Control of the rat angiotensin I converting enzyme gene by CRE-like sequences

We characterized the role of potential cAMP-responsive elements (CRE) in basal and in induced angiotensin converting enzyme (ACE) gene promoter activity in order to shed light on the regulation of somatic ACE expression. We identified stimulators and repressors of basal expression between 122 and 288 bp and between 415 and 1303 bp upstream from the transcription start site, respectively, using a rabbit endothelial cell (REC) line. These regions also contained elements associated with the response to 8BrcAMP. When screening for CRE motifs we found pCRE, a proximal sequence between 209 and 222 bp. dCRE, a distal tandem of two CRE-like sequences conserved between rats, mice and humans, was detected between 834 and 846 bp. Gel retardation analysis of nuclear extracts of REC indicated that pCRE and dCRE bind to the same protein complexes as bound by a canonical CRE. Mutation of pCRE and dCRE in REC established the former as a positive element and the latter as a negative element. In 293 cells, a renal cell line, pCRE and dCRE are negative regulators. Co-transfection of ATF-2 or ATF-2 plus c-Jun repressed ACE promoter activity, suggesting that the ACE gene is controlled by cellular stress. Although mapping of cAMP responsiveness was consistent with roles for pCRE and dCRE, mutation analysis indicated that they were not required for cAMP responsiveness. We conclude that the basal activity of the somatic ACE promoter is controlled by proximal and distal CREs that can act as enhancers or repressors depending on the cell context.

Early effects of estrogen on the rat ventral prostate

Complex interactions between androgen and estrogen (E2) regulate prostatic development and physiology. We analyzed the early effects of a high single dose of E2 (25 mg/kg body weight) and castration (separately or combined) on the adult 90-day-old male Wistar rat ventral prostate. Androgen levels, prostate weight, and the variation in the relative and absolute volume of tissue compartments and apoptotic indices were determined for 7 days. Castration and exogenous E2 markedly reduced ventral prostate weight (about 50% of the control), with a significant reduction in the epithelial compartment and increased stroma. The final volume of the epithelium was identical at day 7 for all treatments (58.5% of the control). However, E2 had an immediate effect, causing a reduction in epithelial volume as early as day 1. An increase in smooth muscle cell volume resulted from the concentration of these cells around the regressing epithelium. The treatments resulted in differential kinetics in epithelial cell apoptosis. Castration led to a peak in apoptosis at day 3, with 5% of the epithelial cells presenting signs of apoptosis, whereas E2 caused an immediate increase (observed on day 1) and a sustained (up to day 7) effect. E2 administration to castrated rats significantly increased the level of apoptosis by day 3, reaching 9% of the epithelial cells. The divergent kinetics between treatments resulted in the same levels of epithelial regression after 7 days (~30% of control). These results show that E2 has an immediate and possibly direct effect on the prostate, and anticipates epithelial cell death before reducing testosterone to levels as low as those of castrated rats. In addition, E2 and androgen deprivation apparently cause epithelial cell death by distinct and independent pathways.

The blockade of cyclooxygenases-1 and -2 reduces the effects of hypoxia on endothelial cells

Hypoxia activates endothelial cells by the action of reactive oxygen species generated in part by cyclooxygenases (COX) production enhancing leukocyte transmigration. We investigated the effect of specific COX inhibition on the function of endothelial cells exposed to hypoxia. Mouse immortalized endothelial cells were subjected to 30 min of oxygen deprivation by gas exchange. Acridine orange/ethidium bromide dyes and lactate dehydrogenase activity were used to monitor cell viability. The mRNA of COX-1 and -2 was amplified and semi-quantified before and after hypoxia in cells treated or not with indomethacin, a non-selective COX inhibitor. Expression of RANTES (regulated upon activation, normal T cell expressed and secreted) protein and the protective role of heme oxygenase-1 (HO-1) were also investigated by PCR. Gas exchange decreased partial oxygen pressure (PaO2) by 45.12 ± 5.85% (from 162 ± 10 to 73 ± 7.4 mmHg). Thirty minutes of hypoxia decreased cell viability and enhanced lactate dehydrogenase levels compared to control (73.1 ± 2.7 vs 91.2 ± 0.9%, P < 0.02; 35.96 ± 11.64 vs 22.19 ± 9.65%, P = 0.002, respectively). COX-2 and HO-1 mRNA were up-regulated after hypoxia. Indomethacin (300 µM) decreased COX-2, HO-1, hypoxia-inducible factor-1alpha and RANTES mRNA and increased cell viability after hypoxia. We conclude that blockade of COX up-regulation can ameliorate endothelial injury, resulting in reduced production of chemokines.

