Inter-relationships between DNA damage, ascorbic acid and glycaemic control in Type 2 diabetes mellitus.

AIMS: The onset of complications in Type 2 diabetes mellitus (DM) patients cannot be predicted in individuals. Evidence suggests a link between complications and hyperglycaemia, oxidative stress and antioxidants, but causality is unclear. This study investigated baseline (entry) fasting plasma ascorbic acid, lymphocytic DNA damage and glycaemic control in Type 2 DM as part of a long-term study, the aim of which is to explore a biomarker profiling approach to identify and improve outcome in high-risk subjects. METHODS: A cross-sectional study, in which DNA damage, glycated haemoglobin (HbA(1c)), fasting plasma glucose (FPG) and ascorbic acid (AA) were measured on fasting blood samples collected from 427 Type 2 DM subjects. RESULTS: DNA damage was significantly (P < 0.0001) and directly correlated to both FPG (r = 0.540) and HbA(1c) (r = 0.282), and was significantly (P < 0.0001), independently and inversely correlated to plasma AA (r = -0.449). In those subjects with both poor glycaemic control and low AA (< 48 microm, the overall mean value for the study group), DNA damage was significantly (P < 0.005) higher compared with those subjects with a similar degree of hyperglycaemia but with AA above the mean. CONCLUSIONS: The novel finding of a significant inverse relationship between plasma AA and DNA damage in Type 2 DM indicates that poorly controlled diabetic subjects might benefit from increased dietary vitamin C. The data also have important implications for biomarker profiling to identify those subjects who might benefit most from intensive therapy. Longer-term follow-up is underway.

The effect of conventional CR39 and Fresnel prisms on high and low contrast acuity.

The effect of conventional CR39 and Fresnel prisms on high and low contrast letter acuity was studied. Visual acuity of the fully corrected better eyes of 15 subjects was measured with the high (90%) and low (10%) contrast logMAR letter charts, while they wore prisms of varying power. The results showed that when the power of the conventional CR39 prism and the Fresnel prism reached 10 prism dioptres and 5 prism dioptres, respectively, significant reduction (1 tail t-test, p < 0.05) of the high and low contrast acuity occurred. The Fresnel prism caused a significantly greater acuity reduction than the conventional CR39 prism for powers ranging from 5 to 30 prism dioptres for both contrasts. The rate of acuity reduction with increasing prism power was greater with the low contrast targets than with the high contrast targets for both prisms. In addition, the rate of acuity reduction with increasing prism power was greater with the Fresnel prism than with the conventional CR39 prism for both contrasts. The conventional CR39 prism reduced acuity by a ratio of about 0.8-0.9 of that of the Fresnel prism for powers ranging from 5 to 20 prism dioptres and by about 0.7 for 30 prism dioptres. These ratios applied for both high and low contrast acuity, and therefore were independent of the level of contrast used.

Vision of low astigmats through thick and thin lathe-cut soft contact lenses.

Distance and near visual acuity of 13 low astigmats were determined in a double-masked experiment through thick and thin (centre thickness 0.12 mm and 0.06 mm, respectively) spherical lathe-cut soft lenses. For each lens type, distance and near LogMAR VA and over-refraction were assessed with different logMAR VA charts. For 70% of the subjects, the residual astigmatism was significantly lower than the refractive astigmatism with thicker lenses. No statistically significant differences in the distance and near logMAR VA was found between the two lens types using any of the charts used, though, in general, logMAR VA obtained through the thicker lens was better than logMAR VA through the thinner lens. The variabilities in distance and near logMAR VA between the two lens types increased with decreased contrast. The variabilities in distance logMAR VA were greater with Chinese charts than with English charts, and LogMAR VA with Chinese charts were significantly worse for both lens types. Based on the results of this study, we concluded that thicker spherical lathe-cut soft lenses provide better vision in low astigmats. The Snellen acuity test is inadequate for vision assessment of soft contact lens wearers. When a patient wearing thin soft contact lenses complains of poor vision in spite of 6/6 or 6/5 Snellen acuity, changing to thicker lenses may be considered.

