Astrocyte Mitochondria in In Vitro Models of Ischemia

There is growing evidence that preservation of mitochondrial respiratory function during cerebral ischemia-reperfusion predicts the ultimate extent of tissue injury. Because neurons are selectively vulnerable to ischemic injury, many studies have focused on neuronal mitochondrial dysfunction in ischemia. However, positron emission tomography (PET) studies in animals and humans suggest that non-neuronal cells such as astrocytes may also experience mitochondrial metabolic compromise that contributes to ischemic necrosis. Astrocytes carry out a number of functions that are critical to normal nervous system function, including uptake of neurotransmitters, regulation of pH and ion concentrations, and metabolic support of neurons. Mitochondria are important for many of these actions. We have used a cell culture model of stroke, oxygen-glucose deprivation (OGD), to study the response of astrocyte mitochondria to ischemia, and to evaluate how changes in astrocyte mitochondrial function might affect neuronal survival and recovery after ischemia.     

Astrocyte Mitochondrial Mechanisms of Ischemic Brain Injury and Neuroprotection

Research on ischemic brain injury has established a central role of mitochondria in neuron death. Astrocytes are also damaged by ischemia, although the participation of mitochondria in their injury is ill defined. As astrocytes are responsible for neuronal metabolic and trophic support, astrocyte dysfunction will compromise postischemic neuronal survival. Ischemic alterations to astrocyte energy metabolism and the uptake and metabolism of the excitatory amino acid transmitter glutamate may be particularly important. Despite the significance of ischemic astrocyte injury, little is known of the mechanisms responsible for astrocyte death and dysfunction. This review focuses on differences between astrocyte and neuronal metabolism and mitochondrial function, and on neuronal-glial interactions. The potential for astrocyte mitochondria to serve as targets of neuroprotective interventions is also discussed.     

线粒体在运动保护心肌免受缺血再灌注损伤中的作用

线粒体是参与心肌缺血再灌注(myocardial ischemia and reperfusion,MI/R)损伤的关键细胞器,线粒体活性氧(reactive oxygen species,ROS)爆发、Ca²⁺失调、线粒体通透性转换孔(mitochondrial permeability transition pore,mPTP)开放、线粒体肿胀、促凋亡蛋白释放等都会导致线粒体功能障碍,心肌功能受损。运动是预防MI/R损伤的有效干预手段,其保护作用可能通过线粒体来实现。运动保护MI/R损伤的线粒体机制由多种因素决定,如线粒体能量学、K_ATP通道、mPTP、线粒体跨膜电位(ΔΨ_m)、线粒体蛋白、线粒体脂质、线粒体质量控制、远程调控因子等。本文综述了MI/R产生的线粒体机制,运动对MI/R的保护作用以及线粒体在其中的作用,以期为MI/R损伤的线粒体治疗策略提供参考。     

Reciprocal Modulation Between Microglia and Astrocyte in Reactive Gliosis Following the CNS Injury

Reactive gliosis, also known as glial scar formation, is an inflammatory response characterized by the proliferation of microglia and astrocytes as well as astrocytic hypertrophy following injury in the central nervous system (CNS). The glial scar forms a physical and molecular barrier to isolate the injured area from adjacent normal nervous tissue for re-establishing the integrity of the CNS. It prevents the further spread of cellular damage but represents an obstacle to regrowing axons. In this review, we integrated the current findings to elucidate the tightly reciprocal modulation between activated microglia and astrocytes in reactive gliosis and proposed that modification of cellular response to the injury or cellular reprogramming in the glial scar could lead advances in axon regeneration and functional recovery after the CNS injury.     

Expression of Dixdc1 and its Role in Astrocyte Proliferation after Traumatic Brain Injury

