High-Fat Diet-Induced Diabetic Conditions Exacerbate Cognitive Impairment in a Mouse Model of Alzheimer's Disease Via a Specific Tau Phosphorylation Pattern.

BACKGROUND: Epidemiological evidence has demonstrated a clear association between diabetes mellitus and increased risk of Alzheimer's disease (AD). Cerebral accumulation of phosphorylated tau aggregates, a cardinal neuropathological feature of AD, is associated with neurodegeneration and cognitive decline. Clinical and experimental studies indicate that diabetes mellitus affects the development of tau pathology; however, the underlying molecular mechanisms remain unknown. OBJECTIVE: In the present study, we used a unique diabetic AD mouse model to investigate the changes in tau phosphorylation patterns occurring in the diabetic brain. DESIGN: Tau-transgenic mice were fed a high-fat diet (n = 24) to model diabetes mellitus. These mice developed prominent obesity, severe insulin resistance, and mild hyperglycemia, which led to early-onset neurodegeneration and behavioral impairment associated with the accumulation of hyperphosphorylated tau aggregates. RESULTS: Comprehensive phosphoproteomic analysis revealed a unique tau phosphorylation signature in the brains of mice with diabetic AD. Bioinformatic analysis of the phosphoproteomics data revealed putative tau-related kinases and cell signaling pathways involved in the interaction between diabetes mellitus and AD. CONCLUSION: These findings offer potential novel targets that can be used to develop tau-based therapies and biomarkers for use in AD.

Diffusion Tensor Imaging MRI With Spin-Echo Sequence and Long-Duration Measurement for Evaluation of Renal Fibrosis in a Rat Fibrosis Model.

BACKGROUND: Renal fibrosis (RF) is a well-known marker for chronic kidney disease (CKD) progression, including chronic renal injury after renal transplantation. However, invasive biopsy is an available examination for evaluation of RF. Diffusion MRI was once recognized as a promising option for RF. However, it is now controversial for RF evaluation in a unilateral ureteral obstruction (UUO) model. METHODS: To seek an optimal imaging method applicable for RF in UUO model kidneys, we attempted a series of MRI methods, including proton density-weighted imaging, T1-weighted imaging, T2-weighted imaging, T2*-weighted imaging, diffusion-weighted imaging, and diffusion tensor imaging (DTI). RESULTS: We identified DTI MRI by spin-echo sequence plus a special kidney attachment as the best option for evaluation of renal UUO fibrosis, compared with normal kidney on the opposite side. To confirm these results, we applied this technique to a rat UUO therapeutic model with the anti-fibrotic reagent Fasudil. Fractional anisotropy values calculated from DTI MRI showed statistically significant linear correlation with the RF area measured by use of Sirius red or Masson trichrome staining of the positive area [cortex (r = 0.6397, P = .0283) and outer stripe of the outer medulla (r = 0.7810, P = .0039)]. CONCLUSIONS: By use of the DTI MRI with spin-echo sequence, it may be possible to accurately evaluate RF in CKD.

Novel microdialysis method to assess neuropeptides and large molecules in free-moving mouse.

Microdialysis is a powerful in vivo technique for the continuous sampling of small molecules within the extracellular fluid space. However, efforts to collect larger molecules have met with little success. To identify biologically active larger molecules in free-moving animals would be of great benefit. For this purpose, we have developed a novel microdialysis method that allows consistent recovery of large molecules from the brain interstitial space in the awake, free-moving mouse. Using a new "vent" probe with a push-pull perfusion system, the present study successfully demonstrated in vivo sampling of pathophysiologically important macromolecules in free-moving mouse brain. This sampling system allowed monitoring of the dynamic changes in their concentrations. Overall, this novel microdialysis system would provide the opportunity to identify the expression patterns of pathophysiologically important proteins in a variety of physiological and pathological processes for a better understanding of various diseases.

Bcl-2 protects tubular epithelial cells from ischemia/reperfusion injury by dual mechanisms.

