Use of Ranibizumab for evaluating focal laser combination therapy for refractory diabetic macular edema patients: an exploratory study on the RELAND trials.

Anti-vascular endothelial growth factor (VEGF) therapy is the first-line treatment for diabetic macular edema (DME), but is less effective in some patients. We conducted a prospective study to determine whether laser combination therapy with anti-VEGF was more effective than Ranibizumab monotherapy in anti-VEGF-resistant DME patients. There was no significant difference in the improvement of the best-corrected visual acuity (BCVA) between the laser combination therapy and Ranibizumab monotherapy groups (3.2 letters and -7.5 letters, p = 0.165). BCVA did not significantly change between visits 1 and 7 (the laser combination group, 64.3 letters 70.3 letters, respectively, p = 0.537; the Ranibizumab monotherapy group, 72.3 letters and 64.8 letters, respectively, p = 0.554), with no significant improvements in central foveal retinal thickness (the laser combination therapy group, 9.3%: the Ranibizumab monotherapy groups, - 7.3%; p = 0.926). There was no significant difference in the number of Ranibizumab intravitreal therapy (IVT) sessions between the groups (laser combination therapy, 5.2; ranibizumab monotherapy, 6.0; p = 0.237). This study did not show that laser combination therapy was significantly more effective for anti-VEGF-resistant DME than anti-VEGF monotherapy alone. Therefore, for anti-VEGF-resistant DME, alternative therapeutic approaches beyond combined laser therapy may be considered.

Safety and Efficacy of Benzalkonium Chloride-optimized Tafluprost in Japanese Glaucoma Patients With Existing Superficial Punctate Keratitis.

PURPOSE: To evaluate the safety and efficacy of benzalkonium chloride (BAK)-optimized tafluprost (with a BAK concentration reduced from 0.01% to 0.001%) in glaucoma patients with existing superficial punctate keratitis (SPK). PATIENTS AND METHODS: A prospective, multicenter, open-label study was designed to compare BAK-optimized tafluprost administered over 12 weeks relative to other preserved prostaglandin analogs previously administered in Japanese glaucoma patients. Thirty patients with SPK graded at <6 points by area density (AD) scoring in 1 eye were recruited. The primary outcome measure was change in AD score at 12 weeks after the switch in treatment compared with that at baseline. Secondary outcome measures included changes in tear film breakup time (TBUT), hyperemia score, and intraocular pressure (IOP). Four patients were excluded from analysis because of treatment discontinuation. RESULTS: Mean AD score±SD decreased significantly from 3.4±0.9 to 1.8±1.8 after the switch (P<0.0001). Mean TBUT increased significantly from 6.3±3.3 to 8.0±4.2 seconds (P<0.01). Mean hyperemia score remained unchanged, whereas mean IOP decreased significantly from 15.6±2.6 to 14.4±2.0 mm Hg (P<0.01). For patients previously treated with BAK-preserved latanoprost (n=17) or bimatoprost (n=2), mean AD score decreased significantly from 3.4±0.9 to 1.8±1.8 (P<0.01) and mean TBUT increased significantly from 6.4±3.6 to 8.2±4.3 seconds (P<0.01); no such changes were apparent for patients previously treated with sofZia-preserved travoprost (n=7). CONCLUSIONS: BAK-optimized tafluprost is a treatment option to improve the condition of the ocular surface and to maintain IOP control in glaucoma patients with existing SPK who have been previously treated with other BAK-preserved prostaglandin analogs.

Comparative proteomic analysis of Methanothermobacter themautotrophicus ΔH in pure culture and in co-culture with a butyrate-oxidizing bacterium.

