Exploring Responsiveness to Highly Challenging Balance and Gait Training in Parkinson's Disease.

BACKGROUND: Exercise potentially improves gait, balance, and habitual physical activity in Parkinson's disease (PD). However, given the heterogeneous nature of the disease, it is likely that people respond differently to exercise interventions. Factors determining responsiveness to exercise interventions remain unclear. OBJECTIVES: To address this uncertainty, we explored the responsiveness to our highly challenging balance and gait intervention (HiBalance) in people with PD. METHODS: Thirty-nine participants with mild-moderate PD who underwent the HiBalance intervention from our randomized controlled trial were included. We defined response in three domains: (1) balance based on Mini-BESTest, (2) gait based on gait velocity, and (3) physical activity based on accelerometry-derived steps per day. In each domain, we explored three responsiveness levels: high, low, or non-responders according to the change from pre- to post-intervention. Separate Random Forests for each responder domain classified these responsiveness levels and identified variable importance. RESULTS: Only the Random Forest for the balance domain classified all responsiveness levels above the chance level indicated by a Cohen's kappa of "slight" agreement. Variable importance differed among the responsiveness levels. Slow gait velocity indicated high responders in the balance domain but showed low probabilities for low and non-responders. For low and non-responders, fall history or no falls, respectively, were more important. CONCLUSIONS: Among three responder domains and responsiveness levels, we could moderately classify responders in the balance domain, but not for the gait or physical activity domain. This can guide inclusion criteria for balance-targeted, personalized intervention studies in people with PD.

MRI subtypes in Parkinson's disease across diverse populations and clustering approaches.

Parkinson's disease (PD) is clinically heterogeneous, which suggests the existence of subtypes; however, there has been no consensus regarding their characteristics. This study included 633 PD individuals across distinct cohorts: unmedicated de novo PD, medicated PD, mild-moderate PD, and a cohort based on diagnostic work-up in clinical practice. Additionally, 233 controls were included. Clustering based on cortical and subcortical gray matter measures was conducted with and without adjusting for global atrophy in the entire PD sample and validated within each cohort. Subtypes were characterized using baseline and longitudinal demographic and clinical data. Unadjusted results identified three clusters showing a gradient of neurodegeneration and symptom severity across the entire sample and the individual cohorts. When adjusting for global atrophy eight clusters were identified in the entire sample, lacking consistency in individual cohorts. This study identified atrophy-based subtypes in PD, emphasizing the significant impact of global atrophy on subtype number, patterns, and interpretation in cross-sectional analyses.

Structural parameters are superior to eigenvector centrality in detecting progressive supranuclear palsy with machine learning & multimodal MRI.

Progressive supranuclear palsy (PSP) is an atypical Parkinsonian syndrome characterized initially by falls and eye movement impairment. This multimodal imaging study aimed at eliciting structural and functional disease-specific brain alterations. T1-weighted and resting-state functional MRI were applied in multi-centric cohorts of PSP and matched healthy controls. Midbrain, cerebellum, and cerebellar peduncles showed severely low gray/white matter volume, whereas thinner cortical gray matter was observed in cingulate cortex, medial and temporal gyri, and insula. Eigenvector centrality analyses revealed regionally specific alterations. Multivariate pattern recognition classified patients correctly based on gray and white matter segmentations with up to 98 % accuracy. Highest accuracies were obtained when restricting feature selection to the midbrain. Eigenvector centrality indices yielded an accuracy around 70 % in this comparison; however, this result did not reach significance. In sum, the study reveals multimodal, widespread brain changes in addition to the well-known midbrain atrophy in PSP. Alterations in brain structure seem to be superior to eigenvector centrality parameters, in particular for prediction with machine learning approaches.

Validation of fNIRS measurement of executive demand during walking with and without dual-task in younger and older adults and people with Parkinson's disease.