Effect of antipyrine on the gastric emptying of liquid in rats

Antipyrine (At) and dipyrone (Dp) delay gastric emptying (GE) in rats. The objective of the present study was to assess the effects of intravenous (iv) and intracerebroventricular (icv) administration of At and Dp on the GE of liquid by rats. GE was assessed in male Wistar rats (5-10 in each group) 10 min after the icv or iv drug injection by measuring percent gastric retention (%GR) of a saline test meal labeled with phenol red 10 min after administration by gavage. The At iv group was significantly higher (64.4 ± 2.6%) compared to control (33.4 ± 1.5%) but did not differ from the Dp group (54.3 ± 3.8%). After icv administration of At, %GR (34.2 ± 2%) did not differ from control (32.6 ± 1.9%), but was significantly higher after Dp (54.5 ± 2.3%). Subdiaphragmatic vagotomy significantly reduced %GR in the At group (30.2 ± 0.7%) compared to the sham group, but was significantly higher than in the controls (23.0 ± 0.5%). In the animals treated with At iv, baclofen significantly reduced %GR (28.3 ± 2.4%) compared to vehicle-treated animals (55.2 ± 3.2%). The same occurred in the animals treated iv with vehicle and icv with baclofen. Although vagotomy and baclofen reduced %GR per se, the reduction was twice more marked in the animals treated with At. The results suggest that At administered iv, but not icv, delays GE of liquid in rats with the participation, at least in part, of the vagus nerve and that this phenomenon is blocked by the activation of GABA B receptors in the central nervous system.

The selective and non-selective cyclooxygenase inhibitors valdecoxib and piroxicam induce the same postoperative analgesia and control of trismus and swelling after lower third molar removal

We compared the clinical efficacy of orally administered valdecoxib and piroxicam for the prevention of pain, trismus and swelling after removal of horizontally and totally intrabony impacted lower third molars. Twenty-five patients were scheduled to undergo removal of symmetrically positioned lower third molars in two separate appointments. Valdecoxib (40 mg) or piroxicam (20 mg) was administered in a double-blind, randomized and crossed manner for 4 days after the surgical procedures. Objective and subjective parameters were recorded for comparison of postoperative courses. Both agents were effective for postoperative pain relief (N = 19). There was a similar mouth opening at suture removal compared with the preoperative values (86.14 ± 4.36 and 93.12 ± 3.70% of the initial measure for valdecoxib and piroxicam, respectively; ANOVA). There was no significant difference regarding the total amount of rescue medication taken by the patients treated with valdecoxib or piroxicam (173.08 ± 91.21 and 461.54 ± 199.85 mg, respectively; Wilcoxon test). There were no significant differences concerning the swelling observed on the second postoperative day compared to baseline measures (6.15 ± 1.84 and 8.46 ± 2.04 mm for valdecoxib and piroxicam, respectively; ANOVA) or on the seventh postoperative day (1.69 ± 1.61 and 2.23 ± 2.09 mm for valdecoxib and piroxicam, respectively; ANOVA). The cyclooxygenase-2 selective inhibitor valdecoxib is as effective as the non-selective cyclooxygenase inhibitor piroxicam for pain, trismus and swelling control after removal of horizontally and totally intrabony impacted lower third molars.

