Ozagrel reverses streptozotocin-induced constriction of arterioles in rat retina

Retinal blood flow decreases early in the progression of diabetic retinopathy; however, the mediators and mechanisms responsible for this decrease have yet to be determined. In this study, diabetes was induced by streptozotocin in rats, and retinal blood flow was measured via intravital microscopy 1 or 3 weeks following the induction of hyperglycemia. Additionally, retinal arteriolar diameters and flow were measured prior to and following acute administration of the thromboxane synthase inhibitor ozagrel to investigate the potential role of thromboxane in the observed constriction. Minimal changes in the retinal diameters and flow were observed at 1 week of diabetes; however, at 3 weeks of diabetes, arteriolar constriction and decreases in blood flow were significant. Notably, the constriction occurred only in the arterioles that were in closer proximity to the venules draining the retina. Acute administration of ozagrel reversed the constriction of the closely venule-paired arterioles. In summary, the results suggest that thromboxane mediates localized, venule-dependent arteriolar constriction induced by streptozotocin-induced diabetes in rats.     

Apoptotic death of photoreceptors in the streptozotocin-induced diabetic rat retina

Aims/hypothesis Neurodegenerative changes in the diabetic retina occurring before diabetic retinopathy could be inevitable by the altered energy (glucose) metabolism, in the sense that dynamic image-processing activity of the retinal neurons is exclusively dependent on glucose. We therefore investigated the morphological changes in the neural retina, including neuronal cell death, of a streptozotocin-induced model of diabetes.Methods Streptozotocin was intravenously injected. Rats were maintained hyperglycaemic without insulin treatment for 1 week and 4, 8, 12, and 24 weeks, respectively. Diabetic retinas were processed for histology, electron microscopy, and immunohistochemistry using the TUNEL method.Results A slight reduction in the thickness of the inner retina was observed throughout the diabetic retinas and a remarkable reduction was seen in the outer nuclear layer 24 weeks after the onset of diabetes. The post-synaptic processes of horizontal cells in the deep invaginations of the photoreceptors showed degeneration changes from 1 week onwards. A few necrotic ganglion cells were observed after 4 weeks. At 12 weeks, some amacrine cells and a few horizontal cells showed necrotic features. Three to seven cellular layers in the outer nuclear layer and nerve terminals, rolled by the fine processes of the Müller cells near the somata of the degenerated ganglion cells, were apparent at 24 weeks. Apoptosis appeared in a few photoreceptor cells at 4 weeks, and the number of apoptotic photoreceptors increased thereafter.Conclusion/interpretation These findings suggest that the visual loss associated with diabetic retinopathy could be attributed to an early phase of substantial photoreceptor loss, in addition to later microangiopathy.Abbreviations TUNEL Terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling - GCL ganglion cell layer - IPL inner plexiform layer - INL inner nuclear layer - OPL outer plexiform layer - ONL outer nuclear layer     

Endothelium-dependent constriction demonstrated in vivo in mouse cerebral arterioles

Endothelium-dependent constriction in mouse pial arterioles was demonstrated by testing contractile responses before and after endothelial injury. All responses were monitored at the same site as the injury. Injury was produced in vivo by exposing an arteriole on the brain surface to the beam of a 6-mW helium-neon laser after first sensitizing the microvascular bed to the laser energy by injecting Evans blue intravenously. The contractile response to serotonin creatinine sulfate (20 micrograms/ml) and to sodium arachidonate (30 micrograms/ml) was monitored in vivo with an image splitter and TV microscope. The responses before laser injury were always constriction; the responses after laser injury were always relaxation. After laser injury, acetylcholine chloride (80 micrograms/ml) constricted every vessel tested at the injured site. Thus, the injured segment had not lost the capacity to constrict even though neither serotonin nor arachidonate remained able to induce a constriction. The endothelium-dependent constrictions to serotonin or arachidonate were also blocked by pretreatment of the mouse with cyclooxygenase inhibitors, acetylsalicylic acid (100 mg/kg i.p.) or indomethacin (5 mg/kg). The data suggests that endothelium-dependent constriction to serotonin is mediated by release of arachidonate from the endothelial cell and conversion of that arachidonate by cyclooxygenase to some constricting prostanoid.     

