Lipid peroxidation in the synaptosomal and mitochondrial fractions of separate brain structures in hypoxia

The paper studies intensification of lipid peroxide oxidation in separate brain structures (the medulla oblongata, cerebellum, visual and sensomotor cortex), synaptosomal and mitochondrial fractions under hypoxia. It has been established that acute hypoxia increases accumulation of lipid peroxidation (LPO) products, hydroperoxide and malonyl dialdehyde. Intensification of LPO in synaptosomes and mitochondria is more pronounced as compared to the whole structures. Preliminary treatment with antioxidants (vitamin E and ionol) considerably suppressed LPO intensity under both hypoxia and hypoxia with reoxygenation. Intensification of LPO in synaptosomes and mitochondria is suggested to be the key point in structural-functional disturbances of the nervous system under hypoxia and ischemia.     

The Na, K-ATPase activity of the separate structural components of the guinea-pig visual system in hypoxia

Different degree of sensitivity to acute hypoxia in vivo has been shown in guinea-pig pigmented epithelium, retina and visual cortex Na, K-ATPase. The highest degree of the enzyme activity inhibition has been noted in the pigmented epithelium (more than 3 times), lower inhibition-in visual cortex (26%) and the lowest-in the retina (18%). In contrast to Na, K-ATPase, hypoxia effect on Mg-ATPase resulted in both inhibition and activation of the enzyme in all 3 structures. Reoxygenation following acute hypoxia increases Na, K-ATPase activity inhibition in the retina and visual cortex. But under reoxygenation the enzyme activity is recovered in the pigmented epithelium. Preliminary administrations of vitamin E completely prevented Na, K-ATPase activity inhibition in all studied structures.     

Activation and damage of endothelial cells upon hypoxia/reoxygenation. Effect of extracellular pH

Disturbances of blood flow upon vascular occlusions and spasms result in hypoxia and acidosis, while its subsequent restoration leads to reoxygenation and pH normalization (re-alkalization) in ischemic sites of the vascular bed. The effect of hypoxia/reoxygenation on activation and stimulation of apoptosis in cultured human endothelial cells was studied. The cells were subjected to hypoxia (2% O₂, 5% CO₂, 93% N₂) for 24 h followed by reoxygenation (21% O₂, 5% CO₂, 74% N₂) for 5 h. Reoxygenation was carried out at different pH-6.4 (preservation of acidosis after hypoxia), 7.0, and 7.4 (partial and complete re-alkalization, respectively). Hypoxia only slightly (by ～30%) increased the cell adhesion molecule ICAM-1 content on the cell surface, whereas reoxygenation more than doubled its expression. The reoxygenation effect depended on the medium acidity, and ICAM-1 increase was more pronounced at pH 7.0 compared to that at pH 6.4 and 7.4. Neither hypoxia nor reoxygenation induced expression of two other cell adhesion molecules, VCAM and E-selectin. Incubation of cells under hypoxic conditions but not reoxygenation stimulated secretion of von Willebrand factor and increased its concentration in the culture medium by more than 4 times. The percentage of cells containing apoptosis marker, activated caspase-3, was increased by approximately 1.5 times upon hypoxia as well as hypoxia/reoxygenation. Maximal values were achieved when reoxygenation was performed at pH 7.0. These data show that hypoxia/reoxygenation stimulate pro-inflammatory activation (ICAM-1 expression) and apoptosis (caspase-3 activation) of endothelial cells, and the extracellular pH influences both processes.     

Effects of adaptation to exposure to short-term stress on indices of resistance of energy metabolism and contractile function of the myocardium to acute hypoxic hypoxia and reoxygenation

Effect of preliminary adaptation to immobilization stress with progressive duration from 15 min. to 1 h (every second day, 8 sessions) on the resistance of indices of myocardial energy metabolism and contractile function to acute hypoxic hypoxia and subsequent reoxygenation was studied. It was shown, that adaptation to short-term stress exposure by some way provided the retention of activities of important enzymes like creatine-phosphokinase and phosphorylase under the harmful action of acute hypoxia and subsequent reoxygenation. At the same time, the ATP restoration and the CP super-restoration were observed during reoxygenation. This effect, in its turn, was accompanied by a more pronounced super-restoration of the heart contractile function than in control.     

