Deficit of somatostatin-like immunoreactivity in the vitreous fluid of diabetic patients: possible role in the development of proliferative diabetic retinopathy

OBJECTIVE: To evaluate the vitreous levels of somatostatin-like immunoreactivity (SLI) in patients with proliferative diabetic retinopathy (PDR). RESEARCH DESIGN AND METHODS: A total of 14 diabetic patients with PDR, in whom a vitrectomy was performed, were included in the study. Sixteen nondiabetic patients, with other conditions requiring vitrectomy, served as a control group. Both venous blood and vitreous samples were collected at the time of vitreoretinal surgery. Patients in whom intravitreous hemoglobin was detectable were excluded. In addition, a correction for plasma levels of SLI and intravitreal proteins was performed. SLI was measured by radioimmunoassay and vitreous hemoglobin by spectrophotometry. RESULTS: SLI in the vitreous fluid was significantly lower in diabetic patients than in the control group (68 +/- 18.7 vs. 193.6 +/- 30.8 pg/ml, P < 0.01). The vitreous SLI-to-plasma SLI ratio was strikingly higher in nondiabetic subjects than in diabetic patients with PDR (5.3 [1.2-71.1] vs. 0.6 [0.03-4.1], P < 0.01). After correcting for total vitreous protein concentration, SLI (pg/mg of proteins) remained significantly higher in nondiabetic control subjects than in diabetic patients with PDR (186 [51-463] vs. 7.5 [0.8-82], P < 0.0001). Remarkably, intravitreous levels of SLI were higher than those obtained in plasma in nondiabetic control subjects (193.6 +/- 30.8 vs. 43.5 +/- 10.7 pg/ml, P < 0.0001). Finally, a lack of relationship between plasma and vitreous levels of SLI was observed in both diabetic patients with PDR and nondiabetic control subjects. CONCLUSIONS: The significantly higher SLI in the vitreous fluid than in plasma detected in nondiabetic control subjects supports the concept that somatostatin plays a relevant role in retinal homeostasis. In addition, the intravitreous deficit of SLI observed in diabetic patients with PDR suggests that it might contribute to the process of retinal neovascularization.     

Erythropoietin is expressed in the human retina and it is highly elevated in the vitreous fluid of patients with diabetic macular edema

OBJECTIVE: Erythropoietin has been recently found to be increased in the vitreous fluid from ischemic retinal diseases such as proliferative diabetic retinopathy (PDR). The aims of the present study were 1) to measure erythropoietin levels in the vitreous fluid from patients with diabetic macular edema (DME), a condition in which the ischemia is not a predominant event, and 2) to compare erythropoietin mRNA expression between human retinas from nondiabetic and diabetic donors without retinopathy. RESEARCH DESIGN AND METHODS: Vitreous samples from 12 type 2 diabetic patients with DME without significant retinal ischemia and 12 PDR patients were prospectively analyzed. Ten nondiabetic patients with macular holes served as the control group. Erythropoietin was assessed by radioimmunoassay (milliunits per milliliter). Erythropoietin mRNA expression was measured by quantitative real-time RT-PCR analysis in the retina from eight nondiabetic and eight age-matched diabetic donors without diabetic retinopathy RESULTS: Intravitreal erythropoietin concentration was higher in both PDR and DME patients than in nondiabetic control subjects (PDR vs. control subjects: median 302 [range 117-1,850] vs. 30 mU/ml [10-75], P < 0.01; DME vs. control subjects: 430 [41-3,000] vs. 30 mU/ml [10-75], P < 0.01). However, no significant differences were found between DME and PDR patients. Erythropoietin mRNA expression was detected in the human retina, and it was higher in the retina from diabetic than from nondiabetic donors. CONCLUSIONS: As occurs in PDR, intravitreous erythropoietin concentrations are strikingly higher in DME. Erythropoietin is expressed in the human retina, and it is upregulated in diabetic patients even without retinopathy. These findings suggest that other factors apart from ischemia are involved in the overexpression of erythropoietin in diabetic retinopathy.     

