Jung-Eun Lee

The effect of biodegradable gelatin microspheres on the neuroprotective effects of high mobility group box 1 A box in the postischemic brain.

High mobility group box 1 (HMGB1) is a family of endogenous molecules that is released by necrotic cells and causes neuronal damages by triggering inflammatory processes. In the cerebral ischemic brain, sustained and regulated suppression of HMGB1 has been emerged as a therapeutic means to grant neuroprotection. HMGB1 consists of two HMG boxes (A and B) and an acidic C-terminal tail, and the A box peptide antagonistically competes with HMGB1 for its receptors. In the middle cerebral artery occlusion (MCAO) in rats, a murine model of transient cerebral ischemia, administration of HMGB1 A box intraparenchymally, after encapsulated in biodegradable gelatin microspheres (GMS), which enhances the stability of peptide inside and allows its sustained delivery, at 1 h, 3 h, or 6 h after MCAO, reduced mean infarct volumes by, respectively, 81.3%, 42.6% and 30.7% of the untreated MCAO-brain, along with remarkable improvement of neurological deficits. Furthermore, the administration of HMGB1 A box/GMS suppressed proinflammatory cytokine inductions more strongly than the injection of non-encapsulated HMGB1 A box. Given that insulted brains-like ischemia have enhanced gelatinase activity than the normal brain, our results suggest that GMS-mediated delivery of therapeutic peptides is a promising means to provide efficient neuroprotection in the postischemic brain.

Antidepressant effects of exercise are produced via suppression of hypocretin/orexin and melanin-concentrating hormone in the basolateral amygdala

Physical exercise is considered beneficial in the treatment of depression, but the underlying mechanism is not clearly understood. In the present study, we investigated the mechanism regulating antidepressant effects of exercise by focusing on the role of the amygdala using a well-defined animal model of depression. C57BL/6 mice treated with repeated restraint showed depression-like behaviors, which was counteracted by post-stress treatment with physical exercise. The two neuropeptides hypocretin/orexin (Hcrt/Orx) and melanin-concentrating hormone (MCH) were transcriptionally upregulated in the BLA after repeated stress, and their enhanced expression was downregulated by treatment with exercise, mirroring stress-induced depression-like behaviors and their reversal by exercise. Stereotaxic injection of either Hcrt/Orx peptide or MCH peptide within the BLA commonly increased phospho-CaMKIIα level and produced depression-like behaviors, mimicking the neural states in the BLA of mice subjected to repeated stress. In contrast, siRNA-mediated suppression of Hcrt/Orx or MCH in the BLA blocked stress-induced depression-like behaviors. Furthermore, siRNA-mediated inhibition of CaMKIIα in the BLA also counteracted stress-induced depression-like behaviors. Local injection of Hcrt/Orx peptide or MCH peptide within the BLA in exercise-treated animals blocked antidepressant-like effects of exercise. Together these results suggest that exercise produces antidepressant effects via suppression of Hcrt/Orx and MCH neural systems in the BLA.

Role of dopamine D2 receptors in optimizing choice strategy in a dynamic and uncertain environment

In order to investigate roles of dopamine receptor subtypes in reward-based learning, we examined choice behavior of dopamine D1 and D2 receptor-knockout (D1R-KO and D2R-KO, respectively) mice in an instrumental learning task with progressively increasing reversal frequency and a dynamic two-armed bandit task. Performance of D2R-KO mice was progressively impaired in the former as the frequency of reversal increased and profoundly impaired in the latter even with prolonged training, whereas D1R-KO mice showed relatively minor performance deficits. Choice behavior in the dynamic two-armed bandit task was well explained by a hybrid model including win-stay-lose-switch and reinforcement learning terms. A model-based analysis revealed increased win-stay, but impaired value updating and decreased value-dependent action selection in D2R-KO mice, which were detrimental to maximizing rewards in the dynamic two-armed bandit task. These results suggest an important role of dopamine D2 receptors in learning from past choice outcomes for rapid adjustment of choice behavior in a dynamic and uncertain environment.

Implementation of a Two-dimensional Behavior Matrix to Distinguish Individuals with Differential Depression States in a Rodent Model of Depression

Animal models of depression are used to study pathophysiology of depression and to advance therapeutic strategies. Stress-induced depression models in rodents are widely used. However, amenable behavioral criteria and experimental procedures that are suitable for animal models have not been established. Given that depression is clinically diagnosed by multiple symptomatic criteria and stress effects are imposed to the brain non-specifically in stress-induced depression models, analyses of depression states in rodents using multiple symptomatic criteria may provide more power than any methods relying on a single symptomatic criterion. To address this, C57BL/6 inbred mice were restrained for 2 h daily for 14 d, and depression states of individual mice were assessed using the U-field test, behavioral assessment developed to measure animals sociability, and the tail suspension test and/or forced swim test, which are the typical methods that measure psychomotor withdrawal states. Although the majority of these mice showed severe depressive behaviors in both tests, a significant proportion of them, which were all inbred mice and received the same amount of restraints, expressed differential depression states in the sociability test and psychomotor withdrawal tests. To easily read-out differential depression states of individuals in two different tests, a standard method and basic parameters required to construct two-way behavior matrix were introduced. The utility and features of this two-way behavior analysis method for studies of different depressive states of individuals were discussed.

