Corneal collagen crosslinking for keratoconus or corneal ectasia without epithelial debridement.

PURPOSE: Corneal collagen crosslinking (CXL) is a relatively new technique to reduce the progression of keratoconus. The technique can be performed with or without complete debridement of the corneal epithelium. We describe a novel intermediate technique involving mechanical disruption of the epithelium, and evaluate its safety and efficacy. METHODS: The case notes of 128 eyes with progressive keratoconus or iatrogenic corneal ectasia who had undergone CXL using the epithelial disruption technique were retrospectively reviewed. Thin corneas were treated with hypotonic riboflavin. All others were treated with an isotonic solution. Note was made of preoperative and postoperative parameters, including uncorrected visual acuity (UCVA), best spectacle-corrected visual acuity (BSCVA), refraction, endothelial cell count, and corneal tomography. Occurrence of procedure-related complications was recorded. Statistical analyses were performed using the paired sample t-test and Wilcoxon signed-rank test, with a level of P<0.05 being accepted as statistically significant. RESULTS: At 12 months, 41.8% of patients treated with isotonic riboflavin had improved UCVA and 29.7% had improved BSCVA. Only 13.4% lost lines of UCVA and 14.9% lost BSCVA. Of the patients treated with hypotonic riboflavin, at 12 months, 75% demonstrated stability of BSCVA and 25% had stable Kmax. In addition, 25% showed improved visual acuity at 12 months, and 58.3% showed regression of their Kmax. Our rate of short-term complications was comparable to studies using complete epithelial removal. CONCLUSIONS: CXL with epithelial disruption is a safe and effective treatment for keratoconus or iatrogenic corneal ectasia, and may be better tolerated by patients than the epithelium-off technique.

Sterile keratitis after combined riboflavin-UVA corneal collagen cross-linking for keratoconus.

PURPOSE: To investigate patient risk factors and to look for potential causes of sterile infiltrates following an unexpected cluster of sterile keratitis after a routine collagen cross-linking (CXL) list. METHODS: The records of all 148 cases of CXL were reviewed retrospectively. The equipment and solutions used and our clinic's standard operating procedure for CXL were reviewed. An in-vitro experiment to explore the variation in ultraviolet A (UVA) irradiance from fluctuations in the working distance of the UVA lamp was conducted. RESULTS: The four patients who developed sterile infiltrates had steeper maximum corneal curvatures (68.0±7.3 D) and thinner pachymetry (389.9±49.0 µm) than the 144 who did not (57.0±8.2 D, P=0.05; 454.6±45.4 µm, P=0.08). A corneal curvature of >60 Dand a pachymetry of <425 µm were significant risk factors. All four affected cases obtained a complete resolution with topical antibiotics and steroids. The unaided VA and the maximum K improved from their pre-operative levels in three out of four patients. A 2-mm reduction in distance of the VEGA C.B.M. X-Linker from a treated surface increased irradiance to 3.5-3.7 mW/cm(2), which is above the threshold for endothelial toxicity. CONCLUSION: Patients with thinner and steeper corneas are at an increased risk of developing sterile keratitis. The visual outcomes despite this complication are good.

Conjunctivitis due to Phthirus pubis: a trap for the unwary.

It is not uncommon for patients to present to eye casualties with a red eye and where treatment has already been started by the general practitioner, usually in the form of antibiotic drops or ointment for presumed bacterial/viral conjunctivitis. Occasionally, the diagnosis is not simply a bacterial conjunctivitis and this can only be clarified by close ocular inspection.

In vitro antidiabetic activities of five medicinal herbs used in Chinese medicinal formulae.

