Report for the Eighth Asian National Control Laboratory Network meeting in 2023: Self-sufficiency strategy of plasma-derived medicinal products and regulatory harmonisation.

The Eighth Asian National Control Laboratory (NCL) Network meeting, entitled "Biological Products Quality Control and Self-Sufficiency Strategy focusing on plasma-derived medicinal products (PDMPs)" was held in Seoul on 31 August 2023. The participants were NCL experts from Indonesia, Japan, Malaysia, the Philippines, Vietnam, and the Republic of Korea. Special lectures included the PDMPs self-sufficiency strategies of the World Health Organization (WHO) and Indonesian Food and Drug Authority, and a case study on Global Benchmarking Tool (GBT) assessment for vaccines by the Korea Ministry of Food and Drug Safety. The NCL delegates shared their current experiences with national lot releases and biological standardisation. The meeting contributed to a mutual understanding of the progress of the PDMPs self-sufficiency among Asian countries, the WHO's support strategies, and the NCL's plan for the preparation of the WHO GBT assessment. In the panel discussion, all participants agreed that building capacity in blood safety in the Asian region and harmonisation of relevant international regulatory requirements will support appropriate emergency preparedness, particularly source materials in the region, and will build the foundation for resolving the PDMPs supply insecurity that has worsened after the COVID-19 pandemic in some countries.

Report on the seventh meeting of national control laboratories for vaccines and biologicals of the WHO Western Pacific and South-East Asia member states.

The Biregional Network of National Control Laboratories (NCLs) of the WHO Western Pacific and South-East Asia Regions has been meeting annually since 2018 to enhance NCLs' voluntary participation capacity. Its seventh meeting was hosted by the Korea National Institute of Food and Drug Safety Evaluation (NIFDS) of the Ministry of Food and Drug Safety (MFDS), in conjunction with the Global Bio Conference, in Seoul on September 6, 2022. Over 60 participants from seven countries, (India, Indonesia, Japan, Korea, Malaysia, the Philippines, and Vietnam) attended the meeting on-site and online. The theme of this meeting was 'Quality Control Issues and International Trends for Biologicals including Vaccines and Plasma-Derived Medicinal Products.' Three special speeches were presented on sharing the quality control system for biologicals, including NCLs' considerations in preparing the WHO Listed Authorities and sharing MFDS experiences. Furthermore, the participating NCLs shared country-specific issues related to national lot releases during the COVID-19 pandemic and acknowledged the meeting's crucial role in response preparedness for pandemic emergencies and enhancing regulatory capacity through coalitions and information exchange among NCLs. The NIFDS will cooperate closely with other Asian NCLs to enhance biological product quality control, aiming to establish regional standards and standardize test methods through collaboration.

A collaborative study to establish the second national standard for hepatitis B immunoglobulin in Korea.

This study aimed to establish a second national standard for hepatitis B immunoglobulin (HBIG) that can be used for potency assays of hepatitis B and normal immunoglobulin. The candidate material was manufactured using a process approved as Good Manufacturing Practice. The freeze-dried candidate preparation was tested for physicochemical and biological properties, including pH, residual moisture, molecular size distribution, and potency. A collaborative study was performed involving four laboratories, including the National Institute of Food and Drug Safety Evaluation, as an official national control laboratory in Korea and manufacturers. The potency was calibrated against the second international standard for HBIG using two enzyme immunoassays: enzyme-linked immunosorbent assay and electrochemiluminescence immunoassay. Results from 240 assays were obtained from four laboratories, and combined potency estimates were obtained by calculating the geometric means. Intra- and inter-laboratory variability showed acceptable geometric coefficients of variation of 1.3-6.0 and 3.2-3.6%, respectively. The candidate preparation showed satisfactory stability in accelerated thermal degradation and real-time stability tests. Based on these results, the potency value of 105 IU/vial was assigned (95% confidence intervals: 100.0-109.2 IU/vial), and it was deemed suitable to serve as the Korean national standard for HBIG.

DJ-1 protects against undernutrition-induced atrophy through inhibition of the MAPK-ubiquitin ligase pathway in myoblasts.

