Core temperature circadian rhythm across aging in Spontaneously Hypertensive Rats.

The purpose of this study was to evaluate the circadian rhythm of core temperature (Tcore) across aging in Spontaneously Hypertensive Rats (SHR) with comparison to the two rat strains often used as their normotensive control animals, namely, Wistar (WIS) and Wistar Kyoto (WKY). METHODS: WIS, WKY and SHR rats were subdivided into three different groups according their age: WIS16, WIS48, WIS72, WKY16, WKY48, WKY72, SHR16, SHR48 and SHR72 weeks-old. Body mass and blood pressure were periodically measured along the experiments. All animal group had their circadian rhythm of Tcore evaluated over three consecutive days (72 h) by telemetry using an implanted temperature sensor. The Tcore circadian rhythm was averaged in 1-h blocks and analyzed using the cosinor method. RESULTS: Sixteen-week-old SHR (SHR16) presented higher Tcore than WIS16 (from 06am to 06pm) and WKY16 (from 07am to 06pm). Both normotensive groups exhibited increases in Tcore during circadian rhythm with aging. The cosinor analysis showed no differences between strains and ages for the acrophase. An age effect on the SHR strain (SHR16 < SHR72) was observed regarding the amplitude. SHR16 had higher values regarding MESOR compared to WIS16 and WKY16. In addition, WIS72 and WKY72 showed higher values than WIS16 and WKY16, respectively. Finally, no differences were observed in the strength rhythm analysis. CONCLUSIONS: SHR presented impaired thermoregulatory control at only 16 weeks of age when showing a higher body temperature during the activity phase, while other circadian rhythm parameters showed no differences across aging. Therefore, in taking our results as a whole we can conclude that WIS and WKY are appropriate Wistar strains to be used as normotensive controls for SHR.

Chemical Characterization and Antimicrobial Activity Evaluation of Natural Oil Nanostructured Emulsions.

The aim of this work was to investigate the antimicrobial activity of nanostructured emulsions based on copaiba (Copaifera langsdorffii) resin-oil, copaiba essential oil, and bullfrog (Rana catesbeiana Shaw) oil against fungi and bacteria related to skin diseases. Firstly, the essential oil was extracted from copaiba resin-oil and these oils, along with bullfrog oil, were characterized by gas chromatography combined with mass spectrometry (GC-MS). Secondly, nanostructured emulsion systems were produced and characterized. The antimicrobial susceptibility assay was performed, followed by the Minimum Inhibitory Concentration (MIC) determination, the bioautography assay, and the antibiofilm determination. Strains of the genera Staphylococcus, Pseudomonas, and Candida were used. The CG-MS analysis was able to identify the components of copaiba resin-oil, copaiba essential oil, and bullfrog oil. The MIC assay in association with the bioautography revealed that some esters of palmitic and oleic acids, a-curcumene, a-himachalene, isothujol, and α-fenchene--probably inhibited some strains. The nanostructured emulsions based on copaiba resin-oil and essential oil improved the antimicrobial activity of the pure oils, especially against Staphylococcus and Candida, resistant to azoles. The bullfrog oil nanostructured emulsion showed a lower antimicrobial effect when compared to the copaiba samples. However, bullfrog oil-based nanostructured emulsion showed a significant antibiofilm activity (p < 0.05). Given the significant antimicrobial and antibiofilm activities of the evaluated oils, it may be concluded that nanostructured emulsions based on copaiba and bullfrog oils are promising candidates for the treatment of infections and also may be used to incorporate other antimicrobial drugs.

Influence of the freeze-drying process on the physicochemical and biological properties of pre-heated amphotericin B micellar systems.

