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  • Morpho‑functional study of the hypothalamic proline‑rich polypeptide apoptotic activity against mouse Ehrlich ascites carcinoma.

Morpho‑functional study of the hypothalamic proline‑rich polypeptide apoptotic activity against mouse Ehrlich ascites carcinoma.

Oncology reports (2020-05-08)
S Abrahamyan, I Sahakyan, N Tumasyan, N Kocharyan, A Simonyan, R Aroutiounian, G Chailyan, S Chailyan, T Davtyan, K Galoian
ABSTRACT

A new type of bioactive polypeptides of the neurosecretory hypothalamus called proline‑rich peptides (PRPs), which are isolated from bovine neurosecretory granules of the neurohypophysis, are synthesized in the form of a common precursor protein (neurophysin vasopressin‑associated glycoprotein). Proline‑rich polypetide 1 (PRP‑1; also known as galarmin) is comprised of 15 amino acids residues, and has been suggested to possess anti‑neurodegenerative, immunoregulatory, hematopoietic, antimicrobial and antitumor properties. The cytostatic, antiproliferative effect of PRP‑1 was demonstrated in the human chondrosarcoma JJ012 and triple negative breast carcinoma MDA MB 231 cell lines. PRP‑1 action is disease and tissue specific. To further explore the antitumorigenic and possible cytotoxic effects of PRP‑1, a morpho‑functional study on the effect of PRP‑1 on a mouse Ehrlich ascites carcinoma (EAC) model was conducted. The PRP‑1‑induced morphological features of EAC cells confirmed the apoptotic nature of PRP‑1, as manifested by cell shrinkage, membrane blebbing, chromosome condensation (pyknosis) and nuclear fragmentation (karyorrhexis). The effect of PRP‑1 on the number of tumor cells incubated for 24 h and their viability in trypan blue‑stained samples lead to a 44% reduction in the number of viable cells on day 11 post‑inoculation vs. 22% inhibition of viable cells after PRP‑1 treatment (0.1 µg/ml) on day 7 post‑inoculation. Apoptosis experiments using an Annexin V‑cyanine 3 apoptosis detection kit indicated that 24 h incubation with 0.1 µg/ml PRP‑1 caused a significant increase in the number of apoptotic cells, reaching 50.33%, compared to 8.33% in the sample control on day 7 post‑inoculation.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
DPX Mountant for histology, slide mounting medium
Sigma-Aldrich
Freund′s Adjuvant, Complete, cell suspension
Sigma-Aldrich
Bovine Serum Albumin, lyophilized powder, crystallized, ≥98.0% (GE)
SAFC
Hanks′ Balanced Salt solution, HBSS Modified, with calcium, with magnesium, without phenol red, liquid, suitable for cell culture
Sigma-Aldrich
3,3′-Diaminobenzidine tetrahydrochloride, tablet, 10 mg substrate per tablet
Sigma-Aldrich
Annexin V-Cy3 Apoptosis Detection Kit
Sigma-Aldrich
p-Xylene-bis(N-pyridinium bromide), ≥95% (TLC)