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Objective This study was realized to evaluate the nutritional value of rice grains as a replacement for corn grains in the diet of growing Hanwoo steers. Methods Two experimental diets were prepared: i) Corn total mixed ration (TMR) consisting of 20% corn grains and ii) Rice TMR consisting of 20% rice grains, in a dry matter (DM) basis. These treatments were used for in vitro rumen fermentation and in vivo growth trials. In the rumen fermentation experiment, the in vitro DM digestibility (IVDMD), in vitro crude protein digestibility (IVCPD), in vitro neutral detergent fiber digestibility, pH, ammonia nitrogen, and volatile fatty acids (VFA) were estimated at 48 h, and the gas production was measured at 3, 6, 12, 24, and 48 h. Twenty four growing Hanwoo steers (9 months old; body weight [BW]: 259±13 kg) were randomly divided into two treatment groups and the BW, dry matter intake (DMI), average daily gain (ADG), and feed conversion ratio (FCR) were measured. Results The in vitro experiment showed that the IVDMD, IVCPD, and VFA production of the Rice TMR were higher than those of the Corn TMR (p0.05) in the final BW, ADG, DMI, and FCR between the two TMRs. Conclusion The use of rice grains instead of corn grains did not exhibit any negative effects on the rumen fermentation or growth performance, thereby rice grains with a DM of less than 20% could be used as a starch source in the diet of growing steers.

The efficacy of ethanol extract ofDiospyros kaki(EEDK) on dry eye (DE) was determined using an experimental mouse model. Experimental groups included three treated with various amounts of EEDK and one treated with omega-3 for 2 weeks. Damage to the ocular surface was evaluated, and the presence of conjunctival goblet cells was determined. Moreover, the inflammatory response was analyzed via RT-PCR analysis and a reporter gene assay. Fluorescein staining intensity decreased in the EEDK treatment group, and goblet cell density increased significantly in a dose-dependent manner. Pro-inflammatory cytokines were upregulated in human corneal epithelial cells treated with Pam3Cys-Ser-(Lys)-4. However, pro-inflammatory cytokines were downregulated at the mRNA level upon treatment with EEDK. Furthermore, EEDK regulated Pam3CSK4-induced gene expression through interferon regulatory factors. EEDK effectively improves the conjunctival goblet cell density and reduces the inflammatory response by reducing interferon regulatory factor activation downstream of Toll-like receptors in DE. Therefore, EEDK could be beneficial agents for preventing and treating DE.

Ethnopharmacological relevance In contrast to other leguminous plants generally used as food, Rhynchosia volubilis Loureiro, a small soybean with a black seed coat, has been used as a traditional oriental remedy for various human diseases in Eastern Asia. In this study, we demonstrated the protective effect of R. volubilis against dry eye disease. Aim of the study We aimed to investigate whether a standardized ethanol extract of R. volubilis (EERV) can protect the cornea in a benzalkonium chloride (BAC)-induced mouse dry eye model. Materials and methods Experimental dry eye was induced by the instillation of 0.2% BAC on mouse cornea. A standardized ethanol extract of R. volubilis (EERV) was orally administered following BAC treatment. The positive control group was treated with commercial eye drops. Fluorescein staining, tear break-up time (BUT), and hematoxylin and eosin staining were evaluated on the ocular surface. Squamous metaplasia and apoptosis in the corneal epithelial layer were detected by immunostaining. Furthermore, the protein expression of cytochrome c, Bcl-2, and Bax was determined. Results EERV treatment significantly improved fluorescein scoring, BUT, and smoothness in the cornea compared to the vehicle group. In addition, EERV inhibited squamous metaplasia and apoptosis in the cornea. The expression of cytochrome c and Bax was upregulated, while that of Bcl-2 was downregulated in the vehicle group compared with that in the control group. However, EERV treatment inhibited the expression of cytochrome c and Bax, while that of Bcl-2 was improved. Conclusion Standardized EERV could be a beneficial candidate for the treatment of dry eye disease.

Purpose: To evaluate the role of intravitreal aflibercept (IVA) for treating persistent central serous chorioretinopathy (CSCR) of more than 6 weeks. Methods: AFlibercept Efficacy in Cscr Treatment (NCT01971190) was a 6-month, phase 2, multicenter, prospective, randomized, controlled clinical trial at 6 clinical sites in Korea. Forty-three patients were randomized into group 1 (29) with 3 monthly IVA administrations or group 2 (14) with sham treatment and followed for 6 months with pro re nata (PRN) IVA. Patients could be retreated with IVA when they met the eligibility criteria for this. The primary end point was a mean change in central subfield thickness (CST) from baseline. Best-corrected visual acuity (BCVA) and CST were checked monthly. Data from a subgroup with long-term follow-up in a real-life practice were also compared. Results: After 3 months of treatment, group 1 had a greater change in mean CST (173.97 vs 80.15 µm; P = .001) and BCVA improvement (10.38 vs 8.62 Early Treatment of Diabetic Retinopathy Study letters; P = .52) than group 2. Among patients whose disease had lasted ≥3 months, BCVA improvement was significantly better in group 1 than in group 2 after 3 months (12.0 vs −2.7 letters; P = .007), and the percentage of eyes requiring PRN IVA was lower in group 1 (50% vs 100%). These differences were attenuated at 6 months but maintained during additional 6-month follow-up. Conclusion: Intravitreal aflibercept may facilitate the anatomical improvement in persistent CSCR and may improve the visual outcomes in relatively chronic cases. The IVA injection may be a therapeutic option for CSCR.

