Tear protein interaction with hydrogel contact lenses

The design and synthesis of biomaterials covers a growing number of biomedical applications. The use of biomaterials in biological environment is associated with a number of problems, the most important of which is biocompatabUity. If the implanted biomaterial is not compatible with the environment, it will be rejected by the biological site. This may be manifested in many ways depending on the environment in which it is used. Adsorption of proteins takes place almost instantaneously when a biomaterial comes into contact with most biological fluids. The eye is a unique body site for the study of protein interactions with biomaterials, because of its ease of access and deceptive complexity of the tears. The use of contact lenses for either vision correction and cosmetic reasons or as a route for the controlled drug delivery, has significantly increased in recent years. It is relatively easy to introduce a contact lens Into the tear fluid and remove after a few minutes without surgery or trauma to the patient. A range of analytical techniques were used and developed to measure the proteins absorbed to some existing commercial contact lens materials and also to novel hydrogels synthesised within the research group. Analysis of the identity and quantity of proteins absorbed to biomaterials revealed the importance of many factors on the absorption process. The effect of biomaterial structure, protein nature in terms of size. shape and charge and pH of the environment on the absorption process were examined in order to determine the relative up-take of tear proteins. This study showed that both lysozyme and lactoferrin penetrate the lens matrix of ionic materials. Measurement of the mobility and activity of the protein deposited into the surface and within the matrix of ionic lens materials demonstrated that the mobility is pH dependent and, within the experimental errors, the biological activity of lysozyme remained unchanged after adsorption and desorption. The study on the effect of different monomers copolymerised with hydroxyethyl methacrylate (HEMA) on the protein up-take showed that monomers producing a positive charge on the copolymer can reduce the spoilation with lysozyme. The studies were extended to real cases in order to compare the patient dependent factors. The in-vivo studies showed that the spoilation is patient dependent as well as other factors. Studies on the extrinsic factors such as dye used in colour lenses showed that the addition of colourant affects protein absorption and, in one case, its effect is beneficial to the wearer as it reduces the quantity of the protein absorbed.

Tear and skin phospholipids:analysis and hydrogel analogues

This thesis is concerned with the analysis of phospholipids in the tear film and with the synthesis of phospholipids analogous to hydrogels. The work consists of two areas. The first area is the study of the phospholipids in the tear film, their nature and fate. The use of liquid chromatography mass spectrometry determined that the concentration of phospholipids in the tear film was less than previously thought. The analysis of the tear film phospholipids continued with thin layer chromatography. This showed the presence of diacylglycerides (DAGs) in the tear film at relatively high concentrations. The activity of an enzyme, phospholipase C, was found in the tear film. It was hypothesised that the low concentration of phospholipids and high concentrations of DAG in the tear film was due to the action of this enzyme. The second area of study was the synthesis of phospholipids analogous materials for use in ocular and dermal applications for use in ocular and dermal applications.For ocular applications the synthesis involved the use of the monomer N,N-dimethyl-N-(2-acryloylethyl)-N-(3-sulfopropyl) ammonium betaine (SPDA) in combination with 2-hdyroxyethyl methacrylate (HEMA). Charge-balanced membranes were also synthesised using potentially anionic monomers in conjunction with cationic monomers in stoichiometrically equivalent ratios also with HEMA as a commoner. Membranes of SPDA copolymers and charge-balanced copolymers proved to have some properties suitable for ocular applications. The dermal materials consisted of one family of partially hydrated hydrogels synthesised from SPDA in combination with ionic monomers: sodium 2-(acrylamido)-2-methyl propane sulfonate and acrylic acid-bis(3-sulfopropyl)-ester, potassium salt. A second family of partially hydrated hydrogels was synthesised from N-vinyl pyrrolidone in combination with the same ionic monomers. Both of the partially hydrated hydrogels synthesised proved to have some properties suitable for use as adhesives for the skin.

