Low Amino Acid Score of Breakfast is Associated with the Incidence of Cognitive Impairment in Older Japanese Adults: A Community-Based Longitudinal Study.

BACKGROUND: The protein digestibility-corrected amino acid score (PDCAAS) represents the degree of utilizable dietary protein, namely the protein quality. The PDCAAS of a diet is required to be evaluated on a meal-by-meal basis, as food digestion and absorption occur with each meal intake. Although a positive association between protein intake and cognitive function has been reported, no study has investigated the association between PDCAAS of a diet and cognitive function. OBJECTIVES: To investigate the relationship between PDCAAS of a diet and cognitive impairment in older adults. DESIGN: Longitudinal epidemiological study. SETTING: Community-based setting. PARTICIPANTS: We analyzed 541 community-dwellers who participated in both baseline and follow-up survey. They were 60-83 years of age without cognitive impairment at baseline. MEASUREMENTS: Cognitive impairment was defined as a Mini-Mental State Examination (MMSE) score ≤27. Individual PDCAASs were calculated for each of three regular meals from the 3-day dietary records at baseline. Participants were classified into two groups according to the sex-specific tertiles (T1-T3) of the PDCAAS for each meal (i.e., T1 as the low score group and T2-T3 as the medium and high score group). The dependent variable was cognitive impairment observed after 4 years, and the explanatory variables were the PDCAAS groups for each meal (the medium and high group as the reference) and covariates (sex, age, body mass index, education, depressive symptoms, medical history, protein intake at each meal, and the MMSE score at baseline). Multivariable logistic regression analysis was performed to evaluate the low PDCAAS group for cognitive impairment after 4 years. RESULTS: A significant association was observed only between a low PDCAAS of breakfast and the incidence of cognitive impairment (the adjusted odds ratios [95% confidence intervals] of low PDCAAS for cognitive impairment for breakfast, lunch, and dinner were 1.58 [1.00-2.50], 0.85 [0.54-1.34], and 1.08 [0.71-1.65], respectively). CONCLUSION: A lower PDCAAS of breakfast, i.e., a diet with poor quality of protein, was associated with the incidence of cognitive impairment in older adults of the community.

Dietary Patterns and Muscle Mass, Muscle Strength, and Physical Performance in the Elderly: A 3-Year Cohort Study.

OBJECTIVES: Nutritional support effectively prevents and treats sarcopenia; however, the influence of overall dietary patterns on sarcopenia parameters is less investigated. This study aimed to determine the association between adherence to Mediterranean-style diet (MD), Dietary Approaches to Stop Hypertension (DASH), Japanese Food Guide Spinning Top (JFG-ST), and modified JFG-ST (mJFG-ST) and muscle mass, muscle strength, and physical performance in community-dwelling Japanese elderly. DESIGN AND SETTINGS: This prospective cohort study recruited individuals aged over 60 years from a community college in Nagoya, Japan. PARTICIPANTS AND MEASUREMENTS: A total of 666 participants were followed up annually from 2014 to 2017. Demographic data, anthropometric measurements, and sarcopenia parameters including walking speed (WS), hand grip strength in the dominant hand (HGS), and skeletal mass index (SMI) were recorded. Self-recall dietary intake was assessed using a validated food frequency questionnaire comprising 29 food groups. Adherence to MD, DASH, JFG-ST, and mJFG-ST was determined by tertiles. RESULTS: At baseline, the mean age of all participants (56.5% women) was 69.4±4.4 years. WS, HGS, and SMI were 1.4±0.2 (m/s), 28.9±8.1 (kg), and 6.7±1.0 (kg/m2), respectively. In longitudinal analysis, participants with higher JFG-ST adherence scores were more likely to have higher SMI (Q3 vs. Q1: mean difference, 0.048; p=0.04) after adjustment, and its benefits were more evident in men (Q2 vs. Q1: mean difference, 0.098; p=0.047; Q3 vs. Q1: mean difference, 0.091; p=0.017) than in women. WS and HGS were not associated with any type of dietary pattern. CONCLUSIONS: Adherence to JFG-ST was positively associated with SMI in Japanese community-dwelling elderly adults aged over 60 years, specifically in men. The country-specific dietary recommendations are required to be developed for sarcopenia prevention.

