Alterations in Cell Motility, Proliferation, and Metabolism in Novel Models of Acquired Temozolomide Resistant Glioblastoma.

Glioblastoma (GBM) is an aggressive and incurable tumor of the brain with limited treatment options. Current first-line standard of care is the DNA alkylating agent temozolomide (TMZ), but this treatment strategy adds only ~4 months to median survival due to the rapid development of resistance. While some mechanisms of TMZ resistance have been identified, they are not fully understood. There are few effective strategies to manage therapy resistant GBM, and we lack diverse preclinical models of acquired TMZ resistance in which to test therapeutic strategies on TMZ resistant GBM. In this study, we create and characterize two new GBM cell lines resistant to TMZ in vitro, based on the 8MGBA and 42MGBA cell lines. Analysis of the TMZ resistant (TMZres) variants in conjunction with their parental, sensitive cell lines shows that acquisition of TMZ resistance is accompanied by broad phenotypic changes, including increased proliferation, migration, chromosomal aberrations, and secretion of cytosolic lipids. Importantly, each TMZ resistant model captures a different facet of the "go" (8MGBA-TMZres) or "grow" (42MGBA-TMZres) hypothesis of GBM behavior. These in vitro model systems will be important additions to the available tools for investigators seeking to define molecular mechanisms of acquired TMZ resistance.

Evaluation of Combination Drug Therapy for Treatment of Antibiotic-Resistant Inhalation Anthrax in a Murine Model.

Bacillus anthracis is considered a likely agent to be used as a bioweapon, and the use of a strain resistant to the first-line antimicrobial treatments is a concern. We determined treatment efficacies against a ciprofloxacin-resistant strain of B. anthracis (Cipr Ames) in a murine inhalational anthrax model. Ten groups of 46 BALB/c mice were exposed by inhalation to 7 to 35 times the 50% lethal dose (LD50) of B. anthracis Cipr Ames spores. Commencing at 36 h postexposure, groups were administered intraperitoneal doses of sterile water for injections (SWI) and ciprofloxacin alone (control groups), or ciprofloxacin combined with two antimicrobials, including meropenem-linezolid, meropenem-clindamycin, meropenem-rifampin, meropenem-doxycycline, penicillin-linezolid, penicillin-doxycycline, rifampin-linezolid, and rifampin-clindamycin, at appropriate dosing intervals (6 or 12 h) for the respective antibiotics. Ten mice per group were treated for 14 days and observed until day 28. The remaining animals were euthanized every 6 to 12 h, and blood, lungs, and spleens were collected for lethal factor (LF) and/or bacterial load determinations. All combination groups showed significant survival over the SWI and ciprofloxacin controls: meropenem-linezolid (P = 0.004), meropenem-clindamycin (P = 0.005), meropenem-rifampin (P = 0.012), meropenem-doxycycline (P = 0.032), penicillin-doxycycline (P = 0.012), penicillin-linezolid (P = 0.026), rifampin-linezolid (P = 0.001), and rifampin-clindamycin (P = 0.032). In controls, blood, lung, and spleen bacterial counts increased to terminal endpoints. In combination treatment groups, blood and spleen bacterial counts showed low/no colonies after 24-h treatments. The LF fell below the detection limits for all combination groups yet remained elevated in control groups. Combinations with linezolid had the greatest inhibitory effect on mean LF levels.

Acquisition of estrogen independence induces TOB1-related mechanisms supporting breast cancer cell proliferation.

Resistance to therapies targeting the estrogen pathway remains a challenge in the treatment of estrogen receptor-positive breast cancer. To address this challenge, a systems biology approach was used. A library of small interfering RNAs targeting an estrogen receptor (ER)- and aromatase-centered network identified 46 genes that are dispensable in estrogen-dependent MCF7 cells, but are selectively required for the survival of estrogen-independent MCF7-derived cells and multiple additional estrogen-independent breast cancer cell lines. Integration of this information identified a tumor suppressor gene TOB1 as a critical determinant of estrogen-independent ER-positive breast cell survival. Depletion of TOB1 selectively promoted G1 phase arrest and sensitivity to AKT and mammalian target of rapmycin (mTOR) inhibitors in estrogen-independent cells but not in estrogen-dependent cells. Phosphoproteomic profiles from reverse-phase protein array analysis supported by mRNA profiling identified a significant signaling network reprogramming by TOB1 that differed in estrogen-sensitive and estrogen-resistant cell lines. These data support a novel function for TOB1 in mediating survival of estrogen-independent breast cancers. These studies also provide evidence for combining TOB1 inhibition and AKT/mTOR inhibition as a therapeutic strategy, with potential translational significance for the management of patients with ER-positive breast cancers.

Sankofan socio-ethical reflections: the Tuskegee University National Bioethics Center's decade of operation, 1999-2009.