Number and Distribution of Mouse Retinal Cone Photoreceptors: Differences between an Albino (Swiss) and a Pigmented (C57/BL6) Strain

We purpose here to analyze and compare the population and topography of cone photoreceptors in two mouse strains using automated routines, and to design a method of retinal sampling for their accurate manual quantification. In whole-mounted retinas from pigmented C57/BL6 and albino Swiss mice, the longwave-sensitive (L) and the shortwave-sensitive (S) opsins were immunodetected to analyze the population of each cone type. In another group of retinas both opsins were detected with the same fluorophore to quantify all cones. In a third set of retinas, L-opsin and Brn3a were immunodetected to determine whether L-opsin⁺cones and retinal ganglion cells (RGCs) have a parallel distribution. Cones and RGCs were automatically quantified and their topography illustrated with isodensity maps. Our results show that pigmented mice have a significantly higher number of total cones (all-cones) and of L-opsin⁺cones than albinos which, in turn, have a higher population of S-opsin⁺cones. In pigmented animals 40% of cones are dual (cones that express both opsins), 34% genuine-L (cones that only express the L-opsin), and 26% genuine-S (cones that only express the S-opsin). In albinos, 23% of cones are genuine-S and the proportion of dual cones increases to 76% at the expense of genuine-L cones. In both strains, L-opsin⁺cones are denser in the central than peripheral retina, and all-cones density increases dorso-ventrally. In pigmented animals S-opsin⁺cones are scarce in the dorsal retina and very numerous in the ventral retina, being densest in its nasal aspect. In albinos, S-opsin⁺cones are abundant in the dorsal retina, although their highest densities are also ventral. Based on the densities of each cone population, we propose a sampling method to manually quantify and infer their total population. In conclusion, these data provide the basis to study cone degeneration and its prevention in pathologic conditions.     

L-Dopa and the Albino Riddle: Content of L-Dopa in the Developing Retina of Pigmented and Albino Mice

Background

The absence or deficiency of melanin as in albinos, has detrimental effects on retinal development that include aberrant axonal projections from eye to brain and impaired vision. In pigmented retinal pigment epithelium (RPE), dihydroxyphenalanine (L-Dopa), an intermediate in the synthetic path for melanin, has been hypothesized to regulate the tempo of neurogenesis. The time course of expression of retinal L-Dopa, whether it is harbored exclusively in the RPE, the extent of deficiency in albinos compared to isogenic controls, and whether L-Dopa can be restored if exogenously delivered to the albino have been unknown.

Methodology/Principal Findings

L-Dopa and catecholamines including dopamine extracted from retinas of pigmented (C57BL/6J) and congenic albino (C57BL/6J-tyr^c2j) mice, were measured throughout development beginning at E10.5 and at maturity. L-Dopa, but not dopamine nor any other catecholamine, appears in pigmented retina as soon as tyrosinase is expressed in RPE at E10.5. In pigmented retina, L-Dopa content increases throughout pre- and postnatal development until the end of the first postnatal month after which it declines sharply. This time course reflects the onset and completion of retinal development. L-Dopa is absent from embryonic albino retina and is greatly reduced in postnatal albino retina compared to pigmented retina. Dopamine is undetectable in both albino and pigmented retinas until after the postnatal expression of the neuronal enzyme tyrosine hydroxylase. If provided to pregnant albino mothers, L-Dopa accumulates in the RPE of the fetuses.

Conclusions

L-Dopa in pigmented RPE is most abundant during development after which content declines. This L-Dopa is not converted to dopamine. L-Dopa is absent or at low levels in albino retina and can be restored to the RPE by administration in utero. These findings further implicate L-Dopa as a factor in the RPE that could influence development, and demonstrate that administration of L-Dopa could be a means to rescue developmental abnormalities characteristic of albinos.     