Synthesis of novel cylic monomers for hydrogel polymers

The aim of the project was to synthesise hydrophilic derivatives of 1,2-dihydroxy-3,5-cyclohexadiene (DHCD) and to copolymerise these derivatives with 2-hydroxyethyl methacrylate (HEMA), to give a completely new range of hydrogel materials. It was thought that hydro gels incorporating hydrophilic derivatives of DHCD could have good mechanical properties and good water binding ability. A model compound for cis-DHCD was sought, as cis-DHCD was expensive and stable under only a narrow range of conditions. Catechol was found to be an excellent model for cis-DHCD, as 1H NMR spectroscopy indicated that both compounds contained eclipsed hydroxy groups and flat rings. A number of catechol derivatives were prepared in good yield, under non-acidic conditions at room temperature. The limited availabilty of cis-DHCD led to an investigation into synthesising hydrophilic derivatives of both cis and trans-DHCD indirectly. Hydrophobic derivatives were easily prepared by indirect routes, but it was found that hydrophilic derivatives were considerably more difficult to synthesise. A number of novel routes to both cis and trans-DHCD were also explored. Copolymerisation of diacetate, dimethylcarbonate and dipivalate derivatives of cis-DHCD with HEMA, to form a hitherto unknown group of hydrogels, is reported. Hydrogels containing these monomers showed significant improvements in both tensile strength and Youngs modulus, at both equivalent composition and water content, over the corresponding HEMA / styrene and HEMA / methyl methacrylate analogues. It was observed that derivatives of trans-DHCD polymerise with difficulty. 1H NMR studies indicated that both faces of the ring were shielded by the pendant groups thereby preventing efficient polymerisation of the trans monomers.

Drug/Water interactions in hydrogel matrices

The primary aim of this research has been the investigation of the role of water structuring effects in the widely different extents of irritancy displayed by certain antibiotics. The compounds involved were members of the Lincomycin group of antibiotics. The aqueous solution behaviour of these co~pounds was studied using techniques such as vapour pressure osmometry end differential scanning calorimetry (D.S.C.). The effects of the antibiotics on water structure in hydrogel membrane preparations In which the equilibrium water content (E.W.C.) and constituent amounts of freezing and non-freezing water ware varied were also investigated using D.S.C. The permeability of water swollen hydrogel preparations to aqueous antibiotic solutions as well as other solutes were studied. A series of hydrogel preparations into which the antibiotics had been incorporated during polymerisation were developed and used in studies of the effects of the antibiotics end their water structure modifications on the permeation of a range of solutes.

Characterisation and modification of hydrogel polymers

The work described in this thesis can be broadly divided into two sections. The first being the characterisation of hydrogel polymers in both their hydrated and dehydrated states and the second some aspects of the structural modification of polymers. The characterisation of hydrogel polymers in their dehydrated state (xerogels) involves such techniques as elemental analysis, pyrolysis gas liquid chromatography, infra-red spectroscopy, density determination and surface characterisation by contact angle measurements. The characterisation of some commercially available hydrogel materials was undertaken using such techniques and the results obtained were compared to laboratory synthesised systems in an attempt to assess the value of the combination of techniques employed. In the characterisation of hydrated polymers the amoumt and nature of water present is the single most important factor. The most convenient method of characterising this water involves the use of differential scanning calorimetry (DSC), coupled with total equilibrium water content measurements. DSC distinguishes between non-freezing and freezing water but in addition provides some information on the continuum of states in the freezing water fraction. Two aspects of the structural modification of hydrogel polymers were studied. The first involved the incorporation of acrylamide and substituted acryamide monomers into a copolymer system and an examination of the effect of this on the amino acid interaction of the polymers. The second was the attempted synthesis of cell surface analogues by the attachment of sugar type molecules to the polymer using a variety of reaction methods.

