Advanced glycation end products (AGEs) include a variety of protein adducts whose accumulation alters the structure and function of tissue proteins and stimulates cellular responses. They have been implicated in tissue damage associated with diabetic complications. To assess the possible link between AGE accumulation and the development of diabetic nephropathy (DN), we have examined the immunohistochemical localization of various AGE structures postulated to date, i.e., pentosidine, Nepsilon-(carboxymethyl)lysine (CML), and pyrraline, in diabetic and control kidneys. CML and pentosidine accumulate in the expanded mesangial matrix and thickened glomerular capillary walls of early DN and in nodular lesions and arterial walls of advanced DN, but were absent in control kidneys. By contrast, pyrraline was not found within diabetic glomeruli but was detected in the interstitial connective tissue of both normal and diabetic kidneys. Although the distribution of pyrraline was topographically identical to type III collagen, distribution of pentosidine and CML was not specific for collagen type, suggesting that difference in matrix protein composition per se could not explain heterogeneous AGE localization. Since oxidation is linked closely to the formation of pentosidine and CML, we also immunostained malondialdehyde (MDA), a lipid peroxidation product whose formation is accelerated by oxidative stress, assuming that local oxidative stress may serve as a mechanism of pentosidine and CML accumulation. Consistent with our assumption, diabetic nodular lesions were stained positive for MDA. These findings show that AGE localization in DN varies according to AGE structure, and suggest that the colocalization of markers of glycoxidation (pentosidine and CML) with a marker of lipid peroxidation reflects a local oxidative stress in association with the pathogenesis of diabetic glomerular lesions. Thus, glycoxidation markers may serve as useful biomarkers of oxidative damage in DN.

Incompletely condensed, fluorinated polyhedral oligomeric silsesquioxane with the highly reactive group of trisodium silanolate was used for the synthesis of an initiator for atom transfer radical polymerization. The initiator was applied to solution polymerization of methyl methacrylate (MMA) in the presence of a copper complex. The polymerization proceeded in a living fashion, providing tadpole-shaped polymers with an “inorganic head” of polyhedral oligomeric silsesquioxane (POSS) and an “organic tail” of well-defined PMMA. A blend film composed of the tadpole-shaped polymer and a matrix PMMA was annealed at 180 °C for 5 days and then analyzed by neutron reflectometry, X-ray photoelectron spectroscopy, and contact angle measurement. These analyses revealed that the tadpole-shaped polymer was preferentially populated at the air/polymer interface, and the outermost layer of the film was almost completely covered by the POSS heads. This was mainly due to the low surface free energy of the fluorinated POSS moiety. Owing to this unique structure, the blend film showed strong resistance against Ar+ ion etching, despite the overall POSS content was only 2 wt %.

Immune attacks are key issues for cell transplantation. To assess the safety and the immune reactions after iPS cells-derived retinal pigment epithelium (iPS-RPE) transplantation, we transplanted HLA homozygote iPS-RPE cells established at an iPS bank in HLA-matched patients with exudative age-related macular degeneration. In addition, local steroids without immunosuppressive medications were administered. We monitored immune rejections by routine ocular examinations as well as by lymphocytes-graft cells immune reaction (LGIR) tests using graft RPE and the patient’s blood cells. In all five of the cases that underwent iPS-RPE transplantation, the presence of graft cells was indicated by clumps or an area of increased pigmentation at 6 months, which became stable with no further abnormal growth in the graft during the 1-year observation period. Adverse events observed included corneal erosion, epiretinal membrane, retinal edema due to epiretinal membrane, elevated intraocular pressure, endophthalmitis, and mild immune rejection in the eye. In the one case exhibiting positive LGIR tests along with a slight fluid recurrence, we administrated local steroid therapy that subsequently resolved the suspected immune attacks. Although the cell delivery strategy must be further optimized, the present results suggest that it is possible to achieve stable survival and safety of iPS-RPE cell transplantation for a year.

Membrane permeability is a significant obstacle facing the development of cyclic peptide drugs. However, membrane permeation mechanisms are poorly understood. To investigate common features of permeable (and nonpermeable) designs, it is necessary to reproduce the membrane permeation process of cyclic peptides through the lipid bilayer. We simulated the membrane permeation process of 100 six-residue cyclic peptides across the lipid bilayer based on steered molecular dynamics (MD) and replica-exchange umbrella sampling simulations and predicted membrane permeability using the inhomogeneous solubility-diffusion model and a modified version of it. Furthermore, we confirmed the effectiveness of this protocol by predicting the membrane permeability of 56 eight-residue cyclic peptides with diverse chemical structures, including some confidential designs from a pharmaceutical company. As a result, a reasonable correlation between experimentally assessed and calculated membrane permeability of cyclic peptides was observed for the peptide libraries, except for strongly hydrophobic peptides. Our analysis of the MD trajectory demonstrated that most peptides were stabilized in the boundary region between bulk water and membrane and that for most peptides, the process of crossing the center of the membrane is the main obstacle to membrane permeation. The height of this barrier is well correlated with the electrostatic interaction between the peptide and the surrounding media. The structural and energetic features of the representative peptide at each vertical position within the membrane were also analyzed, revealing that peptides permeate the membrane by changing their orientation and conformation according to the surrounding environment.

