Judith Flanagan

Role of carnitine in disease

Carnitine is a conditionally essential nutrient that plays a vital role in energy production and fatty acid metabolism. Vegetarians possess a greater bioavailability than meat eaters. Distinct deficiencies arise either from genetic mutation of carnitine transporters or in association with other disorders such as liver or kidney disease. Carnitine deficiency occurs in aberrations of carnitine regulation in disorders such as diabetes, sepsis, cardiomyopathy, malnutrition, cirrhosis, endocrine disorders and with aging. Nutritional supplementation of L-carnitine, the biologically active form of carnitine, is ameliorative for uremic patients, and can improve nerve conduction, neuropathic pain and immune function in diabetes patients while it is life-saving for patients suffering primary carnitine deficiency. Clinical application of carnitine holds much promise in a range of neural disorders such as Alzheimers disease, hepatic encephalopathy and other painful neuropathies. Topical application in dry eye offers osmoprotection and modulates immune and inflammatory responses. Carnitine has been recognized as a nutritional supplement in cardiovascular disease and there is increasing evidence that carnitine supplementation may be beneficial in treating obesity, improving glucose intolerance and total energy expenditure.

Urinary biomarkers involved in type 2 diabetes: a review

Diabetes mellitus is one of the most challenging health concerns of the 21st century. With at least 30% of the diabetic population remaining undiagnosed, effective and early diagnosis is of critical concern. Development of a diagnostic test, more convenient and reliable than those currently used, would therefore be highly beneficial. Urine as a diagnostic medium allows for non‐invasive detection of biomarkers, including some associated with type 2 diabetes and its complications. This review provides a synopsis of those urinary biomarkers that potentially may provide a basis for the development of improved diagnostic tests. Three main pathways for the sourcing of potential makers are identified: kidney damage, oxidative stress and low‐grade inflammation including atherosclerosis/vascular damage. This review briefly presents each pathway and some of the most relevant urinary biomarkers that may be used to monitor the development or progression of diabetes and its complications. In particular, biomarkers of renal dysfunction such as transferrin, type IV collagen and N‐acetyl‐β‐D‐glucosaminidase might prove to be more sensitive than urinary albumin, the current gold standard, in the detection of incipient nephropathy and risk assessment of cardiovascular disease. Inflammatory markers including orosomucoid, tumour necrosis factor‐α, transforming growth factor‐β, vascular endothelial growth factor and monocyte chemoattractant protein‐1, as well as oxidative stress markers such as 8‐hydroxy‐2′deoxyguanosine may also be useful biomarkers for diagnosis or monitoring of diabetic complications, particularly kidney disease. However, the sensitivity of these markers compared with albumin requires further investigation. Copyright

Expression and Localization of Carnitine/Organic Cation Transporter OCTN1 and OCTN2 in Ocular Epithelium

PURPOSE The existence of an organic cation transport process in rabbit cornea and conjunctiva that mediates absorption of carnitine has previously been suggested. This study was conducted to determine the expression and localization of the carnitine/organic cation transporter (OCTN1 and OCTN2) in corneal or conjunctival epithelium. METHODS Reverse transcriptase-polymerase chain reaction (RT-PCR) was used for OCTN1 and OCTN2 mRNA expression in cultured human corneal-limbal epithelial (HCLE) or human conjunctival epithelial (HCjE) cells. Immunofluorescence staining with polyclonal antibody against human OCTN1 or OCTN2 was performed to investigate transporter expression in ocular epithelial cells or rabbit corneal and conjunctival epithelium. Polarity of the transporter expression was determined using Western blot analysis of the apical or basal membrane proteins extracted from the cultured cells. Apical or basal uptake of [H(3)]-L-carnitine was determined using the polarized epithelial cells grown onto collagen-coated porous filter support. RESULTS OCTN1 and OCTN2 mRNA expression was detected in HCLE and HCjE cells of rabbits and humans. OCTN1 and OCTN2 were predominately localized in the apical membranes of the cells. HCLE and HCjE cells were able to take up L-carnitine; most carnitine uptake occurred through the apical surfaces. CONCLUSIONS This report is the first to document OCTN1 and OCTN2 expression in human corneal and conjunctival epithelial cells. These findings suggest potential involvement of OCTN1 and OCTN2 in the transport of carnitine in ocular tissues.

Contact lens deposits, adverse responses, and clinical ocular surface parameters.

