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1.  The Association between Choroidal Thickness Variations and Response to Intravitreal Bevacizumab in Central Serous Chorioretinopathy 
To analyze differences in the subfoveal choroidal thickness (SFChT) between bevacizumab responders (BevRs) and nonresponders (BevNRs) in patients with idiopathic central serous chorioretinopathy (CSC).
The medical records of 30 unilateral chronic CSC patients who were treated with intravitreal bevacizumab (IVB) as a first line treatment were reviewed. Patients were categorized as BevNRs when CSC did not completely resolve after a minimum of 3 IVB treatments. Enhanced depth imaging-optical coherence tomography was used and SFChT was measured before and after treatment. Choroidal hyperpermeability was also evaluated using indocyanine angiography.
Twenty and 10 eyes were classified as BevRs or BevNRs, respectively. The mean number of IVB treatments was 2.22 ± 0.89 in BevRs, and 4.80 ± 1.03 in BevNRs. Compared with BevNRs, BevRs demonstrated significantly greater pretreatment SFChT (441.25 ± 88.09 vs. 364.10 ± 61.97 µm); SFChT reduction following IVB was significantly greater in BevRs than BevNRs. SFChT in the unaffected eyes was also greater in BevRs than BevNRs. Choroidal hyperpermeability was detected less frequently in BevNRs (hypofluorescence on late-phase, 0.0% and 33.3% in BevNRs and BevRs, respectively; p= 0.049).
Compared with CSC eyes that did not respond well to IVB, BevRs demonstrated significantly thicker SFChT at baseline, greater reduction in SFChT after IVB treatment, and hyperfluorescence on late-phase indocyanine green angiography. We recommend IVB injection as the first-line therapy for CSC eyes with relatively high SFChT and hyperfluorescence on late-phase indocyanine green angiography.
PMCID: PMC4446555  PMID: 26028943
Bevacizumab; Central serous chorioretinopathy; Enhanced depth image-optical coherence tomography; Subfoveal choroidal thickness
2.  Structural Recovery of the Detached Macula after Retinal Detachment Repair as Assessed by Optical Coherence Tomography 
To investigate correlations between preoperative and postoperative foveal microstructures in patients with macula-off rhegmatogenous retinal detachment (RRD).
We reviewed the records of 31 eyes from 31 patients with macula-off RRD who had undergone successful re-attachment surgery. We analyzed data obtained from complete ophthalmologic examinations and optical coherence tomography (OCT) before and 9 to 12 months after surgery. All postoperative OCT measurements were taken with spectral-domain OCT, but a subset of preoperative OCT measurements were taken with time-domain OCT.
The mean duration of macular detachment was 15.5 ± 15.2 days, and mean preoperative best-corrected visual acuity (BCVA, logarithm of the minimum angle of resolution) was 1.03 ± 0.68. Preoperative visual acuity was correlated with retinal detachment height (p < 0.001) and the existence of intraretinal separation (IRS) along with outer layer undulation (OLU) (p = 0.022), but not with macula-off duration. The final BCVA was significantly correlated with integrity of the junction between the photoreceptor inner and outer segments (IS/OS) combined with the continuity of external limiting membrane (ELM) (p = 0.025). The presence of IRS and OLU on a detached macula were highly correlated with the final postoperative integrity of the IS/OS junction and the ELM (p = 0.017).
Eyes preoperatively exhibiting IRS and OLU showed a higher incidence of disruption to the photoreceptor IS/OS junction and the ELM at final follow-up. Such a close correlation between preoperative and postoperative structural changes may explain why ultimate visual recovery in such eyes is poor.
PMCID: PMC3663060  PMID: 23730110
Intraretinal layer separation; Junction between the photoreceptor inner and outer segments; Optical coherence tomography; Outer retinal layer undulation; Rhegmatogenous retinal detachment
3.  Macular Gradient Measurement in Myopic Posterior Staphyloma Using Optical Coherence Tomography 
To evaluate clinical characteristics and the macular gradient in myopic posterior staphyloma with time domain (TD) optical coherence tomography (OCT).
Sixty-four staphyloma eyes of 40 patients were examined. Macular gradient (tangent θ) and the location of staphyloma were assessed with OCT imaging. The macular gradient was measured at points 1 mm and 2 mm distant from the fovea. The relationships of the macular gradient with age, axial length, and spherical equivalent were analyzed.
