In vivo degradation of banana starch: Structural characterization of the degradation process

This work reports the first ultrastructural investigation into the degradation process that starch granules isolated from bananas (cv. Nanicao) undergo during ripening. Starch granules from green bananas had a smooth surface, while granules from ripe bananas were more elongated with parallel striations, as revealed by CSLM and SEM. AFM images revealed that the first layer covering the granule surface is composed of a hard material and, as degradation proceeds, hard and soft regions seem to be repeated at regular intervals. WAXD patterns of banana starches were C-type, and the crystalline index was reduced during ripening. The B-/A-type ratio was increased, indicating the preferential degradation of the A-type allomorph. The branch-chain length distribution showed predominantly short chains of amylopectin (A and B1-chain). The fa/fb ratio was reduced during degradation, while amylose content was increased. The results allowed a detailed understanding of the changes that starch granules undergo during banana ripening. (C) 2010 Elsevier Ltd. All rights reserved.

Induction and maintenance of in vivo ventricular fibrillation in rabbits

Aim: To provide new sustainable in vivo models of ventricular fibrillation in rabbits. Methods: New Zealand rabbits were submitted to anaesthesia and mechanical ventilation. after which ventricular fibrillation was induced through electrical stimulation (for 2 min at 100 Hz, with 2-ms pulses, 10 mA. and 10V) directly to the heart. To that end, the animals were divided into two groups: right ventricle (n = 11) and left ventricle (n = 11). In group right ventricle, the thoracic cavity was exposed, and a catheter was introduced into the right ventricle via the right jugular vein. in group left ventricle, the thorax remained closed, and the catheter was introduced into the left ventricle via the left common carotid artery (cervical access). Results: Sustained ventricular fibrillation was achieved in 100% of group right ventricle rabbits (n = 11 and in 82% of group left ventricle rabbits (n = 9). Conclusion: Both models proved appropriate for achieving sustained ventricular fibrillation. However, in view of the invasiveness of the procedure adopted in group right ventricle, the experimental conditions used in group left ventricle seemed more physiological and more effective in inducing sustained ventricular fibrillation. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Penile erection induced in vivo by a purified toxin from the Brazilian spider Phoneutria nigriventer

OBJECTIVES To verify the effect on erectile tissue of mice of two neuropeptides extracted from the poison of a spider, Phoneutric nigriventer, (Tx2-5 and -6. termed `eretina`) after direct injection into the corpus cavernosum, to assess the minimum dosage necessary for effect. the time for initiation of action, the local duration of the erection, histological effects and the presence of local and systemic side-effects. MATERIALS AND METHODS When applied intraperitoneally, eretina promotes the relaxation of cavernous smooth muscle, thus causing penile erection. Thirty-five mice were divided in two groups; 10 control mice were injected 20 mu L of saline solution, and in the treated group, 25 mice were divided into groups of five and each subgroup received eretina in decreasing doses (0.024, 0.012, 0.006, 0.003 and 0.0015 mu g/kg) until the minimum dose that produced an erection was determined. After treatment 211 mice were monitored to determine the response and any collateral effects. RESULTS The minimum dose producing an erection was 0.006 mu g/kg, the five mice in this group having evidence of an erection at 35-45 min after injection. The histology of the cavernosum of mice treated with eretina showed dilatation and congestion of the vascular spaces with more blood than in controls. With the minimum dose there were no local or systemic collateral effects and the erection was lost after 120-140 min. CONCLUSION The minimum dose of eretina producing an erection in mice was determined. and the agent was safe for this use as it did not produce any collateral toxic effects. These studies indicate a possible means of determining the mechanism of action of eretina.

