Comparative Analysis Between Paramecium Strains with Different Syngens Using the RAPD Method.

Paramecium spp. are a genus of free-living protists that live mainly in freshwater environments. They are ciliates with high motility and phagocytosis and have been used to analyze cell motility and as a host model for pathogens. Besides such biological characteristics, apart from the usual morphological and genetic classification of species, the existence of taxonomies (such as syngens) and mating types related to Paramecium's unique reproduction is known. In this study, we attempted to develop a simple method to identify Paramecium strains, which are difficult to distinguish morphologically, using random amplified polymorphic DNA (RAPD) analysis. Consequently, we can observe strain-specific band patterns. We also confirm that the presence of endosymbiotic Chlorella cells affects the band pattern of P. bursaria. Furthermore, the results of the RAPD analysis using several P. caudatum strains with different syngens show that it is possible to detect a band specific to a certain syngen. By improving the reaction conditions and random primers, based on the results of this study, RAPD analysis can be applied to the identification of Paramecium strains and their syngen confirmation tests.

Takotsubo cardiomyopathy after vaccination for coronavirus disease 2019 in a patient on maintenance hemodialysis.

Coronavirus disease-2019 (COVID-19) has affected more than 220 million individuals since the global pandemic began. There is an urgent need for safe and effective vaccines, and vaccinations, such as mRNA vaccines, have been initiated worldwide. However, the adverse effects of these vaccines remain unclear. We herein present a case of an 80-year-old female on maintenance hemodialysis who developed takotsubo cardiomyopathy 4 days after receiving the first dose of the Pfizer-BioNTech COVID-19 vaccine. There was no obvious trigger for the onset of takotsubo cardiomyopathy other than the COVID-19 vaccination, which was the most significant event preceding her presentation. Echocardiograms obtained during her admission allowed us to monitor and show the recovery of left ventricular wall motion. We confirmed the diagnosis of takotsubo cardiomyopathy based on the findings, including transient left ventricular dysfunction, electrocardiographic abnormalities, an elevated troponin level, and the absence of occlusive coronary artery disease. In the present case, the vaccination may have triggered emotional or physical stress. Although difficulties are associated with proving the causal relationship in the present case, the temporal relationship between the vaccination and the onset of takotsubo cardiomyopathy is highly suggestive. The adverse effects associated with the vaccine are typical of COVID-19 vaccines administered to date, most of which are acceptable. Therefore, despite our experience of the present case, we still recommend the vaccination for COVID-19 because takotsubo cardiomyopathy induced by the COVID-19 vaccine is extremely rare and the prognosis of the patient was good. We herein present the first case of a patient on hemodialysis who developed takotsubo cardiomyopathy after receiving COVID-19 vaccination.

Effect of advanced periodontal self-care in patients with early-stage periodontal diseases on endothelial function: An open-label, randomized controlled trial.

Although a significant association between periodontal disease and atherosclerotic cardiovascular disease has been reported, their cause-to-effect relationship remains controversial. This randomized controlled clinical trial aimed to investigate the effect of advanced self-care on atherosclerotic cardiovascular disease-related vascular function markers flow-mediated brachial artery dilatation (FMD) and serum asymmetric dimethylarginine (ADMA) level in patients with early-stage periodontal disease. The study was designed as a parallel group, 3-month follow-up, open-label, randomized controlled trial. The control group received standard care for periodontal diseases, whereas the test group additionally applied disinfectant using a custom-fabricated prescription tray for advanced self-care twice a day. Overall, 110 patients provided data for FMD and serum ADMA level. No significant improvements in FMD were observed in the control (mean increase, -0.1%; 95% confidence interval [CI], -1.0-0.8; P = 0.805) or test (mean increase, -0.3%; 95% CI, -1.1-0.4; P = 0.398) group. No significant changes in serum ADMA levels were observed (mean reduction, 0.01 µmol/L; 95% CI, -0.00-0.02; P = 0.366 and mean reduction, 0.00 µmol/L; 95% CI, -0.01-0.01; P = 0.349, respectively). No significant between-group differences were found in FMD (mean difference, -0.2%; 95% CI, -1.4-0.9; p = 0.708) or serum ADMA levels (mean difference, 0.01 nmol/L; 95% CI, -0.00-0.03; p = 0.122). Significant improvements in the average probing pocket depth were observed in the control and test groups. The bleeding on probing score in the test group was significantly reduced, while that in the control group was reduced, although not significantly. Periodontal care for a 3-month duration did not provide better endothelial function although improvements of periodontal status in patients with early-stage periodontal diseases. This trial is registered in UMIN Clinical Trials Registry (www.umin.ac.jp/ctr/; ID: UMIN000023395).

