Biguanide-induced mitochondrial dysfunction yields increased lactate production and cytotoxicity of aerobically-poised HepG2 cells and human hepatocytes in vitro

As a class, the biguanides induce lactic acidosis, a hallmark of mitochondrial impairment. To assess potential mitochondrial impairment, we evaluated the effects of metformin, buformin and phenformin on: 1) viability of HepG2 cells grown in galactose, 2) respiration by isolated mitochondria, 3) metabolic poise of HepG2 and primary human hepatocytes, 4) activities of immunocaptured respiratory complexes, and 5) mitochondrial membrane potential and redox status in primary human hepatocytes. Phenformin was the most cytotoxic of the three with buformin showing moderate toxicity, and metformin toxicity only at mM concentrations. Importantly, HepG2 cells grown in galactose are markedly more susceptible to biguanide toxicity compared to cells grown in glucose, indicating mitochondrial toxicity as a primary mode of action. The same rank order of potency was observed for isolated mitochondrial respiration where preincubation (40 min) exacerbated respiratory impairment, and was required to reveal inhibition by metformin, suggesting intramitochondrial bio-accumulation. Metabolic profiling of intact cells corroborated respiratory inhibition, but also revealed compensatory increases in lactate production from accelerated glycolysis. High (mM) concentrations of the drugs were needed to inhibit immunocaptured respiratory complexes, supporting the contention that bioaccumulation is involved. The same rank order was found when monitoring mitochondrial membrane potential, ROS production, and glutathione levels in primary human hepatocytes. In toto, these data indicate that biguanide-induced lactic acidosis can be attributed to acceleration of glycolysis in response to mitochondrial impairment. Indeed, the desired clinical outcome, viz., decreased blood glucose, could be due to increased glucose uptake and glycolytic flux in response to drug-induced mitochondrial dysfunction.     

Lipopolysaccharide from the commensal microbiota of the breast enhances cancer growth: role of S100A7 and TLR4

The role of commensal bacterial microbiota in the pathogenesis of human malignancies has been a research field of incomparable progress in recent years. Although breast tissue is commonly assumed to be sterile, recent studies suggest that human breast tissue may contain a bacterial microbiota. In this study, we used an immune‐competent orthotopic breast cancer mouse model to explore the existence of a unique and independent bacterial microbiota in breast tumors. We observed some similarities in breast cancer microbiota with skin; however, breast tumor microbiota was mainly enriched with Gram‐negative bacteria, serving as a primary source of lipopolysaccharide (LPS). In addition, dextran sulfate sodium (DSS) treatment in late‐stage tumor lesions increased LPS levels in the breast tissue environment. We also discovered an increased expression of S100A7 and low level of TLR4 in late‐stage tumors with or without DSS as compared to early‐stage tumor lesions. The treatment of breast cancer cells with LPS increased the expression of S100A7 in breast cancer cells in vitro. Furthermore, S100A7 overexpression downregulated TLR4 and upregulated RAGE expression in breast cancer cells. Analysis of human breast cancer samples also highlighted the inverse correlation between S100A7 and TLR4 expression. Overall, these findings suggest that the commensal microbiota of breast tissue may enhance breast tumor burden through a novel LPS/S100A7/TLR4/RAGE signaling axis.     

Mechanical environment of the supraspinatus tendon: three-dimensional finite element model analysis

BACKGROUND: We analyzed the mechanical environment of the supraspinatus tendon using a three-dimensional finite element model with the software programs MENTAT and MARC. METHODS: The supraspinatus tendon that attaches to the superior facet was extracted and modeled. The geometric shape of the humeral head was determined from computed tomography images, and the shape of the supraspinatus tendon was determined from magnetic resonance images of the shoulder at 0 degrees of abduction in a healthy 27-year-old man. The distal portion of the humeral head was fixed, and 10 N of tensile force was applied to the proximal end of the tendon. The tensile stress was calculated. RESULTS: The tensile stress was 1.8 MPa for the bursal side and 15.0 MPa for the articular side of the anterior portion of the supraspinatus tendon. The intensity was 0 MPa for the bursal side and 4.5 MPa for the articular side of the middle portion of the tendon. The intensity was 0.1 MPa for the bursal side and 5.2 MPa for the posterior edge of the tendon. CONCLUSIONS: Based on the three-dimensional finite element method, the maximal tensile stress was observed on the articular side of the anterior edge of the supraspinatus tendon. Our result may explain the frequent occurrence of rotator cuff tears at this site.