covalent immobilization of WGA on silk films. Then, we manufactured meshes made of silk fibres, tailored them with WGA grafting and finally evaluated the cytocompatibility and the inflammatory response of silk and silk-lectin meshes compared to common polypropylene mesh, using fibroblasts and peripheral blood mononuclear cells, respectively. The in vitro experiments revealed that the cytocompatibility of silk can be enhanced by surface immobilization with lectin WGA without exhibiting negative response in terms of pro-inflammatory reaction. Grafting lectin to silk meshes could bring advantages to facilitate cell-coating of meshes prior to implantation, which is an imperative prerequisite for abdominal wall tissue regeneration using cell-based therapy.

The generation of functional biomimetic skeletal muscle constructs is still one of the fundamental challenges in skeletal muscle tissue engineering. With the notion that structure strongly dictates functional capabilities, a myriad of cell types, scaffold materials and stimulation strategies have been combined. To further optimize muscle engineered constructs, we have developed a novel bioreactor system (MagneTissue) for rapid engineering of skeletal muscle-like constructs with the aim to resemble native muscle in terms of structure, gene expression profile and maturity. Myoblasts embedded in fibrin, a natural hydrogel that serves as extracellular matrix, are subjected to mechanical stimulation via magnetic force transmission. We identify static mechanical strain as a trigger for cellular alignment concomitant with the orientation of the scaffold into highly organized fibrin fibrils. This ultimately yields myotubes with a more mature phenotype in terms of sarcomeric patterning, diameter and length. On the molecular level, a faster progression of the myogenic gene expression program is evident as myogenic determination markers MyoD and Myogenin as well as the Ca2+ dependent contractile structural marker TnnT1 are significantly upregulated when strain is applied. The major advantage of the MagneTissue bioreactor system is that the generated tension is not exclusively relying on the strain generated by the cells themselves in response to scaffold anchoring but its ability to subject the constructs to individually adjustable strain protocols. In future work, this will allow applying mechanical stimulation with different strain regimes in the maturation process of tissue engineered constructs and elucidating the role of mechanotransduction in myogenesis. Statement of Significance Mechanical stimulation of tissue engineered skeletal muscle constructs is a promising approach to increase tissue functionality. We have developed a novel bioreactor-based 3D culture system, giving the user the possibility to apply different strain regimes like static, cyclic or ramp strain to myogenic precursor cells embedded in a fibrin scaffold. Application of static mechanical strain leads to alignment of fibrin fibrils along the axis of strain and concomitantly to highly aligned myotube formation. Additionally, the pattern of myogenic gene expression follows the temporal progression observed in vivo with a more thorough induction of the myogenic program when static strain is applied. Ultimately, the strain protocol used in this study results in a higher degree of muscle maturity demonstrated by enhanced sarcomeric patterning and increased myotube diameter and length. The introduced bioreactor system enables new possibilities in muscle tissue engineering as longer cultivation periods and different strain applications will yield tissue engineered muscle-like constructs with improved characteristics in regard to functionality and biomimicry.

Temporary scaffolds that mimic the extracellular matrix’s structure and provide a stable substratum for the natural growth of cells are an innovative trend in the field of tissue engineering. The aim of this study is to obtain and design porous 2D fibroin-based cell matrices by femtosecond laser-induced microstructuring for future applications in muscle tissue engineering. Ultra-fast laser treatment is a non-contact method, which generates controlled porosity—the creation of micro/nanostructures on the surface of the biopolymer that can strongly affect cell behavior, while the control over its surface characteristics has the potential of directing the growth of future muscle tissue in the desired direction. The laser structured 2D thin film matrices from silk were characterized by means of SEM, EDX, AFM, FTIR, Micro-Raman, XRD, and 3D-roughness analyses. A WCA evaluation and initial experiments with murine C2C12 myoblasts cells were also performed. The results show that by varying the laser parameters, a different structuring degree can be achieved through the initial lifting and ejection of the material around the area of laser interaction to generate porous channels with varying widths and depths. The proper optimization of the applied laser parameters can significantly improve the bioactive properties of the investigated 2D model of a muscle cell matrix.