Selective loss of [3H]kainic acid and [3H]AMPA binding in layer VI of frontal cortex in Huntington's disease

Excitatory amino acid neurotoxicity has been proposed to cause the neostriatal neuronal degeneration of Huntington's disease (HD); N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), and kainate receptors have been hypothesized to play important roles in this process. We have recently reported a loss of neurons in layer VI of the cerebral cortex in HD. Using quantitative autoradiographic methods, we have now measured NMDA, AMPA, and kainate receptor binding in the frontal cerebral cortex of the brains of controls and individuals with HD. We find no change in NMDA receptor binding but a selective decrease in kainate and AMPA receptor binding in layer VI. These data suggest that cerebral cortical neurons possessing kainate or AMPA receptors may be selectively vulnerable in individuals with HD.     

Comprehensive analysis of risk factors for acquisition of Pseudomonas aeruginosa in young children with cystic fibrosis

Racial differences in insulin secretion and insulin sensitivity in healthy children were studied by administering a 2-hour hyperglycemic clamp (225 mg/dL) to 14 black and 16 white healthy adolescents (Tanner II-V), and 12 black and 11 white prepubertal children, matched for age, body mass index, and Tanner I pubertal development. In prepubertal children, fasting and first-phase insulin concentrations were higher in blacks compared with whites (14.7+/-1.3 vs 10.4+/-1.2, P=0.02, and 76.9+/-6.8 vs 52.1+/-6.4 microu/mL, P=0.016). There were no differences in second-phase insulin levels and insulin sensitivity index. In pubertal adolescents, first-phase and second-phase insulin concentrations were higher in blacks compared with whites (first-phase: 157.3+/-18.3 vs 77.0+/-8.7 microu/mL, P=0.0003; second-phase: 175.0+/-24.3 vs 108.7+/-8.8 microu/mL, P=0.012). Insulin sensitivity index was 35% lower in black adolescents compared with whites (P=0.02). These findings indicate that significant differences in insulin secretion and sensitivity are detectable early in childhood in healthy African-American vs American whites. However, genetic (race) vs environmental factors (physical activity/fitness, energy balance) should be carefully scrutinized as potential factors responsible for such differences.     

Anion currents and predicted glutamate flux through a neuronal glutamate transporter

Kinetic properties of a native, neuronal glutamate transporter were studied by using rapid applications of glutamate to outside-out patches excised from Purkinje neurons. Pulses of glutamate activated anion currents associated with the transporter that were weakly antagonized by the transporter antagonist kainate. In addition, kainate blocked a resting anion conductance observed in the absence of glutamate. Transporter currents in response to glutamate concentration jumps under a variety of conditions were used to construct a cyclic kinetic model of the transporter. The model simulates both the anion conductance and the glutamate flux through the transporter, thereby permitting several predictions regarding the dynamics of glutamate transport at the synapse. For example, the concentration-dependent binding rate of glutamate to the transporter is high, similar to binding rates suggested for ligand-gated glutamate receptors. At saturating glutamate concentrations, transporters cycle at a steady-state rate of 13/sec. Transporters are predicted to have a high efficiency; once bound, a glutamate molecule is more likely to be transported than to unbind. Physiological concentrations of internal sodium and glutamate significantly slow net transport. Finally, a fixed proportion of anion and glutamate flux is expected over a wide range of circumstances, providing theoretical support for using net charge flux to estimate the amount and time course of glutamate transport.     

Haemosuccus pancreaticus as a rare initial manifestation of chronic pancreatitis

Twenty-seven painful knee replacements were evaluated arthroscopically. The diagnostic and therapeutic value of these arthroscopic procedures was studied retrospectively. In 5 of the 27 cases, the arthroscopy revealed no diagnosis for the pain. Some form of arthroscopic treatment was performed in 20 cases; in 6 of these 20 cases, however, the treatment did not reduce the pain. Based on these findings, we conclude that the indications for arthroscopic evaluation and treatment of painful knee prostheses are limited.     

Ex vivo expansion of genetically marked rhesus peripheral blood progenitor cells results in diminished long-term repopulating ability