Primarily, this is a Sankofan socio-ethical analysis of the moral foundation of the Tuskegee University National Bioethics Center's decade of operation. The first section of the study will do the following: a) a Sankofan socio-ethical analysis of the Center's raison d'être; and b) definitions of ethical terms and the social world of the infamous syphilis study. The second section, as a result of the analysis, will address the Center's following challenges: c) the Center's challenge of theory and practice; d) the Center's challenge of moral heritage; and e) the Center's challenge of the future.

Nutritional modulation of the cell cycle and breast cancer.

In the USA, breast cancer accounts for approximately 30% of all cancers diagnosed in women and is the second leading cause of cancer death in women. An understanding of the molecular genetic events governing breast cancer lead to both prevention and intervention strategies in an attempt to reduce mortality and morbidity from breast cancer. The last three decades of medical research examining the molecular pathogenesis of cancers have provided compelling evidence for the universal disruption of the cell cycle in human tumors. The importance of cell cycle control in human cancer was recognized by the recent award of the Nobel Prize to Drs Nurse and Hartwell for their discovery of the cyclins. More recent studies have demonstrated a critical interface between hormonal signaling and the cell cycle. In parallel, epidemiological studies have identified as being associated with breast cancer important dietary and environmental components that regulate hormonal signaling. This review describes the intersection of these two fields of study, which together imply a role for dietary prevention and intervention in human breast cancer perhaps through altering cell cycle components.

Functions of the adapter protein Cas: signal convergence and the determination of cellular responses.

Since Cas was first identified as a highly phosphorylated 130 kilodalton protein that associated with the v-Src and v-Crk-oncoproteins, considerable effort has been made to determine its function. Its predicted role as a scaffolding molecule based on its domain structure has been largely confirmed. Through its ability to undergo rapid changes in phosphorylation, subcellular localization and association with heterologous proteins, Cas may spatially and temporally regulate the function of its binding partners. Numerous proteins have been identified that bind to Cas in vitro and/or in vivo, but in only a few cases is there an understanding of how Cas may function in these protein complexes. To date, Cas-Crk and Cas-Src complexes have been most frequently implicated in Cas function, particularly in regards to processes involving regulation of the actin cytoskeleton and proliferation. These and other Cas protein complexes contribute to the critical role of Cas in cell adhesion, migration, proliferation and survival of normal cycling cells. However, under conditions in which these processes are deregulated, Cas appears to play a role in oncogenic transformation and perhaps metastasis. Therefore, in its capacity as an adapter protein, Cas serves as a point of convergence for many distinct signaling inputs, ultimately contributing to the generation of specific cellular responses.

A longitudinal study of otitis media with effusion among 2- to 5-year-old African-American children in child care.

OBJECTIVE: To prospectively document the prevalence of otitis media with effusion (OME) in 86 African-American children between ages 2 and 5 years. STUDY DESIGN: Eighty-six children in center-based child care whose ear status had been followed from infancy continued to be observed. Middle ear status was assessed by pneumatic otoscopy and tympanometry biweekly. RESULTS: The prevalence of OME decreased as children became older. The mean proportion of examinations demonstrating bilateral OME (BOME) ranged from 12% between 24 to 30 months to 4% between 54 to 60 months of age. The mean proportion of exams revealing bilateral normal ears increased from 77% at 24 to 30 months to 88% at 54 to 60 months of age. Although 60 children had experienced BOME that lasted 4 months or longer in the 6- to 24-month age period, only 8 of these children experienced at least 4 months of continuous BOME between 24 to 60 months. CONCLUSIONS: The proportion of time with BOME decreased progressively with increasing age in this population. Only 8 of 60 children who had experienced more than 4 consecutive months of BOME before 2 years of age continued to manifest persistent effusion or experience recurrences of prolonged BOME after 2 years of age.

Fracture resistance of four different restorations for cuspal replacement.

The purpose of this study was to compare the fracture resistance of four posterior restorations involving an entire cusp replacement. Four groups were established, each containing eight caries-free mandibular molars, similar in size and anatomic form. A mesio-occlusal preparation including the lingual cusp was prepared on all teeth. Group A were restored with a pin-retained amalgam. Group B were restored with amalgam and a meta adhesive. Group C were restored with a composite resin with a beta-glass quartz insert. Group D were restored with composite resin and a HEMA adhesive. All specimens were mounted in acrylic and stored in artificial saliva for 30 days. Each specimen was loaded in compression at a 90 degrees angle in an Instron testing machine with a crosshead speed of 0.5 cm/min. Results demonstrated the mean (SD) failure loads in kilograms for each group to be: A, 1155 (388); B, 1232 (436); C, 1345 (375); D, 1595 (373). Analysis of variance indicated no significant difference among groups at P<0.05. Although the values for the composite resin restoration with the adhesive were higher than the other restorative techniques.

