Correlation between radiographic and arthroscopic findings in Asian osteoarthritic knees.

PURPOSE: To evaluate the correlation between radiographic and arthroscopic findings for severity of knee osteoarthritis (OA) in 119 Asians. METHODS: Medical records of 52 men and 67 women aged 33 to 70 (mean, 51) years who had complaints of chronic mechanical knee pain for >3 months and were arthroscopically diagnosed with articular cartilage degeneration of the knee were reviewed. The severity of knee OA was classified using the Kellgren-Lawrence (K&L) scale and the modified Outerbridge classification. The highest grade in any compartment of the knee was used to represent the severity of OA. The correlation between the radiographic and arthroscopic findings for severity of knee OA was evaluated. RESULTS: 17 of the 119 patients with no radiographic evidence of OA (K&L grade 0) had articular cartilage degeneration arthroscopically (mean grade, 1.14). In general, higher K&L grades correlated with more severe articular cartilage degeneration. The Pearson correlation coefficient was 0.32, indicating that the association between the radiographic and arthroscopic findings for severity of knee OA was weak. CONCLUSION: The K&L scale correlated poorly with arthroscopic findings of articular cartilage degeneration in an Asian population with knee OA.

Ablation of central nervous system progenitor cells in transgenic rats using bacterial nitroreductase system.

Specific ablation of central nervous system (CNS) progenitor cells in the brain of live animals is a powerful method to determine the functions of these cells and to reveal novel avenues for the treatment of several CNS-related disorders. To achieve this goal, we generated a line of transgenic rats expressing a bacterial enzyme, Escherichia coli nitroreductase gene (NTR), under control of the nestin promoter. In this system, NTR(+) cells are selectively eliminated upon application of prodrug CB1954, through activation of programmed cell death machineries. At 5 days of age, which is a time when cerebellar development is occurring, transgenic rats bearing the nestin-NTR/green fluorescent protein (GFP) gene are overtly normal and express NTR/GFP in neuronal stem cells, without any toxicity in these cells. The functional consequence of progenitor cell ablation was demonstrated by administering prodrug CB1954 into the cerebellum at this 5-day time point. Stem cell ablation in these neonates resulted in sensorimotor abnormalities, cerebellar degeneration, overall reduction in cerebellar seize, and manifestation of ataxia. In adult rats, GFP expression was not seen in the hippocampal progenitor cells and seen only at very low levels in the lateral ventricles, indicating a different NTR/GFP expression pattern between neonates and adults. In addition, application of CB1954 by intraventricular delivery reduced the number of 5-bromo-2'-deoxyuridine-labeled proliferating cells in the lateral ventricle but not hippocampus of NTR/GFP rats. These findings shows that targeted expression of NTR under a specific promoter might be of significant value in addressing the function of distinct cell population in vivo.

Disruption of ShcA signaling halts cell proliferation--characterization of ShcC residues that influence signaling pathways using yeast.

Shc adapter proteins are thought to regulate cellular proliferation, differentiation and apoptosis by activating the SOS-Grb2-RAS-MAPK signaling cascade. Using the small hairpin RNA (shRNA) technique, we found that decreasing ShcA mRNA reduced the proliferative ability of HEK293 mammalian culture cells. We then recapitulated phosphorylation-dependent Shc-Grb2 complex formation in Saccharomyces cerevisiae. Immunoprecipitation followed by Western analysis demonstrated that activated TrkB, composed of the intracellular domain of TrkB fused to glutathione S-transferase (GST-TrkB(ICD)), promoted the association of ShcC and Grb2 in yeast. The Ras-recruitment system (RRS), in which a myristoylated (Myr)-bait and son of sevenless (hSOS)-prey are brought together to complement the defective Ras-cAMP pathway in a thermosensitive cdc25H mutant yeast strain, was used to validate a phenotypic assay. Yeast cells transformed with both Myr-ShcC and hSOS-Grb2 (referred to as scheme 1) or Myr-Grb2 and hSOS-ShcC (scheme 2) did not grow at non-permissive temperature; the additional transformation of GST-TrkB(ICD) enabled growth. GST-TrkB(ICD) also enabled growth with hSOS-Grb2 and either Myr-ShcA or Myr-SHP2. Mutational analysis of TrkB showed that its kinase activity was essential for complementation, while its docking site for Shc proteins was not. Mutational analysis of ShcC showed that the PTB and SH2 domains were not essential for complementation but phosphorylation at Y304 in the CH1 domain was. Phosphorylation at Y304 could not be substituted by an acidic amino acid. The RRS provides a genetic system to probe Shc proteins and potentially identify member specific protein partners and pharmacological reagents.

An age-related increase in resistance to DNA damage-induced apoptotic cell death is associated with development of DNA repair mechanisms.

Neurons in the developing brain die via apoptosis after DNA damage, while neurons in the adult brain are generally resistant to these insults. The basis for this resistance is a matter of conjecture. We report here that cerebellar granule neurons (CGNs) in culture lose their competence to die in response to DNA damage as a function of time in culture. CGNs at either 1 day in vitro (DIV) or 7 DIV were treated with the DNA damaging agents camptothecin, UV or gamma-irradiation and neuronal survival measured. The younger neurons were effectively killed by these agents, while the older neurons displayed a significant resistance to killing. Neuronal survival did not change with time in culture when cells were treated with C2-ceramide or staurosporine, agents which do not target DNA. The resistance to UV irradiation developed over time in culture and was not due to changes in mitotic rate. Increases in DNA strand breakage, up-regulation of the levels of both p53 and its phosphorylated form and nuclear translocation of p53 were equivalent in both older and younger neurons, indicating a comparable p53 stress response. In addition, we show that treatment of older neurons with pharmacological inhibitors of distinct components of the DNA repair machinery promotes the accumulation of DNA damage and sensitizes these cells to the toxic effects of UV exposure. These data demonstrate that older neurons appear to be more proficient in DNA repair in comparison to their younger counterparts, and that this leads to increased survival after DNA damage.