Facial pigmentation as a biomarker of carotid atherosclerosis in middle-aged to elderly healthy Japanese subjects.

BACKGROUND/PURPOSE: Perceived age may be a better predictor of mortality rate than chronological age. We have demonstrated that perceived age was a significant biomarker for carotid atherosclerosis in Japanese. However, it remains to be determined which skin parameter is associated with atherosclerosis. The purpose of this study is to analyze the relationship between 10 facial skin-aging parameters and atherosclerosis in 169 middle-aged to elderly Japanese women who participated. METHODS: Facial photographs were taken under a shadowless lamp from three directions using a high-resolution digital camera. The digital images of each subject were analyzed using computer software and various parameters of skin aging such as pigmentation, wrinkles, and skin color were quantified. Carotid intima-media thickness (IMT) and brachial-ankle pulse wave velocity (baPWV) were measured as indices for atherosclerosis. RESULTS: Facial pigmentation showed a significant correlation with carotid IMT, even after correction for age (r = 0.13, P = 0.03), and with visceral fat area. Stepwise regression analysis indicated that facial pigmentation was associated with carotid IMT via visceral fat area. CONCLUSION: Facial pigmentation may be a useful biomarker for carotid atherosclerosis in Japanese women.

Missense variants of the alanine: glyoxylate aminotransferase 2 gene correlated with carotid atherosclerosis in the Japanese population.

Alanine:glyoxylate aminotransferase 2 (AGXT2; EC 2.6.1.44) degrades asymmetric dimethylarginine (ADMA), a competitive inhibitor of nitric oxide (NO) synthase. Increased ADMA, reduced NO, and hypertension are shown in Agxt2 knockout mice. There are four single nucleotide polymorphisms (rs37370, rs37369, rs180749, and rs16899974) with which AGXT2 activity changes in humans and may be related to vulnerability of vascular sclerosis. To examine the relationship between them, we studied the functional haplotypes of the AGXT2 gene and decided their relationship with arteriosclerotic changes via carotid intima-media thickness (carotid IMT) in Japanese subjects. Genotyping of those polymorphisms and the carotid IMT in 1,426 Japanese subjects were then evaluated. Subjects with C-A-A-A haplotype (rs37370, rs37369, rs180749, rs16899974) showed low AGXT2 activity (P<0.0001; Pearson’s correlation coefficients: 0.497). The C-A-A-A haplotype was significantly associated with mean carotid IMT (P=0.049) and max carotid IMT (P=0.004). Subjects with two C-A-A-A haplotypes exhibited thicker mean carotid IMT (P=0.022) and maximum carotid IMT (P=0.001). In multiple regression analysis, subjects with two C-A-A-A haplotypes were independently and positively associated with mean carotid IMT (P=0.02) and maximum IMT (P=0.005) after correction. There was a significant correlation between the functional variants in the AGXT2 gene and carotid IMT in Japanese. The AGXT2 genotype may be an important factor underlying atherosclerosis.

Vildagliptin preserves the mass and function of pancreatic ß cells via the developmental regulation and suppression of oxidative and endoplasmic reticulum stress in a mouse model of diabetes.

AIM: We investigated the molecular mechanisms by which vildagliptin preserved pancreatic ß cell mass and function. METHODS: Morphological, biochemical and gene expression profiles of the pancreatic islets were investigated in male KK-A(y) -TaJcl(KK-A(y) ) and C57BL/6JJcl (B6) mice aged 8 weeks which received either vildagliptin or a vehicle for 4 weeks. RESULTS: Body weight, food intake, fasting blood glucose, plasma insulin and active glucagon-like peptide-1 were unchanged with vildagliptin treatment in both mice. In KK-A(y) mice treated with vildagliptin, increased plasma triglyceride (TG) level and islet TG content were decreased, insulin sensitivity significantly improved, and the glucose tolerance ameliorated with increases in plasma insulin levels. Furthermore, vildagliptin increased glucose-stimulated insulin secretion, islet insulin content and pancreatic ß cell mass in both strains. By vildagliptin, the expression of genes involved in cell differentiation/proliferation was upregulated in both strains, those related to apoptosis, endoplasmic reticulum stress and lipid synthesis was decreased and those related to anti-apoptosis and anti-oxidative stress was upregulated, in KK-A(y) mice. The morphological results were consistent with the gene expression profiles. CONCLUSION: Vildagliptin increases ß cell mass by not only directly affecting cell kinetics but also by indirectly reducing cell apoptosis, oxidative stress and endoplasmic reticulum stress in diabetic mice.