Vitreous levels of vascular cell adhesion molecule and vascular endothelial growth factor in patients with proliferative diabetic retinopathy: a case-control study

OBJECTIVE: To evaluate the intravitreous concentration of vascular cell adhesion molecule (VCAM)-1 in diabetic patients with proliferative diabetic retinopathy (PDR) and the relationship of VCAM-1 with vascular endothelial growth factor (VEGF). RESEARCH DESIGN AND METHODS: Serum and vitreous fluid samples were obtained simultaneously at the onset of vitrectomy from 20 diabetic patients with PDR and 20 nondiabetic control subjects with nonproliferative ocular disease. Both groups were matched by serum levels of VCGM-1 and VEGF. VCAM-1 and VEGF were determined by enzyme-linked immunosorbent assay. Statistics were determined using the Mann-Whitney U test and Spearman's rank correlation test. RESULTS: The intravitreous concentration of VCAM-1 was signifcantly elevated in diabetic patients with PDR compared with control subjects (26 ng/ml [19-118] vs. 22 ng/ml [20-47], P < 0.05). A direct correlation between VCAM-1 and total vitreous proteins was detected in diabetic patients (r = 0.64, P = 0.003), but not in control subjects. After adjusting for total intravitreous proteins, VCAM-1 was significantly lower in diabetic patients with PDR than in control subjects (8.2 ng/ml [4-31.4] vs. 43.1 ng/ml [9.7-100], P < 0.001). Intravitreous VEGF concentrations were higher in patients with PDR than in control subjects in absolute terms (1.34 ng/ml [0.16-6.22] vs. 0.009 ng/ml [0.009-0.044], P < 0.0001) and after correcting for total vitreal proteins (0.33 ng/ml [0.01-2.3] vs. 0.013 ng/ml [0.003-0.035], P = 0.0001). Finally, the vitreous ratio of VCAM-1 to proteins correlated with the vitreous ratio of VEGF to proteins in both diabetic patients (r = 0.74, P = 0.001) and control subjects (r = 0.84, P = 0.005). CONCLUSIONS: The low proportion of VCAM-1 in relation to total vitreal proteins observed in diabetic patients with PDR suggests that VCAM-1 is quenched by diabetic retina. In addition, the direct correlation detected between VCAM-1 and VEGF suggests that cellular adhesion and neovascularization may be linked processes.     

Free insulin-like growth factor 1 in the vitreous fluid of diabetic patients with proliferative diabetic retinopathy: a case-control study

The aim of the study was to evaluate the vitreous levels of free insulin-like growth factor 1 (IGF-1) in patients with proliferative diabetic retinopathy (PDR). For this, a total of 36 diabetic patients with PDR (group A) and 28 non-diabetic patients (group B) in whom a vitrectomy was performed were compared. Both groups were matched by age, sex and serum-free IGF-1. In a subgroup of diabetic patients (n =21) and non-diabetic patients (n =13), vitreous and serum total IGF-1, IGF-binding protein 1 (IGFBP-1) and IGFBP-3 were also determined. Serum and vitreous levels of free IGF-1, total IGF-1, IGFBP-1 and IGFBP-3 were measured by immunological methods. Vitreal proteins were assessed by a turbidimetric method and adjusted for vitreous haemoglobin. Vitreous levels of free IGF-1 were elevated in group A (median, 0.16 ng/ml; range 0.06-0.57 ng/ml) in comparison with group B (median, 0.12 ng/ml; range 0.06-0.22 ng/ml; P <0.001); however, after adjusting for vitreal proteins, free IGF-1 levels were significantly lower in group A in comparison with group B [0.05 ng/mg (0.01-0.45 ng/mg) versus 0.15 ng/mg (0.07-0.66 ng/mg); P <0.001]. The relatively lower free IGF-1 level observed in group A could not be attributed to differences in the distribution of intravitreous IGFBP-1 and IGFBP-3 in relation to total IGF-1. Notably, the contribution of free IGF-1 to total IGF-1 in vitreous fluid was 10% in group A and 42% in group B; these percentages largely exceed that obtained in serum (<1%). Our results suggest that although there is an enhancement of intravitreous free IGF-1 in diabetic patients due to serum diffusion, a deficit in its intraocular production also exists. In addition, these findings support the concept that intraocular-produced free IGF-1 plays a relevant role in retinal homoeostasis.     

