Search

Knee osteoarthritis, the most prevalent degenerative joint disorder worldwide, is driven by chronic low-grade inflammation and subsequent cartilage degradation. Clinical data on the role of the Hoffa or infrapatellar fat pad in knee osteoarthritis are, however, scarce. The infrapatellar fat pad is a richly innervated intracapsular, extrasynovial adipose tissue, and an abundant source of adipokines and proinflammatory and catabolic cytokines, which may contribute to chronic synovial inflammation, cartilage destruction, and subchondral bone remodeling during knee osteoarthritis. How the infrapatellar fat pad interacts with neighboring tissues is poorly understood. Here, we review available literature with regard to the infrapatellar fat pad's interactions with cartilage, synovium, bone, menisci, ligaments, and nervous tissue during the development and progression of knee osteoarthritis. Signaling cascades are described with a focus on immune cell populations, pro- and anti-inflammatory cytokines, adipokines, mesenchymal stromal cells, and molecules derived from conditioned media from the infrapatellar fat pad. Understanding the complex interplay between the infrapatellar fat pad and its neighboring articular tissues may help to better understand and treat the multifactorial pathogenesis of osteoarthritis.

AIMS Giant cell tumour of bone (GCTB) is histologically defined as a lesion containing reactive giant cells and a neoplastic mononuclear cell population; in up to 92% of cases, GCTB is characterised by a specific mutation of the histone gene H3F3A. The cellular composition ranges from giant-cell-rich to giant-cell-poor. The diagnosis of GCTB can be challenging, and several other lesions need to be excluded, e.g. aneurysmal bone cysts, non-ossifying fibromas, chondroblastomas, brown tumours, and giant-cell-rich osteosarcomas. Our aim was to analyse the clinical history, imaging, molecular pathology and histology of three H3F3A-mutated bone tumours without detectable giant cells. None of the patients received denosumab therapy. METHODS AND RESULTS Diagnostic material was obtained by curettage or resection and/or biopsy. Common histomorphological features of all three reported lesions were fibrocytic, oval cells in a background of osteoid and an absence of multinuclear giant cells as confirmed with CD68 immunohistochemistry. We used immunohistochemistry and Sanger sequencing to demonstrate positivity for the H3.3 p.G34W mutation. Differential diagnoses were systematically excluded on the basis of histomorphology, immunohistochemistry, and fluorescence in-situ hybridisation. The imaging (radiography, computed tomography, and magnetic resonance imaging) for all three cases is presented and discussed. CONCLUSIONS We believe that these GCTBs without giant cells expand one end of the heterogeneous range of GCTB. Because of the lack of giant cells, correct diagnosis of GCTB is challenging or even impossible on histological grounds alone. In these cases, detection of the characteristic H3F3A mutation (G34W-specific antibody RM263 or sequencing) is extremely helpful for diagnosing those lesions without giant cells as giant cell tumours of bone.

Giant cell tumour of bone (GCTB) comprises the eponymous osteoclastic multinucleated giant cells eliciting bone lysis, an H3F3A-mutated neoplastic mononucleated fibroblast-like cell population, and H3F3A wild-type mononucleated stromal cells. In this study, we characterised four new cell lines from GCTB. Furthermore, we compared the genome-wide DNA methylation profile of 13 such tumours and three further cell lines with giant cell-rich lesions comprising three H3F3B-mutated chondroblastomas, three USP6-rearranged aneurysmal bone cysts, three non-ossifying fibromas, two hyperparathyroidism-associated brown tumours as well as mesenchymal stem cells, osteoblasts, and osteoclasts. In an unsupervised analysis, we delineated GCTB and chondroblastomas from the other analysed tumour entities. Using comparative methylation analysis, we demonstrated that the methylation pattern of the cell lines approximately equals that of H3F3A-mutated stromal cells in tissue. These patterns more resemble that of osteoblasts than that of mesenchymal stem cells, which argues for the osteoblast as the cell of origin of giant cell tumours of bone. Using enrichment analysis, we detected distinct hypermethylated clusters containing histone and collagen genes as well as target genes of the tumour suppressor p53. We found that the promotor regions of CDKN1A, CDKN2A, and IGFBP3 are methylated more strongly in GCTB than in the other giant cell-containing lesions, mesenchymal stem cells, osteoblasts, and osteoclasts (p 0.001). This hypermethylation correlates with the lower gene expression at the mRNA level for these three genes in the cell lines, the lack of p16 and p21 in these cell lines, and the lower expression of p16 and p21 in GCTB. Overall, our analysis reveals characteristic DNA methylation patterns of giant cell tumours of bone and chondroblastomas and shows that cell lines of giant cell tumours of bone are a valid model for further analysis of H3F3A-mutated tumour cells. © 2022 The Authors. The Journal of Pathology published by John WileySons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Donor variation is a prominent critical issue limiting the applicability of cell-based therapies. We hypothesized that batch effects during propagation of bone marrow stromal cells (BMSCs) in human platelet lysate (hPL), replacing fetal bovine serum (FBS), can affect phenotypic and functional variability. We therefore investigated the impact of donor variation, hPL- vs. FBS-driven propagation and exhaustive proliferation, on BMSC epigenome, transcriptome, phenotype, coagulation risk and osteochondral regenerative function. Notably, propagation in hPL significantly increased BMSC proliferation, created significantly different gene expression trajectories and distinct surface marker signatures, already after just one passage. We confirmed significantly declining proliferative potential in FBS-expanded BMSC after proliferative challenge. Flow cytometry verified the canonical fibroblastic phenotype in culture-expanded BMSCs. We observed limited effects on DNA methylation, preferentially in FBS-driven cultures, irrespective of culture duration. The clotting risk increased over culture time. Moreover, expansion in xenogenic serum resulted in significant loss of function during 3D cartilage disk formation and significantly increased clotting risk. Superior chondrogenic function under hPL-conditions was maintained over culture. The platelet blood group and isoagglutinins had minor impact on BMSC function. These data demonstrate pronounced batch effects on BMSC transcriptome, phenotype and function due to serum factors, partly outcompeting donor variation after just one culture passage.

