Discrepant serum creatinine concentrations caused by paraprotein interference preceding diagnosis of monoclonal gammopathy of undetermined significance.

We report a man in his 70s who presented with discrepant serum creatinine concentrations in different hospitals at the same time. Further examinations of these discrepancies revealed turbidity of the serum sample and, thus, a reagent reaction and false hypercreatinine caused by paraprotein interference were suspected. Serum protein electrophoresis revealed a small amount of monoclonal γ globulin (2.9 g/L), which may have been involved in paraprotein interference. Monoclonal λ-type IgG was detected in the serum, resulting in a diagnosis of monoclonal gammopathy of undetermined significance. Previous studies indicated paraprotein interference in serum containing monoclonal IgM or a large amount of IgG (> 25 g/L). Although this case of paraprotein interference induced by a small amount of IgG is rare, a discrepancy in creatinine results may be an indicator leading to the diagnosis of plasma cell proliferative diseases.

Eicosapentaenoic Acid Preserves Mitochondrial Quality and Attenuates Cardiac Remodeling After Myocardial Infarction in Rats.

Eicosapentaenoic acid (EPA) reduces the risk of ischemic heart diseases and is a component of mitochondria. We herein investigated whether dietary EPA mediated mitochondrial fatty acid compositions, dynamics, and functions, resulting in the attenuation of cardiac remodeling after myocardial infarction (MI). The coronary artery of male rats was ligated to induce MI, and they were then treated with or without EPA (1000 mg/kg/day) for 12 weeks. The EPA treatment improved left ventricular systolic function and increased the mitochondrial content of EPA in the non-infarct region 12 weeks after MI. The content of ATP and mitochondrial complex II, III, and IV activities decreased after MI but were maintained by the EPA treatment in association with the preservation of optic atrophy 1, a mitochondrial fusion protein. The present results suggest that dietary EPA increased the mitochondrial content of EPA and preserved the expression of mitochondrial fusion proteins and energy metabolism, which attenuated left ventricular remodeling after MI.

Nicorandil Suppresses Ischemia-Induced Norepinephrine Release and Ventricular Arrhythmias in Hypertrophic Hearts.

PURPOSE: Ventricular arrhythmias (VAs) are a common cause of sudden death in acute myocardial infarction (MI), for which hypertension is a major risk factor. Nicorandil opens ATP-sensitive potassium (KATP) channels, which are expressed by nerve terminals and cardiomyocytes and regulate the release of norepinephrine (NE). However, the effects of nicorandil on ischemic NE release in cardiac tissue remain unclear. Therefore, we herein investigated whether nicorandil suppressed interstitial NE concentrations and VAs during acute MI in pressure overload-induced hypertrophic hearts. METHODS: Rats were divided into two groups: an abdominal aortic constriction (AAC) group and sham-operated (Sham) group. Four weeks after constriction, cardiac geometry and functions were examined using echocardiography and hemodynamic analyses. Myocardial ischemia was induced by coronary artery occlusion for 100 min with or without the administration of nicorandil. VAs were assessed by electrocardiography, and NE concentrations in the ischemic region were measured using a micro-dialysis method. RESULTS: AAC induced left ventricular hypertrophy with diastolic dysfunction. VAs markedly increased in the early phase (0-20 min) of ischemia in both groups and were more frequent in the AAC group. Cardiac interstitial NE concentrations were higher in the AAC group before ischemia and significantly increased during ischemia in both groups. Nicorandil significantly suppressed ischemia-induced VAs and NE increases in the AAC group. CONCLUSION: Ischemia-induced VAs were more frequent in hypertrophic hearts and associated with high interstitial concentrations of NE. The attenuation of ischemia-induced increases in NE through neuronal KATP opening by nicorandil may suppress ischemia-induced VAs in hypertrophic hearts.

Raman Metabolomics of Candida auris Clades: Profiling and Barcode Identification.

