Publications in 2021


Cells. 2021 Feb 1;10(2):298. doi: 10.3390/cells10020298.

 

Mechanical Stress Induce PG-E2 in Murine Synovial Fibroblasts Originating from the Temporomandibular Joint

 

Ute Nazet , Laura Feulner , Dominique Muschter, Patrick Neubert, Valentin Schatz, Susanne Grässel, Jonathan Jantsch, Peter Proff , Agnes Schröder, Christian Kirschneck (SP4)                     

 

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Abstract

Genetic predisposition, traumatic events, or excessive mechanical exposure provoke arthritic changes in the temporomandibular joint (TMJ). We analysed the impact of mechanical stress that might be involved in the development and progression of TMJ osteoarthritis (OA) on murine synovial fibroblasts (SFs) of temporomandibular origin. SFs were subjected to different protocols of mechanical stress, either to a high-frequency tensile strain for 4 h or to a tensile strain of varying magnitude for 48 h. The TMJ OA induction was evaluated based on the gene and protein secretion of inflammatory factors (Icam-1Cxcl-1Cxcl-2Il-1ßIl-1raIl-6Ptgs-2, PG-E2), subchondral bone remodelling (RanklOpg), and extracellular matrix components (Col1a2Has-1, collagen and hyaluronic acid deposition) using RT-qPCR, ELISA, and HPLC. A short high-frequency tensile strain had only minor effects on inflammatory factors and no effects on the subchondral bone remodelling induction or matrix constituent production. A prolonged tensile strain of moderate and advanced magnitude increased the expression of inflammatory factors. An advanced tensile strain enhanced the Ptgs-2 and PG-E2 expression, while the expression of further inflammatory factors were decreased. The tensile strain protocols had no effects on the RANKL/OPG expression, while the advanced tensile strain significantly reduced the deposition of matrix constituent contents of collagen and hyaluronic acid. The data indicates that the application of prolonged advanced mechanical stress on SFs promote PG-E2 protein secretion, while the deposition of extracellular matrix components is decreased.

 

Keywords

temporomandibular joint; osteoarthritis; synovitis; inflammation; mechanical strain

 


Cell Signal 2021 Feb 8;109948.  doi: 10.1016/j.cellsig.2021.109948. Online ahead of print.

 

Adrenergic signalling in osteoarthritis

 

Rebecca Sohn 1Gundula Rösch 1Marius Junker 1Andrea Meurer 1Frank Zaucke 1Zsuzsa Jenei-Lanzl 2

 

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Abstract

Adrenoceptors (ARs) mediate the effects of the sympathetic neurotransmitters norepinephrine (NE) and epinephrine (E) in the human body and play a central role in physiologic and pathologic processes. Therefore, ARs have long been recognized as targets for therapeutic agents, especially in the field of cardiovascular medicine. During the past decades, the contribution of the sympathetic nervous system (SNS) and particularly of its major peripheral catecholamine NE to the pathogenesis of osteoarthritis (OA) attracted growing interest. OA is the most common degenerative joint disorder worldwide and a disease of the whole joint. It is characterized by progressive degradation of articular cartilage, synovial inflammation, osteophyte formation, and subchondral bone sclerosis mostly resulting in chronic pain. The subchondral bone marrow, the periosteum, the synovium, the vascular meniscus and numerous tendons and ligaments are innervated by tyrosine hydroxylase-positive (TH+) sympathetic nerve fibers that release NE into the synovial fluid and cells of all abovementioned joint tissues express at least one out of nine AR subtypes. During the past decades, several in vitro studies explored the AR-mediated effects of NE on different cell types in the joint. So far, only a few studies used animal OA models to investigate the contribution of distinct AR subtypes to OA pathogenesis in vivo. This narrative review shortly summarizes the current background knowledge about ARs and their signalling pathways at first. In the second part, we focus on recent findings in the field of NE-induced AR-mediated signalling in different joint tissues during OA pathogenesis and at the end, we will delineate the potential of targeting the adrenergic signalling for OA prevention or treatment. We used the PubMed bibliographic database to search for keywords such as 'joint' or 'cartilage' or 'synovium' or 'bone' and 'osteoarthritis' and/or 'trauma' and 'sympathetic nerve fibers' and/or 'norepinephrine' and 'adrenergic receptors / adrenoceptors' as well as 'adrenergic therapy'.

 

 

Keywords: Adrenoceptors; Norepinephrine; Osteoarthritis; Sympathetic nervous system.

 


Publications in 2020


Int J Mol Sci 2020 May 30;21(11):3924.  doi: 10.3390/ijms21113924.

 

Norepinephrine Inhibits the Proliferation of Human Bone Marrow-Derived Mesenchymal Stem Cells via β2-Adrenoceptor-Mediated ERK1/2 and PKA Phosphorylation

 

Jessica Hedderich 1Karima El Bagdadi 1Peter Angele 2Susanne Grässel 3Andrea Meurer 1Rainer H Straub 4Frank Zaucke 1Zsuzsa Jenei-Lanzl 1

 

 

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Abstract

Bone marrow-derived mesenchymal stem cells (BMSCs) represent an alternative to chondrocytes to support cartilage regeneration in osteoarthritis (OA). The sympathetic neurotransmitter norepinephrine (NE) has been shown to inhibit their chondrogenic potential; however, their proliferation capacity under NE influence has not been studied yet. Therefore, we used BMSCs obtained from trauma and OA donors and compared the expression of adrenergic receptors (AR). Then, BMSCs from both donor groups were treated with NE, as well as with combinations of NE and α1-, α2- or β1/2-AR antagonists (doxazosin, yohimbine or propranolol). Activation of AR-coupled signaling was investigated by analyzing ERK1/2 and protein kinase A (PKA) phosphorylation. A similar but not identical subset of ARs was expressed in trauma (α2B-, α2C- and β2-AR) and OA BMSCs (α2A-, α2B-, and β2-AR). NE in high concentrations inhibited the proliferation of both trauma and OA BMCSs significantly. NE in low concentrations did not influence proliferation. ERK1/2 as well as PKA were activated after NE treatment in both BMSC types. These effects were abolished only by propranolol. Our results demonstrate that NE inhibits the proliferation and accordingly lowers the regenerative capacity of human BMSCs likely via β2-AR-mediated ERK1/2 and PKA phosphorylation. Therefore, targeting β2-AR-signaling might provide novel OA therapeutic options.