Particles released from cobalt-chromium-molybdenum (CoCrMo) alloys are considered common elicitors of chronic inflammatory adverse effects. There is a lack of data demonstrating particle numbers, size distribution and elemental composition of bone marrow resident particles which would allow for implementation of clinically relevant test strategies in bone marrow models at different degrees of exposure. The aim of this study was to investigate metal particle exposure in human periprosthetic bone marrow of three types of arthroplasty implants. Periprosthetic bone marrow sections from eight patients exposed to CoCrMo particles were analyzed via spatially resolved and synchrotron-based nanoscopic X-ray fluorescence imaging. These analyses revealed lognormal particle size distribution patterns predominantly towards the nanoscale. Analyses of particle numbers and normalization to bone marrow volume and bone marrow cell number indicated particle concentrations of up to 1 × 1011 particles/ml bone marrow or 2 × 104 particles/bone marrow cell, respectively. Analyses of elemental ratios of CoCrMo particles showed that particularly the particles’ Co content depends on particle size. The obtained data point towards Co release from arthroprosthetic particles in the course of dealloying and degradation processes of larger particles within periprosthetic bone marrow. This is the first study providing data based on metal particle analyses to be used for future in vitro and in vivo studies of possible toxic effects in human bone marrow following exposure to arthroprosthetic CoCrMo particles of different concentration, size, and elemental composition.

The influence of a SARS-CoV-2 infection on inflammatory bowel disease (IBD) has not yet been well characterized and it is unclear whether this requires an adaptation of the immunosuppressive therapy.A national register was established for the retrospective documentation of clinical parameters and changes in immunosuppressive therapy in SARS-CoV-2 infected IBD patients.In total, only 3 of 185 IBD patients (1.6 %) were tested for SARS-CoV-2 infection because of abdominal symptoms. In the course of COVID-19 disease, 43.5 % developed diarrhea, abdominal pain or hematochezia (risk of hospitalization with vs. without abdominal symptoms: 20.0 % vs. 10.6 %, p 0.01). With active IBD at the time of SARS-CoV-2 detection, there was an increased risk of hospitalization (remission 11.2 %, active IBD 23.3 % p 0.05). IBD-specific therapy remained unchanged in 115 patients (71.4 %); the most common change was an interruption of systemic therapy (16.2 %).New abdominal symptoms often appeared in SARS-CoV-2 infected IBD patients. However, these only rarely led to SARS-CoV-2 testing. A high IBD activity at the time of SARS-CoV-2 detection was associated with an increased risk of hospitalization.Der Einfluss einer SARS-CoV-2-Infektion auf chronisch entzündliche Darmerkrankungen (CED) ist bislang nicht gut charakterisiert, und es ist unklar, ob diese eine Anpassung der immunsuppressiven Therapie erfordert.Für die retrospektive Dokumentation klinischer Parameter und Veränderungen einer immunsuppressiven Therapie von mit SARS-CoV-2 infizierten CED-Patienten wurde ein nationales Melderegister etabliert.Insgesamt wurden nur 3 von 185 CED-Patienten (1,6 %) wegen abdomineller Symptome auf eine SARS-CoV-2-Infektion getestet. Im COVID-19-Krankheitsverlauf entwickelten 43,5 % Durchfall, abdominelle Schmerzen oder Hämatochezie (Hospitalisierungsrisiko mit vs. ohne abdominelle Symptome: 20,0 % vs. 10,6 %, p 0,01). Bei aktiver CED zum Zeitpunkt des SARS-CoV-2-Nachweises bestand ein erhöhtes Hospitalisierungsrisiko (Remission 11,2 %, aktive CED 23,3 % p 0,05). Die CED-spezifische Therapie blieb bei 115 Patienten (71,4 %) unverändert; die häufigste Änderung bestand in einer Unterbrechung der systemischen Therapie (16,2 %).Bei mit SARS-CoV-2 infizierten CED-Patienten traten häufig neue abdominelle Symptome bei Infektion auf. Diese führten aber nur selten zur SARS-CoV-2-Testung. Eine hohe CED-Aktivität zum Zeitpunkt des SARS-CoV-2-Nachweises war mit einem erhöhten Hospitalisierungsrisiko assoziiert.

The therapeutic efficacy of clinically applied mesenchymal stromal cells (MSCs) is limited due to their injection into harsh in vivo environments, resulting in the significant loss of their secretory function upon transplantation. A potential strategy for preserving their full therapeutic potential is encapsulation of MSCs in a specialized protective microenvironment, for example hydrogels. However, commonly used injectable hydrogels for cell delivery fail to provide the bio-instructive cues needed to sustain and stimulate cellular therapeutic functions. Here we introduce a customizable collagen I-hyaluronic acid (COL-HA)-based hydrogel platform for the encapsulation of MSCs. Cells encapsulated within COL-HA showed a significant expansion of their secretory profile compared to MSCs cultured in standard (2D) cell culture dishes or encapsulated in other hydrogels. Functionalization of the COL-HA backbone with thiol-modified glycoproteins such as laminin led to further changes in the paracrine profile of MSCs. In depth profiling of more than 250 proteins revealed an expanded secretion profile of proangiogenic, neuroprotective and immunomodulatory paracrine factors in COL-HA-encapsulated MSCs with a predicted augmented pro-angiogenic potential. This was confirmed by increased capillary network formation of endothelial cells stimulated by conditioned media from COL-HA-encapsulated MSCs. Our findings suggest that encapsulation of therapeutic cells in a protective COL-HA hydrogel layer provides the necessary bio-instructive cues to maintain and direct their therapeutic potential. Our customizable hydrogel combines bioactivity and clinically applicable properties such as injectability, on-demand polymerization and tissue-specific elasticity, all features that will support and improve the ability to successfully deliver functional MSCs into patients.