In vivo electrochemical characterization and inflammatory response of multiwalled carbon nanotube-based electrodes in rat hippocampus

Stimulating neural electrodes are required to deliver charge to an environment that presents itself as hostile. The electrodes need to maintain their electrical characteristics (charge and impedance) in vivo for a proper functioning of neural prostheses. Here we design implantable multi-walled carbon nanotubes coating for stainless steel substrate electrodes, targeted at wide frequency stimulation of deep brain structures. In well-controlled, low-frequency stimulation acute experiments, we show that multi-walled carbon nanotube electrodes maintain their charge storage capacity (CSC) and impedance in vivo. The difference in average CSCs (n = 4) between the in vivo (1.111 mC cm(-2)) and in vitro (1.008 mC cm(-2)) model was statistically insignificant (p > 0.05 or P-value = 0.715, two tailed). We also report on the transcription levels of the pro-inflammatory cytokine IL-1 beta and TLR2 receptor as an immediate response to low-frequency stimulation using RT-PCR. We show here that the IL-1 beta is part of the inflammatory response to low-frequency stimulation, but TLR2 is not significantly increased in stimulated tissue when compared to controls. The early stages of neuroinflammation due to mechanical and electrical trauma induced by implants can be better understood by detection of pro-inflammatory molecules rather than by histological studies. Tracking of such quantitative response profits from better analysis methods over several temporal and spatial scales. Our results concerning the evaluation of such inflammatory molecules revealed that transcripts for the cytokine IL-1 beta are upregulated in response to low-frequency stimulation, whereas no modulation was observed for TLR2. This result indicates that the early response of the brain to mechanical trauma and low-frequency stimulation activates the IL-1 beta signaling cascade but not that of TLR2.

Testosterone represses ubiquitin ligases atrogin-1 and Murf-1 expression in an androgen-sensitive rat skeletal muscle in vivo

Pires-Oliveira M, Maragno AL, Parreiras-E-Silva LT, Chiavegatti T, Gomes MD, Godinho RO. Testosterone represses ubiquitin ligases atrogin-1 and Murf-1 expression in an androgen-sensitive rat skeletal muscle in vivo. J Appl Physiol 108: 266-273, 2010. First published November 19, 2009; doi:10.1152/japplphysiol.00490.2009.-Skeletal muscle atrophy induced by denervation and metabolic diseases has been associated with increased ubiquitin ligase expression. In the present study, we evaluate the influence of androgens on muscle ubiquitin ligases atrogin-1/MAFbx/FBXO32 and Murf-1/Trim63 expression and its correlation with maintenance of muscle mass by using the testosterone-dependent fast-twitch levator ani muscle (LA) from normal or castrated adult male Wistar rats. Gene expression was determined by qRT-PCR and/or immunoblotting. Castration induced progressive loss of LA mass (30% of control, 90 days) and an exponential decrease of LA cytoplasm-to-nucleus ratio (nuclear domain; 22% of control after 60 days). Testosterone deprivation induced a 31-fold increase in LA atrogin-1 mRNA and an 18-fold increase in Murf-1 mRNA detected after 2 and 7 days of castration, respectively. Acute (24 h) testosterone administration fully repressed atrogin-1 and Murf-1 mRNA expression to control levels. Atrogin-1 protein was also increased by castration up to 170% after 30 days. Testosterone administration for 7 days restored atrogin-1 protein to control levels. In addition to the well known stimulus of protein synthesis, our results show that testosterone maintains muscle mass by repressing ubiquitin ligases, indicating that inhibition of ubiquitin-proteasome catabolic system is critical for trophic action of androgens in skeletal muscle. Besides, since neither castration nor androgen treatment had any effect on weight or ubiquitin ligases mRNA levels of extensor digitorum longus muscle, a fast-twitch muscle with low androgen sensitivity, our study shows that perineal muscle LA is a suitable in vivo model to evaluate regulation of muscle proteolysis, closely resembling human muscle responsiveness to androgens.

