Deciphering the role of FUS::DDIT3 expression and tumor microenvironment in myxoid liposarcoma development.

BACKGROUND: Myxoid liposarcoma (MLS) displays a distinctive tumor microenvironment and is characterized by the FUS::DDIT3 fusion oncogene, however, the precise functional contributions of these two elements remain enigmatic in tumor development. METHODS: To study the cell-free microenvironment in MLS, we developed an experimental model system based on decellularized patient-derived xenograft tumors. We characterized the cell-free scaffold using mass spectrometry. Subsequently, scaffolds were repopulated using sarcoma cells with or without FUS::DDIT3 expression that were analyzed with histology and RNA sequencing. RESULTS: Characterization of cell-free MLS scaffolds revealed intact structure and a large variation of protein types remaining after decellularization. We demonstrated an optimal culture time of 3 weeks and showed that FUS::DDIT3 expression decreased cell proliferation and scaffold invasiveness. The cell-free MLS microenvironment and FUS::DDIT3 expression both induced biological processes related to cell-to-cell and cell-to-extracellular matrix interactions, as well as chromatin remodeling, immune response, and metabolism. Data indicated that FUS::DDIT3 expression more than the microenvironment determined the pre-adipocytic phenotype that is typical for MLS. CONCLUSIONS: Our experimental approach opens new means to study the tumor microenvironment in detail and our findings suggest that FUS::DDIT3-expressing tumor cells can create their own extracellular niche.

Polysaccharides and Structural Proteins as Components in Three-Dimensional Scaffolds for Breast Cancer Tissue Models: A Review.

Breast cancer is the most common cancer among women, and even though treatments are available, efficiency varies with the patients. In vitro 2D models are commonly used to develop new treatments. However, 2D models overestimate drug efficiency, which increases the failure rate in later phase III clinical trials. New model systems that allow extensive and efficient drug screening are thus required. Three-dimensional printed hydrogels containing active components for cancer cell growth are interesting candidates for the preparation of next generation cancer cell models. Macromolecules, obtained from marine- and land-based resources, can form biopolymers (polysaccharides such as alginate, chitosan, hyaluronic acid, and cellulose) and bioactive components (structural proteins such as collagen, gelatin, and silk fibroin) in hydrogels with adequate physical properties in terms of porosity, rheology, and mechanical strength. Hence, in this study attention is given to biofabrication methods and to the modification with biological macromolecules to become bioactive and, thus, optimize 3D printed structures that better mimic the cancer cell microenvironment. Ink formulations combining polysaccharides for tuning the mechanical properties and bioactive polymers for controlling cell adhesion is key to optimizing the growth of the cancer cells.

Personalized tissue-engineered veins - long term safety, functionality and cellular transcriptome analysis in large animals.

Tissue engineering is a promising methodology to produce advanced therapy medicinal products (ATMPs). We have developed personalized tissue engineered veins (P-TEV) as an alternative to autologous or synthetic vascular grafts utilized in reconstructive vein surgery. Our hypothesis is that individualization through reconditioning of a decellularized allogenic graft with autologous blood will prime the tissue for efficient recellularization, protect the graft from thrombosis, and decrease the risk of rejection. In this study, P-TEVs were transplanted to vena cava in pig, and the analysis of three veins after six months, six veins after 12 months and one vein after 14 months showed that all P-TEVs were fully patent, and the tissue was well recellularized and revascularized. To confirm that the ATMP product had the expected characteristics one year after transplantation, gene expression profiling of cells from P-TEV and native vena cava were analyzed and compared by qPCR and sequencing. The qPCR and bioinformatics analysis confirmed that the cells from the P-TEV were highly similar to the native cells, and we therefore conclude that P-TEV is functional and safe in large animals and have high potential for use as a clinical transplant graft.

De- and recellularized urethral reconstruction with autologous buccal mucosal cells implanted in an ovine animal model.