BACKGROUND: Walking with a concurrent cognitive task (dual-task walking) can pose a challenge to some populations due to aging or neurodegenerative disease. These tasks require cognitive resources involving the prefrontal cortex and can be studied using functional near-infrared spectroscopy (fNIRS). An important step in understanding fNIRS measures during such walking tasks is validating that measures reflect the demands of the tasks and not confounding sources or movement artifacts. AIM: This study aimed to investigate the validity of fNIRS measures of prefrontal cortex activity as an indicator of executive demand during usual walking (single-task) and dual-task walking against clinical and objective measures of motor behavior in young adults, older adults, and people with Parkinson's disease (PD), by evaluating several validation hypotheses. METHODS: In total, 133 participants were recruited from younger adults (18-50 years, n = 42), older adults (≥60 years, n = 49) and people with PD (≥60 years, n = 42). Activity in the prefrontal cortex during walking with and without an auditory Stroop task was measured with fNIRS. A combined hemoglobin measure (correlation-based signal improvement, CBSI) was calculated for use in a region of interest analysis in the dorsolateral prefrontal cortex (dlPFC). Pre-registered hypotheses regarding convergent validity, discriminant validity and known group validity were tested. An exploratory analysis of different hemoglobin measures was also performed. RESULTS: Increases in dlPFC activity were found from single- to dual-task walking in the younger adults group and from rest to single-task walking in the older adults and PD groups. In line with hypotheses, a positive relationship was found between between dlPFC activity during dual-task walking and dual-task cost in the younger adults group, as well as a positive relationship to step time variability during single-task walking and a negative relationship to walking speed during single-task walking in the PD group. However, several clinical and gait measures lacked a relationship with dlPFC activity. CONCLUSION: The fNIRS results point towards the CBSI measure of dlPFC activity being a valid measure of executive demand during both single and dual-task walking. Some relationships between clinical and gait measures and brain activity during walking need further investigation.

Speech and neuroimaging effects following HiCommunication: a randomized controlled group intervention trial in Parkinson's disease.

Speech, voice and communication changes are common in Parkinson's disease. HiCommunication is a novel group intervention for speech and communication in Parkinson's disease based on principles driving neuroplasticity. In a randomized controlled trial, 95 participants with Parkinson's disease were allocated to HiCommunication or an active control intervention. Acoustic analysis was performed pre-, post- and six months after intervention. Intention-to-treat analyses with missing values imputed in linear multilevel models and complimentary per-protocol analyses were performed. The proportion of participants with a clinically relevant increase in the primary outcome measure of voice sound level was calculated. Resting-state functional MRI was performed pre- and post-intervention. Spectral dynamic causal modelling and the parametric empirical Bayes methods were applied to resting-state functional MRI data to describe effective connectivity changes in a speech-motor-related network of brain regions. From pre- to post-intervention, there were significant group-by-time interaction effects for the measures voice sound level in text reading (unstandardized b = 2.3, P = 0.003), voice sound level in monologue (unstandardized b = 2.1, P = 0.009), Acoustic Voice Quality Index (unstandardized b = -0.5, P = 0.016) and Harmonics-to-Noise Ratio (unstandardized b = 1.3, P = 0.014) post-intervention. For 59% of the participants, the increase in voice sound level after HiCommunication was clinically relevant. There were no sustained effects at the six-month follow-up. In the effective connectivity analysis, there was a significant decrease in inhibitory self-connectivity in the left supplementary motor area and increased connectivity from the right supplementary motor area to the left paracentral gyrus after HiCommunication compared to after the active control intervention. In conclusion, the HiCommunication intervention showed promising effects on voice sound level and voice quality in people with Parkinson's disease, motivating investigations of barriers and facilitators for implementation of the intervention in healthcare settings. Resting-state brain effective connectivity was altered following the intervention in areas implicated, possibly due to reorganization in brain networks.

Robot-based 6D bioprinting for soft tissue biomedical applications.

