Lethality-based selection of recombinant genes in mammalian cells: application to identifying tumor antigens.

Many biological processes result in either cell death or cessation of cell growth. However, plasmid- and retrovirus-based mammalian expression vectors in which it has been possible to construct representative cDNA libraries cannot be readily recovered from cells that are not actively dividing. This has limited the efficiency of selection of recombinant genes that mediate either lytic events or growth arrest. Examples include genes that encode the target antigens of cytotoxic T cells, genes that promote stem-cell differentiation and pro-apoptotic genes. We have successfully constructed representative cDNA libraries in a poxvirus-based vector that can be recovered from cells that have undergone lethality-based selection. This strategy has been applied to selection of a gene that encodes a cytotoxic T-cell target antigen common to several independently derived tumors.

A gene on the HER2 amplicon, C35, is an oncogene in breast cancer whose actions are prevented by inhibition of Syk.

BACKGROUND: C35 is a 12 kDa membrane-anchored protein endogenously over-expressed in many invasive breast cancers. C35 (C17orf37) is located on the HER2 amplicon, between HER2 and GRB7. The function of over-expressed C35 in invasive breast cancer is unknown. METHODS: Tissue microarrays containing 122 primary human breast cancer specimens were used to examine the association of C35 with HER2 expression. Cell lines over-expressing C35 were generated and tested for evidence of cell transformation in vitro. RESULTS: In primary breast cancers high levels of C35 mRNA expression were associated with HER2 gene amplification. High levels of C35 protein expression were associated with hallmarks of transformation, such as, colony growth in soft agar, invasion into collagen matrix and formation of large acinar structures in three-dimensional (3D) cell cultures. The transformed phenotype was also associated with characteristics of epithelial to mesenchymal transition, such as adoption of spindle cell morphology and down-regulation of epithelial markers, such as E-cadherin and keratin-8. Furthermore, C35-induced transformation in 3D cell cultures was dependent on Syk kinase, a downstream mediator of signalling from the immunoreceptor tyrosine-based activation motif, which is present in C35. CONCLUSION: C35 functions as an oncogene in breast cancer cell lines. Drug targeting of C35 or Syk kinase might be helpful in treating a subset of patients with HER2-amplified breast cancers.

The Influence of Maternal Schizotypy on the perception of Facial Emotional Expressions during Infancy: an Event-Related Potential Study.

Parenting directly affects the developmental and clinical outcomes of children. How parental personality relates to perceptual and cognitive mechanisms during early development is not clear. For parents with traits of the personality dimension schizotypy, would their infant display brain responses similar to those on the schizophrenia-spectrum? This study investigates whether maternal personality influences early social-cognitive awareness during the first 6 postnatal months. Schizotypy is a dimension of personality within the general population. If deficits contribute to the development of schizophrenia-spectrum disorders by influencing the development of symptom-like characteristics, they may be observable in neurotypical individuals with schizotypal characteristics. Parents and their infants were shown standardised positive and negative faces and event-related potential responses were assessed. It was hypothesised that the infants of schizotypic mothers would display differential Negative-central event-related potentials for the happy and fearful expressions when compared to infants of non-schizotypic mothers. Results support prior literature; indicating 6-month-old infants allocate more attentional resources to fearful when contrasted to happy faces. The adult cohort displays this same ability. In addition, schizotypic mothers displayed comparable amplitudes for both expressions in comparison to the control mothers who exhibited larger amplitudes towards the fearful compared to the happy expression. Infants of schizotypic mothers did not show a greater sensitivity to facial expressions at 6-months, but schizotypic mothers showed a generalised response towards facial expressions compared to the typical P600 response illustrated by the control mothers. The present study suggests that development in the higher cognitive domains, such as the allocation of attention to novel stimuli, are not affected at 6 months of age by maternal personality related to schizotypy when examined at the group level. Implications for personality development, maternal-infant interactions and cognitive neuroscience methodologies are discussed.

Amantadine in a multimodal analgesic regimen for alleviation of refractory osteoarthritis pain in dogs.