Purpose. To correlate clinical responses during contact lens wear with the amount of protein or cholesterol extracted from lenses after wear. Methods. Clinical parameters, including adverse response rates and corneal staining, and symptomatology rating during lens wear were collected from a series of clinical tests comprising four different silicone hydrogel lenses with four different multipurpose solutions. To test for correlates, the amount of total protein or cholesterol extracted from lenses after daily wear were compared statistically to clinical parameters. Results. The amount of protein (p = 0.008) or cholesterol (p = 0.01) extracted from lenses was higher for those subjects who showed solution-induced corneal staining. Amount of protein extracted was correlated (p < 0.01) with conjunctival staining (R = −0.23), lens front surface wetting (r = 0.14), and lens fit tightness (R = −0.20). These clinical parameters accounted for 48% of lens protein deposition. The amount of cholesterol extracted from lenses was much more weakly associated with clinical variables. Amount of protein or cholesterol extracted from lenses was not associated with the production of any corneal infiltrative or mechanical adverse event during wear and was only very weakly correlated with insertion comfort of lenses. Conclusions. These results suggest that there may be no physiologically relevant consequence of cholesterol depositing on silicone hydrogel lenses. The amount of protein that deposits onto silicone hydrogel lenses during wear may have more affect on lens performance on-eye. However, the correlations were generally small and may still not indicate any causative relevant physiological response. Further work is required to determine whether there is any direct causative effect to support these correlative findings.

Soft contact lens disinfection solution efficacy: clinical Fusarium isolates vs. ATCC 36031.

Purpose. To compare the disinfecting efficacy of five soft contact lens multipurpose disinfection solutions (MPDS) against Fusarium solani clinical isolates and the ISO standard ATCC 36031 strain. Methods. Three commercially available and two recalled MPDS were tested using the ISO/CD 14,729 stand-alone test for contact lens care products against 10 ocular isolates of F. solani and the ATCC 36031 strain. The effect of filtering the fungal suspension before incubating in MPDS was also tested. An average log reduction in colony forming units at the manufacturer’s minimum recommended disinfection time was determined and compared with criteria for stand-alone disinfection products for each MPDS against each strain. Results. No difference between filtered and unfiltered fungal suspensions was observed for the ISO standard, whereas in one MPDS the representative clinical isolate showed significantly increased resistance when unfiltered. All but one solution met the stand-alone criteria of 1.0-log reduction of colony forming units against the recommended ISO standard strain ATCC 36031. However, there was wide variation in the ability of MPDS to meet the ISO disinfection criteria when tested against clinical isolates. Among the commercially available MPDS, the two polyquaternium-based solutions showed a higher disinfecting efficacy than the biguanide-based solution. The two recalled solutions showed a lower disinfecting efficacy than the polyquaternium-based solutions. Further, the clinical isolates were significantly more resistant to disinfection than was the recommended ISO strain. Conclusions. The effect of filtering the fungal suspension to remove hyphae seems to be relevant in the clinical isolate tested, but not in the ISO strain. Clinical isolates were significantly more resistant to disinfection than the recommended ISO strain in the presence of both the commercially available and the recalled MPDS. The use of clinical isolates in stand-alone disinfection testing is indicated. Because there were significant differences in increased resistance exhibited by clinical isolates and in a mixed (unfiltered) culture the use of a single laboratory strain may be insufficient to provide assurance that the disinfection solution will be effective against clinical isolates.

Carboxymethyl Cellulose Stimulates Rabbit Corneal Epithelial Wound Healing

Purpose: Previously, we reported carboxymethyl cellulose (CMC) binding to human corneal epithelial cells and promoting corneal epithelial wound closure in vitro. Using an animal model, the efficacy of CMC in promoting corneal wound healing was examined. Materials and Methods: Following corneal epithelial wounding of NZ white rabbits, CMC (0.2% or 1.0%) or control vehicle (PBS) was administered topically (4 times daily for 3 days) to wounded and unwounded eyes with or without contact lens wear. Wound healing in response to the treatments was measured as percentage reduction of fluorescein-stained wound area 0 to 72 hr post-wounding. Corneas were examined histologically and expression of zonula occludens-1 (ZO-1) tight-junction was detected by immunohistochemistry. Results: Percentage wound reduction in CMC-treated groups was significantly greater than controls (p < 0.05) at 24 and 32 hr. Complete wound closure was observed by 48 hr in 100% of CMC-treated eyes compared to 45% of vehicle-treated eyes. CMC also promoted wound closure dose-dependently. Epithelial cells formed an intact layer following CMC-treatment whereas vehicle-treated cells were less ordered. Strong ZO-1 expression in corneal epithelia of CMC-treated eyes was observed at 72 hr. Contact lens wear appeared to delay wound closure compared to without lens wear during CMC-treatment (p = 0.001). Conclusions: CMC promoted dose-dependent corneal epithelial wound healing. CMC stimulated ZO-1 expression, indicating accelerated corneal epithelial resistance barrier regeneration.

In vitro adsorption of tear proteins to hydroxyethyl methacrylate-based contact lens materials.

