Centro de Biología Molecular Severo Ochoa

The viability and tolerance of individual ureolytic bacteria are a bottleneck in the remediation of cadmium (Cd) by microbially induced carbonate precipitation (MICP) technology. To solve this issue, strains of Bacillus thuringiensis (B. thuringiensis, BT) and Citrobacter freundii (C. freundii, CF) were isolated from soil and studied for their growth characteristics and metabolism. A cooperation system (BT+CF, 1:1, v/v) was constructed and exposed to 20 mg/kg Cd^2 + for 7 days, compared with individual bacteria. The synergistic mechanism of strains that immobilize Cd²⁺ was explored using characterization techniques. Results showed that the main metabolic pathways leading to urea up-regulation were pyrimidine metabolism, urea cycle, and lysine degradation by metabolomic analysis. The cooperation system can effectively remove Cd²⁺ with an efficiency of 97.68 %, which is higher than BT (66.66 %) and CF (88.61 %). The SEM-EDS, TEM, and XPS results revealed that the calcium carbonate polycrystals (vaterite and calcite) were formed during the MICP process, and the XRD and FTIR confirmed that the BT+CF produces more stable carbonate crystals. The BT+CF cooperation system was efficient at immobilizing Cd²⁺ by synergizing the molecular mechanisms of ureolytic bacteria. These results provide a novel perspective for the application of MICP.

Sorafenib is a tyrosine kinase inhibitor (TKI) that belongs to the landscape of treatments for advanced stages of hepatocellular carcinoma (HCC). The induction of cell death and cell cycle arrest by Sorafenib has been associated with mitochondrial dysfunction in liver cancer cells. Our research aim was to decipher underlying oxidative and nitrosative stress induced by Sorafenib leading to mitochondrial dysfunction in liver cancer cells.

MnTBAP, catalase and the scavenger of peroxynitrite FeTPPs were administered to Sorafenib (0-10 μM)-treated HepG2 cells. Oxygen consumption and glycolytic flux were determined in cultured cells. Mitochondrial complex activities were measured in mitochondrial fraction and cell lysates. The protein and mRNA expression of subunits of electron transport chain (ETC) were assessed by immunoblot and RNA-seq.

Sorafenib (10 μM) increased nitric oxide (NO) and superoxide anion (O₂^.-) leading to peroxynitrite generation, and drastically reduced oxygen consumption. Moreover, Sorafenib led to mitochondrial network disorganization and loss of membrane potential. The administration of FeTPPs influenced the recovery of mitochondrial network and oxygen consumption, as well as associated ATP production. Sorafenib downregulated the mRNA expression of all mitochondrial-encoded subunits of ETC and, at to a lesser extent, nuclear-encoded mitochondrial genes. The protein expression of complex I, complex III and complex IV was greatly affected by Sorafenib. Furthermore, Sorafenib diminished the activity of complex I in in-gel assays, whose expression and activity were restored by FeTPPs. However, Sorafenib did not affect the assembly of mitochondrial supercomplexes. Sorafenib altered glycolysis and reduced Krebs cycle intermediates and increased NAD/NADH ratio.

The induction of cell death by Sorafenib was associated with peroxynitrite generation, which impacted the expression of ETC subunits and mitochondrial functionality in liver cancer cells.