Objective To evaluate prognostic factors associated with good response to intravitreal bevacizumab (IVB) treatment in central serous chorioretinopathy (CSC) patients. Methods We retrospectively reviewed ninety-four eyes of 94 CSC patients who received intravitreal bevacizumab injection(IVB) (0.05 ml, 1.25 mg) as a first line treatment. Patients were divided into two groups as good responders and poor responders based on optical coherence tomography (OCT) findings 1 month after the first injection of intravitreal bevacizumab. Good responders were defined as complete resolution of subretinal fluid (SRF) on OCT 1 month after IVB. We compared baseline visual acuity, duration of symptoms, angiographic findings on fluorescein angiography, central retinal thickness (CRT), macular volume (MV),subfoveal choroidal thickness (SFCT) between two groups. Results 42 eyes were included as good responder and 52 eyes as poor responders. Mean age was 47.8±7.4 year old ingood responders and 50.6±7.8 year old in poor responders. Baseline visual acuity (logMAR) was better in good responders than poor responders, 0.19±0.19 and 0.29±0.19 respectively (p=0.015). SFCT was significantly thicker in good responders than poor responders 419.7±103.2㎛ and 376.0±99.0㎛ respectively (p=0.040). Prevalence of chronic cases was higher in poor responders (33/52) than good responders (15/42), (p=0.012). There was no statistical significance in age, sex, CRT, MVbetween two groups. Conclusion Intravitreal injection of bevacizumab can be potential option for the treatment of CSC patients, especially in case of recent onset, better baseline visual acuity with thicker choroid on OCT.

The purpose of the study was to investigate the protective effects of the ethanol extract of Diospyros kaki (EEDK) persimmon leaves to study N-methyl-N-nitrosourea (MNU)-induced retinal degeneration in mice. EEDK was orally administered after MNU injection. Retinal layer thicknesses were significantly increased in the EEDK-treated group compared with the MNU-treated group. The outer nuclear layer was preserved in the retinas of EEDK-treated mice. Moreover, EEDK treatment reduced the MNU-dependent up-regulation of glial fibrillary acidic protein (GFAP) and nestin expression in Müller and astrocyte cells. EEDK treatment also inhibited MNU-dependent down-regulation of rhodopsin expression. Quercetin exposure significantly attenuated the negative effects of H2O2 in R28 cells, suggesting that quercetin can act in an antioxidative capacity. Thus, EEDK may be considered as an agent for treating or preventing degenerative retinal diseases, such as retinitis pigmentosa and age-related macular degeneration.

This study aimed to investigate the levels of aqueous humour cytokines in the eyes of patients with central serous chorioretinopathy (CSC), who received an intravitreal injection of bevacizumab, and to evaluate the differences in cytokine levels between bevacizumab responders and non-responders.Aqueous humour samples were collected from 39 eyes of 39 patients with CSC and 25 eyes of 25 control patients undergoing cataract surgery and were analysed using Multiplex bead assays. We compared the cytokine levels in patients with CSC (responders and non-responders) and control subjects. After intravitreal injections of bevacizumab in patients with CSC, we evaluated the changes in visual acuity and optical coherence tomographic parameters.Significantly increased levels of interferon gamma-induced protein (IP-10) were detected in the aqueous humour of eyes from patients with CSC compared to controls (p = 0.036). Vascular endothelial growth factor (VEGF) levels were significantly higher in bevacizumab responders than in non-responders (p = 0.021). The concentrations of the other cytokines investigated did not differ significantly between patients with CSC and control subjects or between responders and non-responders.The results of the present study suggest that IP-10 and VEGF may contribute to the pathogenesis of CSC. It is clear that intravitreal bevacizumab was helpful in some of the patients with CSC, who showed higher levels of VEGF in the aqueous humour. However, the true effect and the rationale of anti-VEGF injection treatment in patients with CSC remain to be elucidated. Our results suggest that patients with CSC may be more susceptible to VEGF and/or other cytokines than the normal population.

To determine whether intravitreally-injected baicalin inhibits the growth of choroidal neovascularization (CNV) experimentally induced via laser photocoagulation through analysis of angiogenic factors.Six CNVs were induced in the left eyes of 8-week-old male Brown Norway rats. Immediately after the induction of CNV, 4 μl of baicalin solution (0.1, 1 or 5 nmol) and 4 μl of a solution containing 100 μg of bevacizumab were slowly injected into the vitreous cavity under direct observation with an operating microscope. At 14 days after CNV induction, fluorescein angiography (FA) was performed, and choroidal flat mounts were produced for quantitative assessment of CNV. The levels of the anti-angiogenic proteins vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF) and matrix metalloproteinase-2 (MMP-2) were determined via Western blot analysis.FA of bevacizumab- and baicalin-treated rats showed significantly reduced CNV and leakage from the CNV lesions compared to control rats at day 14. Choroidal flat mounts revealed that baicalin inhibited the growth of CNV lesions in a dose-dependent manner. Western blot analysis demonstrated that baicalin significantly attenuated the up-regulation of VEGF, PDGF and MMP-2.Baicalin suppressed laser-induced CNV formation in rats. These results suggest that baicalin should be considered as a candidate drug for treating exudative age-related macular degeneration.

Piceid is widely used in food, cosmetics, and pharmaceuticals because of its therapeutic benefits. However, the use of piceid as a drug is limited because of its low solubility. To increase solubility, we synthesized piceid glucosides using maltosyltransferase from Caldicellulosiruptor bescii. The MTase gene was cloned and expressed in Escherichia coli. The enzyme had a unique transfer specificity to the transfer of maltosyl units. Four piceid transglycosylation products were present and identified by thin-layer chromatography and recycling preparative high-performance liquid chromatography. The major product was purified by C(18) and gel filtration chromatography, and its molecular structure was determined using nuclear magnetic resonance spectroscopy to be α-D-maltosyl-(1→4)-piceid. The solubility of maltosyl piceid was 8.54 × 10(3) and 1.86 × 10(3) times those of natural resveratrol and piceid, respectively, suggesting that the transglycosylation greatly increased the water solubility. This suggests that dietary intake of this compound can enhance the bioavailability of resveratrol in the human body.

