High productivity of immunostimulatory membrane vesicles of Limosilactobacillus antri using glycine.

Nanosized membrane vesicles (MVs) released by bacteria play important roles in both bacteria-bacteria and bacteria-host interactions. Some gram-positive lactic acid bacteria produce MVs exhibiting immunoregulatory activity in the host. We found that both bacterial cells and MVs of Limosilactobacillus antri JCM 15950, isolated from the human stomach mucosa, enhance immunoglobulin A production by murine Peyer's patch cells. However, the thick cell walls of gram-positive bacteria resulted in low MV production, limiting experiments and applications using MVs. In this study, we evaluated the effects of glycine, which inhibits cell wall synthesis, on the immunostimulatory MV productivity of L. antri. Glycine inhibited bacterial growth while increasing MV production, with 20 g/L glycine increasing MV production approximately 12-fold. Glycine was most effective at increasing MV production when added in the early exponential phase, which indicated that cell division in the presence of glycine increased MV production. Finally, glycine increased MV productivity approximately 16-fold. Furthermore, glycine-induced MVs promoted interleukin-6 production by macrophage-like J774.1 cells, and the immunostimulatory activity was comparable to that of spontaneously produced MVs. Our results indicate that glycine is an effective agent for improving the production of MVs with immunostimulatory activity in gram-positive lactic acid bacteria, which can be applied as mucosal adjuvants and functional foods.

Intracranial Non-Sinus-Type Dural Arteriovenous Fistulas Could Be Curable by Transarterial Embolization or Transvenous Embolization with Liquid Embolic Material.

Objective: Recently, the occlusion rate of transarterial embolization (TAE) for intracranial non-sinus-type dural arteriovenous fistulas (NSDAVFs) has improved after ONYX was introduced. Additionally, when TAE for NSDAVF is unsuccessful, transvenous embolization (TVE) has become available as an alternative treatment. We investigated the factor for the favorable occlusion rate of endovascular treatment for NSDAVF at our institutions. Methods: Two hundred and twenty-seven patients with intracranial dural arteriovenous fistulas (DAVFs) were treated at our institutions between September 2014 and October 2022. The patients diagnosed with NSDAVF in all DAVFs who underwent endovascular treatment were included. The clinical characteristics, angiographical outcomes, and clinical outcomes of patients who underwent endovascular treatment were evaluated. Results: Thirty-eight patients had intracranial NSDAVF (tentorial: 23 cases, parasagittal-convexity: 7, anterior cranial fossa: 6, middle cranial fossa: 2). Our participants' mean age was 64.8 ± 11.3 years, and 31 (81.6%) of them were males. Patients' symptoms were as follows: asymptomatic (24), hemorrhage (10), tinnitus (3), and trigeminal neuralgia (1). TAE and TVE were performed on 35 and 3 patients, respectively. The rate of immediate angiographical occlusion was 84.2% (32/38). The follow-up angiographical occlusion rate in 6 months was 88.5% (31/35). Complications occurred in three cases. There was no morbidity or mortality after 30 days. Conclusion: TAE using the combination of the new microcatheter and microguidewire and TVE in the case of difficult or failed TAE for NSDAVF could achieve high success rates and safety.

Standardisation of the Japanese diet for use in animal experiments.

The aim of this study was to develop a purified diet that mimics the characteristics of the Japanese diet using readily available materials with a simpler composition and a focus on quality, with the goal of facilitating performance of studies on the Japanese diet worldwide. The utility of the new diet was examined as a mimic of the standard Japanese diet for use in animal experiments. We examined whether a key characteristic of the Japanese diet of being less likely to cause obesity could be reproduced. The mimic diet had a balance of protein, fat and carbohydrate based on the 1975 Japanese diet, which is the least likely to cause obesity, and materials chosen with reference to the National Health and Nutrition Survey (NHNS). To examine similarities of the mimic diet with the model 1975 Japanese diet, we created a menu of the 1975 diet based on the NHNS and prepared the freeze-dried and powdered diet. The mimic diet, the 1975 Japanese diet, a control AIN-93G diet and a Western diet were fed to mice for 4 weeks. As a result, the mimic diet and the 1975 diet resulted in less accumulation of visceral fat and liver fat. Mice given these two diets showed similar effects. This indicates that the mimic diet used in this study has characteristics of the 1975 Japanese diet and could be used as a standard Japanese diet in animal experiments.