DIX domain containing 1 (Dixdc1), a positive regulator of Wnt signaling pathway, is recently reported to play a role in the neurogenesis. However, the distribution and function of Dixdc1 in the central nervous system (CNS) after brain injury are still unclear. We used an acute traumatic brain injury (TBI) model in adult rats to investigate whether Dixdc1 is involved in CNS injury and repair. Western blot analysis and immunohistochemistry showed a time-dependent up-regulation of Dixdc1 expression in ipsilateral cortex after TBI. Double immunofluorescent staining indicated a colocalization of Dixdc1 with astrocytes and neurons. Moreover, we detected a colocalization of Ki-67, a cell proliferation marker with GFAP and Dixdc1 after TBI. In primary cultured astrocytes stimulated with lipopolysaccharide, we found enhanced expression of Dixdc1 in parallel with up-regulation of Ki-67 and cyclin A, another cell proliferation marker. In addition, knockdown of Dixdc1 expression in primary astrocytes with Dixdc1-specific siRNA transfection induced G0/G1 arrest of cell cycle and significantly decreased cell proliferation. In conclusion, all these data suggest that up-regulation of Dixdc1 protein expression is potentially involved in astrocyte proliferation after traumatic brain injury in the rat.     

Effect of Ca2+ on In Vitro Astrocyte Injury

Abstract: Current literature suggests that a massive influx of Ca²⁺ into the cells of the CNS induces cell damage associated with traumatic brain injury (TBI). Using an in vitro model for stretch-induced cell injury developed by our laboratory, we have investigated the role of extracellular Ca²⁺ in astrocyte injury. The degree of injury was assessed by measurement of propidium iodide uptake and release of lactate dehydrogenase. Based on results of in vivo models of TBI developed by others, our initial hypothesis was that decreasing extracellular Ca²⁺ would result in a reduction in astrocyte injury. Quite unexpectedly, our results indicate that decreasing extracellular Ca²⁺ to levels observed after in vivo TBI increased astrocyte injury. Elevating the extracellular Ca²⁺ content to twofold above physiological levels (2 m M ) produced a reduction in cell injury. The reduction in injury afforded by Ca²⁺ could not be mimicked with Ba²⁺, Mn²⁺, Zn²⁺, or Mg²⁺, suggesting that a Ca²⁺-specific mechanism is involved. Using ⁴⁵Ca²⁺, we demonstrate that injury induces a rapid influx of extracellular Ca²⁺ into the astrocyte, achieving an elevation in total cell-associated Ca²⁺ content two- to threefold above basal levels. Pharmacological elevation of intracellular Ca²⁺ levels with the Ca²⁺ ionophore A23187 or thapsigargin before injury dramatically reduced astrocyte injury. Our data suggest that, contrary to popular assumptions, an elevation of total cell-associated Ca²⁺ reduces astrocyte injury produced by a traumatic insult.     

Oxidative Activity of Mitochondria Isolated from Plant Tissues Sensitive and Resistant to Chilling Injury

Arrhenius plots of the respiration rates of mitochondria isolated from chilling sensitive plant tissues (tomato and cucumber fruit, and sweet potato roots) showed a linear decrease from 25 C to about 9 to 12 C (with Q(10) values of 1.3 to 1.6), at which point there was a marked deviation with an increased slope as temperatures were reduced to 1.5 C (Q(10) of 2.2 to 6.3). The log of the respiration rate of mitochondria from chilling resistant tissues (cauliflower buds, potato tubers, and beet roots) showed a linear decrease over the entire temperature range from 25 to 1.5 C with Q(10) values of 1.7 to 1.8. Phosphorylative efficiency of mitochondria from all the tissues, as measured by ADP:O and respiratory control ratios, was not influenced by temperatures from 25 to 1.5 C. These results indicate that an immediate response of sensitive plant tissues to temperatures in the chilling range (0 to 10 C) is to depress mitochondrial respiration to an extent greater than that predicted from Q(10) values measured above 10 C. The results are also consistent with the hypothesis that a phase change occurs in the mitochondrial membrane as the result of a physical effect of temperature on some membrane component such as membrane lipids.     

Mitochondria and NMDA Receptor-Dependent Toxicity of Berberine Sensitizes Neurons to Glutamate and Rotenone Injury