 

 

Keywords: bone marrow-derived mesenchymal stem cells (BMSCs); norepinephrine; proliferation; regeneration; sympathicus.


Bone 133 (2020) 115181. doi: 10.1016/j.bone.2019.115181.

 

Sensory neuropeptides are required for bone and cartilage homeostasis in a murine destabilization-induced osteoarthritis model

 

Dominique Muschter, Lutz Fleischhauer, Shahed Taheri, Arndt F.Schilling, Hauke Clausen-Schaumann, Susanne Grässel  (SP4+SP5+SP1)

 

 

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Abstract

Numerous studies identified a role for the sensory neuropeptides substance P (SP) and alpha calcitonin gene-related peptide (αCGRP) in osteoarthritis (OA) pain behavior. Surprisingly, little attention has been paid on how their trophic effects on cartilage and bone cells might affect structural changes of bone and cartilage in OA pathology. Here, we sought to elucidate sensory neuropeptides influence on structural alterations of bone and cartilage during murine OA pathophysiology. OA was induced by destabilization of the medial meniscus (DMM) in the right knee joint of 12 weeks old male C57Bl/6J wildtype (WT) mice and mice either deficient for SP (tachykinin 1 (Tac1)−/−) or αCGRP. By OARSI histopathological grading we observed significant cartilage matrix degradation after DMM surgery in αCGRP-deficient mice after 4 weeks whereas Tac1−/− scores were not different to sham mice before 12 weeks after surgery. Indentation-type atomic force microscopy (IT-AFM) identified a strong superficial zone (SZ) cartilage phenotype in Tac1−/− Sham mice. Opposed to WT and αCGRP−/− mice, SZ cartilage of Tac1−/− mice softened 2 weeks after OA induction. In Tac1−/− DMM mice, bone volume to total volume ratio (BV/TV) increased significantly compared to the Tac1−/− Sham group, 2 weeks after surgery. WT mice had reduced BV/TV compared to αCGRP−/− and Tac1−/− mice after 12 weeks. Increased calcified cartilage thickness and medial condyle diameter were detected in the medial tibia of all groups 8 weeks after OA induction by nanoCT analysis. Meniscal ossification occurred in all OA groups, but was significantly stronger in the absence of neuropeptides. Increased serum concentration of the respective non-deleted neuropeptide was observed in both neuropeptide-deficient mice strains.

Both neuropeptides protect from age-related bone structural changes under physiological conditions and SP additionally demonstrates an anabolic effect on bone structure preservation in a pathophysiological situation. Both neuropeptide deficient mice display an intrinsic structural cartilage matrix phenotype that might alter progression of cartilage degeneration in OA.

 

Keywords

Osteoarthritis, Destabilization of the medial meniscus, Substance P, alpha-Calcitonin gene-related peptide, Bone, Cartilage

 

 

 


Osteologie 2020, DOI: 10.1055/a-1206-6663

 

Knorpel-Knochenmark-Mikro-Konnektoren im subchondralen Knochen

 

Shahed Taheri, Kai O. Böker, Wolfgang Lehmann, Arndt Friedrich Schilling (SP5)                     

 

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Abstract

The interaction of the subchondral bone with the articular cartilage is of fundamental importance for joint physiology. Studies on animal and human models have suggested that the lower margin of the articular cartilage is connected to the medullary cavity via small defects or microchannels. While these cartilage-bone marrow micro-connectors (CMMC) can potentially be crucial for molecular exchange, bone-cartilage crosstalk, and even cartilage nutrition, the current knowledge regarding them is limited and incoherent. This review summarizes the so far described characteristics of this anatomical feature in different species, explores recurring patterns in the literature, and discusses its potential functionality.

 

Keywords

 

Subchondral bone - Bone microstructure - Porosity - Osteoarthritis - Cartilage nutrition

 

 

Keywords: bone marrow-derived mesenchymal stem cells (BMSCs); norepinephrine; proliferation; regeneration; sympathicus.


Review: F1000Res. 2020 May 4;9:F1000 Faculty Rev-325.doi: 10.12688/f1000research.22115.1. eCollection 2020.

 

Recent advances in the treatment of osteoarthritis

 

Susanne Grässel, Dominique Muschter (SP4)                     

 

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Abstract

Osteoarthritis (OA) is one of the most debilitating diseases and is associated with a high personal and socioeconomic burden. So far, there is no therapy available that effectively arrests structural deterioration of cartilage and bone or is able to successfully reverse any of the existing structural defects. Efforts to identify more tailored treatment options led to the development of strategies that enabled the classification of patient subgroups from the pool of heterogeneous phenotypes that display distinct common characteristics. To this end, the classification differentiates the structural endotypes into cartilage and bone subtypes, which are predominantly driven by structure-related degenerative events. In addition, further classifications have highlighted individuals with an increased inflammatory contribution (inflammatory phenotype) and pain-driven phenotypes as well as senescence and metabolic syndrome phenotypes. Most probably, it will not be possible to classify individuals by a single definite subtype, but it might help to identify groups of patients with a predominant pathology that would more likely benefit from a specific drug or cell-based therapy. Current clinical trials addressed mainly regeneration/repair of cartilage and bone defects or targeted pro-inflammatory mediators by intra-articular injections of drugs and antibodies. Pain was treated mostly by antagonizing nerve growth factor (NGF) activity and its receptor tropomyosin-related kinase A (TrkA). Therapies targeting metabolic disorders such as diabetes mellitus and senescence/aging-related pathologies are not specifically addressing OA. However, none of these therapies has been proven to modify disease progression significantly or successfully prevent final joint replacement in the advanced disease stage. Within this review, we discuss the recent advances in phenotype-specific treatment options and evaluate their applicability for use in personalized OA therapy.