Development and control of guinea-pig liver estrone sulfate 16 alpha-hydroxylase

Estrone sulfate 16 alpha-hydroxylase activity is undetectable in liver microsomes from fetal guinea-pigs of the English Shorthair variety. Within 2 days of birth, considerable activity is present in both sexes of pigmented and albino animals. In the pigmented group, maximum activity occurs during the second week of life, with the mean values for each of the first 4 weeks, in both sexes, significantly higher than for the corresponding mature (greater than 12-week-old) animals. Immature levels in the albino group are also significantly higher than those of mature albinos. Pigmented females of all ages possess significantly higher activities than do their male counterparts. There are no such sex-related differences in albinos. Pigmented animals of all ages exhibit higher activities than do their albino counterparts. Castration of either sex, pigmented or albino, results in increased enzyme activities as compared with intact or sham-operated controls. Gestation leads to maternal enzyme values which are significantly above those of non-pregnant females, whether pigmented or albino. Beyond the first few days of life, total liver microsomal cytochrome P450 shows no significant change with age, gestation or pigmentation. These data support the conclusion that estrone sulfate 16 alpha-hydroxylase activity in the guinea-pig is markedly diminished, following sexual maturity, by presently unknown factors. This holds for both pigmented and albino animals but the decrease is greater in the latter. This decrease can be reversed by castration in either sex, or by pregnancy and could possibly relate to gonadal-pituitary relationships as demonstrated by others for rat liver hydroxylases.     

Effect of the antioxidant ionol on energy metabolic indices and heart function during acute hypoxia and subsequent reoxygenation

The effect of preliminary administration of antioxidant ionol on the heart energy metabolism and contractile function was estimated in hypoxic hypoxia and subsequent reoxygenation. The protective effect of ionol on the energy metabolism in hypoxia was shown to occur mainly at the level of glycolysis. In reoxygenation, the protective effect of ionol manifested at the level of creatine kinase system to provide a rapid restoration of the CP synthesis rate. This shift correlated with the velocity of restoration of the developed pressure and the velocities of contraction and relaxation. On the whole the data obtained correspond to the notion that creatine kinase system and ATP play an important role in the depression and subsequent restoration on the heart contractile function in acute hypoxia and reoxygenation and ionol provides more effective performance of this system and correspondingly more rapid restoration of the contractile function in reoxygenation.     

Differential effect of dopamine on mitosis in early postnatal albino and pigmented rat retinae

Insufficient levels of L-DOPA, released from the retinal pigment epithelium (RPE), in albino animals are considered responsible for the abnormal development of the underlying neural retina. L-DOPA normalizes retinal neurogenesis by reducing levels of cell proliferation either by acting on the cells directly or by being converted into dopamine. Here we report the effects of dopamine on mitosis in early postnatal neural retinae from albino and pigmented rats, using 4D (x, y, z and time) confocal microscopy. Exogenous dopamine significantly prolongs mitosis in retinae from albino, but not pigmented, animals. As fewer cells move into and divide in the ventricular zone (VZ) in the presence of dopamine, we conclude that the overall cell cycle is affected. The D1 receptor blocker, SCH 23390, inhibits these effects. Thus, the differential effects of dopamine on neural retinae from pigmented and albino rats in vitro must result from the activation of D1 receptors, which are present in the retina from birth. Immunohistochemical labeling of D1 receptors shows that the pattern of their distribution is similar between pigmentation phenotypes, but levels of expression may be elevated in albinos. Labeling is most intense in the inner plexiform layer but is present throughout the neuroblastic layer. These findings are discussed in light of previous reports of reduced catecholamine levels in the albino retina.     

