Conditioned Medium from Cells Overexpressing TDP-43 Alters the Metabolome of Recipient Cells

Abstract

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease caused by the progressive death of both upper and lower motor neurons. The disease presents a poor prognosis, and patients usually die 2–5 years after the onset of symptoms. The hallmark of this disease is the presence of phosphorylated and ubiquitinated aggregates containing trans-active response DNA-binding protein-43 (TDP-43) in the cytoplasm of motor neurons. TDP-43 pathology has been associated with multiple pathways in ALS, such as metabolic dysfunction found in patients and in in vivo models. Recently, it has been described as a “prion-like” protein, as studies have shown its propagation in cell culture from ALS brain extract or overexpressed TDP-43 in co-culture and conditioned medium, resulting in cytotoxicity. However, the cellular alterations that are associated with this cytotoxicity require further investigation. Here, we investigated the effects of conditioned medium from HEK293T (Human Embryonic Kidney 293T) cells overexpressing TDP-43 on cellular morphology, proliferation, death, and metabolism. Although we did not find evidence of TDP-43 propagation, we observed a toxicity of TDP-43-conditioned medium and altered metabolism. These results, therefore, suggest (1) that cells overexpressing TDP-43 produce an extracellular environment that can perturb other cells and (2) that TDP-43 propagation alone may not be the only potentially cytotoxic cell-to-cell mechanism.

DNAJA1 Dysregulates Metabolism Promoting an Antiapoptotic Phenotype in Pancreatic Ductal Adenocarcinoma

“…A number of prior studies have failed to identify any metabolic signature associated with the overexpression of a protein in a cell line. 42,43 A tree diagram (Figure 1F) derived from the score plot (Figure 1E DNAJA1 expression, indicating metabolome perturbations accumulated over time. However, the tree diagram indicates that the metabolome of WT-Bx cells did not change over time.…”

“…Importantly, these metabolic differences are likely due to the cellular activity of DNAJA1 instead of a generic response to the process of overexpressing a protein. A number of prior studies have failed to identify any metabolic signature associated with the overexpression of a protein in a cell line. , …”

“…Overall, these metabolic changes support DNAJA1 serving a proto-oncogenic role in PDAC and are inconsistent with a simple metabolic signature of protein overexpression. 42,43,67 A future detailed flux analysis may prove valuable for the further characterization of these metabolic changes. D1 cells presented apparent weaker lysosomal membranes, decreased actin filament formation, and increased invasion relative to WT.…”

Mechanisms of TDP-43 Proteinopathy Onset and Propagation

“…It is important to note however, that not all cell studies support the notion of TDP-43 propagation. Two reports have shown that while cells may release TDP-43 into the media, there is no evidence of uptake of this protein into recipient cells treated with the conditioned media [136,137]. The conditioned media did result in cytotoxicity and metabolic changes [137], suggesting that TDP-43 may have extracellular impacts.…”

“…Two reports have shown that while cells may release TDP-43 into the media, there is no evidence of uptake of this protein into recipient cells treated with the conditioned media [136,137]. The conditioned media did result in cytotoxicity and metabolic changes [137], suggesting that TDP-43 may have extracellular impacts. Clearly, more work is required to fully understand the mechanism of spread of TDP-43 pathology, as well as identifying potential inter-versus extracellular effects.…”