Amniotic Membrane Transplantation for the Treatment of Infectious Ulcerative Keratitis Before Elective Penetrating Keratoplasty.

PURPOSE: Emergency keratoplasties for inflamed eyes are considered to have a worse prognosis because of immunologic graft rejection. Amniotic membranes have antiinflammatory and antiangiogenic abilities. Therefore, amniotic membrane transplantation (AMT) was performed to stabilize the situation of eyes with severe infectious keratitis before elective penetrating keratoplasty (PK). METHODS: Retrospective, nonrandomized observational case series. Seven to 41 days (median, 20 days) after the onset of intensive antiinfectious medication, an AMT (6 multigrafts and 6 sandwich) was performed in 12 patients [8 men and 4 women; age 46-80 years (median, 66 years)] with herpetic (n = 5), bacterial keratitis (n = 3), or combinations (n = 4). Three to 12 months (median, 5 months) after cessation of the inflammatory status of the eye, a central elective PK (diameter, 7-8 mm) became feasible in 10 eyes. Follow-up ranged from 4 to 38 months (median, 20 months) after PK. RESULTS: The primary success rate of AMT was 11/12 (92%). Five recurrences (41%) were treated successfully 4 times by repeat AMT (sandwich) and 1 time by emergency PK. In 2 of the 12 eyes, an irreversible endothelial immunologic graft reaction appeared 18 and 21 months after PK. One eye suffered from reversible recurrence of herpetic keratitis on the corneal graft. At the end of the follow-up, 10 of 12 grafts (83%) were clear. CONCLUSIONS: A rapid decrease in the inflammatory reaction and a fast reepithelialization because of AMT after intensive antiinfectious medication in case of severe ulcerative keratitis may help to avoid an emergency keratoplasty and improves the prognosis of the elective keratoplasty.

Aqueous humor glycation marker and plasma homocysteine in macular degeneration.

BACKGROUND: We investigated concentrations of total homocysteine (tHcy) in elderly people without and those with age-related macular degeneration (AMD). In addition, we tested the association between plasma tHcy and one glycation marker in aqueous humor. METHODS: People with cataract only (n=48), patients with dry AMD (n=38) and those with wet AMD (n=31) were studied. Blood concentrations of tHcy, and methylation and vitamin markers were measured in 116 blood samples. The concentrations of the extracellular soluble receptor for advanced glycated end products (esRAGE) were measured in 77 aqueous humor samples. RESULTS: Mean aqueous humor concentration of esRAGE and that of plasma tHcy did not differ significantly between the groups. Arterial hypertension but not eye disease explained the tHcy elevation in plasma in this study. In the cataract group, a significant negative correlation was found between plasma tHcy and that of esRAGE in aqueous humor (r=-0.483, p=0.006). In patients with dry AMD, the concentration of esRAGE in aqueous humor correlated negatively to tHcy and positively to serum folate. CONCLUSIONS: Plasma tHcy levels were positively associated with hypertension, but not with AMD in this study. Higher esRAGE in aqueous humor was related to higher folate and lower tHcy in blood. Following studies may assess whether B-vitamins can protect against age-related ocular diseases by reducing glycation.

Synthesis and properties of (triptycenedicarboxylatio)zinc coordination networks.

(Triptycenedicarboxylato)zinc metal-organic frameworks (MOFs) based on paddle wheel secondary building units (SBUs) with different axial ligands have been prepared. The reproducible formation of the layered paddle-wheel structures from triptycenedicarboxylic acid (H(2)TDC) and zinc nitrate under various conditions seems to be characteristic of this acid and is utilized for the construction of 3D frameworks by a pillaring approach. We attempted to bring additional functionalities into MOFs by employing the appropriate pillaring ligands, for example, bis(4-pyridyl)-s-tetrazine and bis(4-pyridyl)-dimethoxy-p-phenylenedivinylene, and investigated certain properties of some MOF materials, such as guest-exchange behavior, luminescence, microporosity, and stability.

MMP-2 and MMP-9 secretion by rpe is stimulated by angiogenic molecules found in choroidal neovascular membranes.