 

Keywords

 

Osteoarthritis, OA phenotype, therapy, inflammation, subchondral bone, cartilage, pain, metabolic syndrome, senescence

 


Int J Mol Sci 2020 Mar 18;21(6):2085. doi: 10.3390/ijms21062085.

 

Adrenoceptor Expression during Intervertebral Disc Degeneration

 

Johannes Kupka 1Annika Kohler 1Karima El Bagdadi 1Richard Bostelmann 2Marco Brenneis 1Christoph Fleege 1Danny Chan 3Frank Zaucke 1Andrea Meurer 1Marcus Rickert 1Zsuzsa Jenei-Lanzl 1

 

 

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Abstract

Healthy and degenerating intervertebral discs (IVDs) are innervated by sympathetic nerves, however, adrenoceptor (AR) expression and functionality have never been investigated systematically. Therefore, AR gene expression was analyzed in both tissue and isolated cells from degenerated human IVDs. Furthermore, human IVD samples and spine sections of wildtype mice (WT) and of a mouse line that develops spontaneous IVD degeneration (IVDD, in SM/J mice) were stained for ARs and extracellular matrix (ECM) components. In IVD homogenates and cells α1a-, α1b-, α2a-, α2b-, α2c-, β1-, and β2-AR genes were expressed. In human sections, β2-AR was detectable, and its localization parallels with ECM alterations. Similarly, in IVDs of WT mice, only β2-AR was expressed, and in IVDs of SM/J mice, β2AR expression was stronger accompanied by increased collagen II, collagen XII, decorin as well as decreased cartilage oligomeric matrix protein expression. In addition, norepinephrine stimulation of isolated human IVD cells induced intracellular signaling via ERK1/2 and PKA. For the first time, the existence and functionality of ARs were demonstrated in IVD tissue samples, suggesting that the sympathicus might play a role in IVDD. Further studies will address relevant cellular mechanisms and thereby help to develop novel therapeutic options for IVDD.

 

 

Keywords: IVD degeneration (IVDD); adrenoceptors; intervertebral disc (IVD); sympathicus.

 


Publications in 2019


 

 

Int. J. Mol. Sci. 201920(3), 585; https://doi.org/10.3390/ijms20030585

 

Impact of Mechanical Load on the Expression Profile of Synovial Fibroblasts from Patients with and without Osteoarthritis

 

Agnes Schröder, Ute Nazet, Dominique Muschter, Susanne Grässel, Peter Proff  and Christian Kirschneck

(SP4)                     

 

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Abstract

Osteoarthritis (OA) affects the integrity of the entire joint including the synovium. The most abundant cells in the synovium are fibroblasts (SF). Excessive mechanical loading might contribute to OA pathogenesis. Here, we investigate the effects of mechanical loading on SF derived from non-OA (N-SF) and OA patients (OA-SF). We treated N-SF and OA-SF with or without mechanical loading for 48h after 24h of preincubation. Then we assessed gene and protein expression of proinflammatory factors (TNFα, COX-2, PG-E2, IL-6), extracellular matrix (ECM) components (COL1, FN1) and glycosaminoglycans (GAGs) via RT-qPCR, ELISA, DMMB assay and HPLC. Mechanical loading significantly increased TNFα and PG-E2 secretion by N-SF and OA-SF, whereas in OA-SF IL-6 secretion was reduced. COL1 and FN1 secretion were downregulated in N-SF during loading. OA-SF secreted less COL1 compared to N-SF under control conditions. In contrast, OA-SF in general expressed more FN1. GAG synthesis was upregulated in N-SF, but not in OA-SF during loading with OA-SF displaying a higher charge density than N-SF. Mechanical loading enhanced proinflammatory factor expression and GAG synthesis and decreased secretion of ECM components in N-SFs, indicating a contributing role of SF to OA development.

 

Keywords

osteoarthritis; synovial fibroblasts; mechanical loading; synovium; inflammation; synovitis; compressive force

 

 


Int. J. Mol. Sci. 201920(13), 3127; https://doi.org/10.3390/ijms20133127

 

Norepinephrine Inhibits Synovial Adipose Stem Cell Chondrogenesis via α2a-Adrenoceptor-Mediated ERK1/2 Activation

 

Karima El Bagdadi, Frank Zaucke, Andrea Meurer, Rainer H. Straub, Zsuzsa Jenei-Lanzl 

(SP8)                     

 

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Abstract

In recent years, first evidences emerged that sympathetic neurotransmitters influence osteoarthritis (OA) manifestation. Joint-resident stem cells might contribute to cartilage repair, however, their chondrogenic function is reduced. The neurotransmitter norepinephrine (NE) was detected in the synovial fluid of trauma and OA patients. Therefore, the aim of this study was to analyse how NE influences the chondrogenesis of synovial adipose tissue-derived stem cells (sASCs). sASCs were isolated from knee-OA patients synovia. After adrenoceptor (AR) expression analysis, proliferation and chondrogenic differentiation in presence of NE and/or α- and β-AR antagonist were investigated. Cell count, viability, chondrogenic and hypertophic gene expression, sulfated glycosaminoglycan (sGAG) and type II collagen content were determined. Key AR-dependent signaling (ERK1/2, PKA) was analyzed via western blot. sASC expressed α1A-, α1B-, α2A-, α2B-, α2C-, and β2-AR in monolayer and pellet culture. NE did not affect proliferation and viability, but 10−7 and 10−6 M NE significantly reduced sGAG and type II collagen content as well as ERK1/2 phosphorylation. These effects were fully reversed by yohimbine (α2-AR antagonist). Our study confirms the important role of NE in sASC chondrogenic function and provides new insights in OA pathophysiology. Future studies might help to develop novel therapeutic options targeting neuroendocrine pathways for OA treatment

 

Keywords

synovial adipose stem cells; sympathicus; norepinephrin; adrenoceptors; chondrogenesis; osteoarthritis; physioxoa

 

 


 

 

Eur. Cell Mater. 2019 May 6;37:360-381. doi: 10.22203/eCM.v037a22.