OBJECTIVES: Patients with urethral stricture due to any type of trauma, hypospadias or gender dysphoria suffer immensely from impaired capacity to urinate and are in need of a new functional urethra. Tissue engineering with decellularization of a donated organ recellularized with cells from the recipient patient has emerged as a promising alternative of advanced therapy medicinal products. The aim of this pilot study was to develop an ovine model of urethral transplantation and to produce an individualized urethra graft to show proof of function in vivo. METHODS: Donated urethras from ram abattoir waste were decellularized and further recellularized with autologous buccal mucosa epithelial cells excised from the recipient ram and expanded in vitro. The individualized urethral grafts were implanted by reconstructive surgery in rams replacing 2.5 ± 0.5 cm of the native penile urethra. RESULTS: After surgery optimization, three ram had the tissue engineered urethra implanted for one month and two out of three showed a partially regenerated epithelium. CONCLUSIONS: Further adjustments of the model are needed to achieve a satisfactory proof-of-concept; however, we interpret these findings as a proof of principle and a possible path to develop a functional tissue engineered urethral graft with de- and recellularization and regeneration in vivo after transplantation.

Breast Cancer Patient-Derived Scaffolds Can Expose Unique Individual Cancer Progressing Properties of the Cancer Microenvironment Associated with Clinical Characteristics.

Breast cancer is a heterogeneous disease in terms of cellular and structural composition, and besides acquired aggressive properties in the cancer cell population, the surrounding tumor microenvironment can affect disease progression and clinical behaviours. To specifically decode the clinical relevance of the cancer promoting effects of individual tumor microenvironments, we performed a comprehensive test of 110 breast cancer samples using a recently established in vivo-like 3D cell culture platform based on patient-derived scaffolds (PDSs). Cell-free PDSs were recellularized with three breast cancer cell lines and adaptation to the different patient-based microenvironments was monitored by quantitative PCR. Substantial variability in gene expression between individual PDS cultures from different patients was observed, as well as between different cell lines. Interestingly, specific gene expression changes in the PDS cultures were significantly linked to prognostic features and clinical information from the original cancer. This link was even more pronounced when ERα-status of cell lines and PDSs matched. The results support that PDSs cultures, including a cancer cell line of relevant origin, can monitor the activity of the tumor microenvironment and reveal unique information about the malignancy-inducing properties of the individual cancer niche and serve as a future complementary diagnostic tool for breast cancer.

The Effect of Hypoxic and Normoxic Culturing Conditions in Different Breast Cancer 3D Model Systems.

The field of 3D cell cultures is currently emerging, and material development is essential in striving toward mimicking the microenvironment of a native tissue. By using the response of reporter cells to a 3D environment, a comparison between materials can be assessed, allowing optimization of material composition and microenvironment. Of particular interest, the response can be different in a normoxic and hypoxic culturing conditions, which in turn may alter the conclusion regarding a successful recreation of the microenvironment. This study aimed at determining the role of such environments to the conclusion of a better resembling cell culture model to native tissue. Here, the breast cancer cell line MCF7 was cultured in normoxic and hypoxic conditions on patient-derived scaffolds and compared at mRNA and protein levels to cells cultured on 3D printed scaffolds, Matrigel, and conventional 2D plastics. Specifically, a wide range of mRNA targets (40), identified as being regulated upon hypoxia and traditional markers for cell traits (cancer stem cells, epithelial-mesenchymal transition, pluripotency, proliferation, and differentiation), were used together with a selection of corresponding protein targets. 3D cultured cells were vastly different to 2D cultured cells in gene expression and protein levels on the majority of the selected targets in both normoxic and hypoxic culturing conditions. By comparing Matrigel and 3DPS-cultured cells to cells cultured on patient-derived scffolds, differences were also noted along all categories of mRNA targets while specifically for the GLUT3 protein. Overall, cells cultured on patient-derived scaffolds closely resembled cells cultured on 3D printed scaffolds, contrasting 2D and Matrigel-cultured cells, regardless of a normoxic or hypoxic culturing condition. Thus, these data support the use of either a normoxic or hypoxic culturing condition in assays using native tissues as a blueprint to optimize material composition.

