Inhibitory effects of glycoprotein-120 (G-120) from Ulmus davidiana Nakai on cell growth and activation of matrix metalloproteinases

Excessive breakdown of extracellular matrix by metalloproteinases (MMPs) occurs in many pathological conditions. Consequently, methods for inhibiting MMP activity have therapeutic potential. In this study, we investigated the effect of G-120, a 120 kDa glycoprotein purified from the Oriental herbal plant, Ulmus davidiana Nakai (UDN), on the activity and production of several MMPs by evaluating its growth inhibitory effect on NIH 3T3 cells. Tritium uptake assays showed that proliferation of NIH 3T3 cells was strongly suppressed, and G-120-mediated inhibition of DNA synthesis proved to involve a cytostatic, rather than a cytotoxic, effect, as shown by cytotoxicity and apoptosis assays. More importantly, G-120 strongly reduced the gelatinolytic and collagenase activities of MMP proteins, as well as expression of MMP-2 and MMP-9. Electrophoretic mobility shift assays revealed that it suppressed the DNA binding activity of NF-kappaB. Collectively, our observations show that G-120 strongly inhibits the activation of MMPs and NF-kappaB.     

Anti-tumor activity of the farnesyl-protein transferase inhibitors arteminolides, isolated from Artemisa

Members of the Artemisia genus are important medicinal plants found throughout the world. Arteminolides A-D (1-4), isolated from the aerial parts of Artemisia, have an inhibitory activity on farnesyl-protein transferase (FPTase; EC 2.5.1.29) in in vitro assay. This study was carried out with the purpose of validating anti-tumor effects of the compounds in human tumor cells and mouse xenograft model. The arteminolides inhibited tumor cell growth in a dose-dependent manner. Furthermore, arteminolide C (3) blocked in vivo growth of human colon and lung tumor xenograft without the loss of body weight in nude mice.     

Effects of Aroclor 1254 on the expression of rat placental PRL-family genes

This study was performed to investigate the effects of Aroclor 1254 (A1254), a commercial polychlorinated biphenyl mixture, on the expression of rat placental prolactin (PRL) family genes and reproductive activity. Placental lactogen-Iv and -II, and prolactin-like protein-A and -C mRNA levels were significantly decreased in the placentas of A1254-treated rats in a dose-dependent manner. The mRNA levels of Pit-1alpha and beta isotypes, which are involved in the regulation of PRL family gene expression, were also decreased in the A1254-treated rat placenta. In the rat placental junctional zone, high-dose A1254 (25 mg/kg B.W.) treatment reduced the number of spongiotrophoblasts, cells in which the PRL family genes are expressed. Finally, maternal exposure to A1254 was shown to have significant toxic effects on reproductive activity, including embryonic and placental growth retardation, delay of parturition, and reduction of the number of pups per litter. The results of the present study indicated that A1254 has an inhibitory effect on PRL family, Pit-1alpha, and beta gene expression in the rat placenta, leading to significant toxic effects on reproductive activity in rats.     

Activity alters muscle reinnervation and terminal sprouting by reducing the number of Schwann cell pathways that grow to link synaptic sites

In partially denervated rodent muscle, terminal Schwann cells (TSCs) located at denervated end plates grow processes, some of which contact neighboring innervated end plates. Those processes that contact neighboring synapses (termed "bridges") appear to initiate nerve terminal sprouting and to guide the growth of the sprouts so that they reach and reinnervate denervated end plates. Studies conducted prior to knowledge of this potential involvement of Schwann cells showed that direct muscle stimulation inhibits terminal sprouting following partial denervation (Brown and Holland, 1979). We have investigated the possibility this inhibition results from an alteration in the growth of TSC processes. We find that stimulation of partially denervated rat soleus muscle does not alter the length or number of TSC processes but does reduce the number of TSC bridges. Stimulation also reduces the number of TSC bridges that form between end plates during reinnervation of a completely denervated muscle. The nerve processes ("escaped fibers") that normally grow onto TSC processes during reinnervation are also reduced in length. Therefore, stimulation alters at least two responses to denervation in muscles: (1) the ability of TSC processes to form or maintain bridges with innervated synaptic sites, and (2) the growth of axons along processes extended by TSCs.     