Diversity of pathological features other than Lewy bodies in familial Parkinson's disease due to SNCA mutations.

The clinical features of the genetically determined forms of familial Parkinson's disease (PD) have been described in multiple reports, but there have been few comparative neuropathologic studies. Five familial PD cases, with mutations in SNCA, were matched for age, sex, and Alzheimer type pathology with sporadic PD cases. Immunohistochemistry for phospho-tau and α-synuclein was performed in 8 brain regions. The frequency of tau pathology and the morphologic features of α-synuclein pathology in familial PD were compared with sporadic PD using semi-quantitative methods. In familial PD, there were significantly more tau positive extra-perikaryal spheroid-like and thread-like lesions than in the sporadic PD. There was no significant difference in the amount of α-synuclein positive neuronal perikaryal pathology between familial PD and sporadic PD, but α-synuclein positive oligodendroglial and neuritic lesions were significantly greater in familial PD compared to sporadic PD. In the substantia nigra, familial PD had more marked neuronal loss and fewer residential neurons with Lewy bodies than the sporadic PD, suggesting a close relationship between the severity of neuronal loss and Lewy body formation. The results show a diversity of pathological features of genetically determined familial PD, and they draw attention to the possible role of tau protein in neurodegeneration. Moreover, the presence of oligodendroglial inclusions at the light and electron microscopic levels in familial PD suggests that PD and multiple system atrophy form a continuum of α-synuclein pathology.

High serum concentrations of pentosidine, an advanced glycation end product, are associated with low normal value of ankle-brachial index in apparently healthy men.

The ankle-brachial index (ABI) is widely used for peripheral arterial disease screening and is associated with future cardiovascular events. Pentosidine, an advanced glycation end product, accumulates with age and in diabetes and end-stage renal disease; but the significance of elevated serum pentosidine is not well known. We investigated the relationship of the ABI to circulating pentosidine concentrations as well as other atherogenic factors in apparently healthy men. The study group consisted of a total of 170 apparently healthy men (age, 55 ± 9 years). Serum pentosidine concentrations were measured by enzyme-linked immunosorbent assay. The mean ABI and pentosidine concentrations of the whole study group were 1.16 ± 0.07 (range, 0.98-1.35) and 36.1 ± 10.6 ng/mL (range, 11.2-81.0), respectively. Univariate analysis showed that the ABI was inversely correlated with pentosidine (P = .0004), small low-density lipoprotein (LDL) cholesterol (P = .017), LDL cholesterol (P = .019), apolipoprotein B (P = .024), fasting insulin (P = .028), very small LDL cholesterol (P = .036), difference in ABIs between legs (P = .037), malondialdehyde-modified LDL (P = .044), and homeostasis model assessment of insulin resistance (P = .047). Stepwise multiple linear regression analysis revealed that increased pentosidine, fasting insulin, small LDL cholesterol, difference in ABIs between legs, difference in systolic blood pressure between arms, and reduced body mass index were independent determinants of reduced ABI (adjusted R(2) = 0.237, P < .0001). Serum pentosidine was an important, independent determinant of ABI in healthy men. Subjects with an ABI less than 1.10 showed higher pentosidine concentrations.

Very small low-density lipoprotein cholesterol level is a determinant of arterial stiffness in men with impaired glucose metabolism.