 

Cells under pressure - the relationship between hydrostatic pressure and mesenchymal stem cell chondrogenesis.

 

Pattappa Girish, Zellner Johannes, Johnstone Brian, Docheva Denitsa, Angele Peter

(SP7)                     

 

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Abstract

Early osteoarthritis (OA), characterised by cartilage defects, is a degenerative  disease that greatly affects the adult population. Cell-based tissue engineering methods are being explored as a solution for the treatment of these chondral defects. Chondrocytes are already in clinical use but other cell types with chondrogenic properties, such as mesenchymal stem cells (MSCs), are being researched. However, present methods for differentiating these cells into stable articular-cartilage chondrocytes that contribute to joint regeneration are not effective, despite extensive investigation. Environmental stimuli, such as mechanical forces, influence chondrogenic response and are beneficial with respect to matrix formation. In vivo, the cartilage is subjected to multiaxial loading involving compressive, tensile, shear and fluid flow and cellular response. Tissue formation mechanobiology is being intensively studied in the cartilage tissue-engineering research field. The study of the effects of hydrostatic pressure on cartilage formation belongs to the large area of mechanobiology. During cartilage loading, interstitial fluid is pressurised and the surrounding matrix delays pressure loss by reducing fluid flow rate from pressurised regions. This fluid pressurisation is known as hydrostatic pressure,  where a uniform stress around the cell occurs without cellular deformation. In vitro studies, examining chondrocytes under hydrostatic pressure, have described its anabolic effect and similar studies have evaluated the effect of hydrostatic  pressure on MSC chondrogenesis. The present review summarises the results of these studies and discusses the mechanisms through which hydrostatic pressure exerts its effects.

 

 

 


 

 

Osteoarthritis and Cartilage, 2019. 27(8): p. 1208-1218.

 

Mechanosensitive MiRs Regulated by Anabolic and Catabolic Loading of Human Cartilage

 

Hecht Nicole, Johnstone Brian, Angele Peter, Walker Tilman, Richter Wiltrud.

(SP3 + SP7)                     

 

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Elucidation of whether miRs are involved in mechanotransduction pathways by which cartilage is maintained or disturbed has a particular importance in our understanding of osteoarthritis (OA) pathophysiology. The aim was to investigate whether mechanical loading influences global miR-expression in human chondrocytes and to identify mechanosensitive miRs responding to beneficial and non-beneficial loading regimes as potential to obtain valuable diagnostic or therapeutic targets to advance OA-treatment.

 

 

 

 


 

 

Int. J. Mol. Sci. 201920(5), 1212; doi:10.3390/ijms20051212

 

Role of Norepinephrine in IL-1β-Induced Chondrocyte Dedifferentiation under Physioxia

 

Saskia Speichert, Natalie Molotkov, Karima El Bagdadi, Andrea Meurer, Frank Zaucke, Zsuzsa Jenei-Lanzl

(SP8)                     

 

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As part of the pathogenesis of osteoarthritis (OA), chondrocytes lose their phenotype and become hypertrophic, or dedifferentiate, mainly driven by interleukin-1β (IL-1β). The contribution of other factors to the dedifferentiation process is not completely understood. Recent studies suggested a dose-dependent role for the sympathetic neurotransmitter norepinephrine (NE) in OA chondrocyte metabolism. Therefore, the aim of this study was to analyze the contribution of NE (10−8 M, 10−6 M) to human articular OA chondrocyte dedifferentiation in the absence or presence of IL-1β (0.5 ng/mL). Here, we demonstrate that OA chondrocytes express α2A-, α2C- and β2-adrenoceptors (AR) and show the characteristic shift towards a fibroblast-like shape at day 7 in physioxic monolayer culture. NE alone did not affect morphology but, in combination with IL-1β, markedly accelerated this shift. Moderate glycosaminoglycan (GAG) staining was observed in untreated and NE-treated cells, while IL-1β strongly decreased GAG deposition. IL-1β alone or in combination with NE decreased SOX9, type II collagen, COMP, and aggrecan, and induced MMP13 and ADAMTS4 gene expression, indicating an accelerated dedifferentiation. NE alone did not influence gene expression and did not modulate IL-1β-mediated effects. In conclusion, these results indicate that low-grade inflammation exerts a dominant effect on chondrocyte dedifferentiation and should be targeted early in OA therapy. 

 

Keywords

osteoarthritis; chondrocytes; dedifferentiation; norepinephrine; interleukin-1β; adrenoceptors; physioxia

 

 


 

 

Int J Mol Sci. 2019 Feb 12;20(3). pii: E770. doi: 10.3390/ijms20030770.

 

Developmental Transformation and Reduction of Connective Cavities within the Subchondral Bone.