The Association between Dietary Amino Acid Intake and Cognitive Decline 8 Years Later in Japanese Community-Dwelling Older Adults.

OBJECTIVES: Previous studies have reported a relationship between low protein intake and cognitive decline and have suggested that this association may be related to specific amino acid intake. However, the effects of amino acid intake on the maintenance of cognitive function have yet to be clarified. We examined the longitudinal association between dietary amino acid intake and cognitive function in community-dwelling older adults. DESIGN: Longitudinal epidemiological study. SETTING: Community-based setting. PARTICIPANTS: This study comprised 427 study participants aged 60-82 years with no cognitive decline, defined as a Mini-Mental State Examination (MMSE) score of >27 at baseline, who also participated in a follow-up. The average and standard deviation of the follow-up period was 8.2 ± 0.3 years. MEASUREMENTS: Dietary intake was assessed using three-day dietary records at baseline. Participants were classified into quartiles (Q1-Q4) based on the intake of 19 amino acids for males and females. Next, we classified participants into Q1 and Q2-Q4 groups. Cognitive function was assessed using the MMSE both at baseline and at follow-up. Multivariable logistic regression models were used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for the associations between the Q1 group and cognitive decline (MMSE ≤27), using the Q2-Q4 group as a reference group. Covariates were age, sex, body mass index, years of education, severity of depressive symptoms, history of lifestyle diseases (hypertension, dyslipidemia, diabetes mellitus, stroke, and ischemic heart disease), energy intake (kcal/d), protein intake (g/d), and MMSE score at baseline. RESULTS: Cognitive decline was present in 133 (31.1%) participants. After adjustment for covariates, including total protein intake, the ORs (95% CIs) for cognitive decline were 2.40 (1.21-4.75) for lysine, 2.05 (1.02-4.09) for phenylalanine, 2.18 (1.09-4.34) for threonine, and 2.10 (1.06-4.15) for alanine. CONCLUSION: The results suggest that lysine, phenylalanine, threonine, and alanine intake is important for the maintenance of cognitive function in older people, independent of total protein intake.

The Japan-Multimodal Intervention Trial for Prevention of Dementia (J-MINT): The Study Protocol for an 18-Month, Multicenter, Randomized, Controlled Trial.

BACKGROUND/OBJECTIVES: The Japan-multimodal intervention trial for prevention of dementia (J-MINT) is intended to verify the effectiveness of multi-domain interventions and to clarify the mechanism of cognitive improvement and deterioration by carrying out assessment of dementia-related biomarkers, omics analysis and brain imaging analysis among older adults at high risk of dementia. Moreover, the J-MINT trial collaborates with partnering private enterprises in the implementation of relevant interventional measures. This manuscript describes the study protocol. DESIGN/SETTING: Eighteen-month, multi-centered, randomized controlled trial. PARTICIPANTS: We plan to recruit 500 older adults aged 65-85 years with mild cognitive impairment. Subjects will be centrally randomized into intervention and control groups at a 1:1 allocation ratio using the dynamic allocation method with all subjects stratified by age, sex, and cognition. INTERVENTION: The multi-domain intervention program includes: (1) management of vascular risk factors; (2) group-based physical exercise and self-monitoring of physical activity; (3) nutritional counseling; and (4) cognitive training. Health-related information will be provided to the control group every two months. MEASUREMENTS: The primary and secondary outcomes will be assessed at baseline, 6-, 12-, and 18-month follow-up. The primary outcome is the change from baseline to 18 months in a global composite score combining several neuropsychological domains. Secondary outcomes include: cognitive change in each neuropsychological test, incident dementia, changes in blood and dementia-related biomarkers, changes in geriatric assessment including activities of daily living, frailty status and neuroimaging, and number of medications taken. CONCLUSIONS: This trial that enlist the support of private enterprises will lead to the creation of new services for dementia prevention as well as to verify the effectiveness of multi-domain interventions for dementia prevention.

Combined Impact of Physical Frailty and Social Isolation on Rate of Falls in Older Adults.