Ischemia/reperfusion (I/R) injury, which induces extensive loss of tubular epithelial cells, is associated with delayed graft function following kidney transplantation. Recent reports have suggested that cell death by I/R injury occurs by autophagy, a cellular degradation process responsible for the turnover of unnecessary or dysfunctional organelles and cytoplasmic proteins, as well as by apoptosis. Recently, we demonstrated that overexpression of the anti-apoptotic factor, Bcl-2, inhibited tubular apoptosis and subsequent tubulointerstitial damage after I/R injury. Autophagy is also observed in cells undergoing cell death in several diseases. Therefore, we hypothesized that increased Bcl-2 protein may protect tubular epithelial cells by suppressing autophagy and inhibiting apoptosis. In the present study, a transgenic mouse model (LC3-GFP TG) in which autophagosomes are labeled with LC3-GFP and Bcl-2/LC3-GFP double transgenic mice (Bcl-2/LC3-GFP TG) were used to examine the effect of Bcl-2 on I/R-induced autophagy. I/R injury, which is associated with marked disruption of normal tubular morphology, promoted the formation of LC3-GFP dots, representing extensively induced autophagosomes. On electron microscopy, the autophagosomes contained mitochondria in I/R-injured tubular epithelial cells. In contrast, Bcl-2 augmentation suppressed the formation of autophagosomes and there was less tubular damage. In conclusion, Bcl-2 augmentation protected renal tubular epithelial cells from I/R injury by suppressing autophagosomal degradation and inhibiting tubular apoptosis.

Aspirin: recent developments.

Aspirin exerts anti-thrombotic action by acetylating and inactivating cyclooxygenase-1, preventing the production of thromboxane A2 in platelets. Through this inhibition of platelet function, aspirin is considered as a preventative of ischemic diseases such as coronary and cerebral infarction. However, many studies have revealed that aspirin has other beneficial actions in addition to its anti-platelet activity. For example, aspirin may confer some benefit against colorectal cancer. Here, we discuss the involvement of inflammation in atherosclerosis and how aspirin exerts its beneficial actions in atherosclerotic diseases and cancer.

Roles of TGF-beta1 and apoptosis in the progression of glomerulosclerosis in human IgA nephropathy.

Apoptotic glomerular cells have been detected in the severely damaged glomeruli that are a consequence of human IgA nephropathy. Transforming growth factor-(TGF) beta1 is known to induce apoptosis in cultured mesangial cells. To clarify whether TGF-beta1 contributes to the progression of IgA nephropathy by activating apoptosis in glomerular cells, we examined the expression of TGF-beta1 gene and apoptotic changes in kidney biopsy samples, and assessed those relations to the severity of nephropathy. 32 patients with IgA nephropathy, showing proteinuria (> 1 g/day) and serum creatinine less than 1.5 mg/dl were classified according to glomerular sclerosis index (GSI) into 3 groups (Group I: GSI < 0.3,Group 11: 0.3 < or = GSI < 1.0, Group: III GSI > or = 1.0). Computer-aided morphometry of glomeruli and arteries, and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling of fragmented DNA (TUNEL) staining were performed. Expression of TGF-beta1 and caspase-3 mRNAs in renal biopsy samples was analyzed by real-time PCR (Taq Man method). Increased glomerular area, interstitial fibrosis, lymphocytic infiltration, and tubulointerstitial changes were observed to accompany increased severity of GSI. TUNEL index was higher in Group III. The levels of TGF-beta1 and caspase-3 mRNAs were significantly increased in Group III (183 and 190%, respectively). Furthermore, caspase-3 mRNA levels were tightly associated with TGF-beta1 mRNA expression (r = 0.677, p < 0.0001). The present study suggests that the activation of TGF-beta1 plays a role in the progression of IgA nephropathy even in the moderate degree of glomerular injury, in part via activation of apoptosis of glomerular cells.

Hypertension in a patient with Gitelman's syndrome.