Increased alpha2-adrenergic constriction of isolated arterioles in diffuse scleroderma

OBJECTIVE: Vasospasm and ischemic organ injury are important in the pathogenesis of systemic sclerosis (SSc; scleroderma). The present study was performed to determine whether SSc arterioles have an intrinsic disturbance in vasoconstrictor activity. METHODS: Skin biopsy samples were obtained from the upper arm of 11 patients with diffuse SSc (clinically uninvolved skin) and 8 age- and sex-matched control subjects. Dermal arterioles were dissected from the biopsy sample and mounted in a myograph for continuous monitoring of arteriolar diameter. The resting internal diameter of control and SSc arterioles was similar (mean +/- SEM 164+/-15 micro and 166+/-18micro, respectively). RESULTS: Dermal arterioles displayed no spontaneous constrictor activity in the absence of stimulation. Vasoconstriction in response to KCI, a receptor-independent activator of smooth muscle, or to phenylephrine, a selective alpha1-adrenergic receptor (alpha1-AR) agonist, was similar in control and SSc arterioles. However, constrictor responses to UK 14,304, a selective alpha2-AR agonist, were increased in SSc compared with control arterioles (maximal constriction responses of 25+/-5% and 67+/-4% [mean +/- SEM] in control and SSc arterioles, respectively; P = 0.000014). Mechanical denudation of the endothelium did not alter reactivity to alpha2-AR activation, indicating that the enhanced constriction in SSc was not mediated by changes in endothelial dilator activity. Indeed, in arterioles constricted with phenylephrine, the endothelial stimuli acetylcholine or bradykinin evoked endothelium-dependent relaxation that was similar in control and SSc arterioles. CONCLUSIONS: Vascular smooth muscle in SSc arterioles displayed a selective increase in alpha2-AR reactivity. The endothelial dilator function appeared normal. Altered activity of smooth muscle alpha2-ARs may contribute to the vasospastic activity that is a prominent feature of the SSc disease process.     

Venular constriction of submucosal arterioles induced by dextran sodium sulfate

BACKGROUND: Leukocyte and platelet adherence in postcapillary venules has been speculated to induce constriction of closely paired arterioles. This mechanism was investigated in the current study, in a model of intestinal inflammation induced by dextran sodium sulfate (DSS; 5,000 molecular weight). METHODS: Closely paired, parallel arterioles and venules in the submucosa of the mouse intestine were observed using fluorescent intravital microscopy. Arterioles in control mice were compared to arterioles in mice given 5% DSS in drinking water for 11 days. RESULTS: DSS induced an inflammatory response in which fluorescently labeled leukocytes and platelets were observed adhering to venules. Arterioles constricted and flow decreased significantly when the arterioles were paired with venules having adherent leukocytes and platelets, although the decreases in flow and diameter appeared to be more dependent on platelet than leukocyte adherence. No significant constriction was observed in arterioles paired with venules having minimal platelet adherence. Inhibition of thromboxane with 100 mg/kg ozagrel induced a significant dilation of arterioles in DSS mice that was absent in control mice. CONCLUSION: The results are consistent with a proposed mechanism in which thromboxane constricts submucosal arterioles when the arterioles are closely paired with platelet-bearing venules in DSS-induced inflammation.     