OBJECTIVES: The aim of this study was to examine the impact of the combination of physical frailty and social isolation on falling in community-dwelling older adults. DESIGN: A cross-sectional study of data obtained at registration in a randomized control trial. SETTING: Community-based study of participants recruited from Toyota, Japan. PARTICIPANTS: 380 community-dwelling older adults (47.9% women, mean age = 72.3 ± 4.6 years). MEASUREMENTS: Participants were categorized as non-frail or pre-frail/frail based on the Fried frailty criteria (slowness, weakness, exhaustion, low activity, and weight loss). Social isolation was examined using the Lubben Social Network Scale (LSNS-6), and scores lower than 12 points indicated social isolation. Participants were divided into four groups depending on pre-frail/frail status and social isolation, and experiences of multiple falls over the past year were compared between the groups. RESULTS: Participants were classified into robust (n = 193), physical frailty (PF; n = 108), social isolation (SI; n = 43), and PF with SI (PF+SI; n = 36) groups. A total of 38 (10.0%) participants reported multiple falls. Logistic regression analysis showed that PF and SI groups were not independently associated with falling (PF: OR 1.64, 95% CI 0.65-4.16, SI: OR 2.25, 95% CI 0.77-6.58), while PF+SI group was significantly associated with falling compared with the robust group (OR 3.06, 95% CI 1.00-9.34, p = 0.049) after controlling for confounding factors. CONCLUSION: Our findings support the assertion that coexistence with physical frailty and social isolation were associated with falling in the older adults.

Cognitive Dysfunction in Urban-Community Dwelling Prefrail Older Subjects.

OBJECTIVES: A number of studies have reported that frailty is cross-sectionally associated with cognitive decline and is also a risk for future cognitive decline or dementia; however, there have been only a few studies that focus on the association between prefrailty and cognitive dysfunction. In the current study, we investigated the association between prefrailty and cognition. DESIGN: A cross-sectional study of the data obtained at registration in a randomized control trial. SETTING: Toyota, Japan. PARTICIPANTS: Community-dwelling older subjects (male 54.6%) who had cognitive complaints. MEASUREMENTS: A battery of neuropsychological and physical assessments were performed. Prefrailty was defined as exhibiting one or two of the five Fried criteria (weight loss, exhaustion, weakness, slow gait speed and low physical activity). We performed a multiple regression analysis to investigate the associations of cognitive performance with prefrailty, adjusting for the factors that were significantly different between the robust and prefrailty groups. To assess the cognitive attributes that were significantly associated with prefrailty, logistic analysis was performed to see if one specific criterion of the five frailty criteria was associated with cognitive performance. RESULTS: The study subjects included 183 prefrail and 264 robust individuals. The prefrail subjects with cognitive complaints were older, less educated, more depressive, and more likely to have diabetes mellitus than the robust subjects. The prefrail subjects had lower performance in a wide-range of cognitive domains, and after adjustments for age, education, depressive mood, and diabetes mellitus, prefrailty was associated with a decline in delayed memory and processing speed. Among the components of the Fried criteria, slow gait speed and loss of activity were significantly associated with slow processing speed as assessed by the digit symbol substitution test. CONCLUSION: The current results demonstrated that prefrailty was associated with worse memory and processing speed performance, but not with other cognitive domains.

Taxol inhibits neointimal smooth muscle cell accumulation after angioplasty in the rat.

Despite significant improvements in the primary success rate of the medical and surgical treatments for atherosclerotic disease, including angioplasty, bypass grafting, and endarterectomy, secondary failure due to late restenosis continues to occur in 30-50% of individuals. Restenosis and the later stages in atherosclerotic lesions are due to a complex series of fibroproliferative responses to vascular injury involving potent growth-regulatory molecules (such as platelet-derived growth factor and basic fibroblast growth factor) and resulting in vascular smooth muscle cell (VSMC) proliferation, migration, and neointimal accumulation. We show here, based on experiments with both taxol and deuterium oxide, that microtubules are necessary for VSMCs to undergo the multiple transformations contributing to the development of the neointimal fibroproliferative lesion. Taxol was found to interfere both with platelet-derived growth factor-stimulated VSMC migration and with VSMC migration and with VSMC proliferation, at nanomolar levels in vitro. In vivo, taxol prevented medial VSMC proliferation and the neointimal VSMC accumulation in the rat carotid artery after balloon dilatation and endothelial denudation injury. This effect occurred at plasma levels approximately two orders of magnitude lower than that used clinically to treat human malignancy (peak levels achieved in this model were approximately 50-60 nM). Taxol may therefore be of therapeutic value in preventing human restenosis with minimal toxicity.