The moderate heat treatment of amphotericin B (AmB) in its micellar form (M-AmB) results in superaggregates (H-AmB) that present a substantially lower toxicity and similar activity. The aim of this work was to evaluate the H-AmB behavior after a freeze-drying process. H-AmB and M-AmB micelles were evaluated before and after freeze-drying concerning their physicochemical and biological properties by spectrophotometry and activity/toxicity assay, respectively. Four concentrations of M-AmB and H-AmB were studied aiming to correlate their aggregation state and the respective biological behavior: 50 mg L(-1), 5 mg L(-1), 0.5 mg L(-1), and 0.05 mg L(-1). Then, potassium leakage and hemoglobin leakage from red blood cells were used to evaluate the acute and chronic toxicity, respectively. The efficacy of M-AmB and H-AmB formulations was assessed by potassium leakage from Candida albicans and by the broth microdilution method. After heating, in addition to an evident turbidity, a slight blueshift from 327 to 323 nm was also observed at the concentrations of 50 and 5 mg L(-1) for H-AmB. Additionally, an increase in the absorbance at 323 nm at the concentration of 0.5 mg L(-1) was detected. Concerning the toxicity, H-AmB caused significantly lower hemoglobin leakage than M-AmB. These results were observed for H-AmB before and after freeze-drying. However, there was no difference between H-AmB and M-AmB concerning their activity. Accordingly, the freeze-drying cycle did not show any influence on the behavior of heated formulations, highlighting the suitability of such a method to produce a new AmB product with a long shelf life and with both greater efficiency and less toxicity.

Attenuation of Ca2+ homeostasis, oxidative stress, and mitochondrial dysfunctions in diabetic rat heart: insulin therapy or aerobic exercise?

We tested the effects of swimming training and insulin therapy, either alone or in combination, on the intracellular calcium ([Ca(2+)]i) homeostasis, oxidative stress, and mitochondrial functions in diabetic rat hearts. Male Wistar rats were separated into control, diabetic, or diabetic plus insulin groups. Type 1 diabetes mellitus was induced by streptozotocin (STZ). Insulin-treated groups received 1 to 4 UI of insulin daily for 8 wk. Each group was divided into sedentary or exercised rats. Trained groups were submitted to swimming (90 min/day, 5 days/wk, 8 wk). [Ca(2+)]i transient in left ventricular myocytes (LVM), oxidative stress in LV tissue, and mitochondrial functions in the heart were assessed. Diabetes reduced the amplitude and prolonged the times to peak and to half decay of the [Ca(2+)]i transient in LVM, increased NADPH oxidase-4 (Nox-4) expression, decreased superoxide dismutase (SOD), and increased carbonyl protein contents in LV tissue. In isolated mitochondria, diabetes increased Ca(2+) uptake, susceptibility to permeability transition pore (MPTP) opening, uncoupling protein-2 (UCP-2) expression, and oxygen consumption but reduced H2O2 release. Swimming training corrected the time course of the [Ca(2+)]i transient, UCP-2 expression, and mitochondrial Ca(2+) uptake. Insulin replacement further normalized [Ca(2+)]i transient amplitude, Nox-4 expression, and carbonyl content. Alongside these benefits, the combination of both therapies restored the LV tissue SOD and mitochondrial O2 consumption, H2O2 release, and MPTP opening. In conclusion, the combination of swimming training with insulin replacement was more effective in attenuating intracellular Ca(2+) disruptions, oxidative stress, and mitochondrial dysfunctions in STZ-induced diabetic rat hearts.

A single intraoperative sub-Tenon's capsule triamcinolone acetonide injection for the treatment of post-cataract surgery inflammation.

PURPOSE: To compare a single intraoperative sub-Tenon's capsule triamcinolone acetonide injection with steroid drops in the treatment of ocular inflammation after cataract surgery. DESIGN: Randomized, double-masked controlled trial. PARTICIPANTS: A total of 100 patients were randomized prospectively into 2 groups: 50 patients treated with 1% prednisolone eyedrops (control group A) and 50 patients treated with sub-Tenon's capsule triamcinolone (treatment group B). METHODS: All patients underwent phacoemulsification and intraocular posterior lens implantation. After surgery, patients were randomized to receive either (group B) an intraoperative 40 mg triamcinolone acetonide sub-Tenon's capsule injection or (group A) 1% prednisolone acetate eyedrops, according to the following schedule: 1 drop 4 times daily (week 1), 3 times daily (week 2), 2 times daily (week 3), once daily (week 4). To mask the study, group B received vehicle drops administered on a similar schedule, and group A received an intraoperative sub-Tenon's capsule injection of a 1 ml balanced salt solution. MAIN OUTCOME MEASURES: The main outcome measures included inflammation (cell, flare, ciliary flush), intraocular pressure, and lack of response. RESULTS: Triamcinolone was shown to have anti-inflammatory efficacy clinically equivalent to conventional 1% prednisolone eyedrops in reducing intraocular inflammation, as measured by clinical methods. Triamcinolone was found to be as safe as the prednisolone in terms of adverse effects, changes in visual acuity, intraocular pressure, and biomicroscopic and ophthalmoscopic variables. On the third, seventh, fourteenth, and twenty-eighth postoperative days, a significantly lower intraocular pressure (P<0.01) was noted in the triamcinolone group than in the prednisolone group. CONCLUSIONS: A single intraoperative 40-mg triamcinolone acetonide sub-Tenon's capsule injection demonstrated a clinically equivalent therapeutic response and ocular tolerance compared with 1% prednisolone drops in controlling postoperative inflammation after uncomplicated cataract surgery and merits further investigation.