Caldicellulosiruptor bescii DSM 6725 utilizes various polysaccharides and grows efficiently on untreated high-lignin grasses and hardwood at an optimum temperature of 80C. It is a promising anaerobic bacterium for studying high-temperature biomass conversion. Its genome contains 2666 protein- coding sequences organized into 1209 operons. Expression of 2196 genes (83%) was confirmed ex- perimentally. At least 322 genes appear to have been obtained by lateral gene transfer (LGT). Putative functions were assigned to 364 conserved/hypothetical protein (C/HP) genes. The genome contains 171 and 88 genes related to carbo- hydrate transport and utilization, respectively. Growth on cellulose led to the up-regulation of 32 carbohydrate-active (CAZy), 61 sugar transport, 25 transcription factor and 234 C/HP genes. Some C/HPs were overproduced on cellulose or xylan, suggesting their involvement in polysaccharide conversion. A unique feature of the genome is en- richment with genes encoding multi-modular, multi-functional CAZy proteins organized into one large cluster, the products of which are proposed to act synergistically on different components of plant cell walls and to aid the ability of C. bescii to convert plant biomass. The high duplication of CAZy domains coupled with the ability to acquire foreign genes by LGT may have allowed the bacterium to rapidly adapt to changing plant biomass-rich environments.

The thermophilic, cellulolytic, anaerobic bacterium ‘Anaerocellum thermophilum’ strain Z-1320 was isolated from a hot spring almost two decades ago and deposited in the German Collection of Microorganisms and Cell Cultures (DSMZ) as DSM 6725. The organism was classified as representing a new genus, ‘Anaerocellum’, primarily on its growth physiology, cell-wall type and morphology. The results of recent physiological studies and of phylogenetic and genome sequence analyses of strain DSM 6725 of ‘A. thermophilum’ obtained from the DSMZ showed that its properties differed from those originally described for strain Z-1320. In particular, when compared with strain Z-1320, strain DSM 6725 grew at higher temperatures and had an expanded range of growth substrates. Moreover, the 16S rRNA gene sequence of strain DSM 6725 fell within the Caldicellulosiruptor clade. It is therefore suggested that ‘Anaerocellum thermophilum’ should be classified as a member of the genus Caldicellulosiruptor, for which the name Caldicellulosiruptor bescii sp. nov. is proposed (type strain DSM 6725T=ATCC BAA-1888T). C. bescii sp. nov. DSM 6725T is the most thermophilic cellulose-degrading organism known. The strain was able to grow up to 90 °C (pH 7.2) and degraded crystalline cellulose and xylan as well as untreated plant biomass, including potential bioenergy plants such as poplar and switchgrass.

To investigate laser photocoagulation and intravitreal bevacizumab efficacy and safety in patients with moderate-to-severe stage 3 retinopathy of prematurity (ROP), and to evaluate the effects of treatment on the development of peripheral retinal vessels.A retrospective chart review of 15 premature babies, all of whom were diagnosed with stage 3 ROP, was conducted. Patients with moderate-to-severe stage 3 ROP, thus with a vascular-active ROP, received intravitreal injections of bevacizumab (0.5 mg/0.02 ml) and laser photocoagulation, whereas those with relatively inactive ROP received laser photocoagulation only. Patients were examined 1, 2, 4, and 8 weeks after treatment, or until peripheral retinal vessel growth over the laser scar was noted.Both eyes of 15 patients diagnosed with moderate-to-severe stage 3 ROP were evaluated. Eight patients (n=16 eyes) received intravitreal bevacizumab injection and laser photocoagulation, and seven patients (n=14 eyes) received laser photocoagulation only. During the follow-up period, regression of plus disease and peripheral retinal vessel development appeared significantly more rapidly in patients who received both intravitreal bevacizumab injection and laser photocoagulation. Peripheral retinal vessel development over the laser scar was identified 1-2 weeks after treatment. No systemic or significant ocular complications, such as vitreous hemorrhage, retinal detachment, or endophthalmitis, were noted during follow-up after treatment.A combination of laser photocoagulation and intravitreal bevacizumab injection seems to be a safe and effective therapy in patients with moderate-to-severe stage 3 ROP, promulgating rapid development of peripheral retinal vessels.

Very few cultivated microorganisms can degrade lignocellulosic biomass without chemical pretreatment. We show here that “ Anaerocellum thermophilum ” DSM 6725, an anaerobic bacterium that grows optimally at 75°C, efficiently utilizes various types of untreated plant biomass, as well as crystalline cellulose and xylan. These include hardwoods such as poplar, low-lignin grasses such as napier and Bermuda grasses, and high-lignin grasses such as switchgrass. The organism did not utilize only the soluble fraction of the untreated biomass, since insoluble plant biomass (as well as cellulose and xylan) obtained after washing at 75°C for 18 h also served as a growth substrate. The predominant end products from all growth substrates were hydrogen, acetate, and lactate. Glucose and cellobiose (on crystalline cellulose) and xylose and xylobiose (on xylan) also accumulated in the growth media during growth on the defined substrates but not during growth on the plant biomass. A. thermophilum DSM 6725 grew well on first- and second-spent biomass derived from poplar and switchgrass, where spent biomass is defined as the insoluble growth substrate recovered after the organism has reached late stationary phase. No evidence was found for the direct attachment of A. thermophilum DSM 6725 to the plant biomass. This organism differs from the closely related strain A. thermophilum Z-1320 in its ability to grow on xylose and pectin. Caldicellulosiruptor saccharolyticus DSM 8903 (optimum growth temperature, 70°C), a close relative of A. thermophilum DSM 6725, grew well on switchgrass but not on poplar, indicating a significant difference in the biomass-degrading abilities of these two otherwise very similar organisms.