In Vivo Effects of Bothrops jararaca Venom on Metabolic Profile and on Muscle Protein Metabolism in Rats

This study investigated the in vivo effects of the Bothrops Jararaca venom (BjV) on general metabolic profile and, specifically. oil muscle protein metabolism in rats. The crude venom (0.4 mg/kg body weight, IV) was infused in awake rats, and plasma activity of enzymes and metabolites levels were determined after 1, 2, 3, and 4 hours. BjV increased urea, lactate, and activities of creatine kinase. lactate dehydrogenase. and aspartate aminotransferase after 4 hours. The content of liver glycogen was reduced by BjV. Protein metabolism was evaluated by means of microdialysis technique and in isolated muscles. BjV induced increase in the muscle interstitial-arterial tyrosine concentration difference. indicating a high protein catabolism. The myotoxicity induced by this venom is associated with reduction of protein synthesis and increase in rates of overall proteolysis, which was accompanied by activation of lysosomal and ubiquitin-proteasome systems without changes in protein levels of cathepsins and ubiquitin-protein conjugates.

Investigation of the in vivo activity of CYP3A in Brazilian volunteers: comparison of midazolam and omeprazole as drug markers

Objective This study compares midazolam with omeprazole as marker drugs for the evaluation of CYP3A activity in nine healthy self-reported white Brazilian volunteers. Methods Omeprazole was also used to evaluate the CYP2C19 phenotype. The volunteers received p.o. 20 mg omeprazole, and blood samples were collected 3.5 h after drug administration. After a washout period of 10 days, the volunteers received p.o. 15 mg midazolam maleate, and serial blood samples were collected up to 6 h after administration of the drug. CYP2C19 was genotyped for the allelic variants CYP2C19*1, CYP2C19*2, CYP2C19*3, and CYP2C19*17. Analysis of omeprazole, hydroxyomeprazole, omeprazole sulfone, and midazolam in plasma was carried out by LC-MS/MS. Results The volunteers genotyped as CYP2C19*1*17, CYP2C19*17*17, CYP2C19*1*1 (n=8), or CYP2C19*17*2 (n=1) presented a median hydroxylation index (omeprazole/hydroxyomeprazole) of 1.35, indicating that all of them were extensive metabolizers of CYP2C19. The volunteers (n=9) presented a 0.12 log of the omeprazole/sulfone ratio and a median oral clearance of midazolam of 17.89 ml min(-1) kg(-1), suggesting normal CYP3A activity. Conclusions Orthogonal regression analysis between midazolam clearance and log of the plasma concentrations of the omeprazole/omeprazole sulfone ratio (R=-0.7544, P < 0.05) suggests that both midazolam and omeprazole can be used as markers of CYP3A activity in the population investigated.

Leuprolide acetate reduces both in vivo and in vitro ovarian steroidogenesis in infertile women undergoing assisted reproduction

Despite the probable inhibitory effects of GnRH analogues on ovarian steroidogenesis in vitro, their association with assisted reproduction protocols shows favorable results. This suggests that there are important differences in the behaviors of these drugs when administered in vivo versus in vitro. To clarify these differences, this study was designed to analyze the effect of leuprolide acetate (LA) on ovarian steroidogenesis in women undergoing In Vitro Fertilization (IVF). A prospective, randomized open label study was conducted on 14 women (26-35 years): seven receiving only gonadotrophins (Group 1) and seven receiving gonadotrophin plus LA at 1mg/day (Group 2). The LA in vivo effect was determined with serum and follicular fluid (FF) samples and via luteinized granulosa cell cultivation (GCC), where cells were obtained during oocyte retrieval after ovarian hyperstimulation. In vitro analysis was performed via addition of LA to GCC only for Group 1 (without LA) at progressively higher concentrations (0, 10(-12), 10(-9) and 10(-6) M). In vivo, the main observation was a reduction in androgen production in Group 2, represented by lower androstenedione production in FF (G1 = 6479 +/- 3458; G2 = 3021 +/- 1119 ng/ml; p = 0.04) and a lower testosterone peak in GC at 96 h (G1 = 0.64 +/- 0.12 ng/ml; G2 = 0.50 +/- 0.19ng/ml; P = 0.02), but a higher fertilization rate (G1 = 67%; G2 = 83%; p = 0.009). in vitro, testosterone, estradiol and progesterone were also reduced by LA, even though this reduction occurred for progesterone only at the highest LA dosage (10(-6) M; 606.0 +/- 114.3 ng/ml versus 1524.0 +/- 246.5 ng/ml; p=0.02). Results show that LA reduces ovarian steroidogenesis in vivo by essentially inhibiting androgen synthesis; whereas, in vitro, ovarian steroidogenesis is reduced overall. (C) 2008 Elsevier Inc. All rights reserved.