Fructus Corni, Fructus Schisandrae Chinensis, Poria, Rhizoma Alismatis and Rhizoma Dioscoreae are commonly used in traditional Chinese medicine for diabetes treatment. They are also the component herbs of an antidiabetic foot ulcer formula with demonstrated clinical efficacy. Although some of these herbal extracts were previously shown to possess in vivo antidiabetic effects (i.e. lowering blood glucose levels), the underlying mechanisms remain elusive. The objective of this study is to investigate the possible antidiabetic mechanisms of these individual herbs, using a systematic study platform which includes four in vitro tissue models: glucose absorption into intestinal brush border membrane vesicles (BBMV), gluconeogenesis by rat hepatoma cell line H4IIE, glucose uptake by human skin fibroblasts cell line Hs68 and mouse adipocytes 3T3-L1. All tested herbs showed significant in vitro antidiabetic effects in at least two models. Fructus Schisandrae Chinensis, Poria, Rhizoma Alismatis and Rhizoma Dioscoreae showed significant inhibitory effects in the BBMV glucose uptake assay. All tested herbs showed significant stimulatory effects to the glucose uptake of Hs68 and 3T3-L1 cells, except Poria and Rhizoma Dioscoreae which were not effective to Hs68 and 3T3-L1 respectively. However, none of the tested herbs inhibited hepatic gluconeogenesis. In conclusion, the five herbs exhibited distinct antidiabetic mechanisms in vitro and hence our investigations provided scientific evidence to support the traditional usage of these herbs for diabetic treatment in medicinal formulae.

Binocularity and severe uniocular trauma.

BACKGROUND/AIMS: Monocular outcomes following severe ocular trauma are well described, but little is known about the effect of such injuries on binocular visual outcomes. We therefore report the binocular outcomes in a series of patients with such injuries. METHODS: A retrospective analysis was undertaken of 6 patients with binocular problems following ocular injuries that were severe enough to warrant referral to the regional centre for vitreoretinal surgery. RESULTS: Mean age: 24 years. Mean follow-up: 2.9 years. All patients had 1 to 5 vitreoretinal procedures and the final visual acuities ranged from 6/9 to 1/60. Three patients regained some binocularity, 2 had intractable diplopia and 1 was 'suppressing' with prisms. The final visual acuity did not predict the binocular outcome. The time delay between the original injury and presentation at the motility service was between 3 months and 2.25 years. CONCLUSION: A good monocular visual outcome does not necessarily result in a satisfactory binocular visual outcome and a poor monocular visual outcome does not prevent severe binocular symptoms. Patients need to be counselled regarding this risk, which is an under-recognised problem. We also recommend that all such patients be referred promptly for a thorough orthoptic assessment.

An in vivo investigation on the wound-healing effect of two medicinal herbs using an animal model with foot ulcer.

BACKGROUND/AIM: Over 190 million people in the world suffer from diabetes mellitus. Diabetics are 25 times more likely to have a leg amputated because of unhealing foot ulcers. Herbal medicine has been used in China to salvage the ulcerated limb. With the aim to study the efficacy of two commonly used herbs for ulcer healing, namely Radix Astragali and Radix Rehmanniae, a good animal model needs to be developed for a proper in vivo investigation. METHODS: Firstly, a diabetic animal model was established by streptozotocin injection. Then standard wounds were created on the feet of the diabetic rats. Digital photographs were taken and analyzed by a novel image analysis software. RESULTS: The average ulcer area in the Radix Rehmanniae treatment group was 11.45 mm2, which was significantly smaller than the 15.12 mm2 in the water treatment group (p = 0.04). Radix Astragali, on the other hand, was found to have no significant effect on ulcer shrinkage. CONCLUSION: Further investigation is needed for the identification of the active principles of Radix Rehmanniae.

Pharmacological investigations of the anti-diabetic effect of Cortex Moutan and its active component paeonol.