Neointima formation in a restenosis model is suppressed in midkine-deficient mice.

Neointima formation is a common feature of atherosclerosis and restenosis after balloon angioplasty. To find a new target to suppress neointima formation, we investigated the possible role of midkine (MK), a heparin-binding growth factor with neurotrophic and chemotactic activities, in neointima formation. MK expression increased during neointima formation caused by intraluminal balloon injury of the rat carotid artery. Neointima formation in a restenosis model was strongly suppressed in MK-deficient mice. Continuous administration of MK protein to MK-deficient mice restored neointima formation. Leukocyte recruitment to the vascular walls after injury was markedly decreased in MK-deficient mice. Soluble MK as well as that bound to the substratum induced migration of macrophages in vitro. These results indicate that MK plays a critical role in neointima formation at least in part owing to its ability to mediate leukocyte recruitment.

Deficiency of the cysteine protease cathepsin S impairs microvessel growth.

During angiogenesis, microvascular endothelial cells (ECs) secrete proteinases that permit penetration of the vascular basement membrane as well as the interstitial extracellular matrix. This study tested the hypothesis that cathepsin S (Cat S) contributes to angiogenesis. Treatment of cultured ECs with inflammatory cytokines or angiogenic factors stimulated the expression of Cat S, whereas inhibition of Cat S activity reduced microtubule formation by impairing cell invasion. ECs from Cat S-deficient mice showed reduced collagenolytic activity and impaired invasion of collagens type I and IV. Cat S-deficient mice displayed defective microvessel development during wound repair. This abnormal angiogenesis occurred despite normal vascular endothelial growth factor and basic fibroblast growth factor levels, implying an essential role for extracellular matrix degradation by Cat S during microvessel formation. These results demonstrate a novel function of endothelium-derived Cat S in angiogenesis.

Risk Factors for the Discontinuation of Home Medical Care among Low-functioning Older Patients.

OBJECTIVES: Older patients receiving home medical care often have declining functional status and multiple disease conditions. It is important to identify the risk factors for care transition events in this population in order to avoid preventable transitions. In the present study, therefore, we investigated the factors associated with discontinuation of home medical care as a potentially preventable care transition event in older patients. METHODS: Baseline data for participants in the Observational study of Nagoya Elderly with HOme MEdical (ONEHOME) study and data on the mortality, institutionalization, or hospitalisation of the study participants during a 2-year follow-up period were used. Discontinuation of home care was defined as admission to a hospital for any reason, institutionalization, or death. Univariate and multivariate Cox hazard models were used to assess the association of each of the factors with the discontinuation of home care during the observational period. The covariates included in the multivariate analysis were those significantly associated with the discontinuation of home care at the level of P<0.05 in the univariate analysis. RESULTS: The univariate Cox hazard model revealed that a low hemoglobin level (< 11g/dL), low serum albumin level (< 3g/dL), higher Charlson Comorbidity Index score, and low Mini Nutritional Assessment Short Form score (< 7) were significantly associated with the discontinuation of home care. A multivariate Cox hazard model including these four factors demonstrated that all four were independently associated with home-care discontinuation. CONCLUSIONS: The present results demonstrated that anemia, hypoalbuminemia, malnourishment, and the presence of serious comorbidities were associated with the discontinuation of home medical care among low-functioning older patients.

Lipid peroxide and transition metals are required for the toxicity of oxidized low density lipoprotein to cultured endothelial cells.