PURPOSE: Matrix metalloproteinases (MMP)-2 and -9 play an important role in the pathogenesis of choroidal neovascularization (CNV). Retinal pigment epithelial cells (RPE) are an important source of MMPs in the outer retinal environment, however little is known about the local factors that modulate MMP secretion in these cells. The purpose of this study was to determine the effects of CNV involved growth factors and the extracellular matrix molecule fibronectin on MMP-2 and -9 secretion by cultured human RPE. METHODS: MMP-2 and -9 secretion was studied using gelatin zymography, Western blot, and ELISA assay of RPE culture supernatants. The effects of stimulating the cells for 36 hours with vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bGFG), tumor necrosis factor-alpha (TNF-alpha), or fibronectin (FN), all angiogenic factors found in CNV membranes, was determined. RESULTS: Resting RPE cells secreted MMP-2 but not MMP-9. Stimulation with TNF-alpha induced secretion of MMP-9 and increased the secretion of MMP-2. MMP-2 secretion was also increased by stimulation with FN and VEGF, but not bFGF. CONCLUSION: The results indicated that the angiogenic molecules VEGF, FN, and TNF-alpha stimulate MMP-2 and -9 secretion from RPE and thus further promote CNV.

The structure of the strongest Brønsted acid: the carborane acid H(CHB11Cl11).

The strongest and most robust carborane acid, H(CHB11Cl11), has a monomeric structure in the gas phase. IR spectra show two nuH-Cl bands at 2357(br) and 2066(br) cm-1 which, together with DFT calculations, indicate the coexistence of at least two isomers. The acidic proton bridges adjacent chlorine atoms with asymmetric Cl-H...Cl hydrogen bonding. The 12,7 isomer is more stable than the 7,8 isomer. These monomers can be condensed into an amorphous solid phase but are metastable. They quickly decay, first to an amorphous dimeric structure, then to a crystalline polymeric phase that has been characterized by X-ray crystallography. In the polymeric structure, the acidic proton bridges chlorine atoms from the 7-11 positions of carborane anions in linear chains. The dimeric phase (nuCl-H...Cl = 1100-2200 cm-1) and polymeric phase (nuasClHCl ca. 1100 cm-1, v broad) have more nearly symmetrical, low-barrier H-bonding. These findings have implications for the dependency of acid strength upon phase.

Novel weak coordination to silylium ions: formation of nearly linear Si-H-Si bonds.

The weakly coordinating carborane anion in ion-like trialkylsilyl species R3Si(CHB11Cl11) can be displaced by nucleophiles as weak as ortho-dichlorobenzene, SO2 and trialkylsilanes, the latter forming nearly linear hydride bridges in R3Si-H-SiR3+ cations.

Inhibitory effects of verapamil isomers on the proliferation of choroidal endothelial cells.

PURPOSE: To evaluate the effects of verapamil isomers on in vitro proliferation of bovine choroidal endothelial cells (CECs). MATERIALS AND METHODS: CECs were isolated from bovine eyes and cultured in endothelial growth medium (EGM). For the proliferation assays, CECs were exposed to verapamil isomers (0.1-100 microM) in EGM with 2% fetal bovine serum or basic fibroblast growth factor (bFGF) (10 ng/ml). After 72 h of incubation with the desired drug, the cellular proliferation was determined by an MTT assay and a BrdU assay. In addition, the drug toxicity on CECs stimulated with EGM was evaluated by cell counting with trypan blue. RESULTS: All verapamil isomers inhibited the bFGF- or medium-stimulated growth significantly in a concentration range of 10-40 microM without toxicity. No significant differences were seen between the inhibitory effects of the various isomers. Cell toxicity was detected at a concentration of 100 microM verapamil isomers on EGM-stimulated CECs. CONCLUSION: The results demonstrate the efficacy of all verapamil isomers in inhibiting CEC proliferation involved in the process of choroidal neovascularization. D: -(+)-Verapamil may be recommended for further in vivo evaluation in an animal model of exudative AMD; it has fewer systemic and local side effects because calcium channels are not blocked.

Sequential induction of angiogenic growth factors by TNF-alpha in choroidal endothelial cells.

Inflammatory mediators have been proposed to play a critical role in the pathogenesis of choroidal neovascularization, a blinding complication of age-related macular degeneration. We evaluated the expression of TNF-alpha in human choroidal neovascular membranes and found that it colocalized with cells expressing VEGF, angiopoietin (Ang)-1 and Ang2. In cultured choroidal endothelial cells we found that TNF-alpha increased Ang2 mRNA (increased transcription) and protein levels prior to those of Ang1 and VEGF. The results raise the possibility that during neovascularization, TNF-alpha may modulate endothelial plasticity and survival by sequential inactivation of Tie2 followed by activation of Tie2 and VEGF receptors.

