Millifluidic culture improves human midbrain organoid vitality and differentiation

Human midbrain-specific organoids (hMOs) serve as an experimental in vitro model for studying the pathogenesis of Parkinson's disease (PD). In hMOs, neuroepithelial stem cells (NESCs) give rise to functional midbrain dopaminergic (mDA) neurons that are selectively degenerating during PD. A limitation of the hMO model is an under-supply of oxygen and nutrients to the densely packed core region, which leads eventually to a "dead core". To reduce this phenomenon, we applied a millifluidic culture system that ensures media supply by continuous laminar flow. We developed a computational model of oxygen transport and consumption in order to predict oxygen levels within the hMOs. The modelling predicts higher oxygen levels in the hMO core region under millifluidic conditions. In agreement with the computational model, a significantly smaller "dead core" was observed in hMOs cultured in a bioreactor system compared to those ones kept under conventional shaking conditions. Comparing the necrotic core regions in the organoids with those obtained from the model allowed an estimation of the critical oxygen concentration necessary for ensuring cell vitality. Besides the reduced "dead core" size, the differentiation efficiency from NESCs to mDA neurons was elevated in hMOs exposed to medium flow. Increased differentiation involved a metabolic maturation process that was further developed in the millifluidic culture. Overall, bioreactor conditions that improve hMO quality are worth considering in the context of advanced PD modelling

Examining the growth and stable isotopes of phytoplankton and periphyton communities exposed to oil sands reclamation strategies

The impacts of oil sands processed materials (OSPM) on phytoplankton and periphyton community growth and stable carbon and nitrogen isotopes were examined. Estimates of plankton and periphyton community growth, measured as chl a and dry weight, were low and similar in reference and OSPM reclamation wetlands. The use of stable isotope analyses revealed higher δ15N of plankton and periphyton in OSPM wetlands than reference wetlands, possibly due to increased TN concentrations in some OSPM wetlands. In the laboratory, water-soluble fractions (WSF) of two types of OSPM (mature fine tailings, MFT and consolidated tailings, CT) and an amendment material (peat-mineral mixture), potential fill materials in wetland or end pit lake reclamation, were examined for phytoplankton community growth and stable carbon and nitrogen isotopes. All WSF treatments had higher chl a compared to reference water and maximum growth was observed at a 50:50 ratio of peat:CT or peat:MFT. In general, WSFs of peat had the highest concentration of total nitrogen (TN) whereas WSFs of MFT had the highest total phosphorus (TP; 3x higher). The results suggested that the addition of peat as an amendment to OSPM (particularly for MFT), contributing additional TN, could improve phytoplankton community growth in oil sands reclamation. At higher percentages of MFT WSF, there was increased turbidity due to fine clay particles that likely contributed to reduced phytoplankton growth. Turbidity could be an important factor limiting phytoplankton growth and thus reducing dietary resources and biological detritus (via sedimentation) in the initial development of an end pit lake. The WSFs also promoted the unfavourable growth of filamentous algae, highest at intermediate concentrations of peat and CT WSFs and inhibited in MFT WSFs due to light limitation. Stable N isotopes of plankton and filamentous algae suggests that 15N enrichment of algae could be a useful indicator of nutrient inputs, including OSPM seepage into natural aquatic systems, for oil sands regional monitoring programs

The Interplay between Protein L-Isoaspartyl Methyltransferase Activity and Insulin-Like Signaling to Extend Lifespan in Caenorhabditis elegans

The protein L-isoaspartyl-O-methyltransferase functions to initiate the repair of isomerized aspartyl and asparaginyl residues that spontaneously accumulate with age in a variety of organisms. Caenorhabditis elegans nematodes lacking the pcm-1 gene encoding this enzyme display a normal lifespan and phenotype under standard laboratory growth conditions. However, significant defects in development, egg laying, dauer survival, and autophagy have been observed in pcm-1 mutant nematodes when deprived of food and when exposed to oxidative stress. Interestingly, overexpression of this repair enzyme in both Drosophila and C. elegans extends adult lifespan under thermal stress. In this work, we show the involvement of the insulin/insulin-like growth factor-1 signaling (IIS) pathway in PCM-1-dependent lifespan extension in C. elegans. We demonstrate that reducing the levels of the DAF-16 downstream transcriptional effector of the IIS pathway by RNA interference reduces the lifespan extension resulting from PCM-1 overexpression. Using quantitative real-time PCR analysis, we show the up-regulation of DAF-16-dependent stress response genes in the PCM-1 overexpressor animals compared to wild-type and pcm-1 mutant nematodes under mild thermal stress conditions. Additionally, similar to other long-lived C. elegans mutants in the IIS pathway, including daf-2 and age-1 mutants, PCM-1 overexpressor adult animals display increased resistance to severe thermal stress, whereas pcm-1 mutant animals survive less long under these conditions. Although we observe a higher accumulation of damaged proteins in pcm-1 mutant nematodes, the basal level of isoaspartyl residues detected in wild-type animals was not reduced by PCM-1 overexpression. Our results support a signaling role for the protein L-isoaspartyl methyltransferase in lifespan extension that involves the IIS pathway, but that may be independent of its function in overall protein repair

The Role of DJ-1 in Cellular Metabolism and Pathophysiological Implications for Parkinson’s Disease

DJ-1 is a multifunctional protein associated with pathomechanisms implicated in different chronic diseases including neurodegeneration, cancer and diabetes. Several of the physiological functions of DJ-1 are not yet fully understood; however, in the last years, there has been increasing evidence for a potential role of DJ-1 in the regulation of cellular metabolism. Here, we summarize the current knowledge on specific functions of DJ-1 relevant to cellular metabolism and their role in modulating metabolic pathways. Further, we illustrate pathophysiological implications of the metabolic effects of DJ-1 in the context of neurodegeneration in Parkinson´s disease