Association of Chr17q25 with cerebral white matter hyperintensities and cognitive impairment: the J-SHIPP study.

BACKGROUND AND PURPOSE: A recent genome-wide association study has successfully identified several genetic variations in the Chr17q25 locus as susceptible genotypes for white matter hyperintensities. We report the first replication study in subjects of non-European origin. We also investigated possible associations with other asymptomatic cerebrovascular diseases and cognitive function. METHODS: Study subjects were 1190 general Japanese persons (66.0 ± 8.9 years old). Asymptomatic cerebrovascular damage, including lacunar infarctions, microbleeds, periventricular hyperintensity and deep and subcortical white matter hyperintensity (DSWMH), was evaluated by brain magnetic resonance imaging. RESULTS: A polymorphism rs3744028 was significantly associated with DSWMH grade (P = 0.015) but not periventricular hyperintensity, lacunar infarction, and microbleeds. Although age, hypertension, insulin resistance, B-type natriuretic peptide, and carotid atherosclerosis were also correlated with DSWMH, association of the genotype was independent of these environmental risk factors. In contrast, the risk allele had a protective effect against reduced cognitive function. CONCLUSION: Susceptibility of the 17q25 locus may be conserved beyond ethnic differences. Genetic variants may have bipolar effects on brain histological and functional changes.

Adipose tissue reduction in mice lacking the translational inhibitor 4E-BP1.

All nuclear-encoded mRNAs contain a 5' cap structure (m7GpppN, where N is any nucleotide), which is recognized by the eukaryotic translation initiation factor 4E (eIF4E) subunit of the eIF4F complex. The eIF4E-binding proteins constitute a family of three polypeptides that reversibly repress cap-dependent translation by binding to eIF4E, thus preventing the formation of the eIF4F complex. We investigated the biological function of 4E-BP1 by disrupting its gene (Eif4ebp1) in the mouse. Eif4ebp1-/- mice manifest markedly smaller white fat pads than wild-type animals, and knockout males display an increase in metabolic rate. The males' white adipose tissue contains cells that exhibit the distinctive multilocular appearance of brown adipocytes, and expresses the uncoupling protein 1 (UCP1), a specific marker of brown fat. Consistent with these observations, translation of the peroxisome proliferator-activated receptor-gamma co-activator 1 (PGC1), a transcriptional co-activator implicated in mitochondrial biogenesis and adaptive thermogenesis, is increased in white adipose tissue of Eif4ebp1-/- mice. These findings demonstrate that 4E-BP1 is a novel regulator of adipogenesis and metabolism in mammals.

Creatinine to Cystatin C Ratio as a Marker of Bone Property in Older Adults: The J-SHIPP Study.

OBJECTIVES: To clarify whether serum creatinine to cystatin C ratio (CCR), a marker of muscle mass and muscle function may be used as a simple marker of bone property. DESIGN: A cross-sectional analysis. SETTING: A general population-based observation study. PARTICIPANTS: 1,606 middle-aged to elderly (≥50 years, mean age: 66.9 ± 7.5 years old) men (n = 642) and post-menopausal women (n = 964). MEASUREMENT: Speed of sound (SOS) at the calcaneal bone was used as a surrogate marker of bone mineral density. The cross-sectional area of the muscle at the mid-thigh was measured using computed tomography. RESULTS: There was significant linear correlation between the quartiles of CCR and SOS (Q1: 1,495 ± 25, Q2: 1,499 ± 24, Q3: 1,507 ± 26, Q4: 1,511 ± 25 m/sec; P < 0.001) even in a sex-separated analysis. This association was independent of major covariates (Q1: ß = -0.126, P < 0.001; Q2: ß = -0.096, P = 0.001; Q3: ß = -0.022; P = 0.412, Q4: reference) and the mid-thigh muscle mass, while creatinine alone or eGFR did not show clear association with SOS. CONCLUSION: The CCR may be used as a simple marker of bone property independently of muscle mass in a general population with preserved renal function.

Prognostic significance of FTO genotype in the development of obesity in Japanese: the J-SHIPP study.

