Connective tissue growth factor (CTGF/CCN2) in haemophilic arthropathy and arthrofibrosis: a histological analysis.

INTRODUCTION: Joint haemorrhage is the principal clinical manifestation of haemophilia frequently leading to advanced arthropathy and arthrofibrosis, resulting in severe disability. The degree and prevalence of arthrofibrosis in hemophilic arthropathy is more severe than in other forms of arthropathy. Expression of connective tissue growth factor (CTGF) has been linked to many fibrotic diseases, but has not been studied in the context of haemophilic arthropathy. AIM: We aim to compare synovial tissues histologically from haemophilia and osteoarthritis patients with advanced arthropathy in order to compare expression of proteins that are possibly aetiologic in the development of arthrofibrosis. METHODS: Human synovial tissues were obtained from 10 haemophilia and 10 osteoarthritis patients undergoing joint surgery and processed for histology and immunohistochemistry. RESULTS: All samples from haemophilia patients had synovitis with hypertrophy and hyperplasia of synovial villi. Histologically, synovial tissues contained hyperplastic villi with increased cellularity and abundant haemosiderin- and ferritin-pigmented macrophage-like cells (HMCs), with a perivascular localization in the sub-surface layer. CTGF staining was observed in the surface layer and sub-surface layer in all haemophilia patients, exclusively co-localizing with HMCs. Quantification showed that the extent of CTGF-positive areas was correlated with the degree of detection of HMCs. CTGF was not observed in any of the samples from osteoarthritis patients. CONCLUSION: Using histological analysis, we showed that CTGF expression is elevated in haemophilia patients with arthrofibrosis and absent in patients with osteoarthritis. Additionally, we found that CTGF is always associated with haemosiderin-pigmented macrophage-like cells, which suggests that CTGF is produced by synovial A cells following the uptake of blood breakdown products.

Taking small steps towards targets - perspectives for clinical practice in diabetes, cardiometabolic disorders and beyond.

Big changes are hard. When trying to achieve guideline targets in diabetes and cardiometabolic disorders, patients can lack commitment or suffer despondency. It is much easier to make small changes in lifestyle or treatment, which are less noticeable and easier to manage long-term. Obesity is central to the cardiometabolic disorders, and even small weight losses of 2-5% can improve the cardiometabolic risk profile and substantially reduce the risk of developing type 2 diabetes. Likewise, small increases in physical activity, such as 15-30 min of brisk walking per day, can cut the risk of heart disease by 10%. Lifestyle or treatment changes that lead to small improvements in metabolic parameters also impact patient outcome - for example, a 5 mmHg decrease in blood pressure can translate into significant reductions in the rates of myocardial infarction and cardiovascular mortality. Benefits of small changes can also be seen in health economic outcome models. Implementing change at an individual versus a population level has different implications for overall benefit and patient motivation. Even very small steps taken in trying to reach guideline targets should represent a positive achievement for patients. Patient engagement is essential - only when patients commit themselves to change can benefits be maintained, and physicians should recognise their influence. Small changes in individual parameters can result in significant beneficial effects; however, a major impact can occur when small changes are made together in multiple parameters. More research is required to elucidate the full impact of small changes on patient outcome.

Control of cardiovascular risk factors in patients with type 2 diabetes and hypertension in a French national study (Phenomen).

