Integrating biological vasculature into a multi-organ-chip microsystem

Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG geförderten) Allianz- bzw. Nationallizenz frei zugänglich.This publication is with permission of the rights owner freely accessible due to an Alliance licence and a national licence (funded by the DFG, German Research Foundation) respectively.A chip-based system mimicking the transport function of the human cardiovascular system has been established at minute but standardized microsystem scale. A peristaltic on-chip micropump generates pulsatile shear stress in a widely adjustable physiological range within a microchannel circuit entirely covered on all fluid contact surfaces with human dermal microvascular endothelial cells. This microvascular transport system can be reproducibly established within four days, independently of the individual endothelial cell donor background. It interconnects two standard tissue culture compartments, each of 5 mm diameter, through microfluidic channels of 500 μm width. Further vessel branching and vessel diameter reduction down to a microvessel scale of approximately 40 μm width was realised by a two-photon laser ablation technique applied to inserts, designed for the convenient establishment of individual organ equivalents in the tissue culture compartments at a later time. The chip layout ensures physiological fluid-to-tissue ratios. Moreover, an in-depth microscopic analysis revealed the fine-tuned adjustment of endothelial cell behaviour to local shear stresses along the microvasculature of the system. Time-lapse and 3D imaging two-photon microscopy were used to visualise details of spatiotemporal adherence of the endothelial cells to the channel system and to each other. The first indicative long-term experiments revealed stable performance over two and four weeks. The potential application of this system for the future establishment of human-on-a-chip systems and basic human endothelial cell research is discussed.BMBF, 0315569, GO-Bio 3: Multi-Organ-Bioreaktoren für die prädiktive Substanztestung im Chipforma

scAmpi—A versatile pipeline for single-cell RNA-seq analysis from basics to clinics

Single-cell RNA sequencing (scRNA-seq) has emerged as a powerful technique to decipher tissue composition at the single-cell level and to inform on disease mechanisms, tumor heterogeneity, and the state of the immune microenvironment. Although multiple methods for the computational analysis of scRNA-seq data exist, their application in a clinical setting demands standardized and reproducible workflows, targeted to extract, condense, and display the clinically relevant information. To this end, we designed scAmpi (Single Cell Analysis mRNA pipeline), a workflow that facilitates scRNA-seq analysis from raw read processing to informing on sample composition, clinically relevant gene and pathway alterations, and in silico identification of personalized candidate drug treatments. We demonstrate the value of this workflow for clinical decision making in a molecular tumor board as part of a clinical study

Characterization of diabetes following pancreatic surgery in patients with congenital hyperinsulinism

Background: Congenital hyperinsulinism (CHI) is the most common cause of persistent hypoglycaemia in infancy that leads to unfavourable neurological outcome if not treated adequately. In patients with severe diffuse CHI it remains under discussion whether pancreatic surgery should be performed or intensive medical treatment with the acceptance of recurrent episodes of mild hypoglycaemia is justified. Near-total pancreatectomy is associated with high rates of insulin-dependent diabetes mellitus and exocrine pancreatic insufficiency. Little is known about the management and long-term glycaemic control of CHI patients with diabetes after pancreatic surgery. We searched the German/Austrian DPV database and compared the course of 42 CHI patients with diabetes to that of patients with type 1 diabetes mellitus (T1DM). Study groups were compared at diabetes onset and after a follow-up period of 6.1 [3.3–9.7] (median [interquartile range]) years. Results: The majority of CHI patients with diabetes were treated with insulin (85.2% [70.9–99.5] at diabetes onset, and 90.5% [81.2–99.7] at follow-up). However, compared to patients with T1DM, significantly more patients in the CHI group with diabetes were treated with conventional insulin therapy (47.8% vs. 24.4%, p = 0.03 at diabetes onset, and 21.1% vs. 6.4% at follow-up, p = 0.003), and only a small number of CHI patients were treated with insulin pumps. Daily insulin dose was significantly lower in CHI patients with diabetes than in patients with T1DM, both at diabetes onset (0.3 [0.2–0.5] vs. 0.6 IE/kg/d [0.4–0.8], p = 0.003) and follow-up (0.8 [0.4–1.0] vs. 0.9 [0.7–1.0] IE/kg/d, p = 0.02), while daily carbohydrate intake was comparable in both groups. Within the first treatment year, HbA1c levels were significantly lower in CHI patients with diabetes (6.2% [5.5–7.9] vs. 7.2% [6.5–8.2], p = 0.003), but increased to a level comparable to that of T1DM patients at follow-up. Interestingly, in CHI patients, the risk of severe hypoglycaemia tends to be higher only at diabetes onset (14.8% vs. 5.8%, p = 0.1). Conclusions: In surgically treated CHI patients insulin treatment needs to be intensified in order to achieve good glycaemic control. Our data furthermore emphasize the need for improved medical treatment options for patients with diazoxide- and/or octreotide-unresponsive CHI

Chilling and forcing from cut twigs - how to simplify phenological experiments for citizen science

Low-cost phenological experiments with cut twigs are increasingly used to study bud development in response to spring warming and photoperiod. However, a broader variety of species needs to be tackled and in particular the influence of insufficient winter chilling deserves more attention. Therefore, we investigated if and how chilling requirements can be efficiently investigated by cut twigs and how this low-tech approach could be successfully implemented as a citizen science or school project. We conducted an experiment on bud burst and leaf development of Corylus avellana L. twigs, with natural chilling outdoors on a shrub (S) and another chilling treatment as cut twigs in containers (C), and subsequent forcing indoors. Subsampling of the number of cutting dates and number of twigs was used to infer minimum required sample sizes. Apart from insufficiently chilled twigs, ~80% of the twigs (both S and C) reached leaf out. For multiple definitions of chilling and forcing, a negative exponential relationship was revealed between chilling and amount of forcing needed to reach certain developmental stages. At least 5 out of 15 cutting dates or alternatively half of the 10 twig repetitions, but especially those mirroring low chilling conditions, were needed to describe the chilling-forcing relationship with some degree of robustness. In addition, for cutting dates with long chilling, i.e., from January onwards, freshly cut twigs (S) required significantly more forcing to reach bud burst than twigs from containers (C), although the effect was small. In general, chilling conditions of mature shrubs were well captured by cut twigs, therefore opening the possibility of chilling through refrigeration. We conclude that experimental protocols as outlined here are feasible for citizen scientists, school projects, and science education, and would have the potential to advance the research field if carried out on a large scale. We provide an easy-to-use Shiny simulation app to enable citizen scientists to build up a bud development model based on their own experimental data and then simulate future phenological development with winter and/or spring warming. This may encourage them to further study other aspects of climate change and the impacts of climate change