3D Printed Nanocellulose Scaffolds as a Cancer Cell Culture Model System.

Current conventional cancer drug screening models based on two-dimensional (2D) cell culture have several flaws and there is a large need of more in vivo mimicking preclinical drug screening platforms. The microenvironment is crucial for the cells to adapt relevant in vivo characteristics and here we introduce a new cell culture system based on three-dimensional (3D) printed scaffolds using cellulose nanofibrils (CNF) pre-treated with 2,2,6,6-tetramethylpyperidine-1-oxyl (TEMPO) as the structural material component. Breast cancer cell lines, MCF7 and MDA-MB-231, were cultured in 3D TEMPO-CNF scaffolds and were shown by scanning electron microscopy (SEM) and histochemistry to grow in multiple layers as a heterogenous cell population with different morphologies, contrasting 2D cultured mono-layered cells with a morphologically homogenous cell population. Gene expression analysis demonstrated that 3D TEMPO-CNF scaffolds induced elevation of the stemness marker CD44 and the migration markers VIM and SNAI1 in MCF7 cells relative to 2D control. T47D cells confirmed the increased level of the stemness marker CD44 and migration marker VIM which was further supported by increased capacity of holoclone formation for 3D cultured cells. Therefore, TEMPO-CNF was shown to represent a promising material for 3D cell culture model systems for cancer cell applications such as drug screening.

Individualized tissue-engineered veins as vascular grafts: A proof of concept study in pig.

Personalized tissue engineered vascular grafts are a promising advanced therapy medicinal product alternative to autologous or synthetic vascular grafts utilized in blood vessel bypass or replacement surgery. We hypothesized that an individualized tissue engineered vein (P-TEV) would make the body recognize the transplanted blood vessel as autologous, decrease the risk of rejection and thereby avoid lifelong treatment with immune suppressant medication as is standard with allogenic organ transplantation. To individualize blood vessels, we decellularized vena cava from six deceased donor pigs and tested them for cellular removal and histological integrity. A solution with peripheral blood from the recipient pigs was used for individualized reconditioning in a perfusion bioreactor for seven days prior to transplantation. To evaluate safety and functionality of the individualized vascular graft in vivo, we transplanted reconditioned porcine vena cava into six pigs and analyzed histology and patency of the graft at different time points, with three pigs at the final endpoint 4-5 weeks after surgery. Our results showed that the P-TEV was fully patent in all animals, did not induce any occlusion or stenosis formation and we did not find any signs of rejection. The P-TEV showed rapid recellularization in vivo with the luminal surface covered with endothelial cells. In summary, the results indicate that P-TEV is functional and have potential for use as clinical transplant grafts.

Optimized alginate-based 3D printed scaffolds as a model of patient derived breast cancer microenvironments in drug discovery.

The cancer microenvironment influences tumor progression and metastasis and is pivotal to consider when designingin vivo-like cancer models. Current preclinical testing platforms for cancer drug development are mainly limited to 2D cell culture systems that poorly mimic physiological environments and traditional, low throughput animal models. The aim of this work was to produce a tunable testing platform based on 3D printed scaffolds (3DPS) with a simple geometry that, by extracellular components and response of breast cancer reporter cells, mimics patient-derived scaffolds (PDS) of breast cancer. Here, the biocompatible polysaccharide alginate was used as base material to generate scaffolds consisting of a 3D grid containing periostin and hydroxyapatite. Breast cancer cell lines (MCF7 and MDA-MB-231) produced similar phenotypes and gene expression levels of cancer stem cell, epithelial-mesenchymal transition, differentiation and proliferation markers when cultured on 3DPS and PDS, contrasting conventional 2D cultures. Importantly, cells cultured on 3DPS and PDS showed scaffold-specific responses to cytotoxic drugs (doxorubicin and 5-fluorouracil) that were different from 2D cultured cells. In conclusion, the data presented support the use of a tunable alginate-based 3DPS as a tumor model in breast cancer drug discovery.

