CD28-independent, TRAF2-dependent costimulation of resting T cells by 4-1BB ligand

4-1BB ligand (4-1BBL) is a member of the tumor necrosis factor (TNF) family expressed on activated antigen-presenting cells. Its receptor, 4-1BB, is a member of the TNF receptor family expressed on activated CD4 and CD8 T cells. We have produced a soluble form of 4-1BBL using the baculovirus expression system. When coimmobilized on plastic with anti-CD3, soluble 4-1BBL induces interleukin (IL)-2 production by resting CD28+ or CD28- T cells, indicating that 4-1BBL can function independently of other cell surface molecules, including CD28, in costimulation of resting T cell activation. At low concentrations of anti-CD3, 4-1BBL is inferior to anti-CD28 in T cell activation. However, when 4-1BB ligand is provided together with strong TCR signals, then 4-1BBL and anti-CD28 are equally potent in stimulation of IL-2 production by resting T cells. We find that TNF receptor-associated factor (TRAF)1 or TRAF2 associate with a glutathione S-transferase-4-1BB cytoplasmic domain fusion protein in vitro. In T cells, we find that association of TRAF1 and TRAF2 with 4-1BB requires 4-1BB cross-linking. In support of a functional role for TRAF2 in 4-1BB signaling, we find that resting T cells isolated from TRAF2-deficient mice or from mice expressing a dominant negative form of TRAF2 fail to augment IL-2 production in response to soluble 4-1BBL. Thus 4-1BB, via the TRAF2 molecule, can provide CD28-independent costimulatory signals to resting T cells.     

Comparison of two outcome measures for infants with cerebral palsy and infants with motor delays

BACKGROUND AND PURPOSE: The purpose of this study was to compare the Gross Motor Function Measure (GMFM) and the Peabody Developmental Gross Motor Scale (PDMS-GM) as measures of change in infants with cerebr-al palsy (CP) and infants with motor delays. We hypothesized that mean change scores would be greater for the GMFM than for the PDMS-GM. SUBJECTS AND METHODS: Subjects were 42 infants with a mean adjusted age of 13.9 months (SD=6.1, range=4.2-24.2). Twenty-four infants had CP, and 18 infants had motor delays. The GMFM and the PDMS-GM were administered to the infants 3 times over a 6-month period. Raw scores were standardized for data analysis. Data were analyzed using a 3-factor repeated-measures analysis of variance. RESULTS: For the 6-month period, mean PDMS-GM age-equivalent scores increased 3.8 months and mean scaled scores increased 35 points for infants with motor delays compared with increased scores of 1.8 months and 13 points for infants with CP. Mean GMFM scores increased by 12.2% for infants with rmotor delays and by 4.2% for infants with CP. The diagnosis X time interaction was significant. Infants with motor delays had a greater change in motor development compared with the infants with CP. The hypothesis that the GMFM is more responsive to change than the PDMS-GM was not supported. CONCLUSION AND DISCUSSION: The findings suggest that the GMFM and the PDMS-GM are comparable in measuring change in infants with CP or motor delays. Implications for selection and use of either measure are discussed.     

Successful delayed thrombolytic therapy in a patient with massive pulmonary embolism

Pica is the persistent, culturally and developmentally inappropriate ingestion of non-nutritive substances (DSM-IV). AB is a 75-year-old lady with a 40-year history of schizophrenia and a 20-year history of pica who, at emergency laparotomy, had 175.32 Pounds of loose change in her stomach. Although pica has been reported to coexist with schizophrenia, she had had no positive symptoms of schizophrenia for at least 20 years. She has CT evidence of fronto-tempotal atrophy most marked on the left in the temporal lobe and on the right in the frontal lobe. Pica has been found to be related to cognitive deficits and hyperoral behaviour to temporal lesions. Neuropsychological testing reveals deficits closely related to these changes.     