AIMS: The purpose of this study is to explore whether antioxidant DJ-1 protein affects the atrophy of skeletal muscle cell induced by undernutrition. MAIN METHODS: To determine cell atrophic responses, L6 cell line and skeletal primary cells from mouse hind limbs were cultivated under condition of FBS-free and low glucose. Changes of protein expression were analyzed using Western blot. Overexpression and knockdown of DJ-1 was performed in cells to assess its influence on cell atrophic responses. KEY FINDINGS: Undernutrition decreased cell size and increased the abundance of oxidized form and total form of DJ-1 protein in L6 myoblasts. The undernourished cells revealed an elevation in the expression of muscle-specific RING finger-1 (MuRF-1) and atrogin-1, and in the phosphorylations of p38 mitogen-activated protein kinase (MAPK) and stress-activated protein kinase/c-Jun N-terminal kinase compared with control groups. Moreover, DJ-1-knockout mice showed a decrease in cell size and an enhancement in the expression of MuRF-1 and atrogin-1, as well as in the phosphorylation of MAPKs in gastrocnemius muscles; these changes were also observed in L6 cells transfected with siRNA of DJ-1. On the other hand, L6 cells overexpressing full-length DJ-1 did not exhibit the alterations in cell size and ubiquitin ligases seen after undernourished states of control cells. Myotubes differentiated from L6 cells also showed elevated expression of MuRF-1 and atrogin-1 in response to undernutrition. SIGNIFICANCE: These results suggest that DJ-1 protein may contribute to undernutrition-induced atrophy via MAPKs/ubiquitin ligase pathway in skeletal muscle cells.

Proteomic analysis of kidneys from selenoprotein M transgenic rats in response to increased bioability of selenium.

BACKGROUND: To characterize changes in global protein expression in kidneys of transgenic rats overexpressing human selenoprotein M (SelM) in response to increased bioabivility of selenium (Sel), total proteins extracted from kidneys of 10-week-old CMV/hSelM Tg and wild-type rats were separated by 2-dimensional gel electrophoresis and measured for changes in expression. RESULTS: Ten and three proteins showing high antioxidant enzymatic activity were up- and down-regulated, respectively, in SelM-overexpressing CMV/hSelM Tg rats compared to controls based on an arbitrary 2-fold difference. Up-regulated proteins included LAP3, BAIAP2L1, CRP2, CD73 antigen, PDGF D, KIAA143 homolog, PRPPS-AP2, ZFP313, HSP-60, and N-WASP, whereas down-regulated proteins included ALKDH3, rMCP-3, and STC-1. After Sel treatment, five of the up-regulated proteins were significantly increased in expression in wild-type rats, whereas there were no changes in CMV/hSelM Tg rats. Only two of the down-regulated proteins showed reduced expression in wild-type and Tg rats after Sel treatment. CONCLUSIONS: These results show the primary novel biological evidences that new functional protein groups and individual proteins in kidneys of Tg rats relate to Sel biology including the response to Sel treatment and SelM expression.

Nicastrin overexpression in transgenic mice induces aberrant behavior and APP processing.

Nicastrin (NCT) is a component of the presenilin protein complex, which is involved in the cleavage of ß-amyloid precursor protein (ßAPP) and Notch. The aim of this study was to determine the manner in which overexpression of wild-type human nicastrin (hNCTw) or mutant human nicastrin (hNCTm, D336A/Y337A) regulates brain functions and amyloid precusor protein (APP) processing. For this, we created transgenic (Tg) mice expressing neuron-specific enolase (NSE)-controlled hNCTw or hNCTm and measured their phenotypes as time passed. The NSE/hNCTw and NSE/hNCTm Tg groups exhibited greater behavioral dysfunction from 10 months of age than the non-Tg group, although their severities differed. Further, activity and component levels of the γ-secretase complex were significantly elevated in NSE/hNCTw Tg mice, expect for PEN-2. These alterations induced stimulation of APP processing, resulting in overproduction of Aß-42 peptide in the NSE/hNCTw Tg group, whereas the NSE/hNCTm Tg group showed a comparatively weaker effect. Furthermore, the highest expression levels of ß-secretase and NICD were observed in the NSE/hNCTw Tg group, similar to other phenotypes. Especially, a significances interference on the interaction between NCT and γ-secretase substrates was detected in NSE/hNCTm Tg groups compare with NSE/hNCTw Tg group. These results indicate that hNCTw overexpression in Tg mice promoted active assembly of the γ-secretase complex through modulation of APP processing and behavior, whereas the lesser effect in NSE/hNCTm Tg mice was due to reduced expression of hNCTm. These Tg mice could be useful for the development and application of therapeutic drugs in an animal model of Alzheimer's disease.