This study targets on-site/real-time taxonomic identification and metabolic profiling of seven different Candida auris clades/subclades by means of Raman spectroscopy and imaging. Representative Raman spectra from different Candida auris samples were systematically deconvoluted by means of a customized machine-learning algorithm linked to a Raman database in order to decode structural differences at the molecular scale. Raman analyses of metabolites revealed clear differences in cell walls and membrane structure among clades/subclades. Such differences are key in maintaining the integrity and physical strength of the cell walls in the dynamic response to external stress and drugs. It was found that Candida cells use the glucan structure of the extracellular matrix, the degree of α-chitin crystallinity, and the concentration of hydrogen bonds between its antiparallel chains to tailor cell walls' flexibility. Besides being an effective ploy in survivorship by providing stiff shields in the α-1,3-glucan polymorph, the α-1,3-glycosidic linkages are also water-insoluble, thus forming a rigid and hydrophobic scaffold surrounded by a matrix of pliable and hydrated ß-glucans. Raman analysis revealed a variety of strategies by different clades to balance stiffness, hydrophobicity, and impermeability in their cell walls. The selected strategies lead to differences in resistance toward specific environmental stresses of cationic/osmotic, oxidative, and nitrosative origins. A statistical validation based on principal component analysis was found only partially capable of distinguishing among Raman spectra of clades and subclades. Raman barcoding based on an algorithm converting spectrally deconvoluted Raman sub-bands into barcodes allowed for circumventing any speciation deficiency. Empowered by barcoding bioinformatics, Raman analyses, which are fast and require no sample preparation, allow on-site speciation and real-time selection of appropriate treatments.

Roles of autophagy in angiotensin II-induced cardiomyocyte apoptosis.

Autophagy is a self-degradation process of cytoplasmic components and occurs in the failing heart. Angiotensin II plays a critical role in the progression of heart failure and induces autophagy. We investigated the mechanism underlying angiotensin II-enhanced autophagy and examined the role of autophagy in angiotensin II-induced cardiomyocyte injury. Neonatal rat cardiomyocytes were treated with angiotensin II (1-100 nmol/L). Angiotensin II dose-dependently increased autophagy indicators of microtubule-associated protein 1 light chain (LC) 3-II and monodansylcadaverine-labelled vesicles. It also enhanced the intracellular production of reactive oxygen species (ROS), assessed by H2DCFDA, an intracellular ROS indicator. NADPH oxidase- and mitochondria-derived ROS production was increased by angiotensin II, while angiotensin II-induced LC3-II expression was suppressed by inhibitors of these sources of ROS. Confocal microscopy revealed that superoxide-producing mitochondria colocalized with lysosomes after the angiotensin II stimulation. Myocyte apoptosis was assessed by nuclear staining with DAPI and caspase-3 activity. A 6-h stimulation with angiotensin II did not affect myocyte apoptosis, while a co-treatment with 3-methyl-adenine (3MA), an autophagy inhibitor, augmented apoptosis. These results indicate that autophagy suppressed apoptosis because it removed damaged mitochondria in the early stages of the angiotensin II stimulation. A longer angiotensin II stimulation for 24 h induced apoptosis and propidium iodide-positive lethal myocytes, while the co-treatment with 3MA did not lead to further increases. In conclusion, angiotensin II-induced autophagy removes ROS-producing mitochondria. Autophagy is a beneficial phenomenon against myocyte apoptosis in the early phase, but its benefit was limited in the late phase of angiotensin II stimulation.

Raman Study of Pathogenic Candida auris: Imaging Metabolic Machineries in Reaction to Antifungal Drugs.