Identification of Characteristic Components and Foodstuffs in Healthy Japanese Diet and the Health Effects of a Diet with Increased Use Frequency of these Foodstuffs.

SCOPE: Our recent study showed that the 1975 Japanese diet exhibited strong health benefits. In the current study, we aimed to develop a diet with even higher health benefits. METHODS: First, to determine the characteristic components in the 1975 diet, we used mass spectrometry for analysis of Japanese diets from several years and performed principal component analysis. Next, a diet with an increased use frequency of foodstuffs contained characteristic components (the modified diet) was prepared and fed to mice. RESULTS: Performed principal component analysis revealed that the 1975 diet contained 14 characteristic components that were found in fish, fruits, vegetables, seaweed, soybean foods, soup stock "dashi", and fermented seasoning. Based on these, the modified diet was prepared and fed to mice. The liver total cholesterol and serum LDL cholesterol decreased significantly in mice fed the modified diet and serum total cholesterol showed a downward trend, compared to mice fed the 1975 diet. There was no difference between the modified diet and the control groups. In addition, serum adiponectin level increased in mice fed the modified diet and serum TBARS and IL-6 levels decreased. CONCLUSION: By modifying the 1975 diet, it was possible to make a diet with more benefit.

Intake of mulberry 1-deoxynojirimycin prevents colorectal cancer in mice.

The effect of 1-deoxynojirimycin, a caloric restriction mimetic, was examined in ICR mice with azoxymethane dextran sodium sulfate-induced colorectal cancer. Azoxymethane is a carcinogen (10 mg/kg body weight), and 2% dextran sodium sulfate (w/v) used as a colitis-inducing agent. Mice were separated into 5 groups: a group without colorectal cancer fed a normal diet (CO- group), and groups with colorectal cancer fed a normal diet (CO+ group), a calorie-restricted diet (caloric restriction group), and diets including 0.02% and 0.1% 1-deoxynojirimycin (l-1-deoxynojirimycin and H-1-deoxynojirimycin groups). The tumor incidence and number were reduced significantly in the caloric restriction group compared to the CO+ group, and were also suppressed in a dose-dependent manner by 1-deoxynojirimycin. mRNA for anti-apoptotic Bcl-2 was decreased and that for pro-apoptotic Bax was increased in the carcinoma tissue of CR, l-1-deoxynojirimycin and H-1-deoxynojirimycin groups. These results suggest that caloric restriction and 1-deoxynojirimycin inhibit growth of colorectal cancer by inducing apoptosis in an induced cancer model in mice.

Breeding of a new potato variety 'Nagasaki Kogane' with high eating quality, high carotenoid content, and resistance to diseases and pests.

'Nagasaki Kogane' is a new potato variety bred from a cross between 'Saikai 35' as a female parent and 'Saikai 33' as a male. 'Saikai 35' is resistant to bacterial wilt, contains the H1 and Rychc genes for resistance to the potato cyst nematode (PCN) and potato virus Y (PVY), respectively, and has high carotenoid content, while 'Saikai 33' has large and high-yielding tubers and is resistant to both bacterial wilt and PCN. The carotenoid content of 'Nagasaki Kogane' is higher than that of 'Dejima', a common double cropping variety. The taste quality of steamed 'Nagasaki Kogane' is comparable to that of 'Inca-no-mezame' tubers, which has high levels of carotenoid, and superior to 'Nishiyutaka', another popular double cropping variety. 'Nagasaki Kogane' is suitable for French fries, because its tuber has high starch content. The marketable yield of 'Nagasaki Kogane' was higher than that of 'Inca-no-mezame' in spring cropping, although it was lower than that of 'Nishiyutaka' in double cropping regions. 'Nagasaki Kogane' tubers are larger on average than 'Inca-no-mezame' tubers in spring cropping. Moreover, the 'Nagasaki Kogane' variety is resistant to PCN and PVY, and exhibits a high level of resistance to bacterial wilt.