BACKGROUND: Nonsteroidal anti-inflammatory drugs (NSAIDs) do not always provide sufficient pain relief in dogs with osteoarthritis (OA). HYPOTHESIS: The use of amantadine in addition to NSAID therapy will provide improved pain relief when compared with the use of nonsteroidal analgesics alone in naturally occurring OA in dogs. ANIMALS: Thirty-one client-owned dogs with pelvic limb lameness despite the administration of an NSAID. METHODS: The study was randomized, blinded, and placebo controlled with parallel groups (days 21-42). On day 0, analgesic medications were discontinued. On day 7, all dogs received meloxicam for 5 weeks. On day 21, all dogs received amantadine (3-5 mg/kg once daily per os) or placebo for 21 days, in addition to receiving meloxicam. Assessments were performed before the study and on days 7, 21, and 42. Primary outcome measures were blinded owner assessments of activity using client-specific outcome measures (CSOM) on days 0, 7, 21, and 42. Data were analyzed by a mixed model approach. RESULTS: For CSOM activity, there was a significant time by treatment effect (P=.009). On the basis of the planned post hoc t-tests of postrandomization means, there was a significant difference between treatment groups on day 42 (P=.030), with the amantadine group being more active. CONCLUSIONS AND CLINICAL IMPORTANCE: In dogs with osteoarthritic pain refractory to an NSAID, physical activity is improved by the addition of amantadine. Amantadine might be a useful adjunct therapy for the clinical management of canine osteoarthritic pain.

Arachidonic acid potentiates acid-sensing ion channels in rat sensory neurons by a direct action.

Acid-sensing ion channels (ASICs) are activated by a decrease in extracellular pH. ASICs are expressed in nociceptive sensory neurons, and several lines of evidence suggest that they are responsible for signaling the pain caused by extracellular acidification, but little is understood of the modulation of ASICs by pro-inflammatory factors. Using whole-cell patch clamp we demonstrate that low pH evokes three distinct inward currents in rat dorsal root ganglion neurons: a slowly inactivating transient current, a rapidly inactivating transient current, and a sustained current. All three currents were potentiated by arachidonic acid (AA), to 123%, 171%, and 264% of peak current, respectively. Membrane stretch had no effect on proton-gated currents, implying that AA is unlikely to act via local membrane deformation. The current carried by heterologously expressed ASIC1a and ASIC3 was also potentiated by AA. AA potentiates ASIC activation by a direct mechanism, because inhibition of AA metabolism had no effect on potentiation, and potentiation of single ASIC2a channels could be observed in cell-free patches. Potentiation by lipids with the same chain length as AA increased as the number of double bonds was increased. AA is known to be released in inflammation and the results suggest that AA may be an important pro-inflammatory agent responsible for enhancing acid-mediated pain.

Can probiotics enhance vaccine-specific immunity in children and adults?

The growing use of probiotics by the general public has heightened the interest in understanding the role of probiotics in promoting health and preventing disease. General practitioners and specialists often receive inquiries from their patients regarding probiotic products and their use to ward off systemic infection or intestinal maladies. Enhanced immune function is among the touted health benefits conferred by probiotics but has not yet been fully established. Results from recent clinical trials in adults suggest a potential role for probiotics in enhancing vaccine-specific immunity. Although almost all vaccinations are given during infancy and childhood, the numbers of and results from studies using probiotics in pediatric subjects are limited. This review evaluates recent clinical trials of probiotics used to enhance vaccine-specific immune responses in adults and infants. We highlight meaningful results and the implications of these findings for designing translational and clinical studies that will evaluate the potential clinical role for probiotics. We conclude that the touted health claims of probiotics for use in children to augment immunity warrant further investigation. In order to achieve this goal, a consensus should be reached on common study designs that apply similar treatment timelines, compare well-characterised probiotic strains and monitor effective responses against different classes of vaccines.

Comparative evaluation of six rapid serological tests for HIV-1 antibody.