Breast cancer patient-derived scaffolds as a tool to monitor chemotherapy responses in human tumor microenvironments.

Breast cancer is a heterogeneous disease where the tumor microenvironment, including extracellular components, plays a crucial role in tumor progression, potentially modulating treatment response. Different approaches have been used to develop three-dimensional models able to recapitulate the complexity of the extracellular matrix. Here, we use cell-free patient-derived scaffolds (PDSs) generated from breast cancer samples that were recellularized with cancer cell lines as an in vivo-like culture system for drug testing. We show that PDS cultured MCF7 cancer cells increased their resistance against the front-line chemotherapy drugs 5-fluorouracil, doxorubicin and paclitaxel in comparison to traditional two-dimensional cell cultures. The gene expression of the environmentally adapted cancer cells was modulated in different ways depending on the drug and the concentration used. High doses of doxorubicin reduced cancer stem cell features, whereas 5-fluorouracil increased stemness and decreased the proliferative phenotype. By using PDSs repopulated with other breast cancer cell lines, T-47D and MDA-MB-231, we observed both general and cell line specific drug responses. In summary, PDSs can be used to examine the extracellular matrix influence on cancer drug responses and for testing novel compounds in in vivo-like microenvironments.

Patient-derived scaffolds as a model of colorectal cancer.

BACKGROUND: Colorectal cancer is the second most common cause of cancer-related death worldwide and standardized therapies often fail to treat the more aggressive and progressive types of colorectal cancer. Tumor cell heterogeneity and influence from the surrounding tumor microenvironment (TME) contribute to the complexity of the disease and large variability in clinical outcomes. METHODS: To model the heterogeneous nature of colorectal cancer, we used patient-derived scaffolds (PDS), which were obtained via decellularization of surgically resected tumor material, as a growth substrate for standardized cell lines. RESULTS: After confirmation of native cell absence and validation of the structural and compositional integrity of the matrix, 89 colorectal PDS were repopulated with colon cancer cell line HT29. After 3 weeks of PDS culture, HT29 cells varied their gene and protein expression profiles considerably compared to 2D-grown HT29 cells. Markers associated with proliferation were consistently decreased, while markers associated with pluripotency were increased in PDS-grown cells compared to their 2D counterparts. When comparing the PDS-induced changes in HT29 cells with clinically relevant tumor information from individual patients, we observed significant associations between stemness/pluripotency markers and tumor location, and between epithelial-to-mesenchymal transition (EMT) markers and cancer mortality. Kaplan-Meier analysis revealed that low PDS-induced EMT correlated with worse cancer-specific survival. CONCLUSIONS: The colorectal PDS model can be used as a simplified personalized tool that can potentially reveal important diagnostic and pathophysiological information related to the TME.

Characterization of cell-free breast cancer patient-derived scaffolds using liquid chromatography-mass spectrometry/mass spectrometry data and RNA sequencing data.

Patient-derived scaffolds (PDSs) generated from primary breast cancer tumors can be used to model the tumor microenvironment in vitro. Patient-derived scaffolds are generated by repeated detergent washing, removing all cells. Here, we analyzed the protein composition of 15 decellularized PDSs using liquid chromatography-mass spectrometry/mass spectrometry. One hundred forty-three proteins were detected and their relative abundance was calculated using a reference sample generated from all PDSs. We performed heatmap analysis of all the detected proteins to display their expression patterns across different PDSs together with pathway enrichment analysis to reveal which processes that were connected to PDS protein composition. This protein dataset together with clinical information is useful to investigators studying the microenvironment of breast cancers. Further, after repopulating PDSs with either MCF7 or MDA-MB-231 cells, we quantified their gene expression profiles using RNA sequencing. These data were also compared to cells cultured in conventional 2D conditions, as well as to cells cultured as xenografts in immune-deficient mice. We investigated the overlap of genes regulated between these different culture conditions and performed pathway enrichment analysis of genes regulated by both PDS and xenograft cultures compared to 2D in both cell lines to describe common processes associated with both culture conditions. Apart from our described analyses of these systems, these data are useful when comparing different experimental model systems. Downstream data analyses and interpretations can be found in the research article "Patient-derived scaffolds uncover breast cancer promoting properties of the microenvironment" [1].