Comparison of the Effects of the 1975 Japanese Diet and the Modern Mediterranean Diet on Lipid Metabolism in Mice.

The Japanese diet and the Mediterranean diet are both known to be good for health, but there had been no direct comparison of their health benefits. In this study, we compared the 1975 Japanese diet, which has been found to have high health benefits, with the 2010 Italian diet, which contributes to the longest life expectancy in Mediterranean countries. Diets were created using one-week menus of the two diets based on FAOSTAT Food Balance Sheets. The diets were prepared, freeze-dried, powdered and fed to mice for 4 weeks to examine their effects on lipid metabolism. In mice fed the Japanese diet, the visceral fat weight was lower, adipocytes were smaller, the liver weight was lower and liver TG tended to be lower than those fed the Italian diet, and little lipid accumulation was observed in hepatocytes of mice fed the Japanese diet. In addition, in mice fed the Japanese diet, the expression levels of genes related to fatty acid synthesis were lower, whereas those of genes related to catabolism of fatty acids and cholesterol were higher than those fed the Italian diet. Therefore, the Japanese diet reduced accumulation of lipids in the white adipose tissue and liver by suppressing fatty acid synthesis and promoting catabolism of fatty acids and cholesterol in the liver, compared to the Italian diet.

High dietary cholesterol intake during lactation promotes development of fatty liver in offspring of mice.

SCOPE: We have previously demonstrated in mice that a maternal high fat diet during lactation enhances the susceptibility of offspring to obesity and fatty liver. A causative molecule, however, is yet to be identified. Therefore, we examined the role of cholesterol, the dietary intake of which increases with consumption of a high fat diet. Excessive cholesterol intake is involved in the development of fatty liver, which in turn becomes a risk factor for metabolic syndrome, which includes obesity, insulin resistance, and dyslipidemia. However, our knowledge of the influences on offspring of excessive maternal cholesterol intake alone during pregnancy and lactation is limited. We examined how excessive maternal cholesterol intake during lactation influences susceptibility of the offspring to metabolic syndrome in mice. RESULTS: High cholesterol intake promoted triacylglycerol accumulation in the liver of offspring (p < 0.05), and elevated expression of molecules involved in hepatic lipoprotein influx was identified as the underlying mechanism. CONCLUSION: These findings demonstrate that excessive maternal cholesterol intake during lactation enhances the susceptibility of the offspring to development of fatty liver.

Effects of Fatty Acid Quality and Quantity in the Japanese Diet on the Suppression of Lipid Accumulation.

Japan has been known as a healthy country since its life expectancy became among the highest in the world in the 1980s. The influence of the Japanese diet is one of the factors explaining Japan's high life expectancy. Our recent study that fed representative freeze-dried and powdered Japanese diets from 1960, 1975, 1990, and 2005 based on National Health and Nutrition Research to mice showed the 1975 Japanese diet exhibited the strongest visceral fat accumulation suppression and overall health benefits. However, it is unclear why. We investigated the effects of the fatty acid composition in Japanese diets on visceral fat accumulation in mice. ICR mice were fed diets replicating the fatty acid composition and macronutrient ratios of Japanese diets from 1960, 1975, 1990, and 2005 for four weeks. The 1975 diet suppressed visceral fat accumulation and adipocyte hypertrophy. DNA microarray analysis showed the 1975 diet suppressed Acyl-CoA synthetase and prostaglandin D2 synthase mRNA expressions in white adipose tissue. As the effects of the 1975 diet are likely due to differences in fatty acid intake and/or composition, we investigated test diets that replicated only the fatty acid composition of Japanese diets. There were no significant differences in visceral fat mass. Therefore, both the quality and quantity of fatty acids are involved in the anti-obesity effects of the 1975 Japanese diet.

The Japanese diet from 1975 delays senescence and prolongs life span in SAMP8 mice.