Hematopoietic response of rats exposed to the impact of an acute psychophysiological stressor on responsiveness to an in vivo challenge with Listeria monocytogenes: Modulation by Chlorella vulgaris prophylactic treatment

In this study, we investigated the hematopoietic response of rats pretreated with CV and exposed to the impact of acute escapable, inescapable or psychogenical stress on responsiveness to an in vivo challenge with Listeria monocytogenes. No consistent changes were observed after exposure to escapable footshock. Conversely, the impact of uncontrollable stress (inescapable and psychogenical) was manifested by an early onset and increased severity and duration of myrelossuppression produced by the infection. Small size CFU-CM colonies and increased numbers of clusters were observed, concurrently to a greater expansion in the more mature population of bone marrow granulocytes. No differences were observed between the responses of both uncontrollable stress regimens. CV prevented the myelossuppression caused by stress/infection due to increased numbers of CFU-GM in the bone marrow. Colonies of cells tightly packed, with a very condensed nucleus; in association with a greater expansion in the more immature population of bone marrow granulocytes were observed. Investigation of the production of colony-stimulating factors revealed increased colony-stimulating activity (CSA) in the serum of normal and infected/stressed rats treated with the algae. CV treatment restored/enhanced the changes produced by stress/infection in total and differential bone marrow and peripheral cells counts. Further studies demonstrated that INF-gamma is significantly reduced, whereas IL-10 is significantly increased after exposure to Uncontrollable stress. Treatment with CV significantly increased INF-gamma levels and diminished the levels of IL-10. Uncontrollable stress reduced the protection afforded by CV to a lethal dose of L. monocytogenes, with survival rates being reduced from (50%) in infected rats to 20% in infected/stressed rats. All together, our results suggest Chlorella treatment as an effective tool for the prophylaxis of post-stress myelossupression, including the detrimental effect of stress on the course and outcome of infections. (C) 2008 Elsevier Inc. All rights reserved.

Effect of laser phototherapy on recurring herpes labialis prevention: an in vivo study

Alternative treatment for recurrent labial infection by herpes simplex virus (HSV) have been considered. The aim of this study was to evaluate the effectiveness of laser phototherapy in prevention and reduction of severity of labial manifestations of herpes labialis virus. Seventy-one patients, divided into experimental (n = 41) and control (n = 30) groups were followed up for 16 months. Patients in the control group were treated topically with aciclovir and patients in the experimental group were subjected to laser phototherapy (one session per week, 10 weeks): 780 nm, 60 mW, 3.0 J/cm(2) or 4.5 J/cm(2) on healthy (no HSV-1 infection) and affected (with HSV-1 infection) tissues. Patients in the experimental group presented a significant decrease in dimension of herpes labialis lesions (P = 0.013) and inflammatory edema (P = 0.031). The reduction in pain level (P = 0.051) and monthly recurrences (P = 0.076) did not reach statistical significance. This study represents an in vivo indication that this treatment should be further considered as an effective alternative to therapeutic regimens for herpes labialis lesions.

