Search

BACKGROUND: Both propofol use and advanced age are predictors of intraoperative hypotension. We previously demonstrated that propofol enhances vasodilation in mesenteric arteries from aged rats, partly due to increased nitric oxide (NO) bioavailability. Patients chronically treated with angiotensin-converting enzyme (ACE) inhibitors may exhibit refractory hypotension under general anesthesia. We hypothesized that propofol enhances NO-mediated vasodilation in arteries from aged rats chronically treated with ACE inhibitors. METHODS: Sprague-Dawley rats aged 12 to 13 months were treated with or without captopril for 7 to 8 weeks, yielding a final age of 14 to 15 months at the time of experimentation. Before euthanasia, arterial blood pressures were obtained through carotid artery cannulation. Concentration-response curves to propofol (0.1-100 µM) or methacholine (MCh) (0.01-3 µM) were then assessed on isolated resistance mesenteric arteries (100-200 μm diameter) from both treatment (captopril) and control rats. MCh relaxation was also assessed after propofol pretreatment (1 and 10 µM). N(G)-nitro-l-arginine methyl ester (l-NAME) (100 µM) and meclofenamate (10 µM) were used to inhibit NO and prostaglandin synthesis, respectively. Concentration-response data were summarized as 50% of the maximum relaxation response or area under the curve. RESULTS: Mean arterial blood pressure in the captopril-treated rats was lower than in untreated rats (P = 0.049). When comparing relaxation in arteries from captopril-treated versus untreated rats, concentration-response curves revealed that captopril-treated rats display greater direct propofol relaxation (P = 0.018). MCh relaxation in the absence of propofol, however, was not different between captopril-treated and untreated rats (P = 0.80). Propofol pretreatment increased MCh relaxation in arteries from captopril-treated compared with untreated rats (P = 0.029 for 1 µM and P = 0.020 for 10 µM). Meclofenamate did not have an effect in this response (P = 0.22). l-NAME-dependent inhibition of MCh relaxation, however, was greater in arteries from control compared with captopril-treated rats (P = 0.0077). However, propofol increased the proportion of NO-dependent vasodilation to MCh similarly in both groups. This suggests that other vasodilatory pathways are involved in the differential response to MCh in the presence of propofol in captopril-treated rats. CONCLUSIONS: Our results show that mesenteric arterial relaxation in response to propofol, both by direct stimulation and through modulation of endothelium-dependent mechanisms, is, in part, NO-dependent. In captopril-treated rats, propofol further increased arterial relaxation through a non-NO-dependent vasodilating pathway (e.g., endothelium-derived hyperpolarizing factor), which may account for enhanced vasodilation during propofol exposure in patients treated with ACE inhibitors.

Recently, the efficacy of functional electrical stimulation (FES) cycling have been demonstrated on the improvement of strength and motor control in adults with stroke. FES-cycling, providing a repetitive goal-oriented task, could facilitate cortical reorganization and utilization of residual cortico-spinal pathways. These benefits could be more enhanced in children because of the greater plasticity and flexibility of their central nervous system.The aim of the present case report study was to explore the feasibility of FES-cycling in children with cerebral palsy (CP) and to provide a set of instrumental measures able to evaluate the effects of this novel treatment on cycling and walking ability.Interventional study.Two ambulant outpatient children with diplegic CP were recruited by the "E. Medea" Scientific Institute.Patients followed a FES-cycling treatment for 30 minutes a day, 3 days a week for 7 weeks. Pre and post treatment tests were performed, namely clinical measures and electromyographic, kinematic and oxygen expenditure analysis during gait and cycling.The treatment was safe, feasible and well accepted by the 2 children. After treatment both patients achieved a more symmetrical muscular strategy during voluntary cycling and gait and a significant reduction of muscle co-contractions during cycling. These improvements were corroborated by a decrease in oxygen expenditure during the post test for one of the two children, the less impaired, implying a better exploiting of bi-articular muscles.FES-cycling is feasible and safe and it may be an alternative rehabilitation method for diplegic CP patients. The set of instrumental measurements proposed seems to be a valuable tool for functional assessment to identify subclinical anomalies and improvements on cycling and gait in CP patients.