Analysis of differential plaque depositions in the brains of Tg2576 and Tg-APPswe/PS1dE9 transgenic mouse models of Alzheimer disease

Adequate assessment of plaque deposition levels in the brain of mouse models of Alzheimer disease (AD) is required in many core issues of studies on AD, including studies on the mechanisms underlying plaque pathogenesis, identification of cellular factors modifying plaque pathology, and developments of anti-AD drugs. The present study was undertaken to quantitatively evaluate plaque deposition patterns in the brains of the two popular AD models, Tg2576 and Tg-APPswe/PS1dE9 mice. Coronally-cut brain sections of Tg2576 and Tg-APPswe/PS1dE9 mice were prepared and plaque depositions were visualized by staining with anti-amyloid β peptides antibody. Microscopic images of plaque depositions in the prefrontal cortex, parietal cortex, piriform cortex and hippocampus were obtained and the number of plaques in each region was determined by a computer-aided image analysis method. A series of optical images representing a gradual increase of plaque deposition levels were selected in the four different brain regions and were assigned in each with a numerical grade of 1-6, where +1 was lowest and +6, highest, so that plaques per unit in mm2 increased sigmoidally over the grading scales. Analyzing plaque depositions using the photographic plaque reference panels and a computer-aid image analysis method, it was demonstrated that the brains of Tg2576 mice started to accumulate predominantly small plaques, while the brains of Tg-APPswe/PS1dE9 mice deposited relatively large plaques.

G9a-Mediated Regulation of OXT and AVP Expression in the Basolateral Amygdala Mediates Stress-Induced Lasting Behavioral Depression and Its Reversal by Exercise

Chronic stress produces behavioral depression. Conversely, physical exercise is held to be beneficial in the treatment of depression. Although genomic mechanisms are likely involved in these behavioral changes, underlying mechanisms are not clearly understood. In the present study, we investigated whether stress effects and their reversal by exercise occur via genomic mechanisms in the amygdala, a core part of the limbic system important for regulating mood states. Mice treated with chronic restraint showed lasting depression-like behaviors, which were counteracted by treatment with scheduled forceful exercise. Microarray analysis identified a number of genes whose expression in the amygdala was either upregulated or downregulated after repeated stress, and these changes were reversed by exercise. Of these genes, the neuropeptides oxytocin (OXT) and arginine vasopressin (AVP) were selected as representative stress-induced and exercise-responded genes in the BLA. Stereotaxic injection of OXT or AVP receptor agonists within the BLA in normal mice produced depression-like behaviors, whereas small interfering RNA (siRNA)-mediated suppression of the OXT or AVP transcripts in the BLA was sufficient to block stress-induced depressive behaviors. Stress-induced depression-like behaviors were accompanied by a global reduction of G9a histone methyltransferase and H3K9me2 at the OXT and AVP promoters. Conversely, repeated exercise increased the levels of G9a and H3K9me2 at the OXT and AVP promoters in the BLA, which was associated with the suppression of OXT and AVP expressions. These results identify G9a-induced histone methylation at the OXT and AVP promoters in the BLA as a mechanism for mediating stress-induced lasting behavioral depression and its reversal by exercise.

Expression of the plant viral protease NIa in the brain of a mouse model of Alzheimer's disease mitigates Aβ pathology and improves cognitive function.

The plant viral protease, NIa, has a strict substrate specificity for the consensus sequence of Val-Xaa-His-Gln, with a scissoring property after Gln. We recently reported that NIa efficiently cleaved the amyloid-β (Aβ) peptide, which contains the sequence Val-His-His-Gln in the vicinity of the cleavage site by α-secretase, and that the expression of NIa using a lentiviral system in the brain of AD mouse model reduced plaque deposition levels. In the present study, we investigated whether exogenous expression of NIa in the brain of AD mouse model is beneficial to the improvement of cognitive deficits. To address this question, Lenti-NIa was intracerebrally injected into the brain of Tg-APPswe/PS1dE9 (Tg-APP/PS1) mice at 7 months of age and behavioral tests were performed 15-30 days afterwards. The results of the water maze test indicated that Tg-APP/PS1 mice which had been injected with Lenti-GFP showed an increased latency in finding the hidden-platform and markedly enhanced navigation near the maze-wall, and that such behavioral deficits were significantly reversed in Tg-APP/PS1 mice injected with Lenti-NIa. In the passive avoidance test, Tg-APP/PS1 mice exhibited a severe deficit in their contextual memory retention, which was reversed by NIa expression. In the marble burying test, Tg-APP/PS1 mice buried marbles fewer than non-transgenic mice, which was also significantly improved by NIa. After behavioral tests, it was verified that the Tg-APP/PS1 mice with Lenti-NIa injection had reduced Aβ levels and plaque deposition when compared to Tg-APP/PS1 mice. These results showed that the plant viral protease, NIa, not only reduces Aβ pathology, but also improves behavioral deficits.

