Docosahexaenoic Acid Increases Vesicular Glutamate Transporter 2 Protein Levels in Differentiated NG108-15 Cells.

Docosahexaenoic acid (DHA; 22:6n-3), which is enriched in the neuronal membrane, plays a variety of roles in the brain. Vesicular glutamate transporters (VGLUTs) are responsible for incorporating glutamine into synaptic vesicles. We investigated the influence of DHA on the fatty acid profile and the levels of VGLUT1 and VGLUT2 proteins in differentiated NG108-15 cells, a neuroblastoma-glioma hybrid cell line. NG108-15 cells were plated and 24 h later the medium was replaced with Dulbecco's modified Eagle's medium supplemented with 1% fetal bovine serum, 0.2 mM dibutyryl cAMP, and 100 nM dexamethasone, which was added to induce differentiation. After 6 d, the amount of DHA in the cells was increased by addition of DHA to the medium. VGLUT2 levels were increased by the addition of DHA. These data indicate that DHA affected the levels of VGLUT2 in NG108-15 cells under differentiation-promoting conditions, suggesting that DHA affects brain functions involving VGLUT2.

Comparative analysis of age in monocrotaline-induced pulmonary hypertensive rats.

Pulmonary arterial hypertension (PAH) is a rare, progressive, and fatal cardiovascular/lung disease. The incidence rate is affected by age. Monocrotaline (MCT, 60 mg/kg)-treated rats are widely used as an experimental PAH model. Here, we found that young rats died at a mean of 23.4 days after MCT injection, whereas adult rats survived for over 42 days. However, young (7-week-old) and adult (20-week-old) MCT-treated rats developed PAH, and had upregulated Ca2+-sensing receptor and transient receptor potential canonical subfamily 6 channel expression in pulmonary arteries. The present study provides novel information for elucidating the mechanism underlying the age difference in PAH patients.

Fully hydrogenated canola oil extends lifespan in stroke-prone spontaneously hypertensive rats.

BACKGROUND: Canola oil (Can) and several vegetable oils shorten the lifespan of stroke-prone spontaneously hypertensive rats (SHRSP). Although similar lifespan shortening has been reported for partially hydrogenated Can, the efficacy of fully hydrogenated oils on the lifespan remains unknown. The present study aimed to investigate the lifespan of SHRSP fed diets containing 10 % (w/w) of fully hydrogenated Can (FHCO) or other oils. METHODS: Survival test: Upon weaning, male SHRSP were fed a basal diet for rodents mixed with one of the test oils -i.e., FHCO, Can, lard (Lrd), and palm oil (Plm) throughout the experiment. The animals could freely access the diet and drinking water (water containing 1 % NaCl), and their body weight, food intake, and lifespan were recorded. Biochemical analysis test: Male SHRSP were fed a test diet with either FHCO, Can, or soybean oil (Soy) under the same condition, except to emphasize effects of fat, that no NaCl loading was applied. Soy was used as a fat source in the basal diet and was set the control group. Blood pressures was checked every 2 weeks, and serum fat levels and histological analyses of the brain and kidney were examined after 7 or 12 weeks of feeding. RESULTS: During the survival study period, the food consumption of FHCO-fed rats significantly increased (15-20 % w/w) compared with that of rats fed any other oil. However, the body weight gain in the FHCO group was significantly less (10-12 %) than that in the control group at 9-11 weeks old. The FHCO (> 180 days) intervention had the greatest effect on lifespan, followed by the Lrd (115 ± 6 days), Plm (101 ± 2 days), and Can (94 ± 3 days) diets. FHCO remarkably decreased the serum cholesterol level compared with Can and the systolic blood pressure from 12 to 16 weeks of age. In addition, while some rats in the Can group exhibited brain hemorrhaging and renal dysfunction at 16 weeks old, no symptoms were observed in the FHCO group. CONCLUSION: This current study suggests that complete hydrogenation decreases the toxicity of Can and even prolongs the lifespan in SHRSP.

A Critical Review of the Consensus Statement from the European Atherosclerosis Society Consensus Panel 2017.