OBJECTIVE: Susceptibility of fat mass and obesity-associated (FTO) gene polymorphisms to obesity has been reported in various populations. Polymorphisms in the melanocortin 4 receptor (MC4R) gene were recently explored as another susceptible locus. However, prognostic significance of these genetic variations has not been fully elucidated. Here, we investigated the involvement of FTO rs9939609 and MC4R rs17782313 polymorphisms in the development of obesity. Association with type 2 diabetes mellitus (T2DM) was also investigated. SUBJECTS: We analyzed 2806 community-dwelling middle-aged to elderly subjects (61+/-14 years). Clinical parameters were obtained from the subjects' personal health records, evaluated at their annual medical check-up. RESULTS: FTO genotype was significantly associated with current body mass index (BMI; TT 23.2+/-3.2, TA 23.7+/-3.2, AA 24.4+/-3.2 kg m(-2), P=2.5 x 10(-6)) and frequency of obesity (26.6, 32.0, 43.0% respectively, P=2.0 x 10(-4)). Age- and sex-adjusted odds ratio for obesity was 1.30 (P=0.004) in TA and 2.07 (P=0.002) in AA genotype. During the 9.4 years comprising the follow-up period, 214 new cases of obesity were diagnosed among 1718 subjects whose retrospective data were available. A allele frequency of the FTO genotype was significantly higher in subjects who developed obesity (22.2, 15.8%, P=0.001), Age-, sex- and initial BMI-adjusted odds ratio for the development of obesity was 1.46 (95% confidence interval, 1.04-2.04) (P=0.031). However, association studies and meta-analysis of T2DM did not actively support the involvement of FTO genotype. No significant differences were observed between the MC4R genotype and BMI (P=0.015), and the frequency of obesity (P=0.284). CONCLUSION: FTO genotype is an independent risk factor for future development of obesity.

Activity-dependent transfer of brain-derived neurotrophic factor to postsynaptic neurons.

Neurotrophins such as brain-derived neurotrophic factor (BDNF) are thought to be transferred from post- to presynaptic neurons and to be involved in the formation and plasticity of neural circuits. However, direct evidence for a transneuronal transfer of BDNF and its relation to neuronal activity remains elusive. We simultaneously injected complementary DNAs of green fluorescent protein (GFP)-tagged BDNF and red fluorescence protein into the nucleus of single neurons and visualized expression, localization, and transport of BDNF in living neurons. Fluorescent puncta representing BDNF moved in axons in the anterograde direction, though some moved retrogradely, and transferred to postsynaptic neurons in an activity-dependent manner.

Comparative aspects on the role of polypyrimidine tract-binding protein in internal initiation of hepatitis C virus and picornavirus RNAs.

We compared the effects of polypyrimidine tract-binding protein (PTB) on hepatitis C virus (HCV genotype IIa), encephalomyocarditis virus (EMCV) and poliovirus internal ribosome entry site (IRES) activities in vitro. It bound strongly to EMCV IRES, but weakly to PV and HCV RNAs. PV IRES showed the strongest dependency to PTB and it showed less than one-tenth of IRES activity after the immuno-depletion of PTB from HeLa S10 lysate with pre-coated anti-PTB IgG beads, comparing to the normal IgG beads-treated S10 lysate. EMCV IRES activity was approximately 40% of that of normal control after PTB depletion. Especially, HCV IRES activity was approximately 95%, and most weekly affected by the depletion of PTB. Repletion of PTB to depleted S10 lysate restored activities of PV and EMCV IRESs. The data suggest that PTB plays an important role in picornaviral IRESs, but not in HCV IRES.

Durability of protective effect of dulaglutide on pancreatic ß-cells in diabetic mice: GLP-1 receptor expression is not reduced despite long-term dulaglutide exposure.

AIMS: It is well-known that chronic exposure to large amounts of ligand leads to downregulation of its receptor. It is not known, however, whether a GLP-1R agonist downregulates its receptor. For this reason, our study examined whether GLP-1R expression is reduced after long-term exposure to dulaglutide (Dula) in non-diabetic and diabetic mice. METHODS: Seven-week-old male db/db and db/m mice were given either Dula (0.6mg/kg×2/week) or a control vehicle (CTL) for 17 weeks. Various metabolic parameters, such as glucose-stimulated insulin secretion (GSIS), insulin and TG content in islets, were evaluated after the intervention. ß-cell-related gene expression was also analyzed by real-time RT-PCR. RESULTS: In db/m mice, GLP-1R expression in ß-cells did not decrease, not even after long-term administration of Dula, compared with control mice, while GLP-1R expression in 24-week-old db/db mice treated with Dula was augmented, rather than downregulated, compared with 24-week-old CTL db/db mice. This was probably due to improved glycaemic control. In db/db mice treated with Dula, food intake and blood glucose levels were significantly decreased up to 24 weeks of age compared with CTL db/db mice, and their expression levels of various ß-cell-related genes, insulin content and GSIS were also enhanced. In contrast, oxidative and endoplasmic reticulum stress, inflammation, fibrosis and apoptosis were suppressed with Dula treatment. CONCLUSION: Dula exerts beneficial effects on glycaemic control and has long-lasting protective effects on pancreatic ß-cells. GLP-1R expression levels were not reduced at all in non-diabetic as well as diabetic mice despite long-term dulaglutide exposure.