Ocular response to silicone-hydrogel contact lenses

The thesis investigates the ocular response to silicone-hydrogel (SiH) contact lens wear, a relatively new contact lens material that has a higher modulus of rigidity and different surface coating than used in conventional hydrogel materials. The properties of SiH materials differ significantly from conventional hydrogels and, using subjective and objective means of assessment, the thesis examines how these properties affect reflection and biometry, ocular physiology, tear film characteristics, symptomatology, adverse events and complications. A range of standard and newly designed investigative techniques were employed, and latter involving novel imaging techniques, for the objective assessment of physiological changes which occur with contact lens wear. The study is the first to combine these techniques with biochemical analyses of the tear film composition. Forty-seven subjects were fitted with SiH lenses and randomly allocated to one of the two materials currently on the market (Lotrafilcon A or Balafilcon A) on an either daily or continuous wear basis. An additional control group of 14 age-matched non-contact lens wearers were monitored over the same period. Measurements were taken before and 1, 3, 6, 12 and 18 months after initial fitting. The findings reported in this thesis will enable contact lens practitioners and manufacturers to understand further the optical, physiological and biochemical nature of the ocular response to SiH contact lenses and hence facilitate the development of this important generation of contact lens material.

Effect of extrusion and photo-oxidation on polyethylene/clay nanocomposites

Polyethylene (a 1:1 blend of m-LLDPE and z-LLDPE) double layer silicate clay nanocomposites were prepared by melt extrusion using a twin screw extruder. Maleic anhydride grafted polyethylene (PEgMA) was used as a compatibiliser to enhance the dispersion of two organically modified monmorilonite clays (OMMT): Closite 15A (CL15) and nanofill SE 3000 (NF), and natural montmorillonite (NaMMT). The clay dispersion and morphology obtained in the extruded nanocomposite samples were fully characterised both after processing and during photo-oxidation by a number of complementary analytical techniques. The effects of the compatibiliser, the organoclay modifier (quartenary alkyl ammonium surfactant) and the clays on the behaviour of the nanocomposites during processing and under accelerated weathering conditions were investigated. X-ray diffraction, transmission electron microscopy (TEM), scanning electron microscopy (SEM), rheometry and attenuated reflectance spectroscopy (ATR-FTIR) showed that the nanocomposite structure obtained is dependent on the type of clay used, the presence or absence of a compatibiliser and the environment the samples are exposed to. The results revealed that during processing PE/clay nanocomposites are formed in the presence of the compatibiliser PEgMA giving a hybrid exfoliated and intercalated structures, while microcomposites were obtained in the absence of PEgMA; the unmodified NaMMT-containing samples showed encapsulated clay structures with limited extent of dispersion in the polymer matrix. The effect of processing on the thermal stability of the OMMT-containing polymer samples was determined by measuring the additional amount of vinyl-type unsaturation formed due to a Hoffman elimination reaction that takes place in the alkyl ammonium surfactant of the modified clay at elevated temperatures. The results indicate that OMMT is responsible for the higher levels of unsaturation found in OMMT-PE samples when compared to both the polymer control and the NaMMT-PE samples and confirms the instability of the alkyl ammonium surfactant during melt processing and its deleterious effects on the durability aspects of nanocomposite products. The photostability of the PE/clay nanocomposites under accelerated weathering conditions was monitored by following changes in their infrared signatures and mechanical properties. The rate of photo-oxidation of the compatibilised PE/PEgMA/OMMT nanocomposites was much higher than that of the PE/OMMT (in absence of PEgMA) counterparts, the polymer controls and the PE–NaMMT sample. Several factors have been observed that can explain the difference in the photo-oxidative stability of the PE/clay nanocomposites including the adverse role played by the thermal decomposition products of the alkyl ammonium surfactant, the photo-instability of PEgMA, unfavourable interactions between PEgMA and products formed in the polymer as a consequence of the degradation of the surfactant on the clay, as well as a contribution from a much higher extent of exfoliated structures, determined by TEM, formed with increasing UV-exposure times.

