Etanercept decreases tumor necrosis factor-α activity in chronic wound fluid

High levels of tumor necrosis factor-alpha (TNF-alpha), a pro-inflammatory cytokine, are present in the wound fluid of chronic nonhealing wounds. This leads to increased inflammation, cytokine expression, and ultimately results in impaired wound healing and tissue destruction. Etanercept is a recombinant fusion protein that consists of the soluble TNF receptor (p75) linked to the Fc portion of human IgG1. It is an effective inhibitor of TNF-alpha and has been shown to provide rapid and sustained improvement in rheumatoid arthritis by acting as a soluble receptor binding TNF-alpha and preventing its proinflammatory activities. Therefore, the aim of this study was to determine whether Etanercept could inhibit TNF-alpha activity in chronic wound fluid. Wound fluid was collected from the venous leg ulcers of 16 different patients. The effect of Etanercept on TNF-alpha activity was evaluated using both a TNF-alpha bioassay and an enzyme-linked immunoassay. Etanercept was found to reduce the cytotoxic effect of chronic wound fluid on L929 fibroblasts by approximately 30% and neutralized TNF-alpha binding in the enzyme-linked immunoassay by up to 80%. Direct application of Etanercept to chronic wounds may therefore reduce the inflammatory activity of TNF-alpha, which could reduce the chronicity of venous leg ulcers and thus aid in the healing of these wounds.     

Differential role for the striatum and cerebellum in response to novel movements using a motor learning paradigm.

The aim of this pilot study was to examine the role of the striatum and cerebellum in the adaptation to a novel movement within a sequence of practiced movements using a motor learning paradigm. The performance of patients in the early or advanced stages of Parkinson's disease (PD) and of patients with damage to the cerebellum (CE) was compared, respectively to a group of aged and young matched controls on an adapted version of the Mirror-Tracing Test. In this task, subjects were required to trace a series of complex figures in two conditions: (1) a Practiced condition, in which the figures were composed of the juxtaposition of three simple designs that were extensively practiced before; and (2) a Mixed condition in which triads were created by replacing the last simple figure of the triads in the Practiced condition by a new simple figure that had never been traced individually before. Results showed that all clinical groups were slower than controls at tracing the Practiced triads. Most interestingly, however, only patients in the advanced stages of PD showed increased completion time to trace the triads in the Mixed condition. This suggests that a bilateral striatal dysfunction affects the ability to adapt to a novel motion within a sequence of practiced movements. Although exploratory, these results support a functional dissociation between the striatum and cerebellum in acquiring visuomotor skilled behaviors.     

A novel two-step method for the formation of tissue-engineered cartilage by mature bovine chondrocytes: the alginate-recovered-chondrocyte (ARC) method.

Most attempts to tissue-engineer cartilage have involved seeding of cultured cells into a biological or synthetic scaffold. We have developed a novel two-step culture approach that makes possible the in vitro formation of cartilaginous-like tissue by mature adult bovine chondrocytes without the aid of a synthetic matrix. The first step consists of culturing chondrocytes under conditions that maintain their rounded shape and their molecular phenotype as assessed by type II collagen and aggrecan production. This step was accomplished by culturing the isolated chondrocytes in alginate beads until the cells have reestablished a proteoglycan-rich cell-associated matrix (CM). The second step consists of culturing the cells with their CM, after recovery from the beads, on a tissue culture insert with a porous membrane. In this study, young adult bovine articular chondrocytes were cultured in alginate beads in the presence of 10% or 20% fetal bovine serum (FBS). After 7 days of culture, the alginate beads were dissolved by incubating the beads for 20 min in sodium citrate buffer, a calcium chelator. Following a brief centrifugation, the cells with their CM were recovered, resuspended in medium containing 10% or 20% FBS and seeded onto a tissue culture insert. After 1 week of culture on the insert, the individual cells with their CM progressively became incorporated into a mass of cartilaginous tissue. Culture with 20% FBS resulted in the best formation of tissues. These tissues, easily recovered from the insert, were then subjected to biochemical and histological analyses. The biochemical results showed that the chondrocytes remain phenotypically stable in the tissues. The de novo tissue has a relatively high ratio of PG/collagen. Histological examination of the tissue revealed it contained a cartilage-like matrix strongly stained with toluidine blue. This scaffold-free system appears ideal to study, in vitro, the development of transplantable cartilaginous tissue.     

Tumor induction by ras and myc oncogenes in fetal and neonatal brain: modulating effects of developmental stage and retroviral dose

Introduction into fetal rat brain cells of a replication-defective retroviral vector harboring v-Ha-ras and v-gag-myc rapidly causes the induction of highly malignant undifferentiated neuroectodermal tumors following transplantation into the brains of syngeneic hosts [Wiestler, et al. (1992) Cancer Res. 52: 3760–3767]. In the present study, we have investigated the modulating effect of the developmental stage of neural target cells and of the dose of the retroviral vector used in the grafting experiments. Exposure of fetal cells from embryonic day (E)12 or E14 produced a 100% incidence of malignant neuroectodermal tumors which led to the death of recipient animals after a median latency period of 32 days. A 100-fold reduction of the virus dose from 2.062×10⁶ to 2.062×10⁴ focus-forming units/ml resulted in a lower tumor incidence of 25%. Of six neural grafts exposed to v-Ha-ras and v-myc at E16, only one showed evidence of tumorigenesis (low-grade astrocytoma and hemangioma). All other transplants were morphologically normal for observation periods of 26 weeks, indicating a marked loss of transforming activity of ras and myc in more advanced stages of brain development. In retrovirus-exposed donor cells which caused the development of neural tumors in recipient rats, malignant transformation was also evident during culture in vitro, usually after 9–12 days. Oncogene complementation was also studied in the newborn rat brain. After microinjection of the retroviral vector into the brain at postnatal day (P)0, P1 and P3, 5 out of 20 animals (25%) developed a total of seven brain tumors. Histopathologically, three of these neoplasms were malignant neuroectodermal tumors which, in contrast to those induced in fetal brain transplants showed evidence of focal glial and/or neuronal differentiation. In addition, we observed one oligodendroglioma, two hemangiomas and a malignant hemangioendothelioma. These data indicate that neural precursor cells and endothelia of the rat brain represent the major target cells for the complementary action of ras and myc and that the use of target cells from later developmental stages (E16 and postnatal) leads to the induction of both primitive and more differentiated neoplasms.These studies were supported by the Fonds zur Förderung der wissenschaftlichen Forschung in Österreich (Erwin Schrödinger fellowship, JO501-MED), by the Swiss National Science Foundation and by the Cancer League of the Kanton of Zürich