 

Shahed Taheri, Thomas Winkler, Lia Sabrina Schenk , Carl Neuerburg,, Sebastian Felix Baumbach, Jozef Zustin, Wolfgang Lehmann, Arndt Friedrich Schilling

(SP5)                     

 

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Abstract

It is widely accepted that the subchondral bone (SCB) plays a crucial role in the 15 physiopathology of osteoarthritis (OA), although its contribution is still debated. Much of the pre16 clinical research on the role of SCB is concentrated on comparative evaluations of healthy vs. early 17 OA or early OA vs. advanced OA cases, while neglecting how pure maturation could change the 18 SCB’s microstructure. To assess the transformations of the healthy SCB from young age to early 19 adulthood, we examined the microstructure and material composition of the medial condyle of the 20 femur in calves (3 months) and cattle (18 months) for the calcified cartilage (CC) and the 21 subchondral bone plate (SCBP). The entire subchondral zone (SCZ) was significantly thicker in 22 cattle compared to calves, although the proportion of the CC and SCBP thicknesses were relatively 23 constant. The trabecular number (Tb.N.) and the connectivity density (Conn.D) was significantly 24 higher in the deeper region of the SCZ, while the bone volume fraction (BV/TV), and the degree of 25 anisotropy (DA) were more affected by age rather than the region. The mineralization increased 26 within the first 250 µm of the SCZ irrespective of sample type, and became stable thereafter. Cattle 27 exhibited higher mineralization than calves at all depths, with a mean Ca/P ratio of 1.59 and 1.64 for 28 calves and cattle, respectively. Collectively, these results indicate that the SCZ is highly dynamic at 29 early age, and CC is the most dynamic layer of the SCZ.

 

Keywords

Bone histomorphometry; Subchondral bone; Microcomputed tomography; 31 Osteoarthritis; Calcified cartilage

 

 


 

 

Int J Mol Sci. 2019 Feb; 20(3): 503. Published online 2019 Jan 24.

 

Sensory Neuropeptides and their Receptors Participate in Mechano-Regulation of Murine Macrophages

 

Dominique Muschter, Anna-Sophie Beiderbeck, Tanja Späth, Christian Kirschneck, Agnes Schröder and Susanne Grässel

(SP4)                     

 

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Abstract

This study aimed to analyze if the sensory neuropeptide SP (SP) and the neurokinin receptor 1 (NK1R) are involved in macrophage mechano-transduction, similar to chondrocytes, and if alpha-calcitonin gene-related peptide (αCGRP) and the CGRP receptor (CRLR/Ramp1) show comparable activity. Murine RAW264.7 macrophages were subjected to a cyclic stretch for 1–3 days and 4 h/day. Loading and neuropeptide effects were analyzed for gene and protein expression of neuropeptides and their receptors, adhesion, apoptosis, proliferation and ROS activity. Murine bone marrow-derived macrophages (BMM) were isolated after surgical osteoarthritis (OA) induction and proliferation, apoptosis and osteoclastogenesis were analyzed in response to loading. Loading induced NK1R and CRLR/Ramp1 gene expression and altered protein expression in RAW264.7 macrophages. SP protein and mRNA level decreased after loading whereas αCGRP mRNA expression was stabilized. SP reduced adhesion in loaded RAW264.7 macrophages and both neuropeptides initially increased the ROS activity followed by a time-dependent suppression. OA induction sensitized BMM to caspase 3/7 mediated apoptosis after loading. Both sensory neuropeptides, SP and αCGRP, and their receptors are involved in murine macrophage mechano-transduction affecting neuropeptide impact on adhesion and ROS activity. OA induction altered BMM apoptosis in response to loading indicate that OA-associated biomechanical alterations might affect the macrophage population.

 

Keywords

cyclic stretch, mechanoregulation, murine macrophages, substance P, alpha-calcitonin gene-related peptide, neurokinin receptor, CGRP receptor, destabilized medial meniscus

 

 

 

 


 

 

Ann Stem Cell Res 2(1): 001-005. 2019 Feb 09.

 

Integrin α10β1-selected Equine MSCs have Improved Chondrogenic Differentiation, Immunomodulatory and Cartilage Adhesion Capacity

 

Kristina Uvebrant, Linda Reimer Rasmusson, Jan F. Talts, Paolo Alberton, Attila Aszodi and Evy Lundgren-Äkerlund

(SP1)                     

 

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Abstract

Cell therapy based on multipotent, adult mesenchymal stem cells (MSCs) is a promising method for the regeneration of cartilage tissue and treatment of osteoarthritis in both humans and animals. For safe and effective use of MSCs as therapeutic agents, quality control of the isolation and expansion methods as well as the final MSC product is of key importance. The aim of this study was to evaluate integrin α10β1 as a novel MSC biomarker to attest identity, purity and potency of MSC preparations. We found that MSCs, isolated from equine adipose tissue (AT) and selected using an antibody directed to integrin α10, expressed the stem cell markers CD44, CD90 and CD105, differentiated to chondrocytes, osteocytes and adipocytes and demonstrated immunomodulatory capacity as judged by suppression of T-cell proliferation and expression of PGE2 . In addition, we found that integrin α10-selected equine AT-MSCs showed higher chondrogenic differentiation capacity, in pellet mass cultures, compared to unselected MSCs. Moreover, the integrin α10-selected fraction of MSCs showed significantly higher adhesion capacity to cartilage defects in ex vivo osteochondral explants compared to unselected MSCs demonstrating a higher ability of the α10-enriched MSCs to home to an osteochondral damage. Taken together, the results show that integrin α10β1 can identify a homogenous population of multipotent equine AT-MSCs with a high chondrogenic differentiation and immunomodulatory potency as well as homing capacity. Thus, we conclude that integrin α10β1 is a biomarker that can ensure quality, potency and consistency of MSC preparations for cartilage repair.

 

Keywords

Integrin α10β1, MSCs, Selection, Chondrogenesis, Immunomodulation

 

 


 

 

Cell Signal. 2019 Jan;53:212-223. doi: 10.1016/j.cellsig.2018.10.005. Epub 2018 Oct 9.

 

Interleukin-1β signaling in osteoarthritis - chondrocytes in focus.