Platelet-derived growth factor induces apoptosis in vascular smooth muscle cells: roles of the Bcl-2 family.

Apoptosis (programmed cell death) is observed in vascular smooth muscle cells (VSMC) in atherosclerotic lesions and stenotic lesions after injury, and modulates the cellularity of these lesions. It is recognized that cell growth and apoptosis are two linked processes. Platelet-derived growth factor (PDGF) induces VSMC proliferation and migration in vitro. We studied the effect of PDGF on apoptosis in VSMC. Cultured rat VSMC were treated with PDGF-AA or PDGF-BB. PDGF-BB induced cell death in cultured VSMC in a time- and dose-dependent manner, but PDGF-AA did not. Gel electrophoresis of genomic DNA and in situ DNA labeling confirmed that the cell death induced by PDGF-BB is apoptosis. PDGF-BB treatment reduced bcl-2 mRNA and bcl-xl mRNA expression, in contrast, induced bcl-xs mRNA expression, linked with the induction of apoptosis in cultured VSMC.

Establishment of swine interleukin-6 sandwich ELISA.

We established a sandwich enzyme-linked immunosorbent assay (ELISA) for swine interleukin-6 (SwIL-6), which was applied for detection of SwIL-6 in vitro and in vivo. Anti-SwIL-6 rabbit- and goat-polyclonal antibodies, and monoclonal antibody (mAb) were prepared, conforming that all of the antibodies were reactive with recombinant SwIL-6 by Western blotting and indirect ELISA. A sandwich ELISA was developed using the mAb as a capture antibody and biotinylated goat-polyclonal antibody as a detection antibody. The detection limit of the sandwich ELISA for rSwIL-6 was 49pg/ml and did not show cross-reactivity with swine IL-1b, IL-4, IL-8, IL-18, IL-12, and IFN-g. Using the ELISA, SwIL-6 was detected in culture medium of the monocytes stimulated with PHA-P and PMA, and the plasma or the bronchoalveolar lavage fluid (BALF) of pigs experimentally infected with Actinobacillus pleuropneumoniae or Mycoplasma hyopneumoniae. This ELISA for SwIL-6 may be useful for understanding the role of this cytokine in various swine diseases.

Expression and characterization of the recombinant swine interleukin-6.

The swine interleukin-6 (SwIL-6) cDNA was cloned by RT-PCR and each expression system of recombinant SwIL-6 in Escherichia coli, insect cells, and mammalian cells was developed. Recombinant SwIL-6 produced in bacteria was applied for generation of the polyclonal antibodies. The rSwIL-6 was purified from supernatant of insect cells with a Q-sepharose or anti-SwIL-6 monoclonal antibody based immunoaffinity column. The antibodies showed that the molecular weight of rSwIL-6 was approximately 26kDa in E. coli, 25, 26, 30kDa in insect cells, and 26 and 30kDa in mammalian cells. These variations of molecular weight were probably due to the different modifications of glycosylation. All these recombinant proteins retained the antigenicity and biological activity on 7TD1 mouse cells.

The effect of recombinant swine interleukin-4 on swine immune cells and on pro-inflammatory cytokine productions in pigs.