A sulphonate based hydrogel for nucleus pulposus repair

Introduction: Lower back pain treatment and compensation costs >$80 billion overall in the US. 75% of back pain is due to disc degeneration in the lumbar region of the spine. Current treatment comprises of painkillers and bed rest or as a more radical solution – interbody cage fusion. In the early stages of disc degeneration the patient would benefit from addition of an injectable gel which polymerises in situ to support the degenerated nucleus pulposus. This involves a material which is an analogue of the natural tissue capable of restoring the biomechanical properties of the natural disc. The nucleus pulposus of the intervertebral disc is an example of a natural proteoglycan consisting of a protein core with negatively charged keratin and chondroitin sulphate attached. As a result of the high fixed charge density of the proteoglycan, the matrix exerts an osmotic swelling pressure drawing sufficient water into support the spinal system. Materials and Methods: NaAMPs (sodium 2- acrylamido 2-methyl propane sulphonic acid) and KSPA (potassium 3- sulphopropyl acrylate) were selected as monomers, the sulphonate group being used to mimic the natural sulphate group. These are used in dermal applications involving chronic wounds and have acceptably low cytotoxicity. Other hydrophilic carboxyl, amide and hydroxyl monomers such as 2-hydroxyethyl acrylamide, ß-carboxyethyl acrylate, acryloyl morpholine, and polyethylene glycol (meth)acrylate were used as diluents together with polyethyleneglycol di(meth)acrylate and hydrophilic multifunctional macromers as cross-linker. Redox was the chosen method of polymerisation and a range of initiators were investigated. Components were packaged in two solutions each containing a redox pair. A dual syringe method of injection into the cavity was used, the required time for polymerisation is circa 3-7 minutes. The final materials were tested using a Bohlin CVO Rheometer cycling from 0.5-25Hz at 37oC to measure the modulus. An in-house compression testing method was developed, using dialysis tubing to mimic the cavity, the gels were swelled in solutions of various osmolarity and compressed to ~ 20%. The pre-gel has also been injected into sheep spinal segments for mechanical compression testing to demonstrate the restoration of properties upon use of the gel. Results and Discussion: Two systems resulted using similar monomer compositions but different initiation and crosslinking agents. NaAMPs and KSPA were used together at a ratio of ~1:1 in both systems with 0.25-2% crosslinking agent, diacrylate or methacrylate. The two initiation systems were ascorbic acid/oxone, and N,N,N,N - tetramethylethylenediamine (TEMED)/ potassium persulphate. These systems produced gelation within 3-7 and 3-5 minutes respectively. Storage of the two component systems was shown to be stable for approximately one month after mixing, in the dark, refrigerated at 1-4oC. The gelation was carried out at 37oC. Literature values for the natural disc give elastic constants ranging from 3-8kPa. The properties of the polymer can be tailored by altering crosslink density and monomer composition and are able to match those of the natural disc. It is possible to incorporate a radio-opaque (histodenz) to enable x-ray luminescence during and after injection. At an inclusion level of 5% the gel is clearly visible and polymerisation and mechanical properties are not altered. Conclusion: A two-pac injection system which will polymerise in situ, that can incorporate a radio-opaque, has been developed. This will reinforce the damaged nucleus pulposus in degenerative disc disease restoring adequate hydration and thus biomechanical properties. Tests on sheep spine segments are currently being carried out to demonstrate that a disc containing the gel has similar properties to an intact disc in comparison to one with a damaged nucleus.