The possibility of primitive hematopoietic cell ex vivo expansion is of interest for both gene therapy and transplantation applications. The engraftment of autologous rhesus peripheral blood (PB) progenitors expanded 10 to 14 days were tracked in vivo using genetic marking. Stem cell factor (SCF)/granulocyte colony-stimulating factor (G-CSF)-mobilized and CD34-enriched PB cells were divided into two equal aliquots and transduced with one of two retroviral vectors carrying the neomycin-resistance gene (neo) for 4 days in the presence of interleukin-3 (IL-3), IL-6, and SCF in the first 5 animals, IL-3/IL-6/SCF/Flt-3 ligand (FLT) in 2 subsequent animals, or IL-3/IL-6/SCF/FLT plus an autologous stromal monolayer (STR) in the final 2. At the end of transduction period, one aliquot (nonexpanded) from each animal was frozen, whereas the other was expanded under the same conditions but without vector for a total of 14 days before freezing. After total body irradiation, both the nonexpanded and expanded transduced cells were reinfused. Despite 5- to 13-fold higher cell and colony-forming unit (CFU) doses from the expanded fraction of marked cells, there was greater short- and long-term marking from the nonexpanded cells in all animals. In animals receiving cells transduced and expanded in the presence of IL-3/IL-6/SCF/FLT, engraftment by the marked expanded cells was further diminished. This discrepancy was even more pronounced in the animals who received cells transduced and expanded in the presence of FLT and autologous stroma, with no marking detectable from the expanded cells. Despite lack of evidence for expansion of engrafting cells, we found that the addition of FLT and especially STR during the initial brief transduction period increased engraftment with marked cells into a clinically relevant range. Levels of marked progeny cells originating from the nonexpanded aliqouts were significantly higher than that seen in previous 4 animals receiving cells transduced in the presence of IL-3/IL-6/SCF, with levels of 10% to 20% confirmed by Southern blotting from the nonexpanded IL-3/IL-6/SCF/FLT/STR graft compared with 0.01% in the original IL-3/IL-6/SCF cohort. These results suggest that, although expansion of PB progenitors is feasible ex vivo, their contribution towards both short- and long-term engraftment is markedly impaired. However, a brief transduction in the presence of specific cytokines and stromal support allows engraftment with an encouraging number of retrovirally modified cells.     

The Fanconi anemia complementation group C gene (FAC) suppresses transformation of mutant fibroblasts by the SV40 virus

Fanconi anemia (FA) is a heterogeneous genetic syndrome manifested by bone marrow failure and consisting of at least five complementation groups (A, B, C, D, E). Mutations in a gene termed FAC are responsible for the C complementation group, but the function of the FAC protein remains obscure. FA patients are also highly cancer-prone; the molecular basis for this susceptibility is unclear but has led to the hypothesis that the wild-type FA gene may act as a tumor suppressor. In vitro, mutant FA primary fibroblasts are 3- to 50-fold more sensitive than normal fibroblasts to transformation in culture by the SV40 virus. We confirmed this marked susceptibility to transformation of a FAC-mutant primary fibroblast cell line, GM449. We then introduced a copy of the wild-type FAC cDNA into GM449 cells using a recombinant adeno-associated virus (rAAV) vector. We found that GM449 cells transduced with a copy of the normal FAC cDNA by a FAC-rAAV vector were at least 10-fold less prone to form transformed foci. Diminished transformation potential of transduced cells was a specific effect of the FAC cDNA since GM449 cells transduced with a rAAV vector not containing FAC retained marked susceptibility to SV40 transformation.     

Estrogen replacement therapy and worsening of radiographic knee osteoarthritis: the Framingham Study

OBJECTIVE: To examine whether estrogen replacement therapy (ERT) prevents worsening of radiographic knee osteoarthritis (OA) in elderly women. METHODS: A total of 551 women ages 63-91 years (mean age 71) in the Framingham Study were followed up from biennial examination 18 (1983-1985) to examination 22 (1992-1993). Data on postmenopausal ERT were obtained every 2 years. Subjects were classified into 3 groups according to their estrogen use at biennial examination 18: never users (n = 349), past users (n = 162), and current users (n = 40). Women received anteroposterior weight-bearing knee radiographs at examinations 18 and 22. Using the Kellgren and Lawrence criteria, global radiographic knee OA was assessed, (grade range 0-4) and individual radiographic features, such as osteophytes and joint space narrowing, were scored from 0 to 3. Worsening was defined as either development of radiographic OA that was not present at baseline (incident OA) or progression of baseline radiographic OA by > or =1 Kellgren and Lawrence grade (progressive OA). Potential confounding factors included age, body mass index, weight change, smoking, knee injury, physical activity level, and bone mineral density at the femoral neck. RESULTS: During 8 years of followup, 17.4% of knee radiographic scores worsened by 1 grade and 5.8% by 2 or 3 grades among never users of ERT. Among current estrogen users, only 11.7% of knee radiographic scores worsened by 1 grade and none worsened by more than 1 grade. After adjusting for age and other potential confounding factors, the relative risk of incident radiographic knee OA in comparison with never users of estrogen was 0.8 (95% confidence interval [95% CI] 0.5-1.4) in past users and 0.4 (95% CI 0.1-3.0) in current users. Current use of estrogen also showed a trend toward decreased risk of progressive knee OA compared with never use (odds ratio [OR] 0.5, 95% CI 0.1-2.9). When both incident and progressive radiographic knee OA cases were combined, current ERT use had a 60% decreased risk compared with never use (OR 0.4, 95% CI 0.1-1.5). CONCLUSION: This is the first prospective cohort study to examine the effects of ERT on radiographic knee OA. The results indicate that current use of ERT had a moderate, but not statistically significant, protective effect against worsening of radiographic knee OA among elderly white women. These findings corroborate those of cross-sectional studies and point further to a potential benefit of female hormones in OA.