DendroPrime as an adhesion barrier on fracture fixation plates: an experimental study in rabbits.

We tested the anti-adhesional effect of a new thiol-ene-based coating in a rabbit model. In 12 New Zealand white rabbits, the periosteum and cortex of the proximal phalanx of the second toe of both hind paws was scratched. Stainless steel plates were fixated with screws. One plate was coated with DendroPrime and the other left bare. The non-operated second toes of both hind paws of an additional four rabbits served as controls. Seven weeks after surgery, the soft tissue adhesion to the plates was evaluated macroscopically, and joint mobility was measured biomechanically. Toe joint mobility was about 20% greater and statistically significant in specimens with coated plates compared with the bare plates. Soft tissue overgrowth and, in some cases, synovitis or adhesions between the plate and the tendon were observed on all bare plates but not on any of the coated plates. We conclude that the thiol-ene-based coating can improve joint mobility by about 20%. This material has a potential to reduce adhesion around plates in fracture surgery.

Structural Analysis of Experimentally Induced Disc Herniation-Like Changes in the Rat.

INTRODUCTION: A disc herniation has traditionally been considered as disc tissue that has slipped out from an intervertebral disc. However, it was recently suggested that the disc herniation mass is a product of bioactive substances from the disc and that the disc hernia would more likely be scar tissue than herniated disc material. In this study, we aimed to analyze the structural components of experimentally induced disc herniations and compare with scar tissue and nucleus pulposus, in the rat. METHODS: Twenty-eight rats had their L4-5 discs punctured. After three weeks, the nodule that had been formed over the puncture site, scar tissue from the spine musculature, and normal nucleus pulposus were harvested and processed for further analysis. RESULTS: Proteomics analysis demonstrated that the formed nodule was more similar to scar tissue than to nucleus pulposus. Gene expression analysis showed that there was no resemblance between any tissues when looking at inflammatory genes but that, there was a clear resemblance between the nodule and scar tissue when analyzing extracellular matrix-related genes. Analysis of the GAG and polysaccharide size distribution revealed that only the nodule and scar tissue contained the shorter versions, potentially short chain hyaluronic acid that is known to induce inflammatory responses. The hematoxylin and eosin stained sections of the nodule, disc tissue, and scar tissue indicated that the morphology of the nodule and scar tissue was very similar. CONCLUSIONS: The nodule formed after experimental disc puncture, and that resembles a disc hernia, has a more structural resemblance to scar tissue than disc tissue. The nodule is, therefore, more likely to be induced by disc-derived bioactive substances than being formed by herniated disc material.

Patient-derived scaffolds uncover breast cancer promoting properties of the microenvironment.

Tumor cells interact with the microenvironment that specifically supports and promotes tumor development. Key components in the tumor environment have been linked to various aggressive cancer features and can further influence the presence of subpopulations of cancer cells with specific functions, including cancer stem cells and migratory cells. To model and further understand the influence of specific microenvironments we have developed an experimental platform using cell-free patient-derived scaffolds (PDSs) from primary breast cancers infiltrated with standardized breast cancer cell lines. This PDS culture system induced a series of orchestrated changes in differentiation, epithelial-mesenchymal transition, stemness and proliferation of the cancer cell population, where an increased cancer stem cell pool was confirmed using functional assays. Furthermore, global gene expression profiling showed that PDS cultures were similar to xenograft cultures. Mass spectrometry analyses of cell-free PDSs identified subgroups based on their protein composition that were linked to clinical properties, including tumor grade. Finally, we observed that an induction of epithelial-mesenchymal transition-related genes in cancer cells growing on the PDSs were significantly associated with clinical disease recurrences in breast cancer patients. Patient-derived scaffolds thus mimics in vivo-like growth conditions and uncovers unique information about the malignancy-inducing properties of tumor microenvironment.