 

Jenei-Lanzl Zsuzsa, Meurer Andrea, Zaucke Frank

(SP8)                     

 

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Abstract

Osteoarthritis (OA) can be regarded as a chronic, painful and degenerative disease that affects all tissues of a joint and one of the major endpoints being loss of articular cartilage. In most cases, OA is associated with a variable degree of synovial inflammation. A variety of different cell types including chondrocytes, synovial fibroblasts, adipocytes, osteoblasts and osteoclasts as well as stem and immune cells are involved in catabolic and inflammatory processes but also in attempts to counteract the cartilage loss. At the molecular level, these changes are regulated by a complex network of proteolytic enzymes, chemokines and cytokines (for review: [1]). Here, interleukin-1 signaling (IL-1) plays a central role and its effects on the different cell types involved in OA are discussed in this review with a special focus on the chondrocyte.  

 

 

 


 

 

Int J Mol Sci. 2019 Jan 23;20(3). pii: E484. doi: 10.3390/ijms20030484.

 

The Importance of Physioxia in Mesenchymal Stem Cell Chondrogenesis and the Mechanisms Controlling Its Response.

 

Pattappa Girish, Johnstone Brian, Zellner Johannes, Docheva Denitsa, Angele Peter.

(SP7)                     

 

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Abstract

Articular cartilage covers the surface of synovial joints and enables joint movement. However, it is susceptible to progressive degeneration with age that can be accelerated by either previous joint injury or meniscectomy. This degenerative disease is known as osteoarthritis (OA) and it greatly affects the adult population. Cell-based tissue engineering provides a possible solution for treating OA at its earliest stages, particularly focal cartilage lesions. A candidate cell type for treating these focal defects are Mesenchymal Stem Cells (MSCs). However, present methods for differentiating these cells towards the chondrogenic lineage lead to hypertrophic chondrocytes and bone formation in vivo. Environmental stimuli that can stabilise the articular chondrocyte phenotype without compromising tissue formation have been extensively investigated. One factor that has generated intensive investigation in MSC chondrogenesis is low oxygen tension or physioxia (2⁻5% oxygen). In vivo articular cartilage resides at oxygen tensions between 1⁻4%, and in vitro results suggest that these conditions are beneficial for MSC expansion and chondrogenesis, particularly in suppressing the cartilage hypertrophy. This review will summarise the current literature regarding the effects of physioxia on MSC chondrogenesis with an emphasis on the pathways that control tissue formation and cartilage hypertrophy.

 

Keywords

chondrogenesis; early osteoarthritis; hypertrophy; hypoxia; hypoxia inducible factors; mesenchymal stem cells

 

 

 

 


Publications in 2018


 

 

Sci Rep. 2018 Jun 25;8(1):9645. doi: 10.1038/s41598-018-27927-8.

 

TNF inhibits catecholamine production from induced sympathetic neuron-like cells in rheumatoid arthritis and osteoarthritis in vitro

 

Markus Herrmann, Sven Anders, Rainer H. Straub & Zsuzsa Jenei-Lanzl

(SP8)                     

 

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Abstract

Synovial adipose stem cells (sASC) can be differentiated into catecholamine-expressing sympathetic neuron-like cells to treat experimental arthritis. However, the pro-inflammatory tumor necrosis factor (TNF) is known to be toxic to catecholaminergic cells (see Parkinson disease), and this may prevent anti-inflammatory effects in inflamed tissue. We hypothesized that TNF exhibits inhibitory effects on human differentiated sympathetic tyrosine hydroxylase-positive (TH+) neuron-like cells. For the first time, iTH+ neuron-like sympathetic cells were generated from sACSs of rheumatoid arthritis (RA) and osteoarthritis (OA) synovial tissue. Compared to untreated controls in both OA and RA, TNF-treated iTH+ cells demonstrated a weaker staining of catecholaminergic markers in cell cultures of RA/OA patients, and the amount of produced noradrenaline was markedly lower. These effects were reversed by etanercept. Exposure of iTH+ cells to synovial fluid of RA patients showed similar inhibitory effects. In mixed synovial cells, significant effects of TNF on catecholamine release were observed only in OA. This study shows that TNF inhibits iTH+ synovial cells leading to the decrease of secreted noradrenaline. This might be a reason why discovered newly appearing TH+ cells in the synovium are not able to develop their possible full anti-inflammatory role in arthritis.  

 

Keywords

Bone histomorphometry; Subchondral bone; Microcomputed tomography; 31 Osteoarthritis; Calcified cartilage 

 

 

 


 

 

Biochim Biophys Acta Mol Basis Dis. 2018 Mar;1864(3):851-859. doi: 10.1016/j.bbadis.2017.12.024. Epub 2017 Dec 19.

 

Extracellular matrix content and WNT/β-catenin levels of cartilage determine the chondrocyte response to compressive load.

 

Praxenthaler Heiko, Krämer Elisabeth, Weisser Melanie, Hecht Nicole, Fischer Jennifer, Grossner Tobias, Richter Wiltrud.

(SP3)

 

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Abstract

During osteoarthritis (OA)-development extracellular matrix (ECM) molecules are lost from cartilage, thus changing gene-expressionmatrix synthesis and biomechanical competence of the tissue. Mechanical loading is important for the maintenance of articular cartilage; however, the influence of an altered ECM content on the response of chondrocytes to loading is not well understood, but may provide important insights into underlying mechanisms as well as supplying new therapies for OA. Objective here was to explore whether a changing ECM-content of engineered cartilage affects major signaling pathways and how this alters the chondrocyte response to compressive loading. Activity of canonical WNT-, BMP-, TGF-β- and p38-signaling was determined during maturation of human engineered cartilage and followed after exposure to a single dynamic compression-episode. WNT/β-catenin- and pSmad1/5/9-levels declined with increasing ECM-content of cartilage. While loading significantly suppressed proteoglycan-synthesis and ACAN-expression at low ECM-content this catabolic response then shifted to an anabolic reaction at high ECM-content. A positive correlation was observed between GAG-content and load-induced alteration of proteoglycan-synthesis. Induction of high β-catenin levels by the WNT-agonist CHIR suppressed load-induced SOX9- and GAG-stimulation in mature constructs. In contrast, the WNT-antagonist IWP-2 was capable of attenuating load-induced GAG-suppression in immature constructs. In conclusion, either ECM accumulation-associated or pharmacologically induced silencing of WNT-levels allowed for a more anabolic reaction of chondrocytes to physiological loading. This is consistent with the role of proteoglycans in sequestering WNT-ligands in the ECM, thus reducing WNT-activity and also provides a novel explanation of why low WNT-activity in cartilage protects from OA-development in mechanically overstressed cartilage.