Intravascular infusion is the most popular route for therapeutic multipotent mesenchymal stromal/stem cell (MSC) delivery in hundreds of clinical trials. Meta-analysis has demonstrated that bone marrow MSC infusion is safe. It is not clear if this also applies to diverse new cell products derived from other sources, such as adipose and perinatal tissues. Different MSC products display varying levels of highly procoagulant tissue factor (TF) and may adversely trigger the instant blood-mediated inflammatory reaction (IBMIR). Suitable strategies for assessing and controlling hemocompatibility and optimized cell delivery are crucial for the development of safer and more effective MSC therapies.

Porous tantalum components are widely used for complex acetabular reconstructions in revision hip arthroplasty. Multiple other metal alloys such as titanium-aluminum-vanadium or cobalt-chromium-molybdenum are principally used in artificial joint setups. We report a case of tantalum component failure being both cause and effect of a multiple metal exposure. Our aims were to assess and to characterize associated particle exposure and biological consequences. Metal level quantification revealed substantial in vivo exposure to particulate and dissociated tantalum, zirconium, chromium, cobalt, molybdenum, titanium, aluminum and vanadium in periprosthetic compartments. Aside from micron-sized particles, nanoparticles of a broad size range and elemental composition were obtained. Histological exams verified a spectrum of necrotic changes in the periprosthetic tissues. In the presented case tantalum release was accompanied by concomitance of particles originating from other utilized metals. We conclude that an overall in vivo exposure assessment is mandatory for realistic appraisal of metal toxicity and associated risks.

Placenta-derived decidua stromal cells (DSCs) are being investigated as an alternative to other sources of mesenchymal stromal cells (MSCs) for cellular therapy. DSCs are more effective in treating acute inflammatory diseases in human and this is our preclinical safety study of human DSCs in Sprague-Dawley rats and Balb/c mice. Human DSCs were cultured and expanded from fetal membranes obtained from placentas following cesarean section. In rats, 0.5 × 106 cells/kg were injected intravenously (n = 4) or intra-aortal (n = 4). In mice, DSCs were given intravenously at doses ranging from 4-40 × 106 cells/kg (total of n = 120 mice). In vivo tracking of human cells in mice was performed by using transduced DSC with luciferin gene, and in rats by using 18F-FDG PET. Clotting parameters were determined in vitro and in vivo. All intra-arterially DSC-treated rats had normal motility and behavior and histological examination was normal for liver, spleen kidneys and thigh muscles. Mice treated with DSCs showed no immediate or long-term side effects. None of the mice died or showed acute toxicity or adverse reactions 3 and 30 days after DSC infusion. Murine blood biochemistry profiles related to liver, kidney, heart, and inflammatory indices was not influenced by DSC infusion and complete blood counts were normal. In vivo tracking of infused DSCs detected a signal in the lungs for up to 4 days post infusion. Compared to bone marrow derived MSCs, the DSCs had better viability, smaller size, but stronger clotting in human blood and plasma. Both MSC- and DSC-induced coagulation and complement activation markers, thrombin-anti-thrombin complex (TAT) and C3a, and in vitro clotting parameters were decreased by heparin supplementation. In conclusion, DSCs are safe with almost no side effects even with doses 40 times higher than are used clinically, particularly when supplemented with low-dose heparin.

Osteosclerotic metaphyseal dysplasia (OSMD) is a rare autosomal recessive sclerosing skeletal dysplasia. We report on a 34-year-old patient with sandwich vertebrae, platyspondyly, osteosclerosis of the tubular bones, pathologic fractures, and anemia. In the third decade, he developed osteonecrosis of the jaws, which was progressive in spite of repeated surgical treatment over a period of 11 years. An iliac crest bone biopsy revealed the presence of hypermineralized cartilage remnants, large multinucleated osteoclasts with abnormal morphology, and inadequate bone resorption typical for osteoclast-rich osteopetrosis. After exclusion of mutations in TCIRG1 and CLCN7 we performed trio-based exome sequencing. The novel homozygous splice-site mutation c.261G>A in the gene LRRK1 was found and co-segregated with the phenotype in the family. cDNA sequencing showed nearly complete skipping of exon 3 leading to a frameshift (p.Ala34Profs*33). Osteoclasts differentiated from the patient's peripheral blood monocytes were extremely large. Instead of resorption pits these cells were only capable of superficial erosion. Phosphorylation of L-plastin at position Ser5 was strongly reduced in patient-derived osteoclasts showing a loss of function of the mutated LRRK1 kinase protein. Our analysis indicates a strong overlap of LRRK1-related OSMD with other forms of intermediate osteopetrosis, but an exceptional abnormality of osteoclast resorption. Like in other osteoclast pathologies an increased risk for progressive osteonecrosis of the jaws should be considered in OSMD, an intermediate form of osteopetrosis. © 2020 The Authors. Journal of Bone and Mineral Research published by American Society for Bone and Mineral Research.

Adipose tissue engineering aims to provide solutions to patients who require tissue reconstruction following mastectomies or other soft tissue trauma. Mesenchymal stromal cells (MSCs) robustly differentiate into the adipogenic lineage and are attractive candidates for adipose tissue engineering. This work investigates whether pore size modulates adipogenic differentiation of MSCs toward identifying optimal scaffold pore size and whether pore size modulates spatial infiltration of adipogenically differentiated cells. To assess this, extrusion-based 3D printing is used to fabricate photo-crosslinkable gelatin-based scaffolds with pore sizes in the range of 200-600 mu m. The adipogenic differentiation of MSCs seeded onto these scaffolds is evaluated and robust lipid droplet formation is observed across all scaffold groups as early as after day 6 of culture. Expression of adipogenic genes on scaffolds increases significantly over time, compared to TCP controls. Furthermore, it is found that the spatial distribution of cells is dependent on the scaffold pore size, with larger pores leading to a more uniform spatial distribution of adipogenically differentiated cells. Overall, these data provide first insights into the role of scaffold pore size on MSC-based adipogenic differentiation and contribute toward the rational design of biomaterials for adipose tissue engineering in 3D volumetric spaces.