AIM: Pulse wave velocity (PWV), an estimate of arterial stiffness, is an important predictor of cardiovascular risk. The aim of this study was to investigate the relationship between lipoprotein subclasses and brachial-ankle PWV (baPWV). METHODS: A total of 131 apparently healthy Japanese men without a history of cardiovascular disease were divided into two groups: normal glucose metabolism (n =87) and impaired glucose metabolism (n =44). Cholesterol concentrations of major lipoproteins and their subclasses were determined by HPLC with gel permeation columns. RESULTS: In the normal glucose metabolism group, age, systolic blood pressure, and diastolic pressure were associated with increased baPWV, and a stepwise multiple linear regression analysis revealed that age (p =0.022) and systolic blood pressure (p < 0.001) were significantly independent determinants of baPWV. In the impaired glucose metabolism group, age (p =0.002), very small LDL cholesterol (p =0.012), systolic blood pressure (p =0.021), and the fasting plasma glucose concentration (p =0.038) were identified as independent determinants of baPWV, although a univariate analysis revealed significant relationships of several plasma lipid compositions or species to baPWV. CONCLUSIONS: In addition to aging, hypertension and glucose levels, very small LDL cholesterol levels appear to play an important role in the development of arterial stiffness in men with impaired glucose metabolism.

Low folate levels may be an atherogenic factor regardless of homocysteine levels in young healthy nonsmokers.

Low folate and high homocysteine levels are emerging as important risk factors for atherosclerosis and predictors of early coronary heart disease. We evaluated folate and homocysteine levels, compared them with endothelial function, and analyzed their association with the methylenetetrahydrofolate reductase (MTHFR) and endothelial nitric oxide synthase genotypes. We recruited 71 young healthy male nonsmokers without overt cardiovascular or renal disease. Plasma homocysteine levels were enhanced 2-fold in the subjects with the MTHFR 677T/T compared with the others (P = .0001) and also enhanced in the subjects with the endothelial nitric oxide synthase -786C allele (P = .031). Homocysteine levels were independently predicted only by the MTHFR genotype. A relationship between folate and homocysteine levels was not significant. Plasma folate levels were associated independently either with high-density lipoprotein cholesterol levels or with endothelial function in the brachial artery. These results suggest that low folate levels may be a risk factor for cardiovascular diseases regardless of homocysteine levels and that the subjects with lower folate levels should be recommended for dietary folic acid supplementation to elevate endothelial function and probably increase high-density lipoprotein cholesterol levels.

Microsomal triglyceride transfer protein gene polymorphism strongly influences circulating malondialdehyde-modified low-density lipoprotein.

Microsomal triglyceride transfer protein (MTP) plays a critical role in the assembly of lipoproteins. Therefore, we studied whether MTP gene polymorphisms are associated with atherosclerosis-promoting parameters, especially metabolic profiles and endothelial function, in healthy young men. One hundred one healthy men (mean age, 30.3 years) were studied. We analyzed the 2 promoter polymorphisms (-493G/T and -400A/T) of the MTP gene. Linkage disequilibrium analysis revealed a significant but incomplete linkage disequilibrium between the 2 polymorphisms (D' = 0.74). The -493T allele carriers (n = 26) showed marked increases in their levels of malondialdehyde-modified low-density lipoprotein (mean value, 135 vs 99 U/L in the G/G carriers; P = .003) and triglycerides (2.15 vs 1.16 mmol/L, P = .014), and reduced low-density lipoprotein particle size (259.2 vs 264.3 nm, P = .023), whereas there was no difference in apolipoproteins, insulin, adiponectin, homocysteine, folate, and endothelial function assessed using ultrasound measurement of brachial artery flow-mediated vasodilation. In contrast, the -400T allele carriers (n = 61) showed a reduced endothelial function (P = .044), accompanied by elevated apolipoprotein B levels in subjects with higher triglyceride levels. These results indicate that both promoter polymorphisms may be associated with the development of atherosclerosis and cardiovascular diseases, but that the mechanism responsible may be different.

Differentiation capacity of endothelial progenitor cells correlates with endothelial function in healthy young men.