The calibration of a 2.5x Galilean focusable telescope as an optometer for refraction.

A 2.5x Selsi achromatic Galilean focusable telescope was calibrated for refraction at 6 m. In its calibration, minus power trial lens was placed at the objective of the telescope to simulate vergence of a target at a finite distance, before the back vertex power (BVP) of the telescope at each setting (telescope length) was measured by a focimeter. By using a graphical presentation of the results, the BVP of the telescope at each setting could be determined at different selected target distances. For a target vergence of -0.167 D or a distance of 6 m, the common testing distance in clinical practice, this calibrated telescope had a BVP or refraction measuring range of -7.27(-)+7.52 D. When this telescope was used to measure simulated manifest refractive errors at 6 m, it yielded a mean error of +0.13 D with a 95% confidence limit of agreement of -0.38(-)+0.64 D. These results indicated that the accuracy and precision of telescopic refraction were comparable to that of retinoscopy. Therefore, the calibrated telescope could be considered as a reliable and inexpensive instrument for determining spherical refractive errors. Telescopic refraction is applicable in refracting economically disadvantaged population in underserved areas where modern equipment and electricity are not available. In addition, it provides an alternative subjective refraction method for low vision population because the magnification of this calibrated telescope has the advantage of allowing low vision patients to be refracted at the common 6 m testing distance in clinical practice.

Comparison of the caldesmon content of cardiac and smooth muscle.

The high abundance of caldesmon in smooth muscle and its ability to inhibit actomyosin ATPase activity have led to the hypothesis that caldesmon modulates contractile activity. It has also been proposed, however, that caldesmon acts as a structural protein in muscle and non-muscle cells. We have determined the caldesmon content of mammalian cardiac muscle and have found that caldesmon is 200-fold less abundant in cardiac muscle than it is in gizzard smooth muscle. This finding argues against a role for caldesmon in the modulation of cardiac contractility.

Identification and localization of caldesmon in cardiac muscle.

Caldesmon has been detected in smooth muscle and in a number of non-muscle cells. It binds both actin and myosin and may act as a regulator of contraction or a structural element in smooth muscle. The presence of caldesmon in striated muscle has not been well established. To address this issue, polyclonal antibodies and a panel of monoclonal antibodies were raised against chicken gizzard smooth muscle caldesmon and used to demonstrate that caldesmon is present in adult cardiac muscle of a variety of mammalian species. Western-blot analysis revealed the presence of caldesmon in ventricular myocytes isolated from rat heart. The epitopes for the individual monoclonal antibodies were mapped to the caldesmon primary structure using chymotryptic and 2-nitro-5-thiocyanatobenzoic acid fragments. Bovine and rat cardiac caldesmons were recognized only by a subset of these monoclonal antibodies, indicating primary sequence differences from the chicken smooth muscle protein. Immunofluorescence labelling of isolated myocytes from rat, rabbit and guinea pig cardiac muscle revealed a striated pattern of fluorescence labelling. Dual labelling of caldesmon and myosin or caldesmon and alpha-actinin demonstrated that caldesmon was present at the centre of the I-band rather than in the A-band, as might have been expected from the myosin binding properties of the smooth muscle protein. These results suggest a structural role for caldesmon in cardiac muscle cells.

Scattering properties of Bagolini lenses and their effects on spatial vision.

