Changes in renal hemodynamics and excretory function induced by a reduction of ANG II effects during renal development

The aim was to evaluate whether blockade of ANG II effects during renal development modifies the renal response to an increment of plasma amino acid concentration. It was also examined in anesthetized rats whether the reduction of the renal ability to eliminate an acute volume expansion (VE), elicited by blockade of ANG II during renal development, is sex and/or age dependent. Newborn Sprague-Dawley rats were treated with vehicle or an AT(1)-receptor antagonist (ARA) during postnatal nephrogenesis. Amino acid infusion induced increments (P < 0.05) of glomerular filtration rate (31 +/- 6%) and renal plasma flow (26 +/- 5%) in male but not in female vehicle-treated rats. Natriuretic and diuretic responses to amino acid infusion were similar in male and female vehicle-treated rats. These renal hemodynamics and excretory responses to amino acid infusion were abolished in ARA-treated rats. Renal responses to VE were evaluated at 3-4 and 9-10 mo of age in vehicle and ARA-treated rats. VE-induced natriuresis and diuresis were reduced by more than 38% (P < 0.05) in 3- to 4-mo-old male and female ARA-treated rats. An age-dependent reduction (P < 0.05) in the renal ability to eliminate VE was found in male but not in female rats treated with ARA. Our results demonstrate that the renal effects induced by an increment in amino acids are abolished when ANG II effects have been reduced during nephrogenesis. In addition, this reduction of ANG II effects elicits an impairment of the renal ability to eliminate an acute VE in males and females, which is aggravated by age only in male rats.     

Age-, gender-, and species-dependent mutagenicity in T cells of mice and rats exposed by inhalation to 1,3-butadiene

Experiments were performed: (i) to investigate potential age- and gender-dependent differences in mutagenic responses in T cells following exposures of B6C3F1 mice and F344 rats by inhalation for 2 weeks to 0 or 1250 ppm butadiene (BD), and (ii) to determine if exposures for 2 weeks to 62.5 ppm BD produce a mutagenic effect in female rats. To evaluate the effect of age on mutagenic response, mutant manifestation curves for splenic T cells of female mice exposed at 8-9 weeks of age were defined by measuring Hprt mutant frequencies (MFs) at multiple time points after BD exposure using a T cell cloning assay and comparing the resulting mutagenic potency estimate (calculated as the difference of areas under the mutant manifestation curves of treated versus control animals) to that reported for female mice exposed to BD in the same fashion beginning at 4-5 weeks of age. The shapes of the mutant T cell manifestation curves for spleens were different [e.g., the maximum BD-induced MFs in older mice (8.0+/-1.0 [S.D.]x10(-6)) and younger mice (17.8+/-6.1 x 10(-6)) were observed at 8 and 5 weeks post-exposure, respectively], but the mutagenic burden was the same for both age groups. To assess the effect of gender on mutagenic response, female and male rodents were exposed to BD at 4-5 weeks of age and Hprt MFs were measured when maximum MFs are expected to occur post-exposure. The resulting data demonstrated that the pattern for mutagenic susceptibility from high-level BD exposure is female mice>male mice>female rats>male rats. Exposures of female rats to 62.5 ppm BD caused a minor but significant mutagenic response compared with controls (n=16/group; P=0.03). These results help explain part of the differing outcomes/interpretations of data in earlier Hprt mutation studies in BD-exposed rodents.     

Sex differences in circulating and renal angiotensins of hypertensive mRen(2). Lewis but not normotensive Lewis rats

Sex differences in blood pressure are evident in experimental models and human subjects, yet the mechanisms underlying this disparity remain equivocal. The current study sought to define the extent of male-female differences in the circulating and tissue renin-angiotensin aldosterone systems (RAASs) of congenic mRen(2). Lewis and control Lewis rats. Male congenics exhibited higher systolic blood pressure than females [200 +/- 4 vs. 146 +/- 7 mmHg, P < 0.01] or Lewis males and females [113 +/- 2 vs. 112 +/- 2 mmHg, P > 0.05]. Plasma ANG II levels were twofold higher in male congenics [47 +/- 3 vs. 19 +/- 3 pM, P < 0.01] and fivefold higher than in male or female Lewis rats [6 +/- 1 vs. 6 +/- 1 pM]. ANG I levels were also highest in the males; however, plasma ANG-(1-7) was higher in female congenics. Male congenics exhibited greater circulating renin and angiotensin-converting enzyme (ACE) activities, as well as angiotensinogen, than female littermates. Renal cortical and medullary ANG II levels were also higher in the male congenics versus all the other groups; ANG I was lower in the males. Cortical ACE2 activity was higher in male congenics, yet neprilysin activity and protein were greater in the females, which may contribute to reduced renal levels of ANG II. These data reveal that sex differences in both the circulating and renal RAAS are apparent primarily in the hypertensive group. The enhanced activity of the RAAS in male congenics may contribute to the higher pressure and tissue injury evident in the strain.     