Six commercial rapid test kits for HIV-1 antibody were evaluated. Four laboratory technologists tested sera from four groups of U.S. military personnel or dependents: 202 subjects positive for HIV-1 antibody by Western blot, 200 seronegative voluntary blood donors, 199 seronegative obstetrics/gynecology patients, and 99 subjects with sera reactive by ELISA but negative by recombinant protein EIA and indeterminate by Western blot. The three tests using solid-phase immunoassay technology demonstrated the highest mean sensitivity (> 99%) and specificity (> 91%) for all groups tested, including sera indeterminate by Western blot. Two dot-immunoblot assays were less specific, possibly due to indistinct reaction end points, and a latex agglutination assay was also less specific because of difficulty distinguishing reactive results from the granular background. In an "ease-of-use" assessment, solid-phase capture immunoassays required less time and equipment and were easier to interpret than other testing methods. Solid-phase capture immunoassays for HIV-1 antibody may be suitable for use in emergency situations and in developing countries because they are highly sensitive and specific and are rapidly performed with minimal laboratory equipment.

Demographics of aging and skin disease.

What are the dermatologic needs of aging and skin disease in the elderly in the new millennium? This question may be impossible to answer, but predictions may be derived from current sources of data. Data from the US Census Bureau and the National Ambulatory Medical Care Survey are used to characterize current use of dermatologic care in the aging population of the United States and to estimate possible future trends in dermatologic care as the elderly population continues to grow. Dermatologic accompaniments of aging are briefly reviewed, as are issues in health care financing.

Isovolemic hemodilution in stroke. A study in gerbils.

Isovolemic hemodilution has been reported to increase cerebral perfusion in humans and has been advocated as a treatment for acute cerebral infarction. This study examines the effect of isovolemic hemodilution with low-molecular-weight dextran on mortality and the incidence of neurological deficit in gerbils after internal carotid ligation. Sixty-four Mongolian gerbils were anesthetized with pentobarbital and the left internal carotid artery was ligated in both control and experimental animals. In the experimental group, blood was removed and an equal volume of dextran was injected to reestablish normal blood volume and lower hematocrit to a mean of 30.5. Control animals were not so treated. Animals were observed for neurological deficits for 24 hours after carotid ligation. The incidence of neurological deficit in control animals was 67%; it was 64% in the experimental group. Mortality within the first 24 hours was 28% in the controls and 75% in animals that were treated by hemodilution (p less than 0.001). Isovolemic hemodilution with dextran did not reduce the incidence of neurological deficit after carotid ligation in gerbils and was associated with a significant increase in mortality during the first 24 hours.

Adolescent pregnancy: a twenty year review.

In 20 years, from 1957 through 1977, there were 2,280 cases of adolescent pregnancy. The number one problem was low birth weight which occurred in 17 percent of all patients. The second significant obstetrical risk was toxemia of pregnancy, occurring in 15 percent of the cases. The cesarean section rate was 6.7 percent. There were 1.8 percent stillbirths and one maternal death.

An analysis of methods to reform Medicare payment for physician services.

Congress and the administration are changing Medicare's method of paying for physician services, with the intention of reducing the growth in or absolute level of Medicare payment rates. We analyzed the implications of four possible strategies for payment reform: modifications to the present system of paying by customary, prevailing, and reasonable (CPR) charges; payment based on fee schedules; payment for packages of related services; and capitation payment. In some cases, improved quality or access may result. The inherent danger is that constraining program expenditures may impair access to and quality of care to beneficiaries, especially for poor or infirm people, who are more vulnerable. Thus, any payment reform will require careful monitoring to safeguard beneficiaries' quality of care.

Human bite marks. Differential diagnosis.

Human bite marks are common findings in cases of fights among children, child abuse, sexual abuse, among institutionalized persons, and in a number of homicide cases. Human bites can mimic annular or arciform dermatoses. These are reviewed from both a clinical and histologic viewpoint. An example is presented of a 2 year-old girl with several annular lesions that were clinically mistaken for a dermatophyte infection. Antifungal medications were ineffective. After several days, a dermatologist identified the lesions as human bites. Physicians and other health care workers must be able to differentiate the clinical appearance of bite marks from other dermatologic diseases at an early stage so as to initiate proper therapy and counseling and, if indicated, a search for the perpetrator.

Developing a forensic dental code and programme.

An attempt was made to develop a computer-assisted programme to aid in the positive identification of human remains. The first and second generations of a computer code and programme for entering ante-mortem and post-mortem records into a computer data bank are discussed. The initial code is presented along with the results following the computer evaluation of 50 record pairs. The self-evaluating aspects of the programme are discussed and the proposed second generation code is presented. The programme exhibits some promising characteristics but further refinement based on increased data banks is needed.