A role for Fis1 in both mitochondrial and peroxisomal fission in mammalian cells

The mammalian dynamin-like protein DLP1/Drp1 has been shown to mediate both mitochondrial and peroxisomal fission. In this study, we have examined whether hFis1, a mammalian homologue of yeast Fis1, which has been shown to participate in mitochondrial fission by an interaction with DLP1/Drp1, is also involved in peroxisomal growth and division. We show that hFis1 localizes to peroxisomes in addition to mitochondria. Through differential tagging and deletion experiments, we demonstrate that the transmembrane domain and the short C-terminal tail of hFis1 is both necessary and sufficient for its targeting to peroxisomes and mitochondria, whereas the N-terminal region is required for organelle fission. hFis1 promotes peroxisome division upon ectopic expression, whereas silencing of Fis1 by small interfering RNA inhibited fission and caused tubulation of peroxisomes. These findings provide the first evidence for a role of Fis1 in peroxisomal fission and suggest that the fission machinery of mitochondria and peroxisomes shares common components.     

Sonic hedgehog myocardial gene therapy: tissue repair through transient reconstitution of embryonic signaling

Sonic hedgehog (Shh) is a crucial regulator of organ development during embryogenesis. We investigated whether intramyocardial gene transfer of naked DNA encoding human Shh (phShh) could promote a favorable effect on recovery from acute and chronic myocardial ischemia in adult animals, not only by promoting neovascularization, but by broader effects, consistent with the role of this morphogen in embryogenesis. After Shh gene transfer, the hedgehog pathway was upregulated in mammalian fibroblasts and cardiomyocytes. This resulted in preservation of left ventricular function in both acute and chronic myocardial ischemia by enhanced neovascularization, and reduced fibrosis and cardiac apoptosis. Shh gene transfer also enhanced the contribution of bone marrow-derived endothelial progenitor cells to myocardial neovascularization. These data suggest that Shh gene therapy may have considerable therapeutic potential in individuals with acute and chronic myocardial ischemia by triggering expression of multiple trophic factors and engendering tissue repair in the adult heart.     

Tendon proteoglycans: biochemistry and function

Tendon remodeling occurs in response to changes in loading and mobilization. Though the normal mechanical function depends on precise alignment of collagen fibrils, it is proteoglycans that regulate collagen fibrillogenesis and thus, indirectly, tendon function. In this paper we discuss the basic biochemical structure of several members of two proteoglycans families. Decorin, biglycan, fibromodulin and lumican, all members of the small leucine-rich proteoglycans family, bind to collagen fibrils and are active participants in fibrillogenesis. Aggrecan and versican, two members of large modular proteoglycans or lecticans, and their partner hyaluronan likely provide tendon tissues with a high capacity to resist high compressive and tensile forces associated with loading and mobilization. We present data from our laboratory showing that proteoglycans and glycosaminoglycan content increases not only with growth but also with loading of young avian gastrocnemius tendons. Specifically, an increase in the content of keratan sulfate, chondroitin sulfate and hyaluronan was observed. Moderate exercise for several weeks led not only to a further increase in total proteoglycans content but also to qualitative changes in proteoglycan make up.     

Safety assessment of potential lactic acid bacteria Bifidobacterium longum SPM1205 isolated from healthy Koreans

The safety assessment of Bifidobacterium longum SPM1205 isolated from healthy Koreans and this strain's inhibitory effects on fecal harmful enzymes of intestinal microflora were investigated. The overall safety of this strain was investigated during a feeding trial. Groups of SD rats were orally administered a test strain or commercial reference strain B. longum 1 x 10(9) CFU/kg body weight/day for four weeks. Throughout this time, their feed intake, water intake and live body weight were monitored. Fecal samples were periodically collected to test harmful enzyme activities of intestinal microflora. At the end of the four-week observation period, samples of blood, liver, spleen, kidney, and gut tissues were collected to determine for hematological parameters and histological differences. The results obtained in this experiment demonstrated that four weeks of consumption of this Bifidobacterium strain had no adverse effects on rat's general health status, blood biochemical parameters or histology. Therefore, it is likely to be safe for human use. Fecal harmful enzymes such as beta-glucosidase, beta-glucuronidase, tryptophanase and urease, were effectively inhibited during the administration of the B. longum SPM1205. These results suggested that this B. longum SPM 1205 could be used for humans as a probiotic strain.     

Degradation of crystalline cellulose by the brown-rot basidiomycete Fomitopsis palustris

This study demonstrated that the brown rot basidiomycete Fomitopsis palustris was able to degrade crystalline cellulose (Avicel). This fungus could also produce the three major cellulases (exoglucanases, endoglucanases, and beta-glucosidase) when the cells were grown on 2.0% Avicel. Avicel degraded by F. palustris showed a decrease in relative crystallinity from 83% to 78.5% after 14 days of incubation. The characterization study indicated that optimum pH was 4.5 and optimum temperature was 70 degrees C for exoglucanase (cellobiohydrolase) activity. Hydrolysis of Avicel by the crude enzyme from F. palustris yielded 1.6 mg/ml of glucose after 43 h, which corresponded to a cellulose conversion degree of 3.2%. Therefore, this study revealed for the first time that the brown rot basidiomycete F. palustris produces cellulases capable of yielding soluble sugars from crystalline cellulose.     