Abstract:A clinical grade prototype of posterior multifunctional Jones matrix optical coherence tomography (JM-OCT) is presented. This JM-OCT visualized depth-localized birefringence in addition to conventional cumulative phase retardation imaging through local Jones matrix analysis. In addition, it simultaneously provides a sensitivity enhanced scattering OCT, a quantitative polarization uniformity contrast, and OCT-based angiography. The probe beam is at 1-µm wavelength band. The measurement speed and the depth-resolution were 100,000 A-lines/s, and 6.6 µm in tissue, respectively. Normal and pathologic eyes are examined and several clinical features are revealed, which includes high birefringence in the choroid and lamina cribrosa, and birefringent layered structure of the sclera. The theoretical details of the depth-localized birefringence imaging and conventional phase retardation imaging are formulated. This formulation indicates that the birefringence imaging correctly measures a depth-localized single-trip phase retardation of a tissue, while the conventional phase retardation can provide correct single-trip phase retardation only for some specific types of samples.

The purpose of the present study was to evaluate the intraretinal migration of the retinal pigment epithelium (RPE) cells in age-related macular degeneration (AMD) using polarimetry. We evaluated 155 eyes at various AMD stages. Depolarized light images were computed using a polarization-sensitive scanning laser ophthalmoscope (PS-SLO), and the degree of polarization uniformity was calculated using polarization-sensitive optical coherence tomography (OCT). Each polarimetry image was compared with the corresponding autofluorescence (AF) images at 488 nm (SW-AF) and at 787 nm (NIR-AF). Intraretinal RPE migration was defined by the presence of depolarization at intraretinal hyperreflective foci on PS-SLO and PS-OCT images, and by the presence of hyper-AF on both NIR-AF and SW-AF images. RPE migration was detected in 52 of 155 eyes (33.5%) and was observed in drusenoid pigment epithelial detachment (PED) and serous PED with significantly higher frequencies than in other groups (P = 0.015). The volume of the migrated RPE cluster in serous PED was significantly correlated with the volume of the PED (R2 = 0.26; P = 0.011). Overall, our results showed that intraretinal RPE migrations occurred in various AMD stages, and that they occurred more commonly in eyes with serous and drusenoid PED.

Incompletely condensed polyhedral oligomeric silsesquioxane (POSS) with the highly reactive group of trisodium silanolate was used for the synthesis of two initiators for atom transfer radical polymerization, one with a 2-bromoisobutyl group and the other with a chlorosulfonyl group. These initiators were applied to solution polymerizations of styrene and methyl methacrylate in the presence of a copper complex. In both systems, polymerization proceeded in a living fashion, as indicated by the first-order kinetics of monomer consumption, the evolution of molecular weight in direct proportion to monomer conversion, the good agreement of molecular weight with the theoretical one, and the low polydispersity, thus providing tadpole-shaped polymers with an “inorganic head” of POSS and an “organic tail” of well-defined polymer. Thermogravimetric and differential scanning calorimetric studies showed that both thermal degradation and glass transition temperatures of the organic/inorganic hybrid polymers with molecular weights up to about 20 000 were enhanced as compared to those of model polymers without the POSS moiety.

Many studies on uremic toxins have focused on enzymatic biochemistry. Recently, attention has turned to nonenzymatic biochemistry, especially progressive and irreversible modifications of proteins. Two different approaches opened the field of irreversible nonenzymatic modifications of proteins in uremia: the advanced glycation end products (AGEs) derived from the Maillard reaction and the advanced lipoxidation end products (ALEs) derived from lipid peroxidation. They have revealed the accumulation of reactive carbonyl compounds (RCOs) derived from carbohydrates and lipids and the subsequent carbonyl modifications of proteins ("carbonyl stress"). In this article, we describe the causal role of various RCOs and AGEs/ALEs accumulating in uremia, the clinical consequences of carbonyl stress in uremia, and finally, the therapeutic perspectives. We propose carbonyl stress as a new uremic toxicity.