Role of BMP, betaig-h3, and chitosan in early bony consolidation in distraction osteogenesis in a dog model

The purpose of this investigation was to study the effect of bone morphogenetic protein (BMP), transforming growth factor beta-induced gene h3 (betaig-h3), and chitosan on early bony consolidation in distraction osteogenesis in a dog model. Sixteen dogs were used for this study. The lateral surface of the mandibular body was exposed in the subperiosteal plane and the vertical osteotomy on the mandibular body was extended downward. An external distraction device was applied to the mandibular body, and the mandibular distraction was started 5 days after the operation at a rate of 2 mm/day up to a 10-mm distraction after 5 days. The experimental group was then divided into a control group, a BMP group, a betaig-h3 group, and a chitosan group, depending on the type of implantation material used in the distracted area. On the same day after completing the distraction, BMP, betaig-h3, or chitosan was implanted into the distracted area. No material was implanted into the distracted area in the control group. After implanting the materials, the distraction device was left in place for 7 weeks to allow for bony consolidation. Four dogs were allocated to each group. Two dogs in each group, a total of eight dogs, were killed 4 weeks after completing the distraction and the other eight dogs were killed after 7 weeks. Serial radiographs were obtained every week after completing the distraction. New bone was generated in the distracted zone in all groups. In the BMP group, the formation of active woven bone was observed throughout the distracted zone, and the new bone appeared to be nearly normal cortical bone 7 weeks after implantation. In the betaig-h3 and chitosan groups, the development of new bone was observed in the distracted zone after 7 weeks; however, the amount was less than that in the BMP group. In the control group, the new bone was observed at the edges of the distracted zone. These findings suggest that BMP seems to be very effective in early bony consolidation in distraction osteogenesis.     

Monitoring flap for buried free tissue transfer: its importance and reliability

To improve the success rate of microsurgical flap transfers into a buried area, it is important to monitor the circulation of the flap during the early stage. A monitoring flap includes such advantages as simplicity, reliability, noninvasiveness, and the ability to continuously monitor the vascular status of various buried flaps. This article describes experiences related to the importance and reliability of a monitoring flap. A total of 109 flaps in 99 patients were treated with buried free flaps, including a monitoring flap, between 1990 and 1999. Forty-nine patients received a tubed free radial forearm flap with a skin-monitoring flap, and six received a free jejunal flap with a jejunal segment monitoring flap for the reconstruction of the esophagus. Vascularized fibular grafts with a skin monitoring flap or peroneus longus muscle monitoring flap were used for reconstructing the mandible in six patients and for treating osteonecrosis of the femoral head in 48 flaps in 38 patients. Monitoring flap abnormalities were indicated in 14 flaps; therefore, immediate revisions were performed on the pedicle of the monitoring flap and microanastomosis site. Among these 14 flaps, nine showed true thrombosis and five showed false-positive thrombosis. Among the nine flaps that showed true thrombosis, five were salvaged and four were finally lost. The false-positive thrombosis in the five flaps was attributed to torsion or tension of the perforator of the monitoring flap in three flaps, an unclear determination in one flap because the monitoring flap size was too small, and damage to the perforator in the last flap. The total thrombosis rate was 8.3 percent (nine of 109), and the failure rate of the free tissue transfer was 3.7 percent (four of 109). The overall sensitivity of the monitoring flap was 100 percent, the predictive value of a positive test was 64 percent (nine of 14), and false-positive results occurred in 36 percent (five of 14). The salvage rate was 55.6 percent. To improve the reliability of a monitoring flap, it is recommended that the size of the flap be larger than 1 x 2 cm to assess the arterial status, and that a perforator with the appropriate caliber be selected. When a monitoring flap is fixed to a previous incision line or a newly created wound, any torsion or tension of the perforator should be avoided. In conclusion, the current results suggest that a monitoring flap is a simple, extremely useful, and reliable method for assessing the vascular status of a buried free flap.     

