Human cerebrospinal fluid affects chemoradiotherapy sensitivities in tumor cells from patients with glioblastoma.

Cancers in the central nervous system resist therapies effective in other cancers, possibly due to the unique biochemistry of the human brain microenvironment composed of cerebrospinal fluid (CSF). However, the impact of CSF on cancer cells and therapeutic efficacy is unknown. Here, we examined the effect of human CSF on glioblastoma (GBM) tumors from 25 patients. We found that CSF induces tumor cell plasticity and resistance to standard GBM treatments (temozolomide and irradiation). We identified nuclear protein 1 (NUPR1), a transcription factor hampering ferroptosis, as a mediator of therapeutic resistance in CSF. NUPR1 inhibition with a repurposed antipsychotic, trifluoperazine, enhanced the killing of GBM cells resistant to chemoradiation in CSF. The same chemo-effective doses of trifluoperazine were safe for human neurons and astrocytes derived from pluripotent stem cells. These findings reveal that chemoradiation efficacy decreases in human CSF and suggest that combining trifluoperazine with standard care may improve the survival of patients with GBM.

Long-term adherence of human brain cells in vitro is enhanced by charged amine-based plasma polymer coatings.

Advances in cellular reprogramming have radically increased the use of patient-derived cells for neurological research in vitro. However, adherence of human neurons on tissue cultureware is unreliable over the extended periods required for electrophysiological maturation. Adherence issues are particularly prominent for transferable glass coverslips, hindering imaging and electrophysiological assays. Here, we assessed thin-film plasma polymer treatments, polymeric factors, and extracellular matrix coatings for extending the adherence of human neuronal cultures on glass. We find that positive-charged, amine-based plasma polymers improve the adherence of a range of human brain cells. Diaminopropane (DAP) treatment with laminin-based coating optimally supports long-term maturation of fundamental ion channel properties and synaptic activity of human neurons. As proof of concept, we demonstrated that DAP-treated glass is ideal for live imaging, patch-clamping, and optogenetics. A DAP-treated glass surface reduces the technical variability of human neuronal models and enhances electrophysiological maturation, allowing more reliable discoveries of treatments for neurological and psychiatric disorders.

Retromer regulates the lysosomal clearance of MAPT/tau.

The macroautophagy/autophagy-lysosome axis enables the clearance and degradation of cytoplasmic components including protein aggregates, damaged organelles and invading pathogens. Protein aggregation and lysosomal system dysfunction in the brain are common features of several late-onset neurological disorders including Alzheimer disease. Spatial overlap between depletion of the endosomal-sorting complex retromer and MAPT/tau aggregation in the brain have been previously reported. However, whether retromer dysfunction plays a direct role in mediating MAPT aggregation remains unclear. Here, we demonstrate that the autophagy-lysosome axis is the primary mode for the clearance of aggregated species of MAPT using both chemical and genetic approaches in cell models of amyloid MAPT aggregation. We show that depletion of the central retromer component VPS35 causes a block in the resolution of autophagy. We establish that this defect underlies marked accumulation of cytoplasmic MAPT aggregates upon VPS35 depletion, and that VPS35 overexpression has the opposite effect. This work illustrates how retromer complex integrity regulates the autophagy-lysosome axis to suppress MAPT aggregation and spread.

BrainPhys neuronal medium optimized for imaging and optogenetics in vitro.

The capabilities of imaging technologies, fluorescent sensors, and optogenetics tools for cell biology are advancing. In parallel, cellular reprogramming and organoid engineering are expanding the use of human neuronal models in vitro. This creates an increasing need for tissue culture conditions better adapted to live-cell imaging. Here, we identify multiple caveats of traditional media when used for live imaging and functional assays on neuronal cultures (i.e., suboptimal fluorescence signals, phototoxicity, and unphysiological neuronal activity). To overcome these issues, we develop a neuromedium called BrainPhys™ Imaging (BPI) in which we optimize the concentrations of fluorescent and phototoxic compounds. BPI is based on the formulation of the original BrainPhys medium. We benchmark available neuronal media and show that BPI enhances fluorescence signals, reduces phototoxicity and optimally supports the electrical and synaptic activity of neurons in culture. We also show the superior capacity of BPI for optogenetics and calcium imaging of human neurons. Altogether, our study shows that BPI improves the quality of a wide range of fluorescence imaging applications with live neurons in vitro while supporting optimal neuronal viability and function.