Copyright © 2017 Elsevier B.V. All rights reserved.

 

KEYWORDS:

Cell signaling; Chondrocyte; Mechanical loading; Osteoarthritis; SOX9

 

 

 


 

 

Int J Mol Sci. 2018 Jan 26; 19 (2). pii: E367. doi: 10.3390/ijms19020367.

 

Do Neuroendocrine Peptides and Their Receptors Qualify as Novel Therapeutic Targets in Osteoarthritis?

 

Grässel Susanne, Muschter Dominique.

(SP4)

                 

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Abstract

Joint tissues like synovium, articular cartilage, meniscus and subchondral bone, are targets for neuropeptides. Resident cells of these tissues express receptors for various neuroendocrine-derived peptides including proopiomelanocortin (POMC)-derived peptides, i.e., α-melanocyte-stimulating hormone (α-MSH), adrenocorticotropin (ACTH) and β-endorphin (β-ED), and sympathetic neuropeptides like vasoactive intestinal peptide (VIP) and neuropeptide y (NPY). Melanocortins attained particular attention due to their immunomodulatory and anti-inflammatory effects in several tissues and organs. In particular, α-MSH, ACTH and specific melanocortin-receptor (MCR) agonists appear to have promising anti-inflammatory actions demonstrated in animal models of experimentally induced arthritis and osteoarthritis (OA). Sympathetic neuropeptides have obtained increasing attention as they have crucial trophic effects that are critical for joint tissue and bone homeostasis. VIP and NPY are implicated in direct and indirect activation of several anabolic signaling pathways in bone and synovial cells. Additionally, pituitary adenylate cyclase-activating polypeptide (PACAP) proved to be chondroprotective and, thus, might be a novel target in OA. Taken together, it appears more and more likely that the anabolic effects of these neuroendocrine peptides or their respective receptor agonists/antagonists may be exploited for the treatment of patients with inflammatory and degenerative joint diseases in the future.

 

KEYWORDS

NPY; PACAP; VIP; alpha-MSH; neuroendocrine; osteoarthritis; proopiomelanocortin

 

 

 


 

 

J Cell Physiol. 2018 Jan;233(1):699-711. doi: 10.1002/jcp.25933. Epub 2017 May 19.

 

Global chondrocyte gene expression after a single anabolic loading period: Time evolution and re-inducibility of mechano-responses.

 

Scholtes Simone, Krämer Elisabeth, Weisser Melanie, Roth Wolfgang, Luginbühl Reto, Grossner Tobias, Richter Wiltrud. 

(SP3)

 

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Abstract

Aim of this study was a genome-wide identification of mechano-regulated genes and candidate pathways in human chondrocytes subjected to a single anabolic loading episode and characterization of time evolution and re-inducibility of the response. Osteochondral constructs consisting of chondrocyte-seeded collagen-scaffold connected to β-tricalcium-phosphate were pre-cultured for 35 days and subjected to dynamic compression (25% strain, 1 Hz, 9 × 10 min over 3 hr) before microarray-profiling was performed. Proteoglycan synthesis was determined by 35 S-sulfate-incorporation over 24 hr. Cell viability and hardness of constructs were unaltered by dynamic compression while proteoglycan synthesis was significantly stimulated (1.45-fold, p = 0.016). Among 115 significantly regulated genes, 114 were up-regulated, 48 of them twofold. AP-1-relevant transcription factors FOSB and FOS strongly increased in line with elevated ERK1/2-phosphorylation and rising MAP3K4 expression. Expression of proteoglycan-synthesizing enzymes CHSY1 and GALNT4 was load-responsive as were factors associated with the MAPK-, TGF-β-, calcium-, retinoic-acid-, Wnt-, and Notch-signaling pathway which were significantly upregulated SOX9, and BMP6 levels rose significantly also after multiple loading episodes at daily intervals even at the 14th cycle with no indication for desensitation. Canonical pSmad2/3 and pSmad1/5/9-signaling showed no consistent regulation. This study associates novel genes with mechanoregulation in chondrocytes, raising SOX9 protein levels with anabolic loading and suggests that more pathways than so far anticipated apparently work together in a complex network of stimulators and feedback-regulators. Upregulation of mechanosensitive indicators extending differentially into the resting time provides crucial knowledge to maximize cartilage matrix deposition for the generation of high-level cartilage replacement tissue.

© 2017 Wiley Periodicals, Inc.

 

KEYWORDS:

BMP signalling; MAPK; SOX9; TGF-β signaling; proteoglycan synthesis

 

 

 


Publications in 2017


 

 

J Biosci Bioeng. 2017 Apr;123(4):512-522. doi: 10.1016/j.jbiosc.2016.11.002. Epub 2016 Nov 28.

 

Cell compaction influences the regenerative potential of passaged bovine articular chondrocytes in an ex vivo cartilage defect model.

 

Schmutzer Michael, Aszodi Attila.

