Classification and subtype prediction of adult soft tissue sarcoma by functional genomics

Adult soft tissue sarcomas are a heterogeneous group of tumors, including well-described subtypes by histological and genotypic criteria, and pleomorphic tumors typically characterized by non-recurrent genetic aberrations and karyotypic heterogeneity. The latter pose a diagnostic challenge, even to experienced pathologists. We proposed that gene expression profiling in soft tissue sarcoma would identify a genomic-based classification scheme that is useful in diagnosis. RNA samples from 51 pathologically confirmed cases, representing nine different histological subtypes of adult soft tissue sarcoma, were examined using the Affymetrix U95A GeneChip. Statistical tests were performed on experimental groups identified by cluster analysis, to find discriminating genes that could subsequently be applied in a support vector machine algorithm. Synovial sarcomas, round-cell/myxoid liposarcomas, clear-cell sarcomas and gastrointestinal stromal tumors displayed remarkably distinct and homogenous gene expression profiles. Pleomorphic tumors were heterogeneous. Notably, a subset of malignant fibrous histiocytomas, a controversialhistological subtype, was identified as a distinct genomic group. The support vector machine algorithm supported a genomic basis for diagnosis, with both high sensitivity and specificity. In conclusion, we showed gene expression profiling to be useful in classification and diagnosis, providing insights into pathogenesis and pointing to potential new therapeutic targets of soft tissue sarcoma.     

Mechano-active scaffold design of small-diameter artificial graft made of electrospun segmented polyurethane fabrics

To fabricate a "mechano-active" tubular scaffold of nonwoven mesh-type small-diameter artificial graft made of the synthetic durable elastomer, segmented polyurethane, the fabrication technique of electrospinning on a mandrel under a high rotation speed and transverse movement was used. Emphasis was placed on how the rotation speed of the mandrel and the fusion or welding states of fibers at contact points affect the compliance (ease of intraluminal pressure-dependent circumferential inflation) and Young's modulus determined by uniaxial stretching in the longitudinal and circumferential directions. The results showed that a high rotation speed is attributed to exhibit isotropic mechanical properties in the entire range of applied strain but reduces the compliance, and a high fusion state, which is produced using a mixed solvent with a high content of high-boiling-point solvent, reduces the compliance but is expected to exhibit high durability in a continuously loaded pulsatile stress field in an arterial circulatory system.     

Type IV collagen alpha 5 chain. Normal distribution and abnormalities in X-linked Alport syndrome revealed by monoclonal antibody.

Although the evidence indicates that mutation of the gene for the alpha 5 chain of type IV collagen, alpha 5-(IV), is the primary defect in X-linked Alport syndrome, protein data for the alpha 5(IV) chain with regard to its normal distribution and its distribution in patients with Alport syndrome is lacking. We produced a rat monoclonal antibody (H51) by immunizing rats with a synthetic peptide corresponding to the nonconsensus amino acid sequence of alpha 5(IV) NC1 domain. H51 reacted by Western blotting with 26-kd cationic monomers and associated dimers of human type IV collagen NC1 domain. Immunohistochemical studies demonstrated that in normal human kidney alpha 5(IV) was present in the glomerular basement membrane and basement membranes of the Bowman's capsule and in some tubules (collecting ducts). The alpha 5(IV) chain was also detected in the basement membranes of normal skin, eye, and lung. Male patients with X-linked Alport syndrome revealed no reactivity of renal and epidermal basement membranes with H51, whereas alpha 5(IV) staining was normal in the glomerular basement membrane of patients with other types of glomerular diseases, including benign familial hematuria. The staining was also normal in the skin of nonaffected males in X-linked Alport families. Female heterozygous for Alport syndrome exhibited a discontinuous or mosaic pattern in the immunofluorescent staining of the epidermal basement membrane. These findings confirm that in patients with X-linked Alport syndrome there are abnormalities in alpha 5(IV) in renal and epidermal basement membranes at the protein level. Immunofluorescent staining of skin biopsies with this antibody may be of value in making a diagnosis of Alport syndrome, and, furthermore, may aid in detecting carrier females in whom urinary abnormalities are often mild or silent.     

Different Ca2+ dynamics between isolated hippocampal pyramidal cells and dentate granule cells

The hippocampal formation contains a variety of neuronal types. The principal neurons are granule cells in the dentate gyrus and pyramidal cells in Ammon's horn. These two neuron types show distinct cell morphology and display a different vulnerability to ischemic injury or various neurotoxins. In order to illustrate the difference in the pathophysiological properties of these neurons, we established a method for separately culturing granule cells and pyramidal cells. They were prepared from the dentate gyrus and Ammon's horn of 3-day-old Wistar rat pups and maintained for 7–9 days in culture. After transient exposure to N-methyl-D-aspartate or glutamate, both the cultured neuron populations displayed somatic Ca²⁺ transients with similar amplitudes, but the subsequent recovery to baseline was about twice as fast in granule cells than in pyramidal cells. Similar results were obtained for K⁺ depolarization-induced Ca²⁺ elevation, suggesting that the relatively rapid Ca²⁺ clearance in granule cells is independent of Ca²⁺ influx pathways. The present study provides the first evidence for a difference in Ca²⁺ dynamics and homeostasis between granule and pyramidal cells and may represent a cellular basis for the differential vulnerability of hippocampal neurons.     

