Effect of gentamicin on CD3+ T-lymphocyte proliferation for treatment of equine recurrent uveitis: An in vitro study

Objective

The objective of the study was to determine the effect of gentamicin on CD3+ T-lymphocyte proliferation and cell viability using an in vitro cell culture model as a means of investigating the mechanism of action of low-dose intravitreal gentamicin injection.

Animals Studied

Three adult horses with no evidence of ophthalmic or systemic disease.

Procedure

Peripheral blood lymphocytes were treated with gentamicin at concentrations 37.5 μg/mL, 112.5 μg/mL, 187 μg/mL, 375 μg/mL, or 750 μg/mL then stimulated to proliferate with concanavalin A (ConA). 4′,6-diamidino-2-phenylindole (DAPI) and carboxyfluoroscein succinimidyl ester (CSFE) were used as markers of cell viability and cell proliferation, respectively. Following 5-day culture, live cell counts and CSFE fluorescent intensity data were collected via automated cell count and flow cytometry. The experimental design was duplicated using preservative-free gentamicin and a proprietary brand formulation. Statistical analysis was performed using two-way ANOVA with Tukey's multiple comparison test.

Results

No statistically significant comparisons in CD3+ T-lymphocyte live cell counts and geometric mean fluorescent intensity of CSFE were identified between gentamicin concentrations or formulations.

Conclusions

Gentamicin had no effect on equine peripheral blood CD3+ T-lymphocyte cell viability and proliferation in concentrations ranging from “safe” to “retinotoxic” in relation to intravitreal injection volumes. Low-dose intravitreal gentamicin may not suppress the Th1- and Th17-mediated immune response.     

Factors shaping genetic structure within populations ofEurya japonica (Theaceae)

Starch-gel electrophoresis was used to resolve nine polymorphic enzyme loci from leaf tissue collected from 20 Korean populations ofEurya japonica in order to determine differences in allele frequencies between male and female trees. In addition, 84 adults were sampled and mapped in a population located on Naenaro Island in Korea to examine spatial genetic structure using spatial autocorrelation analysis. Allele frequencies between males and females gave few contribution to the genetic structuring within populations. Only nine (5%) of 180 cases were significantly different from both sexes in allele frequencies. On the other hand, Moran'sI was significantly different from the expected value in 31 (23.5%) of 132 cases. In the shortest distance (0<5m),I was significantly positive in 10 (22.7%) of 44 cases. The results indicate that a significant small scale genetic structure was detected in the population and patch widths were inferred to be approximately 5–7 m. A nonrandom distribution of genotypes may be indicative of restricted gene flowvia seed and pollen dispersal, and patchy establishments of genetically distinct individuals. These factors are responsible for shaping population genetic structure ofE. japonica.     

Port‐Orford‐cedar resistant to Phytophthora lateralis

Phytophthora lateralis, an exotic root pathogen, is the primary cause of Port‐Orford‐cedar (Chamaecyparis lawsoniana) mortality throughout its native range in southwest Oregon and northwest California. Most trees in the field are very susceptible, but genetic resistance to this pathogen has been demonstrated. Since the late 1980s, the USDA Forest Service and the Bureau of Land Management in cooperation with Oregon State University have conducted an intensive programme to identify and test resistant trees from the field, and propagate them in a seed orchard with the goal of providing resistant seedlings for regeneration. Susceptible families showed only 0–10% survival using a variety of inoculation techniques. Rooted cuttings of resistant parents are seldom killed, and seedling families of these parents exhibit 25–100% survival, depending on family and inoculation technique. Symptom development on resistant trees, including sunken lesions, and resinosis as well as reduced colonization and re‐isolation success, was consistent with a hypersensitive reaction. In a long‐term field test, five resistant families had 20–80% survival after 16 years, while three susceptible families had 0–8% survival in the same interval.     