(SP1)                     

 

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Abstract

The loss and degradation of articular cartilage tissue matrix play central roles in the process of osteoarthritis (OA). New models for evaluating cartilage repair/regeneration are thus of great value for transferring various culture systems into clinically relevant situations. The repair process can be better monitored in ex vivo systems than in in vitro cell cultures. I have therefore established an ex vivo defect model prepared from bovine femoral condyles for evaluating cartilage repair by the implantation of cells cultured in various ways, e.g., monolayer-cultured cells or suspension or pellet cultures of articular bovine chondrocytes representing different cell compactions with variable densities of chondrocytes. I report that the integrin subunit α10 was significantly upregulated in suspension-cultured bovine chondrocytes at passage P2 compared with monolayer-cultured cells at P1 (p = 0.0083) and P2 (p < 0.05). Suspension-cultured cells did not promote cartilage repair when compared with implanted monolayer-cultured chondrocytes and pellets: 24.0 ± 0.66% for suspension cells, 46.4 ± 2.9% for monolayer cells, and 127.64 ± 0.90% for pellets (p < 0.0001) of the original defect volume (percentage of defect). Additional cultivation with chondrogenesis-promoting growth factors TGF-β1 and BMP-2 revealed an enhancing effect on cartilage repair in all settings. The advantage and innovation of this system over in vitro differentiation (e.g., micromass, pellet) assays is the possibility of examining and evaluating cartilage regeneration in an environment in which implanted cells are embedded within native surrounding tissue at the defect site. Such ex vivo explants might serve as a better model system to mimic clinical situations.

 

Keywords

BMP2; Cartilage repair; Cell compaction; Chondrocytes; Ex vivo defect model; Growth factors; Passaging; Pellets; Suspension culture; TGFbeta1 

 

 


 

 

Int J Mol Sci. 2017 Apr 28; 18 (5). pii: E931. doi: 10.3390/ijms18050931

 

Peripheral Nerve Fibers and Their Neurotransmitters in Osteoarthritis Pathology.

 

Grässel Susanne, Muschter Dominique.

(SP4)

 

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Abstract

The importance of the nociceptive nervous system for maintaining tissue homeostasis has been known for some time, and it has also been suggested that organogenesis and tissue repair are under neuronal control. Changes in peripheral joint innervation are supposed to be partly responsible for degenerative alterations in joint tissues which contribute to development of osteoarthritis. Various resident cell types of the musculoskeletal system express receptors for sensory and sympathetic neurotransmitters, allowing response to peripheral neuronal stimuli. Among them are mesenchymal stem cells, synovial fibroblasts, bone cells and chondrocytes of different origin, which express distinct subtypes of adrenoceptors (AR), receptors for vasoactive intestinal peptide (VIP), substance P (SP) and calcitonin gene-related peptide (CGRP). Some of these cell types synthesize and secrete neuropeptides such as SP, and they are positive for tyrosine-hydroxylase (TH), the rate limiting enzyme for biosynthesis of catecholamines. Sensory and sympathetic neurotransmitters are involved in the pathology of inflammatory diseases such as rheumatoid arthritis (RA) which manifests mainly in the joints. In addition, they seem to play a role in pathogenesis of priori degenerative joint disorders such as osteoarthritis (OA). Altogether it is evident that sensory and sympathetic neurotransmitters have crucial trophic effects which are critical for joint tissue and bone homeostasis. They modulate articular cartilage, subchondral bone and synovial tissue properties in physiological and pathophysiological conditions, in addition to their classical neurological features.

 

KEYWORDS:

CGRP; adrenoceptors; cartilage; neurotransmitters; osteoarthritis; peripheral nervous system; subchondral bone; substance P

 

 

 

 


 

 

Cartilage Vol. 2  Chapter 10 Page 191-227 Springer February 2017 

 

The Sensory and Sympathetic Nervous System in Cartilage Physiology and Pathophysiology

 

Grässel Susanne, Straub Rainer, Jenei-Lanzl Zsuzsa

(SP8)               

 

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ISBN 978-3-319-45801-4

DOI   10.1007/978-3-319-45803-8_9

 

Abstract

The peripheral nervous system is critically involved in metabolism of joint tissue and intervertebral disks (IVD). Nerve fibers of sympathetic and sensory origin innervate synovial tissue and subchondral bone of diarthrodial joints. In pathophysiological situations as in osteoarthritis (OA), rheumatoid arthritis (RA), and IVD degeneration, innervation patterns of sympathetic and sensory nerve fibers are partly altered in joint tissue and IVD.

Various resident cell types of the musculoskeletal system express receptors for sensory and sympathetic neurotransmitters allowing response to neuronal stimuli. Among them are mesenchymal stem cells, synovial fibroblasts, bone cells, and different types of chondrocytes, which express distinct subtypes of adrenoceptors, receptors for vasoactive intestinal peptide (VIP), for substance P (SP), and calcitonin gene-related peptide (CGRP). Some of these cell types even synthesize neuropeptides such as SP, and they are positive for tyrosine hydroxylase (TH), the rate limiting enzyme for biosynthesis of catecholamines. During endochondral ossification in embryonic limb development, sensory and sympathetic neurotransmitters modulate osteo-chondrogenic differentiation of mesenchymal progenitor cells, vascularization, and matrix differentiation indicating a distinct role in skeletal growth and possible limb regeneration processes. In adults, sensory and sympathetic neurotransmitters are involved in pathology of inflammatory diseases as rheumatoid arthritis which manifests mainly in joints. In addition, they might play a role in pathogenesis of a priori degenerative joint disorders, as osteoarthritis and intervertebral disk degeneration.

Altogether it became evident that sensory and sympathetic neurotransmitters have crucial trophic effects which are critical for proper limb formation during embryonic skeletal growth. In adults, they modulate articular cartilage, subchondral bone and synovial tissue homeostasis, and physiological and pathophysiological conditions, in addition to their classical neurological features.

 

Keywords

Nerve Fiber Nerve Growth Factor Articular Cartilage Vasoactive Intestinal Peptide Annulus Fibrosus