A carbocyclic nucleoside analogue is a TNF-alpha inhibitor with immunosuppressive action: role of prostaglandin E2 and protein kinase C and comparison with pentoxifylline

PURPOSE: To compare the activity of topical 0.05% and 0.01% mequitazine versus vehicle in the prevention of allergic conjunctivitis induced by a conjunctival provocation test with allergens. METHODS: Forty subjects with a history of grass pollen allergic conjunctivitis were enrolled in this comparative, randomized, double-masked study. Fifteen minutes before the conjunctival provocation test, subjects received one instillation alone of 0.05% mequitazine eyedrops in one eye and in the fellow eye either one drop of 0.01% dose or vehicle, at the same dosage. Then a specific conjunctival provocation test (CPT) was performed with the allergen threshold concentration previously defined. The therapeutic efficacy was assessed by the decrease in a symptomatic composite score representing the allergic reaction. RESULTS: Topical mequitazine at the dose of 0.05% significantly decreases the intensity of the allergic reaction compared to vehicle. Moreover, the group treated with this dose needed a higher allergen concentration to trigger ocular signs. The 0.01% dose appears efficient only on itching, compared to vehicle. CONCLUSION: The results of this study support the efficacy and the onset of action of 0.05% mequitazine eyedrops versus 0.01% or vehicle in the prevention of allergic conjunctivitis induced by a conjunctival provocation test.     

Biased suppression of hematopoiesis and multiple developmental defects in chimeric mice containing Shp-2 mutant cells

Shp-2 is a cytoplasmic tyrosine phosphatase that contains two Src homology 2 (SH2) domains at the N terminus. Biochemical data suggests that Shp-2 acts downstream of a variety of receptor and cytoplasmic tyrosine kinases. A targeted deletion mutation in the N-terminal SH2 (SH2-N) domain results in embryonic lethality of homozygous mutant mice at midgestation. In vitro embryonic stem (ES) cell differentiation assays suggest that Shp-2 might play an important role in hematopoiesis. By aggregating homozygous mutant (Shp-2(-/-)) ES cells and wild-type (WT) embryos, we created Shp-2(-/-)-WT chimeric animals. We report here an essential role of Shp-2 in the control of blood cell development. Despite the widespread contribution of mutant cells to various tissues, no Shp-2(-/-) progenitors for erythroid or myeloid cells were detected in the fetal liver and bone marrow of chimeric animals by using the in vitro CFU assay. Furthermore, hematopoiesis was defective in Shp-2(-/-) yolk sacs. In addition, the Shp-2 mutation caused multiple developmental defects in chimeric mice, characterized by short hind legs, aberrant limb features, split lumbar vertebrae, abnormal rib patterning, and pathological changes in the lungs, intestines, and skin. These results demonstrate a functional involvement of Shp-2 in the differentiation of multiple tissue-specific cells and in body organization. More importantly, the requirement for Shp-2 is more stringent in hematopoiesis than in other systems.     

Extracellular matrix for a rechargeable cell delivery system

Above a critical concentration, aqueous polymer solutions of N-isopropylacrylamide copolymers with small amounts of acrylic acid, synthesized in benzene by radical polymerization, exhibited four distinct phases as the temperature increased; clear solution, opaque solution, gel and shrunken gel. The transition between the opaque solution phase and the gel phase was in the range of 30-34 degrees C and was reversible without syneresis and noticeable hysteresis under the experimental conditions used in this study. Islets of Langerhans, isolated from Sprague-Dawley rat pancreata and entrapped in the gel matrix, remained viable, with no significant decrease in insulin secretion function in vitro for one month. When islets were encapsulated with the gel matrix in hollow fibers [molecular weight cut-off (MWCO)= approximately 400000] and were exposed to dynamic changes in glucose and theophylline concentrations, their insulin secretion patterns demonstrated a smaller lag time and higher amplitude in insulin release than islets entrapped in a conventional alginate matrix under the same experimental conditions. From these two observations, i.e. gel reversibility and islet functionality in the matrix observed in in vitro experiments, the N-isopropylacrylamide copolymers with acrylic acid synthesized in this study are optimum candidates for the extracellular matrix in a diffusion chamber-type cell delivery system in order to recharge the entrapped cells when cell functionality in the system decreases.     