BACKGROUND: The Consensus Statement from the European Atherosclerosis Society (EAS) Consensus Panel 2017 concludes on the basis of 3 different types of clinical studies that low-density lipoprotein (LDL) causes atherosclerotic cardiovascular disease (ASCVD). In Mendelian randomization studies, rare genetic mutations affecting LDL receptor function were found to cause higher or lower LDL-C levels, which are associated with correspondingly altered ASCVD risk. In prospective cohort studies and randomized controlled trials (RCTs) of statins, a remarkably consistent log-linear association was demonstrated between the absolute magnitude of LDL-C exposure and ASCVD risk. The EAS Statement proposes that any mechanism of lowering plasma LDL concentration should reduce the risk of ASCVD events proportional to the absolute reduction in LDL-C and the cumulative duration of exposure to lower LDL-C. However, as we explain, we do not find this conclusion acceptable. SUMMARY: Our review points out that different interpretations are possible for the results of Mendelian randomization studies. As for prospective cohort studies, many inconsistent reports on the association of LDL-C and ASCVD were disregarded when drafting the Statement, reports with and without genetic factors related to LDL receptor function should be analyzed separately, and the term ASCVD in the Statement is used inappropriately because myocardial infarction and cerebral infarction differ in their association with LDL-C. As for RCTs, clinical reports on statins published before and after the implementation of new regulations affecting clinical trials (2004/2005) should not both be included in meta-analyses because the evaluated efficacy of statins changed markedly, and the irreversible adverse effects of statins need to be evaluated more rigorously now that their mechanisms have been elucidated. Key Messages: Apart from the EAS hypothesis that LDL causes ASCVD, recent pharmacological/biochemical studies, as summarized in this review and elsewhere, have revealed that atherosclerosis is caused by statins taken to lower LDL-C, as well as by warfarin and some types of vegetable fats and oils, in the absence of significantly elevated LDL-C levels. Thus, the promotion of statin treatment by the Statement is rather risky and we do not feel that the conclusions are justified for the prevention of ASCVD.

Medicines and Vegetable Oils as Hidden Causes of Cardiovascular Disease and Diabetes.

BACKGROUND: Positive associations have been observed between cardiovascular disease (CVD) and type 2 diabetes mellitus (DM), but their causal relationship has not been clarified. Nevertheless, guidelines from relevant medical societies recommend using cholesterol lowering medication (statin) for both types of patients. Medicines with several different action mechanisms have been developed, and the effectiveness of different lifestyle modifications has been studied extensively for the prevention of DM, which was successful in improving clinical marker status in relatively short-term treatments, but none have been shown to be effective in improving long-term outcomes (mortality from CVD and all causes). SUMMARY: Statin-induced suppression of prenyl intermediates in the cholesterol biosynthetic pathway has been linked to stimulated atherosclerosis and heart failure. On the other hand, certain types of vegetable oil and hydrogenated oil shortened the survival of stroke-prone spontaneously hypertensive rats by decreasing platelet number, increasing hemorrhagic tendency and damaging kidney functions, which could not be accounted for by their fatty acid and phytosterol compositions. These vegetable oils and medicines such as statin and warfarin share, in part, a common mechanism to inhibit vitamin K2-dependent processes, which was interpreted to lead to increased onset of CVD, DM, chronic kidney disease, bone fracture and even mental disorder. Impaired vitamin K2-dependent processes by some types of vegetable oils and medicines, but not plasma high low density lipoprotein cholesterol, were proposed as the cause of CVD, DM and other lifestyle-related diseases. High n-6/n-3 fatty acid ratio of ingested foods, but not animal fats, was emphasized to be another risk factor for many of the diseases described above. KEY MESSAGES: To date, no randomized controlled trials (RCTs) have been performed to prove the above interpretation. However, the opposite types of RCT trials have been performed by increasing the intake of high-linoleic vegetable oils and reducing that of animal fats, which resulted in increased CVD and all-cause mortality. The amounts of these vegetable oils to exhibit adverse effects in animal studies are not huge (<6 energy %), which should not be overlooked nor disregarded.

Effects of arachidonic acid intake on inflammatory reactions in dextran sodium sulphate-induced colitis in rats.