Effect of caloric restriction on Hprt lymphocyte mutation in aging rats

Caloric restriction (CR) reduces tumor incidence and retards aging in laboratory animals, including non-human primates. Because of the relationships among mutation, disease susceptibility, and aging, we investigated whether or not CR affects the accumulation of somatic cell mutations in aging animals. Starting at approximately 2 months of age, male CD rats (Harlan Sprague-Dawley-derived) were placed on different levels of dietary intake: ad libitum (AL) feeding, and 90% (10% CR), 75% (25% CR) and 60% (40% CR) of the total calories consumed by AL animals. At 3, 6, 12, and 24 months after the beginning of CR, Hprt mutant frequencies (MFs) were determined. The MFs measured in spleen lymphocytes from AL and CR rats sacrificed at 3 months of dietary restriction were similar for all dietary groups. However, the MFs at 6, 12, and 24 months of CR were significantly higher in AL-fed rats compared with animals on 40% CR: (4.5+/-0.4)x10(-6) versus (3.3+/-0.3)x10(-6) (P=0.032) in 6 months CR rats; (10.3+/-2.3)x10(-6) versus (7.3+/-1.2)x10(-6) in 12 months CR rats (P=0.04), and (18.3+/-3.2)x10(-6) versus (7.8+/-1.0)x10(-6) (P=0.001) in 24 months CR rats. In addition, rats receiving 25% CR for 24 months had a MF, (10.7+/-2.0)x10(-6), between the 40% CR and AL rats. Multiplex PCR of the Hprt gene in mutant clones from 12 and 24 months 40% CR rats and the corresponding AL rats detected deletions in 42% of CR mutants and 19% of AL mutants. Because of the difference in Hprt MF in the two groups, the estimated MF associated with deletions in CR rats was similar to the deletion MF in AL rats. This observation implies that the lower MF in CR rats is due to a reduction in smaller Hprt mutations (i.e. base substitutions and frameshifts). The pattern of smaller Hprt mutations from AL rats suggests that many were produced by reactive oxygen species (ROS). The results indicate that CR reduces the accumulation of spontaneous somatic cell mutation in aging rats, especially those caused by base substitutions and frameshifts.     

Sex differences in renal injury and nitric oxide production in renal wrap hypertension

To investigate the faster rate of renal disease progression in men compared with women, we addressed the following questions in the renal wrap (RW) model of hypertension: 1) Do sex differences exist in RW-induced renal injury, which are independent of sex differences in blood pressure? 2) Do sex differences in nitric oxide (NO) production exist in RW hypertension? Male (M) and female (F) rats underwent sham-operated (M-Sham, n = 7; F-Sham, n = 10) or RW (M-RW, n = 13; F-RW, n = 14) surgery for 9 wk. Markers of renal injury, including the glomerulosclerosis index (F-RW, 0.70 +/- 0.1 vs. M-RW, 2.2 +/- 0.6; P < 0.05), mean glomerular volume (F-RW, 1.05 +/- 0.050 x 10(6) vs. M-RW, 1.78 +/- 0.15 x 10(6) microm(3); P < 0.001), and proteinuria (F-RW, 68.7 +/- 15 vs. M-RW, 124 +/- 7.7 mg/day; P < 0.001) were greater in RW males compared with RW females. Endothelial NO synthase protein expression was elevated in the renal cortex (3.2-fold) and medulla (2.2-fold) 9 wk after RW in males, whereas no differences were observed in females. Neuronal NO synthase protein expression was unchanged in the renal cortex in males and in both the renal cortex and medulla in females, whereas in the male medulla, neuronal NOS was decreased by 57%. These data suggest the degree of renal injury is greater in male compared with female rats in RW hypertension despite similar degrees of hypertension and renal function and may involve sex differences in renal NO metabolism.     

