Effects of melatonin on spinal cord injury-induced oxidative damage in mice testis.

This study evaluated the effects of melatonin on spinal cord injury (SCI)-induced oxidative damage in testes. Adult male C57BL/6 mice were randomly divided into sham-, SCI- or melatonin (10 mg/kg, i.p.)-treated SCI groups. To induce SCI, a standard weight-drop method that induced a contusion injury at T10 was used. After 1 week, testicular blood flow velocity was measured using the Laser Doppler Line Scanner. Malondialdehyde (MDA), glutathione (GSH), oxidised glutathione (GSSG) and myeloperoxidase (MPO) were measured in testis homogenates. Microvascular permeability of the testes to Evan's Blue was examined by spectrophotometric and fluorescence microscopic quantitation. The tight junction protein zonula occludens-1 (ZO-1) and occludin in testes were assessed by immunoblot analysis. Melatonin increased the reduced blood flow and decreased SCI-induced permeability of capillaries. MDA levels and MPO activity were elevated in the SCI group compared with shams, which was reversed by melatonin. In contrast, SCI-induced reductions in GSH/GSSG ratio were restored by melatonin. Decreased expression of ZO-1 and occludin was observed, which was attenuated by melatonin. Overall, melatonin treatment protects the testes against oxidative stress damage caused by SCI.

The antioxidant butylated hydroxytoluene prevents early cholesterol-induced microcirculatory changes in rabbits.

Microcirculation was studied during 10 wk in untreated rabbits (n = 13) and in rabbits treated with dietary addition of 1% cholesterol (n = 13), 1% cholesterol + 1% of the antioxidant BHT (butylated hydroxytoluene) (n = 11), or 1% BHT (n = 5). The studies were performed by direct intravital microscopic imaging of the left and right conjunctivae with the use of a stereo microscope and a high resolution television camera. Microvessel diameter, erythrocyte flow velocity, and microhemorheologic conditions were evaluated quantitatively via a computer-assisted digital image processing system. Significant and marked changes occurred in all the above variables as a consequence of cholesterol feeding. After 3 wk of feeding there was a dramatic decrease (approximately 30%) in blood flow velocity in arterioli of the third order (P < 0.0001), accompanied by aggregation of cells in 40-50% of the smaller conjunctival vessels (P < 0.0001). These changes were enhanced further during the following 7 wk of treatment. All the above changes in the microcirculation were markedly reduced by the addition of BHT treatment. The diameter of the above arterioli decreased in the purely cholesterol-fed group (P < 0.005), whereas this did not occur in the group fed both cholesterol and BHT. In rabbits fed BHT in the absence of cholesterol, there was no significant effect on any assessed microcirculatory variable. In conclusion, the results demonstrate that the antioxidant BHT prevented early cholesterol-induced microcirculatory changes. This prevention occurred in the absence of a reduction of blood lipid levels. The results provide strong support for the hypothesis that a considerable part of the effects on microcirculation in hypercholesterolemia may be due to cholesterol-induced oxidations and not to cholesterol itself. The results are discussed in relation to the previously demonstrated antiatherogenic effect of BHT and the possible use of antioxidants in the therapy and prophylaxis of atherosclerosis.

N-acetylcysteine improves microcirculatory flow during smoking: new effects of an old drug with possible benefits for smokers.

BACKGROUND: Cigarette smoking provokes marked acute changes in the microcirculatory vasculature, including a reduced blood flow velocity. In accordance with the hypothesis that the reduced blood flow is due to an imbalance between pro-oxidants and oxidants, we recently showed that most of the reduction could be reversed by a high dose of vitamin C. HYPOTHESIS: In the present work we tested the hypothesis that N-acetylcysteine, a mucolyticum and an antioxidant, may have an effect on the smoking-induced changes observed by vital capillary microscopy of the nailfold. METHODS: In all, 37 healthy volunteers of both genders and with varied smoking habits were treated with N-acetylcysteine 200 mg t.i.d. for 2 weeks. In vivo investigation of the microcirculation by capillaroscopy was performed before and after treatment. RESULTS: Treatment with N-acetylcysteine significantly reduced the smoking-induced relative decrease in capillary blood flow velocity in a group of volunteers with varied smoking habits (p = 0.0016). The preventive effect was clearly significant in smokers (p = 0.003). CONCLUSION: Treatment with N-acetylcysteine has a positive impact on microcirculatory flow during smoking, particularly in habitual smokers.