Effect of the time-of-day of training on explicit memory

Studies have shown a time-of-day of training effect on long-term explicit memory with a greater effect being shown in the afternoon than in the morning. However, these studies did not control the chronotype variable. Therefore, the purpose of this study was to assess if the time-of-day effect on explicit memory would continue if this variable were controlled, in addition to identifying the occurrence of a possible synchronic effect. A total of 68 undergraduates were classified as morning, intermediate, or afternoon types. The subjects listened to a list of 10 words during the training phase and immediately performed a recognition task, a procedure which they repeated twice. One week later, they underwent an unannounced recognition test. The target list and the distractor words were the same in all series. The subjects were allocated to two groups according to acquisition time: a morning group (N = 32), and an afternoon group (N = 36). One week later, some of the subjects in each of these groups were subjected to a test in the morning (N = 35) or in the afternoon (N = 33). The groups had similar chronotypes. Long-term explicit memory performance was not affected by test time-of-day or by chronotype. However, there was a training time-of-day effect [F (1,56) = 53.667; P = 0.009] with better performance for those who trained in the afternoon. Our data indicated that the advantage of training in the afternoon for long-term memory performance does not depend on chronotype and also that this performance is not affected by the synchronic effect.

Effect of exercise on the caloric intake of breast cancer patients undergoing treatment

The purpose of this study was to examine the effects of an exercise intervention on the total caloric intake (TCI) of breast cancer patients undergoing treatment. A secondary purpose was to determine whether or not a relationship existed between changes in TCI, body fat composition (%BF), and fatigue during the study, which lasted 6 months. Twenty females recently diagnosed with breast cancer, scheduled to undergo chemotherapy or radiation, were assigned randomly to an experimental (N = 10) or control group (N = 10). Outcome measures included TCI (3-day food diary), %BF (skinfolds), and fatigue (revised Piper Fatigue Scale). Each exercise session was conducted as follows: initial cardiovascular activity (6-12 min), followed by stretching (5-10 min), resistance training (15-30 min), and a cool-down (approximately 8 min). Significant changes in TCI were observed among groups (F1,18 = 8.582; P = 0.009), at treatments 2 and 3, and at the end of the study [experimental (1973 ± 419), control (1488 ± 418); experimental (1946 ± 437), control (1436 ± 429); experimental (2315 ± 455), control (1474 ± 294), respectively]. A significant negative correlation was found (Spearman rho(18) = -0.759; P < 0.001) between TCI and %BF and between TCI and fatigue levels (Spearman rho(18) = -0.541; P = 0.014) at the end of the study. In conclusion, the results of this study suggest that an exercise intervention administered to breast cancer patients undergoing medical treatment may assist in the mitigation of some treatment side effects, including decreased TCI, increased fatigue, and negative changes in body composition.

The effect of a low dose of clenbuterol on rat soleus muscle submitted to joint immobilization

