The effect of A1 and A2 reactive astrocyte expression on hydrocephalus shunt failure.

BACKGROUND: The composition of tissue obstructing neuroprosthetic devices is largely composed of inflammatory cells with a significant astrocyte component. In a first-of-its-kind study, we profile the astrocyte phenotypes present on hydrocephalus shunts. METHODS: qPCR and RNA in-situ hybridization were used to quantify pro-inflammatory (A1) and anti-inflammatory (A2) reactive astrocyte phenotypes by analyzing C3 and EMP1 genes, respectively. Additionally, CSF cytokine levels were quantified using ELISA. In an in vitro model of astrocyte growth on shunts, different cytokines were used to prevent the activation of resting astrocytes into the A1 and A2 phenotypes. Obstructed and non-obstructed shunts were characterized based on the degree of actual tissue blockage on the shunt surface instead of clinical diagnosis. RESULTS: The results showed a heterogeneous population of A1 and A2 reactive astrocytes on the shunts with obstructed shunts having a significantly higher proportion of A2 astrocytes compared to non-obstructed shunts. In addition, the pro-A2 cytokine IL-6 inducing proliferation of astrocytes was found at higher concentrations among CSF from obstructed samples. Consequently, in the in vitro model of astrocyte growth on shunts, cytokine neutralizing antibodies were used to prevent activation of resting astrocytes into the A1 and A2 phenotypes which resulted in a significant reduction in both A1 and A2 growth. CONCLUSIONS: Therefore, targeting cytokines involved with astrocyte A1 and A2 activation is a promising intervention aimed to prevent shunt obstruction.

A high-resolution real-time quantification of astrocyte cytokine secretion under shear stress for investigating hydrocephalus shunt failure.

It has been hypothesized that physiological shear forces acting on medical devices implanted in the brain significantly accelerate the rate to device failure in patients with chronically indwelling neuroprosthetics. In hydrocephalus shunt devices, shear forces arise from cerebrospinal fluid flow. The shunt's unacceptably high failure rate is mostly due to obstruction with adherent inflammatory cells. Astrocytes are the dominant cell type bound directly to obstructing shunts, rapidly manipulating their activation via shear stress-dependent cytokine secretion. Here we developed a total internal reflection fluorescence microscopy combined with a microfluidic shear device chip (MSDC) for quantitative analysis and direct spatial-temporal mapping of secreted cytokines at the single-cell level under physiological shear stress to identify the root cause for shunt failure. Real-time secretion imaging at 1-min time intervals enabled successful detection of a significant increase of astrocyte IL-6 cytokine secretion under shear stress greater than 0.5 dyne/cm2, validating our hypothesis and highlighting the importance of reducing shear stress activation of cells.

Methotrexate-loaded nitrogen-doped graphene quantum dots nanocarriers as an efficient anticancer drug delivery system.

Graphene quantum dots (GQDs) are new efficient nanomaterials used in therapeutic applications. In this study, blue fluorescent nitrogen-doped GQDs (N-GQDs) were synthesized by a hydrothermal method via pyrolisis of citric acid as the carbon source and urea as the nitrogen source. The existence of doped nitrogen in GQDs was confirmed by FTIR characterization. Here, for the first time, the N-GQDs were loaded with the anticancer drug, methotrexate (MTX), to prepare MTX-(N-GQDs) as an efficient drug delivery system. The establishment of the strong π-π stacking interaction between MTX and N-GQDs was confirmed by FTIR and UV-vis spectroscopies indicating successful loading of MTX to N-GQDs. The in-vitro cytotoxicity of MTX-(N-GQDs) on human breast cancer cells investigated through MTT assay suggested that the drug-free N-GQDs nanocarriers are highly biocompatible, whereas the MTX-loaded ones are more cytotoxic than the free MTX.