Within this interdisciplinary study, we demonstrate the applicability of a 6D printer for soft tissue engineering models. For this purpose, a special plant was constructed, combining the technical requirements for 6D printing with the biological necessities, especially for soft tissue. Therefore, a commercial 6D robot arm was combined with a sterilizable housing (including a high-efficiency particulate air (HEPA) filter and ultraviolet radiation (UVC) lamps) and a custom-made printhead and printbed. Both components allow cooling and heating, which is desirable for working with viable cells. In addition, a spraying unit was installed that allows the distribution of fine droplets of a liquid. Advanced geometries on uneven or angled surfaces can be created with the use of all six axes. Based on often used bioinks in the field of soft tissue engineering (gellan gum, collagen, and gelatin methacryloyl) with very different material properties, we could demonstrate the flexibility of the printing system. Furthermore, cell-containing constructs using primary human adipose-derived stem cells (ASCs) could be produced in an automated manner. In addition to cell survival, the ability to differentiate along the adipogenic lineage could also be demonstrated as a representative of soft tissue engineering.

Investigating underlying brain structures and influence of mild and subjective cognitive impairment on dual-task performance in people with Parkinson's disease.

Cognitive impairment can affect dual-task abilities in Parkinson's disease (PD), but it remains unclear whether this is also driven by gray matter alterations across different cognitive classifications. Therefore, we investigated associations between dual-task performance during gait and functional mobility and gray matter alterations and explored whether these associations differed according to the degree of cognitive impairment. Participants with PD were classified according to their cognitive function with 22 as mild cognitive impairment (PD-MCI), 14 as subjective cognitive impairment (PD-SCI), and 20 as normal cognition (PD-NC). Multiple regression models associated dual-task absolute and interference values of gait speed, step-time variability, and reaction time, as well as dual-task absolute and difference values for Timed Up and Go (TUG) with PD cognitive classification. We repeated these regressions including the nucleus basalis of Meynert, dorsolateral prefrontal cortex, and hippocampus. We additionally explored whole-brain regressions with dual-task measures to identify dual-task-related regions. There was a trend that cerebellar alterations were associated with worse TUG dual-task in PD-SCI, but also with higher dual-task gait speed and higher dual-task step-time variability in PD-NC. After multiple comparison corrections, no effects of interest were significant. In summary, no clear set of variables associated with dual-task performance was found that distinguished between PD cognitive classifications in our cohort. Promising but non-significant trends, in particular regarding the TUG dual-task, do however warrant further investigation in future large-scale studies.

Biofabrication's Contribution to the Evolution of Cultured Meat.

Cultured Meat (CM) is a growing field in cellular agriculture, driven by the environmental impact of conventional meat production, which contributes to climate change and occupies ≈70% of arable land. As demand for meat alternatives rises, research in this area expands. CM production relies on tissue engineering techniques, where a limited number of animal cells are cultured in vitro and processed to create meat-like tissue comprising muscle and adipose components. Currently, CM is primarily produced on a small scale in pilot facilities. Producing a large cell mass based on suitable cell sources and bioreactors remains challenging. Advanced manufacturing methods and innovative materials are required to subsequently process this cell mass into CM products on a large scale. Consequently, CM is closely linked with biofabrication, a suite of technologies for precisely arranging cellular aggregates and cell-material composites to construct specific structures, often using robotics. This review provides insights into contemporary biomedical biofabrication technologies, focusing on significant advancements in muscle and adipose tissue biofabrication for CM production. Novel materials for biofabricating CM are also discussed, emphasizing their edibility and incorporation of healthful components. Finally, initial studies on biofabricated CM are examined, addressing current limitations and future challenges for large-scale production.

The Human Affectome.

Over the last decades, theoretical perspectives in the interdisciplinary field of the affective sciences have proliferated rather than converged due to differing assumptions about what human affective phenomena are and how they work. These metaphysical and mechanistic assumptions, shaped by academic context and values, have dictated affective constructs and operationalizations. However, an assumption about the purpose of affective phenomena can guide us to a common set of metaphysical and mechanistic assumptions. In this capstone paper, we home in on a nested teleological principle for human affective phenomena in order to synthesize metaphysical and mechanistic assumptions. Under this framework, human affective phenomena can collectively be considered algorithms that either adjust based on the human comfort zone (affective concerns) or monitor those adaptive processes (affective features). This teleologically-grounded framework offers a principled agenda and launchpad for both organizing existing perspectives and generating new ones. Ultimately, we hope the Human Affectome brings us a step closer to not only an integrated understanding of human affective phenomena, but an integrated field for affective research.