BACKGROUND: Endothelial progenitor cells (EPCs) have been assumed to maintain vascular endothelial integrity, so the present study investigated whether the functional capacity of EPCs correlates with endothelial function in healthy young subjects, as has been confirmed in aged subjects with atherosclerotic disease. METHODS AND RESULTS: EPCs in 41 healthy, young male nonsmokers (age 33.1 +/-3.9 years, mean +/- SD) were characterized. The correlation between flow-mediated vasodilation (FMD) and the number of EPCs or the plasma concentrations of growth factors, such as vascular endothelial growth factor, did not reach statistical significance. However, FMD was significantly correlated with the EPC differentiation index, defined as the ratio of the number of EPCs to the total number of adherent cells (r=0.391, P=0.011) and the abundance of endothelial nitric oxide synthase mRNA (r=0.340, P=0.030). CONCLUSIONS: In healthy young men, despite a lack of correlation of the number or colony counts of EPCs, the ability of circulating progenitor cells to differentiate into an endothelial lineage is closely correlated with endothelial function. This cell function assay may serve as a novel biomarker for vascular function in healthy subjects in the pre-atherosclerotic stage.

Prevalence of restless legs syndrome in a Japanese elderly population.

OBJECTIVE: To estimate the prevalence of restless leg syndrome (RLS) in elderly Japanese people by means of a population-based survey of subjects aged > or = 65 years. BACKGROUND: Studies conducted worldwide have revealed large variations in the prevalence of RLS among different populations. However, few studies have been done in Japan. METHODS: A population-based survey was carried out from 2003 to 2006 through a local healthcare project in the small town of Ajimu in a rural area of southern Japan. A Japanese translation of the questionnaire covering the four features of RLS as defined by the International RLS Study Group in 1995 was used to confirm the diagnosis of RLS. All participants aged > or = 65 years were invited to fill out the questionnaire. Subjects with positive results underwent face-to-face interviews. RESULTS: A total of 1251 persons (men, 35%; mean age, 75.0+/-6.1 years) answered the questionnaire. Of these 1251 participants, 70 (5.6%) (men, 20%; mean age, 75+/-4.9 years) answered the questions on RLS positively. Face-to-face interviews and examination confirmed the diagnosis of RLS in 12 subjects. Therefore the overall prevalence of RLS in the elderly Japanese population was estimated at 0.96%, with a higher prevalence in women (1.23%) than in men (0.46%). CONCLUSION: The overall prevalence of RLS among inhabitants of Ajimu aged > or = 65 years is 0.96%. Most of the subjects identified were women. The prevalence of RLS is lower in Japan than in studies conducted in European and North American populations.

Circulating malondialdehyde-modified low-density lipoprotein is strongly associated with very small low-density lipoprotein cholesterol concentrations in healthy men.

BACKGROUND: Despite the importance of oxidized LDL and small LDL particles as atherogenic lipoproteins, the relationship between oxidized LDL and the distributions of size subclasses of lipoproteins is not fully proved. We investigated the relationship of circulating malondialdehyde-modified (MDA)-LDL, an oxidized form of LDL, and lipoprotein subclasses in healthy men. METHODS: The study group consisted of a total of 170 healthy Japanese men (55+/-9 y). Plasma cholesterol concentrations in major lipoproteins and their subclasses were determined by HPLC with gel permeation columns. RESULTS: In univariate analysis, body mass index, waist circumference, blood pressure, white blood cell count, C-reactive protein, uric acid, fasting insulin, HOMA-IR, total cholesterol, triglycerides, each VLDL subclass cholesterol, each LDL subclass cholesterol, small HDL cholesterol, and very small HDL cholesterol were positively correlated with MDA-LDL, whereas adiponectin and large HDL cholesterol were inversely correlated with MDA-LDL. In stepwise multiple regression analysis, very small LDL cholesterol, medium VLDL cholesterol, very small HDL cholesterol, small HDL cholesterol, and systolic blood pressure were identified as independent determinants of MDA-LDL (R(2)=0.718, p<0.0001). CONCLUSIONS: Circulating MDA-LDL concentrations are strongly associated with very small LDL cholesterol concentrations in healthy men. HDL size heterogeneity has a biphasic effect on MDA-LDL.

The effects of endothelial nitric oxide synthase gene polymorphisms on endothelial function and metabolic risk factors in healthy subjects: the significance of plasma adiponectin levels.