The effect of a Bagolini lens on spatial vision was investigated by studying its far-field diffraction pattern as produced by a coherent beam of laser light, and its effect on the contrast sensitivity function (CSF) in human subjects. For lenses of the main type studied, which were crossed by a series of slightly-irregular striated bands, each consisting of fine, parallel, etched lines of various widths and separations, the diffraction pattern consisted of undiffracted light giving a bright central spot and wide-angle, diffracted light giving a dim streak. The latter was due to the sum of the diffraction patterns associated by the irregular fine etched lines. The streak produced by a single striated band was modulated by a series of regular maxima and minima related to the width of the band. Analysis of this pattern gave the width of the band as 0.6 mm, in close agreement with direct microscopical measurements. When four bands were illuminated by a beam of about 3 mm diameter, similar to the diameter of the photopic pupil, the diffraction pattern showed no obvious maxima and minima, due to irregularity in the width and separation of the bands. The central spot contained more than 90% of the total light in the diffraction pattern. Thus the Bagolini lens, with its relatively weak far-field diffraction pattern lacking regular maxima and minima when areas > or = 3 mm in diameter were used, was expected to have only a small effect on the apparent contrast of the targets in CSF experiments. This was confirmed by the measurements: Bagolini lenses showed no significant effect on either the monocular or binocular CSF. Further similar measurements with lenses of slightly different design from another manufacturer confirmed these findings. Therefore Bagolini lenses do not disrupt vision when they are used to determine the presence of suppression and anomalous retinal correspondence.

The validity of current clinical tests of contrast sensitivity and their ability to predict reading speed in low vision.

PURPOSE: Contrast sensitivity (CS) testing using chart tests of CS is becoming increasingly common in low vision assessment. Yet we know little about the validity of these charts, i.e. which region of the spatial frequency spectrum is being measured. In this study we aimed to determine the validity of currently available CS charts by comparison against oscilloscope-based CS. We also determined their relative ability to predict reading speed. METHODS: CS was measured with five commercially available charts and the contrast sensitivity function was determined with sinusoidal gratings presented on a Joyce screen using a two-alternative forced choice staircase technique in 36 observers with low vision and 3 with normal vision. Reading rate was also measured with the subject reading with his or her own optical low vision aid. RESULTS: The results show that the Pelli-Robson chart and the Cambridge gratings are good measures of medium to low spatial frequencies, as would be predicted from their design, while the Regan and UW charts correlated with medium to high frequencies. The Vistech chart was a good predictor of CS at each spatial frequency. CONCLUSIONS: The best chart test of CS depends on which region of the CS curve is of interest. All the charts were good predictors of reading rate.

International cooperative exchange programs in clinical optometry.

A reciprocal exchange program between the University of Waterloo and the Hong Kong Polytechnic in clinical optometry is presented. Costs and benefits are described.

Normal values of eye position and head size in Chinese children from Hong Kong.

We measured the eye position and head dimensions of Chinese children in Hong Kong. Values for exophthalmos, interpupillary distance (IPD), interorbital distance (IOD), distance between medial canthi, and head dimensions were found to be larger than those for Caucasian, Black, or other Chinese groups.

Notch in contrast sensitivity function of optical origin: diffraction effects of acrylic filters.

A series of retinal image degrading filters was evaluated by measuring the contrast sensitivity function of four human subjects through the filters (residual CSF). The acrylic filters, with regularly spaced cross-hatches, produced progressively more reduction in the residual CSF as the density of the cross hatching increased. For some of the filters there was a selective loss of a narrow band of spatial frequencies as a result of diffraction effects. This experiment serves to further emphasize the need to rule out optical causes of such notches in the CSF before making a diagnosis of neurological dysfunction.

Eye position and head size in the Chinese population: a comparison of the Chinese from Hong Kong with the Chinese from Guangdong Province.

Measurements obtained in this study include facial and head dimensions. From our findings, it would appear that the Chinese population in Hong Kong have not become different in the two or three generations that separate them from those living in Guangdong province of China.

Normal values of eye position in the Chinese population of Hong Kong.

Exophthalmos, interpupillary distance (IPD), interobital distance (IOD), and inner intercanthal distance (ICD) were measured in an adult Chinese population from Hong Kong (HKC). Mean values and normal range for 95% of the population were determined and the relation with head size and body height examined. Values of exophthalmos and IPD in our Chinese population were similar to those given for Caucasian groups. IOD and ICD were larger in the Chinese than in adult Caucasians. Our findings show generally larger values in the HKC than has been found for other populations in mainland China.