Using functional near-infrared spectroscopy to measure prefrontal cortex activity during dual-task walking and navigated walking: A feasibility study.

INTRODUCTION: While functional near-infrared spectroscopy (fNIRS) can provide insight into motor-cognitive deficits during ecologically valid gait conditions, the feasibility of using fNIRS during complex walking remains unknown. We tested the process and scientific feasibility of using an fNIRS device to measure cortical activity during complex walking tasks consisting of straight walking and navigated walking under single and dual-task (DT) conditions. METHODS: Nineteen healthy people from 18 to 64 years (mean age: 45.7 years) participated in this study which consisted of three complex walking protocols: (i) straight walking, DT walking (walking while performing an auditory Stroop task) and single-task auditory Stroop, (ii) straight and navigated walking, and (iii) navigated walking and navigated DT walking. A rest condition (standing still) was also included in each protocol. Process feasibility outcomes included evaluation of the test procedures and participant experience during and after each protocol. Scientific feasibility outcomes included signal quality measures, and the ability to measure changes in concentration of deoxygenated and oxygenated hemoglobin in the prefrontal cortex. RESULTS: All participants were able to complete the three protocols with most agreeing that the equipment was comfortable (57.9%) and that the testing duration was adequate (73.7%). Most participants did not feel tired (94.7%) with some experiencing pain (42.1%) during the protocols. The signal qualities were high for each protocol. Compared to the rest condition, there was an increase in oxygenated hemoglobin in the prefrontal cortex when performing dual-task walking and navigation. CONCLUSION: We showed that our experimental setup was feasible for assessing activity in the prefrontal cortex with fNIRS during complex walking. The experimental setup was deemed acceptable and practicable. Signal quality was good during complex walking conditions and findings suggest that the different tasks elicit a differential brain activity, supporting scientific feasibility.

Implicit Motor Sequence Learning in People with Mild to Moderate Parkinson's Disease: Behavior and Related Brain Function.

BACKGROUND: Deficits in motor learning could be an important explanation for the balance and gait impairments characteristic of people with Parkinson's disease (PD). Empirical studies often report that so-called implicit motor sequence learning is impaired in people with PD, but the results are inconclusive. Altered brain activity during implicit motor sequence learning has also been reported for people with PD in comparison to healthy individuals. OBJECTIVE: To investigate implicit motor sequence learning and associated neural correlates in individuals with mild to moderate PD. METHODS: Fifty-seven participants with PD and 34 healthy participants, all ≥60 years of age, performed the serial reaction time task (SRTT) during the acquisition of functional magnetic resonance imaging (fMRI) data. We analyzed the SRTT as a measure of implicit motor sequence learning in two complementary ways. We analyzed the task-induced fMRI data within regions of interest (ROIs) as well as functional connectivity between ROIs. RESULTS: We found a significant group difference in SRTT performance indicating that the participants with PD had a somewhat lower level of implicit motor sequence learning than the healthy participants. Exploratory analyses suggested that impairments in implicit motor sequence learning for people with PD might be due to a lower learning rate. We did not find any significant group differences in the fMRI data. CONCLUSION: Our exploratory finding of a lower implicit motor learning rate in PD could have important implications for how people with PD should practice new motor tasks and physical exercise. Future studies need to confirm this finding with hypothesis-driven analyses.

Multiclass prediction of different dementia syndromes based on multi-centric volumetric MRI imaging.