In 8 eyes (12.5%), the bottoms of the staphylomas were in the fovea, and there was no macular gradient. However, in the other 56 eyes (87.5%), the bottoms of the staphylomas were not in the foveal area, and macular gradients existed. Staphylomas were commonly located in the infero-nasal retinal area. The mean macular gradient (tangent θ) was 0.26 ± 0.08 at 1 mm distance from the fovea and 0.28 ± 0.10 at 2 mm. No significant relationships were observed between macular gradient and axial length, patient age, or spherical equivalent.
TD OCT reveals staphyloma location. If the location is outside of the fovea, a macular gradient exists and can be measured by OCT. Axial length measurement error may occur in eyes with poor visual fixation and steep macular gradients.
PMCID: PMC3149134  PMID: 21860570
Macula; Myopia; Optical coherence tomography
4.  Robust, high-throughput solution structural analyses by small angle X-ray scattering (SAXS) 
Nature methods  2009;6(8):606-612.
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, data analysis, and couples structural analysis with automated archiving. We subjected 50 representative proteins, mostly from Pyrococcus furiosus, to this pipeline, revealing 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.
PMCID: PMC3094553  PMID: 19620974
5.  Time-Lag between Subretinal Fluid and Pigment Epithelial Detachment Reduction after Polypoidal Choroidal Vasculopathy Treatment 
The goal of the present research was to study post-treatment changes in polypoidal choroidal vasculopathy (PCV) shown by optical coherence tomography (OCT).
The study included 12 patients with naive PCV. Photodynamic therapy and 3 consecutive intravitreal bevacizumab injections at 6-week intervals were given. Best corrected visual acuity, subretinal fluid (SRF), pigment epithelium detachment (PED), central macular thickness (CMT), and total macular volume (TMV) were measured before and after treatment as assessed by Stratus OCT3.
After treatment, the SRF height decreased earlier than the PED height. The SRF diameter decreased with statistical significance. However, the PED diameter did not show a statistically significant improvement, persisting at pre-treatment levels. Both CMT and TMV decreased significantly after treatment.
After PCV treatment, SRF and PED stabilized, as shown by OCT. However, the PED treatment response was both delayed and refractory compared to the SRF response. The small change in post-treatment PED diameter may suggest the possibility of PCV recurrence.
PMCID: PMC3060400  PMID: 21461221
Optical coherence tomography; Pigment epithelial detachment; Polypoidal choroidal vasculopathy
6.  Insights into plant biomass conversion from the genome of the anaerobic thermophilic bacterium Caldicellulosiruptor bescii DSM 6725 
Nucleic Acids Research  2011;39(8):3240-3254.
Caldicellulosiruptor bescii DSM 6725 utilizes various polysaccharides and grows efficiently on untreated high-lignin grasses and hardwood at an optimum temperature of ∼80°C. 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 experimentally. 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 carbohydrate 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 enrichment 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.
PMCID: PMC3082886  PMID: 21227922
7.  Efficient Degradation of Lignocellulosic Plant Biomass, without Pretreatment, by the Thermophilic Anaerobe “Anaerocellum thermophilum” DSM 6725▿  
Applied and Environmental Microbiology  2009;75(14):4762-4769.
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.
PMCID: PMC2708433  PMID: 19465524
8.  Genome Sequence of the Anaerobic, Thermophilic, and Cellulolytic Bacterium “Anaerocellum thermophilum” DSM 6725▿  
Journal of Bacteriology  2009;191(11):3760-3761.
“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.
PMCID: PMC2681903  PMID: 19346307
9.  Changes in the Catalytic Properties of Pyrococcus furiosus Thermostable Amylase by Mutagenesis of the Substrate Binding Sites▿  
Applied and Environmental Microbiology  2007;73(17):5607-5612.
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 kcat/Km 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.
PMCID: PMC2042082  PMID: 17630303
10.  Enzymatic Analysis of an Amylolytic Enzyme from the Hyperthermophilic Archaeon Pyrococcus furiosus Reveals Its Novel Catalytic Properties as both an α-Amylase and a Cyclodextrin-Hydrolyzing Enzyme 
Applied and Environmental Microbiology  2004;70(10):5988-5995.
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.
PMCID: PMC522074  PMID: 15466542

Results 1-10 (10)