Accumulation of NH2-terminal fragment of connective tissue growth factor in the vitreous of patients with proliferative diabetic retinopathy

OBJECTIVE: To evaluate the expression of connective tissue growth factor (CTGF) and its fragments in the vitreous of patients with proliferative diabetic retinopathy (PDR) and to localize CTGF expression in associated preretinal membranes. RESEARCH DESIGN AND METHODS: Vitreous was obtained from 24 patients with active PDR, 4 patients with quiescent PDR, and 23 patients with other retinal diseases and no diabetes, including 5 patients with vitreous hemorrhage. Enzyme-linked immunosorbent assay was used to determine levels of whole CTGF and its NH2- and COOH-terminal fragments. Preretinal membranes from three patients with active PDR were stained immunohistochemically for the presence of CTGF and cell type-specific markers. RESULTS: A significant increase in NH2-terminal CTGF fragment content was found in vitreous samples from patients with active PDR when compared with samples from nondiabetic patients (P<0.0001) or patients with quiescent PDR (P=0.02). Levels of NH2-terminal CTGF were also greater in vitreous samples from diabetic patients with vitreous hemorrhage compared with samples from nondiabetic patients with vitreous hemorrhage (P=0.02). Vitreous levels of whole CTGF were similar in all groups. COOH-terminal fragments of CTGF were not detected. CTGF immunoreactivity was predominantly localized to smooth muscle actin-positive myofibroblasts within active PDR membranes. CONCLUSIONS:-NH2-terminal CTGF fragment content is increased in the vitreous of patients with active PDR, suggesting that it plays a pathogenic role or represents a surrogate marker of CTGF activity in the disorder. The localization of CTGF in myofibroblasts suggests a local paracrine mechanism for induction of fibrosis and neovascularization.     

Role of circulating somatostatin in regulation of gastric acid secretion, gastrin release, and islet cell function. Studies in healthy subjects and duodenal ulcer patients.

Studies were designed (a) to determine whether somatostatin is released into the circulation after meals in sufficient amounts to regulate gastric or pancreatic islet function in humans and (b) to investigate the possible role of somatostatin in the pathogenesis of duodenal ulcer disease. Mean plasma somatostatin-like immunoreactivity (SLI) increased from 6.2 +/- 1.5 pg/ml to a peak level of 13.8 +/- 1.3 pg/ml in eight healthy subjects after a 1,440-cal steak meal (P less than 0.005). When somatostatin-14 was infused intravenously, basal and food-stimulated gastric acid secretion and also basal and food-simulated plasma insulin and glucagon concentrations were reduced significantly at mean plasma SLI concentrations within the range seen after a meal. Thus, the amount of somatostatin reaching the systemic circulation after a steak meal was sufficient to inhibit gastric acid secretion and islet cell function. On the other hand, basal and food-stimulated plasma gastrin concentrations were reduced by intravenous somatostatin only at plasma SLI concentrations that were several-fold greater than post-prandial SLI concentrations. Although duodenal ulcer patients had significantly higher basal, food-stimulated, and peak pentagastrin-stimulated gastric acid secretion rates than healthy controls, duodenal ulcer patients and controls had nearly identical basal and food-stimulated SLI concentrations. Moreover, food-stimulated gastric acid secretion and gastrin release were inhibited by intravenous somatostatin to the same extent in ulcer patients and controls. These studies suggest that duodenal ulcer patients release normal amounts of somatostatin into the circulation and that target cells controlling acid secretion and gastrin release are normally sensitive to somatostatin in these patients.     

彩色多普勒超声对糖尿病增殖性视网膜病变的诊断价值

目的探讨超声对糖尿病增殖性视网膜病变的诊断价值。方法68例（107只眼）糖尿病患者和60例（60只眼）健康对照组,分析其眼声像图特点。结果107只患眼,检出玻璃体出血32只眼,玻璃体机化膜61只眼,玻璃体下出血14只眼,玻璃体后脱离22只眼,视网膜脱离13只眼;视网膜中央动脉收缩末期和舒张末期血流速度较对照组降低（P〈0．05）,超声诊断符合99只眼,误诊7只眼,漏诊1只眼。结论超声在糖尿病增殖性视网膜病变的诊断中具有重要的价值。     

Radio-immunoassay of somatostatin from isolated rat pancreatic islets

Certain aspects of radio-immunoassay of somatostatin from isolated rat pancreatic islets are described. Somatostatin-14, and not somatostatin-28, is secreted from isolated rat pancreatic islets. Less somatostatin secretion is measured per islet owing to purity of tracer in the radio-immunoassay. Theophylline apparently cross-reacts with somatostatin in the assay described, and this has to be taken into consideration when studying somatostatin release induced by theophylline in isolated islets.     