The aim of this study was to investigate the effects of the administration of oral arachidonic acid (AA) in rats with or without dextran sulphate sodium (DSS)-induced inflammatory bowel disease. Male Wistar rats were administered AA at 0, 5, 35 or 240 mg/kg daily by gavage for 8 weeks. Inflammatory bowel disease was induced by replacing drinking water with 3 % DSS solution during the last 7 d of the AA dosing period. These animals passed loose stools, diarrhoea and red-stained faeces. Cyclo-oxygenase-2 concentration and myeloperoxidase activity in the colonic tissue were significantly increased in the animals given AA at 240 mg/kg compared with the animals given AA at 0 mg/kg. Thromboxane B2 concentration in the medium of cultured colonic mucosae isolated from these groups was found to be dose-dependently increased by AA, and the increase was significant at 35 and 240 mg/kg. Leukotriene B4 concentration was also significantly increased and saturated at 5 mg/kg. In addition, AA at 240 mg/kg promoted DSS-induced colonic mucosal oedema with macrophage infiltration. In contrast, administration of AA for 8 weeks, even at 240 mg/kg, showed no effects on the normal rats. These results suggest that in rats with bowel disease AA metabolism is affected by oral AA, even at 5 mg/kg per d, and that excessive AA may aggravate inflammation, whereas AA shows no effects in rats without inflammatory bowel disease.

Lipase-catalyzed interesterification of Sunite sheep tail fat and flaxseed oil provides a fat having a unique fatty acid content and favorable physicochemical and nutritional properties.

The aim of this study is to combine flaxseed oil (FO), rich in α-linolenic acid (ALA), with Sunite sheep tail fat (STF) through a lipase-catalyzed transesterification reaction, in order to produce an edible oil with a fatty acid ratio suitable for human needs. Initially, the optimal conditions for esterification were determined using the Box-Behnken design, with the measurement criterion being the content of ALA at the sn-2 position. The results indicated that the highest content of sn-2 ALA was obtained under the conditions of using 6.8 wt% Lipozyme®RMIM as the catalyst, a reaction temperature of 57°C, a reaction time of 3.3 h, and a substrate mass ratio of 5.6:4.4 for STF and FO. This led to the rapid breaking and recombining of molecular bonds, resulting in the interesterified fat (IF) with the highest content of ALA at the sn-2 position. Comparing STF and FO, IF exhibited excellent fatty acid composition and content. Furthermore, IF had a lower melting point and crystallization temperature compared to STF, and its solid fat content decreased with increasing temperature, completely melting at temperatures above 30°C. Thus, IF is a synthesized fat with excellent properties from both animal and vegetable sources.

Arachidonic acid supplementation during gestational, lactational and post-weaning periods prevents retinal degeneration induced in a rodent model.

Fatty acids and their derivatives play a role in the response to retinal injury. The effects of dietary arachidonic acid (AA) supplementation on N-methyl-N-nitrosourea (MNU)-induced retinal degeneration was investigated in young Lewis rats during the gestational, lactational and post-weaning periods. Dams were fed 0·1, 0·5 or 2·0% AA diets or a basal (< 0·01% AA) diet. On postnatal day 21 (at weaning), male pups received a single intraperitoneal injection of 50 mg MNU/kg or vehicle, and were fed the same diet as their mother for 7 d. Retinal apoptosis was analysed by the terminal deoxynucleotidyl transferase-mediated dUTP digoxigenin nick-end labelling (TUNEL) assay 24 h after the MNU treatment, and retinal morphology was examined 7 d post-MNU. Histologically, all rats that received MNU and were fed the basal and 0·1% AA diets developed retinal degeneration characterised by the loss of photoreceptor cells (disappearance of the outer nuclear layer and the photoreceptor layer) in the central retina. The 0·5 and 2·0% AA diets rescued rats from retinal damage. Morphometrically, in parallel with the AA dose (0·5 and 2·0% AA), the photoreceptor ratio significantly increased and the retinal damage ratio decreased in the central retina, compared with the corresponding ratios in basal diet-fed rats. In parallel with the increase in serum and retinal AA levels and the AA:DHA ratio, the apoptotic index in the central retina was dose-dependently decreased in rats fed the 0·5 and 2·0% AA diets. In conclusion, an AA-rich diet during the gestation, lactation and post-weaning periods rescued young Lewis rats from MNU-induced retinal degeneration via the inhibition of photoreceptor apoptosis. Therefore, an AA-enriched diet in the prenatal and postnatal periods may be an important strategy to suppress the degree of photoreceptor injury in humans.

Docosahexaenoic acid contributes to increased CaMKII protein expression and a tendency to increase nNOS protein expression in differentiated NG108-15 cells.