Biological evaluation of neopeptins isolated from a Streptomyces strain

BACKGROUND: Microbial secondary metabolites are a rich source of antifungal agents and have merit as alternatives to synthetic fungicides. To develop disease control agents against powdery mildew, the lipopeptide antibiotic neopeptins were identified from the culture broth of a Streptomyces sp., and in vivo control efficacy of the compounds was evaluated on cucumber plants under glasshouse conditions. RESULTS: The Streptomyces sp. KNF2047 antagonistic against powdery mildew development in cucumber plants was isolated from a soil sample. Antifungal compounds were purified from the culture broth and identified as neopeptin A and B. In vitro microtitre assays revealed the inhibitory activities of the compounds in the range 128-512 microg mL(-1) against the mycelial growth of Alternaria mali, Botrytis cinerea, Cladosporium cucumerinum, Colletotrichum lagenarium, Didimella bryoniae and Magnaporthe grisea. Although neither compound showed remarkable in vitro antifungal activity against other plant pathogenic fungi, a mixture of neopeptins (484 mg of neopeptin A and 290 mg of neopeptin B per gram of partially purified powder) showed potent protective and curative activity against cucumber powdery mildew in vivo. The disease control activity of the neopeptins at a concentration of 2.4 mg L(-1) was 92.1%, which was similar to that of the commercial fungicide fenarimol (89.3% at 63 mg L(-1)) and that of the commercial biocontrol agent Actinovate (67.4% at 2 x 10(7) cfu L(-1)). CONCLUSION: Neopeptin mixtures isolated from Streptomyces sp. KNF2047 showed potent disease control activity against powdery mildew development on cucumber plants. .     

Photoinduced generation of 2,3-butanedione from riboflavin

The volatile compound formation from riboflavin solution of a phosphate buffer (0.1 M, pH 6.5) under light for 15 h was studied by SPME-GC and SPME-GC/MS analysis. Only one major compound in the riboflavin solution was formed and increased as the light exposure time increased. The light-exposed riboflavin solution had a buttery odor. The compound of riboflavin solution under light was analyzed by gas chromatography and olfactometry. The major volatile compound eluted from the gas chromatograph had a buttery odor. The buttery odor compound was positively identified as 2,3-butanedione by a combination of gas chromatographic retention time, mass spectrum, and odor evaluation of authentic 2,3-butanedione. The addition of sodium azide, a singlet oxygen quencher, to riboflavin solution minimized the formation of the buttery odor compound. Singlet oxygen was involved in the formation of the buttery odor. The 2,3-butanedione was produced from the reaction between riboflavin and singlet oxygen. Singlet oxygen was formed from triplet oxygen by riboflavin photosensitization mechanism. This is the first reported oxidation reaction between riboflavin and singlet or triplet in food and biological systems.     

Age-related changes in calretinin-immunoreactive periglomerular cells in the rat main olfactory bulb

The changes of calretinin (CR)-immunoreactive periglomerular cells in the glomerular layer of the main olfactory bulb (MOB) were investigated in rats differing ages from postnatal month 1 (PM 1) to PM 24. The number of cresyl violet-positive periglomerular cells was similar between PM 1 and PM 12, but they decreased slightly in the PM 24 group. The size of CR-immunoreactive periglomerular cells in the glomerular layer increased with age, while their numbers did not change significantly in the PM 6-PM 24 groups. In the PM 24 group, numbers of CR-positive periglomerular cell bodies and their processes decreased, while the size of CR-positive cell bodies in the glomeruli was larger than that of the previous groups. These results suggest that CR-immunoreactive periglomerular cells in the rat MOB are well-developed in the PM 6 group, and that periglomerular cells in the PM 24 group show poor CR-immunoreactivity compared to those in the PM 6 group.     

Manufacture and properties of Miscanthus–wood particle composite boards

Miscanthus sacchariflorus straw was used as a raw material for the manufacture of Miscanthus–wood particle composite board with Douglas-fir particles in ratios of 100/0, 80/20, 60/40, 40/60, 20/80, and 0/100. A commercial phenol–formaldehyde resin was used as a binder at 9 and 11 % for target densities of 0.50 and 0.65 g/cm³, respectively. The effects of the Miscanthus/wood particle ratio on the composite board properties were investigated. In addition, the density profile was also examined to improve the understanding of the composite board manufacturing process. Results indicate that the internal bonding value increased drastically in the board containing up to 50 % wood particles, providing a valuable parameter for subsequent research. The board properties were greatly improved with increasing density and binder addition level.     