Gitelman's syndrome is an autosomal recessive disorder characterized by sodium wasting and hypotension. A middle-aged woman was diagnosed with Gitelman's syndrome because of typical clinical manifestations in the youth and homozygous mutations of 18-base-pair insertion in exon 6 of thiazide-sensitive NaCl-cotransporter gene. It was unusual that she showed hypertension with advancing age. Her serum potassium levels remained low at around 3.5 mEq/l despite potassium supplementation. This case demonstrates that hypertension could result in spite of the extremely decreased sodium reabsorption in Gitelman's syndrome and that essential hypertension is genetically heterogeneous, and abnormality of all genes may not be necessarily required to cause blood pressure rise.

Additive effects of nicorandil on coronary blood flow during continuous administration of nitroglycerin.

OBJECTIVES: We examined whether patients with ischemic heart disease (IHD) should be treated with nicorandil, an adenosine triphosphate-sensitive potassium channel opener, in addition to the regular use of nitrates. BACKGROUND: It has been reported that nicorandil possibly has additive effects on nitroglycerin (NTG) treatment for angina, but the mechanism is not clear. METHODS: We directly measured anterograde coronary blood flow (CBF) with a Doppler guide wire to examine the effects of intravenous administration of NTG (0.3 mg) and nicorandil (6 mg) during continuous administration of NTG at a sufficient dose (25 microg/min) in subjects with normal and stenotic coronary arteries. RESULTS: Additional systemic administration of NTG decreased anterograde CBF (normal -19.7%; stenotic -21.2%). In contrast, nicorandil increased anterograde CBF in both normal (54.6%) and stenotic (89.6%) coronary arteries, without the coronary steal phenomenon. There was a tendency toward nicorandil-dilated diameters in the patients with stenotic arteries (p = 0.06). There were no effects of additional administration on pulmonary artery wedge pressure. There was no difference in changes in heart rate and mean aortic blood pressure between NTG and nicorandil therapy. CONCLUSIONS: These results suggest that in patients treated with nitrates, additional administration of nicorandil is more useful, in terms of increasing CBF, than additional administration of nitrates. Adjunctive use of nicorandil with nitrates may provide the further benefit of myocardial protection and may improve the prognosis of patients with IHD.

[A case of severe systemic edema in an elderly hypertensive patient with systemic lupus erythematodes during long-term treatment with anti-hypertensive drugs].

A 71-year-old woman receiving both angiotensin II receptor antagonist and calcium antagonist suffered severe systemic edema. She had been treated for essential hypertension with amlodipine for 2 years and candesartan for 3 months, and systemic lupus erythematodes (SLE) with steroids. During treatment, severe systemic edema appeared, mainly on her face, arms, and legs. At first, we suspected drug-induced edema by candesartan, so it was halted, but the edema still continued. We then considered amlodipine to be the culprit, and finally, the severe systemic edema disappeared after cessation of amlodipine. To control her blood pressure, we recommended candesartan, but 3 months late she suffered severe systemic edema again, thus the causative we drugs of her edema were thought to be both amlodipine and candesartan. Edema is a common symptom in elderly patients and we frequently observe drug-induced edema. In this case, there was underlying acceleration of blood vessel permeability induced by SLE and steroids and moreover, vasodilatation by candesartan and/or amlodipine further accelerated blood vessel permeability, and thus might have caused severe edema. It is very difficult to determine the cause of edema, especially in elderly patients, but we should consider not only one but also two or more drugs as being involved in drug-induced edema.

[Review of randomized trials of angiotensin receptor blockers in the treatment of patients with congestive heart failure].

Angiotensin-converting enzyme (ACE) inhibitors are the first-line drugs for the treatment of congestive heart failure, but many patients could not receive those benefits because of intolerance. Recent randomized trials of angiotensin receptor blockers (ARB) in the treatment of patients with congestive heart failure elucidated clinical benefits of ARB as well as ACE inhibitors. ELITE II showed equivalent effect on mortality and morbidity between losartan and captopril and less adverse events in losartan. Val-HeFT showed additive benefits of valsartan on standard treatment with ACE inhibitors, diuretics and digitalis in patients with heart failure. CHARM would be elucidate the clinical usefulness of candesartan cilexetil in a broad spectrum of patients with symptomatic heart failure including patients whose LVEF greater than 40%.