Docosahexaenoic acid (DHA; 22:6n-3), an n-3 polyunsaturated fatty acid, has various important roles in brain functions. Nitric oxide (NO) produced by neuronal NO synthase (nNOS) and Ca2+/calmodulindependent protein kinase II (CaMKII) is also involved in brain functions. We investigated the influence of DHA on nNOS and CaMKII protein expression in differentiated NG108-15 cells. NG108-15 cells were seeded in 12-well plates, and after 24 h, the medium was replaced with Dulbecco's modified Eagle's medium containing 1% fetal bovine serum, 0.2 mM dibutyryl cyclic adenosine monophosphate and 100 nM dexamethasone as differentiation-inducing medium. When cells were cultured in differentiation-inducing medium, neurite-like outgrowths were observed on days 5 and 6. However, no significant difference in morphology was observed in cells with or without DHA treatment. With or without DHA addition, nNOS protein expression was increased on days 5 and 6 compared with day 0. This increase tended to be enhanced by DHA. CaMKII protein expression did not change after differentiation without DHA, but was significantly increased on day 6 compared with day 0 with DHA addition. These data indicate that DHA is involved in brain functions by regulating CaMKII and nNOS protein expression.

Rapeseed (canola) oil aggravates metabolic syndrome-like conditions in male but not in female stroke-prone spontaneously hypertensive rats (SHRSP).

This study was conducted to investigate whether or not there are sex differences in canola oil (CAN)-induced adverse events in the rat and to understand the involvement and the role of testosterone in those events, including life-shortening. Stroke-prone spontaneously hypertensive rats (SHRSP) of both sexes were fed a diet containing 10 wt/wt% soybean oil (SOY, control) or CAN as the sole dietary fat. The survival of the males fed the CAN diet was significantly shorter than that of those fed the SOY diet. In contrast, the survival of the females was not affected by CAN. The males fed the CAN diet showed elevated blood pressure, thrombopenia and insulin-tolerance, which are major symptoms of metabolic syndrome, whereas such changes by the CAN diet were not found in the females. Plasma testosterone was significantly lower in animals of both sexes fed the CAN diet than in those fed the SOY diet, but interestingly, the lowered testosterone was accompanied by a marked increase in plasma aldosterone only in the males. These results demonstrate significant sex differences in CAN-toxicity and suggest that those sex differences may be attributable to the increased aldosterone level, which triggers aggravation of the genetic diseases specific to SHRSP, that is, metabolic syndrome-like conditions, but only in the males. The present results also suggest that testosterone may negatively regulate aldosterone production in the physiology of the males, and the inhibition of that negative regulation caused by the CAN diet is one of the possible causes of the adverse events.

Identification and characterization of PRG-1 as a neuronal calmodulin-binding protein.

Intracellular Ca2+-dependent cellular responses are often mediated by the ubiquitous protein CaM (calmodulin), which, upon binding Ca2+, can interact with and alter the function of numerous proteins. In the present study, using a newly developed functional proteomic screen of rat brain extracts, we identified PRG-1 (plasticity-related gene-1) as a novel CaM target. A CaM-overlay and an immunoprecipitation assay revealed that PRG-1 is capable of binding the Ca2+/CaM complex in vitro and in transfected cells. Surface plasmon resonance and zero-length cross-linking showed that the C-terminal putative cytoplasmic domain (residues 466-766) of PRG-1 binds equimolar amounts of CaM in a Ca2+-dependent manner, with a relatively high affinity (a Kd value for Ca2+/CaM of 8 nM). Various PRG-1 mutants indicated that the Ca2+/CaM-binding region of PRG-1 is located between residues Ser554 and Gln588, and that Trp559 and Ile578 potentially anchor PRG-1 to CaM. This is supported by pronounced changes in the fluorescence emission spectrum of Trp559 in the PRG-1 peptide (residues 554-588) upon binding to Ca2+/CaM, showing the stoichiometrical binding of the PRG-1 peptide with Ca2+/CaM. Immunoblot analyses revealed that the PRG-1 protein is abundant in brain, but is weakly expressed in the testes. Immunohistochemical analysis revealed that PRG-1 is highly expressed in forebrain structures and in the cerebellar cortex. Furthermore, PRG-1 localizes at the postsynaptic compartment of excitatory synapses and dendritic shafts of hippocampal neurons, but is not present in presynaptic nerve terminals. The combined observations suggest that PRG-1 may be involved in postsynaptic functions regulated by intracellular Ca2+-signalling.