Exposure to acute hypoxia induces a transient DNA damage response which includes Chk1 and TLK1

Severe hypoxia has been demonstrated to induce a replication arrest which is associated with decreased levels of nucleotides. Chk1 is rapidly phosphorylated in response to severe hypoxia and in turn deactivates TLK1 through phosphorylation. Loss of Chk1 has been shown to sensitize cells to hypoxia/reoxygenation. After short (acute) exposure to hypoxia this is due to an increased rate of reoxygenation-induced replication restart and subsequent p53-dependent apoptosis. After longer (chronic) exposure to hypoxia S phase cells do not undergo reoxygenation-induced replication restart. Cells exposed to these levels of hypoxia are however sensitive to loss of Chk1. This suggests a new role for Chk1 in the cell cycle response to reoxygenation.Key words: hypoxia, reoxygenation, replication restart, Chk1, TLK1     

Exposure to acute hypoxia induces a transient DNA damage response which includes Chk1 and TLK1

Severe hypoxia has been demonstrated to induce a replication arrest which is associated with decreased levels of nucleotides. Chk1 is rapidly phosphorylated in response to severe hypoxia and in turn deactivates TLK1 through phosphorylation. Loss of Chk1 has been shown to sensitize cells to hypoxia/reoxygenation. After short (acute) exposure to hypoxia this is due to an increased rate of reoxygenation-induced replication restart and subsequent p53-dependent apoptosis. After longer (chronic) exposure to hypoxia S phase cells do not undergo reoxygenation-induced replication restart. Cells exposed to these levels of hypoxia however are sensitive to loss of Chk1. This suggests a new role for Chk1 in the cell cycle response to reoxygenation.     

Short-term hypoxia/reoxygenation activates the angiogenic pathway in rat caudate putamen

In response to hypoxia, tissues have to implement numerous mechanisms to enhance oxygen delivery, including the activation of angiogenesis. This work investigates the angiogenic response of the hypoxic caudate putamen after several recovery times. Adult Wistar rats were submitted to acute hypoxia and analysed after 0 h, 24 h and 5 days of reoxygenation. Expression of hypoxia-inducible factor-1 alfa (HIF-1α) and angiogenesis-related genes including vascular endothelial growth factor (VEGF), adrenomedullin (ADM) and transforming growth factor-beta 1 (TGF-β1) was determined by both RT-PCR and ELISA. For vessel labelling, lectin location and expression were analysed using histochemical and image processing techniques (fractal dimension). Expression of Hif-1α, Vegf, Adm and Tgf-β1 mRNA rose immediately after hypoxia and this increase persisted in some cases after 5 days post-hypoxia. While VEGF and TGF-β1 protein levels increased parallel to mRNA expression, ADM remained unaltered. The quantification of the striatal vessel network showed a significant augmentation at 24 h of reoxygenation. These results reveal that not only short-term hypoxia, but also the subsequent reoxygenation period, up-regulate the angiogenic pathway in the rat caudate putamen as a neuroprotective mechanism to hypoxia that seeks to maintain a proper blood supply to the hypoxic tissue, thereby minimizing the adverse effects of oxygen deprivation.     

丹参对心肌低氧/复氧损伤的保护作用的研究

目的：研究中药丹参（ＳＭ）对心肌低氧／复氧损伤的保护作用。方法：运用＾３１Ｐ－ＮＭＲ技术对离体灌流大鼠心脏的高能磷酸化合物含量及细胞内的ｐＨ值（ｐＨｉ）进行动态跟踪。结果：丹参注射液能明显减轻低氧期间心肌高能磷酸合物含量的下降,促使复氧期间ＰＣｒ、ＡＴＰ相对含量的恢复,减少低氧及复氧阶段心肌ｐＨｉ的下降。结论：丹参参改善低氧及复氧期间心肌能量代谢水平,减轻心肌低氧／复氧损伤,并能显著改善细胞内酸碱     