To understand the physiological basis of methanogenic archaea living on interspecies H(2) transfer, the protein expression of a hydrogenotrophic methanogen, Methanothermobacter thermautotrophicus strain ΔH, was investigated in both pure culture and syntrophic coculture with an anaerobic butyrate oxidizer Syntrophothermus lipocalidus strain TGB-C1 as an H(2) supplier. Comparative proteomic analysis showed that global protein expression of methanogen cells in the model coculture was substantially different from that of pure cultured cells. In brief, in syntrophic coculture, although methanogenesis-driven energy generation appeared to be maintained by shifting the pathway to the alternative methyl coenzyme M reductase isozyme I and cofactor F(420)-dependent process, the machinery proteins involved in carbon fixation, amino acid synthesis, and RNA/DNA metabolisms tended to be down-regulated, indicating restrained cell growth rather than vigorous proliferation. In addition, our proteome analysis revealed that α subunits of proteasome were differentially acetylated between the two culture conditions. Since the relevant modification has been suspected to regulate proteolytic activity of the proteasome, the global protein turnover rate could be controlled under syntrophic growth conditions. To our knowledge, the present study is the first report on N-acetylation of proteasome subunits in methanogenic archaea. These results clearly indicated that physiological adaptation of hydrogenotrophic methanogens to syntrophic growth is more complicated than that of hitherto proposed.

Specific DNA binding of a potential transcriptional regulator, inosine 5'-monophosphate dehydrogenase-related protein VII, to the promoter region of a methyl coenzyme m reductase I-encoding operon retrieved from Methanothermobacter thermautotrophicus strain DeltaH.

Two methyl coenzyme M reductases (MCRs) encoded by the mcr and mrt operons of the hydrogenotrophic methanogen Methanothermobacter thermautotrophicus DeltaH are expressed in response to H(2) availability. In the present study, cis elements and trans-acting factors responsible for the gene expression of MCRs were investigated by using electrophoretic mobility shift assay (EMSA) and affinity particle purification. A survey of their operator regions by EMSA with protein extracts from mrt-expressing cultures restricted them to 46- and 41-bp-long mcr and mrt upstream regions, respectively. Affinity particle purification of DNA-binding proteins conjugated with putative operator regions resulted in the retrieval of a protein attributed to IMP dehydrogenase-related protein VII (IMPDH VII). IMPDH VII is predicted to have a winged helix-turn-helix DNA-binding motif and two cystathionine beta-synthase domains, and it has been suspected to be an energy-sensing module. EMSA with oligonucleotide probes with unusual sequences showed that the binding site of IMPDH VII mostly overlaps the factor B-responsible element-TATA box of the mcr operon. The results presented here suggest that IMPDH VII encoded by MTH126 is a plausible candidate for the transcriptional regulator of the mcr operon in this methanogen.

Application of pseudomurein endoisopeptidase to fluorescence in situ hybridization of methanogens within the family Methanobacteriaceae.

In situ detection of methanogens within the family Methanobacteriaceae is sometimes known to be unsuccessful due to the difficulty in permeability of oligonucleotide probes. Pseudomurein endoisopeptidase (Pei), a lytic enzyme that specifically acts on their cell walls, was applied prior to 16S rRNA-targeting fluorescence in situ hybridization (FISH). For this purpose, pure cultured methanogens within this family, Methanobacterium bryantii, Methanobrevibacter ruminantium, Methanosphaera stadtmanae, and Methanothermobacter thermautotrophicus together with a Methanothermobacter thermautotrophicus-containing syntrophic acetate-oxidizing coculture, endosymbiotic Methanobrevibacter methanogens within an anaerobic ciliate, and an upflow anaerobic sludge blanket (UASB) granule were examined. Even without the Pei treatment, Methanobacterium bryantii and Methanothermobacter thermautotrophicus cells are relatively well hybridized with oligonucleotide probes. However, almost none of the cells of Methanobrevibacter ruminantium, Methanosphaera stadtmanae, cocultured Methanothermobacter thermautotrophicus, and the endosymbiotic methanogens and the cells within UASB granule were hybridized. Pei treatment was able to increase the probe hybridization ratio in every specimen, particularly in the specimen that had shown little hybridization. Interestingly, the hybridizing signal intensity of Methanothermobacter thermautotrophicus cells in coculture with an acetate-oxidizing H(2)-producing syntroph was significantly improved by Pei pretreatment, whereas the probe was well hybridized with the cells of pure culture of the same strain. We found that the difference is attributed to the differences in cell wall thicknesses between the two culture conditions. These results indicate that Pei treatment is effective for FISH analysis of methanogens that show impermeability to the probe.