Corneal advanced glycation end products increase in patients with proliferative diabetic retinopathy

OBJECTIVE: To evaluate corneal advanced glycation end product (AGE) fluorescence in patients with diabetes and in healthy control subjects. RESEARCH DESIGN AND METHODS: Corneal autofluorescence was measured in 26 eyes of 26 patients with type 2 diabetes (mean age 57.0 years; mean disease duration 12.2 years; mean HbA1c 7.1%) and 13 eyes of 13 healthy age-matched control subjects (mean age 57.9 years). The patients with type 2 diabetes were divided into the following groups: patients without diabetic retinopathy (DR), patients without proliferative diabetic retinopathy (PDR), and patients with PDR. Corneal autofluorescence was measured by fluorophotometry with the wavelength that is characteristic of AGE fluorescence (excitation and emission 360-370 nm and 430-450 nm, respectively). We defined peak corneal autofluorescence levels as corneal AGE fluorescence values. We compared the corneal AGE fluorescence values in the four groups. RESULTS: In the PDR group (11.9 +/- 3.9 arbitrary units [mean +/- SD]), the corneal AGE fluorescence values were significantly higher compared with the control subjects (6.9 +/- 1.3 arbitrary units), the patients without DR (7.4 +/- 2.1 arbitrary units), and the patients without PDR (6.9 +/- 2.2 arbitrary units) (P < 0.05). CONCLUSIONS: We found that corneal AGEs may increase in patients with diabetes and PDR compared with control subjects, patients without DR, and patients without PDR. In the patients with PDR, increased corneal AGEs may play a role in diabetic keratopathy.     

Plasma somatostatin 28 increases in response to feeding in man.

Tissue somatostatin-like immunoreactivity (SLI) consists of a number of molecular species including the cyclic tetradecapeptide or SRIF, an N-terminally extended form of SRIF termed somatostatin-28, as well as larger precursor peptides. The function and nature of circulating SLI is not well understood. In this report, we describe techniques for the definition of the components of plasma SLI in normal human plasma. Plasma SLI measured after gel filtration on Bio-gel P-6 columns was found to consist of from 1-3 peaks. The void volume peak was present in greatest concentration (34.2 +/- 8.9 pg/ml) and did not increase in response to a mixed meal. Very low levels of two additional peaks of SLI activity were found. To further characterize these peaks, 10-ml plasma samples were extracted and concentrated on octadecylsilyl silica (C-18) cartridges with subsequent fractionation on Bio-gel P-6 columns. The two peaks that coeluted with synthetic SRIF and S-28 markers, respectively, were present in concentrations of 5.4 +/- 1.4 and 4.8 +/- 1.9 pg/ml in fasting plasma. In response to a mixed meal, the SLI14 peak doubled (12.9 +/- 2.4 pg/ml) while the SLI28 peak increased to 29.9 +/- 7.2 pg/ml at 120 min. These results provide evidence that S-28 circulates in human plasma and its increase after feeding is consistent with a possible biological role for this peptide.     

Biosynthesis of immunoreactive somatostatin by hypothalamic neurons in culture.

The neuronal biosynthesis of somatostatin-like immunoreactivity (SLI) was investigated using mechanically dispersed neonatal rat hypothalamic cells kept in culture for up to 6 wk. Immunohistochemically, SLI was specifically localized to a small subpopulation of parvicellular neurons and their cell processes. By radioimmunoassay the cellular SLI content declined steadily during the first 2 wk in culture (nadir value of 60 fmol/dish at day 15) but then increased progressively to reach a maximum value of 381 fmol/dish at day 46. Gel chromatographic analysis showed this immunoreactivity to consist of forms corresponding to tetradecapeptide somatostatin (S-14), somatostatin-28 (S-28), and a 15,000-mol-wt molecule. After incubation of the cells with [3H]phenylalanine, the cellular extracts, purified by adsorption to C18 silica, contained material that bound specifically to an immobilized antisomatostatin antibody. Analysis by gel chromatography and high performance liquid chromatography of the specifically bound label provided evidence for the presence of labeled S-14, S-28, and the 15,000-mol-wt molecule. Pulse-chase experiments (20-min pulse, 20-min chase) demonstrated a transfer of radioactivity from the 15,000-mol-wt form to material corresponding to S-14 as well as to S-28. These studies demonstrate that cultured hypothalamic neurons are capable of synthesizing three somatostatin-like peptides (15,000-mol-wt SLI, S-28, S-14), one of which (15,000-mol-wt SLI) serve as a biosynthetic precursor for both S-28 and S-14. This in vitro system should provide a powerful tool for further investigation of the biosynthesis and regulation of biosynthesis of somatostatin in the hypothalamus.     