UNLABELLED: For some years now, the recommendations of scientific societies have significantly reduced the therapeutic targets for blood pressure, glycaemia and lipid levels in diabetic patients. However, little is known regarding the synchronization between effective risk factor management and the guidelines. To examine this issue, the Phenomen survey was conducted between January and July 2001 on 16358 patients suffering from hypertension followed by a general practitioner in France. AIM: To evaluate the control of cardiovascular risk factors in patients with diabetes and hypertension according to the French guidelines. METHODS AND PATIENTS: 8177 general practitioners, selected from a national database according to quotas, taking into account age, practice and area, had to include the first two hypertensive patients they came across in their practice and to collect their demographic data, cardiovascular risk factors and medications. RESULTS: 2346 out of 16358 hypertensive patients presented with type 2 diabetes (14.3% of the cohort). The number of GP consultations in the last 12 months averaged 8.31. According to the French guidelines, 6.5% had a blood pressure<140/80 mmHg, a total of 38.7% patients met the goal of LDL cholesterol level and 26.6% of patients had an HbA1c<6.5%, 53.4% of patients had an HbA1c between 6.6 and 8%. 37.1% of patients continued to receive antihypertensive monotherapy but only 3% in this monotherapy group reached the target of 130/85 mmHg. 29% of the patients were on antiplatelet therapy. 64.6% of these hypertensive diabetic patients presented with more than three other cardiovascular risk factors. Based on WHO recommendations, 0.3% of the patients met all of the blood pressure, lipid and glycaemic treatment objectives. CONCLUSION: Despite frequent monitoring by a general practitioner, the overall management of modifiable risk factors in this diabetic hypertensive population is clearly inadequate. The impact of the guidelines on effective management remains limited and additional information is required to understand why physicians are not more aggressive in managing modifiable risk factors in diabetic patients.

[Is assessment of kidney involvement in patients with essential hypertension satisfactory? General medical observation results in a national study (PHENOMEN)]. / L'évaluation de l'atteinte rénale est-elle satisfaisante chez l'hypertendu essentiel? Observatoire en médecine générale résultats d'une enquête nationale (PHENOMEN).

UNLABELLED: According to ANAES guidelines (2000) adapted from the international guidelines, kidney function should be regularly monitored in hypertensive patients in order to detect vascular nephropathy at an early stage and prevent kidney failure which is a major cardiovascular factor. The PHENOMEN survey was conducted between January and July 2001 on a representative sample of 16,358 patients suffering from hypertension being followed by a General Practitioner (GP). The population included patients from all 22 administrative regions in France. AIMS: To determine the prevalence of renal involvement in patients with hypertension (both treated and untreated); to compare the GP's subjective evaluation of the extent of renal involvement with a more objective assessment made according to ANAES guidelines. METHODS: Kidney function was evaluated by measuring plasma creatinine levels to derive a value for the creatinine clearance rate (CCR) using the Cockeroft-Gault formula. RESULTS: Out of the 16,358 patients included, plasma creatinine levels could be measured in 11,586 (76%) [53% men; 47% women, mean age = 63 +/- 12 years]: mean plasma creatinine = 90 +/- 49 mumol/l, i.e. a mean CCR of 83 +/- 32 ml/min. For 8,650 of the patients (75%), kidney function had been monitored in the preceding twelve months. The physicians defined their patient's kidney function as normal in 10,080 (87%) of cases whereas in reality, 4,411 of 8,650 (51%) were suffering from some degree of renal insufficiency (RI): in 2,422 (28%), RI was mild (> or = 60 CCR < 80 ml/min): in 1,903 (22%) RI was moderate (> or = 30 CCR < 60 ml/min). A minority of patients (44 i.e. 1%) showed severe RI (CCR < 30 ml/min). CONCLUSION: Most GPs apply ANAES guidelines and order plasma creatinine assays. However, there is a discrepancy between how the physician "perceives" the seriousness of the renal insufficiency and a more objective measure based on the CCR figure. It would seem that the CCR derivative is insufficiently exploited despite the widespread availability of converters. Thus, the PHENOMEN survey has revealed that the extent of renal involvement in patients with hypertension still tends to be underestimated by GPs despite the major implications of this complication in terms of therapeutic strategy and global cardiovascular risk.

[Management of essential arterial hypertension and cardiovascular risk levels. Observation in general medicine: national PHENOMEN study. Evaluation and stratification of cardiovascular risk based on the 2000 ANAES recommendations in a population of hypertensive patients]. / Prise en charge de l'HTA essentielle et du niveau de risque cardiovasculaire. Observatoire en médecine générale: enquête nationale PHENOMEN. Evaluation et stratification du risque cardiovasculaire selon les recommandations de l'ANAES 2000 dans une population d'hypertendus.