Cortex Moutan (CM, root bark of Paeonia suffruticosa Andr.) is one of the common herbs found in anti-diabetic traditional Chinese medicine formulae. To study the potential anti-diabetic mechanisms of CM, four in vitro models (intestinal brush border membrane vesicles (BBMV), rat hepatoma cell line H4IIE, human skin fibroblasts cell line Hs68 and mouse adipocytes 3T3-L1) were used. CM showed significant in vitro anti-diabetic effects by inhibiting glucose uptake of BBMV and enhancing glucose uptake into Hs68 and 3T3-L1 cells. Using bioassay-guided fractionation, paeonol was confirmed to be one of the active constituents for inhibiting BBMV glucose uptake. With neonatal-streptozotocin diabetic rats, paeonol (200 and 400mg/kgbody wt.) was found to improve oral glucose tolerance in vivo. To the best of our knowledge, this is the first report on the anti-diabetic effect of paeonol.

Influence of an anti-diabetic foot ulcer formula and its component herbs on tissue and systemic glucose homeostasis.

Complications of diabetes impose major public health burdens worldwide. The positive effect of a Radix Astragali-based herbal preparation on healing diabetic foot ulcers in patients has been reported. Formula 1 is also referred as the 'Herbal drink to strengthen muscle and control swelling'. This formula contains six Chinese medical herbs, including Radix Astragali, Radix Rehmanniae, Rhizoma Smilacis Chinensis, Rhizoma Atractylodis Macrocephalae, Radix Polygoni Multiflori Preparata, and Radix Stephania Tetrandrae. Three of these herbs (Radix Astragali, Radix Rehmanniae, Rhizoma Atractylodis Macrocephalae) are commonly used in different anti-diabetic formulae of Chinese medicine. The objective of the current study is to use an interdisciplinary approach to test the hypothesis that Formula 1 and its components influence tissue and systemic glucose homeostasis. In vitro and in vivo models have been established including: (1) glucose absorption into intestinal brush border membrane vesicles (BBMV); (2) gluconeogenesis by H4IIE hepatoma cells; (3) glucose uptake by 3T3-L1 adipocytes and Hs68 skin fibroblasts; (4) normalization of glycaemic control in a diabetic rat model. The results of in vitro studies indicated that all herbal extracts can modify cellular glucose homeostasis. Since Formula 1 and Rhizoma Smilacis Chinensis extracts demonstrated potent effects on modifying glucose homeostasis in multiple tissues in vitro, they were further studied for their anti-diabetic activities in vivo using a streptozotocin (STZ)-induced diabetic rat model. The results showed that Formula 1 and Rhizoma Smilacis Chinensis extracts did not significantly improve oral glucose tolerance or basal glycaemia in diabetic rats. In conclusion, the anti-diabetic foot ulcer Formula 1 contains ingredients active in modifying tissue glucose homeostasis in vitro but these biological activities could not be associated with improved glycaemic control of diabetes in vivo.

Traumatic wound dehiscence after penetrating keratoplasty-a cause for concern.

AIM: We report the incidence, causes, characteristics, and the outcome of traumatic corneal graft ruptures in a tertiary referral centre in the UK. METHOD: A retrospective analysis of all graft ruptures secondary to trauma that were treated at our centre between 1999 and 2005 was undertaken. Statistical analysis of possible prognostic factors was undertaken using the Fisher's test. RESULTS: Nineteen eyes in 18 patients sustained eye trauma resulting in graft rupture (median age of 53 years; range 27-82; 15 men and four women). Seventeen cases were accidental and two were from violence. The median time interval between grafting and rupture was 8.3 months (range 3 days to 15 years). The 6-year incidence was 3.8%. All graft ruptures occurred at the host-graft junction and ranged from 45 to 270 degrees . Iris prolapse/loss was noticed in 89% and lens loss in 53%. The most common posterior segment complication was vitreous loss (74%), followed by vitreous haemorrhage (32%) and retinal detachment (21%). Grafts with 180 degrees or more of dehiscence were more likely to fail (P<0.001), had more extensive posterior segment damage, and a poorer visual outcome. Grafts without sutures had a more extensive dehiscence (P<0.01). Final visual acuity was worse than 6/60 in 58%. CONCLUSIONS: The risk of traumatic corneal graft rupture is significant and is associated with a poor visual outcome. This fact needs to be clearly emphasised during preoperative counselling and protective measures encouraged.

beta-Amyloid efflux mediated by p-glycoprotein.