INTRODUCTION: Dementia syndromes can be difficult to diagnose. We aimed at building a classifier for multiple dementia syndromes using magnetic resonance imaging (MRI). METHODS: Atlas-based volumetry was performed on T1-weighted MRI data of 426 patients and 51 controls from the multi-centric German Research Consortium of Frontotemporal Lobar Degeneration including patients with behavioral variant frontotemporal dementia, Alzheimer's disease, the three subtypes of primary progressive aphasia, i.e., semantic, logopenic and nonfluent-agrammatic variant, and the atypical parkinsonian syndromes progressive supranuclear palsy and corticobasal syndrome. Support vector machine classification was used to classify each patient group against controls (binary classification) and all seven diagnostic groups against each other in a multi-syndrome classifier (multiclass classification). RESULTS: The binary classification models reached high prediction accuracies between 71 and 95% with a chance level of 50%. Feature importance reflected disease-specific atrophy patterns. The multi-syndrome model reached accuracies of more than three times higher than chance level but was far from 100%. Multi-syndrome model performance was not homogenous across dementia syndromes, with better performance in syndromes characterized by regionally specific atrophy patterns. Whereas diseases generally could be classified vs controls more correctly with increasing severity and duration, differentiation between diseases was optimal in disease-specific windows of severity and duration. DISCUSSION: Results suggest that automated methods applied to MR imaging data can support physicians in diagnosis of dementia syndromes. It is particularly relevant for orphan diseases beside frequent syndromes such as Alzheimer's disease.

Bioprinting of 3D Adipose Tissue Models Using a GelMA-Bioink with Human Mature Adipocytes or Human Adipose-Derived Stem Cells.

Adipose tissue is related to the development and manifestation of multiple diseases, demonstrating the importance of suitable in vitro models for research purposes. In this study, adipose tissue lobuli were explanted, cultured, and used as an adipose tissue control to evaluate in vitro generated adipose tissue models. During culture, lobule exhibited a stable weight, lactate dehydrogenase, and glycerol release over 15 days. For building up in vitro adipose tissue models, we adapted the biomaterial gelatin methacryloyl (GelMA) composition and handling to homogeneously mix and bioprint human primary mature adipocytes (MA) and adipose-derived stem cells (ASCs), respectively. Accelerated cooling of the bioink turned out to be essential for the homogeneous distribution of lipid-filled MAs in the hydrogel. Last, we compared manual and bioprinted GelMA hydrogels with MA or ASCs and the explanted lobules to evaluate the impact of the printing process and rate the models concerning the physiological reference. The viability analyses demonstrated no significant difference between the groups due to additive manufacturing. The staining of intracellular lipids and perilipin A suggest that GelMA is well suited for ASCs and MA. Therefore, we successfully constructed physiological in vitro models by bioprinting MA-containing GelMA bioinks.

Unraveling Parkinson's disease heterogeneity using subtypes based on multimodal data.

BACKGROUND: Parkinson's disease (PD) is a clinically and neuroanatomically heterogeneous neurodegenerative disease characterized by different subtypes. To this date, no studies have used multimodal data that combines clinical, motor, cognitive and neuroimaging assessments to identify these subtypes, which may provide complementary, clinically relevant information. To address this limitation, we subtyped participants with mild-moderate PD based on a rich, multimodal dataset of clinical, cognitive, motor, and neuroimaging variables. METHODS: Cross-sectional data from 95 PD participants from our randomized EXPANd (EXercise in PArkinson's disease and Neuroplasticity) controlled trial were included. Participants were subtyped using clinical, motor, and cognitive assessments as well as structural and resting-state MRI data. Subtyping was done by random forest clustering. We extracted information about the subtypes by inspecting their neuroimaging profiles and descriptive statistics. RESULTS: Our multimodal subtyping analysis yielded three PD subtypes: a motor-cognitive subtype characterized by widespread alterations in brain structure and function as well as impairment in motor and cognitive abilities; a cognitive dominant subtype mainly impaired in cognitive function that showed frontoparietal structural and functional changes; and a motor dominant subtype impaired in motor variables without any brain alterations. Motor variables were most important for the subtyping, followed by gray matter volume in the right medial postcentral gyrus. CONCLUSIONS: Three distinct PD subtypes were identified in our multimodal dataset. The most important features to subtype PD participants were motor variables in addition to structural MRI in the sensorimotor region. These findings have the potential to improve our understanding of PD heterogeneity, which in turn can lead to personalized interventions and rehabilitation.