The aim of the present study was to evaluate the effect of joint immobilization on morphometric parameters and glycogen content of soleus muscle treated with clenbuterol. Male Wistar (3-4 months old) rats were divided into 4 groups (N = 6 for each group): control, clenbuterol, immobilized, and immobilized treated with clenbuterol. Immobilization was performed with acrylic resin orthoses and 10 µg/kg body weight clenbuterol was administered subcutaneously for 7 days. The following parameters were measured the next day on soleus muscle: weight, glycogen content, cross-sectional area, and connective tissue content. The clenbuterol group showed an increase in glycogen (81.6%, 0.38 ± 0.09 vs 0.69 ± 0.06 mg/100 g; P < 0.05) without alteration in weight, cross-sectional area or connective tissue compared with the control group. The immobilized group showed a reduction in muscle weight (34.2%, 123.5 ± 5.3 vs 81.3 ± 4.6 mg; P < 0.05), glycogen content (31.6%, 0.38 ± 0.09 vs 0.26 ± 0.05 mg/100 mg; P < 0.05) and cross-sectional area (44.1%, 2574.9 ± 560.2 vs 1438.1 ± 352.2 µm²; P < 0.05) and an increase in connective tissue (216.5%, 8.82 ± 3.55 vs 27.92 ± 5.36%; P < 0.05). However, the immobilized + clenbuterol group showed an increase in weight (15.9%; 81.3 ± 4.6 vs 94.2 ± 4.3 mg; P < 0.05), glycogen content (92.3%, 0.26 ± 0.05 vs 0.50 ± 0.17 mg/100 mg; P < 0.05), and cross-sectional area (19.9%, 1438.1 ± 352.2 vs 1724.8 ± 365.5 µm²; P < 0.05) and a reduction in connective tissue (52.2%, 27.92 ± 5.36 vs 13.34 ± 6.86%; P < 0.05). Statistical analysis was performed using Kolmogorov-Smirnov and homoscedasticity tests. For the muscle weight and muscle glycogen content, two-way ANOVA and the Tukey test were used. For the cross-sectional area and connective tissue content, Kruskal-Wallis and Tukey tests were used. This study emphasizes the importance of anabolic pharmacological protection during immobilization to minimize skeletal muscle alterations resulting from disuse.

Central actions of glucocorticoids in the control of body fluid homeostasis: Review

The involvement of the hypothalamic-pituitary-adrenal axis in the control of body fluid homeostasis has been extensively investigated in the past few years. In the present study, we reviewed the recent results obtained using different approaches to investigate the effects of glucocorticoids on the mechanisms of oxytocin and vasopressin synthesis and secretion in response to acute and chronic plasma volume and osmolality changes. The data presented here suggest that glucocorticoids are not only involved in the mechanisms underlying the fast release but also in the transcriptional events that lead to decreased synthesis and secretion of these neuropeptides, particularly oxytocin, under diverse experimental conditions of altered fluid volume and tonicity. The endocannabinoid system, through its effects on glutamatergic neurotransmission within the hypothalamus and the nuclear factor κB-mediated transcriptional activity, seems to be also involved in the specific mechanisms by which glucocorticoids exert their central effects on neurohypophyseal hormone synthesis and secretion.

Evidence that blood pressure remains under the control of arterial baroreceptors in renal hypertensive rats

The purpose of the present study was to determine the range of the influence of the baroreflex on blood pressure in chronic renal hypertensive rats. Supramaximal electrical stimulation of the aortic depressor nerve and section of the baroreceptor nerves (sinoaortic denervation) were used to obtain a global analysis of the baroreceptor-sympathetic reflex in normotensive control and in chronic (2 months) 1-kidney, 1-clip hypertensive rats. The fall in blood pressure produced by electrical baroreceptor stimulation was greater in renal hypertensive rats than in normotensive controls (right nerve: -47 ± 8 vs -23 ± 4 mmHg; left nerve: -51 ± 7 vs -30 ± 4 mmHg; and both right and left nerves: -50 ± 8 vs -30 ± 4 mmHg; P < 0.05). Furthermore, the increase in blood pressure level produced by baroreceptor denervation in chronic renal hypertensive rats was similar to that observed in control animals 2-5 h (control: 163 ± 5 vs 121 ± 1 mmHg; 1K-1C: 203 ± 7 vs 170 ± 5 mmHg; P < 0.05) and 24 h (control: 149 ± 3 vs 121 ± 1 mmHg; 1K-1C: 198 ± 8 vs 170 ± 5 mmHg; P < 0.05) after sinoaortic denervation. Taken together, these data indicate that the central and peripheral components of the baroreflex are acting efficiently at higher arterial pressure in renal hypertensive rats when the aortic nerve is maximally stimulated or the activity is abolished.