ProNGF induces p75-mediated death of oligodendrocytes following spinal cord injury

The neurotrophin receptor p75 is induced by various injuries to the nervous system, but its role after injury has remained unclear. Here, we report that p75 is required for the death of oligodendrocytes following spinal cord injury, and its action is mediated mainly by proNGF. Oligodendrocytes undergoing apoptosis expressed p75, and the absence of p75 resulted in a decrease in the number of apoptotic oligodendrocytes and increased survival of oligodendrocytes. ProNGF is likely responsible for activating p75 in vivo, since the proNGF from the injured spinal cord induced apoptosis among p75(+/+), but not among p75(-/-), oligodendrocytes in culture, and its action was blocked by proNGF-specific antibody. Together, these data suggest that the role of proNGF is to eliminate damaged cells by activating the apoptotic machinery of p75 after injury.     

Drosophila lacking dfmr1 activity show defects in circadian output and fail to maintain courtship interest

Fragile X mental retardation is a prominent genetic disorder caused by the lack of the FMR1 gene product, a known RNA binding protein. Specific physiologic pathways regulated by FMR1 function have yet to be identified. Adult dfmr1 (also called dfxr) mutant flies display arrhythmic circadian activity and have erratic patterns of locomotor activity, whereas overexpression of dFMR1 leads to a lengthened period. dfmr1 mutant males also display reduced courtship activity which appears to result from their inability to maintain courtship interest. Molecular analysis fails to reveal any defects in the expression of clock components; however, the CREB output is affected. Morphological analysis of neurons required for normal circadian behavior reveals subtle abnormalities, suggesting that defects in axonal pathfinding or synapse formation may cause the observed behavioral defects.     

Green fluorescent protein-labeled recombinant fluobody for detecting the picloram herbicide

A green fluorescent protein-labeled fluobody was designed to develop a simple immunoassay method for detecting picloram herbicide in an environmental sample. The gfp gene was successfully inserted into the pSJF2 vector harboring the picloram-specific antibody fragment to yield pSJF2GFP. Picloram spiking in an environmental river sample could be indirectly detected by observing the fluorescence intensity value of the gfp-fluobody, exhibiting specific sensitivity to free picloram with an IC50 value of 50 ppb. Using the gfp-fluobody immunoassay avoids the enzyme-substrate reaction for calorimetric detection that is required in an enzyme-linked immunosorbent assay (ELISA).     

Anticoagulant from Taraxacum platycarpum

An anticoagulant was purified from a Chinese herb, Taraxacum platycarpum. Its activity was heat-labile, and was decreased by incubation with subtilisin Carlburg or proteinase K, indicating that the active component was a protein. The protein had a molecular mass of 31 kDa by gel filtration and 33 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis, so it probably was a monomer. When present at the concentration of 70, 255, and 873 nM, respectively, the protein doubled the thrombin time, prothrombin time, and activated partial thromboplastin time. It inhibited thrombin and kallikrein, but did not hydrolyze fibrinogen. The protein bound the anion-binding exosite of thrombin, competing with the fibrinogen binding site. In addition, the protein caused the murine macrophage cell line Raw 264.7 to produce cyclooxygenase-2, nitric oxide synthase, nitric oxide, and tumor necrosis factor-alpha.     

Endophytic bacillus sp. isolated from the interior of balloon flower root

A bacterial strain, designated CY22, was isolated from the interior of balloon flower (Platycodon grandiflorum) root in the Republic of Korea. The isolate coproduced an iturin-like antifungal compound and a surfactin-like potent biosurfactant. Analysis of the 16S-rDNA of strain CY22 showed that the isolate was a member of Bacillus. High similarities were observed between strain CY22 and Bacillus sp. TKSP 24, and between strain CY22 and B. subtilis 168. Phylogenetic analysis based on 16S-rDNA sequences showed that strain CY22 was closely related to Bacillus sp. The main whole-cell fatty acids were anteiso-C15:0 (37%), C17:0 (5.1%), and iso-C15:0 (27.7%). DNA G+C content was 54 mol%. Based on phylogenetic inference, phenotypic and chemotaxonomic characteristics, this endophytic strain Bacillus sp. CY22 was assigned to the genus Bacillus.     