Mediators of CD18/P-selectin-dependent constriction of venule-paired arterioles in hypercholesterolemia

This study addresses the role of venule-derived mediators in the arteriolar constriction that accompanies hypercholesterolemia. Constriction was assessed by measuring the tone of small arterioles closely paired with venules in the mesentery of normal cholesterol rats (NC), high cholesterol rats (HC), HC rats injected with antibodies against CD18 and P-selectin (HC/mAbs), HC rats treated with the thromboxane synthase inhibitor, ozagrel (HC/ozagrel), and HC rats pretreated with anti-platelet serum (HC/APS). Venule-paired arterioles in the untreated HC group demonstrated enhanced tone compared with arterioles in the NC group, while no difference was found between unpaired arterioles of the two groups. Perivascular nitric oxide (NO) concentrations were found to be significantly decreased in venule-paired arterioles of HC rats (238+/-14 nM) compared with those of NC rats (426+/-42 nM). The injection of anti-adhesion antibodies successfully attenuated the enhanced arteriolar tone and venular leukocyte adherence in the HC group, and tended to increase levels of NO in venule-paired arterioles by 33% (to 326+/-19 nM; still lower than that of the NC group). Ozagrel and platelet depletion attenuated the enhanced arteriolar tone by 53% and 33%, respectively, without affecting NO concentrations. These findings indicate that the mechanism of blood cell-dependent arteriolar constriction during hypercholesterolemia may be dependent on thromboxane, a decrease in NO, and the proximity of the arterioles to postcapillary venules.     

Insulin reverses anxiety-like behavior evoked by streptozotocin-induced diabetes in mice

Clinical and preclinical data suggest that diabetes is often associated with anxiety. Insulin, a peptide hormone has been reported to have key functions in the brain and in alleviating several psychological impairments, occur as a consequence of diabetes. However, its effects in diabetes-induced anxiety are scanty. The present study examined whether; insulin can reverse the anxiety-like behavior in streptozotocin (STZ)-induced diabetes in mice. After 8-weeks of diabetes induced by STZ (200 mg/kg, intraperitoneally (i.p.)), mice were given insulin (1–2 IU/kg/day, i.p.)/ diazepam (1 mg/kg/day, i.p.)/ vehicle for 14 days and evaluated for behavioral effects in three validated models of anxiety viz. elevated plus maze (EPM), light–dark (L/D) and hole board (HB) tests. STZ-induced diabetic mice elicited significant behavioral effects which include, decreased percentage open arm entries and time in EPM, reduced latency and time spent in light chamber in L/D, decreased number of head dips, squares crossed and rearings in HB tests respectively. Insulin treatment attenuated the behavioral effects evoked by STZ-induced diabetes in mice as indicated by increased open arms activity in EPM, decreased aversion in light chamber during L/D test and increased exploratory behavior in HB test. In conclusion, this study revealed that insulin can reverse anxiety-like behavior in STZ-induced diabetes in mice.     

链脲佐菌素诱导的糖尿病大鼠视网膜的免疫损伤

分别用免疫组化、免疫荧光方法和计算机图像分析技术研究糖尿病（DM）大鼠视网膜组织中IgA、IgG和IgM的沉积情况。与对照组相比，DM大鼠视网膜组织中IgA、IgG和IgM的沉积增加，有统计学意义（均P〈0．05），说明免疫损伤可能参与了糖尿病视网膜病变的发生发展。     

Inhibition of barium-induced constriction of cerebral surface arterioles by blockers of calcium channels

Barium ion, applied as BaCl2 to the cerebral surface, produces marked constriction of pial arterioles. The present data demonstrate that this constriction is inhibited by the calcium channel blockers verapamil and nimodipine. Verapamil in concentrations as low as 10)-7) M was effective in local administration. Verapamil in milligram per kilogram doses and nimodipine in microgram per kilogram doses were effective 30 min after intraperitoneal injection without producing dilation. The basis for the inhibitory action of the calcium channel blockers is not established by the data. However, in view of barium's known ability to depolarize membranes, it may be that the constriction which results is related to inward passage of ions and is facilitated by inward movements of calcium.     