Simultaneous multiple immunoassays in a compact disc-shaped microfluidic device based on centrifugal force.

BACKGROUND: We explored the potential of a microfluidic device based on centrifugal force as an immunoassay platform by examining the imprecision of assays carried out with 200 nL of sample. METHODS: Biotinylated antibodies against alpha-fetoprotein (AFP), interleukin-6 (IL-6), and carcinoembryonic antigen [(CEA); 0.1 g/L each in 15 mmol/L phosphate-buffered saline (PBS) containing 0.1 mL/L Tween 20] were attached to a microcolumn packed with streptavidin-coated particles. A 200-nL sample was then allowed to pass through the microcolumn for 240 s, followed by Alexa 647-labeled detection antibody (7.5 mg/L in 15 mmol/L PBS containing 10 g/L bovine serum albumin). The flow rate was controlled by altering the rotational speed. Up to 104 sandwich type immunoassays were completed within 50 min. RESULTS: For AFP, IL-6, and CEA the detection limits were, respectively, 0.15, 1.25, and 1.31 pmol/L. Inter- and intraassay imprecisions (CVs) were <10% and <20%, respectively, for analyte concentrations >5 pmol/L. The CEA antibody had the lowest affinity according to fluorescence image analysis of the microcolumn region. The result was confirmed in a comparative study using BIAcore 3000. CONCLUSIONS: Day-to-day (total) imprecision (CV) of immunoassays on the compact disc-shaped device are <20%. Analysis of fluorescence images allows rapid ranking of antibodies according to their affinities.

Impact of endostatin on bFGF-induced proliferation, migration, and matrix metalloproteinase-2 expression/secretion of bovine choroidal endothelial cells.

PURPOSE: To investigate the potential role of endostatin, an endogenous angiogenesis inhibitor, in the prevention of choroidal angiogenesis-related disorders. METHODS: Bovine choroidal endothelial cells (CEC) were cultured and treated with basic fibroblast growth factor (bFGF) alone or combined with endostatin at concentrations ranging from 0.1 to 10 microg/ml. The proliferation and migration of CECs were evaluated by using 3, (4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) colorimetric assay and modified Boyden chamber assay, respectively. For evaluating expression and secretion of matrix metalloproteinase-2 (MMP-2), CEC-conditioned media were subjected to zymography and/or Western blot analysis, and the cells were used for semiquantitative reverse transcription polymerase chain reaction (RT-PCR) analysis. RESULTS: Endostatin did not inhibit bFGF-induced or nonstimulated CEC proliferation (p > 0.05). The bFGF-induced migration was significantly inhibited by endostatin at concentrations of 1 and 10 microg/ml (p < 0.05). The bFGF-upregulated expression of mRNA in CECs and the secretion of MMP-2 protein of CECs were both suppressed by endostatin. CONCLUSIONS: Inhibitory effect of endostatin on expression and secretion of MMP-2 and cell migration, but not on proliferation of CECs, could respond to its therapeutic action for choroidal neovascularization-dependent disorders.

A selective cyclic integrin antagonist blocks the integrin receptors alphavbeta3 and alphavbeta5 and inhibits retinal pigment epithelium cell attachment, migration and invasion.