Procedures for the axenic isolation of conchocelis and monospores from the red seaweed Porphyra yezoensis

In order to maintain axenic seedstock cultures axenically of thecommercially important red seaweed, Porphyra yezoensis, aprocedure was developed for axenic isolation and culture of conchocelis andmonospores. For axenic isolation of the conchocelis, contaminated microalgaewere most effectively removed by filtering contaminated samples through a100-m mesh after sonication. Removal of bacteria and otheralgaewas accomplished using a mixture of 5 agents (0.02% chitosan, 100 gml^–1 GeO₂, 10 gml^–1 ampicillin, 40 gml^–1 kanamycin and 200 gml^–1 streptomycin). Axenic single colonies wereisolatedfrom a semi-solid medium prepared from 1% transfer gel. After collectingmonospores from the 40–50% density layer on a percoll-gradient, removalofbacteria and fungi from the monospores was accomplished using a mixture of 5antibiotics (3.5 g ml^–1 nystatin, 2 mgml^–1 ampicillin, 400 gml^–1 kanamycin, 50 gml^–1 neomycin and 800 gml^–1 streptomycin). Axenic single juvenile blades wereisolated from a semi-solid medium prepared from 0.5% transfer gel.     

A procedure for axenic isolation of the marine microalga Isochrysis galbana from heavily contaminated mass cultures

Isochrysis galbana, one of the most widely usedmarine microalgae in the rearing of finfish and shellfish larvae, is masscultured frequently in outdoor tanks. Under prolonged and repeated culture,severe contamination occurs. Axenic isolation of I.galbanafrom such cultures was best achieved by using a ternary procedure involvingpercoll-gradient centrifugation, treatment with antibiotics, and growth on agarmedium. Protozoa and other algae were removed most effectively by isolation ofI. galbana at the 30–40% density layer on apercoll-gradient. Removal of bacteria was accomplished using a mixture of 5antibiotics (250 g mL^–1 ampicillin, 50g mL^–1 gentamycin, 100 gmL^–1 kanamycin, 500 gmL^–1 neomycin, 50 gmL^–1 streptomycin). Axenic colonies were isolated fromasolid medium prepared from 1% purified agar. The ternary procedure isconsideredapplicable to the isolation of other axenic single-celled microalgae fromheavily contaminated cultures.     

Proinflammatory cytokine profile in Vibrio vulnificus septicemic patients' sera

Vibrio vulnificus causes a fulminant and frequently fatal septicemia in susceptible hosts. The present study was designed to evaluate the proinflammatory cytokine profile in V. vulnificus septicemia patients' sera and the effect of doxycycline therapy on the levels of proinflammatory cytokines. Levels of proinflammatory cytokines, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta and IL-6, were measured in the sera of V. vulnificus septicemic patients and normal healthy volunteers using colorimetric sandwich ELISA. The mean values of TNF-alpha, IL-1beta and IL-6 in the sera of V. vulnificus patients (n=33) increased by 210-, 232- and 40-fold in comparison with those of normal healthy volunteers (n=5), but only the IL-6 level showed a statistically significant difference (P<0.05) between the two groups. Sera from the cases for which doxycycline treatment histories were obvious were designated 'before-treatment' (TX). All the others were included in the after-TX group. In the before-TX group (n=5), the levels of TNF-alpha and IL-1beta significantly increased (P<0.05) in comparison with the after-TX group (n=5). IL-6 levels in the two groups showed no difference. In conclusion, the levels of the well known proinflammatory cytokines TNF-alpha, IL-1beta and IL-6 increased in the V. vulnificus septicemic patients' sera, and the levels of TNF-alpha and IL-1beta decreased significantly after doxycycline treatment. These data indicate that proinflammatory cytokines might play a critical role in V. vulnificus septicemia like in other endotoxemic shocks. The use of doxycycline as an effective bactericidal agent and as an effective modulator of proinflammatory cytokines is supported.