Purification and characterization of calmodulin-dependent multifunctional protein kinase from smooth muscle: isolation of caldesmon kinase.

Previously, it was reported that smooth muscle caldesmon is a protein kinase and is autophosphorylated [Scott-Woo, G.C., & Walsh, M.P. (1988) Biochem. J. 252, 463-472]. We separated a Ca2+/calmodulin-dependent protein kinase from caldesmon in the presence of 15 mM MgCl2. The Ca2+/calmodulin-dependent caldesmon kinase was purified by using a series of liquid chromatography steps and was characterized. The subunit molecular weight (MW) of the kinase was 56K by SDS gel electrophoresis and was autophosphorylated. After the autophosphorylation, the kinase became active even in the absence of Ca2+/calmodulin. The substrate specificity of caldesmon kinase was similar to the rat brain calmodulin-dependent multifunctional protein kinase II (CaM PK-II) and phosphorylated brain synapsin and smooth muscle 20-kDa myosin light chain. The purified kinase bound to caldesmon, and the binding was abolished in the presence of high MgCl2. Enzymological parameters were measured for smooth muscle caldesmon kinase, and these were KCaM = 32 nM, KATP = 12 microM, Kcaldesmon = 4.9 microM, and KMg2+ = 1.1 mM. Optimum pH was 7.5-9.5. The observed properties were similar to brain CaM PK-II, and, therefore, it was concluded that smooth muscle caldesmon kinase is the isozyme of CaM PK-II in smooth muscle.

Current methods of treating and preventing myopia.

In this review, we discuss and compare current methods of treating and preventing myopia including radial keratotomy, keratomileusis, keratophakia, epikeratoplasty, keratokyphosis, scleral reinforcement, phakoemulsification, and heat application. Among the visual training methods are such procedures as biofeedback and behavior modification. The use of drugs, orthokeratology, spectacles, bifocals, prisms, intraocular lenses, contact lenses, and ultrasound are described.

Kinase activity associated with caldesmon is Ca2+/calmodulin-dependent kinase II.

The relationship of the kinase which co-purifies with caldesmon to Ca2+/calmodulin-dependent protein kinase II (CaM-kinase II) was investigated by studying the phosphorylation of bovine brain synapsin I, as well-characterized substrate of CaM-kinase II. Synapsin I is a very good substrate (Km = 90 nM) of the co-purifying kinase, which phosphorylates two sites in synapsin I, both of which are distinct from the single site phosphorylated by cyclic-AMP-dependent protein kinase. Phosphorylation of synapsin I is Ca2(+)- and calmodulin-dependent: half-maximal activation occurs at 0.13 microM-Ca2+ and maximal activity at 0.4 microM-Ca2+. Phosphorylation of the co-purifying kinase slightly enhances the rate, but does not alter the stoichiometry, of subsequent synapsin I phosphorylation; it does, however, circumvent the requirement for Ca2+ and calmodulin. The properties of this kinase therefore closely resemble those of CaM-kinase II, and we conclude that it is probably a smooth-muscle isoenzyme of CaM-kinase II.

Characterization of the autophosphorylation of chicken gizzard caldesmon.