Sound absorption capability and mechanical properties of a composite rice hull and sawdust board

Rice hull–sawdust composite boards were manufactured for sound-absorbing boards in construction. The manufacturing parameters were target density (400, 500, 600, and 700?kg/m³) and rice hull content as percent weight of rice hull/sawdust/phenol resin (10/80/10, 20/70/10, 30/60/10, and 40/50/10). Commercial gypsum board and fiberboard were also used as comparative sound-absorbing materials. The average modulus of rupture (MOR) of the board with a density of 700?kg/m³ and rice hull mixing ratio of 10% was 8.6?MPa, and that of the board with a 400?kg/m³ board density and a rice hull mixing ratio of 40% was 2.2?MPa. The MOR increased with increasing board density or decreasing rice hull mixing ratio. The sound absorption coefficients of some boards (400?kg/m³ and 10%, 500?kg/m³ and 30%, and 500?kg/m³ and 40%) were better than those of the commercial 11-mm-thick gypsum board. Thus, it is concluded that rice hull–sawdust composite boards may be implemented as sound-absorbing barriers in construction due to their high sound absorption coefficients.     

Pro-apoptotic effects of tepoxalin, a cyclooxygenase/lipoxygenase dual inhibitor, on canine synovial fibroblasts

Canine osteoarthritis occurs frequently and causes secondary synovitis. Administration of nonsteroidal anti-inflammatory drugs (NSAIDs) is one of the major therapeutic options for pain management of joint diseases. Tepoxalin has an unique property as an NSAIDs that suppresses both cyclooxygenase and lipoxygenase. The purpose of this study was to evaluate antiproliferative effects of tepoxalin on cultured canine synovial cells. Cytotoxic effects of tepoxalin, carprofen, meloxicam and AA-861 on cultured canine synoviocytes were evaluated by MTT colorimetric assay. Apoptosis was detected by morphological observations with Giemsa or annexin V/Hoechst 33342 staining and by the inhibition of caspase-3 activity with N-Ac-Asp-Glu-Val-Asp-CHO (Ac-DEVD-CHO). Cytotoxic effects of tepoxalin were evident in comparison with the effects of carprofen or meloxicam. The same tendency of cytotoxicity was observed when 5-lipoxygenase was inhibited by AA-861. The morphological findings and contradictory effects of Ac-DEVD-CHO with regard to the cytotoxicity proved the proapoptotic effects of tepoxalin. In conclusion, tepoxalin might control osteoarthritic synovitis by inducing apoptosis in proliferating synoviocytes, while most NSAIDs that selectively inhibit cyclooxygenase-2 most likely would not suppress synovial proliferation.     

Silk fibroin hydrolysate exerts an anti-diabetic effect by increasing pancreatic β cell mass in C57BL/KsJ-db/db mice

Components of silk including silk fibroin have long been used as anti-diabetic remedies in oriental medicine. However, detailed mechanisms underlying these anti-diabetic effects remain unclear. In this study, we examined the anti-diabetic activity of silk fibroin hydrolysate (SFH) in C57BL/KsJ-db/db (db/db) mice, a well-known animal model of non-insulin dependent diabetes mellitus. When the db/db mice were administered SFH in drinking water for 6 weeks, hyperglycemia in the animals gradually disappeared and the level of glycosylated hemoglobin decreased, indicating that SFH plays important role in reducing the symptoms of diabetes. In addition, SFH-treated db/db mice exhibited improved glucose tolerance with increased plasma insulin levels. Immunohistochemical and morphological analyses showed that SFH up-regulated insulin production by increasing pancreatic β cell mass in the mice. In summary, our results suggest that SFH exerts anti-diabetic effects by increasing pancreatic β cell mass in a non-insulin dependent diabetes mellitus mouse model.