Effects of antihypertensive drugs and gene variants in the renin-angiotensin system.

Many genes and environmental factors are involved in the pathogenesis of hypertension, but the exact cause of essential hypertension has not yet been clarified. Gene polymorphism of the renin-angiotensin system (RAS) is one of the candidates. In the current study, we examined whether there was a correlation between the gene polymorphisms in RAS and either the choice of antihypertensive drugs or their efficacy. Subjects with essential hypertension (n=299) were recruited from among the outpatients of Osaka University Hospital and provided their informed consent for genetic analysis. Physicians freely chose the antihypertensive drugs and adjusted its dose until the patient's blood pressure was well controlled. The efficacy of each antihypertensive drug was estimated using the following formula: ABP=BP 1 (before treatment) - BP 2 (after treatment)/BP 1 x 100 (%). Gene variants in RAS were determined using PCR or PCR-RFLP (restriction fragment of polymorphism). The gene polymorphisms of RAS were not associated with delta SBP or ADBP. However, the mean ASBP in subjects with a deletion homozygote of the angiotensin converting enzyme gene (ACE/DD) was significantly lower (p<0.05) than that in patients with an insertion I allele of the ACE gene. The gene polymorphisms of RAS did not significantly affect the choice of antihypertensive drugs. Even though gene polymorphism in the renin angiotensin system was not a major factor in the antihypertensive therapy, the determination of genotype might be of help in the management of essential hypertension.

Lack of correlation between Mbo I restriction fragment length polymorphism of renin gene and essential hypertension in Japanese.

Predisposition to essential hypertension is associated with gene polymorphisms of the renin angiotensin system (RAS). Gene polymorphisms of the angiotensinogen and angiotensin converting enzyme genes are known to be risk factors for hypertension, while few studies concerning the renin gene polymorphism have been published. In the present investigation, we carried out a case control study using a Japanese population to examine the genetic influence of the renin gene on the predisposition to hypertension. Patients (n=235) recruited from outpatients at Osaka University Hospital and diagnosed with essential hypertension or receiving long-term antihypertensive medication participated in the study. Normotensive control subjects (n=510) without a history of hypertension and without diabetes mellitus were recruited from the same population, and were sex-matched with experimental subjects. A polymorphism in intron 9 of the human renin gene was determined as the Mbo I restriction fragment length polymorphism (Mbo I-RFLP). There was no significant association between Mbo I-RFLP of the renin gene and predisposition to essential hypertension in Japanese (p>0.05, chi2=2.1). These results suggest that the Mbo I (+) allele of the renin gene does not increase the risk for hypertension in Japanese.

A common mutation of low-density lipoprotein receptor gene is associated with essential hypertension among Japanese.

Candidate genes offer one approach to the identification of the genetic susceptibility to hypertension. A common gene variant of the low-density lipoprotein (LDL) receptor gene (LDLR) that affects plasma LDL metabolism within the normolipidaemic range, may be such a candidate gene. A common mutation of LDLR, C1773T, was associated with lipid metabolism such that the T1773 allele increased plasma LDL levels in a Caucasian population. The present study examined whether C1773T/LDLR was associated with essential hypertension in a Japanese population. Subjects with essential hypertension (EHT, n = 300) with a family history of hypertension, and controls (NT, n = 310, sex- and age-matched with EHT) were recruited from among out-patients at Osaka University Hospital. A C1773T substitution at codon 570 in LDLR was determined using PCR-Hinc II-RFLP. It was revealed that the C1773 allele was significantly more frequent (0.89) among hypertensive patients (chi2 = 9.58, P < 0.01) than normotensives (0.83), the calculated odds ratio being 1.7 (95% CI: 1.2-2.4). The effect of the T1773 allele on increasing cholesterol was significant in normotensives without antihyperlipidaemic medication, but not in hypertensives. After adjustments of confounding factors, the estimated odds ratio for hypertension in the subjects with C1773 homozygote increased to 2.1 (95% CI: 1.3-3.5), suggesting that this polymorphism is an independent risk factor for hypertension. Our results show that the C1773 mutant of LDLR increases susceptibility to hypertension, but not via hypercholesterolaemia.