OBJECTIVE: Life expectancy in Japan is high, suggesting that the Japanese diet, Nihon shoku (Japanese food), has significant health benefits. However, these benefits have been called into question over the past 50 y, during which time the Japanese diet has become increasingly Westernized. The aim of the present study was to focus on senescence delay and to examine the effects of Japanese diets from different years to identify which Japanese diet is most effective in enhancing life expectancy and delaying senescence. METHODS: Weekly menus from the years 1960, 1975, 1990, and 2005 were reproduced based on the National Health and Nutrition Survey in Japan and prepared as powdered foods. The senescence-accelerated mouse prone 8 (SAMP8) mice were fed standard laboratory chow supplemented with a 30% mix of Japanese meals from various years ad libitum throughout their lifetime. Additionally, the control group was given standard laboratory chow only, to examine the development of mice reared under standard conditions. RESULTS: In the group that ingested the traditional 1975 Japanese diet, life span was prolonged, senescence was delayed, and learning and memory capacities were maintained compared with the group fed the 2005 Japanese diet. The life span of the group that ingested the 1990 Japanese diet showed a tendency to be longer than SAMP8 mice fed the 2005 diet. CONCLUSION: The results of the present study suggested that the traditional Japanese diet is more effective in enhancing life expectancy and delaying senescence than the current Japanese diet.

High-fat diet intake from senescence inhibits the attenuation of cell functions and the degeneration of villi with aging in the small intestine, and inhibits the attenuation of lipid absorption ability in SAMP8 mice.

We examined the effect of a high-fat diet from senescence as a means of preventing malnutrition among the elderly. The senescence-accelerated mouse P8 was used and divided into three groups. The 6C group was given a normal diet until 6 months old. The 12N group was given a normal diet until 12 months old. The 12F group was given a normal diet until 6 months old and then a high-fat diet until 12 months old. In the oral fat tolerance test, there was a decrease in area under the curve for serum triacylglycerol level in the 12N group and a significant increase in the 12F group, suggesting that the attenuation of lipid absorption ability with aging was delayed by a high-fat diet from senescence. To examine this mechanism, histological analysis in the small intestine was performed. As a result, the degeneration of villi with aging was inhibited by the high-fat diet. There was also a significant decrease in length of villus in the small intestine in the 12N group and a significant increase in the 12F group. The high-fat diet from senescence inhibited the degeneration of villi with aging in the small intestine, and inhibited the attenuation of lipid absorption ability.

High Dietary Fat Intake during Lactation Promotes the Development of Social Stress-Induced Obesity in the Offspring of Mice.

This study examined how a maternal high-fat diet (HD) during lactation and exposure of offspring to isolation stress influence the susceptibility of offspring to the development of obesity. C57BL/6J mice were fed a commercial diet (CD) during pregnancy and a CD or HD during lactation. Male offspring were weaned at three weeks of age, fed a CD until seven weeks of age, and fed a CD or HD until 11 weeks of age. Offspring were housed alone (isolation stress) or at six per cage (ordinary circumstances). Thus, offspring were assigned to one of eight groups: dams fed a CD or HD during lactation and offspring fed a CD or HD and housed under ordinary circumstances or isolation stress. Serum corticosterone level was significantly elevated by isolation stress. High-fat feeding of offspring reduced their serum corticosterone level, which was significantly elevated by a maternal HD. A maternal HD and isolation stress had combined effects in elevating the serum corticosterone level. These findings suggest that a maternal HD during lactation enhances the stress sensitivity of offspring. White adipose tissue weights were significantly increased by a maternal HD and isolation stress and by their combination. In addition, significant adipocyte hypertrophy was induced by a maternal HD and isolation stress and exacerbated by their combination. Thus, a maternal HD and isolation stress promote visceral fat accumulation and adipocyte hypertrophy, accelerating the progression of obesity through their combined effects. The mechanism may involve enhanced fatty acid synthesis and lipid influx from blood into adipose tissue. These findings demonstrate that a maternal HD during lactation may increase the susceptibility of offspring to the development of stress-induced obesity.