Elevated remnant-like particle cholesterol and triglyceride levels in diabetic men and women in the Framingham Offspring Study

OBJECTIVE: Remnants of triglyceride-rich lipoproteins are thought to be atherogenic. A new antibody-based assay allows for the isolation of remnant-like particles (RLPs) from plasma or serum, and the subsequent measurement of RLP cholesterol (RLPC) and triglycerides (RLPTGs). We hypothesized that diabetic patients would have higher remnant levels than nondiabetic patients. DESIGN AND METHODS: We compared RLPC and RLPTG levels of diabetic subjects (68 women, 121 men) participating in the Framingham Heart Study with those of nondiabetic subjects (1,499 women, 1,357 men). RESULTS: Mean RLPC values for diabetic women were 106% higher than those for nondiabetic women (0.367 +/- 0.546 mmol/l [14.2 +/- 21.1 mg/dl] vs. 0.179 +/- 0.109 mmol/l [6.9 +/- 4.2 mg/dl]; P < 0.0001), and RLPTG values for diabetic women were 385% higher than those for nondiabetic women (1.089 +/- 2.775 mmol/l [93.1 +/- 245.6 mg/dl] vs. 0.217 +/- 0.235 mmol/l [19.2 +/- 20.8 mg/dl]; P < 0.0001). Similar but less striking differences were observed in diabetic men, who had mean RLPC values 28% higher than those seen in nondiabetic men (0.285 +/- 0.261 mmol/l [11.0 +/- 10.1 mg/dl] vs. 0.223 +/- 0.163 mmol/l [8.6 +/- 6.3 mg/dl]; P < 0.001) and mean RLPTG values 70% higher than those seen in nondiabetic men (0.606 +/- 1.019 mmol/l [53.6 +/- 90.2 mg/dl] vs. 0.357 +/- 0.546 mmol/l [31.6 +/- 48.3 mg/dl]; P < 0.001). Moreover, diabetic men and women had significantly higher total triglycerides and lower HDL cholesterol levels than nondiabetic subjects. CONCLUSIONS: The data indicate that RLP particles are elevated in diabetic subjects. To achieve optimal reduction of risk for cardiovascular disease, treatment of elevated RLP values, along with the control of LDL cholesterol levels, should be considered.     

Vitreous levels of IGF-I, IGF binding protein 1, and IGF binding protein 3 in proliferative diabetic retinopathy: a case-control study

OBJECTIVE: To evaluate vitreous levels of IGF-I and its binding proteins IGFBP-1 and IGFBP-3 in patients with proliferative diabetic retinopathy (PDR). Because intravitreal proteins are elevated in patients with PDR due to the disruption of the blood-retinal barrier, we have corrected vitreal IGF-I and IGFBPs by total vitreal proteins to avoid this confounding factor. RESEARCH DESIGN AND METHODS: We compared 21 diabetic patients with proliferative retinopathy (group A) and 13 nondiabetic patients (group B) in whom a vitrectomy was performed. Both groups were matched by age, serum IGF-I, IGFBP-1, and IGFBP-3 levels. Serum and vitreous levels of IGF-I, IGFBP-1, and IGFBP-3 were measured by immunological methods. Vitreal proteins were assessed by turbidimetric method. RESULTS: Vitreal levels of IGF-I were elevated in group A (median 1.35 ng/ml [range 0.3-8.7]) in comparison with group B (median 0.25 ng/ml [range 0-1.38]), P<0.001. After adjusting by vitreal proteins [ratio IGF-I (ng/ml)/protein (mg/ml)], the differences remain significant (P<0.005). Vitreal levels of IGFBP-1 and IGFBP-3 were also elevated in diabetic patients (IGFBP-1: group A, median 1.6 ng/ml [range 0.6-20.7]; group B, median 0.4 ng/ml [range 0.3-1.9], P<0.001. IGFBP-3: group A, median 102.6 ng/ml [range 53.9-350.8]; group B, median 29.0 ng/ml [range 3.2-87.8], P<0.001). However, when the ratio IGFBP/protein was considered, the differences were not significant. CONCLUSIONS: Intraocular synthesis contributes to elevated vitreous concentrations of IGF-I found in PDR. By contrast, unspecific increase of intravitreal proteins is the main factor explaining the elevated vitreous levels of IGFBP-1 and IGFBP-3 found in diabetic patients.     