The toxicity of oxidized low density lipoprotein (Ox-LDL) to cultured vascular endothelial cells was investigated. The modification of low density lipoprotein (LDL) by copper led to the production of thiobarbituric acid-reacting substance (TBARS) and lipid hydroperoxide (LPO). TBARS was distributed not only in lipoprotein, but also in the aqueous phase, whereas LPO was observed only in the lipoprotein particle. During the incubation of LDL with copper, the copper bound to lipoprotein and formed a complex. The toxicity of products resulting from the oxidation of LDL to endothelial cells was recognized in Ox-LDL particles, not in the aqueous phase. Following dialysis of Ox-LDL against EDTA, copper which had bound to the Ox-LDL particle was released and the toxicity of Ox-LDL disappeared. The addition of copper to the dialyzed Ox-LDL restored the cytotoxicity. To a lesser extent this effect was also observed with the addition of iron. A study of the time-course of LDL oxidation showed that the toxicity of Ox-LDL depends upon the level of LPO, not upon the content of TBARS, the extent of negative charge or the protein adduct of aldehydes. These results demonstrate that transition metal is required for Ox-LDL toxicity and that the toxic moiety of the products resulting from LDL oxidation is LPO associated with the Ox-LDL particle.

Oxidation of low-density lipoprotein by copper and iron in phosphate buffer.

We examined the effect of phosphate buffer on the iron- and copper-catalyzed peroxidation of low-density lipoprotein (LDL). The incubation of LDL with CuSO4 in 0.15 M NaCl led to the peroxidation of LDL as evidenced by the detection of thiobarbituric acid-reactive substances (TBARS) and lipid hydroperoxides (LPO). The peroxidation of LDL was also observed with FeSO4 and FeCl3 in 0.15 M NaCl, although there was a lag phase with FeCl3. In 10 mM phosphate buffer, the peroxidation of LDL was observed with CuSO4 to an extent similar to that in 0.15 M NaCl. However, the peroxidation induced by incubation with FeSO4 and FeCl3 was significantly inhibited in phosphate buffer. Iron and copper each formed a complex with lipoprotein during incubation with LDL in 0.15 M NaCl. Although no effect on the formation of copper-LDL complex was observed in phosphate buffer, the formation of iron-LDL complex was reduced in the buffer. These observations suggest there are marked differences in the peroxidation of LDL and in the formation of complexes with LDL between iron and copper in phosphate buffer.

Role of lipoprotein-copper complex in copper catalyzed-peroxidation of low-density lipoprotein.

The oxidative modification of low-density lipoprotein (LDL) is suggested to play an important role in the pathogenesis of atherosclerosis. The present study examined the role of the formation of LDL-copper (Cu) complex in the peroxidation of LDL. The amount of copper bound to LDL increased during incubation performed with increasing concentrations of CuSO4. More than 80% of the copper bound to the LDL particle was observed in the protein phase of LDL, suggesting that most of the copper ions formed complexes with the ligand-binding sites of apoprotein. The addition of histidine (1 mM), known to form a high affinity complex with copper, and EDTA (1 mM), a metal chelator, during the incubation of LDL with CuSO4 prevented the formation of both thiobarbituric acid-reactive substances (TBARS) and LDL-Cu complexes. EDTA inhibited the copper-catalyzed ascorbate oxidation whereas histidine had no effect, suggesting that the copper within the complex with histidine is available to catalyze the reaction, in contrast to EDTA. These observations indicate that the preventive effect of histidine on the copper-catalyzed peroxidation of LDL is not simply mediated by chelating free copper ions in aqueous phase. Evidence that copper bound to LDL particle still has a redox potential was provided by the observed increase in TBARS content during incubation of LDL-Cu complexes in the absence of free copper ions. The addition of either histidine or EDTA to LDL-Cu complexes inhibited the formation of TBARS by removing copper ions from the LDL forming the corresponding complexes. However, there still remained small amounts of copper in the LDL particles following the treatment of LDL-Cu complexes with histidine or EDTA. The copper ions remaining in the LDL particle lacked the ability to catalyze LDL peroxidation, suggesting that there may be two types of copper binding sites in LDL: tight-binding sites, from which the copper ions are not removed by chelation, and weak-binding sites, from which copper ions are easily removed by chelators. The formation of TBARS in the LDL preparation during incubation with CuSO4 was comparable to the incubation with FeSO4. In contrast, the formation of TBARS in the LDL-lipid micelles by CuSO4 was nearly eliminated even in the presence of ascorbate to promote metal-catalyzed lipid peroxidation, although a marked increase in TBARS content was observed in the LDL-lipid micelles with FeSO4, and with FeCl3 in the presence of ascorbate.(ABSTRACT TRUNCATED AT 400 WORDS)

VEGF protects against oxidized LDL toxicity to endothelial cells by an intracellular glutathione-dependent mechanism through the KDR receptor.