Decreased lipid absorption due to reduced pancreatic lipase activity in aging male mice.

Malnutrition due to aging is partly caused by decreased absorption of nutrients by the gastrointestinal tract. However, the underlying mechanism is unclear and changes in lipid absorption with aging are poorly understood. In this study, changes in lipid absorption with aging were examined in mice aged 3 and 25 months. After overnight fasting, blood samples were collected from snipped tails and then soybean oil was administered orally. Three hours later, mice were sacrificed by decapitation and the liver, pancreas, small intestine and blood were collected. The increase in serum triacylglycerol after soybean oil administration was significantly lower in the older mice, indicating a decrease in lipid absorption with aging. Measurement of mRNA levels for triacylglycerol absorption-related genes showed that mRNA for pancreatic lipase tended to decrease in 25-month-old mice. There was no significant difference in the protein level of pancreatic lipase, but the enzyme activity showed a significant decrease in the older mice. To examine this mechanism, expression levels of mRNA for protein turnover-related genes in the pancreas were measured. The level of a proteasomal mRNA showed a significant decrease in 25-month-old mice. This suggests that the ability to degrade unfolded protein decreases in the aging pancreas, and that this leads to reduction of pancreatic lipase activity and a decrease in lipid absorption.

Kcna1-mutant rats dominantly display myokymia, neuromyotonia and spontaneous epileptic seizures.

Mutations in the KCNA1 gene, which encodes for the α subunit of the voltage-gated potassium channel Kv1.1, cause episodic ataxia type 1 (EA1). EA1 is a dominant human neurological disorder characterized by variable phenotypes of brief episodes of ataxia, myokymia, neuromyotonia, and associated epilepsy. Animal models for EA1 include Kcna1-deficient mice, which recessively display severe seizures and die prematurely, and V408A-knock-in mice, which dominantly exhibit stress-induced loss of motor coordination. In the present study, we have identified an N-ethyl-N-nitrosourea-mutagenized rat, named autosomal dominant myokymia and seizures (ADMS), with a missense mutation (S309T) in the voltage-sensor domain, S4, of the Kcna1 gene. ADMS rats dominantly exhibited myokymia, neuromyotonia and generalized tonic-clonic seizures. They also showed cold stress-induced tremor, neuromyotonia, and motor incoordination. Expression studies of homomeric and heteromeric Kv1.1 channels in HEK cells and Xenopus oocytes, showed that, although S309T channels are transferred to the cell membrane surface, they remained non-functional in terms of their biophysical properties, suggesting a dominant-negative effect of the S309T mutation on potassium channel function. ADMS rats provide a new model, distinct from previously reported mouse models, for studying the diverse functions of Kv1.1 in vivo, as well as for understanding the pathology of EA1.

Nitric oxide-induced downregulation of leptin production by 3T3-L1 adipocytes.

Leptin secreted mainly by adipocytes plays an important role in insulin sensitivity in metabolic syndrome. Inducible nitric oxide synthase (iNOS) in 3T3-L1 adipocytes is induced by lipopolysaccharide (LPS) and several proinflammatory cytokines such as tumor necrosis factor-alpha and interferon-gamma (IFN-gamma). Because the role of iNOS-derived nitric oxide (NO) in adipocyte function has not been fully clarified, the question that we addressed in the present study was whether iNOS-derived NO is involved in regulation of leptin secretion by adipocytes. Incubation of 3T3-L1 adipocytes for 12h with a mixture of IFN-gamma and LPS caused not only a 55% reduction in leptin secretion and a 52% reduction in leptin mRNA, but also significant induction of iNOS at both protein and mRNA levels. Inhibition of leptin secretion that had been induced by the IFN-gamma-LPS mixture was completely nullified by NOS inhibitors such as Nomega-monomethyl-L-arginine and aminoguanidine. Treatment of adipocytes with NO donors such as an NONOate and S-nitrosoglutathione produced an effect on leptin secretion similar to that of the IFN-gamma-LPS mixture. It is likely therefore that NO mediates downregulation of leptin caused by the IFN-gamma-LPS mixture in 3T3-L1 adipocytes, which suggests an important role for NO in adipocyte functions.