Background and Objective:To compare the safety and effectiveness of 25-gauge transconjunctival sutureless vitrectomy (TSV) with conventional 20-gauge pars plana vitrectomy (PPV) for the treatment of vitreoretinal complications in patients with diabetic retinopathy.Patients and Methods:Forty-five eyes of 44 consecutive patients with diabetes mellitus who had undergone 25-gauge TSV for the treatment of vitreous hemorrhage, traction retinal detachment, or diabetic macular edema and 45 control patients with diabetes mellitus who had undergone conventional 20-gauge PPV were studied.Results:The mean duration of surgery in the 25-gauge TSV group was statistically significantly shorter than that in the 20-gauge PPV group. The mean visual acuity at both 1 week and 1 month after surgery was statistically significantly better in the 25-gauge TSV group than in the 20-gauge PPV group. Complications in the 25-gauge TSV group were similar to those in the 20-gauge PPV group.Conclusion:A shorter operation time and faster recovery of postoperative visual acuity were the main advantages of using 25-gauge TSV. Despite its limitations, 25-gauge TSV can be safely used for the treatment of vitreoretinal complications in patients with diabetes mellitus.

OBJECTIVE: To review the epidemiology and prognostic factors related to visual outcome after open globe injuries. METHODS: A retrospective case review during a 14-year period. RESULTS: The study was based on a sample of 321 patients (321 eyes) with open globe injuries. Gender of the sample was predominantly male (76.6%) and the average age was 38.8 years. Injuries occurred most frequently in autumn (37.7%) and in the work place (45.8%). The anatomic site of the wound was corneal (zone I) in 148 eyes, corneoscleral (zone II) in 61 eyes, and scleral (zone III) in 92 eyes. Initial visual acuity was 4/200 or worse in 77.9% of patients and was significantly correlated with the zone of injury (linear by linear association p = 0.000). The elapsed time from injury to surgery was shorter than 48 hours in 90.7%. Primary repair was performed in 301 eyes and, subsequently, pars plana vitrectomy in 165 eyes, and cataract extraction in 67 eyes. Anatomic success was achieved in 91% of eyes and evisceration was performed in 7.8% (25 eyes). Final visual acuity of 5/200 or better was achieved in 58.3% of patients, and vision of 20/40 or better in 36.5%. The zone of injury was the most significant predictor of functional outcome (linear by linear association p = 0.000) as well as anatomic outcome (chi p = 0.000). Associated injuries such as retinal detachment, vitreous hemorrhage, uveal prolapse, and adnexal injury were also important prognostic factors (linear by linear association, p = 0.000). CONCLUSIONS: In this study, we identified the epidemiology and prognostic factors of open globe injuries in a Korean urban community. The visual as well as anatomic outcome for open globe injury was shown to be reasonably favorable when prompt and appropriate surgical management was applied. Language: en

Purpose In this study, the beneficial effects of the oral administration of ethanol extract of Diospyros kaki (EEDK) were tested on a mouse dry eye model induced by benzalkonium chloride (BAC). Methods A solution of 0.2% BAC was administered topically to mouse eyes for 14 days, twice daily, to induce dry eye. Various concentrations of EEDK were administrated daily by oral gavage for 14 days after BAC treatment. Preservative-free eye drops were instilled in the positive-control group. The tear secretion volume (Schirmer’s test), tear break-up time (BUT), and fluorescein score were measured on the ocular surface. BAC-induced corneal damage was tested with hematoxylin-eosin staining. Moreover, apoptotic cell death in the corneal epithelial layer was investigated with terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining. The protein expression level of interleukin-1alpha (IL-1α), IL-1β, IL-6, tumor necrosis factor- alpha (TNF-α), and monocyte chemotactic protein-1 (MCP-1) was determined with western blot analysis. Furthermore, squamous metaplasia in the corneal epithelial layer was detected with immunofluorescent staining for cytokeratine-10. The cellular proliferation in the cornea was examined with immunohistochemical staining for Ki-67. Results EEDK treatment resulted in prolonged BUT, decreased fluorescein score, increased tear volume, and smoother epithelial cells compared with BAC treatment alone in the cornea. Moreover, EEDK treatment inhibited the inflammatory response and corneal epithelial cell death in a BAC-induced murine dry eye model, and changes in squamous cells were inhibited. Proliferative activity in the corneal epithelium cells was improved with EEDK. Conclusions EEDK could be a potential therapeutic agent in the clinical treatment of dry eye.

Aim. To analyze the subfoveal choroid thickness and choroidal volume in unilateral ocular ischemic syndrome (OIS).Methods. A retrospective review was conducted for all patients with unilateral OIS from October 2010 through June 2014. The subfoveal choroidal thickness (SFChT) and choroidal volume of both eyes were compared.Results. 19 unilateral OIS patients were included in this study. The mean SFChT of OIS eyes was significantly lower than that of fellow eyes (OIS eyes: 208.89 ± 82.62 μm and fellow eyes: 265.31 ± 82.77 μm,P<0.001). The choroidal volume of OIS eyes was significantly smaller than that of fellow eyes (OIS eyes: 0.16 ± 0.05 mm3and fellow eyes: 0.21 ± 0.05 mm3,P<0.001).Conclusion. The choroidal thickness and volume of OIS eyes were smaller than those of unaffected fellow eyes. Decreased choroidal circulation caused by carotid artery stenosis might affect the discordance of choroidal thickness and choroidal volume.