急性低氧/复氧胁迫对克氏原螯虾抗氧化-能量代谢的影响

为研究低氧/复氧胁迫对克氏原螯虾(Procambarus clarkii)抗氧化及能量代谢的影响,将克氏原螯虾暴露于(1.0±0.1) mg/L急性低氧胁迫和后续(6.8±0.2) mg/L复氧环境中,于低氧胁迫1h、6h及复氧1h、12h分别采集肝胰腺、鳃和血淋巴,研究低氧/复氧胁迫下克氏原螯虾抗氧化-能量代谢酶的活力变化,分析鳃和肝胰腺组织的超微结构改变。在低氧胁迫下,肝胰腺和血淋巴中SOD酶活力显著下降(P<0.05);复氧以后,肝胰腺、血淋巴及鳃组织中SOD酶活力均出现了显著上升(P<0.05)。SOD酶活力变化可能与复氧过程中超氧阴离子自由基的过量产生有关。在复氧12h后,血淋巴和鳃组织中MDA含量均出现了显著性增加(P<0.01),提示机体细胞在复氧胁迫下产生了脂质过氧化。在低氧胁迫下,肝胰腺、鳃和血淋巴中ACP、AKP酶活力显著上升(P<0.05);在复氧12h后,肝胰腺和鳃组织中ACP酶活力显著降低(P<0.01)。显示低氧/复氧胁迫影响了机体的非特异性免疫应答。在急性低氧胁迫下,肝胰腺、血淋巴与鳃组织中的LDH含量和总ATPase活力均显...     

Nitrites can be reduced in retinal vessels during hypoxia and protect the retina against ischemia and apoptosis

The possibility and the mechanism of the reduction of nitrites in retinal vessels under acute hypoxia in vivo have been investigated. An experimental model of rat retinal ischemia was elaborated using laser coagulation of retinal vessels. It was demonstrated that vessel thrombosis does not occur if the nitrite concentration in the vessels is increased. It was proposed that, under acute hypoxia, nitrites are reduced to NO, which results in drastic vasodilatation. Considering that the effect takes less than a minute, this reduction cannot be due to hypoxic acidosis but is more likely associated with NO reduction by heme proteins. It was found that increased concentration of nitrites protects the retina from the development of ischemia progress and that preliminary administration of nitrites prevents apoptosis in the retina and a decrease in its photoelectric activity.     

Role of catalase on the hypoxia/reoxygenation stress in the hypoxia-tolerant Nile tilapia

The specific contribution of each antioxidant enzyme to protection against the reoxygenation-associated oxidative stress after periods of hypoxia is not well understood. We assessed the physiological role of catalase during posthypoxic reoxygenation by the combination of two approaches. First, catalase activity of Nile tilapias (Oreochromis niloticus) was 90% suppressed by intraperitoneal injection of 3-amino-1,2,4-triazole (ATZ, 1g/kg). In ATZ-injected fish, liver GSH levels, oxidative stress markers, and activities of other antioxidant enzymes remained unchanged. Second, animals with depleted catalase activity (or those saline-injected) were subjected to a cycle of severe hypoxia (dissolved O(2) = 0.28 mg/l for 3 h) followed by reoxygenation (0.5 to 24 h). Hypoxia did not induce changes in the above-mentioned parameters, either in saline- or in ATZ-injected animals. Reoxygenation increased superoxide dismutase activity in saline-injected fish, whose levels were similar to ATZ-injected animals. The activities of glutathione S-transferase, selenium-dependent glutathione peroxidase, and total-GPX and the levels of GSH-eq, GSSG, and thiobarbituric acid reactive substances remained unchanged during reoxygenation in both saline- and ATZ-injected fish. The GSSG/GSH-eq ratio in ATZ-injected fish increased at 30 min of reoxygenation compared with saline-injected ones. Reoxygenation also increased carbonyl protein levels in saline-injected fish, whose levels were similar to the ATZ-injected group. Our work shows that inhibition of liver tilapia catalase causes a redox imbalance during reoxygenation, which is insufficient to induce further oxidative stress. This indicates the relevance of hepatic catalase for hypoxia/reoxygenation stress in tilapia fish.     