Green tea catechin leads to global improvement among Alzheimer's disease-related phenotypes in NSE/hAPP-C105 Tg mice.

Amyloid ß (Αß) has been reported to be responsible for the functional and structural abnormalities of Alzheimer's disease (AD) through the induction of oxidative stress. The aim of this study was to determine whether or not treatment of transgenic (Tg) mice with green tea catechin (GTC), a radical scavenger, improves AD phenotypes. To test this, 7-month-old Tg mice were treated with a low (1 mg) or high (10 mg) dose of GTC for 6 months. Surprisingly, GTC-treated Tg mice exhibited significant decreases in behavioral impairment, Aß-42 production, APP-C99/89 expression, γ-secretase component and Wnt protein levels, γ-secretase activity and MAPK activation. In contrast, the levels of APP-C83 protein and enzyme activities (α-secretase, neprilysin and Pin1) were elevated in the GTC-treated groups. Moreover, GTC-treated groups showed lower levels of total cholesterol and low-density lipoprotein cholesterol, whereas the level of high-density lipoprotein cholesterol increased. These results provide the first experimental evidence that GTC improves AD phenotypes, thereby suggesting that GTC can be used in the prevention of AD or treatment of AD patients.

DJ-1/park7 protects against neointimal formation via the inhibition of vascular smooth muscle cell growth.

AIMS: DJ-1/park7 is a ubiquitously expressed multifunctional protein that plays essential roles in a variety of cells. However, its function in the vascular system has not been determined. We investigated the protective roles of DJ-1/park7 in vascular disorders, especially in neointimal hyperplasia. METHODS AND RESULTS: DJ-1/park7 was strongly expressed in the neointimal layer, in which its oxidized form was predominant. Treatment of vascular smooth muscle cells (VSMCs) from the mouse aorta with H(2)O(2) increased the oxidation of DJ-1/park7 visualized on two-dimensional electrophoresis gels. The growth of VSMCs in FBS-containing media and the release of H(2)O(2) were significantly increased in DJ-1/park7(-/-) knockout mice compared with DJ-1/park7(+/+) wild-type mice. The expression of cyclin D1 and the phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 were greater in VSMCs from the DJ-1/park7(-/-) aorta than from the DJ-1/park7(+/+) aorta. Both of these measures were inhibited by treatment with an ERK1/2 inhibitor or antioxidants and in DJ-1/park7-overexpressing cells. VSMC proliferation, cyclin D1 expression, and ERK1/2 phosphorylation in response to platelet-derived growth factor-BB were upregulated in DJ-1/park7(-/-) compared with DJ-1/park7(+/+) mice. VSMCs of DJ-1/park7(-/-) mice exhibited higher levels of sprout outgrowth of aortic strips and neointimal plaque formation elicited by carotid artery ligation compared with those of DJ-1/park7(+/+) mice. CONCLUSION: These results indicate that DJ-1/park7 is involved in the growth of VSMCs, thereby inhibiting neointimal hyperplasia, and suggest that it might play protective roles in vascular remodelling.

Monitoring of antibiotic resistance in bacteria isolated from laboratory animals.

The drug resistance of microorganisms isolated from laboratory animals never treated with antibiotics is being reported consistently, while the number of laboratory animals used in medicine, pharmacy, veterinary medicine, agriculture, nutrition, and environmental and health science has increased rapidly in Korea. Therefore, this study examined the development of antimicrobial resistance in bacteria isolated from laboratory animals bred in Korea. A total of 443 isolates (7 species) containing 5 Sphingomonas paucimobilis, 206 Escherichia coli, 60 Staphylococcus aureus, 15 Staphylococcus epidermidis, 77 Enterococcus faecalis, 27 Citrobacter freundii, 35 Acinetobacter baumannii were collected from the nose, intestine, bronchus and reproductive organs of ICR mice and SD rats. Of these species, Acinetobacter baumannii and Enterococcus faecalis showed significant antimicrobial resistance according to the minimum inhibition concentration (MIC) in E-test. In case of Acinetobacter baumannii, several isolates showed MIC values 16-128 µg/mL for cefazolin and cefoxitin, and higher resistance (128-512 µg/mL) to nitrofurantoin than that of standard type. Resistance to cefazolin, cefoxitin and nitrofurantoin was detected in 17.14, 20.00, and 8.57% of the Acinetobacter baumannii isolates, respectively. In addition, 44.1% of the Enterococcus faecalis isolates collected from the laboratory animals were resistant to oxacillin concentration of 16-32 µg/mL range, while MIC value of standard type was below oxacillin concentration of 6 µg/mL. These results suggest that in rodent species of laboratory animals, Acinetobacter baumannii are resistance to cefazolin, cefoxitin and nitrofurantoin, whereas those of Enterococcus faecalis were resistance to oxacillin.