Anti-infective efficacy of the lactoferrin-derived antimicrobial peptide HLR1r.

Antimicrobial peptides (AMPs) have emerged as a new class of drug candidates for the treatment of infectious diseases. Here we describe a novel AMP, HLR1r, which is structurally derived from the human milk protein lactoferrin and demonstrates a broad spectrum microbicidal action in vitro. The minimum concentration of HLR1r needed for killing ≥99% of microorganisms in vitro, was in the range of 3-50µg/ml for common Gram-negative and Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA), and for the yeast Candida albicans, when assessed in diluted brain-heart infusion medium. We found that HLR1r also possesses anti-inflammatory properties as evidenced by inhibition of tumor necrosis factor alpha (TNF-α) secretion from human monocyte-derived macrophages and by repression of interleukin-6 (IL-6) and plasminogen activator inhibitor-1 (PAI-1) secretion from human mesothelial cells, without any cytotoxic effect observed at the concentration range tested (up to 400µg/ml). HLR1r demonstrated pronounced anti-infectious effect in in vivo experimental models of cutaneous candidiasis in mice and of excision wounds infected with MRSA in rats as well as in an ex vivo model of pig skin infected with S. aureus. In conclusion, HLR1r may constitute a new therapeutic alternative for local treatment of skin infections.

Anti-infectious and anti-inflammatory effects of peptide fragments sequentially derived from the antimicrobial peptide centrocin 1 isolated from the green sea urchin, Strongylocentrotus droebachiensis.

Bacterial resistance against antibiotic treatment has become a major threat to public health. Antimicrobial peptides (AMPs) have emerged as promising alternative agents for treatment of infectious diseases. This study characterizes novel synthetic peptides sequentially derived from the AMP centrocin 1, isolated from the green sea urchin, for their applicability as anti-infective agents.The microbicidal effect of centrocin 1 heavy chain (CEN1 HC-Br), its debrominated analogue (CEN1 HC), the C-terminal truncated variants of both peptides, i.e. CEN1 HC-Br (1-20) and CEN1 HC (1-20), as well as the cysteine to serine substituted equivalent CEN1 HC (Ser) was evaluated using minimal microbicidal concentration assay. The anti-inflammatory properties were assessed by measuring the inhibition of secretion of pro-inflammatory cytokines. All the peptides tested exhibited marked microbicidal and anti-inflammatory properties. No difference in efficacy was seen comparing CEN1 HC-Br and CEN1 HC, while the brominated variant had higher cytotoxicity. C-terminal truncation of both peptides reduced salt-tolerability of the microbicidal effect as well as anti-inflammatory actions. Also, serine substitution of cysteine residue decreased the microbicidal effect. Thus, from the peptide variants tested, CEN1 HC showed the best efficacy and safety profile. Further, CEN1 HC significantly reduced bacterial counts in two different animal models of infected wounds, while Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) failed to develop resistance against this peptide under continued selection pressure. In summary, CEN1 HC appears a promising new antimicrobial agent, and clinical studies are warranted to evaluate the applicability of this AMP for local treatment of infections in man.

Effect of lactoferrin peptide (PXL01) on rabbit digit mobility after flexor tendon repair.