Cellular localization of inflammatory cytokines in human glomerulonephritis

We evaluated the expression of inflammatory cytokines in renal tissues obtained from 45 patients with several types of glomerulonephritis. Immunofluorescence studies with specific antibodies to interleukin (IL)-1, IL-1, IL-6, tumour necrosis factor (TNF)-, and TNF- showed intense cytoplasmic staining in the glomeruli and interstitium. Cells positive for these cytokines were found frequently in tissue from patients with lupus nephritis (WHO Class IV) and membranoproliferative glomerulonephritis, and, to a lesser extent, in tissue from patients with mesangial proliferative glomerulonephritis, Henoch-Schönlein purpura nephritis, and minimal change nephrotic syndrome. Most of these cells were dual-stained with a monoclonal antibody to monocytes-macrophages. In situ hybridization for cytokine mRNA, combined with immunoperoxidase staining for monocytes-macrophages, detected IL-1, IL-6, and TNF- mRNA in monocytes-macrophages infiltrating the glomeruli and interstitium. Occasionally, there was weak or moderate immunostaining for IL-1, IL-6, and TNF- in the glomerular mesangial and epithelial cells, but in situ hybridization signals were rarely found in these loci. These findings suggest that infiltrating monocytes-macrophages, rather than resident glomerular cells, are the major source of inflammatory cytokines in human glomerulonephritis.     

Spatial memory of children and adults assessed in the radial maze

To assess the development of accurate spatial memory and its relationship to organized search strategies, children (18-71 months old) and college students searched for rewards hidden at the ends of the arms of an eight-arm radial maze. Subjects made their first four choices in random order selected by the experimenter (forced choice) or in whatever order they wished (free choice). In some conditions, a retention interval was imposed between arm choices 4 and 5, followed by a resumption of the task. With free choice, accuracy of spatial memory increased linearly with age, and the proportion of tests on which the child entered four adjacent arms in sequence during the first four choices, a measure of organized searching, also increased linearly with age. Disrupting this sequential search strategy (forced choice) reduced accuracy for both children and college students, although the reduction with college students was small. Delays had minimal effects for children who met criterion at 0-delay and for college students. Although response patterns seem unimportant in accurate spatial memory in rats, systematic search strategies mature together with, and probably contribute to, the improvement of spatial memory with age in humans.     

Effects of dopamine agonists on hypothalamic defensive attack in cats

The effects of methamphetamine (MAT) and apomorphine (APO), dopamine agonists, were studied in 16 cats to evaluate their effects on threshold for defensive attack behavior elicited by electrical stimulation of the ventromedial hypothalamic nucleus (VMH). Directed attack and hissing were selected from elementary responses as constituting a defensive attack. Hissing threshold was measured in two situations, one with human provocation and the other without provocation. MAT administered systemically lowered the thresholds for all three types of responses in a dose-related manner (0.5, 1.0, and 3.0 mg/kg). The effects of 1.0 mg/kg of APO were almost identical to those observed with 0.5 or 1.0 mg/kg of MAT. These results suggest that MAT-induced aggressive behavior may be mediated by a dopamine-induced increase in the excitability of the VMH.     

Prostaglandin F(2alpha), cytokines and cyclic mechanical stretch augment matrix metalloproteinase-1 secretion from cultured human uterine cervical fibroblast cells

Human uterine cervical tissue is composed mainly of fibroblast cells and the extracellular matrix in which collagen types I and III predominate. It is hypothesized that these collagens are degraded by matrix metalloproteinases (MMPs) in the initial step of uterine cervical ripening during parturition. Among the MMPs, MMP-1, -8 and -13 have substrate selectivity for collagen types I and III. In the present study, we examined the regulation of MMP-1 secretion from the human uterine cervix. Immunohistochemistry detected strong staining of MMP-1, but not of MMP-8 or -13, in stromal cells of the pregnant uterine cervix. The MMP-1 expression in the pregnant uterine cervix was further confirmed by Western blot analysis and RT-PCR. To clarify the regulation of MMP-1 production, we subsequently investigated the effects of prostaglandins, inflammatory cytokines and cyclic mechanical stretch on the secretion of MMP-1 from cultured human uterine cervical fibroblast cells. Treatment with prostaglandin (PG)F(2alpha) (10(-7) to 10(-5) mol/l) or interleukin (IL)-1alpha (0.01-1.0 ng/ml) or stimulation with cyclic mechanical stretch increased MMP-1 secretion from cultured human uterine cervical fibroblast cells, with maximal increases of 3.4-, 4.5- and 1.9-fold respectively (24 h of treatment, P < 0.05 for all comparisons). These data suggest that MMP-1 may play a significant role in the degradation of extracellular collagen types I and III in the pregnant uterine cervix during the process of cervical ripening, in response to various stimulations such as PGF(2alpha), IL-1alpha and mechanical stretch.     

Single-equation models for the tear film in a blink cycle: realistic lid motion.

We consider model problems for the tear film over multiple blink cycles that utilize a single equation for the tear film; the single non-linear partial differential equation that governs the film thickness arises from lubrication theory. The two models that we consider arise from considering the absence of naturally occurring surfactant and the case when the surfactant is strongly affecting the surface tension. The film is considered on a time-varying domain length with specified film thickness and volume flux at each end; only one end of the domain is moving, which is analogous to the upper eyelid moving with each blink. Realistic lid motion from observed blinks is included in the model with end fluxes specified to more closely match the blink cycle than those previously reported. Numerical computations show quantitative agreement with in vivo tear film thickness measurements under partial blink conditions. A transition between periodic and non-periodic solutions has been estimated as a function of closure fraction and this may be a criterion for what is effectively a full blink according to fluid dynamics.