VGF Protein and Its C-Terminal Derived Peptides in Amyotrophic Lateral Sclerosis: Human and Animal Model Studies

Abstract

VGF mRNA is widely expressed in areas of the nervous system known to degenerate in Amyotrophic Lateral Sclerosis (ALS), including cerebral cortex, brainstem and spinal cord. Despite certain VGF alterations are reported in animal models, little information is available with respect to the ALS patients. We addressed VGF peptide changes in fibroblast cell cultures and in plasma obtained from ALS patients, in parallel with spinal cord and plasma samples from the G93A-SOD1 mouse model. Antisera specific for the C-terminal end of the human and mouse VGF proteins, respectively, were used in immunohistochemistry and enzyme-linked immunosorbent assay (ELISA), while gel chromatography and HPLC/ESI-MS/MS were used to identify the VGF peptides present. Immunoreactive VGF C-terminus peptides were reduced in both fibroblast and plasma samples from ALS patients in an advanced stage of the disease. In the G93A-SOD1 mice, the same VGF peptides were also decreased in plasma in the late-symptomatic stage, while showing an earlier down-regulation in the spinal cord. In immunohistochemistry, a large number of gray matter structures were VGF C-terminus immunoreactive in control mice (including nerve terminals, axons and a few perikarya identified as motoneurons), with a striking reduction already in the pre-symptomatic stage. Through gel chromatography and spectrometry analysis, we identified one form likely to be the VGF precursor as well as peptides containing the NAPP- sequence in all tissues studied, while in the mice and fibroblasts, we revealed also AQEE- and TLQP- peptides. Taken together, selective VGF fragment depletion may participate in disease onset and/or progression of ALS.

TLQP Peptides in Amyotrophic Lateral Sclerosis: Possible Blood Biomarkers with a Neuroprotective Role

“…2F). TLQP peptide/s immunoreactivity was revealed in a region of cytoplasm close to the nucleus, bona fide the Golgi area, as previously shown (Brancia et al, 2016) and in agreement with the role of VGF as precursor of secretory products (Brancia et al, 2005). In both ALS patients and controls ( Fig.…”

“…In fact, measurement of most neurotrophic factors, including BDNF (Tremolizzo et al, 2016), failed to selectively detect ALS patients and changes at an early phase (Turner et al, 2009). In a previous study, we revealed changes in peptides derived from a different part of VGF, namely the region encompassing the C-terminal end of the VGF precursor (Brancia et al, 2016).…”

“…Transgenic animals were grouped according to age (days postnatal: P) and stage of motor dysfunctions (Brancia et al, 2016), as follows: (i) pre-symptomatic (around P45);…”

“…In ALS, despite the reported evidence that VGF expression is modulated in the animal model and humans (Pasinetti et al, 2006;Zhao et al, 2008), limited information is available regarding TLQP peptides. We have previously reported the involvement of the VGF C-terminal peptides in ALS, modulated in the SOD1 mutant mice and patient's plasma, but exclusively at the final disease phase (Brancia et al, 2016). Afterwards, we aimed at specifically investigating the role of the TLQP peptides in ALS, by studing their expression and changes (using ELISA and immunohistochemistry) in transgenic mice (SOD1-G93A) and the mouse motor neuron-like hybrid cell line (NSC-34), as experimental models.…”

“…Afterwards, we aimed at specifically investigating the role of the TLQP peptides in ALS, by studing their expression and changes (using ELISA and immunohistochemistry) in transgenic mice (SOD1-G93A) and the mouse motor neuron-like hybrid cell line (NSC-34), as experimental models. In parallel, we also investigated, by ELISA, ALS patients' plasma and primary fibroblast cultures, the latter being considered a good cellular model used in human ALS research (Sabatelli et al, 2015;Yang et al, 2015) and also, contain VGF (Brancia et al, 2016). Moreover, in the NSC-34 cells, the neuroprotective role of the TLQP-21 was addressed in parallel with the presence of its two known receptors (gC1q-R and C3a-R), examined by both western blot and immunocytochemistry.…”