PURPOSE: Restoration of digital function after flexor tendon injuries remains a clinical challenge. Complications such as adhesion formation and tendon rupture can lead to limited hand function. The aim of this study was to compare the effects of the lactoferrin-derived peptide, PXL01, formulated in sodium hyaluronate (SH), with SH alone on joint mobility as an indirect measure of postsurgical adhesion prevention and healing strength of the tendon and to elucidate the most optimal concentration of PXL01. METHODS: Using a rabbit flexor tendon repair model, in which the deep flexor tendon was fully transected and repaired, PXL01 in SH or SH alone was administered between the repaired tendon and the tendon sheath before closure of the surgical wound. Three concentrations of PXL01 in SH (5, 20, or 40 mg/mL) were compared to determine the lowest effective concentration. The repaired tendons were evaluated 7 weeks after surgery by measuring the proximal interphalangeal joint mobility by full range of flexion assessment and the tendon repair strength. RESULTS: Treatment with PXL01 formulated in SH resulted in improved mobility of the proximal interphalangeal joint with an average of 10°, corresponding to improvement of approximately 25% to 60% of the flexion of nonoperated toes at the different measuring points compared with SH alone. The difference was statistically significant in 5 out of 6 measuring points (0.5, 1, 2, 3, and 4 N; P < .05). The dose-response study indicated that the lowest effective concentration of PXL01 was 20 mg/mL. There was no difference in healing strength of the tendon between the groups as assessed by load-to-failure breaking strength. CONCLUSIONS: PXL01 in SH significantly improved the mobility compared with the carrier SH alone, without any negative effect on healing strength, and PXL01 at 20 mg/mL was the lowest effective concentration. CLINICAL RELEVANCE: The result provides a valuable basis for a clinical trial to assess efficacy and safety of PXL01 in clinical hand surgery.

N-CAM exhibits a regulatory function in pathological angiogenesis in oxygen induced retinopathy.

BACKGROUND: Diabetic retinopathy and retinopathy of prematurity are diseases caused by pathological angiogenesis in the retina as a consequence of local hypoxia. The underlying mechanism for epiretinal neovascularization (tuft formation), which contributes to blindness, has yet to be identified. Neural cell adhesion molecule (N-CAM) is expressed by Müller cells and astrocytes, which are in close contact with the retinal vasculature, during normal developmental angiogenesis. METHODOLOGY/PRINCIPAL FINDINGS: Notably, during oxygen induced retinopathy (OIR) N-CAM accumulated on astrocytes surrounding the epiretinal tufts. Here, we show that N-CAM ablation results in reduced vascular tuft formation due to reduced endothelial cell proliferation despite an elevation in VEGFA mRNA expression, whereas retinal developmental angiogenesis was unaffected. CONCLUSION/SIGNIFICANCE: We conclude that N-CAM exhibits a regulatory function in pathological angiogenesis in OIR. This is a novel finding that can be of clinical relevance in diseases associated with proliferative vasculopathy.

Model for assessment of mobility of toes and healing of tendons in rabbits.

Repair of a transected flexor tendon will, despite careful technique and early rehabilitation, usually result in a restricted range of movement. This is mainly because adhesions form between the tendon and the surrounding structures. Our aim was to establish an experimental model in rabbits for future studies on new techniques to reduce the formation of adhesions after zone II repair of flexor tendons. In rabbits' hind paws the metatarsal bones II, IV, and V were removed and the flexor tendon was freed to the metatarsophalangeal (MTP) joint. The digits were secured in a specifically-designed biomechanical testing device comprising a servo-hydraulic actuator that was designed to apply controlled force or displacement. The tests were videotaped with a digital force-monitor behind the tested digit. Paper printouts from the recordings were obtained for 0, 0.5, 1, 2, 3, 4, and 5 Newton (N) and metatarsophalangeal, proximal interphalangeal, and distal interphalangeal, angles and distances between metatarsophalangeal joints and claws were measured. The tensile strength of the tendon was evaluated by a load-to-failure test. The continuous data obtained from the experiments were used to calculate functional stiffness at the selected forces. The model allows for unique continuous recordings of mobility of toes, thereby indirectly quantifying the presence of adhesions and the assessment of tensile strength. The data are reproducible, and there is little variation between the digits tested. The model is primarily intended to compare data among treated and non-treated digits of methods to limit the formation of adhesions after tendons have been repaired.