Immortalization of neuroendocrine pinealocytes from transgenic mice by targeted tumorigenesis using the tryptophan hydroxylase promoter

Tryptophan hydroxylase (TPH) is the first enzyme in both serotonin and melatonin biosynthesis in neuroendocrine cells of the pineal gland. The lack of immortalized neuroendocrine pineal cell lines has been a major obstacle to the study of the tissue-specific and circadian regulation of TPH gene expression in the pineal gland. Previously, we demonstrated that a 6.1 kb 5' upstream region of the mouse TPH gene directs the restricted expression of a lacZ reporter gene to the pineal gland and the raphe nuclei of transgenic mice. Therefore, to develop TPH-expressing pineal cell lines we first established transgenic mice carrying a construct consisting of 6.1 kb of 5' flanking region fused to the SV40 T-antigen. These animals developed highly invasive pineal tumors and died at 12-15 weeks of age. The pineal tumors obtained from the transgenic mice were utilized to establish the immortalized pinealocyte-derived cell lines. These cells express two marker enzymes, TPH and serotonin N-acetyltransferase (NAT). In pineal gland TPH and NAT expressions have been known to be regulated during circadian cycle. The two established cell lines therefore promise to be a valuable in vitro model system for the study of the rhythmic nature of the pineal function at molecular level in mammal.     

6beta-Acetoxynortropane: a potent muscarinic agonist with apparent selectivity toward M2-receptors

A series of tropane derivatives, related in structure to baogongteng A (1), an alkaloid from a Chinese herb, were synthesized. 6beta-Acetoxynortropane (5) had weak affinity (Ki 22 microM) for central (M1) muscarinic receptors in a [3H]quinuclidinyl benzilate binding assay but had extremely high affinity (Ki 2.6 nM) and selectivity for M2-muscarinic receptors expressed in CHO cells. It had 13-fold lower affinity for M4-receptors, 260-fold lower affinity for M3-receptors, and 8200-fold lower affinity for M1-receptors expressed in CHO cells. The 6beta-carbomethoxy analogue (14) of baogongteng A had only weak affinity for M2-muscarinic receptors, as did 6beta-carbomethoxynortropane (13) and 6beta-acetoxytropane (4). In transfected CHO cells, 6beta-acetoxynortropane (5) was an agonist at M2-receptors, based on a GTP-elicited decrease in affinity, and a full agonist with an IC50 of 11 nM at M4-receptors, based on inhibition of cyclic AMP accumulation, while being a full agonist at M1-receptors with an EC50 of 23 nM and a partial agonist at M3-receptors with an EC50 of 3.6 nM, based in both cases on stimulation of phosphoinositide breakdown. All of the 16 tropane derivatives had weak affinities for central alpha4beta2-nicotinic receptors with 6beta-carbomethoxynortropane (13) having the highest affinity, which was still 150-fold less than that of nicotine. 6beta-Acetoxynortropane (5) represents a potent muscarinic agonist with apparent selectivity toward M2-receptors.     

Biomechanical and histologic alteration of facial recipient bone after reconstruction with autogenous bone grafts and alloplastic implants: a 1-year study

Potential alteration of the underlying recipient bone resulting from a graft or implant has significant clinical relevance. The present study was designed to evaluate the biomechanical and histologic alteration of facial recipient bone with autogenous bone graft and alloplastic implants over a 1-year period. The bilateral arches of 15 rabbits were randomized between four groups: (1) control (n = 6), subperiosteal exposure of the zygomatic arch was made; (2) onlay (n = 12), bone graft was placed as an onlay to the zygomatic arch; (3) inlay (n = 6), bone graft was placed as an inlay within the zygomatic arch; (4) implant (n = 6), a stainless steel plate was placed as an onlay to the zygomatic arch. Animals were killed 1 year after grafting. In the onlay groups, all steel implants and half of the onlay bone grafts (n = 6) were separated from the zygomatic arch; the remaining onlay bone grafts (n = 6) were left on the zygomatic arch. Three-point breaking strength was measured through the center of the graft/implant site on the zygomatic arch, followed by histologic evaluation and histometric assessment of residual bone density. The findings demonstrated no difference in the breaking strength per unit bone area between the control zygomatic arch group and the onlay group in which the bone graft was left in place. Breaking strength of the zygomatic arch in the former two groups was significantly greater than that in either group in which the onlay bone graft or implant had been removed, and was also greater than the breaking strength in that group in which inlay bone had been placed (p < 0.05). Histologic assessment showed full-thickness conversion in architecture of the zygomatic arch from compact to woven bone beneath onlays of either autogenous bone graft or steel implant; histometric assessment demonstrated an accompanying decrease in bone density in the latter groups relative to the control zygoma (p < 0.05). We conclude that onlay autogenous bone graft and alloplastic implants to the facial skeleton induce transformation of both graft and recipient bone from compact to woven architecture, accompanied by a reduction in bone density. The biomechanical strength of recipient facial bone is significantly weakened if an onlay bone graft or implant is removed. Weakening occurs per unit area of remaining bone, and is therefore independent of any thinning that may occur within the recipient bone because of graft/implant placement. These findings may impact upon decisions to augment stress-bearing regions of the facial skeleton with bone graft or implants, particularly if the graft/implant may eventually require removal.     