OBJECTIVE: To define the prevalence of cardiovascular risk (CVR) levels in a population of hypertensive patients (whether treated or not) monitored by General Practitioners, using the stratification system proposed by the ANAES in 2000. METHODS: Between January and July 2001, a nation-wide survey was carried out based on a representative sample of 8,177 General Practitioners spread evenly throughout all of the 22 administrative regions in France. The evaluation of CVR levels took into account blood pressure readings as well as CVR factors and indicators. RESULTS: This survey covered 16,358 patients (53.2% men, 46% women; mean age = 62.5 +/- 11.9 years; BMI = 27.3 +/- 4.5 kg/m2) with a mean history of hypertension of 7 +/- 7 years. The most common CVR factors were dyslipidemia (59.5%), smoking (19%) and diabetes (16%). Concomitant target organ damage was recorded in 17% and heart disease in 21.8%. One patient in four had more than three CVR factors; respectively 56.5%, 30.9% and 12.5% of the patients had mild, moderate or severe hypertension. More than 50% of the patients were classified as being at "high" or "very high" CV risk. The distribution of risk levels was similar throughout the country with a North-South gradient of the high/very high levels. Only 17% of the patients being treated had completely normal blood pressure (< 140/190 mmHg), although 19% more count as normal if the limit values are included. CONCLUSION: The main objective of this large-scale, nation-wide epidemiological survey was to evaluate how well CVR is being managed in a representative sample of patients with high blood pressure. In general, none of the risk factors is adequately controlled, especially in populations considered as being at high CVR. Moreover, the notion of CVR has only been partially assimilated by General Practitioners.

[Differences between management guidelines and global health strategies for arterial hypertension with metabolic disorders in France in 1999. Ohara study]. / Différences entre recommandations de prise en charge et stratégie globale médicale dans l'hypertension artérielle avec désordres métaboliques en France en 1999. Etude Ohara.

Taking into account all the risk factors and blood pressure levels, as indicated by several American and European recommendations available since 1997, is a leading strategy to reduce mortality and morbidity of hypertensive patients. The aim of this study was to quantify how, in 1999. French practitioners applied recommendations on hypertension (HTN), diabetes and hypercholesterolemia in recently diagnosed hypertensive patients and to evaluate whether or not the recommended targets were met. 1639 French GPs and cardiologists included 5831 recently diagnosed (7.5 +/- 3.6 months) hypertensives (57 +/- 12 years of age, M/F = 55/45%). Initial BP was 173 +/- 15/99 +/- 9 mmHg. 56% had no concomitant disease, 36% had either diabetes, dyslipidemia or coronary heart disease, 8% had at least two concomitant diseases. At the time of the study corresponding to 6.3 +/- 3.8 months after initiation of diet and/or medical treatment, their BP was 148 +/- 17/86 +/- 11 mmHg (-25/-19 mmHg). At that time only 37% of patients with stage 2 HTN were encouraged to adopt lifestyle modifications without any medical treatment as recommended by the JNC VI. Among these hypertensives, measurement of plasma cholesterol was performed in only 61%, HDL-C/LDL-C in 26% and blood glucose in 51%. In the patients with dyslipidemia, LDL-C was measured in only 47%. In the 677 diabetic patients only 27% had a glycated hemoglobin measurement. The percentage of patients reaching target BP was 59% as regard DBP < 90 mmHg, 25% as regard SBP < 140 mmHg, and 23% reached both target values of BP. In addition, 30% of patients with dyslipidemia reached the target LDL-C as defined by French recommendations (ANAES 1996) and 30% of the diabetic patients reached the target value for glycemia recommended by ADA (1997). In 1999 in France, a minority of patients reaches the national or international recommended target values for blood pressure, glycemia and plasma LDL-cholesterol. In spite of that, French practitioners do not implement all the available diagnostic tools to improve the treatment of metabolic disorders in hypertensive patients. As a conclusion, to improve the prognosis of hypertensive patients, it is mandatory to raise the awareness of physicians about multiple risk factor management and help them implement the recommendations in their daily practice.

Phosphorothioate oligonucleotides inhibit the intrinsic tenase complex by an allosteric mechanism.