A large body of evidence suggests that an increase in the brain beta-amyloid (Abeta) burden contributes to the etiology of Alzheimer's disease (AD). Much is now known about the intracellular processes regulating the production of Abeta, however, less is known regarding its secretion from cells. We now report that p-glycoprotein (p-gp), an ATP-binding cassette (ABC) transporter, is an Abeta efflux pump. Pharmacological blockade of p-gp rapidly decrease extracellular levels of Abeta secretion. In vitro binding studies showed that addition of synthetic human Abeta1-40 and Abeta1-42 peptides to hamster mdr1-enriched vesicles labeled with the fluorophore MIANS resulted in saturable quenching, suggesting that both peptides interact directly with the transporter. Finally, we were able to directly measure transport of Abeta peptides across the plasma membranes of p-gp enriched vesicles, and showed that this phenomenon was both ATP- and p-gp-dependent. Taken together, our study suggests a novel mechanism of Abeta detachment from cellular membranes, and represents an obvious route towards identification of such a mechanism in the brain.

Relative bioavailability of oral sustained-release and regular-release oxprenolol tablets at steady-state.

The relative bioavailability of a test sustained-release (SR) oxprenolol tablet against an approved regular-release (RR) tablet has been investigated at steady-state. In a randomized two-way crossover study, one tablet of 160 mg SR oxprenolol once every 24 h and one tablet of 80 mg RR oxprenolol once every 12 h were given to 12 healthy volunteers for 5 days. Blood samples were collected from each subject just prior to each dose-administration on days 1 through 4, and at scheduled time points on day 5 and analysed for oxprenolol concentration using HPLC. The SR tablet resulted in 42 per cent reduction in mean peak drug levels (p = 0.0341) and a statistically non-significant 14 per cent increase in mean trough levels (p = 0.8357) than the RR tablet. However it required 160 per cent longer time to reach average steady-state concentrations (Css) on day 5 (1.38 h for SR versus 0.53 h for RR; p = 0.0205). The mean area under the plasma drug concentration-time curve at steady state (AUC96-120) with the SR tablet was approximately 18 per cent lower than that observed with the RR tablet, and the degree of fluctuation (DF) was reduced by 30 per cent (2.81 for SR versus 4.11 for RR; p = 0.0069). On average, a single dose of SR tablet and two doses of RR tablets maintained the drug levels above a constant Css of 204.6 ng ml-1 for 7.88 and 7.65 h, respectively (p = 0.3513).

Moment method for the estimation of mass transfer coefficients for physiological pharmacokinetic models.

In vitro and in vivo techniques have been utilized to estimate mass transfer coefficients for physiological pharmacokinetic models. No single method has been adopted for estimating this parameter, in part, due to the different model structures with which this parameter may be associated. A specific method has been derived to calculate mass transfer coefficients for non-eliminating membrane-limited tissue compartments. The present method is based on observed concentration-time data, and requires the calculation of the areas under the zero and first moment curves for plasma, and the first moment curve for the tissue. A Monte Carlo simulation technique was used to determine the percentage biases of the method based on a published model for streptozoticin and adriamycin. For the latter model, the method was compared to a non-linear regression parameter estimation technique.

Application of regression analysis in the evaluation of tumor response following the administration of adriamycin either as a solution or via albumin microspheres to the rat.

Regression analysis has been applied to compare the tumor response following the administration of free and microsphere-loaded adriamycin to the rats. A nonmetastasizing sarcoma was induced in male A.S. inbred rats. When the sarcoma was in its fourth passage and 12 days old, four groups of animals received intratumoral injections of 2 or 8 mg/kg of adriamycin as a solution or via albumin microspheres. The animals in each group were monitored over a period of three weeks for changes in body weight and tumor size and their survival, and the results were statistically compared with the data obtained from an untreated group of animals (control). The use of regression analysis in conjunction with partial F-test revealed that the microsphere-delivered drug exhibited a meager but significantly greater response than the free drug. The implications of intratumoral administration of microsphere-loaded drug in the treatment of sarcomas is discussed.