Overexpression of Insulin Degrading Enzyme could Greatly Contribute to Insulin Down-regulation Induced by Short-Term Swimming Exercise.

Exercise training is highly correlated with the reduced glucose-stimulated insulin secretion (GSIS), although it enhanced insulin sensitivity, glucose uptake and glucose transporter expression to reduce severity of diabetic symptoms. This study investigated the impact of short-term swimming exercise on insulin regulation in the Goto-Kakizaki (GK) rat as a non-obese model of non-insulin-dependent diabetes mellitus. Wistar (W/S) and GK rats were trained 2 hours daily with the swimming exercise for 4 weeks, and then the changes in the metabolism of insulin and glucose were assessed. Body weight was markedly decreased in the exercised GK rats compare to their non-exercised counterpart, while W/S rats did not show any exercise-related changes. Glucose concentration was not changed by exercise, although impaired glucose tolerance was improved in GK rats 120 min after glucose injection. However, insulin concentration was decreased by swimming exercise as in the decrease of GSIS after running exercise. To identify the other cause for exercise-induced insulin down-regulation, the changes in the levels of key factors involved in insulin production (C-peptide) and clearance (insulin-degrading enzyme; IDE) were measured in W/S and GK rats. The C-peptide level was maintained while IDE expression increased markedly. Therefore, these results showed that insulin down-regulation induced by short-term swimming exercise likely attributes to enhanced insulin clearance via IDE over-expression than by altered insulin production.

Pen-2 overexpression induces Aß-42 production, memory defect, motor activity enhancement and feeding behavior dysfunction in NSE/Pen-2 transgenic mice.

Pen-2 is a key regulator of the γ-secretase complex, which is involved in the production of the amyloid ß (Aß)-42 peptides, which ultimately lead to Alzheimer's disease (AD). While Pen-2 has been studied in vitro, Pen-2 function in vivo in the brains of transgenic (Tg) mice overexpressing human Pen-2 (hPen-2) protein has not been studied. This study aimed to determine whether Pen-2 overexpression could regulate the AD-like phenotypes in Tg mice. NSE/hPen-2 Tg mice were produced by the microinjection of the NSE/hPen-2 gene into the pronucleus of fertilized eggs. The expression of the hPen-2 gene under the control of the NSE promoter was successfully detected only in the brain and kidney tissue of NSE/hPen-2 Tg mice. Also, 12-month-old NSE/hPen-2 Tg mice displayed behavioral dysfunction in the water maze test, motor activity and feeding behavior dysfunction in food intake/water intake/motor activity monitoring system. In addition, tissue samples displayed dense staining with antibody to the Aß-42 peptide. Furthermore, NSE/hPen-2 Tg mice exhibiting feeding behavior dysfunction were significantly more apt to display symptoms related to diabetes and obesity. These results suggest that Pen-2 overexpression in NSE/hPen-2 Tg mice may induce all the AD-like phenotypes, including behavioral deficits, motor activity and feeding behavior dysfunction, Aß-42 peptide deposition and chronic disease induction.

GATA binding protein 3 overexpression and suppression significantly contribute to the regulation of allergic skin inflammation.