Role of estrogen in modulating EDHF-mediated dilations in the female rat middle cerebral artery

We tested the hypothesis that endothelium-derived hyperpolarizing factor (EDHF) plays a less dominant role in the female cerebrovasculature. The contribution of EDHF to the ATP-mediated dilation was determined in middle cerebral arteries (MCAs) isolated from male and female rats. Four groups of rats were tested: intact male (n = 12), intact female (n = 13), estrogen-treated ovariectomized female (n = 13), and vehicle-treated ovariectomized female (n = 20) rats. Maximal dilation to ATP was similar in all groups. However, in the presence of N(omega)-nitro-L-arginine methyl ester (L-NAME, 3 x 10(-5) M) and indomethacin (10(-5) M), the maximal dilation to ATP was significantly reduced in intact female (24 +/- 9%) and estrogen-treated ovariectomized female (29 +/- 9%) rats compared with intact male (95 +/- 4%) and vehicle-treated ovariectomized female (96 +/- 2%) rats. The ATP-mediated dilation in L-NAME- and indomethacin-treated MCAs isolated from male and ovariectomized female rats was inhibited by charybdotoxin (10(-7) M), an inhibitor of large-conductance Ca2+-sensitive K+ channels. We have defined EDHF as the L-NAME- and indomethacin-insensitive component of the ATP-mediated dilation. Our findings indicate that EDHF-mediated dilations are negligible in the female rat MCA; these dilations can be significantly enhanced after ovariectomy, suggesting that this effect is mediated by estrogen.     

Mouse Complete Stasis Model of Inferior Vena Cava Thrombosis

Venous thromboembolism (VTE) includes both deep vein thrombosis (DVT) and pulmonary embolism (PE). In the United States (U.S.), the high morbidity and mortality rates make VTE a serious health concern ^1-2. After heart disease and stroke, VTE is the third most common vascular disease ³. In the U.S. alone, there is an estimated 900,000 people affected each year, with 300,000 deaths occurring annually ³. A reliable in vivo animal model to study the mechanisms of this disease is necessary.The advantages of using the mouse complete stasis model of inferior vena cava thrombosis are several. The mouse model allows for the administration of very small volumes of limited availability test agents, reducing costs dramatically. Most promising is the potential for mice with gene knockouts that allow specific inflammatory and coagulation factor functions to be delineated. Current molecular assays allow for the quantitation of vein wall, thrombus, whole blood, and plasma for assays. However, a major concern involving this model is the operative size constraints and the friability of the vessels. Also, due to the small IVC sample weight (mean 0.005 grams) it is necessary to increase animal numbers for accurate statistical analysis for tissue, thrombus, and blood assays such as real-time polymerase chain reaction (RT-PCR), western blot, enzyme-linked immunosorbent (ELISA), zymography, vein wall and thrombus cellular analysis, and whole blood and plasma assays ^4-8.The major disadvantage with the stasis model is that the lack of blood flow inhibits the maximal effect of administered systemic therapeutic agents on the thrombus and vein wall.     

Nitric oxide increases fluid extravasation from the splenic circulation of the rat

We hypothesized that nitric oxide (NO) contributes to intrasplenic fluid extravasation by inducing greater relaxation in splenic resistance arteries than veins such that intrasplenic microvascular pressure (P(C)) rises. Fluid efflux was estimated by measuring the difference between splenic blood inflow and outflow. Intrasplenic infusion of the NO donor S-nitroso-N-acetylpenicillamine (SNAP) (0.3 microg. 10 microl(-1). min(-1)) caused a significant increase in intrasplenic fluid efflux (baseline: 0.8 +/- 0.4 ml/min, n = 10 vs. peak rise during SNAP infusion: 1.3 +/- 0.4 ml/min, n = 10; P < 0.05). Intrasplenic P(C) was measured in the isolated, blood-perfused rat spleen. Intrasplenic infusion of SNAP (0.1 microg. 10 microl(-1). min(-1)) caused a significant increase in P(C) (saline: 10.9 +/- 0.2 mmHg, n = 3 vs. SNAP: 12.2 +/- 0.2 mmHg, n = 3; P < 0.05). Vasoreactivity of preconstricted splenic resistance vessels to sodium nitroprusside (SNP) (1 x 10(-12)-1 x 10(-4) M) and SNAP (1 x 10(-10)-3 x 10(-4) M) was investigated with the use of a wire myograph system. Significantly greater relaxation of arterioles than of venules occurred with both SNP (%maximal vasorelaxation: artery 96 +/- 2.3, n = 9 vs. vein 26 +/- 1.9, n = 10) and SNAP (%maximal vasorelaxation: artery 50 +/- 3.5, n = 11 vs. vein 32 +/- 1.7, n = 8). These results are consistent with our proposal that differential vasoreactivity of splenic resistance arteries and veins to NO elevates intrasplenic P(C) and increases fluid extravasation into the systemic lymphatic system.     