Severe injury to the skeletal muscle often results in the formation of scar tissue, leading to a decline in functional performance. Traditionally, tissue engineering strategies for muscle repair have focused on substrates that promote myogenic differentiation of transplanted cells. In the current study, the reported data indicates that mesenchymal stromal cells (MSCs) transplanted via porous alginate cryogels promote muscle regeneration by secreting bioactive factors that profoundly influence the function of muscle progenitor cells. These cellular functions, which include heightened resistance of muscle progenitor cells to apoptosis, migration to site of injury, and prevention of premature differentiation are highly desirable in the healing cascade after acute muscle trauma. Furthermore, stimulation of MSCs with recombinant growth factors IGF-1 and VEGF165 was found to significantly enhance their paracrine effects on muscle progenitor cells. Multifunctional alginate cryogels were then utilized as synthetic niches that facilitate local stimulation of seeded MSCs by providing a sustained release of growth factors. In a clinically relevant injury model, the modulation of MSC paracrine signaling via engineered niches significantly improved muscle function by remodeling scar tissue and promoting the formation of new myofibers, outperforming standalone cell or growth factor delivery.

Diseases that jeopardize the musculoskeletal system and cause chronic impairment are prevalent throughout the Western world. In Germany alone, ~1.8 million patients suffer from these diseases annually, and medical expenses have been reported to reach 34.2bn Euros. Although musculoskeletal disorders are seldom fatal, they compromise quality of life and diminish functional capacity. For example, musculoskeletal disorders incur an annual loss of over 0.8 million workforce years to the German economy. Among these diseases, traumatic skeletal muscle injuries are especially problematic because they can occur owing to a variety of causes and are very challenging to treat. In contrast to chronic muscle diseases such as dystrophy, sarcopenia, or cachexia, traumatic muscle injuries inflict damage to localized muscle groups. Although minor muscle trauma heals without severe consequences, no reliable clinical strategy exists to prevent excessive fibrosis or fatty degeneration, both of which occur after severe traumatic injury and contribute to muscle degeneration and dysfunction. Of the many proposed strategies, cell‐based approaches have shown the most promising results in numerous pre‐clinical studies and have demonstrated success in the handful of clinical trials performed so far. A number of myogenic and non‐myogenic cell types benefit muscle healing, either by directly participating in new tissue formation or by stimulating the endogenous processes of muscle repair. These cell types operate via distinct modes of action, and they demonstrate varying levels of feasibility for muscle regeneration depending, to an extent, on the muscle injury model used. While in some models the injury naturally resolves over time, other models have been developed to recapitulate the peculiarities of real‐life injuries and therefore mimic the structural and functional impairment observed in humans. Existing limitations of cell therapy approaches include issues related to autologous harvesting, expansion and sorting protocols, optimal dosage, and viability after transplantation. Several clinical trials have been performed to treat skeletal muscle injuries using myogenic progenitor cells or multipotent stromal cells, with promising outcomes. Recent improvements in our understanding of cell behaviour and the mechanistic basis for their modes of action have led to a new paradigm in cell therapies where physical, chemical, and signalling cues presented through biomaterials can instruct cells and enhance their regenerative capacity. Altogether, these studies and experiences provide a positive outlook on future opportunities towards innovative cell‐based solutions for treating traumatic muscle injuries—a so far unmet clinical need.

Vascularization of the fracture site and cell-mediated deposition of the mineralized matrix are crucial determinants for successful bone regeneration after injury. Ceramic biomaterials such as bioactive glasses (BAGs) that release bioactive ions have shown promising results in bone defect regeneration. However, it remains unclear how the dosage and composition of bioactive ions influence the angiogenic and osteogenic behavior of primary human mesenchymal stromal cells (MSCs). Here, we show that exposure to ionic dissolution products from 1393 and 45S5 BAGs can evoke distinct angiogenic and osteogenic responses from primary MSCs in a dose- and composition-dependent manner. Significantly higher concentrations of the pro-angiogenic factors VEGF, HGF, PIGF, angiopoietin, and angiogenin were detected in conditioned media (CM) from MSCs exposed to 45S5, but not 1393, BAGs. Application of this CM to human umbilical vein endothelial cells (HUVECs) resulted in robust 2D tube formation in vitro. Osteogenic differentiation of MSCs was assessed by gene expression analysis and mineralization assays. Low concentrations (0.1% w/v) of 1393 BAGs significantly enhanced the gene expression of RUNX2 and ALP and induced an earlier onset of matrix mineralization compared to all other groups. We further tested whether simultaneous exposure to both BAGs would improve both angiogenic secretion and osteogenic differentiation of MSCs, and did not find evidence to support this hypothesis. Our results provide evidence of BAG composition-dependent enhancement of primary human MSCs' regenerative function, besides also underlining the importance of an in vitro evaluation of the dose-response relationship to translate BAG based approaches into safe and effective clinical therapies. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2827-2837, 2018., 2018.

An induced pluripotent stem cell line was generated from primary human bone marrow derived mesenchymal stromal cells of a 74year old donor using retroviruses harboring OCT4, SOX2, KLF4 and c-MYC in combination with the following inhibitors TGFβ receptor-SB 431542, MEK-PD325901, and p53-Pifithrin α. Pluripotency was confirmed both in vitro and in vivo.

An imbalance between matrix metalloproteases (MMPs) and the tissue inhibitors of metalloproteases (TIMPs) may have a negative impact on the healing of rotator cuff tears. The aim of the project was to assess a possible relationship between clinical and radiographic characteristics of patients such as the age, sex, as well as the degenerative status of the tendon and the MMPs and TIMPs in their tenocyte-like cells (TLCs). TLCs were isolated from ruptured supraspinatus tendons and quantitative Real-Time PCR and ELISA was performed to analyze the expression and secretion of MMPs and TIMPs. In the present study, MMPs, mostly gelatinases and collagenases such as MMP-2, -9 and -13 showed an increased expression and protein secretion in TLCs of donors with higher age or degenerative status of the tendon. Furthermore, the expression and secretion of TIMP-1, -2 and -3 was enhanced with age, muscle fatty infiltration and tear size. The interaction between MMPs and TIMPs is a complex process, since TIMPs are not only inhibitors, but also activators of MMPs. This study shows that MMPs and TIMPs might play an important role in degenerative tendon pathologies.