A case of lower-extremity deep burn wounds with periosteal necrosis successfully treated by use of allogenic cultured dermal substitute.

In patients with burns, bone exposure accompanies serious problems which occasionally lead to amputation. We present a case of an 82-year-old woman who sustained 22% of total body surface area flame burns on her bilateral lower extremities with bone exposure. Despite fascial excision and mesh skin graft, muscles, bones, and tendons were widely exposed on her right leg. The wound was infected by methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa. To promote wound healing, we applied an allogeneic cultured dermal substitute (CDS) to the wound surface once weekly, resulting in healthy granulation except for the exposed bone area of the right anterior tibia. We then shaved the cortex of the exposed bone surface until bone marrow bleeding, and grafted mesh skin in combination with CDS. Finally, all wounds healed without osteomyelitis. The use of CDS to treat deep burns exposing bone surface may expand reconstructive options for extremities that otherwise might have been amputated.

Dietary soybean oil, canola oil and partially-hydrogenated soybean oil affect testicular tissue and steroid hormone levels differently in the miniature pig.

The present study was conducted to examine the influence of dietary canola oil (CAN) and partially-hydrogenated soybean oil (HSO) compared to soybean oil (SOY, control) on the morphology and function of testes using miniature pigs as the test subject. Male miniature pigs were fed a diet containing 10%SOY, 9%CAN+1%SOY, or 9%HSO+1%SOY for 18 months. The scheduled autopsies revealed no abnormalities in histopathological examination of the major organs, except the testes. Atrophy of the seminiferous tubules and hyperplasia in the Leydig cells were found in the SOY and CAN groups. DNA microarray analysis indicated downregulation in the CAN and the HSO groups of genes encoding for gonadotropins in the pituitary gland and of enzymes and proteins involved in steroid hormone metabolism in the testes, compared to the SOY group. Plasma levels of sex hormones in the CAN and HSO groups tended to be higher and testosterone and dihydrotestosteorne in the HSO group were significantly higher than in the SOY group. These results demonstrate that testes are morphologically and functionally affected by the dietary oils, while the plasma steroid hormone levels do not necessarily reflect the gene expression, probably owing to feedback regulation via the gonadal hormones in the hypothalamus-pituitary-gonadal axis.

IQ-ArfGEF/BRAG1 is associated with synaptic ribbons in the mouse retina.

IQ-ArfGEF/BRAG1 is a guanine nucleotide exchange factor for ADP ribosylation factors (Arfs), which are implicated in membrane trafficking and actin cytoskeleton dynamics. In this study, we examined the immunohistochemical localization of IQ-ArfGEF/BRAG1 in the adult mouse retina using light and electron microscopy. IQ-ArfGEF/BRAG1 was distributed in a punctate manner and colocalized well with RIBEYE in both the outer and inner plexiform layers. Immunoelectron microscopic analysis showed that IQ-ArfGEF/BRAG1 was localized at the synaptic ribbons of photoreceptors. When heterologously expressed in HeLa cells, IQ-ArfGEF/BRAG1 was recruited to RIBEYE-containing clusters and formed an immunoprecipitable complex with RIBEYE. Furthermore, immunoprecipitation analysis showed that anti-IQ-ArfGEF/BRAG1 antibody efficiently pulled down RIBEYE from retinal lysates. These findings indicate that IQ-ArfGEF/BRAG1 is a novel component of retinal synaptic ribbons and forms a protein complex with RIBEYE.

Different effects of 26-week dietary intake of rapeseed oil and soybean oil on plasma lipid levels, glucose-6-phosphate dehydrogenase activity and cyclooxygenase-2 expression in spontaneously hypertensive rats.

We intended to determine whether or not dietary canola oil (CO) elevates plasma lipids and oxidative stress, since both of these are, possibly, related to the CO-induced life shortening through exacerbation of hypertension-associated vascular lesions found in stroke-prone spontaneously hypertensive rats (SHRSP). Spontaneously hypertensive rats (SHR) were used in this study to avoid a potential bias in the results due to the irregular death by stroke seen in SHRSP. SHR were fed for 26 weeks on a chow containing either, 10 wt/wt% of CO or soybean oil (SO), i.e., the control. Elevated plasma lipids and glucose-6-phosphate dehydrogenase (G6PD) activation in the liver and erythrocyte were found in SHR fed CO compared to that fed SO, while anti-oxidative enzymes other than G6PD were not activated. The CO diet brought about significant vascular lesions in the kidney, in which abundant cyclooxygenase-2 (COX-2) positive foci were immunochemically located in the juxtaglomerular apparatus. These results suggest that dietary CO induces a hyperlipidemic condition, in which G6PD may serve as an NADPH provider, and aggravates genetic diseases in SHR (also, probably, in SHRSP). The increased COX-2 expression indicates a role of renin-angiotensin-aldosterone system activation in the increased vascular lesions, whereas the effects of oxidative stress remain unclear.