The multidrug-resistant Candida auris often defies treatments and presently represents a worldwide public health threat. Currently, the ergosterol-targeting Amphotericin B (AmB) and the DNA/RNA-synthesis inhibitor 5-flucytosine (5-FC) are the two main drugs available for first-line defense against life-threatening Candida auris infections. However, important aspects of their mechanisms of action require further clarification, especially regarding metabolic reactions of yeast cells. Here, we applied Raman spectroscopy empowered with specifically tailored machine-learning algorithms to monitor and to image in situ the susceptibility of two Candida auris clades to different antifungal drugs (LSEM 0643 or JCM15448T, belonging to the East Asian Clade II; and, LSEM 3673 belonging to the South African Clade III). Raman characterizations provided new details on the mechanisms of action against Candida auris Clades II and III, while also unfolding differences in their metabolic reactions to different drugs. AmB treatment induced biofilm formation in both clades, but the formed biofilms showed different structures: a dense and continuous biofilm structure in Clade II, and an extra-cellular matrix with a "fluffy" and discontinuous structure in Clade III. Treatment with 5-FC caused no biofilm formation but yeast-to-hyphal or pseudo-hyphal morphogenesis in both clades. Clade III showed a superior capacity in reducing membrane permeability to the drug through chemically tailoring chitin structure with a high degree of acetylation and fatty acids networks with significantly elongated chains. This study shows the suitability of the in situ Raman method in characterizing susceptibility and stress response of different C. auris clades to antifungal drugs, thus opening a path to identifying novel clinical solutions counteracting the spread of these alarming pathogens.

Raman Spectroscopy of Oral Candida Species: Molecular-Scale Analyses, Chemometrics, and Barcode Identification.

Oral candidiasis, a common opportunistic infection of the oral cavity, is mainly caused by the following four Candida species (in decreasing incidence rate): Candida albicans, Candida glabrata, Candida tropicalis, and Candida krusei. This study offers in-depth Raman spectroscopy analyses of these species and proposes procedures for an accurate and rapid identification of oral yeast species. We first obtained average spectra for different Candida species and systematically analyzed them in order to decode structural differences among species at the molecular scale. Then, we searched for a statistical validation through a chemometric method based on principal component analysis (PCA). This method was found only partially capable to mechanistically distinguish among Candida species. We thus proposed a new Raman barcoding approach based on an algorithm that converts spectrally deconvoluted Raman sub-bands into barcodes. Barcode-assisted Raman analyses could enable on-site identification in nearly real-time, thus implementing preventive oral control, enabling prompt selection of the most effective drug, and increasing the probability to interrupt disease transmission.

Angelica acutiloba Exerts Antihypertensive Effect and Improves Insulin Resistance in Spontaneously Hypertensive Rats Fed with a High-Fat Diet.

INTRODUCTION: Angelica acutiloba is one of the crude drugs used in Chinese herbal medicine, and its intake is expected to improve metabolic syndrome-associated disorders. Here, we examined the effects of A. acutiloba extract (AAE) on hypertension and insulin resistance induced by the treatment of high-fat diet (HFD) to spontaneously hypertensive rats (SHRs). Then, we investigated the mechanisms associated with the effects of AAE. METHODS: AAE was administered to HFD-fed SHRs. Systolic blood pressure (SBP), sympathetic nerve activity, hypothalamic angiotensin-converting enzyme (ACE) activity, blood glucose level, plasma insulin concentration, visceral fat mass, and gene expression of tumor necrosis factor-alpha (TNF-α) in the visceral fat were evaluated. RESULTS: AAE reduced the increases in SBP and hypothalamic ACE activity observed in the HFD-fed SHRs, whereas the suppressive effect on sympathetic nerve activity was slight. Environmental stress-induced pressure and sympathetic overactivity were suppressed by the treatment of AAE. It also decreased the increase in the blood glucose level, plasma insulin concentration, homeostasis model assessment for the insulin resistance, and TNF-α gene expression in the visceral fat, but not the increase in the visceral fat mass. CONCLUSION: AAE has an antihypertensive effect, suppresses stress-induced hypertension, and improves insulin resistance in HFD-fed SHRs. The suppression of brain ACE activity, sympathetic nerve activity, and inflammation are partly involved in the effects of AAE.

A Glucagon-like Peptide 1 Analog Protects Mitochondria and Attenuates Hypoxia-Reoxygenation Injury in Cultured Cardiomyocytes.