GATA binding protein 3 (GATA3) is a key molecule regulating the balance in the ratio of type 1 helper T (Th1) cells to type 2 helper T (Th2) cells, which is thought to be indicative of the pathogenesis of allergic diseases such as asthma and atopic dermatitis. The aim of this study was to investigate the role of GATA3 in allergic skin inflammation. Transgenic (Tg) mice overexpressing human GATA3 (hGATA3) were produced by the microinjection of pCMV/hGATA3 constructs into fertilized mouse eggs. The hGATA3 gene was successfully expressed at the protein level in the lymph node and thymus of CMV/hGATA3 transfected cells and Tg mice. CMV/hGATA3 Tg mice showed a significant increase in the allergic skin inflammation response such as ear thickness, draining auricular lymph node (aLN) weight, epidermal thickness, inflammatory cell number and Th2 immunoglobulin (Ig) concentration compared to wild-type (WT) mice after phthalic anhydride (PA) treatment. Furthermore, the secretion of Th2 type cytokines was increased by PA treatment in CMV/hGATA3 Tg mice, while the secretion of Th1 type cytokine was suppressed under the same conditions. However, the increased levels of Th2 type cytokines in CMV/hGATA3 Tg mice were almost recovered by the down-regulation of GATA3 expression with D-pinitol treatment. Therefore, these findings suggest that GATA3 could be considered as a potential target regulating the mechanism responsible for the differences in allergic skin inflammation.

Aß-42 deposition significantly increases the insolubility of synaptophysin in the brains of hAPPsw/hPS2m double transgenic mice.

Synaptophysin is a synaptic vesicle glycoprotein involved in the regulation process for neurotransmitter release, which is distributed throughout neuroendocrine cells and all neurons in the brain and spinal cord. In an effort to determine whether amyloid ß (Aß)-42 peptides could influence the quantity and biochemical properties of synaptophysin, alterations in the levels of the synaptophysin protein in various soluble fractions were detected in the brains of four genotypes of transgenic mice (Tg) including Non-Tg, neuron-specific enolase (NSE)-hPS2m, NSE-hAPPsw and hAPPsw/hPS2m double Tg mice. Among the four genotypes of Tg mice, the highest levels of Aß-42 peptides were noted in hAPPsw/hPS2m, followed by NSE-hAPPsw, NSE-hPS2m and Non-Tg mice. In the brains of these mice displaying different levels of Aß-42 peptides, the levels of soluble synaptophysin were reduced significantly only in the hAPPsw/hPS2m double Tg mice compared to the Non-Tg mice. However, immunohistochemical analysis revealed no differences in the levels of total synaptophysin protein between the neocortex and hippocampus of the four different genotypes of mice. Western blot analysis using four-step fractions with differing solubility revealed a marked decrease in synaptophysin levels in the Tris-buffer saline fraction of hAPPsw/hPS2m double Tg mice and a significant increase in the formic acid fraction, relative to the Non-Tg mice. The results obtained from our in vivo experiments in mice are identical to the results observed in SK-N-MC cells treated with 100 nM Aß-42 peptides. Therefore, our experiments collectively suggest that Aß-42 peptides may alter the solubility without changing the total amount of synaptophysin.

Detection of allergenic compounds using an IL-4/luciferase/CNS-1 transgenic mice model.

The interleukin-4 (IL-4) signaling cascade has been identified as a potentially important pathway in the development of allergies. The principal objective of this study was to produce novel transgenic (Tg) mice harboring the luciferase gene under the control of the human IL-4 promoter and the enhancer of IL-4 (CNS-1), in an effort to evaluate three types of allergens including a respiratory sensitizer, vaccine additives, and crude extracts of natural allergens in vivo. A new lineage of Tg mice was generated by the microinjection of pIL-4/Luc/CNS-1 constructs into a fertilized mice egg. The luciferase activity was successfully regulated by the IL-4 promoter in splenocytes cultured from IL-4/Luc/CNS-1 Tg mice. From the first five founder lines, one (#57) evidencing a profound response to ovalbumin was selected for use in evaluating the allergens. Additionally, the lungs, thymus, and lymph nodes of IL-4/Luc/CNS-1 Tg mice evidenced high luciferase activity in response to allergens such as phthalic anhydride (PA), trimellitic anhydride, ovalbumin, gelatin, Dermatophagoides pteronyssinus extracts, and Japanese cedar pollen, whereas key allergy-related indicators including ear thickness, Immunoglobulin E concentration, and the infiltration of inflammatory leukocytes in response to PA were unaltered in the Tg mice relative to the non-Tg mice. Furthermore, the expression levels of endogenous type 2 helper T cells cytokines and proinflammatory cytokines were similarly increased in these organs of IL-4/Luc/CNS-1 Tg mice in response to allergens. These results indicate that IL-4/Luc/CNS-1 Tg mice may be used as an animal model for the evaluation and prediction of the human body response to a variety of allergens originating from the environment and from certain industrial products.