A 90-day ad libitum administration toxicity study of oligoglucosamine in F344 rats.

A 90-day ad libitum administration toxicity study of oligoglucosamine (OG) was carried out using F344 rats of both sexes. The animals were divided into four groups of 20 animals each, 10 of each sex, and fed a diet containing 0, 0.04, 0.2 or 1.0 (w/w)% OG. During the administration period, no animals of either sex died or exhibited abnormal signs in the 0.04% OG and 0.2% OG groups. In the 1% OG group, in both sexes, erythema and swelling of the snout and forelimbs and loss of fur in the forelimbs were observed. On macroscopic observation, emaciation, swelling of the snout, auricles and forelimbs and alopecia of the forelimbs were also observed in 2-3 males of the 1% OG group. It was suggested that these topical abnormalities might be due to dermal responses to OG adhering to the skin and fur, which are easily soiled with saliva during grooming. In the animals of the 1% OG group, food consumption decreased, resulting in body weight gain being suppressed. This was found concomitantly with the abnormal findings mentioned above. Thus, feeding difficulties due to the topical lesions on the snout and forelimbs were thought to affect body weight. In hematology, platelet count, lymphocyte count and differential neutrophil count increased in males of the 1% OG group. These changes might be related to the dermal inflammation. Abnormalities in urinalysis and blood chemistry, as well as a small thymus, small spleen, dark spots or areas on the glandular stomach mucosa, pale Harderian glands and small testes in histopathology, were also observed in males in the 1% OG group. Whether or not all these changes were related only to the malnutrition remains to be elucidated. From these results, OG gave rise to no adverse effects in rats up to the dose level of 0.2 (w/w)%. Thus, the no observed adverse effect level was determined to be 0.2 (w/w)% for rats of either sex (124.0mg/kg/day in males, 142.0mg/kg/day in females).

A ninety-day feeding, subchronic toxicity study of oligo-N-acetylglucosamine in Fischer 344 rats.

Oligo-N-acetylglucosamine (OAG) is a hydrolyzed derivative of chitin that has been used as a sweetener in foods. Since, no information has been published about the safety of OAG, a 90-day feeding study was conducted, using F344 Fischer rats of both sexes, to characterize and evaluate the toxicity of OAG, and the results of the study are presented here. Dietary levels of 0% (control), 0.2%, 1%, and 5% OAG did not change any measurements in ophthalmological examinations, clinical signs, body weights, food consumption, hematology, blood biochemistry, urinalysis, necropsies, organ weights or histological examinations. The sole finding, which could not be clarified to be attributed to OAG or not, was a decrease in the relative weight of the submaxillary gland to body weight in the male animals given the 5% OAG diet. Although no lesions were found in either gross or histological examination in the present study, further studies using OAG levels higher than 5% might provide a clue to the mechanisms underlying the decreased organ weight observed here. Taken together, under the conditions in the present study, the No Observed Adverse Effect Level (NOAEL) for males was found to be 1% (0.641 g/kg/day); and that for females, 5% (3.64 /kg/day) or more, based on the lack of toxicological effects.

Dietary lipids impacts on healthy ageing.

Healthy ageing is gaining attention in the lipid nutrition field. As in vivo biomarkers of healthy ageing, we have evaluated the survival, learning/memory performance, and physical potencies in rodents fed a diet supplemented with high-linoleic acid (LNA, omega6) safflower oil or high-alpha-linolenic acid (ALA, omega3) perilla oil for long periods. The results suggested that perilla oil with a low omega6/omega3 ratio is beneficial for healthy ageing. In order to address this issue further, we determined the survival of stroke-prone SHR (SHRSP) rats fed a conventional rodent diet supplemented with 10% fat or oil. Survival was longer with omega3-rich oils compared with omega6-rich oils. However, some kinds of vegetable oils and hydrogenated oils shortened the survival of SHRSP rats to an unusual degree (ca. 40% compared with that of omega6-rich oil) that could not be accounted for by the fatty acid and phytosterol composition of the oils. The observed decrease in platelet counts was associated with pathological changes in the kidney and other organs. Dihydro-vitamin K1 is proposed as a likely candidate as a stroke-stimulating factor in hydrogenated oils. Thus, factors other than fatty acids (omega6/omega3 balance) and phytosterols must be taken into account when fats and oils are evaluated in relation to healthy ageing.