ABSTRACT: Glucagon-like peptide 1 (GLP-1) analogs improve glycemic control in diabetes and protect the heart against ischemia-reperfusion injury. However, the mechanisms underlying this protection remain unclear. Mitochondria are essential for myocyte homeostasis. Therefore, we herein examined the effects of a GLP-1 analog on mitochondria after the hypoxia-reoxygenation of rat neonatal cultured cardiomyocytes. Cardiomyocytes were subjected to hypoxia for 5 hours followed by reoxygenation for 30 minutes in the presence or absence of exendin-4 (50 nmol/L), a GLP-1 analog. Hypoxia-reoxygenation increased lactate dehydrogenase and caspase-3 activities, indicators of lethal myocyte injury and apoptosis, respectively, and exendin-4 attenuated these increases. The content of ATP in myocytes decreased after hypoxia-reoxygenation but was preserved by exendin-4. The membrane potential and shape of mitochondria were assessed using a fluorescent probe. Exendin-4 attenuated the hypoxia-reoxygenation-induced disruption of the mitochondrial membrane potential and shortening. Mitochondrial quality control-related factors, such as optic atrophy protein 1, mitofusin 2, dynamin-related protein 1, and parkin, were examined by Western blotting. Exendin-4 significantly increased the expression of the fusion proteins, optic atrophy protein 1 and mitofusin 2, and decreased that of the mitophagy-related protein, parkin, without altering dynamin-related protein 1 expression levels. Exendin-4 also preserved Akt phosphorylation levels after hypoxia-reoxygenation, whereas wortmannin, an inhibitor of the phosphoinositide 3-kinase-Akt pathway, blunted exendin-4-induced myocyte protection and its effects on mitochondrial quality control factors. In conclusion, exendin-4 protected mitochondria by preserving the phosphorylation of Akt and fusion proteins, leading to the attenuation of hypoxia-reoxygenation-induced injury in cultured myocytes.

Secreted protein acidic and rich in cysteine (SPARC) and a disintegrin and metalloproteinase with thrombospondin type 1 motif (ADAMTS1) increments by the renin-angiotensin system induce renal fibrosis in deoxycorticosterone acetate-salt hypertensive rats.

Secreted protein acidic and rich in cysteine (SPARC), an extracellular matrix (ECM) protein, was recently shown to induce collagen deposition through the production of a disintegrin and metalloproteinase with thrombospondin type 1 motif (ADAMTS1) in the aging heart. ADAMTS1 regulates ECM turnover by degrading ECM components, and its excessive activation contributes to various pathological states, including fibrosis. The present study investigated the pathophysiological regulation and role of SPARC and ADAMTS1 in renal fibrosis using uninephrectomized rats treated with deoxycorticosterone acetate (DOCA, 40 mg/kg/week, subcutaneously) and salt (1% in drinking water). The administration of DOCA and salt gradually and significantly elevated systolic blood pressure during the 3-week treatment period, induced proteinuria, decreased creatinine clearance, and increased NADPH oxidase-derived superoxide production, malondialdehyde concentrations, and monocyte chemoattractant protein-1 and osteopontin expression in the kidneys. Glomerulosclerosis, fibrillar collagen deposition, and transforming growth factor-ß expression increased in a time-dependent manner, and SPARC and ADAMTS1 expression showed a similar pattern to these changes. The angiotensin II type-1 receptor blocker losartan suppressed the overexpression of SPARC and ADAMTS1, and an in vitro exposure to angiotensin II induced the production of both SPARC and ADAMTS1 in renal fibroblast NRK-49F cells. Knockdown of the SPARC gene with small interfering RNA reduced all forms (the 110-kDa latent and 87- and 65-kDa bioactive forms) of ADAMTS1 expression as well as collagen production. These results suggest that SPARC is induced by the renin-angiotensin system and may be a fibrogenic factor, at least in part, by producing ADAMTS1 in hypertensive renal disease.

Raman Imaging of Pathogenic Candida auris: Visualization of Structural Characteristics and Machine-Learning Identification.