To the EditorMany reports exist which advocate the use of intralipidto reverse local anesthetic cardiovascular toxicity. Theprecise mechanism of action, while unknown at the presenttime, may include a combination of reduced tissue bindingof local anesthetic by a plasma-lipid phase and a beneficialmetabolic effect involving cardiac mitochondria.

Compared with the most commonly used gene-specific hybridization probes (subcloned DNA fragments, PCR amplimer products or DNA oligonucleotides), the overgo hybridization method (Ross et al., 1999) has several advantages (better hybridization kinetics, higher specificity labeling, low background of hybridization, low rate of false positives, high throughput, and ease of handling). The foundation of these advantages is the short sequence that is needed for an overgo probe, which increases the likelihood of finding suitable single copy regions from EST sequences or unigenes.

Hind III BAC library of Citrus sinensis cv "Vanoglia" has been constructed.

One of the most important goal in Citrus genetic improvement is the obtainment of seedless cultivars. Among the available strategies, we used the accomplishment of suitable interploid crosses (2X x 4X). To this purpose, three allotetraploid somatic hybrids have been used as pollen parent in sexual crosses with diploid 'Femminello' lemon, and autotetraploid 'Dancy' mandarin as pollen parent in crosses with diploid grapefruit and mandarin.

Repetitive sequences constitute a significant component of most eukaryotic genomes, and the isolation and characterization of repetitive DNA sequences provide an insight into the organization of the genome of interest. Here, we report the isolation and the molecular analysis and methylation status of a novel tandemly organized repetitive DNA sequence from the genome of Poncirus trifoliata. Digestion of P. trifoliate DNA with Afa I produced a prominent fragment of approximately 400 bp. Southern blotting analysis of genomic DNA digested with the same enzyme revealed a ladder composed of DNA fragments that are multimers of the 400-bp Afa I band, indicating that the repetitive DNA is arrayed in tandem. This suggests that Afa I isolated a novel satellite that we have called Poncirus trifoliata satellite DNA 400 (PN400). This satellite composes 25% of the genome and it is also present in lemon, sour orange and kumquat. Analysis of the methylation status demonstrated that the cytosines in CCGG sequences in this satellite were methylated.

In order to obtain new somatic hybrids for genetic improvement of mandarin, several protoplast fusion experiments have been carried out. Isozyme banding pattern analysis and flow cytometry were used for early nuclear characterization. DNA of the hybrids was then extracted and subjected to ISSR-PCR using different primers anchored at 5’ and 3’ends. Successively, for cytoplasmic characterization of the hybrids, CAPs using universal mtDNA and cpDNA primers were performed. Our results showed that these new genotypes are definitely cybrids resulting asymmetric hybrids, in which the nucleus is inherited from the leaf parent and mitochondria from callus parent.

Female sexual function is under-studied, and mechanisms of clitoral engorgement-relaxation are incompletely understood. Penile erection results from nitric oxide (NO) -induced cyclic guanosine monophosphate (cGMP) accumulation. cGMP-dependent protein kinase (PKG) activates large-conductance, calcium-activated potassium channels (BK(Ca)), thereby hyperpolarizing and relaxing vascular and trabecular smooth muscle cells, allowing engorgement. We hypothesize rat clitorises relax by a similar mechanism. Rat clitorises express components of the proposed pathway: neuronal and endothelial NO synthases, soluble guanylyl cyclase (sGC), type 5 phosphodiesterase (PDE-5), and BK(Ca) channels. The NO donor diethylamine NONOate (DEANO), the PKG activator 8-pCPT-cGMP, and the PDE-5 inhibitor sildenafil, cause dose-dependent clitoral relaxation that is inhibited by antagonists of PKG (Rp-8-Br-cGMPS) or BK(Ca) channels (iberiotoxin). Electrical field stimulation induces tetrodotoxin-sensitive NO release and relaxation that is inhibited by the Na+ channel blocker tetrodotoxin or sGC inhibitor 1H-(1,2,4)oxadiozolo(4,3-a)quinoxalin-1-one. Human BK(Ca) channels, transferred to Chinese hamster ovary cells via an adenoviral vector, and endogenous rat clitoral smooth muscle K+ current are activated by this PKG-dependent mechanism. Laser confocal microscopy reveals protein expression of BK(Ca) channels on clitoral smooth muscle cells; these cells exhibit BK(Ca) channel activity that is activated by both DEANO and sildenafil. We conclude that neurovascular derived NO causes clitoral relaxation via a PKG-dependent activation of BK(Ca) channels. The BK(Ca) channel is an appealing target for drug therapy of female erectile dysfunction.