Glucocorticoids act in the dorsal hindbrain to modulate baroreflex control of heart rate

Systemic corticosterone (Cort) modulates arterial baroreflex control of both heart rate and renal sympathetic nerve activity. Because baroreceptor afferents terminate in the dorsal hindbrain (DHB), an area with dense corticosteroid receptor expression, we tested the hypothesis that prolonged activation of DHB Cort receptors increases the midpoint and reduces the gain of arterial baroreflex control of heart rate in conscious rats. Small (3-4 mg) pellets of Cort (DHB Cort) or Silastic (DHB Sham) were placed on the surface of the DHB, or Cort was administered systemically by placing a Cort pellet on the surface of the dura (Dura Cort). Baroreflex control of heart rate was determined in conscious male Sprague Dawley rats on each of 4 days after initiation of treatment. Plots of arterial pressure vs. heart rate were analyzed using a four-parameter logistic function. After 3 days of treatment, the arterial pressure midpoint for baroreflex control of heart rate was increased in DHB Cort rats (123 +/- 2 mmHg) relative to both DHB Sham (108 +/- 3 mmHg) and Dura Cort rats (109 +/- 2 mmHg, P < 0.05). On day 4, baseline arterial pressure was greater in DHB Cort (112 +/- 2 mmHg) compared with DHB Sham (105 +/- 2 mmHg) and Dura Cort animals (106 +/- 2 mmHg, P < 0.05), and the arterial pressure midpoint was significantly greater than mean arterial pressure in the DHB Cort group only. Also on day 4, maximum baroreflex gain was reduced in DHB Cort (2.72 +/- 0.12 beats x min(-1) x mmHg(-1)) relative to DHB Sham and Dura Cort rats (3.51 +/- 0.28 and 3.37 +/- 0.27 beats x min(-1) x mmHg(-1), P < 0.05). We conclude that Cort acts in the DHB to increase the midpoint and reduce the gain of the heart rate baroreflex function.     

Effect on ejaculatory performance and semen parameters of sexually-satiated male goats (Capra hircus) after changing the stimulus female

The association between stimulation of sexual activity and semen characteristics of sexually mature goats was evaluated. Nine 2-year-old criollo bucks were used. Each buck was subjected to 8-weekly trials in which one of the following two treatments was alternately applied. In treatment 1, males were individually exposed to the same estrus-induced female during a 4-h period, and ejaculates were collected and analyzed throughout. Treatment 2 was the same, except that a different doe replaced the stimulus animal for the third and fourth hours. The total number of ejaculations and total sperm number for four sets of data was different (P<0.01) for treatments 1 and 2 (184 versus 252 ejaculations with a total of 354.3 versus 477.1 billion sperm, respectively). In treatment 2, when first and last ejaculation of each male with the (original) stimulus animal were compared, total sperm per ejaculate decreased (P=0.08) from 4.14+/-3.8 x 10(9) to 0.77+/-0.7 x 10(9), while this value increased (P=0.05) to 3.04+/-2.3 x 10(9) after a new female was introduced, which represented a recovery of 67.35%. All males achieved a first ejaculation with the original stimulus animal while on average, only three achieved a seventh service. After changing the stimulus animal, all males ejaculated again. It is concluded that changing the stimulus animal after a 2-h continuous exposure to an estrous female stimulates sexual activity and increases sperm output in male goats.     

Enhanced renal expression of preproendothelin mRNA during chronic angiotensin II hypertension

The purpose of this study was to determine the role of endothelin in mediating the renal hemodynamic and arterial pressure changes observed during chronic ANG II-induced hypertension. ANG II (50 ng x kg(-1) x min(-1)) was chronically infused into the jugular vein by miniosmotic pump for 2 wk in male Sprague-Dawley rats with and without endothelin type A (ET(A))-receptor antagonist ABT-627 (5 mg x kg(-1) x day(-1)) pretreatment. Arterial pressure increased in ANG II rats compared with control rats (149 +/- 5 vs. 121 +/- 6 mmHg, P < 0.05, respectively). Renal expression of preproendothelin mRNA was increased by approximately 50% in both the medulla and cortex of ANG II rats. The hypertensive effect of ANG II was completely abolished in rats pretreated with the ET(A)-receptor antagonist (114 +/- 5 mmHg, P < 0.05). Glomerular filtration rate was decreased by 33% in ANG II rats, and this response was attenuated in rats pretreated with ET(A)-receptor antagonist. These data indicate that activation of the renal endothelin system by ANG II may play an important role in mediating chronic renal and hypertensive actions of ANG II.     