Additional reduction in intraocular pressure achieved with latanoprost in normal-tension glaucoma patients previously treated with unoprostone.

PURPOSE: To determine whether treatment with latanoprost eye drops is able to further reduce intraocular pressure (IOP) in normal-tension glaucoma (NTG) patients whose IOP has been well controlled with unoprostone. PATIENTS AND METHODS: A total of 34 eyes (34 individuals) with NTG that had been treated with 0.12% unoprostone eye drops twice daily for >or=3 months were switched to treatment once daily with eye drops containing 0.005% latanoprost. IOP was measured before and 1, 2, and 3 months after the switch to latanoprost. RESULTS: The mean IOP of all eyes was decreased significantly by 1.8, 2.9, and 2.3 mmHg at 1, 2, and 3 months after the switch from unoprostone to latanoprost treatment. The IOP of patients with an initial IOP of 12 mmHg was reduced by 11.0 or 19.9%, respectively, after 3 months on latanoprost. The IOP of 30 (88.2%) of the 34 eyes was further reduced by the switch from unoprostone to latanoprost. CONCLUSIONS: Latanoprost reduced the IOP of NTG patients who had already been treated with unoprostone, even though both drugs are prostaglandin-related. Switching to latanoprost might thus achieve a maximal decrease in IOP and thereby better prevent damage to the optic nerve and loss of visual field in NTG patients.

Reconstruction and regulation of the central catabolic pathway in the thermophilic propionate-oxidizing syntroph Pelotomaculum thermopropionicum.

Obligate anaerobic bacteria fermenting volatile fatty acids in syntrophic association with methanogenic archaea share the intermediate bottleneck step in organic-matter decomposition. These organisms (called syntrophs) are biologically significant in terms of their growth at the thermodynamic limit and are considered to be the ideal model to address bioenergetic concepts. We conducted genomic and proteomic analyses of the thermophilic propionate-oxidizing syntroph Pelotomaculum thermopropionicum to obtain the genetic basis for its central catabolic pathway. Draft sequencing and subsequent targeted gap closing identified all genes necessary for reconstructing its propionate-oxidizing pathway (i.e., methylmalonyl coenzyme A pathway). Characteristics of this pathway include the following. (i) The initial two steps are linked to later steps via transferases. (ii) Each of the last three steps can be catalyzed by two different types of enzymes. It was also revealed that many genes for the propionate-oxidizing pathway, except for those for propionate coenzyme A transferase and succinate dehydrogenase, were present in an operon-like cluster and accompanied by multiple promoter sequences and a putative gene for a transcriptional regulator. Proteomic analysis showed that enzymes in this pathway were up-regulated when grown on propionate; of these enzymes, regulation of fumarase was the most stringent. We discuss this tendency of expression regulation based on the genetic organization of the open reading frame cluster. Results suggest that fumarase is the central metabolic switch controlling the metabolic flow and energy conservation in this syntroph.

Correlation of corneal sensation, but not of basal or reflex tear secretion, with the stage of diabetic retinopathy.

PURPOSE: To examine the possible relation between corneal sensation or tear secretion and the stage of diabetic retinopathy in diabetic patients. METHODS: Total reflex or basal tear secretion and corneal sensation were determined in 95 patients with type II diabetes mellitus and 58 nondiabetic control subjects. Tear secretion was measured by the Schirmer test and corneal sensation with a Cochet-Bonnet esthesiometer. RESULTS: Corneal sensation and total or reflex tear secretion were significantly reduced in diabetic patients compared with nondiabetic controls. The loss of corneal sensation, but not that of tear secretion, was significantly correlated with stage of diabetic retinopathy in diabetic patients who were diagnosed with no diabetic retinopathy, simple diabetic retinopathy, preproliferative retinopathy, or proliferative retinopathy. CONCLUSION: Both corneal sensation and total or reflex tear secretion are reduced in individuals with diabetes. The decrease in corneal sensation, but not that in each tear secretion, was correlated with the stage of diabetic retinopathy. Given that loss of corneal sensation is a manifestation of diabetic polyneuropathy, these results are consistent with the notion that both diabetic retinopathy and polyneuropathy result from a basement membrane abnormality.