Skeletal muscles have a robust capacity to regenerate, but under compromised conditions, such as severe trauma, the loss of muscle functionality is inevitable. Research carried out in the field of skeletal muscle tissue engineering has elucidated multiple intrinsic mechanisms of skeletal muscle repair, and has thus sought to identify various types of cells and bioactive factors which play an important role during regeneration. In order to maximize the potential therapeutic effects of cells and growth factors, several biomaterial based strategies have been developed and successfully implemented in animal muscle injury models. A suitable biomaterial can be utilized as a template to guide tissue reorganization, as a matrix that provides optimum micro-environmental conditions to cells, as a delivery vehicle to carry bioactive factors which can be released in a controlled manner, and as local niches to orchestrate in situ tissue regeneration. A myriad of biomaterials, varying in geometrical structure, physical form, chemical properties, and biofunctionality have been investigated for skeletal muscle tissue engineering applications. In the current review, we present a detailed summary of studies where the use of biomaterials favorably influenced muscle repair. Biomaterials in the form of porous three-dimensional scaffolds, hydrogels, fibrous meshes, and patterned substrates with defined topographies, have each displayed unique benefits, and are discussed herein. Additionally, several biomaterial based approaches aimed specifically at stimulating vascularization, innervation, and inducing contractility in regenerating muscle tissues are also discussed. Finally, we outline promising future trends in the field of muscle regeneration involving a deeper understanding of the endogenous healing cascades and utilization of this knowledge for the development of multifunctional, hybrid, biomaterials which support and enable muscle regeneration under compromised conditions.

Soft tissue trauma of skeletal muscle is one of the most common side effects in surgery. Muscle injuries are not only caused by accident-related injuries but can also be of an iatrogenic nature as they occur during surgical interventions when the anatomical region of interest is exposed. If the extent of trauma surpasses the intrinsic regenerative capacities, signs of fatty degeneration and formation of fibrotic scar tissue can occur, and, consequentially, muscle function deteriorates or is diminished. Despite research efforts to investigate the physiological healing cascade following trauma, our understanding of the early onset of healing and how it potentially determines success or failure is still only fragmentary. This review focuses on the initial physiological pathways following skeletal muscle trauma in comparison to bone and tendon trauma and what conclusions can be drawn from new scientific insights for the development of novel therapeutic strategies. Strategies to support regeneration of muscle tissue after injury are scarce, even though muscle trauma has a high incidence. Based on tissue specific differences, possible clinical treatment options such as local immune-modulatory and cell therapeutic approaches are suggested that aim to support the endogenous regenerative potential of injured muscle tissues.

Background Vascular calcification is enhanced in uraemic chronic haemodialysis patients, likely due to the accumulation of midsize uraemic toxins, such as interleukin 6 (IL-6) and tumor necrosis factor-alpha (TNF-α). Here we have assessed the impact of uraemia on vascular smooth muscle cell (VSMC) calcification and examined the role of IL-6 and TNF-α as possible mediators and, most importantly, its underlying signalling pathway in VSMCs. Methods VSMCs were incubated with samples of uraemic serum obtained from patients treated with haemodialysis for renal failure in the Permeability Enhancement to Reduce Chronic Inflammation-I clinical trial. The VSMCs were assessed for IL-6 gene regulation and promoter activation in response to uraemic serum and TNF-α with reporter assays and electrophoretic mobility shift assay and for osteoblastic transition, cellular calcification and cell viability upon osteogenic differentiation. Results Uraemic serum contained higher levels of TNF-α and IL-6 compared with serum from healthy individuals. Exposure of VSMCs to uraemic serum or recombinant TNF-α lead to a strong upregulation of IL-6 mRNA expression and protein secretion, which was mediated by activator protein 1 (AP-1)/c-FOS-pathway signalling. Uraemic serum induced osteoblastic transition and calcification of VSMCs could be strongly attenuated by blocking TNF-α, IL-6 or AP-1/c-FOS signalling, which was accompanied by improved cell viability. Conclusion These results demonstrate that uraemic serum contains higher levels of uraemic toxins TNF-α and IL-6 and that uraemia promotes vascular calcification through a signalling pathway involving TNF-α, IL-6 and the AP-1/c-FOS cytokine-signalling axis. Thus treatment modalities aiming to reduce systemic TNF-α and IL-6 levels in chronic haemodialysis patients should be evaluated in future clinical trials.

Mesenchymal stromal cells (MSCs) harbor great therapeutic potential for numerous diseases. From early clinical trials, success and failure analysis, bench-to-bedside and back-to-bench approaches, there has been a great gain in knowledge, still leaving a number of questions to be answered regarding optimal manufacturing and quality of MSCs for clinical application. For treatment of many acute indications, cryobanking may remain a prerequisite, but great uncertainty exists considering the therapeutic value of freshly thawed (thawed) and continuously cultured (fresh) MSCs. The field has seen an explosion of new literature lately, outlining the relevance of the topic. MSCs appear to have compromised immunomodulatory activity directly after thawing for clinical application. This may provide a possible explanation for failure of early clinical trials. It is not clear if and how quickly MSCs recover their full therapeutic activity, and if the "cryo stun effect" is relevant for clinical success. Here, we will share our latest insights into the relevance of these observations for clinical practice that will be discussed in the context of the published literature. We argue that the differences of fresh and thawed MSCs are limited but significant. A key issue in evaluating potency differences is the time point of analysis after thawing. To date, prospective double-blinded randomized clinical studies to evaluate potency of both products are lacking, although recent progress was made with preclinical assessment. We suggest refocusing therapeutic MSC development on potency and safety assays with close resemblance of the clinical reality.