[Angiogenesis induced by SP2/0 and HeLa tumor cells].

Cheek pouches of Syrian golden hamsters and transparent access chambers of BALB/C mice were implanted with SP2/0 and HeLa tumor cells separately. Observation was done by continual photo and video from D 1 through the D 10 after implantation. It was found by the development of angiogenesis and the density of microvessels that both kinds of tumor cells could induce angiogenesis. On D 2 after implantation, there appeared leakage and hemorrhage from the microvessels near the tumor cells. On D 3 and D 4, there was an increased density of new capillaries which formed a very fine, tortuous and basketlike vascular plexus of irregular diameter. Most of the new microvessels came from the venules on the edge of the implant mass and they grew toward the tumor cells to penetrate the tumor tissue on D 4 to D 5. On D 7, the density of new microvessels reached the maximum and they began to extend outside the tumor which was surrounded by dense new capillaries. Compound 36 has been proved an effective substance against some tumors in clinical applications in this country. Also proved by our experiments, a drug effective in inhibition of angiogenesis induced by SP2/0 tumor cells.

[An animal experiment and clinical investigation on the protective effect of selenium on the microcirculation induced by free radical damaged RBCs].

UNLABELLED: Fluorescent labeling image analysis was used to evaluate the changes in cerebral arteriole and veinlet diameters (D), circulation velocities (FV) and permeability (VP) in rats; while in clinics, a laser-doppler device was used for assessing changes of skins and muscles microcirculation. The results show that in control rats, equal volume perfusion of free radical damaged RBCs resulted in decreases of D and FV significantly but VP was increased, whereas in the case when free radical damaged RBCs were perfused together with selenium, no disturbances in the D and VP were observed with FV even improved. In the human control group, either average skin microcirculative perfusions (ASMP) at 25 degrees C or maximal skin microcirculative perfusions (MSMP) at 42 degrees C were evidently decreased during myocardial ischemia/reperfusion period, while ASMP at 24th hour of post-surgery was kept reducing. At the same time, the changes in muscles microcirculative perfusions (MMP) tended to be similar to the skin, but overloaded than the pre-surgery levels at 24th hour of post-surgery period. In the selenium group before surgery, the ratio of MSMP to ASMP was obviously increased than the control group (3.95 in Se group vs 1.74 in control group, P < 0.05), but did not have significantly differences in ASMP, MSMP and MMP between the two groups during surgery period. RBC deformabilities were not changed. At 24th hour post-surgery, the ASMP were almost restored to pre-surgery levels. However, MMP were still lower than the pre-surgery levels. CONCLUSIONS: (1) free radical damaged RBC perfusion leads to damage of microcirculation; (2) selenium is highly efficient in protecting microcirculation from free radical damaged RBC disturbance; and (3) Oral administration of selenium may improve pre-surgery maximal skin microcirculative perfusion and promote recovery of the worsened skin microcirculation in addition to prevent the occurrence of RBC deformability.

Enhanced matrix metalloproteinases-2 activates aortic endothelial hypermeability, apoptosis and vascular rarefaction in spontaneously hypertensive rat.

Microvascular rarefaction with endothelial cells apoptosis is a common characteristic of various microvascular complications in the spontaneously hypertensive rat (SHR). Elevated levels of proteolytic (e.g. matrix metalloproteinase, MMPs) activity and apoptosis in aortic endothelial cells of SHR were found when compared to its normotensive control. However, the exact mechanisms of microvascular rarefaction and the role of MMPs in this process remain poorly understood. Besides cleavage of VEGFR2 via unbalanced MMPs, we hypothesize that selected cleavage of Beta-Catenin and VE-cadherin by MMPs could induce apoptosis of rat aortic endothelial cells (RAECs) and rarefaction. Primary RAECs were isolated, identified and used in a in-vitro model. Transwell system was used to analyze the permeability of Wistar RAECs, SHR RAECs and SHR RAECs with pretreatment by doxycycline. Qualitative and semi-quantitative analysis of major endothelial adhesion molecules were detected by immunofluorescence technique and Western blot, respectively. MMP-2 activity of SHR RAECs was increased significantly and doxycycline (50 µM) effectively reduced the level of MMP-2 and hyper-permeability in SHR RAECs. SHR RAECs showed enhanced cleavage of VEGFR2, VE-cadherin and B-catenin, which could be prevented by doxycycline (50 µM). Doxycycline (50 µM) attenuated hyper-permeability via decreased MMP-2 by protecting VEGFR2, VE-cadherin, Beta-catenin from cleavage and inhibited the reduction of mitochondrial transmembrane potential (MTP), thus prevented mitochondria-mediated apoptotic signaling and capillary rarefaction in the SHR. It might be a novel insight into the mechanisms of SHR microvascular rarefaction that is independent of pressure but relevant to MMP-2.