Cortistatin, a new antiinflammatory peptide with therapeutic effect on lethal endotoxemia

Cortistatin is a recently discovered cyclic neuropeptide related to somatostatin that has emerged as a potential endogenous antiinflammatory factor based on its production by and binding to immune cells. Because human septic shock involves excessive inflammatory cytokine production, we investigated the effect of cortistatin on the production of inflammatory mediators and its therapeutic action in various murine models of endotoxemia. Cortistatin down-regulated the production of inflammatory mediators by endotoxin-activated macrophages. The administration of cortistatin protected against lethality after cecal ligation and puncture, or injection of bacterial endotoxin or Escherichia coli, and prevented the septic shock-associated histopathology, such as infiltration of inflammatory cells and intravascular disseminated coagulation in various target organs. The therapeutic effect of cortistatin was mediated by decreasing the local and systemic levels of a wide spectrum of inflammatory mediators, including cytokines, chemokines, and acute phase proteins. The combined use of cortistatin and other antiinflammatory peptides was very efficient treating murine septic shock. This work provides the first evidence of cortistatin as a new immunomodulatory factor with the capacity to deactivate the inflammatory response. Cortistatin represents a potential multistep therapeutic agent for human septic shock, to be used in combination with other immunomodulatory agents or as a complement to other therapies.     

Overexpression of Hemopexin in the Diabetic Eye: A new pathogenic candidate for diabetic macular edema

OBJECTIVE

Hemopexin is a well-recognized permeability factor in the kidney, but its potential role in blood-retinal barrier (BRB) breakdown has not been explored. The main aims of this study were as follows: 1) to determine hemopexin expression in the retina and its content in the vitreous fluid from diabetic patients with diabetic macular edema (DME) and nondiabetic patients, 2) to evaluate the effect of hemopexin on BRB permeability, and 3) to determine whether dexamethasone prevents an eventual hemopexin-induced hyperpermeability.

RESEARCH DESIGN AND METHODS

Biological material included 1) retinas from 10 diabetic donors with nonproliferative retinopathy and from 10 nondiabetic donors and 2) vitreous fluid from 14 patients with DME and 14 nondiabetic patients. Hemopexin and hemopexin receptor mRNA levels were measured by quantitative RT-PCR and hemopexin concentrations by ELISA. The effect of hemopexin on permeability in culture was evaluated in human retinal pigment epithelial (ARPE)-19 cells and bovine retinal endothelial cells. The experiments were repeated in the presence of hemopexin-neutralizing antibodies and dexamethasone.

RESULTS

A higher expression of hemopexin was detected in the retinal pigment epithelium (RPE) from diabetic patients in comparison with nondiabetic control subjects. Intravitreal hemopexin concentration was higher in patients with DME than in nondiabetic subjects. Hemopexin significantly increased permeability in ARPE-19 cells, which was prevented by both hemopexin-neutralizing antibodies and dexamethasone.

CONCLUSIONS

Hemopexin is overexpressed in the RPE of diabetic patients with DME and induces the breakdown of RPE cells in vitro. Dexamethasone was able to prevent hemopexin-induced hyperpermeability. Our results suggest that hemopexin can be considered a new pathogenic candidate for DME.Diabetic retinopathy remains the leading cause of preventable blindness among working-age individuals in developed countries (1). Whereas proliferative diabetic retinopathy (PDR) is the commonest sight-threatening lesion in type 1 diabetes, diabetic macular edema (DME) is the primary cause of poor visual acuity in type 2 diabetes. Because of the high prevalence of type 2 diabetes, DME is the main cause of visual impairment in diabetic patients (2). When clinically significant DME appears, laser photocoagulation is currently indicated. However, the optimal period of laser treatment is frequently passed and, moreover, is not uniformly successful in halting visual decline. In addition, photocoagulation is not without side effects, with visual field loss and impairment of either adaptation or color vision being the most frequent. Intravitreal corticosteroids have been successfully used in eyes with persistent DME and loss of vision after the failure of conventional treatment. However, reinjections are commonly needed, and there are substantial adverse effects such as infection, glaucoma, and cataract formation. Intravitreal anti–vascular endothelial growth factor (VEGF) agents have also found an improvement of visual acuity and decrease of retinal thickness in DME, even in nonresponders to conventional treatment (3). However, apart from local side effects such as endophthalmitis and retinal detachment, the response to treatment of DME by VEGF blockade is not prolonged and is subject to significant variability. For all these reasons, new pharmacological treatments based on the understanding of the pathophysiological mechanisms of DME are needed.Vascular leakage due to the breakdown of the blood-retinal barrier (BRB) is the main event involved in the pathogenesis of DME (4). However, little is known regarding the molecules primarily involved in this event. By means of a proteomic analysis, we have found that hemopexin was significantly increased in the vitreous fluid of patients with DME in comparison with PDR and nondiabetic control subjects (5). Hemopexin is the best characterized permeability factor in steroid-sensitive nephrotic syndrome (6,7). In addition, hemopexin could be involved in either causing or perpetuating enhanced glomerular permeability in minimal change nephrotic syndrome (7). T cell–associated cytokines like tumor necrosis factor-α are able to enhance hemopexin production in mesangial cells in vitro, and this effect is prevented by corticosteroids (8). However, whether hemopexin also acts as a permeability factor in the BRB and its potential response to corticosteroids remains to be elucidated.On these bases, the aims of the current study were 1) to compare hemopexin and hemopexin receptor (LDL receptor–related protein [LRP1]) levels in retina and in vitreous fluid from diabetic and nondiabetic patients, 2) to evaluate the effect of hemopexin on the permeability of outer and inner BRB in cell cultures, and 3) to determine whether anti-hemopexin antibodies and dexamethasone were able to prevent an eventual hemopexin-induced hyperpermeability.     