Effect of curcumin on the proliferation and apoptosis of hepatic stellate cells

This study was designed to investigate the effect of curcumin (diferuloylmethane) on the proliferation and apoptosis of hepatic stellate cells (HSC). The cell line HSC-T6 (1.25 x 10(5) cells/mL) was incubated with curcumin and HSC proliferation was detected by a methyl thiazolyl tetrazolium colorimetric assay. HSC apoptosis was detected by flow cytometry, transmission electron microscope and agarose gel electrophoresis. HSC proliferation was significantly inhibited in a concentration-dependent manner (10.6 to 63.5%) after incubation with 20-100 μM curcumin, compared with a control group. At 20, 40, and 60 μM, after 24 h of incubation, curcumin was associated with a significant increase in the number of HSC in the G2/M phase, and a significant decrease in cell numbers in the S phase (P < 0.05). At these concentrations, curcumin was also associated with an increase in the apoptosis index of 15.3 ± 1.9, 26.7 ± 2.8, and 37.6 ± 4.4%, respectively, compared to control (1.9 ± 0.6%, P < 0.01). At 40 μM, the curcumin-induced apoptosis index at 12, 24, 36, and 48 h of incubation was 12.0 ± 2.4, 26.7 ± 3.5, 33.8 ± 1.8, and 49.3 ± 1.6%, respectively (P < 0.01). In conclusion, curcumin inhibits the in vitro proliferation of HSCs in the G2/M phase of the cell cycle and also induces apoptosis in a concentration- and time-dependent manner. The in vivo effect of curcumin on HSCs requires further investigation.

Follow-up of patients treated with retinoic acid for the control of radioiodine non-responsive advanced thyroid carcinoma

During thyroid tumor progression, cellular de-differentiation may occur and it is commonly accompanied by metastatic spread and loss of iodine uptake. Retinoic acid (RA) administration might increase iodine uptake in about 40% of patients, suggesting that RA could be a promising therapeutic option for radioiodine non-responsive thyroid carcinoma, although a prospective study with a long-term follow-up has not been reported. This was a clinical prospective study assessing the value of 13-cis-RA in patients with advanced thyroid carcinoma and its impact on major outcomes such as tumor regression and cancer-related death with a long-term follow-up of patients submitted to radioiodine (131I) therapy after RA administration. Sixteen patients with inoperable disease and no significant radioiodine uptake on post-therapy scan were selected. Patients were treated orally with 13-cis-RA at a dose of 1.0 to 1.5 mg·kg-1·day-1 for 5 weeks and then submitted to radioiodine therapy (150 mCi) after thyroxine withdrawal. A whole body scan was obtained 5 to 7 days after the radioactive iodine therapy. RECIST criteria were used to evaluate the response. An objective partial response rate was observed in 18.8%, a stable disease rate in 25% and a progression disease rate in 56.2%. Five patients died (62.5%) in the group classified as progression of disease. Progression-free survival rate (PFS) ranged from 72 to 12 months, with a median PFS of 26.5 months. RA may be an option for advanced de-differentiated thyroid cancer, due to the low rate of side effects.

The effects of stretching on the flexibility, muscle performance and functionality of institutionalized older women

Stretching has been widely used to increase the range of motion. We assessed the effects of a stretching program on muscle-tendon length, flexibility, torque, and activities of daily living of institutionalized older women. Inclusion/exclusion criteria were according to Mini-Mental State Examination (MMSE) (>13), Barthel Index (>13) and Lysholm Scoring Scale (>84). Seventeen 67 ± 9 year-old elderly women from a nursing home were divided into 2 groups at random: the control group (CG, N = 9) participated in enjoyable cultural activities; the stretching group (SG, N = 8) performed active stretching of hamstrings, 4 bouts of 1 min each. Both groups were supervised three times per week over a period of 8 weeks. Peak torque was assessed by an isokinetic method. Both groups were evaluated by a photogrammetric method to assess muscle-tendon length of uni- and biarticular hip flexors and hamstring flexibility. All measurements were analyzed before and after 8 weeks by two-way ANOVA with the level of significance set at 5%. Hamstring flexibility increased by 30% in the SG group compared to pre-training (76.5 ± 13.0° vs 59.5 ± 9.0°, P = 0.0002) and by 9.2% compared to the CG group (76.5 ± 13.0° vs 64.0 ± 12.0°, P = 0.0018). Muscle-tendon lengths of hip biarticular flexor muscles (124 ± 6.8° vs 118.3 ± 7.6°, 5.0 ± 7.0%, P = 0.031) and eccentric knee extensor peak torque were decreased in the CG group compared to pre-test values (-49.4 ± 16.8 vs -60.5 ± 18.9 Nm, -15.7 ± 20%, P = 0.048). The stretching program was sufficient to increase hamstring flexibility and a lack of stretching can cause reduction of muscle performance.