Controlled revascularization and inflammation are key elements regulating endogenous regeneration after (bone) tissue trauma. Peripheral blood-derived cell subsets, such as regulatory T-helper cells and circulating (endothelial) progenitor cells, respectively, can support endogenous tissue healing, whereas effector T cells that are associated with an aged immune system can hinder bone regeneration. CD31 is expressed by diverse leukocytes and is well recognized as a marker of circulating endothelial (precursor) cells; however, CD31 is absent from the surface of differentiated effector T cells. Thus, we hypothesized that by separating the inhibitory fractions from the supportive fractions of circulating cells within the peripheral blood (PB) using the CD31 marker, bone regeneration in biologically compromised conditions, such as those observed in aged patients, could be improved. In support of our hypothesis, we detected an inverse correlation between CD31+ cells and effector T cells in the hematomas of human fracture patients, dependent on the age of the patient. Furthermore, we demonstrated the regenerative capacity of human PB-CD31+ cells in vitro. These findings were translated to a clinically relevant rat model of impaired bone healing. The transplantation of rat PB-CD31+ cells advanced bone tissue restoration in vivo and was associated with an early anti-inflammatory response, the stimulation of (re)vascularization, and reduced fibrosis. Interestingly, the depletion or enrichment of the highly abundant CD31+/14+ monocytes from the mixed CD31+ cell population diminished tissue regeneration at different levels, suggesting combined effects within the PB-CD31+ subsets. In summary, an intraoperative enrichment of PB-CD31+ cells might be a novel option to facilitate endogenous regeneration under biologically impaired situations by supporting immunomodulation and vascularization. © 2016 American Society for Bone and Mineral Research.

Because of the multiple possible aetiologies of painful total knee arthroplasty (TKA), the diagnosis and treatment of such patients are challenging. In a considerable number of patients, an intraarticular pathology is present, although not verifiable with clinical and diagnostic imaging techniques as in cases of primary arthrofibrosis. In these patients, the differentiation between intra- and extraarticular causes of pain remains difficult. Until now, little attention has been paid to changes of the synovial fluid and tissue in these knees. The objective of this study was to analyse the changes of the synovial environment in patients suffering from arthrofibrosis after TKA in comparison with knees with referred pain suffering from hip arthritis. The changes of the synovial environment probably provide additional diagnostic information to verify an intraarticular pathology. The synovial fluid of 10 consecutive knees in 10 patients presenting with a primary arthrofibrosis after TKA without signs of infection, instability, malalignment, or loosening was analysed and compared to the synovial fluid of 10 knees with referred pain serving as controls. The BMP-2 concentration was measured in the synovial fluid, and the presence of cytokines leading to an overexpression of BMP-2 was detected by measuring the change of BMP-2 expression in a synoviocyte cell line following exposing to the synovial fluid of the patients. The concentration of BMP-2 in the synovial fluid was significantly higher in arthrofibrotic TKA knees (24.3 ± 6.9 pg/mL), compared with the control group 5.9 ± 4.8 pg/mL (P

Angiogenesis is essential to tissue reconstitution, is sensitive to mechanical stresses, and currently represents one of the major challenges in tissue engineering. The pro-angiogenic matrix metalloprotease-2 (MMP-2) is upregulated in mechanically loaded mesenchymal stem cells (MSCs). Therefore, MMP-2 may provide a regulating link between angiogenesis and the surrounding mechanical conditions. This study aimed to modulate MMP-2 levels by mechanical loading of MSCs embedded in a three-dimensional matrix as well as to investigate the mechanism of MMP-2 regulation along with its contribution to angiogenesis stimulation. MMP-2-inducing conditions (30% compression, 1#x2009;Hz, 72#x2009;h) were defined after varying loading parameters. Addition of the Golgi-disturbing agent Brefeldin A suppressed this mechanical upregulation of MMP-2. Analysis of enzymatic activities demonstrated an enhancement of pro-MMP-2, mature MMP-2, and tissue inhibitor of metalloproteases-2. Further, mechano-regulation of MMP-14 and mature MMP-2 was dependent upon the activity of furin, a proprotein processing endoprotease. Angiogenesis was stimulated by conditioned media from MSCs loaded at inducing conditions. This augmentation of angiogenesis was hindered by inhibition of pro-MMP-2 and mature MMP-2. In conclusion, mechanical stimulation of MSCs in a three-dimensional matrix induces pro-MMP-2 secretion and MMP-2 activation, potentially via the activation complex consisting of MMP-2/-14/tissue inhibitor of metalloproteases-2. Mechano-regulated pro-MMP-2 and mature MMP-2 seem to contribute to angiogenesis stimulation. Thus, an application of these loading parameters could augment vascularization of tissue-engineered constructs based on the described MMP-2-dependent mechanism.

Progenitor cells are involved in the regeneration of the musculoskeletal system, which is known to be influenced by mechanical boundary conditions. Furthermore, matrix metalloproteases (MMPs) and tissue-specific inhibitors of metalloproteases (TIMPs) are crucial for matrix remodelling processes that occur during regeneration of bone and other tissues. This study has therefore investigated whether MMP activity affects mesenchymal stem cell (MSC) behavior and how MMP activity is influenced by the mechanical stimulation of these cells. Broad spectrum inhibition of MMPs altered the migration, proliferation, and osteogenic differentiation of MSCs. Expression analysis detected MMP-2, -3, -10, -11, -13, and -14, as well as TIMP-2, in MSCs at the mRNA and protein levels. Mechanical stimulation of MSCs led to an upregulation of their extracellular gelatinolytic activity, which was consistent with the increased protein levels seen for MMP-2, -3, -13, and TIMP-2. However, mRNA expression levels of MMPs/TIMPs showed no changes in response to mechanical stimulation, indicating an involvement of post-transcriptional regulatory processes such as alterations in MMP secretion or activation. One potential regulatory molecule might be the furin protease. Specific inhibition of MMP-2, -3, and -13 showed MMP-13 to be involved in osteogenic differentiation. The results of this study suggest that MSC function is controlled by MMP activity, which in turn is regulated by mechanical stimulation of cells. Thus, MMP/TIMP balance seems to play an essential role in transferring mechanical signals into MSC function. Disclosure of potential conflicts of interest is found at the end of this article.

