Hepes sodium salt

Once the docking studies yielded results that allowed us to select four compounds for experimental studies, we evaluated their effect on Aβ_1–42 pathological aggregation. For this purpose, lyophilized wild-type human Aβ_1–42 peptide (chloride salt) was purchased from Calbiochem (Mexico). HEPES sodium salt (>99.5% purity) and ThT were obtained from Sigma–Aldrich (Mexico). A freshly prepared Aβ_1–42 solution (50 μM in fresh MilliQ water) was incubated alone and in the presence of the selected compounds (100 μM) [26 (link)]. The samples were incubated at 37°C in a 0.5-cm path length quartz cell and stirred at 250 rpm. The increase in ThT fluorescence [31 (link)] was measured using a Perkin–Elmer LS-55 fluorescence spectrophotometer equipped with a water-jacketed cell holder for temperature control. The emission and excitation wavelengths were 445 and 480 nm, respectively. The fluorescence emission recordings were made using path-length quartz cuvettes. The buffer solution was 20 mM HEPES and 100 mM NaCl, pH 7.4, containing 3.3 μM ThT [26 (link)]. The inhibition of Aβ_1–42 pathological aggregation for compounds 5, 8, 14 and 19 was calculated after 24 h of incubation.

Deionized water Milli-Q (conductivity < 0.1 μS cm⁻¹) (Millipore system, Molsheim, France) and analytical grade chemicals were used without further purification unless otherwise stated. PSU in pellets with average molecular weight (MW) 35,000 Da (CAS Number 25135-51-7); 1-methyl-2-pyrrolidone (NMP) (CAS Number 872-50-4); Polyvinylpyrrolidone (PVP) K30 (CAS Number 9003-39-8); HEPES sodium salt (CAS Number 75277-39-3); phorbol-12-myristate-13-acetate (PMA) (CAS Number 16561-29-8); acetonitrile (CAS Number 75-05-8); Elastase, Human Neutrophil (CAS 9004-06-2); Elastase Substrate V, Fluorogenic (CAS 72252-90-5); and single-side polished N-type silicon wafer (CAS Number 7440-21-3) were all purchased from Sigma-Aldrich Co., Saint Louis, MO, USA. Sivelestat, neutrophil elastase inhibitor (CAS Number 127373-66-4), and Human PMN Elastase ELISA Kit were acquired from Abcam, Cambridge, UK. Dimethyl sulfoxide (DMSO) (CAS Number 127373-66-4) was purchased from VWR Chemicals, Radnor, PA, USA. Monoclonal antibodies CD42a (GRP-P), PE, and CD62P (P-Selectin), and APC, used for the determination of platelet activation, were purchased from Invitrogen, ThermoFisher Scientific, Waltham, MA, USA.

From Sigma-Aldrich (St. Louis, MO, USA): Adenosine 5′-triphosphate disodium salt hydrate (ATP, A1852), DMEM (D6429), Hanks′ Balanced Salt Solution (HBSS, H4641), HEPES sodium salt (H7006), Monensin (M5273), Nystatin (N6261), Amiloride (PHR1839), Xestospongin C (X2628), MRS-2179 (M3808), U73122 (U6756), Thapsigargin (T9033), Ryanodine (559276), Carbenoxolone (C4790), ¹⁰Panx1 (SML2152), Bafilomycin A1 (19-148), Apyrase (A6237), Gap26 (SML3074). From Molecular probes (Eugene, OR, USA): Hoechst 33,342 (H1399), Propidium iodide (P1304MP). From Evrogene (Moscow, Russia): MMLV reverse transcriptase (SK022S), SYBR Green I PCR Master Mix (PK147L); Triethylammonium salt (TNP-ATP, Ann Arbor, MI, USA, 20902). From Thermo Fisher Scientific (Waltham, MA, USA): Fura-2AM (Cat. #F1221), Fetal Bovine Serum (10099141). Selenium nanoparticles (kindly provided by Dr. S.V. Gudkov, Prokhorov General Physics Institute of the Russian Academy of Sciences, Moscow, Russia).