Although the accumulation of vascular endothelial growth factor (VEGF) has been observed in human atherosclerotic lesions, the exact role of this growth factor in atherogenesis remains unknown. We hypothesized that VEGF in the vascular wall might have a preventive effect on endothelial cell damage during atherosclerosis. To test our hypothesis, we examined whether VEGF protects against the toxicity of oxidized low density lipoprotein (Ox-LDL) in cultured endothelial cells derived from bovine aortas (BAECs). Preincubation of BAECs with VEGF prevented Ox-LDL-induced toxicity in a preincubation time- and VEGF concentration-dependent manner. Addition of N(omega)-nitro-L-arginine methyl ester, a nitric oxide synthase inhibitor, did not reverse the protective effect of VEGF on Ox-LDL toxicity. Incubation of BAECs with VEGF increased intracellular glutathione (GSH) content in a time-dependent manner. Combined addition of VEGF and L-buthionine sulfoximine, a GSH synthesis inhibitor, reversed both GSH levels and the protective effect of VEGF on Ox-LDL-induced cytotoxicity. Placenta growth factor, which ligates to the VEGF Flt-1 receptor but not KDR/Flk-1, failed to prevent Ox-LDL toxicity and had no effect on intracellular GSH levels. An anti-KDR antibody completely blocked these beneficial activities of VEGF. These results suggest that VEGF prevents Ox-LDL-induced endothelial cell damage via an intracellular GSH-dependent mechanism through the KDR/Flk-1 receptor.

Probucol as an antioxidant and antiatherogenic drug.

Recently, interest has increased in the hypothesis that low-density lipoprotein (LDL) modified by oxidation may lead to the initiation and to the development of atherosclerosis. In vitro studies of cellular interactions with LDL have revealed that various cells, including endothelial cells and smooth muscle cells, can oxidize LDL. The biochemical changes in LDL may further enhance its atherogenic potential. In addition to these in vitro studies, there is in vivo evidence for oxidized LDL in atherosclerotic lesions and for circulating antibodies against oxidized LDL. Probucol, 4,4'-(isopropylidenedithio)bis(2,6-di-tert-butylphenol), is a widely used cholesterol-lowering drug. Recently, there has been accumulating evidence for other mechanisms of probucol's antiatherogenic effects apart from cholesterol-lowering action. Attention has especially focused on probucol's antioxidant action in the mechanism of antiatherogenesis. In the present article, we will summarize the antiatherogenic and antioxidant actions of probucol.

Fibrinogen is chemotactic for vascular smooth muscle cells.

We studied the effect of fibrinogen on the migration of bovine aortic smooth muscle cells in culture, using a Neuro Probe 48-well micro chemotaxis chamber. Fibrinogen stimulated the migration of the cells dose-dependently at concentrations from 30 to 1000 micrograms/ml. A modified checkerboard analysis of the response demonstrated that the effect was largely chemotactic in nature. The present results suggest that fibrinogen may play an important role in the pathogenesis of arterial intimal thickening and atherosclerosis.

Low level hyperlipidemia impairs endothelium-dependent relaxation of porcine coronary arteries by two mechanisms. Functional change in endothelium and impairment of endothelium-dependent relaxation by two mediators.

