Dendritic Polyglycerol Sulfate Hydrogels as a Potential Viscosupplement for Treatment of Osteoarthritis

The aim of this thesis was to investigate the potential of a fully synthetic, slowly degradable, heparin sulfate mimetic hydrogel as an alternative viscosupplement for OA management and thereby compare it to the current standard viscosupplement HA. A previous study revealed a short half-life of HA, ranging from half a day up to 9 days in vivo.[65] To avoid several injections, which may incur higher costs and infection risks, a fully synthetic dendritic polyglycerol sulfate (dPGS) hydrogel was evaluated for its bioorthogonality. The rheological properties of this slow-degradable hydrogel were then investigated to determine a suitable concentration for intra-articular injections that mimicked HA in terms of its viscoelastic and mechanical properties. Therefore, different concentrations of dPGS ranging from 3.6 to 4.8 wt% were investigated by means of oscillating and flow rheology, thereby yielding storage (G') and loss modulus (G''), as well as yield stress and shear viscosity. Additionally, blends of commercially available HAs, which varied in respect to their molecular weight, were used as references. As a result, a pronounced coupling of the molecular weight and the rheological properties for the HAs was observed. The zero shear viscosity of the studied HAs ranged between 5 and 1600 Pa⋅s, depending strongly on the molecular weight. Besides, all four HA samples exhibited pronounced shear thinning behavior. Furthermore, the dPGS hydrogel formed more compact networks with increasing concentrations. From a broader comparison, the current findings suggest that an overall polymer concentration of 4.0 wt% dPGS has viscoelastic properties that are comparable to HA in the medically relevant frequency range. The third part of the thesis was focused on the evaluation of dPGS effects on normal and OA-like tissue-engineered cartilage. To overcome the low availability of human primary tissue and high costs of animal models an established in vitro OA model has been used. It is based on porcine cartilage sources and offers a high-throughput analysis of potential active substances in a reproducible and very well characterized approach under standardized conditions.[140, 144] In this model, micromass cultures were treated with 2.5 wt% dPGS hydrogel for 7 days under normal and OA conditions (treated with TNF-α). Live/dead staining of micromasses revealed a majority of viable cells embedded in ECM after 7 days of treatment with the hydrogel in normal and OA conditions. This confirmed previous findings and suggested that dPGS was not harmful for different cell types and even in vivo.[145] Safranin-O staining demonstrated a typical depletion of GAGs in OA-like micromasses but not in the presence of the dPGS hydrogel. There was no distinct difference in immunolabeling for type II collagen. The microarray data showed that rheumatoid arthritis and TNF signaling pathways were downregulated in hydrogel-treated OA-like micromasses in comparison to non-treated OA-like micromasses. Furthermore, the dPGS hydrogel alone did not affect genes related to OA such as ANPEP, COMP, CXCL12, COX2, and TNFSF10, but it could prevent their regulation caused by TNF-α. These findings proved the potential of this hydrogel to prevent the development of TNF-α-induced OA with regard to PG loss and TNF-α-induced expression pattern without additional signs of differentiation and inflammation. In the fourth part of this work, the HA-related modifications were investigated on cellular and molecular level in the same in vitro system to serve as a control for comparisons with the dPGS hydrogel. The data showed no inhibiting or activating effect of HA on normal or OA-like tissue-engineered cartilage on cellular level. Microarray data demonstrated a minor impact of HA on gene expression level. The upregulation of VEGFA and ANKRD37 genes confirmed the chondroprotective potential of HA. It could regulate the cartilage anabolism by stabilizing the chondrocyte phenotype in pathological conditions. In conclusion, the evaluation of the dPGS hydrogel showed that it is a potential alternative for HA as an intra-articular injectable lubricant for osteoarthritis. Moreover, in contrast to HA, dPGS can prevent the development of TNF-α-induced OA with regard to proteoglycan loss and TNF-α-induced expression pattern. Although interactions of dPGS-hydrogels with biological systems have been elucidated to a certain extent, still a lot of open questions remain, especially concerning the in vivo effect on synovial joints. To follow up these promising results, further investigation needs to be performed in animal models. In particular, the localization of this hydrogel in the synovial joint should be further investigated by fluorescent dye conjugation and its anti-inflammatory properties by measuring the related cytokine ratios in the synovial fluid. Since it is known that hydrogels can be used as a delivery system, this hydrogel can also be further optimized with biologics to trigger in situ regeneration

Hyaluronic Acid Influence on Normal and Osteoarthritic Tissue-Engineered Cartilage

The aim of this study is to identify gene expression profiles associated with hyaluronic acid (HA) treatment of normal and osteoarthritis (OA)-like tissue- engineered cartilage. 3D cartilage micromasses were treated with tumour necrosis factor-α (TNF-α) (OA-inducer) and/or HA for 7 days. Viability was examined by PI/FDA staining. To document extracellular matrix (ECM) formation, glycosaminoglycans (GAG) were stained with Safranin-O and cartilage-specific type II collagen was detected immunohistochemically. Genome-wide gene expression was determined using microarray analysis. Normal and OA-like micromasses remained vital and showed a spherical morphology and homogenous cell distribution regardless of the treatment. There was no distinct difference in immunolabeling for type II collagen. Safranin-O staining demonstrated a typical depletion of GAG in TNF-α-treated micromasses (−73%), although the extent was limited in the presence of HA (−39%). The microarray data showed that HA can influence the cartilage metabolism via upregulation of TIMP3 in OA-like condition. The upregulation of VEGFA and ANKRD37 genes implies a supportive role of HA in cartilage maturation and survival. The results of this study validate the feasibility of the in vitro OA model for the investigation of HA. On the cellular level, no inhibiting or activating effect of HA was shown. Microarray data demonstrated a minor impact of HA on gene expression level