Relative magnitude of vascular reactivity in the major arterioles of the retina

The relative magnitude of vascular reactivity to inhaled gas stimuli in the major retinal arterioles has not been systematically investigated. The purpose of this study was to compare the magnitude of retinal vascular reactivity in response to inhaled gas provocation at equivalent measurement sites in the superior-, and inferior-, temporal retinal arterioles (STA, ITA). One randomly selected eye of each of 17 healthy volunteers (age 24.4 ± 4.7) was prospectively enrolled. Volunteers were connected to a sequential gas delivery circuit and a computer-controlled gas blender (RespirAct?, Thornhill Research Inc., Canada) and underwent an isocapnic hyperoxic challenge i.e. P_ETO₂ of 500 mm Hg with P_ETCO₂ maintained at 38 mm Hg during baseline and hyperoxia. Four retinal hemodynamic measurements were acquired using bi-directional laser Doppler velocimetry and simultaneous vessel densitometry (Canon Laser Blood Flowmeter, CLBF-100, Japan) at equivalent positions on the STA and ITA. Statistical analysis was performed using linear mixed-effect models. During the hyperoxic phase, the vessel diameter (STA p = 0.004; ITA p = 0.003), blood velocity (STA p < 0.001; ITA p < 0.001) and flow (STA p < 0.001; ITA p < 0.001) decreased in both the STA and the ITA relative to baseline. The diameter, velocity and flow were equivalent between STA and ITA at baseline and during hyperoxia; and their magnitude of change remained comparable with hyperoxia (p > 0.05). The magnitude of retinal arteriolar vascular reactivity in response to isocapnic hyperoxic inhaled gas challenge was not significantly different between the STA and ITA. However, the correlation analysis did not reveal a significant relationship between the percentage changes in diameter, velocity and flow of the STA and ITA and did not demonstrate equal responses from the STA and ITA to gas provocation.     

Inhibition of ceramide synthesis reverses endothelial dysfunction and atherosclerosis in streptozotocin-induced diabetic rats

Objectives

To explore the effect of myriocin on EDVD and atherosclerosis in diabetic rats.

Methods

Rats were fed with a high-fat/high-sucrose/high-cholesterol diet (20% sucrose, 10% animal oil, 1.0% bile salt and 2.5% cholesterol) (hereinafter defined as diabetic groups) or Purina Rodent Chow (NC group), the former was intervened with low dose streptozotocin (30 mg/kg) after feeding 1 month to make diabetic model. The NC group was intervened with citrate buffer and the diabetic rats were intervened with myriocin (0.3 mg/kg Qod) (MTD group) or just solvent (DC group) for 14 weeks. The EDVD, thickness of fatty deposition under endothelium, ceramide, PI3K/PKB/eNOS, NO and other vital parameters were measured after the rats sacrificed.

Results

In DC group, the ceramide contents in serum and aorta increased, the EDVD was impaired, the fatty deposition under endothelium increased, and the phosphorylation of PI3K/PKB/eNOS and NO release decreased all compared with the NC group (P < 0.05). Compared with the DC group, the ceramide contents in MTD group decreased, the EDVD ameliorated, the fatty deposition diminished, and PI3K/PKB/eNOS phosphorylation and NO release (P < 0.05) increased.

Conclusions

After treated with myriocin, the EDVD in diabetic rats has been improved by increasing PI3K/PKB/eNOS phosphorylation and NO release, and meanwhile the atherosclerosis has reversed.     

