Pyrroloquinoline quinone

The activation of AMPK by this compound is required for the regulation of mitochondrial biogenesis in rotenone-injured SH-SY5Y cells, the upregulation of PGC-1α induced by this chemical is reduced by inhibition of AMPK.[2]

SH-SY5Y human neuroblastoma cells were cultured in DMEM supplemented with 10% FBS at 37 °C in a humidified atmosphere of 95% air and 5% CO2. The day before treatment, cells were plated in a 10-cm culture dish at a density of 1 × 106 cells/mL. The cells were divided into four groups: the control group (no rotenone or PQQ), this compound group (100 μM PQQ), Rotenone group (100 μM rotenone), and this compound + Rotenone group (100 μM PQQ + 100 μM rotenone). Thirty minutes after treatment, the cells were washed once with cold PBS and scraped in urea lysis buffer (9 M sequanal-grade urea, 20 mM HEPES [pH 8.0], 1 mM β-glyceropho_x005f?sphate, 1 mM sodium vanadate, 2.5 mM sodium pyrophosphate). The cells were sonicated in lysis buffer and centrifuged for 15 min at 4 °C and 20,000 × g. Supernatants were collected and reduced with 4.5 mM DTT for 30 min at 55 °C. Reduced lysates were alkylated with iodoacetamide (0.019 g/mL of H2O) for 15 min in the dark. The samples were diluted 1:4 with 20 mM HEPES [pH 8.0] and digested with 10 g/mL trypsin-tosylphenylalanyl chloromethyl ketone in 1 mM HCl overnight. Digested peptide lysates were acidified with 1% TFA, and the peptides were desalted over 360- mg SEP PAK Classic C18 columns. Peptides were eluted with 40% acetonitrile in 0.1% TFA, dried under a vacuum, and stored at −80 °C. Differentially expressed phosphoproteins were identified and analyzed.

Pyrroloquinoline quinone dose-dependently alleviates the locomotor deficits and nigral dopaminergic neuron loss in PD mice, significantly diminishes the reduction of mitochondria number and their pathological change in the midbrain, also dose-dependently blocks rotenone-caused reduction in the expression of PGC-1α and TFAM, two key activators of mitochondrial gene transcription, in the midbrain.[2]