Protein-tyrosine phosphatase Shp-2 regulates cell spreading, migration, and focal adhesion

Shp-2, a widely expressed cytoplasmic tyrosine phosphatase with two SH2 domains, is believed to participate in signal relay downstream of growth factor receptors. We show here that this phosphatase also plays an important role in the control of cell spreading, migration, and cytoskeletal architecture. Fibroblast cells lacking a functional Shp-2 were impaired in their ability to spread and migrate on fibronectin compared with wild-type cells. Furthermore, Shp-2 mutant cells displayed an increased number of focal adhesions and condensed F-actin aggregation at the cell periphery, properties reminiscent of focal adhesion kinase (FAK)-deficient cells. This is consistent with our previous observations in vivo that mice homozygous for the Shp-2 mutation died at midgestation with similar phenotype to FAK and fibronectin-deficient embryos, having severe defects in mesodermal patterning, particularly the truncation of posterior structures. Biochemical analysis demonstrated that FAK dephosphorylation was significantly reduced in Shp-2 mutant cells in suspension. Furthermore, regulated association of Src SH2 domain with FAK and paxillin during cell attachment and detachment on fibronectin was disrupted in Shp-2 mutant cells. This report defines a unique role of the Shp-2 tyrosine phosphatase in cell motility, which might guide the design of a new strategy for pharmaceutical interference of tumor metastasis.     

Axonal injury alters alternative splicing of the retinal NR1 receptor: the preferential expression of the NR1b isoforms is crucial for retinal ganglion cell survival

Cellular-specific splicing of the retinal NMDAR1 receptor (NR1) and expression of NMDAR2 receptor (NR2) subunits in response to optic nerve injury was investigated by in situ hybridization in adult rats. A controlled optic nerve crush led to a clear alteration in the expression of alternatively spliced NR1 variants in the retinal ganglion cell layer (GCL). The NR1-2b and NR1-4b isoforms were preferentially expressed between 2 d and 1 week after injury, whereas expression for all other isoforms remained either unchanged or decreased to barely detectable levels within 4 weeks. Cellular silver grain density for NR2 subunits also declined in the GCL after trauma. To directly test the hypothesis that NR1b expression is crucial for cell survival after axonal trauma, we administered intraocularly an antisense oligonucleotide against the NR1b isoform 2 and 3 d after injury. This led to a drastic loss of retrogradely labeled retinal ganglion cells (RGCs). Antisense targeting clearly reduced retinal NR1 protein levels, as judged by Western blot analysis, but had no effect on the cell number in control retinas. These findings point toward injury-specific changes in alternative splicing of the NR1 receptor, which are crucial for the survival of RGCs after partial axonal trauma. We therefore propose that this reflects an adaptive, rather than a pathogenic, cellular response to neurotrauma.     

Enhanced post-receptor insulin effects in women following dehydroepiandrosterone infusion

While treatment for the neonate continues to be challenging, current technical advances offer more options. Before the 1980s chronic dialysis was technically so difficult for neonates and infants that most considered it impossible; in the 1990s renal replacement therapy is a viable choice. The purpose of this article is neither to advocate active intervention nor passive supportive care, but to help the reader consider questions frequently faced when deciding what to do for a neonate with end-stage renal disease (ESRD). Today ethical issues are tied closely to health care reform, so this bioethical dilemma has only begun. As health care reform addresses benefits of care, the bioethical dilemmas raised by neonates with ESRD will need to carefully considered.     

Paracrine expression of a native soluble vascular endothelial growth factor receptor inhibits tumor growth, metastasis, and mortality rate

Vascular endothelial growth factor (VEGF) is a potent and selective vascular endothelial cell mitogen and angiogenic factor. VEGF expression is elevated in a wide variety of solid tumors and is thought to support their growth by enhancing tumor neovascularization. To block VEGF-dependent angiogenesis, tumor cells were transfected with cDNA encoding the native soluble FLT-1 (sFLT-1) truncated VEGF receptor which can function both by sequestering VEGF and, in a dominant negative fashion, by forming inactive heterodimers with membrane-spanning VEGF receptors. Transient transfection of HT-1080 human fibrosarcoma cells with a gene encoding sFLT-1 significantly inhibited their implantation and growth in the lungs of nude mice following i.v. injection and their growth as nodules from cells injected s.c. High sFLT-1 expressing stably transfected HT-1080 clones grew even slower as s.c. tumors. Finally, survival was significantly prolonged in mice injected intracranially with human glioblastoma cells stably transfected with the sflt-1 gene. The ability of sFLT-1 protein to inhibit tumor growth is presumably attributable to its paracrine inhibition of tumor angiogenesis in vivo, since it did not affect tumor cell mitogenesis in vitro. These results not only support VEGF receptors as antiangiogenic targets but also demonstrate that sflt-1 gene therapy might be a feasible approach for inhibiting tumor angiogenesis and growth.