OBJECTIVE: Genetic variants of the endothelial nitric oxide synthase (eNOS) gene, Glu298Asp and T-786C, have been reported to be associated with cardiovascular disease. Adiponectin is an adipocyte-derived plasma protein with insulin-sensitizing and vascular protective effects; its levels are typically low in metabolic syndrome. Therefore, eNOS gene polymorphisms may also be associated with specific metabolic profiles, including plasma adiponectin levels and atherogenic lipids. METHODS: We evaluated the functional significance of eNOS gene Glu298Asp and T-786C polymorphisms on endothelial function and metabolic profiles in 101 healthy young men (mean age 30.3 years) before the progression of atherosclerotic lesions. RESULTS: No linkage disequilibrium was found between the two genotypes. The Asp298 allele carriers of the eNOS gene presented significantly higher plasma low density lipoprotein (LDL) cholesterol, LDL particle size, malondialdehyde-modified LDL (MDA-LDL), and fasting insulin levels and lower plasma high density lipoprotein (HDL) cholesterol, apolipoprotein A-I levels, and endothelium-dependent vasodilation when compared with noncarriers. In spite of higher MDA-LDL levels, Asp298 carriers had significantly larger LDL particle size. By contrast, in C-786 allele carriers, systolic blood pressure was significantly higher, and plasma high-molecular-weight adiponectin levels and endothelium-dependent vasodilation were significantly lower than those in non-carriers. CONCLUSIONS: Although both eNOS polymorphisms induced endothelial dysfunction, the eNOS T-786C polymorphism may be associated with adiponectin levels, whereas the Glu298Asp polymorphism may be associated with atherogenic lipid levels.

MR imaging of brainstem atrophy in progressive supranuclear palsy.

BACKGROUND: To enhance the sensitivity and specificity of the clinical diagnosis of progressive supranuclear palsy (PSP), neuroradiological parameters established in pathologically proven cases are needed. METHODS: We examined brainstem atrophy in five pathologically confirmed PSP patients (three men, mean age at death 77 years, range 64-84 years). Time interval between symptom onset and MRI ranged from 1 to 5 years, and between MRI and death from 33 to 52 months. Only one patient had clinical diagnosis of PSP at the time of MRI. Control group consisted of 19 age- and gender-matched healthy subjects. Seventeen morphometric parameters of the midbrain and pons were measured on T1-weighted midsagittal and T2-weighted axial MRI scans with Image Analyzer. Measurements of superior cerebellar peduncle (SCP) width were performed on PSP autopsy specimens. RESULTS: Mean SCP width on MRI in PSP (2.7 +/- 0.8 mm, 95%CI: 2.1-3.3) was smaller than in controls (3.7 +/- 0.5 mm, 95%CI: 3.5-3.9). Mean SCP width at autopsy was 8% smaller than mean SCP width on MRI. Midsagittal midbrain area in PSP (99.1 +/- 6.9 mm(2), 95%CI: 90.5-107.6) was smaller than in controls (141.0 +/- 18.1 mm(2), 95%CI: 132.2-149.7). Midbrain/pons area ratio in PSP was 1:5 and in controls was 1:4 (p < 0.01). Repeat MRI 17 months later in one PSP case revealed 30% decrease of SCP width. CONCLUSIONS: MR imaging with quantitative analysis may be useful in the diagnosis of early PSP and in monitoring disease course.

[Effect of high dose pergolide mesilate on restless legs syndrome associated with Parkinson disease].

Restless legs syndrome (RLS) is one of the common nocturnal disturbance seen in Parkinson's disease (PD) patients. The prevalence of RLS with PD is greater than that of general populations; however, etiology of RLS in patients with PD is still controversial. We report a 63-year-old man with PD, who was admitted to our hospital with uncontrollable unpleasant feeling in both legs leading to sleep disturbance. At age 59, he experienced numbness and nocturnal myoclonus in his right foot. One year later, he developed resting tremor and bradykinesia in his right hand, and was diagnosed as PD. Levodopa was initiated with favorable response for his resting tremor and bradykinesia, however, his dysesthesia of the legs spread to both side and associated with an urge to move which occurs at rest and was ameliorated by walking. On admission, his parkinsonism was well controlled by 400 mg/ day of levodopa/benserazide. Polysomnography (PSG) revealed periodic limb movements in sleep (PLMS). Secondary RLS such as drug-induced, iron deficiency and uraemia, was excluded in this patient. Because levodopa did not improve his RLS, additional symptomatic RLS treatment was initiated. Oral dosage with 150 microg pergolide did not have any effect on his RLS symptoms. An increase up to 750 microg pergolide led to a marked reduction of symptoms. Repeated PSG showed significant reduction of PLMS and improved sleep efficacy. Usually, low dose of dopamine agonist is enough to treat RLS occurred in general populations. However, moderate to high dose of dopamine agonists were needed for our patient with RLS, indicating that pharmacological responses might be different between RLS in general and that associated with PD. It is important to consider that PD-related RLS can be treated with high dose dopamine agonist to obtain favorable management of nocturnal disturbances.