Invasive fungal infections caused by yeasts of the genus Candida carry high morbidity and cause systemic infections with high mortality rate in both immunocompetent and immunosuppressed patients. Resistance rates against antifungal drugs vary among Candida species, the most concerning specie being Candida auris, which exhibits resistance to all major classes of available antifungal drugs. The presently available identification methods for Candida species face a severe trade-off between testing speed and accuracy. Here, we propose and validate a machine-learning approach adapted to Raman spectroscopy as a rapid, precise, and labor-efficient method of clinical microbiology for C. auris identification and drug efficacy assessments. This paper demonstrates that the combination of Raman spectroscopy and machine learning analyses can provide an insightful and flexible mycology diagnostic tool, easily applicable on-site in the clinical environment.

Induction of autophagy has protective roles in imatinib-induced cardiotoxicity.

Cardiotoxicity is one of the severe adverse effects of chemotherapeutic agents. Imatinib was previously reported to induce cardiotoxicity. Autophagy is an intracellular bulk protein and organelle degradation process, but its roles in cardiac diseases are unclear. We examined whether imatinib induces cardiomyocyte autophagy, and the role of autophagy in imatinib-induced cardiotoxicity using in vitro and in vivo experiments. In in vitro experiments, neonatal rat cardiomyocytes were treated with imatinib (1, 5, or 10 µM; 6 h). Myocyte autophagy was assessed by microtubule-associated protein light chain (LC) 3-II, beclin 1, mature cathepsin D, and acridine orange-stained mature autolysosome expression. Imatinib increased their expression, suggesting that it induced autophagy. Consequently, imatinib altered the production of mitochondria-derived reactive oxygen species (ROS) and loss of mitochondrial membrane potential, which were assessed by the fluorescent indicator MitoSOX and JC-1, respectively, leading to cardiomyocyte apoptosis. 3-methyl-adenine (3MA), an autophagic inhibitor, exacerbated imatinib-induced apoptosis by 30 %. In in vivo experiments, C57BL/6 mice were treated with imatinib (50 and 200 mg/kg/day) for 5 weeks in the presence or absence of 3MA. Echocardiographic measurement revealed that imatinib (200 mg) caused dilatation of the left ventricle (LV) and reduced LV fractional shortening. Apoptosis and LC3-II expression in cardiac tissue were increased by imatinib. Co-treatment with 3MA and imatinib further impaired imatinib-induced cardiac apoptosis and LV dysfunction. This study suggests that imatinib induces cardiomyocyte apoptosis, leading to cardiac dysfunction. Imatinib increases cardiomyocyte autophagy as a consequence of apoptosis and autophagy has a pro-survival role in imatinib-induced cardiac impairment.

Preconditioning with Short-term Dietary Restriction Attenuates Cardiac Oxidative Stress and Hypertrophy Induced by Chronic Pressure Overload.

Left ventricular (LV) hypertrophy and associated heart failure are becoming a more prevalent and critical public health issue with the aging of society, and are exacerbated by reactive oxygen species (ROS). Dietary restriction (DR) markedly inhibits senescent changes; however, prolonged DR is difficult. We herein investigated whether preconditioning with short-term DR attenuates chronic pressure overload-induced cardiac hypertrophy and associated oxidative stress. Male c57BL6 mice were randomly divided into an ad libitum (AL) diet or 40% restricted diet (DR preconditioning, DRPC) group for 2 weeks prior to ascending aortic constriction (AAC), and all mice were fed ad libitum after AAC surgery. Two weeks after surgery, pressure overload by AAC increased LV wall thickness in association with LV diastolic dysfunction and promoted myocyte hypertrophy and cardiac fibrosis in the AL+AAC group. Oxidative stress in cardiac tissue and mitochondria also increased in the AL+AAC group in association with increments in cardiac NADPH oxidase-derived and mitochondrial ROS production. LV hypertrophy and associated cardiac dysfunction and oxidative stress were significantly attenuated in the DRPC+AAC group. Moreover, less severe mitochondrial oxidative damage in the DRPC+AAC group was associated with the suppression of mitochondrial permeability transition and cardiac apoptosis. These results indicate that chronic pressure overload-induced cardiac hypertrophy in association with cardiac and mitochondrial oxidative damage were attenuated by preconditioning with short-term DR.