DNA shuffling was used to improve the thermostability of maltogenic amylase from Bacillus thermoalkalophilus ET2. Two highly thermostable mutants, III-1 and III-2, were generated after three rounds of shuffling and recombination of mutations. Their optimal reaction temperatures were all 80 degrees C, which was 10 degrees C higher than that of the wild-type. The mutant enzyme III-1 carried seven mutations: N147D, F195L, N263S, D311G, A344V, F397S, and N508D. The half-life of III-1 was about 20 times greater than that of the wild-type at 78 degrees C. The mutant enzyme III-2 carried M375T in addition to the mutations in III-1, which was responsible for the decrease in specific activity. The half-life of III-2 was 568 min while that of the wild-type was < 1 min at 80 degrees C. The melting temperatures of III-1 and III-2, as determined by differential scanning calorimetry, increased by 6.1 degrees C and 11.4 degrees C, respectively. Hydrogen bonding, hydrophobic interaction, electrostatic interaction, proper packing, and deamidation were predicted as the mechanisms for the enhancement of thermostability in the enzymes with the mutations.

To compare clinical outcomes between the IntraLase femtosecond laser and the mechanical microkeratome for creating flaps during laser in situ keratomileusis.Nonrandomized clinical trial.The study involved a total of 55 eyes of 30 patients, with 27 eyes of 16 patients comprising the microkeratome group and 28 eyes of 14 patients comprising the IntraLase group. Refractive errors, visual acuity, higher-order aberrations, contrast sensitivity, and corneal sensitivity were compared between the two groups.There were no statistically significant differences between the two groups in terms of refractive errors and postoperative uncorrected visual acuity. There was no loss of best-corrected visual acuity in either group. The higher-order aberrations were similar in both groups (P.05), except for spherical aberration, which was greater in the microkeratome group (P.05). In the IntraLase group, the contrast sensitivity value at 12 and 18 cycles per degree under mesopic conditions was significantly improved at three months postoperatively (P.05). The IntraLase group showed faster corneal sensitivity recovery compared with the microkeratome group, and corneal sensitivity in the peripheral area was nearly normalized at three months postoperatively in the IntraLase group.The femtosecond laser may have advantages over the microkeratome in the flap-making procedure. However, the IntraLase femtosecond laser failed to have significant superiority over the mechanical microkeratome in clinical outcomes, except for faster recovery of corneal sensation, lesser degree of spherical aberration, and some contrast sensitivity value.

Specific-length maltooligosaccharides, particularly maltohexaose, maltoheptaose, and maltooctaose, were prepared from cyclomaltooligosaccharides (cyclodextrins, CDs) by the preferential cyclodextrin ring-opening reaction of an amylolytic enzyme from Pyrococcus furiosus. The enzyme primarily produces maltohexaose, maltoheptaose, and maltooctaose by hydrolyzing alpha-, beta-, and gamma-CD, respectively. This study aims to develop a high-efficiency synthesis of specific maltooligosaccharides at high-purity. [formula: see text]

Genomic analysis of the hyperthermophilic archaeon Pyrococcus furiosus revealed the presence of an open reading frame (ORF PF1939) similar to the enzymes in glycoside hydrolase family 13. This amylolytic enzyme, designated PFTA ( Pyrococcus furiosus thermostable amylase), was cloned and expressed in Escherichia coli . The recombinant PFTA was extremely thermostable, with an optimum temperature of 90°C. The substrate specificity of PFTA suggests that it possesses characteristics of both α-amylase and cyclodextrin-hydrolyzing enzyme. Like typical α-amylases, PFTA hydrolyzed maltooligosaccharides and starch to produce mainly maltotriose and maltotetraose. However, it could also attack and degrade pullulan and β-cyclodextrin, which are resistant to α-amylase, to primarily produce panose and maltoheptaose, respectively. Furthermore, acarbose, a potent α-amylase inhibitor, was drastically degraded by PFTA, as is typical of cyclodextrin-hydrolyzing enzymes. These results confirm that PFTA possesses novel catalytic properties characteristic of both α-amylase and cyclodextrin-hydrolyzing enzyme.

Purpose To investigate the effect of cataract extraction on the results of frequency doubling technology (FDT) perimetry in healthy subjects. Design Single-center, prospective, case series. Methods We performed FDT threshold C20-1 and Humphrey Swedish Interactive Threshold Algorithm (SITA)–fast programs within 1 month before and 2 months after phacoemulsification in 52 consecutive nonglaucomatous patients. Global indexes, including mean deviation and mean sensitivity of FDT and mean deviation of SITA-fast, were compared before and after cataract surgery. Mean sensitivity of FDT at various visual field grid locations was also compared before and after cataract surgery, as were localized indexes of both FDT and SITA-fast. Results Mean deviation improved after cataract surgery in both FDT threshold (from −7.09 dB–−2.16 dB) and SITA-fast program (from −6.14 dB–−2.87 dB). The mean sensitivity at 17 grid locations when the FDT threshold program was used significantly improved postoperatively (from 19.6 dB–26.13 dB). Postoperative improvements in mean sensitivity were also significant at all different sectors, but there were no significant differences of improvement among them. Postoperative pattern standard deviations from FDT and SITA-fast did not differ significantly from their preoperative values. Conclusions Cataracts induced a generalized reduction of sensitivity in FDT threshold as well as SITA-fast perimetry. Cataract removal did not change pattern standard deviation of FDT or of SITA-fast. Caution should be taken when interpreting the results of FDT perimetry in the eyes with cataract.