Human urotensin II accelerates foam cell formation in human monocyte-derived macrophages.

Human urotensin II (U-II), the most potent vasoconstrictor peptide identified to date, and its receptor (UT) are involved in hypertension and atherosclerosis. Acyl-coenzyme A:cholesterol acyltransferase-1 (ACAT-1) converts intracellular free cholesterol into cholesterol ester (CE) for storage in lipid droplets and plays an important role in the formation of macrophage-derived foam cells in atherosclerotic lesions. We examined the effects of U-II on ACAT-1 expression and CE accumulation in human monocyte-derived macrophages. U-II increased ACAT activity in a concentration-dependent manner after 7 days in monocyte primary culture. Immunoblotting analysis showed that U-II at 25 nmol/L increased ACAT-1 protein expression level by 2.5-fold, which was completely abolished by anti-U-II antibody, selective UT receptor antagonists (urantide and 4-aminoquinoline), a G-protein inactivator (GDP-beta-S), a c-Src protein tyrosine kinase inhibitor (PP2), a protein kinase C (PKC) inhibitor (rottlerin), a mitogen-activated protein kinase kinase (MEK) inhibitor (PD98059), or a Rho kinase (ROCK) inhibitor (Y27632). Northern blotting analysis indicated that among the 4 ACAT-1 mRNA transcripts (2.8-, 3.6-, 4.3-, and 7.0-kb), the 2.8- and 3.6-kb transcript levels were selectively upregulated by approximately 1.7-fold by U-II (25 nmol/L). Further, U-II (25 nmol/L) significantly increased acetylated LDL (acetyl-LDL)-induced CE accumulation in monocyte-derived macrophages but not scavenger receptor class A (SR-A) function as assessed by endocytic uptake of [(125)I]acetyl-LDL. Our results suggest that U-II may play a novel role in the formation of macrophage-derived foam cells by upregulating ACAT-1 expression via the UT receptor/G-protein/c-Src/PKC/MEK and ROCK pathways but not by SR-A, thus contributing to the relatively rapid development of atherosclerosis in hypertension.

Pathological roles of advanced glycation end product receptors SR-A and CD36.

The pathological significance of advanced glycation end product (AGE)-modified proteins deposited in several lesions is generally accounted for by their cellular interaction via the AGE receptors and subsequent acceleration of the inflammatory process. In this study, we focused on two AGE receptors-specifically, the role of SR-A in pathogenesis of diabetic nephropathy and the role of CD36 in AGE-induced downregulation of leptin by adipocytes. In terms of SR-A, diabetic wild-type mice exhibited increased urinary albumin excretion, glomerular hypertrophy, and mesangial matrix expansion, whereas SR-A-knockout mice showed reduced glomerular size and mesangial matrix area. In these diabetic SR-A-knockout mice, the number of macrophages that infiltrated into glomeruli was remarkably reduced (P < 0.05), suggesting that SR-A-dependent glomerular migration of macrophages plays an important role in the pathogenesis of diabetic nephropathy. In terms of CD36, incubation of glycolaldehyde-modified bovine serum albumin (GA-BSA) with 3T3-L1 adipocytes reduced leptin secretion by these cells. The binding of GA-BSA to these cells and subsequent endocytic degradation were effectively inhibited by a neutralizing anti-CD36 antibody. AGE-induced downregulation of leptin was protected by N-acetyl-cysteine, an antioxidant. These results indicate that the interaction of AGE ligands with 3T3-L1 adipocytes via CD36 induces oxidative stress and leads to inhibition of leptin expression by these cells, suggesting a potential link of this phenomenon to exacerbation of the insulin sensitivity in metabolic syndrome.

Glycolaldehyde-modified bovine serum albumin downregulates leptin expression in mouse adipocytes via a CD36-mediated pathway.