Febuxostat Attenuates the Progression of Periodontitis in Rats.

INTRODUCTION: Periodontitis is a lifestyle-related disease that is characterized by chronic inflammation in gingival tissue. Febuxostat, a xanthine oxidase inhibitor, exerts anti-inflammatory and antioxidant effects. OBJECTIVE: The present study investigated the effects of febuxostat on periodontitis in a rat model. METHODS: Male Wistar rats were divided into 3 groups: control, periodontitis, and febuxostat-treated periodontitis groups. Periodontitis was induced by placing a ligature wire around the 2nd maxillary molar and the administration of febuxostat (5 mg/kg/day) was then initiated. After 4 weeks, alveolar bone loss was assessed by micro-computed tomography and methylene blue staining. The expression of osteoprotegerin (OPG), a bone resorption inhibitor, was detected by quantitative RT-PCR and immunological staining, and the number of osteoclasts in gingival tissue was assessed by tartrate-resistant acid phosphatase staining. The mRNA and protein expression levels of the proinflammatory cytokines, tumor necrosis factor-alpha (TNF-α), and interleukin-1 beta (IL-1ß), in gingival tissue were measured using quantitative RT-PCR and immunological staining. Oxidative stress in gingival tissue was evaluated by the expression of 4-hydroxy-2-nonenal (4-HNE), and 8-hydroxy-2-deoxyguanosine (8-OHdG). To clarify the systemic effects of periodontitis, blood pressure and glucose tolerance were examined. RESULTS: In rats with periodontitis, alveolar bone resorption was associated with reductions in OPG and increases in osteoclast numbers. The gingival expression of TNF-α, IL-1ß, 4-HNE, and 8-OHdG was up-regulated in rats with periodontitis. Febuxostat significantly reduced alveolar bone loss, proinflammatory cytokine levels, and oxidative stress. It also attenuated periodontitis-induced glucose intolerance and blood pressure elevations. CONCLUSION: Febuxostat prevented the progression of periodontitis and associated systemic effects by inhibiting proinflammatory mediators and oxidative stress.

Comparison of effects of L/N-type and L-type calcium channel blockers on post-infarct cardiac remodelling in spontaneously hypertensive rats.

Hypertension and coronary events are becoming more prevalent in aging societies, and myocardial infarction usually occurs in calcium channel blocker (CCB)-treated hypertensive patients. We herein compared the effects of cilnidipine, an L/N-type CCB and amlodipine, an L-type CCB, on post-infarct left ventricular (LV) remodelling in spontaneously hypertensive rats (SHRs). Male SHRs were subjected to 30 minutes of left coronary artery occlusion followed by reperfusion (MI group). The administration of cilnidipine (10 mg/kg/d; MI + Cil group) or amlodipine (10 mg/kg/d; MI + Aml group) was initiated one week before surgery and continued for five weeks. Both CCBs decreased blood pressure. Four weeks after surgery, cilnidipine, but not amlodipine, attenuated LV dilatation, fractional shortening impairments, end-diastolic pressure elevations, and tau elongation. In the non-infarct region, myocyte hypertrophy and brain natriuretic peptide (BNP) mRNA levels were similarly attenuated by both CCBs. On the other hand, interstitial fibrosis, the mRNA expression of collagen type III and transforming growth factor (TGF) ß and immunohistological TGF ß protein expression in the non-infarct region were reduced more in the MI + Cil group than in the MI + Aml group. Additionally, elevated angiotensin-converting enzyme activity and interstitial noradrenaline concentrations in the non-infarct region were reduced by cilnidipine. These results suggest that cilnidipine reduced cardiac noradrenaline concentrations and inhibited the renin-angiotensin system, which attenuated post-infarct remodelling more than amlodipine in hypertensive rats.

Antihypertensive and Renoprotective Effects of Dietary Flaxseed and its Mechanism of Action in Deoxycorticosterone Acetate-Salt Hypertensive Rats.