Portal vein hypoglycemia is essential for full induction of hypoglycemia-associated autonomic failure with slow-onset hypoglycemia

Antecedent hypoglycemia leads to impaired counterregulation and hypoglycemic unawareness. To ascertain whether antecedent portal vein hypoglycemia impairs portal vein glucose sensing, thereby inducing counterregulatory failure, we compared the effects of antecedent hypoglycemia, with and without normalization of portal vein glycemia, upon the counterregulatory response to subsequent hypoglycemia. Male Wistar rats were chronically cannulated in the carotid artery (sampling), jugular vein (glucose and insulin infusion), and mesenteric vein (glucose infusion). On day 1, the following three distinct antecedent protocols were employed: 1) HYPO-HYPO: systemic hypoglycemia (2.52 +/- 0.11 mM); 2) HYPO-EUG: systemic hypoglycemia (2.70 +/- 0.03 mM) with normalization of portal vein glycemia (portal vein glucose = 5.86 +/- 0.10 mM); and 3) EUG-EUG: systemic euglycemia (6.33 +/- 0.31 mM). On day 2, all groups underwent a hyperinsulinemic-hypoglycemic clamp in which the fall in glycemia was controlled so as to reach the nadir (2.34 +/- 0.04 mM) by minute 75. Counterregulatory hormone responses were measured at basal (-30 and 0) and during hypoglycemia (60-105 min). Compared with EUG-EUG, antecedent hypoglycemia (HYPO-HYPO) significantly blunted the peak epinephrine (10.44 +/- 1.35 vs. 15.75 +/- 1.33 nM: P = 0.01) and glucagon (341 +/- 16 vs. 597 +/- 82 pg/ml: P = 0.03) responses to next-day hypoglycemia. Normalization of portal glycemia during systemic hypoglycemia on day 1 (HYPO-EUG) prevented blunting of the peak epinephrine (15.59 +/- 1.43 vs. 15.75 +/- 1.33 nM: P = 0.94) and glucagon (523 +/- 169 vs. 597 +/- 82 pg/ml: P = 0.66) responses to day 2 hypoglycemia. Consistent with hormonal responses, the glucose infusion rate during day 2 hypoglycemia was substantially elevated in HYPO-HYPO (74 +/- 12 vs. 49 +/- 4 micromol x kg(-1) x min(-1); P = 0.03) but not HYPO-EUG (39 +/- 7 vs. 49 +/- 4 micromol x kg(-1) x min(-1): P = 0.36). Antecedent hypoglycemia local to the portal vein is required for the full induction of hypoglycemia-associated counterregulatory failure with slow-onset hypoglycemia.     

Cytophotometric assessment of Feulgen-DNA and coomassie-total cell protein in adrenocortical fasciculata cells of noise-exposed Wistar rats.

The present investigation was undertaken to examine possible noise-induced alterations in adrenal fasciculata cell (AFC) metabolism, and also to determine if the magnitude of these changes differs in male versus female rats. Wistar rats approximately 3 months old were exposed to intense noise for 60 min (100 dB, re 2 x 10(-5) N(m2)-1, 350-20,000 Hz); control rats were housed under identical conditions, at an ambient noise level of 40-60 dB. Adrenal fasciculata cells (AFC) from each animal were examined for noise-induced alterations in Feulgen-DNA reactivity (as an indicator of chromatin template activity) and Coomassie-total cell protein levels using scanning-integrating cytophotometry. The results provide evidence that intense noise elicited a marked AFC metabolic enhancement in both male and female rats; the degree of this enhancement was more pronounced in males. This disparity may be due to pre-existing differences in male versus female AFC enzymatic capability and subsequent responsiveness to noise-induced activation.     