Pressure-dependent contraction of rat juxtamedullary afferent arterioles

Pressure-diameter relations were studied in rat afferent arterioles using an isolated, juxtamedullary nephron preparation perfused with a saline solution containing 5% albumin. Angiotensin I (10 microM), angiotensin II (0.1 microM), and norepinephrine (10 microM) increased perfusion pressure, and norepinephrine, but not angiotensin I or II, contracted afferent arterioles, indicating that the vessels are reactive. The control diameter of the afferent arterioles that exhibited pressure-dependent contraction (n = 58) averaged 30.8 +/- 1.1 micron at perfusion pressure of 80 mm Hg. When pressure was increased from 80 to 120 and then to 180 mm Hg, the diameter of these arterioles decreased by 16.4 +/- 2.1%. Glomerular capillary pressure was well autoregulated and averaged 45.2 +/- 2.2, 50.2 +/- 2.4, and 53.0 +/- 3.0 mm Hg, respectively, at perfusion pressures of 80, 120, and 180 mm Hg. Administration of vasodilators or a Ca2+-free solution eliminated the contractile response to pressure elevations; rather, the diameter of these vessels increased significantly by 17.5 +/- 5.1% and 32.0 +/- 9.4%, respectively, when pressure was increased from 80 to 180 mm Hg. Blocking tubuloglomerular feedback mechanism, with furosemide or by removal of the renal papilla (which interrupts the delivery of fluid to the macula densa), eliminated the pressure-dependent contraction of the afferent arterioles. Instead the diameter of these vessels increased by 27.0 +/- 7.8% and 36.0 +/- 5.6%, respectively, when the pressure was increased from 80 to 120 and then to 180 mm Hg. These results demonstrate that juxtamedullary nephrons perfused in vitro autoregulate glomerular capillary pressure.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lack of flow-mediated dilation and enhanced angiotensin II-induced constriction in skeletal muscle arterioles of lupus-prone autoimmune mice

Systemic lupus erythematosus (SLE) is associated with disturbances in the microcirculation of various tissues, yet the nature of arteriolar dysfunction has not been characterized. Thus, changes in diameter of isolated, pressurized skeletal muscle arterioles of mice with systemic autoimmune disease (lupus prone, MRL/lpr four-month old female) and control (MRL) mice were investigated by video-microscopy. Arteriolar responses to changes in intraluminal pressure, flow, and to vasoactive agents with known mechanisms of action were compared. The active and passive (in Ca2+ free solution) diameter of MRL/lpr arterioles were not significantly different compared to MRL and morphometric changes were not apparent. Compared to MRL mice the endothelium-dependent dilations to increase in flow, acetylcholine and bradykinin were markedly reduced in arterioles of MRL/lpr mice. Endothelium-independent dilations to sodium-nitroprusside and adenosine were similar in MRL and MRL/lpr arterioles. Furthermore, angiotensin II elicited greater constrictions in MRL/lpr arterioles, whereas serotonin-induced constrictions were similar in both groups. Thus, in arterioles of MRL/lpr mice endothelium-dependent dilator mechanisms are impaired and constriction to angiotensin II is enhanced, suggesting specific alterations in the vasomotor function of microvessels that are likely contribute to the disturbance of skeletal muscle blood flow observed in systemic lupus erythematosus.     

Ileal brake failure in streptozotocin-induced diabetic rat

BACKGROUND: Diabetes mellitus frequently alters gastrointestinal function, but the pathophysiology of the diabetic gut has not been fully elucidated. Our aim was to characterize the enterogastric modulation of gastric emptying in an experimental model of diabetic rat and to determine the putative consequences of impaired regulation on glycaemic control. METHODS: Studies were performed in streptozotozin-induced diabetic and control groups of male Sprague-Dawley rats. In rats fitted with chronic ileal cannulae, gastric emptying of a peptide meal was measured during ileal infusion of either lipids (ileal brake) or saline. The influence of the ileal brake mechanism on blood glucose levels after oral administration of a glucose solution was also evaluated. RESULTS: Diabetic rats exhibited a precipitous gastric emptying (80% +/- 3% versus 57% +/- 3% in controls; P < 0.05). Ileal lipids delayed gastric emptying in control (38 +/- 4%; P < 0.05 versus ileal saline) but not in diabetic animals (77 +/- 5%; N.S. versus ileal saline). As the ileal brake contributes to the management of postprandial blood glucose levels (114 +/- 4.9 mg/dL after ileal lipids versus 134 +/- 7.8 mg/dL after ileal saline in control rats; P < 0.05), the failure of this mechanism in diabetic rats worsens glycaemic control after feeding (455 +/- 20.4 mg/dL after ileal lipids versus 399 +/- 8.7 mg/dL after ileal saline; P < 0.05). CONCLUSION: Experimental diabetes impairs the ileal brake mechanism and disturbs gastric emptying. These abnormalities may contribute to difficult glycaemic control.     