¹H NMR-based metabolic profiling of naproxen-induced toxicity in rats.

The dose-dependent perturbations in urinary metabolite concentrations caused by naproxen toxicity were investigated using ¹H NMR spectroscopy coupled with multivariate statistical analysis. Histopathologic evaluation of naproxen-induced acute gastrointestinal damage in rats demonstrated a significant dose-dependent effect. Furthermore, principal component analysis (PCA) of ¹H NMR from rat urine revealed a dose-dependent metabolic shift between the vehicle-treated control rats and rats treated with low-dose (10 mg/kg body weight), moderate-dose (50 mg/kg), and high-dose (100 mg/kg) naproxen, coinciding with their gastric damage scores after naproxen administration. The resultant metabolic profiles demonstrate that the naproxen-induced gastric damage exhibited energy metabolism perturbations that elevated their urinary levels of citrate, cis-aconitate, creatine, and creatine phosphate. In addition, naproxen administration decreased choline level and increased betaine level, indicating that it depleted the main protective constituent of the gastric mucosa. Moreover, naproxen stimulated the decomposition of tryptophan into kynurenate, which inhibits fibroblast growth factor-1 and delays ulcer healing. These findings demonstrate that ¹H NMR-based urinary metabolic profiling can facilitate noninvasive and rapid diagnosis of drug side effects and is suitable for elucidating possible biological pathways perturbed by drug toxicity.

IL-6, VEGF, KC and RANTES are a major cause of a high irritant dermatitis to phthalic anhydride in C57BL/6 inbred mice.

BACKGROUND: In previous studies, several strains of mice were used as chemical-induced skin irritation models to identify immunological hazards and elucidate the molecular and cellular mechanisms by which irritant dermatitis disease occur. BALB/c and C57BL/6 mice have been used for most of these experiments. Although there are some differences in the immune response to chemical allergens between these strains, few studies have been conducted to determine what regulatory factors contribute to these variations. METHODS: To investigate the cause of high responses to skin irritation in C57BL/6 mice that are widely used to study atopic dermatitis, changes in various immune-related factors such as ear thickness, myeloperoxidase activity, lymph node weight, IgE concentration and cytokine concentration were measured in C57BL/6 and BALB/c mice following phthalic anhydride (PA) treatment. RESULTS: Based on analysis of the skin irritation, C57BL/6 mice showed a greater skin irritation to PA than BALB/c mice, although the IgE concentration and auricular lymph node weight did not contribute to this difference in the response. However, the concentration of several cytokines and chemokines (interleukin [IL]-6 and vascular endothelial growth factor [VEGF], keratinocyte-derived chemokine [KC] and regulated on activation normal T cell expressed and secreted [RANTES]) were significantly higher in C57BL/6 mice than BALB/c mice following treatment with PA. CONCLUSIONS: Our results suggest that several of the cytokines and chemokines secreted from irritant site could contribute to the regulation mechanism responsible for the difference in the skin irritation among various strains of mice following exposure to PA.

Characterization of changes in global gene expression in the brain of neuron-specific enolase/human Tau23 transgenic mice in response to overexpression of Tau protein.

Tau is a neuronal phosphoprotein responsible for the formation of the neurofibrillary tangles in Alzheimer's disease. To characterize the changes in global gene expression in the brain of transgenic mice that overexpress human Tau23 protein in response to the increase of Tau23 phosphorylation, total RNA extracted from the hippocampus of 12-month-old transgenic and wild-type mice was converted to cDNA, labeled with biotin and hybridized to oligonucleotide microarrays. The microarray results were confirmed by real-time RT-PCR and Western blotting method. It was determined that 43 genes were up-regulated and 8 genes were down-regulated by Tau23 in transgenic mice compared to controls, based on the arbitrary difference in the 2-fold change. Among the up-regulated transcripts, those encoding for transporter and oxidoreductase were dramatically over-represented, followed by those related to regulatory molecule, cytoskeletal protein, signaling molecule, and extracellular matrix protein. Genes encoding for transcription factor, regulatory molecule, miscellaneous function, and chaperone were significantly reduced in the down-regulated group. The major genes in the up-regulated categories included Ecrg4, Folr1, Defb11, Aqp1 and Soctdc1. The major genes in the down-regulated categories were Ncor1, Gpm6a, and Hspd1. These results indicate that the microarray analysis identifies several gene functional groups and individual genes that respond to a sustained increase in Tau23 phosphorylation levels in the brain of transgenic mice. In addition, the results suggest the microarray test is a useful tool for increased understanding of the role of Tau23 protein in regulating neurodegenerative disorders.