Comparative Analysis between Chamomilla recutita and Corticosteroids on Wound Healing. An In Vitro and In Vivo Study

The comparison of chamomile and corticosteroids for treating ulcers was done in vitro and in vivo. The experimental groups were: control; chamomile recutita; triamcinolone acetonide and clobetasol propionate. For the in vitro study the cell viability of fibroblasts cultured for 24 h in media conditioned by the substances was obtained by the MITT reduction analysis. For the in vivo study, 125 male rats were submitted to experimental ulcers treated or not (control) by the substances tested. At 1, 3, 5, 7 and 14 days later 5 animals of each group were sacrificed. The lesions were analyzed by means of clinical observation and histological wound-healing grading. Data were compared by ANOVA (p <= 0.05). All experimental groups presented positive cell viability in 24 h. The cultures treated with chamomile presented the smallest cell viability. All animals of the chamomile group exhibited complete wound healing 9 days before the other groups. Complete repaired lesions were observed after 5 days of treatment only in the chamomile group. Animals treated with chamomile presented significantly faster wound healing in comparison to those treated with corticosteroids. Based on the conditions of this study, we concluded that chamomile in comparison to corticosteroids promotes faster wound healing process. Copyright (C) 2008 John Wiley & Sons, Ltd.

In vivo Dentin Microhardness beneath a Calcium-Phosphate Cement

A minimally invasive caries-removal technique preserves potentially repairable, caries-affected dentin. Mineral-releasing cements may promote remineralization of soft residual dentin. This study evaluated the in vivo remineralization capacity of resin-based calcium-phosphate cement (Ca-PO(4)) used for indirect pulp-capping. Permanent carious and sound teeth indicated for extraction were excavated and restored either with or without the Ca-PO(4) base (control), followed by adhesive restoration. Study teeth were extracted after 3 months, followed by sectioning and in vitro microhardness analysis of the cavity floor to 115-mu m depth. Caries-affected dentin that received acid conditioning prior to Ca-PO(4) basing showed significantly increased Knoop hardness near the cavity floor. The non-etched group presented results similar to those of the non-treated group. Acid etching prior to cement application increased microhardness of residual dentin near the interface after 3 months in situ.

In vivo Dentin Remineralization by Calcium-Phosphate Cement

Minimally invasive caries-removal procedures remove only caries-infected dentin and preserve caries-affected dentin that becomes remineralized. Dental cements containing calcium phosphate promote remineralization. This study evaluated the in vivo remineralization capacity of resin-based calcium-phosphate cement (Ca-P) used for indirect pulp-capping. Carious and sound teeth indicated for extraction were randomly restored with the Ca-P base or without base (control), followed by adhesive restoration. Study teeth were extracted after three months, followed by elemental analysis of the cavity floor. Mineral content of affected or sound dentin at the cavity floor was quantified by electron probe micro-analysis to 100-mu m depth. After three months, caries-affected dentin underneath the Ca-P base showed significantly increased calcium and phosphorus content to a depth of 30 mu m. Mineral content of treated caries-affected dentin was in the range of healthy dentin, revealing the capacity of Ca-P base to promote remineralization of caries-affected dentin.

In vivo Acid Etching Effect on Bacteria within Caries-Affected Dentin

Acid etching procedures may disrupt residual bacteria and contribute to the success of incomplete caries removal followed by adhesive restoration. This study evaluated the in vivo effect of acid etching on cariogenic bacterial activity within affected dentin after minimally invasive treatment of caries lesions. Twenty-eight carious permanent teeth received standardized selective caries removal and random acid etch treatment (E) or not (NE) prior to adhesive restoration. Baseline and 3-month dentin biopsies were collected. The number of bacteria and activity of total bacterial cells and Streptococcus mutans were determined by quantitative PCR and RT-PCR. No statistically significant differences were observed in total bacterial number and activity between E and NE treatments (p > 0.3008). For NE, however, the residual S. mutans bacterial cells were reduced (p = 0.0027), while the activity per cell was significantly increased (p = 0.0010) after reentry at 3 months after restoration. This effect was not observed in group E. Although no significant differences were found between groups, this study suggests that acid etching of affected dentin prior to adhesive restoration may directly or indirectly have an inhibitive effect on the activity of residual cariogenic bacteria. Further research is required to investigate this potential effect. Copyright (C) 2010 S. Karger AG, Basel