The amniotic epithelium consists of cells exhibiting mature epitelial cell characteristics, but also varying degrees of stemness. We tested the hypothesis that induction of epithelial-to-mesenchymal transition (EMT) in amniotic epitelial cells (AECs) derived from human placenta enhances their capacity to support the ischemic myocardium. In response to incubation with transforming growth factor-β1 (TGF-β1) protein, AECs lost their cobblestone morphology and acquired a fibroblastoid shape, associated with downregulation of E-cadherin, upregulation of N-cadherin, Akt phosphorylation, and intracellular periostin translocation. EMT—AECs displayed greatly enhanced mobility and secreted gelatinase activity compared with naive AECs. The surface presentation of CD105 and CD73 decreased, and RNA microarray analysis mirrored the loss of epithelial characteristics and transcriptional profile. Unmodified AECs and EMT—AECs were then injected intramyocardially in fully immunocompetent mice after permanent LAD ligation, and heart function was followed by MRI as well as 2D speckle tracking echocardiography after 4 weeks. EMT—AEC-treated infarct hearts displayed better global systolic function and improved longitudinal strain rate in the area of interest. Although no signals of human cells were detectable by histology, infarct size was smaller in EMT—AEC-treated hearts, associated with fewer TUNEL-positive cells and upregulation of periostin, while blood vessel density was increased in both ACE- and EMT—AEC-treated hearts. We conclude that EMT enhances the cardioprotective effects of human AECs.

Mesenchymal stromal cells (MSCs) are promising candidates in regenerative cell-therapies. However, optimizing their number and route of delivery remains a critical issue, which can be addressed by monitoring the MSCs' bio-distribution in vivo using super-paramagnetic iron-oxide nanoparticles (SPIONs). In this study, amino-polyvinyl alcohol coated (A-PVA) SPIONs are introduced for cell-labeling and visualization by magnetic resonance imaging (MRI) of human MSCs. Size and surface charge of A-PVA-SPIONs differ depending on their solvent. Under MSC-labeling conditions, A-PVA-SPIONs have a hydrodynamic diameter of 42 +/- 2 nm and a negative Zeta potential of 25 +/- 5 mV, which enable efficient internalization by MSCs without the need to use transfection agents. Transmission X-ray microscopy localizes A-PVA-SPIONs in intracellular vesicles and as cytosolic single particles. After identifying non-interfering cell-assays and determining the delivered and cellular dose, in addition to the administered dose, A-PVA-SPIONs are found to be non-toxic to MSCs and non-destructive towards their multi-lineage differentiation potential. Surprisingly, MSC migration is increased. In MRI, A-PVA-SPION-labeled MSCs are successfully visualized in vitro and in vivo. In conclusion, A-PVA-SPIONs have no unfavorable influences on MSCs, although it becomes evident how sensitive their functional behavior is towards SPION-labeling. And A-PVA-SPIONs allow MSC-monitoring in vivo.

Regenerative medicine aims to replace lost cells and to restore damaged tissues and organs by either tissue-engineering approaches or stimulation of endogenous processes. Due to their biological properties, stem cells promise to be an effective source for such strategies. Especially adult multipotent stem cells (ASCs) are believed to be applicable in a broad range of therapies for the treatment of multifactorial diseases or age-related degeneration, although the molecular and cellular mechanisms underlying their regenerative function are often hardly described. Moreover, in some demanding clinical situations their efficiency remains limited. Thus, a basic understanding of ASCs regenerative function, their complex interplay with their microenvironment and how compromising conditions interfere with their efficiency is mandatory for any regenerative strategy. Concerning this matter, the impact of patient-specific constraints are often underestimated in research projects and their influence on the study results disregarded. Thus, researchers are urgently depending on well-characterized tissue samples or cells that are connected with corresponding donor information, such as secondary diseases, medication. Here, we outline principle pitfalls during experimental studies using human samples, and describe a potential strategy to overcome these challenges by establishing a core unit for cell and tissue harvesting. This facility aims to bridge the gap between clinic and research laboratories by the provision of a direct link to the clinical operating theatres. Such a strategy clearly supports basic and clinical research in the conduct of their studies and supplies highly characterized human samples together with the corresponding donor information.

There is growing evidence that adaptive immunity contributes to endogenous regeneration processes: For example, endogenous bone fracture repair is modulated by T cells even in the absence of infection. Because delayed or incomplete fracture healing is associated with poor long-term outcomes and high socioeconomic costs, we investigated the relationship between an individual’s immune reactivity and healing outcome. Our study revealed that delayed fracture healing significantly correlated with enhanced levels of terminally differentiated CD8 + effector memory T (T EMRA ) cells (CD3 + CD8 + CD11a ++ CD28 − CD57 + T cells) in peripheral blood. This difference was long lasting, reflecting rather the individual’s immune profile in response to lifelong antigen exposure than a post-fracture reaction. Moreover, CD8 + T EMRA cells were enriched in fracture hematoma; these cells were the major producers of interferon-γ/tumor necrosis factor–α, which inhibit osteogenic differentiation and survival of human mesenchymal stromal cells. Accordingly, depletion of CD8 + T cells in a mouse osteotomy model resulted in enhanced endogenous fracture regeneration, whereas a transfer of CD8 + T cells impaired the healing process. Our data demonstrate the high impact of the individual adaptive immune profile on endogenous bone regeneration. Quantification of CD8 + T EMRA cells represents a potential marker for the prognosis of the healing outcome and opens new opportunities for early and targeted intervention strategies.