Phosphorothioate oligonucleotides (PS ODNs) prolong the activated partial thromboplastin time in human plasma by inhibition of intrinsic tenase (factor IXa-factor VIIIa) activity. This inhibition was characterized using ISIS 2302, a 20-mer antisense PS ODN. ISIS 2302 demonstrated hyperbolic, mixed-type inhibition of factor X activation by the intrinsic tenase complex. The decrease in V(max(app)) was analyzed by examining complex assembly, cofactor stability, and protease catalysis. ISIS 2302 did not inhibit factor X activation by the factor IXa-phospholipid complex, or significantly affect factor VIII-phospholipid affinity. Inhibitory concentrations of ISIS 2302 modestly decreased the affinity of factor IXa-factor VIIIa binding in the presence of phospholipid (K(D) = 11.5 vs 4.8 nM). This effect was insufficient to explain the reduction in V(max(app)). ISIS 2302 did not affect the in vitro half-life of factor VIIIa, suggesting it did not destabilize cofactor activity. In the presence of 30% ethylene glycol, the level of factor X activation by the factor IXa-phospholipid complex increased 3-fold, and the level of chromogenic substrate cleavage by factor IXa increased more than 50-fold. ISIS 2302 demonstrated partial inhibition of factor X activation by the factor IXa-phospholipid complex, and chromogenic substrate cleavage by factor IXa, only in the presence of ethylene glycol. Like the intact enzyme complex, ISIS 2302 demonstrated hyperbolic, mixed-type inhibition of chromogenic substrate cleavage by factor IXa (K(I) = 88 nM). Equilibrium binding studies with fluorescein-labeled ISIS 2302 demonstrated a similar affinity (K(D) = 92 nM) for the PS ODN-factor IX interaction. These results suggest that PS ODNs bind to an exosite on factor IXa, modulating catalytic activity of the intrinsic tenase complex.

Combinatorial synthesis of small-molecule libraries using 3-amino-5-hydroxybenzoic acid.

A non-peptide library of 2001 compounds has been prepared utilizing solid-phase techniques. The split/combine method was demonstrated to work well to form mixtures of compounds based on 3-amino-5-hydroxybenzoic acid as a core structure. The benzoic acid of the core structure served as the attachment point for the resin and the amino and hydroxy positions were variably substituted.

Topical cyclosporine A and corneal wound healing.

The effect of 2% topical cyclosporine A (CsA) ointment on corneal epithelial and stromal wound healing was evaluated in a masked, placebo-controlled study in rabbits. The difference in epithelial would healing rate in the treatment and placebo group was not statistically significant. There was also no difference in the values of bursting strength of 2-mm central penetrating wounds and in the measurement of hydroxyproline content in the central corneal buttons from the wounded area in both the groups. Clinical evaluation of the quality of regenerating epithelium, stromal edema, haze, and iritis did not reveal a difference between the two groups. There was a significantly higher incidence of conjunctival injection in the CsA-treated group. We conclude that topical cyclosporine A 2% does not significantly inhibit epithelial or stromal corneal wound healing.

Topical fibronectin in an alkali burn model of corneal ulceration in rabbits.

We studied the effect of topical fibronectin on epithelial wound healing and ulceration in alkali-burned rabbit corneas. After the first 56 hours, fibronectin accelerated complete surface reepithelialization to 4.3 +/- 2.3 days. Control alkali-burned corneas treated with phosphate-buffered saline or albumin did not resurface for 6.7 +/- 3.7 days and 6.2 +/- 2.5 days, respectively. When recurrent epithelial defects occurred, the time required for healing was also significantly accelerated by fibronectin treatment. Corneal ulceration developed in 25 of 28 and 15 of 18 saline and albumin-treated control eyes, respectively; only nine of 18 fibronectin-treated eyes ultimately ulcerated. Immunohistologic studies showed that the initially deposited fibronectin-fibrinogen matrix on the surface of burned corneas had disintegrated by 72 to 96 hours after wounding, corresponding clinically to the time of secondary epithelial breakdown. A prominent fibronectin-fibrinogen matrix remained on the surface of fibronectin-treated corneas, presumably aiding surface reepithelialization and decreasing corneal ulceration.

Ophthalmic beta-blockers: determination of plasma and aqueous humor levels by a radioreceptor assay following multiple doses.

