Early secondary succession in bottomland hardwood forests of southeastern Virginia.

Addressing the need for reference sites that permit wetland managers to evaluate the relative success of wetland restoration efforts, this project examines the early successional properties of a chronosequence of 17 forested wetlands that have been clear-cut and allowed to naturally revegetate. Ordinations performed on the data using CANOCO software indicated three general types of communities- one dominated by bald cypress (Taxodium distichum) and water tupelo (Nyssa aquatica), one dominated by black willow (Salix nigra), and one with a species composition similar to that of a mature stand of bottomland hardwoods. These divisions were correlated with the percentage of stems originating as coppice on stumps leftover from the clear-cut. In particular, the bottomland hardwood stands were regenerating predominantly as coppice, while the cypress/tupelo and black willow stands were regenerating primarily as seedlings. As indicated by the earlier development of overstory basal area, coppice sites were also regenerating much faster. The hydrology of a site also exhibited a strong impact on the rate of regeneration, with the semipermanently to permanently flooded portions of sites often exhibiting little or no regeneration. The results indicate that, because of the overwhelming reliance on coppice sprouts as the main source of stems and the concomitant enhanced rates of regeneration, certain vegetative parameters of clear-cut bottomiand hardwood stands would not be effective benchmarks by which to judge the relative success of creation and restoration efforts.

Cysteine protease activity is up-regulated in inflamed ankle joints of rats with adjuvant-induced arthritis and decreases with in vivo administration of a vinyl sulfone cysteine protease inhibitor.

OBJECTIVE: Cysteine proteases are postulated to play a role in tissue destruction in the joints of animals with arthritis. The purpose of the present study was to confirm the concept that cysteine proteases are enzymes involved in the pathology of rheumatoid arthritis (RA). METHODS: Arthritis was induced in Lewis rats by adjuvant injection (adjuvant-induced arthritis [AIA] model) and scored for inflammation. At necropsy, the rear paws were either fixed in formalin and assigned a histologic score (based on synovial cell proliferation, cartilage erosion, bone erosion, and fibroproliferative pannus) or frozen, cryosectioned, and assayed for enzyme activity either by in situ cytochemical staining with a post-azo-coupling method using a chromogenic substrate (Z-arg-arg-MNA) or by a novel assay placing the tissue section directly in a cuvette using the fluorogenic substrate Z-arg-arg-AMC. RESULTS: Enzymatic activity, measured either in frozen sections in situ or in the cuvette assay, was positively correlated with joint destruction (r = 0.7) and inflammation (r = 0.8). Activity was not inhibited significantly by Pefabloc (a serine protease inhibitor), EDTA (a metalloprotease inhibitor), or pepstatin A (an aspartyl protease inhibitor) but was inhibited by E-64 and vinyl sulfone irreversible inhibitors of cysteine proteases. The effect of one of the vinyl sulfone cysteine protease inhibitors, Mu-Leu-HomoPhe-vinylsulfone, was tested in vivo by dietary administration at 2.2 mg/kg/day in the AIA model; this resulted in a significant decrease in inflammation and in the amount of cysteine protease activity measured in the joint tissue. CONCLUSION: Cysteine protease activity levels increase in the diseased state and may be an important target for designing small molecule inhibitors to reduce the inflammation and tissue destruction associated with RA.

Development of an in vitro extracellular matrix assay for studies of brain tumor cell invasion.

Invasion of brain by tumor cells is an inherent feature of the malignant phenotype. Assays to quantitate invasiveness should provide a powerful tool to investigate this phenomenon. We have developed a modified in vitro assay to measure tumor cell invasion, attachment, and chemotaxis using a barrier of the complex basement membrane Matrigel on gelatin-coated filters. Within 5 hours, 7.8% of U251MGp and 2.6% of SF126 human malignant glioma cells invaded the Matrigel and filter, compared with 0.8% of normal human leptomeningeal cells. The extent of invasion was directly proportional to incubation time and filter pore size and inversely proportional to the Matrigel concentration. Cells from exponentially growing U251MGp cultures invaded more readily (10.9%) than cells from plateau-phase cultures (2.3%); however, labeling studies with bromodeoxyuridine showed that quiescent cells and rapidly dividing cells were equally capable of invading. This suggests that the mechanisms underlying invasion by malignant glioma cells are distinct from those underlying proliferation and indicates the need for therapy aimed specifically at invasive behavior. In a practical application of this assay to test a potential anti-invasive strategy, monoclonal antibodies to the beta subunit of an integrin receptor mediating attachment to the extracellular matrix inhibited invasion by U251MGp cells in a dose-dependent manner. This assay should allow evaluation of the cellular and molecular basis of brain tumor progression and perhaps aid the development of rationally designed drugs that limit tumor invasion. It may also allow prediction of the clinical behavior of neoplasms in individual patients.

Experimental basis for increasing the therapeutic index of carboplatin in brain tumor therapy by pretreatment with WR compounds.

We tested an experimental strategy to decrease the dose-limiting hematotoxicity of carboplatin without compromising its activity against brain tumors. The effect of pretreatment with WR-1065, a chemomodifier that penetrates brain poorly, on carboplatin's cytotoxicity was evaluated in human hematopoietic granulocyte-monocyte progenitor cells and in three human glioblastoma cell lines. WR-1065 reduced bone marrow toxicity without decreasing carboplatin's activity against glioblastoma cells. These results suggest that the therapeutic index of carboplatin might be increased in the treatment of malignant brain tumors.