Phosphorylation of threonine 10 on CKBBP1/SAG/ROC2/Rbx2 by protein kinase CKII promotes the degradation of IkappaBalpha and p27Kip1

In eukaryotic cells, protein kinase CKII is required for progression through the cell division cycle. We recently reported that CKBBP1/SAG/ROC2/Rbx2 associates with the beta-subunit of CKII and is phosphorylated by purified CKII in the presence of ATP in vitro. In this report, we demonstrate that CKBBP1 is efficiently phosphorylated in vitro by purified CKII in the presence of GTP and by heparin-sensitive protein kinase in HeLa cell extract. Mutational analysis indicates that CKII phosphorylates threonine at residue 10 within CKBBP1. Furthermore, CKBBP1 is phosphorylated in vivo and threonine to alanine mutation at residue 10 abrogates the phosphorylation of CKBBP1 observed in vivo, indicating that CKII is a major kinase that is responsible for in vivo phosphorylation of CKBBP1. As compared with the wild-type CKBBP1 or CKBBP1T10E (in which threonine 10 is replaced by glutamate), overexpression of nonphosphorylatable CKBBP1 (CKBBP1T10A) results in accumulation of IkappaBalpha and p27Kip1. Experiments using proteasome inhibitor MG132 and CKII inhibitor 5,6-dichloro-1-beta-d-ribofuranosylbenzimidazole suggest that the accumulation of IkappaBalpha and p27Kip1 results primarily from the reduction of proteasomal degradation in cells expressing CKBBP1T10A, and that CKII-mediated CKBBP1 phosphorylation is required for efficient degradation of IkappaBalpha and p27Kip1. Overexpression of CKBBP1T10A in HeLa cells suppresses cell proliferation and causes accumulation of G1/G0 peak of the cell cycle. Taken together, our results indicate that CKII may control IkappaBalpha and p27Kip1 degradation and thereby G1/S phase transition through the phosphorylation of threonine 10 within CKBBP1.     

Cloning and expression analysis of a Parkinson's disease gene, uch-L1, and its promoter in zebrafish

Three genes, alpha-synuclein, parkin, and ubiquitin C-terminal hydrolase L1 (UCH-L1), have been associated with inherited forms of Parkinson's disease (PD), although their in vivo functions have remained largely unknown. To develop an animal model for the molecular study of PD, we cloned zebrafish uch-L1 cDNA and its gene promoter. Sequence analysis revealed that the zebrafish Uch-L1 is highly homologous (79%) to the human UCH-L1, which is a member of the deubiquitinating enzymes. By whole-mount in situ hybridization, we examined the spatiotemporal expression of uch-L1 mRNA in developing zebrafish embryos. The uch-L1 mRNAs are detected in neuronal cells at the first day of embryo development. The expression domain of uch-L1 overlaps with that of tyrosine hydroxylase, a molecular marker for dopaminergic neurons, in the ventral diencephalon, an equivalent structure to the substantia nigra where PD progresses in human. To further analyze the tissue-specific regulation of uch-L1 gene expression, we also tested its gene promoter activity and showed a preferential neuronal expression in transient transgenic zebrafish embryos. These results suggest that uch-L1 may have an important role in the development of neuronal cells in early embryos as well as in the degeneration and disease of neuronal cells in late adult brain.     

Allometry and biomass of Korean pine (Pinus koraiensis) in central Korea

Aboveground tree biomass of Korean pine (Pinus koraiensis Sieb. et Zucc.) was determined for a natural forest of Korean pine and mixed deciduous trees and seven age classes of plantation forests in central Korea. Regression analyses of the dry weights of stem wood, stem bark, branches, and needles versus diameter at breast height were used to calculate regression equations of the form of log Y = a + b log X. Biomass of Korean pine in the mixed forest was 118 Mg ha(-1), and biomass in the plantations was linearly related to stand age, ranging from 52.3 Mg ha(-1) in 11 to 20-year-old stands to 317.9 Mg ha(-1) in 71 to 80-year-old stands. The proportions of stem wood and stem bark in the total aboveground biomass decreased with stand age while those of branch and needle increased. Specific leaf area of Korean pine ranging from 35.2 to 52.1 cm2 g(-1) was significantly different among crown positions and needle ages; in general, lower crown position and current needles had the greatest surface area per unit dry weight.