Background:Patient age has a decisive impact on the short- term postoperative results in surgery for carcinoma. Patients and methods: This prospective multicenter study involved 75 German hospitals and 3756 patients undergoing treatment in 1999: 1447 aged under 65 years, 1847 aged 65-79 years, and 458 aged over 80 years. Results: In the oldest patient group, there was a significant- ly higher proportion of extensive, localized tumors (UICC stage II: 25.9%, 28.4%, and 36.1%, respec- tively) and significant differences were found among the three groups in operation rates (98.8%, 98.6%, and 96.5%), resection rate (94.2%, 93.2%, and 83.9%), general postop- erative complications (21.5%, 28.6%, and 41.2%), morbidity (36.5%, 42.6%, and 50.0%) and mortality (2.7%, 6.6%, and 11.8%). Conclusion: In the elderly, locally advanced tumors, but not metastasiz- ing tumors, are to be expected. The increase in postoperative morbidity and mortality rates with increasing age was due to the increase in gener- al postoperative complications. Sur- gery for colorectal carcinoma in pa- tients of advanced age is not associat- ed with any increase in intraoperative or specific postoperative complica- tions.

The interest on applying mesenchymal stromal cells (MSCs) in orthopedic disorders has risen tremendously in the last years due to scientific successes in preclinical in vitro and animal model studies. In a wide range of diseases and injuries of the musculoskeletal system, MSCs are currently under evaluation, but so far have found access to clinical use only in few cases. The current assignment is to translate the acquired knowledge into clinical practice. Therefore, this review aims at presenting a synopsis of the up-to-date status of the use of MSCs and MSC related cell products in musculoskeletal indications. Clinical studies were included, whereas preclinical and animal study data not have been considered. Most studies published so far investigate the final outcome applying bone marrow derived MSCs. In fewer trials the use of adipose tissue derived MSCs and allogenic MSCs was investigated in different applications. Although the reported results are equivocal in the current literature, the vast majority of the studies shows a benefit of MSC based therapies depending on the cell sources and the indication in clinical use. In summary, the clinical use of MSCs in patients in orthopedic indications has been found to be safe. Standardized protocols and clear definitions of the mechanisms of action and the mode and timing of application as well as further coordinated research efforts will be necessary for finally adding MSC based therapies in standard operating procedures and guidelines for the clinicians treating orthopedic disorders.

Background No regenerative approach has thus far been shown to be effective in skeletal muscle injuries, despite their high frequency and associated functional deficits. We sought to address surgical trauma‐related muscle injuries using local intraoperative application of allogeneic placenta‐derived, mesenchymal‐like adherent cells (PLX‐PAD), using hip arthroplasty as a standardized injury model, because of the high regenerative and immunomodulatory potency of this cell type. Methods Our pilot phase I/IIa study was prospective, randomized, double blind, and placebo‐controlled. Twenty patients undergoing hip arthroplasty via a direct lateral approach received an injection of 3.0 × 108 (300 M, n = 6) or 1.5 × 108 (150 M, n = 7) PLX‐PAD or a placebo (n = 7) into the injured gluteus medius muscles. Results We did not observe any relevant PLX‐PAD‐related adverse events at the 2‐year follow‐up. Improved gluteus medius strength was noted as early as Week 6 in the treatment‐groups. Surprisingly, until Week 26, the low‐dose group outperformed the high‐dose group and reached significantly improved strength compared with placebo [150 M vs. placebo: P = 0.007 (baseline adjusted; 95% confidence interval 7.6, 43.9); preoperative baseline values mean ± SE: placebo: 24.4 ± 6.7 Nm, 150 M: 27.3 ± 5.6 Nm], mirrored by an increase in muscle volume [150 M vs. placebo: P = 0.004 (baseline adjusted; 95% confidence interval 6.0, 30.0); preoperative baseline values GM volume: placebo: 211.9 ± 15.3 cm3, 150 M: 237.4 ± 27.2 cm3]. Histology indicated accelerated healing after cell therapy. Biomarker studies revealed that low‐dose treatment reduced the surgery‐related immunological stress reaction more than high‐dose treatment (exemplarily: CD16+ NK cells: Day 1 P = 0.06 vs. placebo, P = 0.07 vs. 150 M; CD4+ T‐cells: Day 1 P = 0.04 vs. placebo, P = 0.08 vs. 150 M). Signs of late‐onset immune reactivity after high‐dose treatment corresponded to reduced functional improvement. Conclusions Allogeneic PLX‐PAD therapy improved strength and volume of injured skeletal muscle with a reasonable safety profile. Outcomes could be positively correlated with the modulation of early postoperative stress‐related immunological reactions.

Numerous clinical trials of mesenchymal stromal/stem cells (MSCs) as a new treatment for coronavirus-induced disease (COVID-19) have been registered recently, most of them based on intravenous (IV) infusion. There is no approved effective therapy for COVID-19, but MSC therapies have shown first promise in the treatment of acute respiratory distress syndrome (ARDS) pneumonia, inflammation, and sepsis, which are among the leading causes of mortality in COVID-19 patients. Many of the critically ill COVID-19 patients are in a hypercoagulable procoagulant state and at high risk for disseminated intravascular coagulation, thromboembolism, and thrombotic multi-organ failure, another cause of high fatality. It is not yet clear whether IV infusion is a safe and effective route of MSC delivery in COVID-19, since MSC-based products express variable levels of highly procoagulant tissue factor (TF/CD142), compromising the cells' hemocompatibility and safety profile. Of concern, IV infusions of poorly characterized MSC products with unchecked (high) TF/CD142 expression could trigger blood clotting in COVID-19 and other vulnerable patient populations and further promote the risk for thromboembolism. In contrast, well-characterized products with robust manufacturing procedures and optimized modes of clinical delivery hold great promise for ameliorating COVID-19 by exerting their beneficial immunomodulatory effects, inducing tissue repair and organ protection. While the need for MSC therapy in COVID-19 is apparent, integrating both innate and adaptive immune compatibility testing into the current guidelines for cell, tissue, and organ transplantation is critical for safe and effective therapies. It is paramount to only use well-characterized, safe MSCs even in the most urgent and experimental treatments. We here propose three steps to mitigate the risk for these vulnerable patients: (1) updated clinical guidelines for cell and tissue transplantation, (2) updated minimal criteria for characterization of cellular therapeutics, and (3) updated cell therapy routines reflecting specific patient needs.