Effects of consecutive supervised core stability training on pain and disability in women with nonspecific chronic low back pain

Introduction: Low back pain is a very common health care problem affecting 85% of population. Spine rehabilitation programs are very various in subjects with chronic low back pain but core stability training is often the chosen treatment. However, the aim of this study was to examine the effects of consecutive supervised core stability training on pain intensity and disability of nonspecific chronic low back pain. Materials and Methods: Twenty-four non-athletic females with chronic low back pain were participated in the study. They were randomly divided into two groups: experimental group (12 days consecutively under physical therapist supervision) and control group (no intervention). The subjects in both groups were not allowed to perform any sport activity during 12 days of study. Before and after the intervention, visual analogue scale and the ososetry disability index were used to assess pain intensity and disability, respectively. Results: The findings show that the disability and intensity pain variables were significantly decreased in the experimental group (P=0.0001 and P=0.003, respectively), while no significant changes were found with respect to those variables in the control group. Conclusions: Consecutive supervised core stability training may decrease the pain intensity and disability of the patients with chronic low back pain

Chia seeds as a potential cognitive booster in the APP23 Alzheimer’s disease model

Glucose hypometabolism potentially contributes to Alzheimer’s disease (AD) and might even represent an underlying mechanism. Here, we investigate the relationship of diet-induced metabolic stress and AD as well as the therapeutic potential of chia seeds as a modulator of glucose metabolism in the APP23 mouse model. 4–6 (pre-plaque stage, PRE) and 28–32 (advanced-plaque stage, ADV) weeks old APP23 and wild type mice received pretreatment for 12 weeks with either sucrose-rich (SRD) or control diet, followed by 8 weeks of chia seed supplementation. Although ADV APP23 mice generally showed functioning glucose homeostasis, they were more prone to SRD-induced glucose intolerance. This was accompanied by elevated corticosterone levels and mild insulin insensitivity. Chia seeds improved spatial learning deficits but not impaired cognitive flexibility, potentially mediated by amelioration of glucose tolerance, attenuation of corticosterone levels and reversal of SRD-induced elevation of pro-inflammatory cytokine levels. Since cognitive symptoms and plaque load were not aggravated by SRD-induced metabolic stress, despite enhanced neuroinflammation in the PRE group, we conclude that impairments of glucose metabolism do not represent an underlying mechanism of AD in this mouse model. Nevertheless, chia seeds might provide therapeutic potential in AD as shown by the amelioration of cognitive symptoms

A Novel Method Facilitating the Simple and Low-Cost Preparation of Human Osteochondral Slice Explants for Large-Scale Native Tissue Analysis

For in vitro modeling of human joints, osteochondral explants represent an acceptable compromise between conventional cell culture and animal models. However, the scarcity of native human joint tissue poses a challenge for experiments requiring high numbers of samples and makes the method rather unsuitable for toxicity analyses and dosing studies. To scale their application, we developed a novel method that allows the preparation of up to 100 explant cultures from a single human sample with a simple setup. Explants were cultured for 21 days, stimulated with TNF-α or TGF-β 3, and analyzed for cell viability, gene expression and histological changes. Tissue cell viability remained stable at >90% for three weeks. Proteoglycan levels and gene expression of COL2A1, ACAN and COMP were maintained for 14 days before decreasing. TNF-α and TGF-β 3 caused dose-dependent changes in cartilage marker gene expression as early as 7 days. Histologically, cultures under TNF-α stimulation showed a 32% reduction in proteoglycans, detachment of collagen fibers and cell swelling after 7 days. In conclusion, thin osteochondral slice cultures behaved analogously to conventional punch explants despite cell stress exerted during fabrication. In pharmacological testing, both the shorter diffusion distance and the lack of need for serum in the culture suggest a positive effect on sensitivity. The ease of fabrication and the scalability of the sample number make this manufacturing method a promising platform for large-scale preclinical testing in joint research

Dietary-challenged mice with Alzheimer-like pathology show increased energy expenditure and reduced adipocyte hypertrophy and steatosis

Alzheimer's disease (AD) is frequently accompanied by progressing weight loss, correlating with mortality. Counter-intuitively, weight loss in old age might predict AD onset but obesity in midlife increases AD risk. Furthermore, AD is associated with diabetes-like alterations in glucose metabolism. Here, we investigated metabolic features of amyloid precursor protein overexpressing APP23 female mice modeling AD upon longterm challenge with high-sucrose (HSD) or high-fat diet (HFD). Compared to wild type littermates (WT), APP23 females were less prone to mild HSD-induced and considerable HFD-induced glucose tolerance deterioration, despite unaltered glucose tolerance during normal-control diet. Indirect calorimetry revealed increased energy expenditure and hyperactivity in APP23 females. Dietary interventions, especially HFD, had weaker effects on lean and fat mass gain, steatosis and adipocyte hypertrophy of APP23 than WT mice, as shown by 1H-magnetic resonance-spectroscopy, histological and biochemical analyses. Proteome analysis revealed differentially regulated expression of mitochondrial proteins in APP23 livers and brains. In conclusion, hyperactivity, increased metabolic rate, and global mitochondrial dysfunction potentially add up to the development of AD related body weight changes in APP23 females, becoming especially evident during diet-induced metabolic challenge. These findings emphasize the importance of translating this metabolic phenotyping into human research to decode the metabolic component in AD pathogenesis