Effect of triiodothyronine on the maxilla and masseter muscles of the rat stomatognathic system

The maxilla and masseter muscles are components of the stomatognathic system involved in chewing, which is frequently affected by physical forces such as gravity, and by dental, orthodontic and orthopedic procedures. Thyroid hormones (TH) are known to regulate the expression of genes that control bone mass and the oxidative properties of muscles; however, little is known about the effects of TH on the stomatognathic system. This study investigated this issue by evaluating: i) osteoprotegerin (OPG) and osteopontine (OPN) mRNA expression in the maxilla and ii) myoglobin (Mb) mRNA and protein expression, as well as fiber composition of the masseter. Male Wistar rats (~250 g) were divided into thyroidectomized (Tx) and sham-operated (SO) groups (N = 24/group) treated with T3 or saline (0.9%) for 15 days. Thyroidectomy increased OPG (~40%) and OPN (~75%) mRNA expression, while T3 treatment reduced OPG (~40%) and OPN (~75%) in Tx, and both (~50%) in SO rats. Masseter Mb mRNA expression and fiber type composition remained unchanged, despite the induction of hypo- and hyperthyroidism. However, Mb content was decreased in Tx rats even after T3 treatment. Since OPG and OPN are key proteins involved in the osteoclastogenesis inhibition and bone mineralization, respectively, and that Mb functions as a muscle store of O2 allowing muscles to be more resistant to fatigue, the present data indicate that TH also interfere with maxilla remodeling and the oxidative properties of the masseter, influencing the function of the stomatognathic system, which may require attention during dental, orthodontic and orthopedic procedures in patients with thyroid diseases.

Involvement of β3-adrenergic receptors in the control of food intake in rats

This study examined the food intake changes evoked by intracerebroventricular (icv) injection of a selective agonist (BRL37344, 2 and 20 nmol) or antagonist (SR59230A, 10 and 50 nmol) of β3-adrenergic receptors in 24-h fasted rats (adult male Wistar rats, 200-350 g, N = 6/treatment). The animals were also pretreated with saline icv (SAL) or SR59230A (50 nmol) followed by BRL37344 (20 nmol) or SAL in order to determine the selectivity of the effects evoked by BRL37344 on food intake or the selectivity of the effects evoked by SR59230A on risk assessment (RA) behavior. The highest dose of BRL37344 (N = 7) decreased food intake 1 h after the treatment (6.4 ± 0.5 g in SAL-treated vs 4.2 ± 0.8 g in drug-treated rats). While both doses of SR59230A failed to affect food intake (5.1 ± 1.1 g for 10 nmol and 6.0 ± 1.8 g for 50 nmol), this treatment reduced the RA frequency (number/30 min) (4 ± 2 for SAL-treated vs 1 ± 1 for 10 nmol and 0.5 ± 1 for 50 nmol SR59230A-treated rats), an ethological parameter related to anxiety. While pretreatment with SR59230A (7.0 ± 0.5 g) abolished the hypophagia induced by BRL37344 (3.6 ± 0.9 g), BRL37344 suppressed the reduction in RA frequency caused by SR59230A. These results show that the hypophagia caused by BRL37344 is selectively mediated by β3-adrenergic receptors within the central nervous system. Moreover, they suggest the involvement of these receptors in the control of anxiety.