Central serous chorioretinopathy (CSC) is characterized by well-demarcated serous neurosensory retinal detachment in the posterior pole. CSC is a self-limiting disease that is usually associated with a good visual prognosis. In some cases, however, persistent CSC results in permanent damage to the retina or retinal pigment epithelium [1,2,3,4,5]. Therefore, if the disease continues beyond the acute phase, active treatment must be considered to prevent irreversible damage to retinal function. The pathogenesis of CSC is associated with abnormal choroidal circulation and/or abnormal barrier function in the retinal pigment epithelium. Indocyanine green angiography (ICGA) shows dilated choroidal vessels and choroidal hyperpermeability in CSC patients [6]. The recent development of enhanced depth image-optical coherence tomography (EDI-OCT) has improved the accuracy of choroidal examinations with better quantitative results and increased reliability [7,8,9]. Several EDI-OCT investigations have analyzed the choroidal thickness of CSC eyes. The results of these investigations reveal greater choroidal thickness in CSC eyes relative to age-matched normal eyes. Furthermore, choroidal thickness in unaffected contralateral eyes is also greater than in age-matched controls [10,11,12,13,14]. Intravitreal bevacizumab (IVB) injection is a therapeutic option for chronic CSC. Studies that evaluated the efficacy of IVB injection in CSC eyes have found good visual and anatomical outcomes [15,16,17,18,19,20]. However, IVB administration is not effective in all cases of chronic CSC, which necessitates consideration of alternative treatments such as focal argon laser photocoagulation or photodynamic therapy with verteporfin. Inoue et al. [21] demonstrated that photodynamic therapy for chronic CSC was not effective in eyes without intense hyperfluorescence on late-phase ICGA. Moreover, Jirarattanasopa et al. [22] documented a connection between choroidal thickness and choroidal vascular hyperpermeability on ICGA. They reported that choroidal thickness is greater in areas with choroidal vascular hyperpermeability on ICGA than in unaffected areas [21,22]. Based on these observations, we hypothesized that the responsiveness of specific CSC patients to IVB injection might be related to late-phase ICGA findings and pretreatment choroidal thickness. Given the absence of prior studies on this issue, we investigated differences in pretreatment and posttreatment subfoveal choroidal thickness (SFChT) and late-phase ICGA findings between bevacizumab responders (BevRs) and nonresponders (BevNRs) using Heidelberg Spectralis EDI-OCT and ICGA.

Gusakov critiques our methodology for comparing the cellulolytic activity of the bacterial cellulase CelA with the fungal cellulase Cel7A. We address his concerns by clarifying some misconceptions, carefully referencing the literature, and justifying our approach to point out that the results from our study still stand.

Purpose We evaluated the association of angiogenic and inflammatory cytokine levels in the aqueous humor with development of retinal ischemia and recurrent macular edema in retinal vein occlusion (RVO) patients. Methods This was a retrospective cross-sectional study, and patients with RVO (n = 41) and age-matched control subjects (n = 25) were included. The concentrations of angiogenic and inflammatory cytokines, including VEGF, PDGF-AA, IL-1a, IL-6, IL-8, MCP-1, TNF-α, and IP-10, in the aqueous humor were measured before intravitreal injection of bevacizumab using suspension array technology. After retinal hemorrhage disappeared, fluorescein angiography (FA) images were obtained. Based on FA data, RVO patients were divided into a nonischemic group and an ischemic group. We investigated the presence of recurrent macular edema using optical coherent tomography (OCT) during the follow-up period. We compared the levels of cytokines between RVO patients and control subjects, between nonischemic and ischemic groups, and between patients with and without recurrent macular edema. Results The levels of VEGF, PDGF-AA, IL-1a, IL-6, IL-8, MCP-1, TNF-α, and IP-10 in the aqueous humor were higher in the RVO group than in the control group. The levels of IL-8, PDFGF-AA, TNF-α, and VEGF in the aqueous humor were significantly higher in the ischemic RVO group than in the nonischemic RVO group. We did not observe any association between cytokine levels and recurrent macular edema. Conclusions Angiogenic and inflammatory cytokines were overexpressed in RVO patients. Additionally, increased levels of IL-8, PDFGF-AA, TNF-α, and VEGF in the aqueous humor at the onset of RVO were associated with the development of future retinal ischemia in RVO patients.

Microbacterium sp. AL-210 producing a novel levan fructotransferase (LFTase) was screened from soil samples. The LFTase was purified to homogeneity by (NH4)2SO4 fractionation, column chromatography on Resource Q, and Superdex 200HR. The molecular weight of the purified enzyme was estimated to be approximately 46 kDa by both SDS-PAGE and gel filtration, and the enzyme's isoelectric point was pH 4.8. The major product produced from the levan hydrolysis by the enzyme reaction was identified by atmospheric pressure ionization mass spectrometry and NMR analysis as di-D-fructose-2,6':6,2'-dianhydride (DFA IV). The optimum pH and temperature for DFA IV production were 7.0 and 40 degrees C, respectively. The enzyme was stable at a pH range 7.0-8.0 and up to 40 degrees C. The enzyme activity was inhibited by FeCl2 and AgNO3. The enzyme converted the levan to DFA IV, with a conversion yield of approximately 44%. A gene encoding the LFTase (lftM) from Microbacterium sp. AL-210 was cloned and sequenced. The nucleotide sequence included an ORF of 1593 nucleotides, which is translated into a protein of 530 amino acid residues. The predicted amino acid sequence of the enzyme shared 79% of the identity and 86% of the homology with that of Arthrobacter nicotinovorans GS-9.

An Enzyme Drill Cellulase enzymes degrade the cell walls of plants by breaking down cellulose into its constituent sugar fragments and thus have attracted interest for biofuels production. Using transmission electron microscopy Brunecky et al. (p. 1513 ; see the Perspective by Berlin ) discovered that an especially active cellulase, CelA, from Caldicellulosiruptor bescii bacteria does not move along the surface of the substrate, but drills into the cellulose to form cavities.