Repression of tau hyperphosphorylation by chronic endurance exercise in aged transgenic mouse model of tauopathies.

The present study was undertaken to investigate whether chronic endurance exercise affects tau phosphorylation levels in the brain with Alzheimer's disease (AD)-like pathology. To address this, the transgenic (Tg) mouse model of tauopathies, Tg-NSE/htau23, which expresses human tau23 in the brain, was chosen. Animals were subjected to chronic exercise for 3 months from 16 months of age. The exercised Tg mouse groups were treadmill run at speeds of 12 m/min (intermediate exercise group) or 19 m/min (high exercise group) for 1 hr/day, 5 days/week, during the 3-month period. Chronic endurance exercise in Tg mice increased the expression of Cu/Zn-superoxide dismutase (SOD) and catalase, and also their enzymatic activities in the brain. In parallel, chronic exercise in Tg mice up-regulated the expression of phospho-PKCalpha, phospho-AKT, and phospho-PI3K, and down-regulated the expressions of phospho-PKA, phosphor-p38, phospho-JNK, and phospho-ERK. Moreover, chronic exercise up-regulated both cytosolic and nuclear levels of beta-catenin, and the expression of T-cell factor-4 (Tcf-4) and cyclin D1 in the brain. As a consequence of such changes, the levels of phospho-tau in the brain of Tg mice were markedly decreased after exercise. Immunohistochemical analysis showed an exercised-induced decrease of the phospho-tau levels in the CA3 subregion of the hippocampus. These results suggest that chronic endurance exercise may provide a therapeutic potential to alleviate the tau pathology.

The effects of long-duration, low-temperature ground transportation on physiological and biochemical indicators of stress in mice.

Transportation can cause stress to laboratory animals and alter physiological characteristics that may confound experimental results. The authors investigated stress-related effects of 3-4 h of transportation by truck in two strains of mice (C57BL/6, which are known to be aggressive, and ICR, which are less aggressive). Transported mice had sufficient space and access to water, though temperature in the truck was lower than what is usually recommended. Transportation affected the following parameters in both strains of mice: (i) serum corticosterone concentrations, (ii) expression of the chaperone proteins Hsp70 and Grp78 in various tissues and (iii) concentrations of serological enzymes that are associated with liver disease. These parameters also differed substantially between the two strains of mice.

Overexpression of human selenoprotein M differentially regulates the concentrations of antioxidants and H2O2, the activity of antioxidant enzymes, and the composition of white blood cells in a transgenic rat.

Selenoprotein is associated with a variety of serious diseases, including infectious diseases, neurodegenerative disorders, cancer and cardiovascular disease. The aim of this study was to produce a new transgenic (Tg) rat expressing human selenoprotein M (SelM) in order to examine the protective function of the antioxidant status in vivo. To achieve this, a new lineage of Tg rats was produced by the microinjection of pCMV/GFP-hSelM constructs into a fertilized rat egg. Several conclusions can be drawn based on the results of the present study. The human SelM gene was successfully expressed at both the transcription and protein levels in the CMV/GFP-hSelM Tg rats. This Tg rat showed a different enzyme activity for the antioxidant protein in the various tissues examined. In response to the 2,2'-azobiz(2-amidinopropane) dihydrochloride (AAPH) injection, the Tg rats showed a lower level of antioxidant and H2O2 concentration as the activity of the antioxidant enzyme was maintained at a higher level in the Tg rats than in the non-Tg rats. Also, the neutrophil-to-lymphocyte ratio was significantly increased in this Tg rat, even though the level of corticosterone remained unchanged in both genotypes. Thus, the results of this study demonstrated that the CMV/GFP-hSelM Tg rat can serve as an animal model for the maintenance of a high level of antioxidant status and can be used to study the biological function of selenoprotein in infectious diseases, cardiovascular disease and cancer.