We evaluated the effect of a low level of hyperlipidemia and the effects of in vitro exposure to atherogenic lipoproteins (LDL, VLDL) on the vascular responsiveness of isolated porcine coronary arteries. Firstly we studied the change in vascular responsiveness induced by feeding a cholesterol-rich diet to pigs for 4 and 9 weeks (C4 and C9 pigs). The serum cholesterol level in pigs fed a cholesterol-rich diet reached 218.5 +/- 32.9 mg/dl compared with 85.5 +/- 8.4 mg/dl in the controls. Segments of the left descending coronary artery were examined. The contraction induced by KCl or prostaglandin F2 alpha was not altered significantly by hypercholesterolemia nor was the relaxation induced by the Ca2+ ionophore, A23187, or by nitroglycerin. Endothelium-dependent relaxation (EDR) evoked by high, but not low, concentrations of bradykinin was reduced in the C4 pigs as compared with those in normal animals. EDRs evoked by bradykinin, substance P, and serotonin were significantly reduced in C9 pigs. Histologically, as observed by light and electron microscopy, fatty changes or intimal thickenings were not seen in the coronary arteries of the C4 pigs. Minimal changes (intimal thickening and fragmentation of internal elastic lamina) were observed only in parts of arteries of the C9 pigs. Secondly, the direct effects of LDL and VLDL on vascular responsiveness were studied. Although preincubation with LDL inhibited the EDR caused by exposure to bradykinin and A23187 in the coronary arteries of normal and cholesterol-fed pigs, preincubation with LDL inhibited the arterial relaxation induced by exposure to substance P or serotonin in both the C4 and the C9 pigs, but not in the control animals. The degree of inhibition was especially marked in the C9 pigs. The inhibitory effect of VLDL on EDR was weaker than that of LDL. Indomethacin (5 microM) did not alter this inhibitory effect of lipoproteins. Neither LDL nor VLDL had any effect on the vascular relaxation induced by nitroglycerin. These results are consistent with the idea that endothelium-dependent arterial relaxation is attenuated even at the very early stage of cholesterol-induced atherosclerosis. Atherogenic lipoproteins may further impair the decreased EDR in the arteries of hyperlipidemic pigs by two factors: one released on stimulation with bradykinin and the calcium ionophore A23187, the other released on stimulation with substance P and serotonin.

Effects of fibrinogen and fibrin on the migration of vascular smooth muscle cells in vitro.

The migration of vascular smooth muscle cells from the media into the intima and their proliferation in the intima play an important role in the pathogenesis of atherosclerosis. We examined the effects of fibrinogen and fibrin on the migration of cultured bovine aortic smooth muscle cells using a modified Boyden chamber assay. The cells migrated to a gradient of soluble fibrinogen. Checkerboard analysis indicated that the effect was largely directional in nature (chemotaxis). The cells also migrated in a dose-dependent manner to a gradient of substrate-bound fibrinogen (haptotaxis). Fibrin, converted from substrate-bound fibrinogen by thrombin, also induced haptotaxis of smooth muscle cells. These observations suggest that, by recruiting smooth muscle cells from the media into the intima, fibrinogen and fibrin may be involved in the pathogenesis of arterial intimal thickening, atherosclerosis, and the organization of a thrombus.

Rho-Rho kinase is involved in smooth muscle cell migration through myosin light chain phosphorylation-dependent and independent pathways.

Although Rho, a small GTPase, has been demonstrated to play an important role in the smooth muscle contraction and relaxation, little is known about the involvement of Rho protein in smooth muscle cell (SMC) migration. In this study the role of Rho-Rho kinase pathway was examined in SMC migration induced by platelet-derived growth factor (PDGF) and lysophosphatidic acid (LPA). C3 transferase, a specific inhibitor of Rho, blocked SMC migration induced by PDGF and LPA. Y-27632, a specific inhibitor of Rho kinase, a direct target molecule of Rho, inhibited PDGF and LPA-induced SMC migration in a concentration dependent manner. Although rapid increase in myosin light chain (MLC) phosphorylation in SMC treated with LPA was observed, no enhanced MLC phosphorylation was detected in response to PDGF. Y-27632 suppressed LPA-induced as well as basal level of MLC phosphorylation. ML-9, a specific inhibitor of myosin light chain kinase (MLCK), inhibited PDGF and LPA-induced SMC migration without the suppression of MLC phosphorylation at 5 min incubation, suggesting that MLCK may contribute to SMC migration via mechanism other than MLC phosphorylation. These results suggest that Rho-Rho kinase pathway is implicated in SMC migration and that different signaling pathways downstream of Rho-Rho kinase may be involved in LPA and PDGF-induced SMC migration. MLC phosphorylation via Rho-Rho kinase pathway appears to be implicated in LPA-dependent SMC migration. Whereas PDGF-mediated SMC migration is independent of increased MLC phosphorylation and other target molecules downstream of Rho-Rho kinase seem to be involved.