Gellan Gum Is a Suitable Biomaterial for Manual and Bioprinted Setup of Long-Term Stable, Functional 3D-Adipose Tissue Models.

Due to its wide-ranging endocrine functions, adipose tissue influences the whole body's metabolism. Engineering long-term stable and functional human adipose tissue is still challenging due to the limited availability of suitable biomaterials and adequate cell maturation. We used gellan gum (GG) to create manual and bioprinted adipose tissue models because of its similarities to the native extracellular matrix and its easily tunable properties. Gellan gum itself was neither toxic nor monocyte activating. The resulting hydrogels exhibited suitable viscoelastic properties for soft tissues and were stable for 98 days in vitro. Encapsulated human primary adipose-derived stem cells (ASCs) were adipogenically differentiated for 14 days and matured for an additional 84 days. Live-dead staining showed that encapsulated cells stayed viable until day 98, while intracellular lipid staining showed an increase over time and a differentiation rate of 76% between days 28 and 56. After 4 weeks of culture, adipocytes had a univacuolar morphology, expressed perilipin A, and secreted up to 73% more leptin. After bioprinting establishment, we demonstrated that the cells in printed hydrogels had high cell viability and exhibited an adipogenic phenotype and function. In summary, GG-based adipose tissue models show long-term stability and allow ASCs maturation into functional, univacuolar adipocytes.

Characterization of Mild and Moderate Dysarthria in Parkinson's Disease: Behavioral Measures and Neural Correlates.

Purpose: Alterations in speech and voice are among the most common symptoms in Parkinson's disease (PD), often resulting in motor speech disorders such as hypokinetic dysarthria. We investigated dysarthria, verbal fluency, executive functions, and global cognitive function in relation to structural and resting-state brain changes in people with PD. Methods: Participants with mild-moderate PD (n = 83) were recruited within a randomized controlled trial and divided into groups with varying degrees of dysarthria: no dysarthria (noDPD), mild dysarthria (mildDPD), moderate dysarthria (modDPD), and also combined mildDPD and modDPD into one group (totDPD). Voice sound level and dysphonia, verbal fluency, motor symptoms, executive functions, disease severity, global cognition, and neuroimaging were compared between groups. Gray matter volume and intensity of spontaneous brain activity were analyzed. Additionally, regressions between behavioral and neuroimaging data were performed. Results: The groups differed significantly in mean voice sound level, dysphonia, and motor symptom severity. Comparing different severity levels of dysarthria to noDPD, groups differed focally in resting-state activity, but not in brain structure. In totDPD, lower scores on semantic verbal fluency, a composite score of executive functions, and global cognition correlated with lower superior temporal gyrus volume. Conclusion: This study shows that severity of dysarthria may be related to underlying structural and resting-state brain alterations in PD as well as behavioral changes. Further, the superior temporal gyrus may play an important role in executive functions, language, and global cognition in people with PD and dysarthria.

Scaffolds for Cultured Meat on the Basis of Polysaccharide Hydrogels Enriched with Plant-Based Proteins.

The world population is growing and alternative ways of satisfying the increasing demand for meat are being explored, such as using animal cells for the fabrication of cultured meat. Edible biomaterials are required as supporting structures. Hence, we chose agarose, gellan and a xanthan-locust bean gum blend (XLB) as support materials with pea and soy protein additives and analyzed them regarding material properties and biocompatibility. We successfully built stable hydrogels containing up to 1% pea or soy protein. Higher amounts of protein resulted in poor handling properties and unstable gels. The gelation temperature range for agarose and gellan blends is between 23-30 °C, but for XLB blends it is above 55 °C. A change in viscosity and a decrease in the swelling behavior was observed in the polysaccharide-protein gels compared to the pure polysaccharide gels. None of the leachates of the investigated materials had cytotoxic effects on the myoblast cell line C2C12. All polysaccharide-protein blends evaluated turned out as potential candidates for cultured meat. For cell-laden gels, the gellan blends were the most suitable in terms of processing and uniform distribution of cells, followed by agarose blends, whereas no stable cell-laden gels could be formed with XLB blends.