Quantitative ultrasonic tissue characterization can identify high-risk atherosclerotic alteration in human carotid arteries.

BACKGROUND: Recently, ultrasonic tissue characterization of the composition of plaques has been performed in a quantitative fashion on the basis of integrated backscatter (IBS) analysis, but most of those studies have used high-frequency ultrasound to obtain microscopic images. METHODS AND RESULTS: We performed B-mode measurement and IBS signal analysis with acoustic densitometry with a 7.5-MHz linear-array transducer in freshly excised human aortas (n=58) (normal, atheromatous, and fibrous tissue) obtained at autopsy. Atheromatous and fibrous tissue had a similar intima-media thickness (IMT), but the IBS value in atheromatous specimens was lower than that in fibrous specimens. We further applied this method to human carotid ultrasonography. The subjects were young (80 regions), middle aged with 1 or no coronary risk factors (low risk) (120 regions), middle aged with >/=2 coronary risk factors (high risk) (240 regions), or elderly (80 regions) or were patients with myocardial infarction (MI) with multivessel disease (90 regions). The IMT was similar in middle-aged, elderly, and MI subjects. In contrast, the IBS value was significantly higher in elderly subjects and lower in high-risk middle-aged and MI subjects compared with that in low-risk middle-aged subjects. The percent of regions diagnosed as atheromatous (IBS less than mean minus 2-SD value of IBS in young subjects) was 11% in low-risk middle-aged subjects, 29% in high-risk middle-aged subjects, and 63% in the MI group. CONCLUSIONS: In conjunction with conventional B-mode imaging, IBS analysis with carotid ultrasonography appeared to provide prognostic information to identify a high-risk group with systemic atherosclerosis, which could lead to coronary heart disease in individuals with early-stage disease.

Deletion polymorphism of angiotensin-converting enzyme gene is associated with postprandial hyperglycaemia in individuals undergoing general check-up.

1. Deletion polymorphism, DD, of the angiotensin-converting enzyme (ACE) gene is reported to be related to cardiovascular disease, which is frequently based on insulin resistance. 2. To clarify the relationship between the ACE genotype DD and plasma glucose increases after an oral glucose load, we performed 75 g oral glucose tolerance test (OGTT) in 301 nondiabetic men (age range 30-60 years) undergoing general check-up. 3. Insertion/deletion (I/D) polymorphism of the ACE gene was explored using a polymerase chain reaction. The frequency of the II, ID and DD genotypes was 0.43, 0.43 and 0.14, respectively. 4. There were no differences in baseline clinical characteristics between subjects with each ACE genotype. 5. The mean (+/- SEM) plasma glucose level at 60 min of the OGTT was significantly higher in subjects with the DD genotype (170.8 +/- 6.9 mg/dL) than in subjects with either the II or ID genotype (mean value for two groups 156.6 +/- 2.7 mg/dL; P < 0.05). Moreover, the mean percentage change of plasma glucose after 60 min of the OGTT, a marker of plasma glucose increase, was significantly higher in individuals with the DD genotype than in individuals with either the II or ID genotypes. 6. In contrast, the mean fasting plasma glucose level, the plasma glucose level at 120 min, the glucose response area and the fasting insulin level were not different between individuals with the DD genotype and individuals with other genotypes. 7. In conclusion, subjects with the DD genotype showed transiently higher levels of plasma glucose after an oral glucose load than subjects with other genotypes. Further studies are required to determine whether the association between ACE genotype and postprandial hyperglycaemia influences the incidence of cardiovascular disease and diabetes mellitus.