Integration of a 3D hydrogel matrix within a hollow core photonic crystal fibre for DNA probe immobilization

In this paper, we demonstrate the integration of a 3D hydrogel matrix within a hollow core photonic crystal fibre (HC-PCF). In addition, we also show the fluorescence of Cy5-labelled DNA molecules immobilized within the hydrogel formed in two different types of HC-PCF. The 3D hydrogel matrix is designed to bind with the amino groups of biomolecules using an appropriate cross-linker, providing higher sensitivity and selectivity than the standard 2D coverage, enabling a greater number of probe molecules to be available per unit area. The HC-PCFs, on the other hand, can be designed to maximize the capture of fluorescence to improve sensitivity and provide longer interaction lengths. This could enable the development of fibre-based point-of-care and remote systems, where the enhanced sensitivity would relax the constraints placed on sources and detectors. In this paper, we will discuss the formation of such polyethylene glycol diacrylate (PEGDA) hydrogels within a HC-PCF, including their optical properties such as light propagation and auto-fluorescence.

A decade of silicone hydrogel development:surface properties, mechanical properties, and ocular compatibility

Since the initial launch of silicone hydrogel lenses, there has been a considerable broadening in the range of available commercial material properties. The very mobile silicon–oxygen bonds convey distinctive surface and mechanical properties on silicone hydrogels, in which advantages of enhanced oxygen permeability, reduced protein deposition, and modest frictional interaction are balanced by increased lipid and elastic response. There are now some 15 silicone hydrogel material variants available to practitioners; arguably, the changes that have taken place have been strongly influenced by feedback based on clinical experience. Water content is one of the most influential properties, and the decade has seen a progressive rise from lotrafilcon-A (24%) to efrofilcon-A (74%). Moduli have decreased over the same period from 1.4 to 0.3 MPa, but not solely as a result of changes in water content. Surface properties do not correlate directly with water content, and ingenious approaches have been used to achieve desirable improvements (e.g., greater lubricity and lower contact angle hysteresis). This is demonstrated by comparing the hysteresis value of the earliest (lotrafilcon-A, >40°) and most recent (delefilcon-A, <10°) coated silicone hydrogels. Although wettability is important, it is not of itself a good predictor of ocular response because this involves a much wider range of physicochemical and biochemical factors. The interference of the lens with ocular dynamics is complex leading separately to tissue–material interactions involving anterior and posterior lens surfaces. The biochemical consequences of these interactions may hold the key to a greater understanding of ocular incompatibility and end of day discomfort.

Predicting success with silicone-hydrogel contact lenses in new wearers

Purpose: to evaluate changes in tear metrics and ocular signs induced by six months of silicone-hydrogel contact lens wear and the difference in baseline characteristics between those who successfully continued in contact lens wear compared to those that did not. Methods: Non-invasive Keratograph, Tearscope and fluorescein tear break-up times (TBUTs), tear meniscus height, bulbar and limbal hyperaemia, lid-parallel conjunctival folds (LIPCOF), phenol red thread, fluorescein and lissamine-green staining, and lid wiper epitheliopathy were measured on 60 new contact lens wearers fitted with monthly silicone-hydrogels (average age 36 ± 14 years, 40 females). Symptoms were evaluated by the Ocular Surface Disease Index (OSDI). After six months full time contact lens wear the above metrics were re-measured on those patients still in contact lens wear (n= 33). The initial measurements were also compared between the group still wearing lenses after six months and those who had ceased lens wear (n= 27). Results: There were significant changes in tear meniscus height (p= 0.031), bulbar hyperaemia (p= 0.011), fluorescein TBUT (p= 0.027), corneal (p= 0.007) and conjunctival (p= 0.009) staining, LIPCOF (p= 0.011) and lid wiper epitheliopathy (p= 0.002) after six months of silicone-hydrogel wear. Successful wearers had a higher non-invasive (17.0 ± 8.2. s vs 12.0 ± 5.6. s; p= 0.001) and fluorescein (10.7 ± 6.4. s vs 7.5 ± 4.7. s; p= 0.001) TBUT than drop-outs, although OSDI (cut-off 4.2) was also a strong predictor of success. Conclusion: Silicone-hydrogel lenses induced significant changes in the tear film and ocular surface as well as lid margin staining. Wettability of the ocular surface is the main factor affecting contact lens drop-out. © 2013 British Contact Lens Association.