Impact of low-density lipoprotein particle size on carotid intima-media thickness in patients with type 2 diabetes mellitus.

Small low-density lipoprotein (LDL) particles and modifications to LDL such as glycation and oxidation have been linked to the pathogenesis of atherosclerosis in patients with diabetes. We investigated whether LDL particle size, or the levels of glycated LDL or malondialdehyde-modified LDL (MDA-LDL) are associated with carotid intima-media thickness (IMT) in patients with type 2 diabetes mellitus. One hundred seventy-two patients with type 2 diabetes mellitus were enrolled. Carotid IMT was measured by high-resolution ultrasound, and LDL particle size and serum glycated LDL and MDA-LDL levels were determined. The 3 variables were significantly correlated with one another. Univariate analyses defined statistically significant correlations of carotid IMT with LDL size, hemoglobin A(1c), glycated LDL, MDA-LDL, high-density lipoprotein (HDL) cholesterol, and age. The strongest association of IMT was with LDL size (r = -0.406, P < .0001), followed by that with HDL cholesterol (r = -0.225, P = .004). A stepwise multiple regression analysis revealed that LDL size and HDL cholesterol are independent predictors of carotid IMT. Neither glycated LDL nor MDA-LDL had a significant independent contribution to the severity of carotid IMT in the multivariate model. Low-density lipoprotein particle size, but not the glycated LDL or MDA-LDL level, was independently associated with carotid IMT in patients with type 2 diabetes mellitus regardless of antidiabetic and lipid-lowering medications. These results suggest that the measurement of LDL size may be more useful than quantification of modified LDLs for assessing atherosclerosis in patients with type 2 diabetes mellitus. Small LDL particles may be the most important predictor for the risk of cardiovascular disease in diabetic patients.

Dopamine agonist therapy for Parkinson disease and pathological gambling.

We report 6 patients with pathological gambling during pharmacologic treatment for Parkinson disease (PD). Four patients were treated with levodopa/carbidopa and dopamine agonist (DA), and 2 patients received DA monotherapy. We reviewed several published reports regarding pathological gambling and antiparkinsonian therapy and suggest that more advanced PD and antiparkinsonian combination therapy (e.g., levodopa/carbidopa and DA) increases the risk for development of pathological gambling behavior compared with monotherapy with either DA or levodopa/carbidopa.

Klotho gene polymorphism may be a genetic risk factor for atherosclerotic coronary artery disease but not for vasospastic angina in Japanese.

BACKGROUND: The klotho gene, originally identified by insertional mutagenesis in mice, suppresses multiple aging phenotypes, including atherosclerosis. We tested the hypothesis that the G-395A polymorphism of the klotho gene is associated with increased risk for 2 types of ischemic heart disease in Japanese. METHODS: The study population consisted of 197 patients with coronary heart disease (CAD) who had >75% luminal diameter narrowing, 77 patients with vasospastic angina (VSA) without significant fixed coronary artery disease, and 331 healthy control subjects. RESULTS: The frequency of the A allele carriers of the klotho gene was significantly higher in the CAD group than in the control group (29.9% vs. 19.0%). The unadjusted odds ratio for CAD in the A allele carriers compared with the control group was 1.82 (p=0.004) and a traditional risk-adjusted logistic regression model revealed that the A allele was an independent predictor of CAD (odds ratio, 1.76; p=0.03). In contrast, the frequency of the A allele carriers was not significantly different in the VSA group (23.4%; adjusted odds ratio, 1.18. CONCLUSIONS: The -395A polymorphism of the human klotho gene may be a genetic risk factor for IHD and not for VSA.