Behavioural and neuroplastic effects of a double-blind randomised controlled balance exercise trial in people with Parkinson's disease.

Balance dysfunction is a disabling symptom in people with Parkinson's disease (PD). Evidence suggests that exercise can improve balance performance and induce neuroplastic effects. We hypothesised that a 10-week balance intervention (HiBalance) would improve balance, other motor and cognitive symptoms, and alter task-evoked brain activity in people with PD. We performed a double-blind randomised controlled trial (RCT) where 95 participants with PD were randomised to either HiBalance (n = 48) or a control group (n = 47). We found no significant group by time effect on balance performance (b = 0.4 95% CI [-1, 1.9], p = 0.57) or on our secondary outcomes, including the measures of task-evoked brain activity. The findings of this well-powered, double-blind RCT contrast previous studies of the HiBalance programme but are congruent with other double-blind RCTs of physical exercise in PD. The divergent results raise important questions on how to optimise physical exercise interventions for people with PD.Preregistration clinicaltrials.gov: NCT03213873.

Olive Oil Extracts and Oleic Acid Attenuate the LPS-Induced Inflammatory Response in Murine RAW264.7 Macrophages but Induce the Release of Prostaglandin E2.

Olive oil contains high amounts of oleic acid (OA). Although OA has been described to inhibit inflammatory processes, the effects of olive oil on cellular mechanisms remain poorly understood. Therefore, we compared the effects of major fatty acids (FA) from olive oil with those of olive oil extracts (OOE) on inflammatory mediators and alterations in the cellular phospholipid composition in murine macrophages. Upon treatment with different OOE, FA compositions of lipopolysaccharide (LPS)-stimulated murine RAW264.7 macrophages were analyzed using gas chromatography. Olive oil extracts and OA significantly reduced the LPS-induced expression of inducible nitric oxide synthase (iNos), cyclooxygenase (Cox2), and interleukin-6 mRNA. In addition, a significant decrease in Cox2 and iNos protein expression was observed. The formation of nitric oxide was significantly reduced, while the formation of prostaglandin (PG) E2 from arachidonic acid significantly increased after treatment with OOE or OA. The latter was associated with a shift in the phospholipid FA composition from arachidonic acid to OA, resulting in an elevated availability of arachidonic acid. Together, OOE and OA mediate anti-inflammatory effects in vitro but increase the release of arachidonic acid and hereinafter PGE2, likely due to elongation of OA and competitive incorporation of fatty acids into membrane phospholipids.

Homogeneous and Reproducible Mixing of Highly Viscous Biomaterial Inks and Cell Suspensions to Create Bioinks.

Highly viscous bioinks offer great advantages for the three-dimensional fabrication of cell-laden constructs by microextrusion printing. However, no standardised method of mixing a high viscosity biomaterial ink and a cell suspension has been established so far, leading to non-reproducible printing results. A novel method for the homogeneous and reproducible mixing of the two components using a mixing unit connecting two syringes is developed and investigated. Several static mixing units, based on established mixing designs, were adapted and their functionality was determined by analysing specific features of the resulting bioink. As a model system, we selected a highly viscous ink consisting of fresh frozen human blood plasma, alginate, and methylcellulose, and a cell suspension containing immortalized human mesenchymal stem cells. This bioink is crosslinked after fabrication. A pre-crosslinked gellan gum-based bioink providing a different extrusion behaviour was introduced to validate the conclusions drawn from the model system. For characterisation, bioink from different zones within the mixing device was analysed by measurement of its viscosity, shape fidelity after printing and visual homogeneity. When taking all three parameters into account, a comprehensive and reliable comparison of the mixing quality was possible. In comparison to the established method of manual mixing inside a beaker using a spatula, a significantly higher proportion of viable cells was detected directly after mixing and plotting for both bioinks when the mixing unit was used. A screw-like mixing unit, termed "HighVisc", was found to result in a homogenous bioink after a low number of mixing cycles while achieving high cell viability rates.