Pharmacokinetic and pharmacodynamic interactions between simvastatin and diltiazem in patients with hypercholesterolemia and hypertension

Pharmacokinetic and pharmacodynamic interactions between simvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, and diltiazem, a calcium antagonist, were investigated in 7 male and 4 female patients with hypercholesterolemia and hypertension. The patients were given, for one in a three consecutive 4-week periods, oral simvastatin (5 mg/day), oral simvastatin (5 mg/day) combined with diltiazem (90 mg/day), and then oral diltiazem (90 mg/day), respectively. The area under the plasma concentration versus time curve up to 6 hours post-dose (AUC0-6h) and maximum plasma concentrations (Cmax) of the drugs, serum lipid profiles, blood pressures and liver functions were assessed on the last day of each of the three 4-week periods. After the combined treatment period, Cmax of HMG-CoA reductase inhibitor was elevated from 7.8 +/- 2.6 ng/ml to 15.4 +/- 7.9 ng/ml (P < 0.01) and AUC0-6h from 21.7 +/- 4.9 ng x hr/ml to 43.3 +/- 23.4 ng x hr/ml (P < 0.01), while Cmax of diltiazem was decreased from 74.2 +/- 36.4 ng/ml to 58.6 +/- 18.9 ng/ml (P < 0.05) and its AUC0-6h from 365 +/- 153 ng x hr/ml to 287 +/- 113 ng x hr/ml (P < 0.01). Compared to simvastatin monotherapy, combined treatment further reduced LDL-cholesterol levels by 9%, from 129 +/- 16 mg/dl to 119 +/- 17 mg/dl (P < 0.05). No adverse events were observed throughout the study. These apparent pharmacokinetic interactions, namely the increase of HMG-CoA reductase inhibitor concentration by diltiazem and the decrease of diltiazem concentration by simvastatin, enhance the cholesterol-lowering effects of simvastatin during combined treatment.     

High volume hydrodynamic injection of plasmid DNA via the hepatic artery results in a high level of gene expression in rat hepatocellular carcinoma induced by diethylnitrosamine

BACKGROUND: Hydrodynamic injection of naked plasmid DNA (pDNA) via the tail vein is a safe and effective method of gene transfer to the liver. However, successful gene transfer has yet to be shown for hepatocellular carcinoma (HCC); therefore, we investigated the feasibility and efficacy of hydrodynamic injection via the tail vein and hepatic artery in a diethylnitrosamine (DEN)-induced HCC model in rats. METHODS: HCC was induced in Sprague-Dawley rats by 100 ppm DEN in drinking water. pCMV-SPORT-beta-galactosidase (beta-gal, 400 microg) was injected (i) via the tail vein in a volume of 0.1 ml/g in 30 s or (ii) via the hepatic artery in a volume of 5 or 10 ml at 1 ml/s, either with or without temporary occlusion of the inferior vena cava (IVC) and portal vein (PV). The liver was harvested 24 h after administration, and beta-gal expression was evaluated with X-gal staining and measurement of enzymatic activity in tissue homogenates. RESULTS: Hydrodynamic injection via the tail vein achieved transgene expression only in non-cancerous tissue (tumor: 0.16 +/- 0.04%, non-tumor: 5.07 +/- 1.66%). Hydrodynamic injection via the hepatic artery was tolerated, but failed to produce efficient transgene expression in tumor and non-tumor cells. On the other hand, concomitant use of temporary IVC/PV occlusion with hydrodynamic injection via the hepatic artery dramatically increased transgene expression in cancer cells, but tumor-selective gene transfer was not achieved with this procedure (tumor: 7.38 +/- 3.66%, non-tumor: 7.77 +/- 1.06%). CONCLUSIONS: High-volume hydrodynamic injection of a pDNA solution via the hepatic artery with IVC/PV occlusion achieved a high level of gene expression in a HCC rat model. This gene transfer technique may have potential in clinical gene therapy for HCC.     

Zymosan-induced luminol-dependent chemiluminescence response of circulating and extravasated leukocytes in experimental sepsis

This study examines a concurrent profiling of circulating and extravasated polymorphonuclear leukocytes (PMNs) in a rat model of experimental sepsis. Fecal peritonitis was induced in Wistar male rats by intraperitoneal instillation of a fecal suspension in saline (1:1 w/v). Blood and peritoneal fluid were collected 8 h following fecal inoculation for the evaluation of inflammatory response of PMNs using zymosan-induced luminol-dependent chemiluminescence. Fifty microliters of pre-diluted blood or peritoneal fluid samples were mixed with 150 microl of reaction mixture (4 x 10(-4) M luminol+50 microg opsonized zymosan+0.1% gelatin in Hank's balanced salt solution) and the chemiluminescence signal was measured in a luminometer at 37 degrees C. Fecal peritonitis caused a significant leukocytopenia (3540+/-297 mm(-3) versus control value of 7525+/-711 mm(-3), p < 0.001) accompanied by massive infiltration of PMNs in the peritoneal cavity (34700+/-4006 versus 7325+/-425 mm(-3), p < 0.001). The phagocytic activity of circulating blood PMNs was down-regulated whereas a significant up-regulation was observed in the activity of PMNs from peritoneal fluid. In conclusion, this study clearly demonstrates sepsis-induced alterations in both blood and peritoneal fluid PMNs and their quantitative assessment may be helpful in disease evaluation and designing effective therapies.     