Objective— Angiotensin II (AII) has been shown to increase endothelial NAD(P)H oxidase activity, which is a source of superoxide anion that in turn may induce the formation of peroxynitrite. Estrogen (E 2 ) has been reported to have vascular protective effects. In this study, we hypothesized that E 2 reduces the AII-induced expression of NAD(P)H oxidase and peroxynitrite in endothelial cells. Methods and Results— Endothelial cells were cultured and stimulated with AII in the absence or presence of E 2 . Western blots were used to assess nitric oxide synthase (NOS) and NAD(P)H oxidase expression. Immunofluorescence of nitrotyrosine provided evidence of peroxynitrite formation. Our data indicate that AII increased the expression of endothelial NOS, inducible NOS, and NAD(P)H oxidase in a dose-dependent manner, which was attenuated by incubation with either E 2 , superoxide dismutase, or the AII type 1 receptor (AT 1 R) inhibitor candesartan. Estrogen as well as superoxide dismutase also inhibited AII-induced AT 1 R expression and nitrotyrosine staining. The effects of E 2 on the AII responses were not inhibited by the E 2 receptor antagonist ICI-182,780. Conclusions— AII stimulation of endothelial cells increases expression of NAD(P)H oxidase and NOS, which may contribute to oxidative stress, as evidenced by peroxynitrite formation. E 2 inhibits these AII effects, possibly through reduced AT 1 R expression.

One of the most important goals in genetic improvement of lemon in the Mediterranean area is certainly the obtainment of new genotypes tolerant or resistant to mal secco disease. Two lemon cybrids (one diploid and one autotetraploid) with an intermediate degree of resistance to mal secco disease, spontaneously obtained by symmetric protoplast fusion between ‘Valencia’ orange and ‘Femminello’ lemon, were used as mother plants in backcrosses with a diploid clone of ‘Femminello’ Lemon Tolerant to the Mal Secco disease (LTMS) to improve tolerance and fruit quality. Since ‘Femminello’ lemon normally reproduces apomictically by nucellar embryony, an embryo-rescue technique was applied 105 days after pollination to recover zygotic embryos. A total of 66 plantlets were regenerated as follows: 20 from the 4nX2n cross, and 46 from the 2nX2n cross. In order to distinguish zygotic embryos from nucellars, flow cytometry was applied on all the 66 plantlets; and a modified AFLP (Amplified Fragment-Length Polymorphism) technique and microsatellite analysis were carried out on a sample of 20 seedlings (6 triploids and 14 diploids). Here we report the usefulness and reliability of the tested techniques and discuss the results obtained. Flow cytometry (FCM) was useful only in inter-ploid crosses, identifying 6 triploids, 4 of which were also detected by SSR (Simple Sequence Repeat) analysis and only 2 by modified AFLP. Among the 14 tested diploids, 5 zygotic genotypes were identified by SSR and 4 by modified AFLP. In our hands, the SSR technique appeared to be more suitable than modified AFLP.

Somatic hybrids between Redblush and Duncan grapefruit (Citrus paradisi Macfadyen) with Avana, Tardivo di Ciaculli mandarin (Citrus deliciosa Tenore) and Fortune mandarin (Citrus reticulata Blanco) were obtained by symmetric protoplast fusion. Isozyme banding pattern analysis and flow cytometry were used for early screening. DNA of the hybrids was then extracted and subjected to inter-simple sequence repeat polymerase chain reaction (ISSR-PCR) using different primers anchored at the 5’ and 3’ ends. PCR products were resolved on 1.5% agarose gel and stained with ethidium bromide. All the bands from the parental cultivars were present in the somatic hybrids. The usefulness and efficiency of the ISSR-PCR method as a screening technique for Citrus somatic hybrids are discussed.