不同发育时期的鸡胚色素上皮对体外视网膜细胞生长的作用(简报)

眼正常发育中,色素上皮层和视刚膜之间在结构上存在着紧密的接触,功能上也相互依赖,但色素上皮的作用不甚清楚。很多脊椎动物的视网膜如果失去了与色素上皮层的紧密     

Effect of prior exercise on maximal short-term power output in humans

The effect of prior exercise (PE) on subsequent maximal short-term power output (STPO) was examined during cycling exercise on an isokinetic ergometer. In the first series of experiments the duration of PE at a power output equivalent to 98% maximum O2 uptake (VO2max) was varied between 0.5 and 6 min before measurement of maximal STPO. As PE duration increased subsequent STPO fell to approximately 70% of control values after 3-6 min. In series ii the effect of varying the intensity of PE of fixed 6-min duration was studied in five subjects. After PE less than 60% VO2max there was an increase of 12% in STPO, but after greater than 60% VO2max there was a progressive fall in STPO as PE intensity increased, indicating a reduction of approximately 35% at 100% VO2max compared with control values. In series iii we examined the effect on STPO of allowing a recovery period after a fixed intensity (mean = 87% VO2max) of 6 min PE before measurement of STPO. This indicated a rapid recovery of dynamic function with a half time of approximately 32 s, which is similar to the kinetics of PC resynthesis and taken with the other findings suggests the dominant role that PC exerts on the STPO under these conditions.     

Effect of pigmented epithelium from different developmental stages of chick embryo on cell growth of retinal explants

The model system to investigate the effect of retinal pigmented epithelium (PE) on the retinal development in vitro has been established in this laboratory. Chick retina separated from 5-day-old embryo (E 5) were cut into strips and explanted on the collagen substratum either in close contact with retinal PE (RPE), or without PE (R). The rates of cell proliferation of retinal strips cultured for 48 hr were measured by the uptake of radioactive thymidine and DNA contents. Both parameters in RPE were increased to values ranging from 137 to 167% when PE was taken from E5 and E6. However PE taken from E7, E8 and E9 had no effect on cell proliferation. The rate of cell proliferation of retina were increased both when separated retina and PE of E5 either from same or from an other eye closely contact again and when retina and PE of E5 were explanted together without separation. However the rates of cell proliferation were remained without much change when a millipore filter existed between retina and PE of E5 as well as the retina was inverted, the ganglion cell layer contacted with PE. The neurite outgrowth from retina explant with and without PE of E5 or E6 were also different. After culture for 24 hr the fiber length of neurite growing in RPE was only 36-39% of that in R. After 48 hr it was about 70% of that in R. This results suggested that the developmental stage of PE and the direct cell-to-cell contact of PE from E5 with photoreceptor layer of retina was important for the retinal cell proliferation. But PE had negative influence on neurite growth of retina in culture.     

Hypoxia and recovery perturb free radical processes and antioxidant potential in common carp (Cyprinus carpio) tissues

The effects of hypoxia exposure and subsequent normoxic recovery on the levels of lipid peroxides (LOOH), thiobarbituric acid reactive substances (TBARS), carbonylproteins, total glutathione levels, and the activities of six antioxidant enzymes were measured in brain, liver, kidney and skeletal muscle of the common carp Cyprinus carpio. Hypoxia exposure (25% of normal oxygen level) for 5h generally decreased the levels of oxidative damage products, but in liver TBARS content were elevated. Hypoxia stimulated increases in the activities of catalase (by 1.7-fold) and glutathione peroxidase (GPx) (by 1.3-fold) in brain supporting the idea that anticipatory preparation takes place in order to deal with the oxidative stress that will occur during reoxygenation. In liver, only GPx activity was reduced under hypoxia and reoxygenation while other enzymes were unaffected. Kidney showed decreased activity of GPx under aerobic recovery but superoxide dismutase (SOD) and catalase responded with sharp increases in activities. Skeletal muscle showed minor changes with a reduction in GPx activity under hypoxia exposure and an increase in SOD activity under recovery. Responses by antioxidant defenses in carp organs appear to include preparatory increases during hypoxia by some antioxidant enzymes in brain but a more direct response to oxidative insult during recovery appears to trigger enzyme responses in kidney and skeletal muscle.