Previous observations by us have clarified that proteins modified by advanced glycation end products (AGEs) are recognized as effective ligands by CD36-overexpressed CHO cells and undergo receptor-mediated endocytosis. CD36, a member of the class B scavenger receptor family, also acts as a fatty acid transporter in adipocytes. Oxidized low-density lipoprotein (Ox-LDL), a ligand for CD36, is known to upregulate CD36 by activating peroxisome proliferator-activated receptor gamma (PPAR-gamma) in macrophages, whereas PPAR-gamma ligands such as troglitazone and 15-deoxy-delta12,14-prostaglandin J2 decrease leptin secretion from adipocytes. The purpose of this study was to examine effects of AGE ligands on leptin expression in adipocytes. Glycolaldehyde-modified bovine serum albumin (GA-BSA) decreased leptin expression at both the protein and mRNA levels in 3T3-L1 adipocytes and mouse epididymal adipocytes. The binding to and subsequent endocytic degradation of GA-BSA by 3T3-L1 adipocytes were effectively inhibited by a neutralizing anti-CD36 antibody. These results indicate that the ligand interaction of GA-BSA with CD36 leads to downregulation of leptin expression in 3T3-L1 adipocytes, suggesting that AGE-induced leptin downregulation is linked to reduction of the insulin sensitivity in metabolic syndrome.

Specific interaction of oxidized low-density lipoprotein with thrombospondin-1 inhibits transforming growth factor-beta from its activation.

Oxidized LDL (Ox-LDL) plays atherogenic roles, whereas thrombospondin-1 (TSP-1) is thought to be anti-atherogenic through activation of TGF-beta that contributes to plaque stabilization. Ox-LDL was prepared by incubating of human LDL with CuSO4. Effect of Ox-LDL on TSP-1-induced TGF-beta activation was examined in the present study. Incubation of Ox-LDL with mouse peritoneal macrophages for 3 days resulted in reduction in amounts of active TGF-beta in the culture medium by 70-78% when compared with that of parallel incubation without Ox-LDL. TSP-1 could enhance conversion of latent TGF-beta1 into active TGF-beta1 in a cell-free system. This TSP-1-mediated latent TGF-beta1 activation was inhibited by 30% by Ox-LDL, suggesting the possible interaction of Ox-LDL with TSP-1. Incubation of TSP-1 with [125I]Ox-LDL or [125I]LDL, followed by immunoprecipitation with an anti-TSP-1 antibody demonstrated that a significant amount of [125I]Ox-LDL was co-precipitated with TSP-1 while precipitation of [125I]LDL was negligible. Furthermore, upon TSP-1-conjugated Sepharose 4B affinity chromatography, both [125I]Ox-LDL and [125I]latent TGF-beta1 bound to the affinity gel were eluted by unlabeled Ox-LDL. These findings indicate that Ox-LDL interacts with TSP-1 and suppresses subsequent TSP-1-dependent TGF-beta activation, revealing a novel atherogenic function of Ox-LDL.

Narrowed abrogation of the Angelman syndrome critical interval on human chromosome 15 does not interfere with epigenotype maintenance in somatic cells.

Human chromosome 15q11-q13 involves a striking imprinted gene cluster of more than 2 Mb that is concomitant with multiple neurological disorders manifested by Prader-Willi syndrome (PWS) and Angelman syndrome (AS). PWS and AS patients with imprinting mutation have microdeletions, which share a 4.3 kb short region of overlap (SRO) at the 5' end of the paternal SNURF-SNRPN gene in PWS, or on the maternal allele, which shares a 880 bp SRO located at the 35 kb upstream of the SNURF-SNRPN promoter in AS. Recent studies have revealed an essential role of PWS-SRO in the postzygotic maintenance of the appropriate epigenotype on the paternal chromosome. For AS-SRO, however, there is insufficient experimental evidence exists to determine the direct functions. Here we show that the complete deletion of AS-SRO does not cause any anomalies of imprinted gene expression or DNA methylation on the mutated human chromosome 15, further supporting the idea that AS-SRO is dispensable for post implantation imprint maintenance. This implies that AS-SRO is not essential for the robust epigenotype preservation in somatic cells.