Effect of a 12-hour subcutaneous infusion of somatostatin-14 on blood glucose, insulin and glucagon levels in healthy subjects and insulin dependent diabetics

We compared the influence of a 12-hour subcutaneous infusion of somatostatin-14 on glucose homeostasis in two normal subjects and two insulin-dependent diabetics (IDD). In all cases, somatostatin infusion led to a decrease of blood glucose during the first hours. The diabetogenic effect of somatostatin was confirmed in normal subjects. Postprandial blood glucose response was exaggerated in one insulin-dependent diabetic while in the other, glucose tolerance was improved. Unexpected high levels of immunoreactive somatostatin were measured in insulin-dependent diabetic patients. They might be due to a decreased somatostatin catabolism.     

Changes in vitreous concentrations of human hepatocyte growth factor (hHGF) in proliferative diabetic retinopathy: implications for intraocular hHGF production

We measured human hepatocyte growth factor (hHGF) concentrations in the original vitreous and in the artificial vitreous after vitrectomy in 13 patients with proliferative diabetic retinopathy (PDR) undergoing repeated pars plana vitrectomy, in order to investigate whether the vitreous hHGF concentration is related to the recurrence of PDR after vitrectomy as well as to the original occurrence of PDR. We also examined the relationship between vitreous concentrations of hHGF and transforming growth factor-beta(2) (TGF-beta(2)), the predominant TGF-beta isoform in the vitreous, in 14 patients with PDR. For the original vitreous, mean hHGF concentrations were higher (P<0.05) in that from patients with severe PDR (vitreous haemorrhage, fibrovascular proliferation and tractional retinal detachment) than in that from patients with vitreous haemorrhage alone. In the artificial vitreous, mean vitreous hHGF concentrations were higher (P<0.05) in that from patients with severe PDR than in that from patients with vitreous haemorrhage alone or with vitreous haemorrhage plus fibrovascular proliferation. No correlation was found between the hHGF concentration in the artificial vitreous and time between vitrectomies. Vitreous hHGF concentrations were directly proportional to vitreous concentrations of latent TGF-beta(2) (r=0. 831; P=0.0002), but inversely proportional to vitreous concentrations of active TGF-beta(2) (r=0.495; P=0.072), which inhibits hHGF production. A decreased conversion of latent into active TGF-beta(2) in ocular disorders such as PDR is likely to result in an increased concentration of hHGF in the vitreous. Thus intraocular hHGF may be involved in pathological mechanisms causing not only the occurrence, but also the recurrence, of PDR.     

Foot pressure measurements in diabetic and nondiabetic amputees.

OBJECTIVE--Foot problems are common in the remaining foot of diabetic amputees. Because high foot pressures are associated with foot ulceration, we studied foot pressures of the remaining foot of diabetic and nondiabetic unilateral amputees. RESEARCH DESIGN AND METHODS--Four age-matched groups of 11 subjects were studied. The groups were comprised of diabetic subjects with previous major amputation, nondiabetic nonneuropathic amputees, diabetic nonamputee patients with similar peripheral nerve involvement as the diabetic amputees, and nondiabetic control subjects. Vibration perception threshold (VPT) was assessed by biothesiometry and foot pressures with an optical pedobarograph. RESULTS--Mean +/- SE VPT in the diabetic amputees was significantly higher than the nondiabetic amputees (40.2 +/- 3.7 vs. 17.7 +/- 2.8 V, P less than 0.002) and similar to diabetic nonamputees (43.4 +/- 3.3 V, NS). VPT was abnormal in 9 (82%) diabetic amputees, 2 (18%) nondiabetic amputees, and 10 (91%) nonamputee diabetic patients. The mean peak foot pressure in the diabetic amputees was higher compared with nondiabetic amputees (18.3 +/- 2.2 vs. 11.3 +/- 1.4 kg/cm2, P less than 0.05) and control subjects (10.0 +/- 1.5 kg/m2, P less than 0.01), but no difference existed with diabetic nonamputees. Abnormally high foot pressures (greater than 12.3 kg/cm2) were present in 8 (73%) diabetic amputees, 3 (27%) nondiabetic amputees, 8 (73%) diabetic nonamputees, and 4 (36%) healthy subjects. CONCLUSIONS--We conclude that high pressures are present under the remaining foot in diabetic amputees, and that these pressures are associated with diabetic neuropathy. Prosthetic usage does not increase the pressures under the remaining foot in nondiabetic amputees.     