Prospective surveillance for otitis media with effusion among black infants in group child care.

OBJECTIVE: To document the prevalence of otitis media with effusion (OME) in 102 black children observed prospectively between 6 and 24 months of age. METHODS: Study children attended nine different center-based child care facilities. Middle ear status was assessed by pneumatic otoscopy and tympanometry every 2 weeks. RESULTS: All children, except one, had OME during the period of observation. The proportion of child-examinations revealing bilateral OME ranged from 76% between 6 and 12 months of age to 30% between 21 and 24 months of age. Effusions were considered purulent in only 13% of examinations revealing middle ear fluid. The mean incidence of purulent OME was 2.13 episodes per child per year. Sixty-six children had at least 4 months of continuous bilateral OME during the period of observation; 57 were followed without placement of tympanostomy tubes. Bilateral OME had resolved before the second birthday in 95% of these children, and within 3 months of achieving the 4-month criterion in 50% of subjects. CONCLUSIONS: Persistent bilateral OME occurs commonly between 6 and 18 months of age in infants who enter group child care during the first year of life. In this study, spontaneous resolution of bilateral effusion by 2 years of age was typical.

Influence of internal fixation on wound infections.

With increasing frequency trauma surgeons are advocating early internal fixation in open fractures. The effect of the fixation devices on the infection rate in contaminated wounds remains a concern as our clinical experience in this area has been mixed. To study the effects of internal fixation on bone infections a 3.5-mm stainless steel screw was inserted into rabbit femurs and the wounds contaminated with Staphylococcus aureus. The controls had the screw hole drilled and taped but the screw was not inserted. Thirty of 49 rabbits receiving the screw subsequently became infected whereas 19 of 56 control animals developed an infection. The difference was significant at the 0.05 confidence level.

Perioperative ischemic injury after coronary bypass graft surgery.

Two hundred twelve patients who underwent isolated coronary bypass graft surgery were prospectively evaluated for perioperative ischemic injury. All patients underwent preoperative and postoperative testing with technetium 99m pyrophosphate first-pass ventriculography combined with myocardial uptake scans, 12-lead electrocardiography, and serial creatinine phosphokinase MB determination. Fifteen percent of the patients had ischemic injury with at least two test results positive, but only 4 percent had positive results of all three tests. No single test proved adequate. Enzyme levels were highly sensitive and had value as a screening test. The electrocardiogram was specific but only moderately sensitive. The single best test was the radionuclide scan with good sensitivity and no false-positive results. All three tests are required to rigorously diagnose ischemic injury.

Weight bearing: its lack of effect on fracture healing.

The load transmitted through fresh fractures of the tarsometatarsus of adult chickens was varied by 40% more, or less, than the body weight to determine the effect of load-bearing on fracture healing. The healing fracture was examined for size and strength at 14 days, but no significant differences were detected in any group. The technique was sensitive enough to detect the differences between healing fractures at 7, 10 and 14 days. In this experiment load transmission was not found to be an important factor in fracture healing.

Hip dislocations in myelodysplasia: a functional assessment.

We reviewed the records of 281 patients with myelodysplasia to ascertain the effect of hip dislocation on their ability to walk. There was no difference in the prevalence of myelodysplasia by gender. When adjusted for neurologic level and patient age, no statistical differences were found in the methods of locomotion between patients with hip dislocations and those without. Data (from this report and other published investigations) show that treatment for hip dislocation in myelodysplasia is unnecessary and fraught with many complications.

Treatment of open fractures: a prospective study.

A double-blind prospective study was done to assess the benefit of delaying closure of the wounds associated with open fractures. An additional double-blind study compared the effectiveness of clindamycin versus cefazolin for prophylactic antibiotic coverage. Quantitative cultures of the wounds were accomplished at the time of debridement and again at the time of closure if the wound was not closed initially. Almost half of the wounds were contaminated (46%) at the time of debridement, although the incidence of wound infection was low (6.5%). Gram-negative organisms resistant to the prophylactic antibiotic were recovered initially only eight times, but four of these (50%) became infected. The contaminating organisms in each case were present in high concentration (greater than 10(5) CFU/gm of tissue) at initial culture. The time of wound closure, cefazolin versus clindamycin, and internal fixation of the fracture were not followed by significant differences in the development of clinical infection in this series.

Mechanical properties of the tibia from chickens with idiopathic scoliosis.

The tibia from six-week old chickens that develop idiopathic scoliosis were studied with stress relaxation experiments and torsional strength testing. Most parameters observed did not show any significant differences between tibias obtained from chickens with scoliosis and tibias from the control birds; however, the rate of stress relaxation of the tibia from the birds with scoliosis was minimally increased over the controls. There were no significant differences noted in ultimate torsional strength, maximum angular deformity or modulae of torsional rigidity of the tibias from scoliotic chickens when compared to tibias from control chickens.