Factorial design based optimization of the formulation of albumin microspheres containing adriamycin.

The use of factorial design in the formulation of adriamycin-associated albumin microspheres, using the heat-stabilization technique, is illustrated. The effect of stabilization temperature, protein concentration and stabilization time on the entrapment and recovery of adriamycin in microspheres have been investigated using a 2 x 4 x 4 factorial design. The associated drug content in unwashed and four times washed microspheres was determined using HPLC. Maximum drug association and drug recovery were obtained from microspheres synthesised using 25 per cent w/v albumin solution and stabilized at 120 degrees C for 2.5 min. Under these conditions, the entrapped and total associated drug content of the microspheres was about 4 per cent and 12 per cent w/w respectively, and the drug recovery was about 75 per cent. The in vitro dissolution study carried out using dynamic dialysis revealed that the release of adriamycin from these particles follows a bi-phasic pattern. The results demonstrate that use of short stabilization time, low protein concentration and low stabilization temperature are required for the formulation of microspheres with high adriamycin content.

Interaction of L-ascorbic acid on the disposition of lead in rats.

L-Ascorbic acid (As-Ac) has been investigated for its potential in the prophylaxis of lead poisoning. Using rats as the animal model, the pharmacokinetics of As-Ac was determined following a single intravenous dose (100 mg/kg) administered through a jugular vein. Plasma As-Ac levels were monitored using an enzyme assisted UV spectrophotometric method. The pharmacokinetic parameters of As-Ac obtained were used to establish a dosage regimen which could maintain its average plasma concentration of about 11 micrograms/ml at steady state in rats. The disposition of lead acetate (1 mg/kg) in rats was studied in the absence and presence of As-Ac at steady state. Lead concentrations were monitored in femur, kidney, liver and plasma over a 120 hr period using flameless atomic absorption spectrophotometry. In presence of As-Ac, femur, kidney and liver demonstrated 56, 22 and 41% respective reductions in their exposure towards lead. In addition, the half-life of lead in femur and plasma was found to be reduced by 27 and 51% respectively.

Optimized synthesis of polyglutaraldehyde nanoparticles using central composite design.

A central composite design was applied to the optimization of the synthesis of polyglutaraldehyde nanoparticles (PGNP). The effects of monomer concentration, surfactant concentration, pH, oxygen level, and stirring rate on the particle size, polydispersity, surface carboxyl group concentration, and yield of PGNP were investigated. The optimal conditions for the synthesis of PGNP were found to be: 7% (w/v) glutaraldehyde, 2.5% (w/v) dextran, pH 12, 70% (v/v) oxygen, and a stirring rate of 1080 rpm. Under these conditions, the values of the dependent variables adequately resembled those predicted by the model. The usefulness of these particles in the targeted delivery of cytotoxic drugs is discussed.

Optimization of high-performance liquid chromatographic analysis for isoxazolyl penicillins using factorial design.

A 3 X 3 factorial design has been used to study the effects of pH and acetonitrile concentration of the eluents on the retention and resolution of cloxacillin, flucloxacillin and dicloxacillin on a C18 column. The logarithm of the capacity factors of these solutes have been found to vary linearly with the pH and quadratically with the acetonitrile content. The equations generated have been employed to predict experimental conditions necessary for an optimum separation. The chromatographic condition selected has been applied to the quantitation of flucloxacillin in human plasma using dicloxacillin as the interval standard. Sample preparation consists of protein precipitation and solid-phase extraction. The detection limit of the assay at 220 nm for flucloxacillin is in the region of 0.1 microgram/ml. This assay has been employed in a study of the relative bioavailability of two commercial flucloxacillin sodium capsules in ten healthy volunteers.