Objectives: Investigations of polymer interactions in single protein solutions is a necessary step in the elucidation of in vivo early binding events during protein deposition on hydroxyethyl methacrylate-based contact lens materials. Quantity and tenacity of binding of significant tear components to groups I and IV contact lenses was assessed. Competitive binding by these components was also examined. Methods: Adsorption on FDA groups I and IV hydrogel lenses was monitored using 125I-labeled protein. Lenses were incubated in increasing concentrations of radiolabeled single species proteins in solution. For competition experiments, concentration of each radiolabeled protein was held constant and the adsorption/sorption challenged with increasing concentrations of nonlabeled proteins. Lenses were soaked in phosphate-buffered saline to determine desorption. Results: Group IV lenses bound large amounts of lysozyme, whereas group I lenses bound highest amounts of albumin. Albumin binding to both lens types was relatively strong and could not be competed from binding by other proteins lysozyme, lactoferrin, and mucin. Mucin at high concentrations tended to positively cooperate with the binding of lactoferrin and albumin to all lenses. Conclusions: Binding of proteins to hydroxyethyl methacrylate-based hydrogel lens surfaces is affected by charge and polymer components, and perhaps manufacturing processes. Albumin binds strongly to lens surfaces, and this may play an adverse role during contact lens wear.

The impact of diabetes on corneal nerve morphology and ocular surface integrity

Diabetes mellitus is a chronic disease that results from inadequate insulin production or ineffective insulin utilization. It is one of the most common systemic diseases worldwide with increasing prevalence. Diabetes mellitus is associated with premature mortality, macrovascular complications such as cardiovascular disease, and microvascular complications, including nephropathy leading to kidney failure, potentially blinding diabetic retinopathy, and diabetic neuropathy. While the retinal complications of diabetes are well recognized by eye care professionals, the effects on the ocular surface are poorly understood. Recent studies have reported on the association between peripheral neuropathy and corneal neuropathy, showing the latter to be of predictive value for the systemic disease. Corneal neuropathy can lead to loss of corneal sensation and can ultimately result in neurotrophic ulcers and significant visual morbidity. The epithelial fragility and poor wound healing that result from reduced epithelial adhesion to the underlying basement membrane in diabetes, together with corneal neuropathy, are thought to increase the susceptibility to persistent corneal erosions and infection, as well as to increase the risk of post-surgical complications. The aim of this article is to review the impact of diabetes on corneal nerve morphology and ocular surface integrity. Changes in the tear film and ocular surface microbiome are highlighted in discussion of the mechanisms that underpin ocular surface changes that increase the susceptibility to corneal erosion and infection.

Glycerol monolaurate inhibits lipase production by clinical ocular isolates without affecting bacterial cell viability

PURPOSE We sought to determine the relative lipase production of a range of ocular bacterial isolates and to assess the efficacy of glycerol monolaurate (GML) in inhibiting this lipase production in high lipase-producing bacteria without affecting bacterial cell growth. METHODS Staphylococcus aureus,Staphylococcus epidermidis,Propionibacterium acnes, and Corynebacterium spp. were inoculated at a density of 10(6)/mL in varying concentrations of GML up to 25 μg/mL for 24 hours at 37 °C with constant shaking. Bacterial suspensions were centrifuged, bacterial cell density was determined, and production of bacterial lipase was quantified using a commercial lipase assay kit. RESULTS Staphylococcus spp. produced high levels of lipase activity compared with P. acnes and Corynebacterium spp. GML inhibited lipase production by Staphylococcal spp. in a dose-dependent manner, with S. epidermidis lipase production consistently more sensitive to GML than S. aureus. Glycerol monolaurate showed significant (P < 0.05) lipase inhibition above concentrations of 15 μg/mL in S. aureus and was not cytotoxic up to 25 μg/mL. For S. epidermidis, GML showed significant (P < 0.05) lipase inhibition above 7.5 μg/mL. CONCLUSIONS Lipase activity varied between species and between strains. Staphylococcal spp. produced higher lipase activity compared with P. acnes and Corynebacterium spp. Glycerol monolaurate inhibited lipase production by S. aureus and S. epidermidis at concentrations that did not adversely affect bacterial cell growth. GML can be used to inhibit ocular bacterial lipase production without proving detrimental to commensal bacteria viability.

Genetic factors and molecular mechanisms in dry eye disease

Dry eye disease (DED) is a complex condition with a multifactorial etiology that can be difficult to manage successfully. While external factors are modifiable, treatment success is limited if genetic factors contribute to the disease. The purpose of this review is to compile research describing normal and abnormal ocular surface function on a molecular level, appraise genetic studies involving DED or DED-associated diseases, and introduce the basic methods used for conducting genetic epidemiology studies.