Crucial role of the terminal complement complex in chondrocyte death and hypertrophy after cartilage trauma.

OBJECTIVE: Innate immune response and particularly terminal complement complex (TCC) deposition are thought to be involved in the pathogenesis of posttraumatic osteoarthritis. However, the possible role of TCC in regulated cell death as well as chondrocyte hypertrophy and senescence has not been unraveled so far and was first addressed using an ex vivo human cartilage trauma-model. DESIGN: Cartilage explants were subjected to blunt impact (0.59 J) and exposed to human serum (HS) and cartilage homogenate (HG) with or without different potential therapeutics: RIPK1-inhibitor Necrostatin-1 (Nec), caspase-inhibitor zVAD, antioxidant N-acetyl cysteine (NAC) and TCC-inhibitors aurintricarboxylic acid (ATA) and clusterin (CLU). Cell death and hypertrophy/senescence-associated markers were evaluated on mRNA and protein level. RESULTS: Addition of HS resulted in significantly enhanced TCC deposition on chondrocytes and decrease of cell viability after trauma. This effect was potentiated by HG and was associated with expression of RIPK3, MLKL and CASP8. Cytotoxicity of HS could be prevented by heat-inactivation or specific inhibitors, whereby combination of Nec and zVAD as well as ATA exhibited highest cell protection. Moreover, HS+HG exposition enhanced the gene expression of CXCL1, IL-8, RUNX2 and VEGFA as well as secretion of IL-6 after cartilage trauma. CONCLUSIONS: Our findings imply crucial involvement of the complement system and primarily TCC in regulated cell death and phenotypic changes of chondrocytes after cartilage trauma. Inhibition of TCC formation or downstream signaling largely modified serum-induced pathophysiologic effects and might therefore represent a therapeutic target to maintain the survival and chondrogenic character of cartilage cells.

The functional role of chondrogenic stem/progenitor cells: novel evidence for immunomodulatory properties and regenerative potential after cartilage injury.

Considering the poor intrinsic healing potential of articular cartilage, resident chondrogenic stem/progenitor cells (CSPCs) have gained attention in recent years. Although, CSPCs are attracted by a cartilage injury, knowledge about the post-traumatic behaviour and functional role of this cell population is fairly basic. The present study, not only elaborated on the regenerative capacities of CSPCs, but also illuminated potential immunomodulatory properties after cartilage trauma. Estimation of the CSPC population size within previously impacted cartilage explants by flow-cytometry revealed an increased percentage of CSPC-marker positive cells as compared to unimpacted tissue. In line with this, proliferation, chemotactic migration and in vitro wound healing activity of isolated CSPCs was similarly enhanced after stimulation with trauma-conditioned (TC) medium. Further, a significant increase in pro- and anti-inflammatory gene expression, as well as IL-6 secretion due to TC-medium-stimulation was measured. In this context, antioxidative or chondroanabolic therapeutic intervention alleviated the post-traumatic response of TC-medium-activated CSPCs and substantially influenced CSPC chondrogenic differentiation in different ways. Overall, this study provided novel insights concerning the functional role of CSPCs after cartilage trauma and the effects of a therapeutic intervention in order to improve regenerative processes and prevent cartilage degeneration following trauma.

Antioxidative therapy in an ex vivo human cartilage trauma-model: attenuation of trauma-induced cell loss and ECM-destructive enzymes by N-acetyl cysteine.

OBJECTIVE: Mechanical trauma of articular cartilage results in cell loss and cytokine-driven inflammatory response. Subsequent accumulation of reactive oxygen (ROS) and nitrogen (RNS) species enhances the enzymatic degradation of the extracellular matrix (ECM). This study aims on the therapeutic potential of N-acetyl cysteine (NAC) in a human ex vivo cartilage trauma-model, focusing on cell- and chondroprotective features. DESIGN: Human full-thickness cartilage explants were subjected to a defined impact trauma (0.59 J) and treated with NAC. Efficiency of NAC administration was evaluated by following outcome parameters: cell viability, apoptosis rate, anabolic/catabolic gene expression, secretion and activity of matrix metalloproteinases (MMPs) and proteoglycan (PG) release. RESULTS: Continuous NAC administration increased cell viability and reduced the apoptosis rate after trauma. It also suppressed trauma-induced gene expression of ECM-destructive enzymes, such as ADAMTS-4, MMP-1, -2, -3 and -13 in a dosage- and time-depending manner. Subsequent suppression of MMP-2 and MMP-13 secretion reflected these findings on protein level. Moreover, NAC inhibited proteolytic activity of MMPs and reduced PG release. CONCLUSION: In the context of this ex vivo study, we showed not only remarkable cell- and chondroprotective features, but also revealed new encouraging findings concerning the therapeutically effective concentration and treatment-time regimen of NAC. Its defense against chondrocyte apoptosis and catabolic enzyme secretion recommends NAC as a multifunctional add-on reagent for pharmaceutical intervention after cartilage injury. Taken together, our data increase the knowledge on the therapeutic potential of NAC after cartilage trauma and presents a basis for future in vivo studies.

The Impact of Histological Clot Composition in Embolic Stroke.

PURPOSE: Thrombus composition has been suggested to have a decisive impact on the outcome of patients treated by mechanical thrombectomy because of embolic stroke. The recent development of stent retrievers allows collection and, hence, histopathological analysis of fresh thrombus material. Against this background, the aim of this prospective study was to assess the impact of thrombus composition on mechanical recanalization, clinical outcome and stroke etiology. METHODS: Thirty-four patients suffering from acute ischemic stroke due to occlusion of the distal internal carotid artery/carotid-T, anterior cerebral artery, or middle cerebral arteries were mechanically recanalized, and thrombus material was obtained. Histological thrombus composition was compared with imaging, clinical, and neurointerventional data. RESULTS: The main findings were that a higher percentage of white blood cells (WBCs) in the thrombus was associated with (i) cardioembolic etiology, (ii) extended mechanical recanalization time, and (iii) less favorable recanalization (Thrombolysis in Cerebral Infarction score) and clinical outcome (National Institute of Health Stroke Scale). CONCLUSION: Our results suggest that thrombi with a high WBC fraction are related to more organized thrombi of cardioembolic origin associated with less favorable recanalization and clinical outcome in acute ischemic anterior circulation stroke. WBC-mediated immunological and coagulatory processes may play a key role in thrombus formation and pathogenesis of stroke warranting further investigation.