To evaluate whether intravitreal ranibizumab injection at cataract surgery prevents postoperative diabetic macular edema (PME) in patients with stable diabetic retinopathy without significant macular edema.Eighty patients with cataract, stable diabetic retinopathy, and no significant macular edema were randomized to a sham group (cataract surgery only) or a group undergoing cataract surgery plus intraoperative ranibizumab injection. Best-corrected visual acuities, central subfield thickness, and total macular volume were assessed at baseline and 1 week, 1, 3, and 6 months postoperatively by spectral domain optical coherence tomography. Clinically meaningful PME (central subfield thickness increase60 μm relative to baseline) was computed.The groups did not differ in baseline best-corrected visual acuity, central subfield thickness, and total macular volume. Compared with the ranibizumab injection group, the sham group had significantly larger central subfield thickness increases relative to baseline at 1 week and 1 month; larger total macular volume increases at all time points (P = 0.012, P = 0.005, P0.001, P0.001, P = 0.005, P = 0.017, respectively); higher PME frequency at 1 month (P = 0.019); and poorer best-corrected visual acuity improvement from baseline to 6 months after surgery (P = 0.046).In patients with stable diabetic retinopathy without significant macular edema, intravitreal ranibizumab injection at cataract surgery may prevent the postoperative worsening of macular edema and may improve the final visual outcome without affecting safety.

In this paper, we present the results of laboratory tests conducted to investigate ultrasonically enhanced flow rate using specially designed and fabricated equipment. The influencing factors αi are verified to investigate the effect of ultrasound on a soil matrix and a flowing liquid. The conditions tested were soil type, temperature and ultrasonic energy. The test results indicate that ultrasound enhances flow rate significantly. The degree of enhancement and the values of influencing factors, αi, however, vary with test conditions.

The most common cause of visual disturbance in patients with diabetic retinopathy is diabetic macular edema (DME) [1]. Laser photocoagulation is the standard treatment for DME [2,3]. Antivascular endothelial growth factor (anti-VEGF) therapy was recently accepted as a first-line treatment for DME because numerous trials have reported its beneficial effects [4,5]. However, despite these dramatic outcomes, not all DME patients respond to anti-VEGF therapy. Nearly 50% of patients treated with ranibizumab in the RESTORE study [4] had an average central subfield thickness (CST) >275 µm at 12 months post-baseline. This means that nearly half of ranibizumab-treated patients still had thickened CST. Triamcinolone acetonide (TA) has been used to treat DME [6,7], and intravitreal triamcinolone acetonide (IVTA) injection has shown efficacy against DME [7,8,9,10]. However, use of IVTA injection is limited in clinical settings because it has been linked to the development of cataracts, elevated intraocular pressure (IOP), sterile pseudo endophthalmitis, and infectious endophthalmitis [7,11,12]. Posterior subtenon triamcinolone acetonide (stTA) injection has been used to treat pseudophakic macular edema and uveitis [7,13,14]. Contrastingly, some authors report that posterior stTA injection for DME treatment does not result in the previously described severe adverse events [7,15,16]. In this study, we investigated the effects of stTA injection for refractory DME after 6 months of failed intravitreal bevacizumab (IVB) injection. We also evaluated potential adverse effects, including elevated IOP and cataract formation.

The aim was to evaluate changes in choroidal thickness, intraocular pressure (IOP) and other parameters of optical coherence tomography, such as central foveal thickness (CFT), macular volume, peripapillary retinal nerve fibre layer (RNFL) thickness after haemodialysis (HD).Thirty-four eyes of 34 chronic renal failure patients (9 eyes with diabetic patients) who underwent haemodialysis in the Dialysis Unit of Gangneung Asan Hospital were included. CFT, macular volume, peripapillary RNFL and choroidal thickness were measured before and after haemodialysis by optical coherence tomography (OCT, Spectralis; Heidelberg Engineering). Changes in the IOP were evaluated with Goldmann applanation tonometry before and after haemodialysis.The average choroidal thickness decreased significantly, from 233.1 ± 77.5 μm to 219.1 ± 76.8 μm (paired t-test, p0.001). This change correlated with the amount of body weight lost (Pearson's correlation coefficient = 0.348, p = 0.044). The mean IOP decreased from 15.1 ± 2.6 mmHg to 13.9 ± 2.2 mmHg after haemodialysis (paired t-test, p = 0.03). The change in macular volume and the small decrease in CFT (214.0 ± 21.0 μm to 213.8 ± 21.8 μm) were not significant. The measured overall change in RNFL thickness from 93.6 ± 15.5 μm to 94.7 ± 14.6 μm was not significant.Haemodialysis can affect various ocular parameters, particularly choroidal thickness and IOP.

PURPOSE To describe the incidence, risk factors for, and long-term visual outcomes of recurrent submacular hemorrhage in the context of age-related macular degeneration. METHODS Medical records of patients with neovascular age-related macular degeneration with or without polypoidal choroidal vasculopathy showing submacular hemorrhage at their first visit to our institution were reviewed. The required minimum follow-up period was 24 months, and any newly developed submacular hemorrhage larger than 1 disk area after near-complete resolution of initial hemorrhage was defined as recurrence. RESULTS A total of 47 eyes of 47 patients were eligible for inclusion. Twenty-four patients showed recurrent submacular hemorrhage during the follow-up period (Group I). Patients without recurrent submacular hemorrhage were included in Group II. The time to recurrent submacular hemorrhage in Group I patients was 21.4 ± 9.2 months. Polypoidal choroidal vasculopathy was present in 50% of Group I patients (n = 12) and 13% of Group II patients (n = 3) (P = 0.025). Intravitreal anti-vascular endothelial growth factor injection was performed during the follow-up period in 70.8% of Group I patients (n = 17) and 95.7% of Group II patients (n = 22) (P = 0.048). Visual acuity change during the follow-up period did not significantly differ between the two groups. CONCLUSION In patients with neovascular age-related macular degeneration presenting with submacular hemorrhage at their first visit, the incidence of recurrent submacular hemorrhage was 51.1% in our retrospective long-term follow-up study. The presence of polypoidal choroidal vasculopathy was associated with an increased risk of recurrent submacular hemorrhage. Use of anti-vascular endothelial growth factor agents was correlated with a reduced risk of such hemorrhage. Visual acuity was stably maintained over 2 years regardless of hemorrhage recurrence.