Tadalafil induces antiproliferation, apoptosis, and phosphodiesterase type 5 downregulation in idiopathic pulmonary arterial hypertension in vitro.

Idiopathic pulmonary arterial hypertension (IPAH) is a fatal disease of the pulmonary artery resulting from a currently unidentified etiology. IPAH is pathologically characterized as sustained vasoconstriction and vascular remodeling of the pulmonary artery. Vascular remodeling is mediated by enhanced proliferation and reduced apoptosis in pulmonary arterial smooth muscle cells (PASMCs). Based on its pathological mechanism, specific phosphodiesterase type 5 (PDE5) inhibitors have been used in the treatment of IPAH. In addition to sildenafil, tadalafil has been approved for the treatment of IPAH. However, the effects of tadalafil on excessive proliferation of IPAH-PASMCs currently remain unknown. In the present study, the in vitro pharmacological profiles of tadalafil for cell proliferation and apoptosis were assessed in IPAH-PASMCs using MTT, BrdU incorporation, and caspase 3/7 assays. Expression analyses revealed that PDE5 mRNA and protein expression levels were markedly higher in IPAH-PASMCs than in normal-PASMCs. The treatment with tadalafil inhibited the excessive proliferation of IPAH-PASMCs in a concentration-dependent manner with an IC50 value of 4.5µM. On the other hand, tadalafil (0.03-100µM) did not affect cell growth of PASMCs from normal subjects and patients with chronic thromboembolic pulmonary hypertension (CTEPH). In addition, tadalafil induced apoptosis in IPAH-PASMCs. The antiproliferative and apoptotic effects of tadalafil were markedly stronger than those of sildenafil and vardenafil. The upregulated expression of PDE5 in IPAH-PASMCs was significantly attenuated by a long-term treatment with tadalafil. Taken together, these results indicate that tadalafil attenuates vascular remodeling by inhibiting cell proliferation, promoting apoptosis, and downregulating PDE5 in IPAH-PASMCs, thereby ameliorating IPAH.

Exploration for unknown substances in rapeseed oil that shorten survival time of stroke-prone spontaneously hypertensive rats. Effects of super critical gas extraction fractions.

To identify the causative substances for the shortening of survival time by rapeseed (Canola) oil in stroke-prone spontaneously hypertensive rats (SHRSP), SHRSP were fed on a standard chow supplemented with 10 w/w% soybean oil (control), rapeseed oil, one of the fractions of rapeseed oil obtained by super critical gas extraction (SCE) under a pressure of 180-bar or 350-bar, at 40 degrees C, or the residue from the extraction (with 0.5% NaCl in drinking water). In another series of experiment, SHRSP were fed for 8 weeks on the above-mentioned diets without salt loading and autopsied. Fatty acid compositions in these diets were similar, except in the soybean oil diet, and phytosterol contents were: (diet containing) 180-bar fraction>residue>rapeseed oil>350-bar fraction>soybean oil. Survival times in the rapeseed oil, 350-bar fraction and residue groups were shorter than, whereas that in the 180-bar fraction was similar to in the soybean oil group. In the 8-week feeding experiment, chronic nephropathy was found frequently in the groups other than the soybean oil group. The heart weights were higher in the rapeseed oil and residue groups. Cerebral necrosis was found in the residue group. Taken together, the followings are concluded, (1) Neither the fatty acid composition, nor the amount of phytosterols in the diets appeared to be decisive in the shortening of life. (2) SCE appeared to produce a safe (180-bar) fraction, though it failed to separate clearly the causative substances into specific fractions. (3) The factors that facilitate the genetic disease of SHRSP appear to exist in rapeseed oil. However, they might not be identical to those responsible for the life-shortening, since there were no findings common across the rapeseed oil, 350-bar and residue groups, which showed similar life-shortening.