Caldesmon, an actin- and calmodulin-binding protein of smooth muscle, is a protein serine/threonine kinase capable of Ca2+/calmodulin-dependent autophosphorylation [Scott-Woo & Walsh (1988) Biochem. J. 252, 463-472]. Phosphorylation nullifies the inhibitory effect of caldesmon on the actin-activated Mg2+-ATPase activity of smooth-muscle myosin [Ngai & Walsh (1987) Biochem. J. 244, 417-425]. We have characterized the kinase activity of caldesmon of chicken gizzard smooth muscle. Autophosphorylation requires Ca2+/calmodulin, but is unaffected by other second messengers (Ca2+/phospholipid/diacylglycerol, cyclic AMP or cyclic GMP), and is inhibited by the calmodulin antagonists chlorpromazine and compound 48/80, with 50% inhibition at 39.8 microM and 12.0 ng/ml respectively. Half-maximal activation of autophosphorylation occurs at 60-80 nM-Ca2+ and 0.14 microM-calmodulin, and maximal activity at 0.14-0.18 microM-Ca2+ and 1 microM-calmodulin; activation is gradually lost at higher Ca2+ and calmodulin concentrations. Autophosphorylation is pH-dependent, with maximal activity over the range pH 7-9, and requires free Mg2+ in addition to the MgATP2- substrate. The Km for ATP is 15.6 +/- 4.1 microM (mean +/- S.D., n = 4), and kinase activity is inhibited by increasing ionic strength [half-maximal inhibition at I = 0.094 +/- 0.009 M (mean +/- S.D., n = 4)]. Autophosphorylation does not affect the rate of hydrolysis of caldesmon (free or bound to calmodulin) by alpha-chymotrypsin. However, a slight difference in peptides generated from phospho- and dephospho-forms of caldesmon is observed. The binding of phospho- or dephospho-caldesmon to F-actin protects the protein against chymotryptic digestion, but does not alter the pattern of peptide generation. Characterization of proteolytic fragments of caldesmon generated by alpha-chymotrypsin and Staphylococcus aureus V8 protease enables localization of the phosphorylation sites and the kinase active site within the caldesmon molecule.

Autophosphorylation of smooth-muscle caldesmon.

Caldesmon, a major actin- and calmodulin-binding protein of smooth muscle, has been implicated in regulation of the contractile state of smooth muscle. The isolated protein can be phosphorylated by a co-purifying Ca2+/calmodulin-dependent protein kinase, and phosphorylation blocks inhibition of the actomyosin ATPase by caldesmon [Ngai & Walsh (1987) Biochem. J. 244, 417-425]. We have examined the phosphorylation of caldesmon in more detail. Several lines of evidence indicate that caldesmon itself is a kinase and the reaction is an intermolecular autophosphorylation: (1) caldesmon (141 kDa) and a 93 kDa proteolytic fragment of caldesmon can be separated by ion-exchange chromatography: both retain caldesmon kinase activity, which is Ca2+/calmodulin-dependent; (2) chymotryptic digestion of caldesmon generates a Ca2+/calmodulin-independent form of caldesmon kinase; (3) caldesmon purified to electrophoretic homogeneity retains caldesmon kinase activity, and elution of enzymic activity from a fast-performance-liquid-chromatography ion-exchange column correlates with caldesmon of Mr 141,000; (4) caldesmon is photoaffinity-labelled with 8-azido-[alpha-32P]ATP; labelling is inhibited by ATP, GTP and CTP, indicating a lack of nucleotide specificity; (5) caldesmon binds tightly to Affi-Gel Blue resin, which recognizes proteins having a dinucleotide fold. Autophosphorylation of caldesmon occurs predominantly on serine residues (83.3%), with some threonine (16.7%) and no tyrosine phosphorylation. Autophosphorylation is site-specific: 98% of the phosphate incorporated is recovered in a 26 kDa chymotryptic peptide. Complete tryptic/chymotryptic digestion of this phosphopeptide followed by h.p.l.c. indicates three major phosphorylation sites. Caldesmon exhibits a high degree of substrate specificity: apart from autophosphorylation, brain synapsin I is the only good substrate among many potential substrates examined. These observations indicate that caldesmon may regulate its own function (inhibition of the actomyosin ATPase) by Ca2+/calmodulin-dependent autophosphorylation. Furthermore, caldesmon may regulate other cellular processes, e.g. neurotransmitter release, through the Ca2+/calmodulin-dependent phosphorylation of other proteins such as synapsin I.