Cultured human skin keloid fibroblasts (KFs) showed bioenergetics similar to cancer cells in generating ATP mainly from glycolysis as demonstrated by increased lactate production. Activities of hexokinase, glyceraldehyde-3-phosphate dehydrogenase, and lactate dehydrogenase were also significantly higher compared with normal fibroblasts (NFs). Inhibitors of glycolysis decreased the rate of ATP biosynthesis more significantly in KFs suggesting their reliance on glycolysis. In contrast, ATP generation in NFs was derived mainly from oxidative phosphorylation (OXPHOS), which was more compromised by mitochondrial/respiratory inhibitors. However, when fortified with excess exogenous respiratory substrates, ATP production was increased to a similar maximal level in both types of fibroblasts. In spite of this seemingly equal total capacity, ATP biosynthesis and intracellular ATP concentration were significantly higher in KFs, which further increased their ATP production when exposed to hypoxia and hypoxia-mimetics: desferrioxamine and cobalt chloride. This upregulation was again significantly compromised by glycolytic inhibitors. The rate of generation of reactive oxygen species was lower in KFs possibly due to their switch to aerobic glycolysis from mitochondrial OXPHOS. Thus, cultured skin KFs could provide a human cell model to study the de-regulation of bioenergetics of proliferative cells and their response to the HIF (hypoxia-inducible factor) signaling.

No abstract

In this study, we sought to better define the limit of spatial resolution at the fingertips of elderly participants (n = 30, age 60-95 years) using an extended set of JVP grating domes, incorporating four new grating dimensions (2.5-, 3.5-, 4.0- and 4.5-mm width). A secondary aim was to examine whether deficits in tactile acuity could be related to hand dysfunction in older adults. Spatial resolution thresholds were determined by the finest grating whose orientation (dominant index finger) could be reported reliably. Manual dexterity was assessed with the Grooved Pegboard Test (GPT). The extended set of domes improved threshold measurements in a majority of participants (21/30). Still, accurate threshold estimates could not be obtained in one third of the participants, mostly in the older age group (8/9, 74-95 years). Grating resolution thresholds at the index finger were strongly correlated (r = 0.66, p<0.01) with dexterity scores derived from the GPT. From these results, we conclude that the 2.5- and 3.5-mm grating domes are suitable additions when assessing spatial acuity at the fingertips of older subjects between 60 and 70 years of age (mean threshold, 2.7+/-0.6 mm). For the older ones, the 4.0- and 4.5-mm domes can improve threshold measurements but interpretation of values can be complicated by the presence of undiagnosed pathologies (e.g., diffuse polyneuropathy) as people advance in age. The strong relationship between grating resolution thresholds and dexterity scores indicates that an impaired spatial acuity at the fingertips may translate into great difficulties in tasks requiring fine manipulations. These findings have important implications for the assessment of hand function in older adults.

No abstract

Akt, a serine/threonine kinase has been shown to stimulate glycolysis in cancer cells but its role in mitochondrial respiration is unknown. Using PTEN-knockout mouse embryonic fibroblasts (MEFPTEN−/−) with hyper-activated Akt as a cell model, we observed a higher respiratory capacity in MEFPTEN−/− compared to the wildtype (MEFWT). The respiratory phenotype observed in MEFPTEN−/− was reproduced in MEFWT by gene silencing of PTEN which substantiated its role in regulating mitochondrial function. The increased activities of the respiratory complexes (RCs) I, III and IV were retained in the same relative proportions as those present in MEFWT, alluding to a possible co-ordinated regulation by PTEN/Akt. Using LY294002 (a PI3K inhibitor) and Akt inhibitor IV, we showed that the regulation of enzyme activities and protein expressions of the RCs was dependent on PI3K/Akt. There was insignificant difference in the protein expressions of mitochondrial transcription factor: peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) and its downstream targets, the nuclear respiratory factor 1 (NRF-1) and mitochondrial transcription factor A (mtTFA) between MEFPTEN−/− and MEFWT. Similarly, mRNA levels of the same subunits of the RCs detected in Western blots were not significantly different between MEFPTEN−/− and MEFWT suggesting that the regulation by Akt on mitochondrial function was probably not via gene transcription. On the other hand, a decrease of total 4E-BP1 with a higher expression of its phosphorylated form relative to total 4E-BP1 was found in MEFPTEN−/−, which inferred that the regulation of mitochondrial respiratory activities by Akt was in part through this protein translation pathway. Notably, gene silencing of 4E-BP1 up-regulated the protein expressions of all RCs and the action of 4E-BP1 appeared to be specific to these mitochondrial proteins. In conclusion, PTEN inactivation bestowed a bioenergetic advantage to the cells by up-regulating mitochondrial respiratory capacity through the 4E-BP1-mediated protein translation pathway.