Ethambutol-mediated cell wall modification in recombinant Corynebacterium glutamicum increases the biotransformation rates of cyclohexanone derivatives.

The effects of structural modification of cell wall on the biotransformation capability by recombinant Corynebacterium glutamicum cells, expressing the chnB gene encoding cyclohexanone monooxygenase of Acinetobacter calcoaceticus NCIMB 9871, were investigated. Baeyer–Villiger oxygenation of 2-(2′-acetoxyethyl) cyclohexanone (MW 170 Da) into R-7-(2′-acetoxyethyl)-2-oxepanone was used as a model reaction. The whole-cell biotransformation followed Michaelis–Menten kinetics. The Vmax and KS values were estimated as 96.8 U g−1 of dry cells and 0.98 mM, respectively. The Vmax was comparable with that of cyclohexanone oxygenation, whereas the KS was almost eightfold higher. The KS value of 2-(2′-acetoxyethyl) cyclohexanone oxygenation was reduced by ca. 30% via altering the cell envelop structure of C. glutamicum with ethambutol, which inhibits arabinosyl transferases involved in the biosynthesis of cell wall arabinogalactan and mycolate layers. The higher whole-cell biotransformation rate was also observed in the oxygenation of ethyl 2-cyclohexanone acetate upon ethambutol treatment of the recombinant C. glutamicum. Therefore, it was assumed that the biotransformation efficiency of C. glutamicum-based biocatalysts, with respect to medium- to large-sized lipophilic organic substrates (MW > ca. 170), can be enhanced by engineering their cell wall outer layers, which are known to function as a formidable barrier to lipophilic molecules.

Robust Protective Effects of a Novel Multimodal Neuroprotectant Oxopropanoyloxy Benzoic Acid (a Salicylic Acid/Pyruvate Ester) in the Postischemic Brain

Cerebral ischemia leads to brain injury via a complex series of pathophysiological events. Therefore, multidrug treatments or multitargeting drug treatments are attractive options in efficiently limiting brain damage. Here, we report a novel multifunctional compound oxopropanoyloxy benzoic acid (OBA-09), a simple ester of pyruvate and salicylic acid. This protective effect was manifested by recoveries from neurological and behavioral deficits. OBA-09 exhibited antioxidative effects in the postischemic brain, which was evidenced by remarkable reduction of lipid peroxidation and 4-hydroxy-2-nonenal staining in OBA-09-administered animals. Reactive oxygen species generation was markedly suppressed in primary cortical cultures under oxygen-glucose deprivation. More interestingly, OBA-09 was capable of scavenging hydroxyl radical in cell-free assays. High-performance liquid chromatography results demonstrated that OBA-09 was hydrolyzed to salicylic acid and pyruvate with t1/2 = 43 min in serum and 4.2 h in brain parenchyma, indicating that antioxidative function of OBA-09 is executed by itself and also by salicylic acid after the hydrolysis. In addition to antioxidative function, OBA-09 exerts anti-excitotoxic and anti-Zn2+-toxic functions, which might be attributed to attenuation of ATP and nicotinamide adenine dinucleotide depletion and to the suppression of nuclear factor-κB activity induction. Together these results indicate that OBA-09 has a potent therapeutic potential as a multimodal neuroprotectant in the postischemic brain and these effects were conferred by OBA-09 itself and subsequently its hydrolyzed products.

TRH and TRH receptor system in the basolateral amygdala mediate stress-induced depression-like behaviors.

Chronic stress is a potent risk factor for depression, but the mechanism by which stress causes depression is not fully understood. To investigate the molecular mechanism underlying stress-induced depression, C57BL/6 inbred mice were treated with repeated restraint to induce lasting depressive behavioral changes. Behavioral states of individual animals were evaluated using the forced swim test, which measures psychomotor withdrawals, and the U-field test, which measures sociability. From these behavioral analyses, individual mice that showed depression-like behaviors in both psychomotor withdrawal and sociability tests, and individuals that showed a resiliency to stress-induced depression in both tests were selected. Among the neuropeptides expressed in the amygdala, thyrotropin-releasing hormone (TRH) was identified as being persistently up-regulated in the basolateral amygdala (BLA) in individuals exhibiting severe depressive behaviors in the two behavior tests, but not in individuals displaying a stress resiliency. Activation of TRH receptors by local injection of TRH in the BLA in normal mice produced depressive behaviors, mimicking chronic stress effects, whereas siRNA-mediated suppression of either TRH or TRHR1 in the BLA completely blocked stress-induced depressive symptoms. The TRHR1 agonist, taltirelin, injection in the BLA increased the level of p-ERK, which mimicked the increased p-ERK level in the BLA that was induced by treatment with repeated stress. Stereotaxic injection of U0126, a potent inhibitor of the ERK pathway, within the BLA blocked stress-induced behavioral depression. These results suggest that repeated stress produces lasting depression-like behaviors via the up-regulation of TRH and TRH receptors in the BLA.