The global incidence of metabolic and age-related diseases, including type 2 diabetes and Alzheimer''s disease, is on the rise. In addition to traditional pharmacotherapy, drug candidates from complementary and alternative medicine are actively being pursued for further drug development. Berberine, a nutraceutical traditionally used as an antibiotic, has recently been proposed to act as a multi-target protective agent against type 2 diabetes, dyslipidemias, ischemic brain injury and neurodegenerative diseases, such as Parkinson''s and Alzheimer''s disease. However, the safety profile of berberine remains controversial, as isolated reports suggest risks with acute toxicity, bradycardia and exacerbation of neurodegeneration. We report that low micromolar berberine causes rapid mitochondria-dependent toxicity in primary neurons characterized by mitochondrial swelling, increased oxidative stress, decreased mitochondrial membrane potential and depletion of ATP content. Berberine does not induce caspase-3 activation and the resulting neurotoxicity remains unaffected by pan-caspase inhibitor treatment. Interestingly, inhibition of NMDA receptors by memantine and MK-801 completely blocked berberine-induced neurotoxicity. Additionally, subtoxic nanomolar concentrations of berberine were sufficient to sensitize neurons to glutamate excitotoxicity and rotenone injury. Our study highlights the need for further safety assessment of berberine, especially due to its tendency to accumulate in the CNS and the risk of potential neurotoxicity as a consequence of increasing bioavailability of berberine.     

Mitochondria in plants

The Botanical Review - It is plain from this survey that the opinions of the most competent investigators are in conflict on many important points. The tendency of many workers to dogmatize upon...     

Succinoxidase Activity of Avocado Fruit Mitochondria in Relation to Temperature and Chilling Injury throughout the Climacteric Cycle

Mitochondria were isolated from `Fuerte' avocado fruit (Persea americana Mill.) at four different stages of the respiratory climacteric. Preclimacteric fruit had the highest rate of succinate oxidation and the postclimacteric mitochondria the lowest. Subsequently, successive additions of ADP increased the respiratory control ratio.     

Acclimation,Hydrogen Peroxide,and Abscisic Acid Protect Mitochondria against Irreversible Chilling Injury in Maize Seedlings

Our previous results indicated that 3-d-old dark-grown chilling-sensitive maize (Zea mays L.) seedlings did not survive 7 d of 4[deg]C chilling stress, but 69% of them survived similar stress when the seedlings were either preexposed to 14[deg]C for 3 d or pretreated with 0.1 mM H2O2 for 4 h at 27[deg]C (T.K. Prasad, M.D. Anderson, B.A. Martin, C.R. Stewart [1994] Plant Cell 6: 65-74) or 1 mM abscisic acid (ABA) for 24 h at 27[deg]C (M.D. Anderson, T.K. Prasad, B.A. Martin, C.R. Stewart [1994] Plant Physiol 105: 331-339). We discovered that chilling imposed oxidative stress on the seedlings. Since H2O2 accumulated during the periods of both acclimation and nonacclimation, we concluded that H2O2 had dual effects at low temperature: (a) During acclimation, its early transient accumulation signals the induction of antioxidant enzymes such as catalase 3 and peroxidase to scavenge H2O2; and (b) at 4[deg]C in nonacclimated seedlings, it accumulates to damaging levels in the tissues because of low levels of these and perhaps other antioxidant enzymes. Three-day-old seedlings pretreated with H2O2 (a mild oxidative stress) or ABA showed induced chilling tolerance. In the present study, we investigated whether mitochondria are a target for chilling-induced oxidative stress and, if so, what differences do acclimation, H2O2, or ABA make to protect mitochondria from irreversible chilling injury. The results indicated that chilling, in general, impairs respiratory activity, the cytochrome pathway of electron transport, and ATPase activity regardless of the treatment. In pretreated seedlings, the activities of catalase 3 and peroxidase in the mitochondria increased severalfold compared with control and nonacclimated seedlings. The increases in these antioxidant enzymes imply that mitochondria are under oxidative stress and such increases could initiate a protective mechanism in the mitochondria. Mitochondrial respiration is partially cyanide resistant during chilling stress and also after the 1st d of recovery. Upon further recovery over 3 d, in contrast to nonacclimated seedlings, the mitochondria of acclimation-, H2O2-, and ABA-treated seedlings showed the following recovery features. (a) The mitochondrial respiration changed from a cyanide-resistant to a cyanide-sensitive cytochrome pathway, (b) cytochrome oxidase activity recovered to control levels, (c) the ability of mitochondria to generate ATP was regained, and (d) the antioxidant enzyme activities remained at or above control levels. Based on these results, we conclude that chilling impairs mitochondrial function and that chilling-induced oxidative stress seems to be a factor, at least in part, for causing possible irreversible damage to the mitochondrial membrance components. Acclimation, H2O2, and ABA provide a protective mechanism by inducing antioxidant enzymes to protect mitochondria from irreversible oxidative damage that is absent in nonacclimated seedlings. Therefore, we conclude that the ability of the seedlings to recover from chilling injury is, at least in part, due to the ability of the mitochondria to resume normal function.     