The in vitro effect and the in vivo influence of recombinant swine IL-4 (rSwIL-4) were characterized in various swine cells and in nursery pigs on LPS-induced endotoxic shock and pro-inflammatory cytokine productions. In in vitro experiment, the rSwIL-4 induced a proliferation of CD4 positive T cells in mitogen-prestimulated peripheral blood mononuclear cell (PBMC). In addition, the rSwIL-4, which was produced from insect cells, promoted the differentiation of monocytes into immature dendritic cells in combination with granulocyte macrophage-colony stimulating factor (GM-CSF). Furthermore, the rSwIL-4 successfully suppressed the LPS-induced secretion of TNF-alpha, IL-1alpha, IL-6, IL-8, and IL-18 from swine alveolar macrophages when rSwIL-4 was treated at the same time with LPS. In in vivo experiment in nursery pigs, subcutaneous pretreatment of rSwIL-4, which was produced from baculovirus expression system, enhanced the severity of respiratory failure with endotoxic shock, and increased the production of TNF-alpha and IL-18 in response to inoculation with LPS. These results indicate that the rSwIL-4 is biologically active in both in vitro and in vivo treatments. Depending on the administration time, pro-inflammatory cytokine productions by IL-4 can cause either inhibitory or stimulatory regulation.

Expression and purification of recombinant swine interleukin-4.

The swine interleukin-4 (SwIL-4) cDNA was cloned by RT-PCR. It was expressed using an expression vector pQE30 in E. coli, a baculovirus AcNPV vector pVL1392 in insect cells, and a pCAGGS vector in mammalian cells. The rSwIL-4 proteins expressed from bacteria and insect cells were purified using a chelating affinity column and a mAb-coupled immunoaffinity column. The amount of the products and their bioactivities were compared. All recombinant cytokines were efficiently reacted with the specific antibodies and the molecular weight of rSwIL-4 was approximately 16 kDa in E. coli, 15 and 18 kDa in insect cells, and 15 and 20 kDa in mammalian cells. Variations of molecular weight observed in insect and mammalian cells were probably due to different modification ways of glycosylation. All these recombinant proteins retained their antigenicity and were biologically active in inducing human TF-1 cell proliferation in vitro. The simple purification method will make it possible to evaluate the in vitro and in vivo effects of IL-4 in pigs.

Usefulness of mandibular third molar coronectomy assessed through clinical evaluation over three years of follow-up.

The aim of this study was to investigate the 3-year morbidity of coronectomy of the lower third molar and to monitor the behaviour and migration pattern of the retained roots postoperatively. A total of 92 patients (111 teeth) who had undergone a coronectomy between October 2005 and July 2009 were investigated. Patients were followed up at 3 months and 1, 2, and 3 years for clinical evaluation and dental computed tomography imaging of the coronectomy sites. In total, 10 cases (9%) required tooth root extraction within the 3 years after coronectomy. In seven of them, the distal pocket of the lower second molars remained connected to the roots within the first year. Of the cases in whom a pocket did not remain at an early stage, none showed peri-apical lesions on transmission images of the retained roots in the apical area, which usually result from necrosis of the pulp. Root migration increased in the first 2 years after coronectomy but stabilized between the second and third years. In addition, a significant difference was noted in root migration between patients of different ages and sex. Retained roots after coronectomy in the lower third molars led to no complications in terms of infection or the development of pathologies within the first 3 years postoperatively.

Association between risk factors for atherosclerosis and mechanical forces in carotid artery.

BACKGROUND AND PURPOSE: Mechanical stresses on the arterial wall participate in the pathogenesis of atherosclerosis as local factors. The relationships between local mechanical forces and risk factors for atherosclerosis were investigated. METHODS: Mechanical forces on the arterial wall were evaluated in the carotid artery in 117 patients with risk factors for atherosclerosis including hypertension, dyslipidemia, diabetes mellitus, and smoking, as well as in 20 age- and sex-matched normal controls. Circumferential wall tension and shear stress were evaluated with Laplace's law and a poiseuillean parabolic model of velocity distribution. Circumferential wall strain was also evaluated as carotid mechanical force. RESULTS: Mechanical forces in subjects with risk factors were characterized by low wall shear stress, high circumferential wall tension, and reduced strain. Systolic blood pressure was significantly negatively associated with shear stress and circumferential wall strain. HDL cholesterol showed a significant positive correlation with shear stress and a negative correlation with wall tension. Fasting blood glucose was significantly associated with shear stress, while smoking showed a negative correlation with shear stress and a positive correlation with wall tension. Accumulation of risk factors was associated with further deterioration of mechanical forces. Furthermore, stepwise regression analysis showed that the number of risk factors was significantly associated with mechanical forces independently of carotid intima-media thickness. CONCLUSIONS: These findings suggest that risk factors for atherosclerosis were associated with alteration of mechanical forces. Consequent alteration in mechanical forces could be an underlying local mechanism for the progression of atherosclerosis.