BACKGROUND/AIMS: Flaxseed contains alpha-linolenic acid (ALA), lignans, and dietary fiber, and its intake lowers blood pressure in hypertensive patients. Here, we examined the effects of flaxseed powder, which includes all flaxseed components, flaxseed oil, composed mainly of ALA, flaxseed lignan, and flaxseed fiber, on hypertension and renal damage induced by deoxycorticosterone acetate (DOCA)-salt. Then, we investigated the mechanisms of action associated with the effects of flaxseed. METHODS: Flaxseed powder, oil, lignan, or fiber was administered to DOCA-salt rats. Systolic blood pressure (SBP), urinary protein excretion, renal angiotensin converting enzyme (ACE) activity, sympathetic nerve activity, and gene expression of inflammatory mediators in the kidney and hypothalamus were measured. RESULTS: Flaxseed powder and oil reduced the increases in SBP and urinary protein excretion induced by DOCA-salt treatment, whereas lignan and fiber had no effects. Flaxseed oil suppressed the increase in renal ACE activity, sympathetic nerve activity, and gene expression of renal and hypothalamic inflammatory mediators. CONCLUSION: Flaxseed has antihypertensive and renoprotective effects in DOCA-salt rats. These effects are likely principally exerted by ALA. Furthermore, the suppression of renal ACE activity, sympathetic nerve activity, and inflammation is partly involved in the effects of flaxseed.

Transiently proliferating perivascular microglia harbor M1 type and precede cerebrovascular changes in a chronic hypertension model.

BACKGROUND: Microglia play crucial roles in the maintenance of brain homeostasis. Activated microglia show a biphasic influence, promoting beneficial repair and causing harmful damage via M2 and M1 microglia, respectively. It is well-known that microglia are initially activated to the M2 state and subsequently switch to the M1 state, called M2-to-M1 class switching in acute ischemic models. However, the activation process of microglia in chronic and sporadic hypertension remains poorly understood. We aimed to clarify the process using a chronic hypertension model, the deoxycorticosterone acetate (DOCA)-salt-treated Wistar rats. METHODS: After unilateral nephrectomy, the rats were randomly divided into DOCA-salt, placebo, and control groups. DOCA-salt rats received a weekly subcutaneous injection of DOCA (40 mg/kg) and were continuously provided with 1% NaCl in drinking water. Placebo rats received a weekly subcutaneous injection of vehicle and were provided with tap water. Control rats received no administration of DOCA or NaCl. To investigate the temporal expression profiles of M1- and M2-specific markers for microglia, the animals were subjected to the immunohistochemical and biochemical studies after 2, 3, or 4 weeks DOCA-salt treatment. RESULTS: Hypertension occurred after 2 weeks of DOCA and salt administration, when round-shaped microglia with slightly shortened processes were observed juxtaposed to the vessels, although the histopathological findings were normal. After 3 weeks of DOCA and salt administration, M1-state perivascular and parenchyma microglia significantly increased, when local histopathological findings began to be observed but cerebrovascular destruction did not occur. On the other hand, M2-state microglia were never observed around the vessels at this period. Interestingly, prior to M1 activation, about 55% of perivascular microglia transiently expressed Ki-67, one of the cell proliferation markers. CONCLUSIONS: We concluded that the resting perivascular microglia directly switched to the pro-inflammatory M1 state via a transient proliferative state in DOCA-salt rats. Our results suggest that the activation machinery of microglia in chronic hypertension differs from acute ischemic models. Proliferative microglia are possible initial key players in the development of hypertension-induced cerebral vessel damage. Fine-tuning of microglia proliferation and activation could constitute an innovative therapeutic strategy to prevent its development.

Pitavastatin suppresses hyperglycaemia-induced podocyte injury via bone morphogenetic protein-7 preservation.