An increase in intraocular pressure after intravitreal steroid injection facilitates reduction of macular edema

“ Anaerocellum thermophilum ” DSM 6725 is a strictly anaerobic bacterium that grows optimally at 75°C. It uses a variety of polysaccharides, including crystalline cellulose and untreated plant biomass, and has potential utility in biomass conversion. Here we report its complete genome sequence of 2.97 Mb, which is contained within one chromosome and two plasmids (of 8.3 and 3.6 kb). The genome encodes a broad set of cellulolytic enzymes, transporters, and pathways for sugar utilization and compared to those of other saccharolytic, anaerobic thermophiles is most similar to that of Caldicellulosiruptor saccharolyticus DSM 8903.

We present an efficient pipeline enabling high-throughput analysis of protein structure in solution with small angle X-ray scattering (SAXS). Our SAXS pipeline combines automated sample handling of microliter volumes, temperature and anaerobic control, rapid data collection and data analysis, and couples structural analysis with automated archiving. We subjected 50 representative proteins, mostly from Pyrococcus furiosus, to this pipeline and found that 30 were multimeric structures in solution. SAXS analysis allowed us to distinguish aggregated and unfolded proteins, define global structural parameters and oligomeric states for most samples, identify shapes and similar structures for 25 unknown structures, and determine envelopes for 41 proteins. We believe that high-throughput SAXS is an enabling technology that may change the way that structural genomics research is done.

Pyrococcus furiosus thermostable amylase (TA) is a cyclodextrin (CD)-degrading enzyme with a high preference for CDs over maltooligosaccharides. In this study, we investigated the roles of four residues (His414, Gly415, Met439, and Asp440) in the function of P. furiosus TA by using site-directed mutagenesis and kinetic analysis. A variant form of P. furiosus TA containing two mutations (H414N and G415E) exhibited strongly enhanced α-(1,4)-transglycosylation activity, resulting in the production of a series of maltooligosaccharides that were longer than the initial substrates. In contrast, the variant enzymes with single mutations (H414N or G415E) showed a substrate preference similar to that of the wild-type enzyme. Other mutations (M439W and D440H) reversed the substrate preference of P. furiosus TA from CDs to maltooligosaccharides. Relative substrate preferences for maltoheptaose over β-CD, calculated by comparing k cat / K m ratios, of 1, 8, and 26 for wild-type P. furiosus TA, P. furiosus TA with D440H, and P. furiosus TA with M439W and D440H, respectively, were found. Our results suggest that His414, Gly415, Met439, and Asp440 play important roles in substrate recognition and transglycosylation. Therefore, this study provides information useful in engineering glycoside hydrolase family 13 enzymes.

A gene that encodes the enzyme Pyrococcus furiosus cyclodextrin glucanotransferase (PFCGT) was cloned in Escherichia coli. PFCGT was highly expressed in recombinant E. coli after compensation for codon usage bias using the pRARE plasmid. Purified PFCGT was extremely thermostable with an optimal temperature and pH of 95 degrees C and 5.0, respectively, retaining 97% of its activity at 100 degrees C. Incubation at 60 degrees C for 20 min during the purification process led to a 1.5-fold increase in enzymatic activity. A time course assay of the PFCGT reaction with starch indicated that cyclic alpha-1,4-glucans with DPs greater than 20 were produced at the beginning of the incubation followed by an increase in beta-CD. The major final product of PFCGT cyclization was beta-CD, and thus the enzyme is a beta-CGTase.

A W229H mutant of 4-alpha-glucanotransferase (4-alpha-GTase) from Pyrococcus furiosus was constructed and its catalytic properties were studied to investigate the role of W229 in the catalytic specificities of the enzyme. Various activities and kinetic parameters were determined for the wild-type and W229H mutant enzymes. The transglycosylation factor and transglycosylation activity of the mutant enzyme markedly decreased, but its hydrolysis activity was scarcely affected. It was discovered that the k(cat)/K(m) value of transglycosylation activity significantly decreased to about 15% of that of the wild type, while k(cat)/K(m) value of hydrolysis activity changed little for the mutant enzyme. The hydrophobicity of W229 was thought to be critical to the transglycosylation activity of the enzyme based on the enzyme's modeled tertiary structures.

The aim of this study was to evaluate the efficacy of 0.005% latanoprost in lowering intraocular pressure (IOP) in patients with chronic angle-closure glaucoma (CACG) and no visible ciliary-body face. Fourteen eyes of 14 Korean patients with CACG with 360 degrees of peripheral anterior synechiae (PAS) and an IOP greater than 21 mmHg without medication were treated with 0.005% latanoprost once-daily. All patients completed 3 months of treatment with latanoprost. The IOP, which was 30.3 +/- 4.5 (mean +/- standard deviation) mmHg at baseline, decreased to 22.6 +/- 4.9 mmHg after 1 week, 19.6 +/- 5.5 mmHg after 1 month, 19.4 +/- 4.9 mmHg after 2 months, and 21.5 +/- 5.9 mmHg after 3 months of treatment with latanoprost (P < 0.01 for each). Ultrasound biomicroscopy of the anterior chamber angle showed anterior bowing of the iris with total occlusion of the angle by PAS, except for 5 eyes with focal microscopic openings to the ciliary-body face at various angles. Adverse ocular events were well-tolerated and transient. In this preliminary study, treatment with 0.005% latanoprost once-daily resulted in a significant reduction in IOP in CACG patients with 360 degrees of PAS on gonioscopy. Our results suggest that latanoprost may be considered as a therapy of choice in these rare cases.