Aqueous extract of Paeoniae Radix Rubra prevents deep vein thrombosis by ameliorating inflammation through inhibiting GSK3β activity

BackgroundDeep vein thrombosis (DVT) is a kind of blood stasis syndrome. Paeoniae Radix Rubra (PRR) has long been widely used for eliminating blood stasis in China, but its effect on DVT has not yet been reported.PurposeThe present study aimed to assess the potential inhibitory effect of the aqueous extract of PRR (i.e.,PRR dispensing granule, PRRDG) on DVT and explore the underlying mechanism.Study design/MethodsThe chemical profile of PRRDG was analyzed by high-performance liquid chromatography. Sprague-Dawley rats were intragastrically treated with PRRDG (0.625, 1.25 and 1.875 g crude drug/kg/d) once daily for 7 consecutive days. On the sixth day, a model of inferior vena cava (IVC) stenosis-induced DVT was established. All rats were sacrificed on the seventh day. Serum was collected for enzyme-linked immunosorbent assay. Thrombus-containing IVC was weighed and further processed for histopathologic examination, immunohistochemical analysis and western blotting. LiCl and LY294002 were adopted to block and increase the activity of glycogen synthase kinase 3β (GSK3β), respectively.ResultsThe chemical profile analysis showed that paeoniflorin, benzoylpaeoniflorin, albiflorin, gallic acid and catechin were the main constituents of PRRDG. LiCl decreased thrombus weight, reduced the number of inflammatory cells in thrombus and vein wall, down-regulated phosphorylated NF-κB p65 (p-p65) protein expression. Similarly, PRRDG decreased thrombus weight and tissue factor (TF) protein expression. PRRDG reduced the protein expression levels of P-selectin, monocyte chemoattractant protein-1 (MCP-1), intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in venous endothelium, serum levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), and the number of inflammatory cells in thrombus and vein wall. Moreover, PRRDG down-regulated p-p65 protein expression and up-regulated phosphorylated GSK3β (p-GSK3β) protein expression. LY294002 abrogated the inhibitory effects of PRRDG on thrombus weight, TF protein expression, TNF-α and IL-1β serum levels, inflammatory cells influxes, and p-p65 protein expression.ConclusionPRRDG prevents DVT by ameliorating inflammation through inhibiting GSK3β activity.     

Gender difference in baroreflex-mediated bradycardia in young rats: role of cardiac sympathetic and parasympathetic components.

In a previous clinical study we have demonstrated a significantly lower baroreflex-mediated bradycardic response in young women compared with men. The present study determined whether sexual dimorphism in baroreflex sensitivity in young rats also covers the reflex tachycardic response. The study was then extended to test the hypothesis that an attenuated cardiac cholinergic component of the baroreflex heart rate response in females may account for the gender difference. Baroreflex sensitivity (BRS) was expressed as the regression coefficient of the reciprocal relationship between evoked changes in blood pressure and heart rate. BRS measured in conscious rats with phenylephrine (BRS(PE)) and nitroprusside (BRS(NP)) represented the reflex bradycardic and tachycardic responses, respectively. Female rats exhibited significantly lower BRS(PE) compared with male rats (-1.53+/-0.1 vs. -2.36+/-0.13 beats x min(-1) x mmHg(-1); p < 0.05) but similar BRS(NP) (-2.60+/-0.20 vs. -2.29+/-0.17 beats x min(-1) x mmHg(-1)). Blockade of cardiac muscarinic receptors with atropine methyl bromide elicited greater attenuation of BRS(PE) in male than in female rats (72+/-4.6 vs. 53+/-6.7% inhibition; p < 0.01) and abolished the gender difference. In male rats cardiac muscarinic blockade attenuated BRS(PE) significantly more than did cardiac beta-adrenergic receptor blockade with propranolol (72+/-4.6 vs. 43+/-2.7; p < 0.01), which suggests greater dependence of BRS(PE) on the parasympathetic component. In females, muscarinic and beta-adrenergic blockade elicited similar attenuation of BRS(PE). The findings suggest that (i) BRS is differentially influenced by gender; female rats exhibit substantially lower BRS(PE) but similar BRS(NP) compared with age-matched male rats and (ii) the sexual dimorphism in BRS(PE) results, at least partly, from a smaller increase in vagal outflow to the heart in response to baroreceptor activation.     