Prevalence of carotid atherosclerosis in diabetic patients. Ultrasound high-resolution B-mode imaging on carotid arteries.

OBJECTIVE--To quantitatively assess atherosclerosis of the carotid artery in subjects with and without diabetes. RESEARCH DESIGN AND METHODS--Ultrasound high resolution B-mode imaging of carotid arteries was conducted on 71 nondiabetic subjects without hyperlipidemia or hypertension and 295 diabetic patients to determine IMT of the arterial wall. RESULTS--IMT was linearly related with age in nondiabetic (IMT = [0.0087 x age] + 0.3318) and diabetic subjects (IMT = [0.0155 x age] + 0.32450). The regression coefficient for age was significantly greater in diabetic than nondiabetic subjects. IMT (mean +/- SD) of diabetic subjects aged 20-29 was significantly greater than that of nondiabetic subjects aged 20-29 (0.73 +/- 0.27 vs. 0.52 +/- 0.07 mm, P less than 0.01). Multivariate regression analysis of 275 NIDDM patients indicated smoking, hyperlipidemia, duration of diabetes, hypertension, and age were factors determining thickness of the carotid arterial wall. CONCLUSIONS--Diabetes, along with age, hyperlipidemia, smoking, and hypertension, aggravates carotid atherosclerosis.     

Measurement, Characterization, and Source of Somatostatin-like Immunoreactivity in Human Amniotic Fluid

Somatostatin-like immunoreactivity (SLI) is widely distributed in tissues and biological fluids. To determine whether SLI is also present in amniotic fluid, samples obtained by amniocentesis from 30 normal and 27 abnormal pregnancies were studied by radioimmunoassay. Direct incubation of [(125)I-Tyr(1)]tetradecapeptide somatostatin (SRIF) with amniotic fluid resulted in 89% tracer degradation. Damage was reduced to <5% when samples were acidified and boiled before the assay. With this technique, SLI was detectable in all normal amniotic fluid samples; the mean level at 15-20 wk of gestation (320+/-55 pg/ml, n = 15) being 4.5 times higher than the mean at 32-43 wk (70+/-12 pg/ml, n = 15) (P < 0.001). In cases of preeclampsia (n = 6), gestational diabetes (n = 5), anencephaly (n = 1), and meningomyelocele (n = 1), SLI values were in the normal range, but in one juvenile diabetic and one patient with chronic renal failure, SLI was undetectable (<10 pg/ml). In a pair of monochorionic diamniotic twins, SLI levels were very different (33 and 197 pg/ml), which suggests that fetal factors are more important than materno-placental ones in determining amniotic fluid SLI. Serial dilutions of amniotic fluid showed parallelism with standard SRIF. When concentrates of pooled amniotic fluid were chromatographed on Sephadex G-25 columns, all SLI eluted in the void volume ahead of SRIF even after treatment with 8 M urea and dithiothreitol. This "big" SLI incubated in amniotic fluid showed 100% stability over 24 h at 37 degrees C, whereas SRIF was rapidly inactivated (t((1/2)) congruent with 7 min). Extracts of placenta and fetal membranes contained no SLI, but small amounts (6-20% of total amniotic fluid SLI) were found in cells from fresh fluid. Radioimmunoassay of SLI in extracts of seven paired cord arterial and venous plasma samples showed no arteriovenous gradient consistent with fetal origin of cord blood SLI. It is concluded that (a) amniotic fluid contains SLI which is of fetal origin and (b) normal levels vary with gestational age. The SLI has a higher molecular weight (>/=5,000) and is more stable in amniotic fluid than SRIF.