BACKGROUND: Proliferative vitreoretinopathy (PVR) is a leading cause of blindness after failed retinal reattachment surgery. PVR is characterized by the proliferation, migration and contraction of retinal pigmented epithelial cells (RPE), and these cellular responses are influenced by the expression and function of integrin receptors. The effect of a cyclic integrin antagonist containing the amino acid sequence Arg-Gly-Asp-D-Phe-Val (RGDfV), specific for the integrin receptors alphavbeta3 and alphavbeta5, was investigated on basic fibroblast growth factor (bFGF), platelet derived growth factor-BB (PDGF-BB), and serum induced human RPE proliferation, migration, invasion and attachment to the extracellular matrix. Furthermore, the effects of bFGF and PDGF-BB regulated expression of integrins alphavbeta3 and alphavbeta5 on RPE cells was examined. METHODS: The effect of a cyclic integrin antagonist and a control peptide (0.01 microg/ml to 300 microg/ml) was investigated on serum or cytokine (bFGF or PDGF-BB pretreatment) induced human fetal RPE cell proliferation by H3-thymidine uptake. The effect of the cyclic integrin antagonist on RPE cell attachment onto different extracellular matrices (laminin, collagen IV, fibronectin), RPE cell invasion stimulated by PDGF-BB or serum, and migration stimulated by PDGF-BB, vascular endothelial growth factor (VEGF) or serum was explored. PDGF-BB and bFGF modulation of the integrin receptors alphavbeta3 and alphavbeta5 was evaluated by flow cytometry. RESULTS: The integrin antagonist did not inhibit DNA synthesis stimulated by serum, bFGF, or PDGF-BB treatment. RPE attachment onto fibronectin was inhibited in a concentration range of 1-10 microg/ml (p < 0.05). Attachment of the RPE cells onto collagen IV and laminin was inhibited in a range of 3-10 microg/ml (p < 0.05). Serum and PDGF-BB stimulated migration was inhibited by the cyclic integrin antagonist in a concentration range of 1-10 microg/ml (p < 0.05). Furthermore, the cyclic integrin antagonist inhibited PDGF-BB stimulated RPE cell invasion through fibronectin (3 microg/ml: 66% inhibition, p < 0.001). In each of these experiments, the control peptides had no significant effects. PDGF-BB and bFGF pretreatment of RPE cells increased the expression of integrin receptors alphavbeta3 (bFGF: 1.9 fold, PDGF-BB: 2.3 fold) and alphavbeta5 (bFGF: 2.9 fold, PDGF-BB: 1.5 fold). CONCLUSION: A selective inhibition of the integrin receptors alphavbeta3 and alphavbeta5 through a cyclic integrin antagonist is able to inhibit RPE cell attachment, migration and invasion. Since these steps are of importance for the progression of PVR, a cyclic integrin antagonist should be further evaluated for the treatment of this disease.

The structure of the H3O+ hydronium ion in benzene.

Infrared, X-ray structural, 1H NMR, and computational evidence for pi-solvation of H3O+ by benzene molecules is presented. A salt with a discrete [H3O.3benzene]+ cation can be isolated using a very weakly interacting carborane counterion, CHB11Cl11-. pi-Arene solvation of H3O+ explains the solubility of this salt in benzene solution. Similar results are indicated for the "Zundel-type" H5O2+ ion. These findings suggest structures for the active protonating species when strong acids are used as catalysts in arene solvents containing trace water. They are also relevant to structures that may be present in biological proton transport.

Carboxyamido-triazole modulates retinal pigment epithelial and choroidal endothelial cell attachment, migration, proliferation, and MMP-2 secretion of choroidal endothelial cells.

PURPOSE: To determine the effect of the calcium signaling modulating drug carboxyamido-triazole (CAI) on substeps of exudative age-related macular degeneration (AMD) in vitro. MATERIALS AND METHODS: Zymography and ELISA determined the effect of CAI on MMP-2 production of choroidal endothelial cells (CECs) stimulated by bFGF and VEGF. The effects of CAI on attachment of retinal pigment endothelial (RPE) cells/CECs onto fibronectin, laminin, collagen IV, and migration toward fibronectin were investigated. Proliferation induced by serum and bFGF (10 microg/ml) with and without CAI (0.1-10 microM) was measured by cell counting and 3H-uptake. Viability and apoptosis of the exposed cells was assessed by an MTT and an apoptosis assay. RESULTS: CAI inhibited serum- and bFGF-induced proliferation, cell attachment onto fibronectin and collagen IV, but only CEC attachment onto laminin. Inhibition of MMP-2 production was observed (10 microM CAI). CAI reduced the cellular viability by apoptosis induction. CONCLUSIONS: CAI inhibits substeps of exudative macular degeneration and may be of value for the treatment of the disease.

Endohedral zintl ions: intermetalloid clusters.

Tetrels can be regarded as most promising candidates for the construction of larger clusters. Recent examples have shown that larger clusters are particularly stable if they contain interstitial atoms (e.g. [Pt@Pb12]2-). Many salts of the polyhedral anions are soluble, but a number of examples-usually those with higher charges-occur only as quasi-discrete units in saltlike crystals (Zintl phases) or as building blocks in intermetallic phases. In this Minireview, the chemistry of intermetalloid clusters is reviewed with reference to the endohedral Zintl ions, Zintl phases, and polyhedral building blocks of intermetallic compounds, including heteroatomic species in the gas phase. We focus on selected examples and discuss the new findings in the context of recent advances in the field of metalloid clusters and (endohedral) fullerenes and fullerides.