Oxidative DNA base modifications as factors in carcinogenesis

Reactive oxygen species can cause extensive DNA modifications including modified bases. Some of the DNA base damage has been found to possess premutagenic properties. Therefore, if not repaired, it can contribute to carcinogenesis. We have found elevated amounts of modified bases in cancerous and precancerous tissues as compared with normal tissues. Most of the agents used in anticancer therapy are paradoxically responsible for induction of secondary malignancies and some of them may generate free radicals. The results of our experiments provide evidence that exposure of cancer patients to therapeutic doses of ionizing radiation and anticancer drugs causes base modifications in genomic DNA of lymphocytes. Some of these base damages could lead to mutagenesis in critical genes and ultimately to secondary cancers such as leukemias. This may point to an important role of oxidative base damage in cancer initiation. Alternatively, the increased level of the modified base products may contribute to genetic instability and metastatic potential of tumor cells.     

Telomerase activity in human development is regulated by human telomerase reverse transcriptase (hTERT) transcription and by alternate splicing of hTERT transcripts

Short mobile elements are present in different recombined forms as interspersed GC-rich islands between AT rich centromeric 155 bp tandem repeats in the dipteran Chironomus pallidivittatus . The basic element is 80 bp long, has a pronounced invert repeat structure and contains a 17 bp segment similar to the CENP-B box in mammals. The element inserts into a specific site of the 155 bp repeat in a defined orientation surrounded by 2 bp direct repeats. The total number per genome of the main variant is <20. Elements can be present in all centromeres from C.pallidivittatus and the sibling species Chironomus tentans with pronounced differences in distribution within and between species.     

Subcellular effects and localization of binding sites of phytohemagglutinin in the potato leafhopper, Empoasca fabae (Insecta: Homoptera: Cicadellidae)

To identify the means by which phytohemagglutinin (PHA) exerts its toxicity on the potato leafhopper, four different methods (thick and semi-thin sectioning combined with immunofluorescent staining, in vitro receptor autoradiography, and immunoelectron microscopy) were used to elucidate the PHA target tissue, binding site, and its effects on this tissue. Sixteen 1- or 2-day-old female potato leafhoppers were fed for 36 h on each of three treatments: a control, diet or a diet containing either the PHA-E subunit or the PHA-L subunit. The PHA-E subunit, but not PHA-L, had previously been shown to be lethal. The insects were then prepared for both light and confocal microscopy. Analysis of images showed that PHA bound only to the surface of midgut epithelial cells of the potato leafhopper. PHA-E caused severe disruption, disorganization, and elongation of the brush border microvilli, and swelling of the epithelial cells into the lumen of the gut, leading to complete closure of the lumen. Furthermore, PHA-E stimulated the division of midgut epithelial cell nuclei, leading to two nuclei in each cell. Nuclei later elongated and degraded. In contrast, PHA-L had little effect on the epithelial cells of the midgut. It did not strongly bind to the surface of epithelial cells and caused much less disruption of brush-border microvilli, less disorganization of the cells and less elongation of nuclei. Strong binding of PHA occurred solely on the cell membrane of the brush border microvilli of epithelial cells. In contrast, the controls (i.e., midgut tissue, blocking agent, PHA, and antibodies) showed that midgut tissue was not autofluorescent and showed no fluorescent binding signal. Analysis of both bright- and dark-field images obtained by autoradiography and immunoelectron microscopy confirmed these findings.     

Immunohistochemical localisation of alpha1B-adrenergic receptors in the rat iris

This article reports results from a meta-analysis on adult age differences in the negative priming effect (21 studies on identity negative priming and 8 on location negative priming). Both younger and older adults were found to be susceptible to the negative priming effect in identity and location tasks. Effect sizes were homogeneous for both tasks, indicating that the data are adequately described without reference to moderator variables. State trace analysis on identity tasks, in which mean latencies in negative priming conditions were regressed onto mean latencies in baseline conditions, showed (a) that in both age groups the negative priming effect is proportional rather than additive and (b) that the negative priming effect is smaller in older adults as compared with younger adults.