Isolation-Inflicted Injury to Mitochondria from Fresh Pollen Gradually Overcome by an Active Strengthening during Germination

Activities of segments of the electron transport pathway of mitochondria isolated from pollen of Typha latifolia L. during the course of germination in vitro were compared with those of mitochondria in intact grains. For this purpose, suitable inhibitors and artificial substrates were selected for their ability to penetrate through the exine, intine, and plasmalemma. In contrast to their counterparts in vivo, mitochondria isolated during the initial stages of germination exhibited low rates of electron transport, resulting from loss of NAD⁺ and displacement of cytochrome c from its site of action. The phosphorylative capacity was also impaired. Great caution must be exercised therefore, before interpreting results obtained with isolated mitochondria.

The gradually acquired resistance of mitochondria to injury during isolation as germination proceeds was shown to depend on an energy-requiring process and not solely on a rearrangement at the membrane level, or imbibitional differences. De novo syntheses of proteins or fatty acids were not required for the strengthening of mitochondria since cycloheximide, chloramphenicol, and cerulenin did not prevent this change. The nature of the energy-requiring process remains obscure. It is probable that strengthening of mitochondrial membranes during seed germination has been misinterpreted due to similar effects of isolational injury.

     

Mechanism of Chilling Injury in Sweet Potato: X. Change in Lipid-Protein Interaction in Mitochondria from Cold-stored Tissue

Seventy per cent of the phospholipid in mitochondria from sweet potato roots was removed by aqueous acetone treatment. The amount of phospholipid that could be rebound to these lipid-depleted mitochondria roughly corresponded to the amount of phospholipid in untreated mitochondria. The activities of NADH-cytochrome c oxidoreductase, succinate-cytochrome c oxidoreductase, cytochrome oxidase, and succinoxidase in lipid-depleted mitochondria were restored by addition of mitochondrial phospholipid to about 60, 50, 15, and 35%, respectively, in comparison to untreated mitochondria. The capacity of lipid-depleted mitochondria from 14-day cold-stored tissue to bind mitochondrial phospholipid from healthy tissue was lower than that from healthy tissue. However, there was no large difference in activities of NADH-cytochrome c oxidoreductase and succinate-cytochrome c oxidoreductase between both phospholipid rebound lipid-depleted mitochondria from healthy and 14-day cold-stored tissues. On the other hand, activity of succinoxidase in phospholipid rebound lipid-depleted mitochondria from 14-day cold-stored tissue was decreased by about 50% of that from healthy tissue. Furthermore, the capacity of lipid-depleted mitochondria from 2-day cold-stored tissue to bind mitochondrial phospholipid from healthy tissue was higher than that from healthy tissue.     

Mitochondria Revisited. Alternative Functions of Mitochondria

This review explores the alternative functions of mitochondria inside the cell. In a general picture of mitochondrial functioning, the importance and uniqueness of these intrinsic functions make them irreplaceable by other intracellular compartments. Among these are, participation in apoptosis and cellular proliferation, regulation of the cellular redox state and level of second messengers, heme and steroid syntheses, production and transmission of a transmembrane potential, detoxication and heat production. In most of the listed functions, reactive oxygen species modulate a number of non-destructive cellular activities. Some of the mitochondrial functions are reviewed in detail.     

Transporters and Channels in Cytotoxic Astrocyte Swelling

Brain edema is a severe clinical complication in a number of pathologies and is a major cause of increased morbidity and death. The swelling of astrocytes caused by a disruption of water and ion homeostasis, is the primary event contributing to the cytotoxic form of brain edema. Astrocyte cytotoxic swelling ultimately leads to transcapillary fluxes of ions and water into the brain parenchyma. This review focuses on the implication of transporters and channels in cytotoxic astrocyte swelling in hyponatremia, ischemia, trauma and hepatic encephalopathy. Emphasis is put on some salient features of the astrocyte physiology, all related to cell swelling, i.e. predominance of aquaporins, control of K⁺ homeostasis and ammonia accumulation during the brain ammonia-detoxifying process.