Microcirculatory disturbances in AIDS patients--a first report.

The aim of this study is to explore the vascular perturbations associated with AIDS infections as a means of developing strategies to help in the treatment of these patients. Closed-circuit television microscopic observation of the microcirculation in the nail fold was carried out on 11 AIDS patients and 11 healthy European adults. A striking pattern of severe microvascular disturbances was demonstrated in these patients: (i) a suppression of the spontaneous, rhythmic adjustments of capillary blood perfusion, and (ii) damage to the endothelium of the capillary wall. These findings suggest a substantial involvement of the microcirculation in the pathogenetic development of the AIDS syndrome, as well as a close relationship between microvascular perfusion and the immune condition of the organism. The data indicate that the treatment of the AIDS patient should be directed not only toward the inhibition or killing of the virus, but also toward the improvement of microcirculatory perfusion.

Inhibition of histamine-induced protein leakage in rat skeletal muscle by blockade of prostaglandin synthesis.

The effects of two prostaglandin synthesis blockers, indomethacin and mefenemic acid, on both histamine-induced extravasation of albumin and arteriole dilation were studied using intravital fluorescent microscopy. Sprague-Dawley rats (140-180 g) were anesthetized with pentobarbital (50mg/Kg) and the cremaster muscle was positioned in a Krebs bath (pH = 7.4, PCO2 = 40 mm Hg, PO2 = 35 mm Hg, temp = 34 degrees). Fluorescein isothiocyanate-labeled albumin was injected intravascularly and the fluorescent image of the cremaster microcirculation was produced by illumination from an argon laser (488 nm). Videotape analysis of the experiments showed that increasing concentrations of histamine in the bath produced both a concentration-dependent arteriole dilation and a concentration-dependent leakage of labeled albumin into the interstitium. Adding indomethacin (10 or 100 micrograms/ml) or mefenemic acid (10 micrograms/ml) to the Krebs bath significantly diminished the protein leakage induced by histamine but did not alter the arteriole dilation. These results indicate that prostaglandins may be directly involved in the histamine-induced increase in vascular permeability to macromolecules but not in the arteriole dilation histamine produces.

Dietary cholesterol induces transient changes in plasma nitrate levels in rabbits that are correlated to microcirculatory changes.

Dietary treatment of rabbits with 1% cholesterol resulted in a transient rise in their plasma nitrate levels. After 3 weeks of treatment the nitrate levels were about 50% higher than those of the controls (p<0.005). After 10 weeks of treatment the nitrate levels were similar to those at the start of the study. In accordance with previous work (Xiu et al., J. Clin. Invest., 1994, 93, 2732-2737), the cholesterol treatment let to a decreased blood flow velocity in arterioli of the third order in the conjunctiva, and a decreased diameter of these arterioli. There was a significant correlation between plasma nitrate levels and the two microcirculatory variables (p<0.0001). Nitrate is the major metabolic end product of nitric oxide (NO), and plasma nitrate levels may be used as an index of the endogenous formation of NO. The present results suggest that dietary cholesterol induces a transient increase in the synthesis of NO. Such an increased synthesis may compensate for part of a cholesterol-induced degradation of NO.

Anisodamine inhibits thromboxane synthesis, granulocyte aggregation, and platelet aggregation. A possible mechanism for its efficacy in bacteremic shock.

Anisodamine hydrochloride is a vasoactive drug produced in the People's Republic of China that appears efficacious in clinical and experimental bacteremic shock, and about whose mode of action little is known. Suspecting that the drug might work by inhibition of platelet or granulocyte aggregation, or both, we tested it in these systems. Anisodamine proved a modest inhibitor of granulocyte aggregation and a powerful inhibitor of platelet aggregation; thromboxane synthesis was inhibited in anisodamine-treated platelets, further suggesting that the biochemical mode of action might be inhibition of cyclo-oxygenase or thromboxane synthetase.