Prevalence of putative periodontopathogens in subgingival dental plaques from gingivitis lesions in Korean orthodontic patients

The objective of this study was to detect and compare the presence of periodontopathogens in the subgingival plaques of gingivitis lesions in adults who wore fixed orthodontic appliances, as opposed to adults who did not wear any orthodontic appliances. Thirty-six individuals participated in this study. Nineteen of these subjects did not wear any orthodontic appliances, and these subjects comprised the control group. The other 17 individuals had been wearing fixed orthodontic appliances for at least 3 months each. After a periodontal examination, we collected subgingival plaque samples from the gingivitis lesions of each patient. Using PCR based on 16S rDNA, we detected the presence of 6 putative periodontopathogenic species, Treponema denticola, Porphyromonas gingivalis, Tannerella forsythia (formerly Bacteroides forsythus), Prevotella nigrescens, Prevotella intermedia, and Actinobacillus actinomycetemcomitans. With regard to the presence of individual periodontopathogens, we found that T. forsythia, T. denticola, and P. nigrescens were significantly more common in the samples obtained from the orthodontic patients than in the samples obtained from the non-orthodontic patient controls. Our results indicate that the local changes associated with the wearing of fixed orthodontic appliances may affect the prevalence of periodontopathogens in subgingival dental plaques.     

Production and characterization of monoclonal antibodies against human ceruloplasmin

Ceruloplasmin (CP) is the major plasma antioxidant and copper transport protein. Monoclonal antibodies (mAbs) against human CP were produced and characterized. A total of five hybridoma cell lines were established (CP2, CP10, CP20, CP25, CP30). From the epitope mapping analysis, two subgroups of mAbs recognize different peptide fragments were identified. When the purified CP was incubated with the mAbs, the ferroxidase activity of CP was inhibited up to a maximum 57 %. Immunoblotting with various tissue homogenates indicated that all the mAbs specifically recognize a single protein band of 130 kDa. They also appear to be extensively cross-reactive among different mammalian including human and avian sources. These results demonstrated that only one type of immunologically similar CP is present in all of the mammalian tissues including human. The CP mAbs could be of great benefit to design the diagnostic kit for CP-related diseases such as Wilson's disease.     

Karyotype of North American shortnose sturgeon Acipenser brevirostrum with the highest chromosome number in the Acipenseriformes

The shortnose sturgeon Acipenser brevirostrum was revealed to have a larger number of chromosomes than previously reported for other sturgeon species. Its chromosome number ranged from 362 to 372 (of ten specimens examined), showing intraindividual variation. The karyotype of metaphase with the highest chromosome number (372) consisted of 89 pairs of macrochromosomes and 97 pairs of microchromosomes (fundamental number; NF=550). Although the microchromosomes were relatively shorter than the macrochromosomes, most of them had discernible arms and centromeres. Silver-stained nucleolar organizer regions (Ag-NORs) were localized on the telomeric regions of 5 pairs of chromosomes (Ag-NORs=10): 4 were made up of small meta/submetacentrics and 1 of acrocentrics. Polyploidy of A. brevirostrum should be hexaploid based on the karyotype, numerous chromosomes, Ag-NORs, and previously reported large genome size (ca. 13pg DNA/cell).Supplementary material to this paper is available in electronic format at http://dx.doi.org/10.1007/s10228-004-0257-z     

Investigation of cell cycle arrest effects of actinomycin D at G1 phase using proteomic methods in B104-1-1 cells

Actinomycin D was previously reported as an inhibitor of Shc/Grb2 interaction in B104-1-1 cells. Actinomycin D arrested the cell cycle at the G1 phase at 1nM, which is about 10 times lower than the inhibition of Shc/Grb2 interactions in B104-1-1 cells. To evaluate other mechanisms of actinomycin D affected suppression of tumors and cell growth, except inhibition of Shc/Grb2 interactions, we examined the proteomic expression profile by proteomic technology. We found up-regulation of MEKK3 and down-regulation of Hsp70 expression from proteomic analysis, which is a very interesting observation because MEKK3 is strongly related with G1 arrest of cell cycle and Hsp70 is also involved in cell cycle regulation. These results indicate that the anti-tumor effects of actinomycin D is due to synergic effects of various proteins regulated by the compound including inhibition of the Shc/Grb2 interaction and other signaling pathways in the cytoplasm. Here we provide a mechanism-based explanation for growth inhibition by actinomycin D using proteomic technology. Thus, this approach may be a potentially useful method to reveal new mechanisms of active compounds or drugs with unknown cellular function.