Autonomic nervous function in non-dipper essential hypertensive subjects. Evaluation by power spectral analysis of heart rate variability.

Autonomic nervous function was evaluated by means of power spectral analysis of heart rate variability in hospitalized dipper (n = 31) and non-dipper (n = 31) essential hypertensive subjects. Twenty-four-hour blood pressure (BP) measurement was performed by the cuff-oscillometric method to evaluate the nocturnal decrease of BP. The non-dipper subjects were defined as those whose nocturnal decrease of systolic BP was < 10% of daytime BP. Power spectral analysis of RR interval was performed from Holter ECG every 10 minutes by the maximum entropy method to obtain the low-frequency band (LFB, 0.04 to 0.15 Hz), which is an index of both parasympathetic and sympathetic nervous activities, and the high frequency band (HFB, 0.15 to 0.4 Hz), which reflects parasympathetic nervous activity. LFB and HFB were averaged every hour to obtain hourly LFB and HFB values. Total LFB and total HFB were calculated as the mean values of 24 hourly averaged LFBs and HFBs. Both LFB and HFB were significantly lower in non-dipper hypertensives than in dipper subjects throughout the day. In dipper hypertensives, LFB showed a nocturnal decrease, whereas HFB was significantly increased during the nighttime. However, these diurnal changes in LFB and HFB were significantly blunted in non-dipper subjects. These findings indicate that non-dipper hypertensive subjects were characterized with a decreased physiological circadian fluctuation on autonomic functions compared with dipper subjects. This alteration in the autonomic nervous function may explain the non-dipper phenomenon in essential hypertension.

Angiotensin blockade and the progression of renal damage in the spontaneously hypertensive rat.

The pathophysiological role of angiotensin II in the development of renal sclerosis was investigated in 5/6-nephrectomized, 12-week-old male spontaneously hypertensive rats. After 1 week of a control period, nephrectomized rats received one of the following treatments for 4 weeks: the selective nonpeptide angiotensin II type 1 receptor antagonist TCV-116 (1 mg/kg per day), the angiotensin converting enzyme inhibitor delapril (30 mg/kg per day), hydralazine (15 mg/kg per day), or vehicle. Urinary protein and albumin excretions and systolic blood pressure were determined every week. Rats with reduced renal mass treated with vehicle had a poor survival rate (30%). Although TCV-116, delapril, and hydralazine treatment significantly improved the survival rate for 4 weeks, hydralazine failed to improve proteinuria and albuminuria as well as the decline in renal function compared with delapril or TCV-116. Histological examination revealed that both TCV-116 and delapril protected glomeruli from sclerosis, whereas hydralazine did not improve histological findings (5%, 7%, and 30% of glomeruli were affected, respectively). These results indicate that angiotensin II plays a dominant role through its type 1 receptor in the pathogenesis of renal deterioration by hypertension.

Vasoconstriction and hypersensitivity to vasoactive substances after acute volume expansion in dogs.

In a search for factors contributing to the sustained blood pressure (BP) elevation in acutely volume-loaded animals, dextran dissolved in lactated Ringer's solution (20 ml/kg) was infused into 34 mongrel dogs over a period of 1 hour under pentobarbital anesthesia and changes in hemodynamic and humoral variables were monitored during its infusion and for 3 hours after its infusion. BP elevation during volume loading (from 114 +/- 3 to 128 +/- 3 [SEM] mm Hg) was attributed to an increase in cardiac output. After volume loading, some dogs maintained BP elevation whereas others did not. The former group showed an increase in total peripheral resistance, demonstrating a transformation of cardiac output to total peripheral resistance as a responsible factor in maintenance of the elevated BP. The plasma levels of norepinephrine, vasopressin, and plasma renin activity were not elevated, indicating that these vasoactive factors were not responsible for elevation of the BP or total peripheral resistance. The changes in the hematocrit, atrial natriuretic factor, urine volume, and urinary sodium excretion were identical in the two groups, and natriuresis was not prominent when total peripheral resistance was high. Pressor responses to norepinephrine and angiotensin II were potentiated 3 hours after stopping infusion in both groups, but this potentiation was not correlated with the increase in total peripheral resistance or mean BP. Thus, acute volume expansion produced resistance-dependent hypertension following the initial volume-dependent hypertension. It is unlikely that a vascular sensitizing natriuretic factor plays a role in the resistance-dependent BP elevation. The mechanism and physiological importance of hypersensitivity to vasoactive substances remain to be elucidated.