We determined the binding affinities of multiple doses of four ophthalmic beta-blockers, timolol, betaxolol, levobunolol and carteolol, to the beta-1 and beta-2 receptors. With a Ki value of 0.39 nM and 0.36 nM for the beta-1 and beta-2 receptors, respectively, levobunolol shows the highest binding affinity to both beta receptors. The Ki values of timolol (1.97 nM for the beta-1 receptor and 2.0 nM for the beta-2 receptor) and of carteolol (0.83 nM and 0.85 nM for the beta-1 and beta-2 receptors, respectively) are characteristic of a nonspecific beta-blocker. On the contrary, betaxolol is a beta-1 specific antagonist (Ki of 23.33 nM) and has a very low binding affinity to the beta-2 receptor (Ki of 200.00 nM). With a radioreceptor assay, levels of beta-antagonist were measured in the plasma and aqueous humor 1 hour and 12 hours after instillation of 50 microliters of 0.5% or 2% each of the four beta-blockers into the rabbit eye. At 1 hour after administration, the plasma levels of timolol, levobunolol, and carteolol are 9.89 ng/ml, 1.60 ng/ml and 8.00 ng/ml, respectively; such levels of 11.82 to 29.22 times the respective Ki values cause a virtually total blockade of both beta-1 and beta-2 receptors and suggest significant systemic absorption. Betaxolol has a peak 1 hour plasma level of 22.28 ng/ml, which is equivalent to only 3.08 times its Ki for the beta-1 receptor and 0.36 times its Ki for the beta-2 receptor; it has less systemic beta-1 blocking activity than the other three drugs and very minimal systemic beta-2 blocking activity. Peak aqueous humor concentrations of all 4 beta-blockers are extremely elevated at 1 hour after administration (timolol 1613.58 ng/ml; betaxolol 866.06 ng/ml; levobunolol 750.89 ng/ml; and carteolol 859.18 ng/ml). Such levels of 14 to 7192 times the respective Ki values should cause a virtually complete blockade of both beta-1 and beta-2 receptors in the iris-ciliary body complex. At 12 hours after administration, plasma levels of all four beta blockers remain moderately elevated (timolol 0.94 ng/ml; betaxolol 9.43 ng/ml; levobunolol 0.66 ng/ml; and carteolol 1.61 ng/ml). Trough aqueous humor levels of levobunolol (43.38 ng/ml) and carteolol (92.81 ng/ml) remain elevated at least 300 times their Ki value. On the contrary, 12-hour trough aqueous humor levels of timolol (33.67 ng/ml) and betaxolol (94.86 ng/ml) have decreased to 2 to 40 times their Ki value, which may explain their requirement for twice daily administration clinically.

Circadian variations and reference intervals for some enzymes in urine of healthy children.

Circadian variations of alanine aminopeptidase (EC 3.4.11.2), gamma-glutamyltransferase (EC 2.3.2.2), and N-acetyl-beta-glucosaminidase (EC 3.2.1.30) in urine were studied in 10 healthy children, ages six to 15 years. Urine specimens were collected during 24 h, grouped into four time intervals. Enzymes were measured spectrophotometrically, with automation. These enzymes all showed diurnal variation, with morning (8 a.m.-12 noon) excretion being highest. We also analyzed timed urinary specimens (8 a.m.-12 noon) from 136 healthy children, ages two to 11 years. Reference intervals are presented for these enzymes. High excretion of the three enzymes was observed in children two and three years old.

Role of fibronectin and fibrinogen in healing of corneal epithelial scrape wounds.