Consumption of an oral carbohydrate-protein gel improves cycling endurance and prevents postexercise muscle damage

Investigators have reported improved endurance performance and attenuated post-exercise muscle damage with carbohydrate-protein beverages (CHO+P) versus carbohydrate-only beverages (CHO). However, these benefits have been demonstrated only when CHO+P was administered in beverage-form, and exclusively in male subjects. Thus, the purposes of this study were to determine if an oral CHO+P gel improved endurance performance and post-exercise muscle damage compared to a CHO gel, and determine if responses were similar between genders. Thirteen cyclists (8 men, 5 women; VO(2)peak = 57.9 +/- 7.0 ml x kg(-1) x min(-1)) completed two timed cycle-trials to volitional exhaustion at 75% of VO(2)peak. At 15-minute intervals throughout these rides, subjects received CHO or CHO+P gels, which were matched for carbohydrate content (CHO = 0.15 g CHO x kg BW(-1); CHO+P = 0.15 g CHO + 0.038 g protein x kg BW(-1)). Trials were performed using a randomly counterbalanced, double-blind design. Subjects rode 13% longer (p < 0.05) when utilizing the CHO+P gel (116.6 +/- 28.5 minutes) versus the CHO gel (102.8 +/- 25.0 minutes). In addition, men (101.8 +/- 24.6; 114.8 +/- 26.2) and women (104.4 +/- 28.6; 119.6 +/- 34.9) responded similarly to the CHO and CHO+P trials, with no significant treatment-by-gender effect. Postexercise creatine kinease (CK) was not significantly different between treatments. However, CK increased significantly following exercise in the CHO trial (183 +/- 116; 267 +/- 214 U x L(-1)), but not the CHO+P trial (180 +/- 133; 222 +/- 141 U x L(-1)). Therefore, to prolong endurance performance and prevent increases in muscle damage, it is recommended that male and female cyclists consume CHO+P gels rather than CHO gels during and immediately following exercise.     

Thermogenic responsiveness to nonspecific beta-adrenergic stimulation is not related to genetic variation in codon 16 of the beta2-adrenergic receptor

Stimulation of beta-adrenergic receptors (beta-AR) by the sympathetic nervous system (SNS) modulates energy expenditure (EE), but substantial interindividual variability is observed. We determined whether the thermogenic response to beta-AR stimulation is related to genetic variation in codon 16 of the beta(2)-AR, a biologically important beta-AR polymorphism, and whether differences in SNS activity (i.e., the stimulus for agonist-promoted downregulation) are involved. The increase in EE (DeltaEE, indirect calorimetry, ventilated hood) above resting EE in response to nonspecific beta-AR stimulation [iv isoproterenol: 6, 12, and 24 ng/kg fat-free mass (FFM)/min] was measured in 46 healthy adult humans [Arg16Arg: 9 male, 7 female, 48 +/- 5 yr; Arg16Gly: 11 male, 4 female, 53 +/- 5 yr; Gly16Gly: 3 male, 12 female, 48 +/- 5 yr (means +/- SE)]. Neither FFM-adjusted baseline resting EE (P = 0.83) nor the dose of isoproterenol required to increase EE 10% above resting (P = 0.87) differed among the three groups (Arg16Arg: 5,409 +/- 209 kJ/day, 11.2 +/- 2.1 ng x kg FFM(-1) x min(-1); Arg16Gly: 5,367 +/- 272 kJ/day, 11.1 +/- 2.1 ng x kg FFM(-1) x min(-1); Gly16Gly: 5,305 +/- 159 kJ/day, 10.5 +/- 1.4 ng x kg FFM(-1) x min(-1)). Consistent with this, muscle sympathetic nerve activity and plasma norepinephrine concentrations were not different among the groups. Group differences in sex composition did not influence the results. Our findings indicate that the thermogenic response to nonspecific beta-AR stimulation, an important mechanistic component of overall beta-AR modulation of EE, is not related to this beta(2)-AR polymorphism in healthy humans. This may be explained in part by a lack of association between this gene variant and tonic SNS activity.