Podocytes form the essential components of the glomerular filtration barrier and play a critical role in diabetic nephropathy. Recent evidence suggests that HMG-CoA reductase inhibitors (statins) exert renoprotective effects. We investigated whether pitavastatin directly suppresses hyperglycaemia-induced podocyte injury using cultured podocytes and, if so, the mechanism of the beneficial effects. Cultured podocytes were exposed to media containing normal (NG; 5 mmol/L) or high (HG; 25 mmol/L) glucose for 1 week. HG increased the lethal injury of podocytes and disruption of F-actin fibers, and reduced the mRNA expression of novel podocyte markers, synaptopodin and Wilms tumor-1 (WT-1), in association with decreased bone morphogenetic protein-7 (BMP-7) expression. Pitavastatin (100 nmol/L) reduced podocyte injury and restored the mRNA expression of synaptopodin and WT1; however, these protective effects were abolished by BMP-7 siRNA. Additionally, pitavastatin suppressed HG-induced Rho kinase activation, as assessed by the phosphorylation level of myosin phosphatase targeting subunit 1 (MYTP1), and C3 exotoxin, a Rho inhibitor, mimicked the effect of pitavastatin on BMP-7 preservation. Pitavastatin attenuates hyperglycaemia-induced podocyte injury via Rho-Rho kinase-dependent BMP-7 preservation.

Pitavastatin Exhibits Protective Effects on Podocytes Accompanied by BMP-7 Up-Regulation and Rho Suppression.

BACKGROUND/AIMS: Podocytes injury is involved in the development of diabetic nephropathy. This study was designed to confirm the reno- and podocyte-protective effects of pitavastatin in diabetic rats and clarify its mechanisms. METHODS: Wistar rats were divided into 4 treatment groups: control, streptozotocin (STZ; 55 mg/kg)-induced diabetes, STZ with pitavastatin (10 mg/kg/day), and STZ with tempol (1 mmol/l). RESULTS: STZ-induced diabetic rats exhibited increases in urinary protein excretion and plasma creatinine, and a decrease in creatinine clearance. Pitavastatin significantly improved these parameters without reducing cholesterol levels, whereas tempol did not. The treatment with STZ-enhanced renal fibrosis, mesangial proliferation, transforming growth factor (TGF)-ß, MCP-1 and suppressed Rho in association with decrement of bone morphogenetic protein (BMP)-7 expression in renal cortex. Moreover, STZ decreased podocyte related factors, podocin and nephrin, and BMP-7 in podocytes. Pitavastatin significantly ameliorated all these indices. On the other hand, improvement by tempol was found only in TGF-ß, MCP-1 and histological changes. CONCLUSION: Pitavastatin exhibited reno- and podocyte-protective effects accompanied by BMP-7 preservation and Rho suppression.

Short-Term Caloric Restriction Suppresses Cardiac Oxidative Stress and Hypertrophy Caused by Chronic Pressure Overload.

BACKGROUND: Caloric restriction (CR) prevents senescent changes, in which reactive oxygen species (ROS) have a critical role. Left ventricular (LV) hypertrophy is a risk factor for cardiovascular diseases. We examined whether CR alters cardiac redox state and hypertrophy from chronic pressure overload. METHODS AND RESULTS: Male c57BL6 mice were subjected to ascending aortic constriction (AAC) with ad libitum caloric intake (AL + AAC group) or 40% restricted caloric intake (CR + AAC group). CR was initiated 2 weeks before AAC and was continued for 4 weeks. Two weeks after constriction, AAC increased LV wall thickness, impaired transmitral flow velocity, and augmented myocyte hypertrophy and fibrosis, in association with enhancement of BNP and collagen III expressions in the AL + AAC group. In the AL + AAC group, oxidative stress in cardiac tissue and mitochondria were enhanced, and NADPH oxidase activity and mitochondrial ROS production were elevated. These changes were significantly attenuated in the CR + AAC group. Additionally, in antioxidant systems, myocardial glutathione peroxidase and superoxide dismutase activities were enhanced in the CR + AAC group. CONCLUSIONS: Chronic pressure overload increased cardiac oxidative damage, in association with cardiac hypertrophy and fibrosis. Short-term CR suppressed oxidative stress and improved cardiac function, suggesting that short-term CR could be a useful strategy to prevent pressure overload-induced cardiac injury.