环孢素A对链脲菌素诱导的糖尿病大鼠视网膜免疫球蛋白异常沉积的影响

目的 探讨糖尿病视网膜病变(DR)发病过程中的免疫机制,观察环孢素A(CsA)对糖尿病大鼠视网膜病变的防治作用.方法 以链脲菌素诱导的糖尿病大鼠为模型,分别于造模前或成模后给予低、中、高剂量CsA皮下注射,并设立正常、单纯糖尿病和胰岛素治疗组为对照,通过HE染色观察糖尿病大鼠视网膜病变,通过免疫组化和免疫荧光检测视网膜免疫球蛋白的沉积.结果 8周时,糖尿病大鼠视网膜已经出现DR的病理学表现,且有大量IgG、IgA、IgM沉积.胰岛素治疗组免疫球蛋白沉积情况与单纯糖尿病组差异无统计学意义.CsA干预各组的视网膜免疫球蛋白沉积情况与单纯糖尿病组相比差异有统计学意义(P＜0.05),尤以高剂量组为著(P＜0.01).结论 免疫机制参与DR的发生,免疫抑制治疗能有效地减少视网膜免疫球蛋白沉积,延缓DR的发生.     

Low-dose streptozotocin-induced diabetes in the spontaneously hypertensive rat

Streptozotocin (STZ, 35 mg/kg body weight) was injected into spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats, and plasma glucose and triglyceride concentrations measured 10 days later. Neither mean (+/- SEM) plasma glucose (130 +/- 3 v 136 +/- 3 mg/dL) nor triglyceride (93 +/- 6 v 108 +/- 7 mg/dL) concentrations increased in WKY rats. In contrast, both plasma glucose (141 +/- 3 v 262 +/- 36) and triglyceride (121 +/- 8 v 196 +/- 7 mg/dL) concentrations increased significantly (P less than .01) following administration of STZ in SHR. Furthermore, when SHR previously injected with STZ were fed a diet enriched in fructose, they had a further increase (P less than .01) in both plasma glucose (343 +/- 38 mg/dL) and triglyceride (774 +/- 57 mg/dL) concentrations. Plasma triglyceride concentration also increased significantly (P less than .05) when STZ-injected WKY rats ingested the fructose-enriched diet, but plasma glucose levels still remained within the normal range (152 +/- 5 mg/dL). These results indicate that SHR were more sensitive to the effects of a decrease in pancreatic beta-cell function (STZ) and an increase in insulin resistance (fructose feeding) than WKY rats.     

Influence of streptozotocin-induced diabetes on the concentration of immunoreactive somatostatin in the retina and peripheral blood of the rat: effect of insulin treatment

Changes in somatostatin-like immunoreactivity (SLI) were examined in the retina and peripheral blood of diabetic rats treated with streptozotocin (STZ) and insulin. There was no change in retinal SLI content at 4 and 11 days after administration of STZ but, thereafter, SLI increased progressively in the diabetic animals by 220% at 18 days and 300% at 27 days. Plasma SLI levels increased by 500% at 11 days and maintained similar levels thereafter. Diabetic animals treated with insulin (3-5 i.u. daily) for 27 days showed a significant (P less than 0.01) decrease of retinal and plasma SLI levels compared with untreated diabetic animals. It is concluded that there is a significant increase of retinal and plasma SLI levels in diabetic rats which tends to normalize after several days of insulin treatment.