A provisional fibronectin (Fn)-fibrinogen (Fg) matrix forms de novo on the bare basement membrane (BM) surface of superficial epithelial scrape wounds in rabbit and guinea pig (GP) corneas. We determined whether such a substrate is essential for epithelial cell adhesion and migration, using three different approaches. (1) Polyclonal and monoclonal IgG reactive against GP Fn were topically administered to inhibit Fn formation in a GP epithelial scrape wound model. Immunofluorescence studies showed that deposition of both Fn and Fg was inhibited in antibody-treated corneas. During the first 38 hr after wounding, the healing rates were 0.46 +/- 0.06 mm2/hr in control eyes, 0.43 +/- 0.05 mm2/hr in those treated with rabbit polyclonal IgG anti-GP Fn and 0.45 +/- 0.09 mm2/hr in those treated with murine monoclonal IgG anti-GP Fn (P greater than 0.4). (2) In a rabbit epithelial scrape wound model, ancrod was administered intravenously to induce systemic Fg depletion. Fg deposition was completely inhibited on the wound surface, but Fn deposition was not suppressed. The healing rate was 1.24 +/- 0.41 mm2/hr in ancrod-treated corneal wounds and 1.19 +/- 0.29 mm2/hr in control eyes during the first 48 hr after wounding (P greater than 0.5). These data from the antibody and ancrod inhibition indicate that Fg binds to Fn and that Fn binds to components other than Fg.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of fibronectin in the healing of superficial keratectomies in vitro.

A fibronectin (Fn)-fibrinogen (Fg) surface matrix is not essential for epithelial cell migration in a corneal epithelial scrape wound model, in which the basement membrane is preserved. We have therefore tested whether such a provisional scaffolding becomes more critical in a superficial keratectomy model, when the basement membrane is surgically removed. Exogeneous Fn at 0.3 mg/ml was added to the medium of organ cultures of rabbit superficial keratectomies. At 48 hr after wounding, the healing rate was 1.12 +/- 0.03 mm2/hr in control corneas and 1.11 +/- 0.03 mm2/hr in those cultured with Fn. At 64 hr after wounding, the healing rates were also not significantly different (P greater than 0.5). Immunofluorescence studies showed that Fn was not detectable on the surface of control corneas but could be depicted under the migrating epithelium. In corneas cultured with Fn, it diffused throughout the entire stroma but did not deposit as a surface matrix. We therefore attempted to obtain formation of a provisional Fn-Fg surface matrix before establishment of the in vitro organ culture by leaving the superficial keratectomies in vivo for 8 hr, 24 hr or 64 hr. At 64 hr after wounding, their healing rate was 0.80 +/- 0.04 mm2/hr, 0.86 +/- 0.04 mm2/hr and 0.85 +/- 0.06 mm2/hr, respectively, which were not significantly different from that of contralateral ex vivo-wounded cultured corneas (0.83 +/- 0.04 mm2/hr, P greater than 0.5). Immunofluorescence studies revealed a Fn matrix on the bare surface of in vivo-wounded specimen, which was not detectable on ex vivo-wounded cultured corneas; there was also no diffuse stromal Fn.(ABSTRACT TRUNCATED AT 250 WORDS)

Topical fibronectin in the treatment of persistent corneal epithelial defects and trophic ulcers.

Six patients with persistent corneal epithelial defects that failed to improve with standard therapy were treated with topical fibronectin in a phase I uncontrolled trial. Fibronectin was purified from autologous plasma by affinity chromatography and administered topically, 0.5 mg/ml five times a day, for three weeks. Complete reepithelialization was achieved in five patients: three healed three to 17 days after initiation of treatment while two healed one to two weeks after discontinuation of topical fibronectin. No side effects were observed. Over a follow-up period of three to 18 months, two recurrences were noted.

Ascorbic acid therapy in a thermal burn model of corneal ulceration in rabbits.

We studied the effect of topical and systemic ascorbic acid on a rabbit model of corneal ulceration produced by thermal burn. Central stromal ulceration occurred in 77% to 100% of eyes in all treatment groups. The rate of ulceration was accelerated in animals treated with systemic ascorbic acid. Ulceration began after 5.80 +/- 2.14 days in control eyes and 2.75 +/- 1.26 days in the eyes of rabbits treated with both topical and systemic ascorbic acid (P less than .02). Subconjunctival ascorbic acid alone did not affect the rate of ulceration (average onset, 6 +/- 1 days). Once ulceration occurred in animals treated with both topical and systemic ascorbic acid, the incidence of perforation and descemetocele formation was increased and that of healing was decreased. The time required for healing was not significantly altered by treatment (9.50 +/- 2.28 days for control eyes vs 5.75 +/- 2.78 days for treated eyes) (P greater than .05).