Recombinant alpha interferon in metastatic renal cell carcinoma. A cooperative phase II study.

A total of 61 evaluable patients with advanced renal cell carcinoma have been treated with 3 x 10(6) IU per square meter of body surface with recombinant alpha 2b interferon three times a week within a Cooperative Phase II Study. Toxic death for terminal renal failure occurred in 1 patient (1.63%), and toxicities greater than WHO grade 2 were present in 10 cases (16%). The overall response rate after six months of treatment was 13.1% (partial response 4, minor response 4). Two complete responses were obtained at nine and fifteen months (3.3%). Long-lasting stabilization of disease was 13.1 percent.

Production of epidermal growth factor in human prostatic cells cultured in vitro.

The Epidermal Growth Factor (EGF) plays an important role in the regulation of in vitro growth of prostate cells inducing a strong mitogenic effect. Nevertheless in our previous study we observed that the treatment of human hypertrophic prostate cell line U285 with exogenous EGF produces a restricted effect on the cellular growth rate. This phenomenon could be due to the capacity of the cells to produce EGF. In this study we aimed to verify this hypothesis by evaluating the presence of mRNA of EGF and EGF receptor (EGF-R) and of their translation products in U285 cells, before and after the treatment with suramin and exogenous EGF. Moreover we studied the effects exerted by these substances on the proliferative rate of the cells U285 after different treatment protocols. The presence in the cells of mRNA for EGF and EGF-R and of their translation products was demonstrated by means of reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemical methods respectively. The modification of growth rate induced by these drugs was studied by FRAME Cytotoxicity Test. The operative modalities adopted to carry out these growth assays tended to 1) focus the effects of suramin in relation to in vitro cellular growth phase; 2) verify the reversibility of its effects; 3) ascertain if it was possible to antagonize the action of suramin by adding exogenous EGF. The results obtained from the RT-PCR showed the presence, in the control cells and in the treated ones, of mRNA coding for EGF and EGF-R. The immunocytochemical analysis indicated that 20% of the control cells are EGF positive, and 83% are EGF-R positive, confirming the results obtained with RT-PCR. Moreover, these stainings showed that the treatment with EGF does not significantly modify the percentage of cells marked by the anti-EGF antibody, while treatments with suramin and suramin plus EGF double this percentage. None of the treatments modifies the percentage of EGF-R positive cells. The growth assays showed that the exposition to highest doses of suramin in the first 24 h of cultures causes a decrease (p < 0.05) of the cellular proliferation during the following 48 h and 72 h and that these effects are irreversible. Moreover, a contemporaneous exposition of the cells to EGF and suramin at seeding strengthens the cytotoxic action of the last drug. To sum up, the demonstration of the presence in the U285 cells of mRNA coding for EGF and EGF-R and of the corresponding proteins, confirms the hypothesis that these cells can produce EGF. Moreover, the cytotoxicity experiments allowed a focusing of the role of the endogenous EGF in the regulation of the U285 cells proliferation and confirmed the importance of biological events that take place in U285 cells during the first 24 h of culture.

Effects of DHT and EGF on human hyperplastic prostate cells cultured in vitro: growth, morphology and phenotype characterisation.

This work studies the effects of dihydrotestosterone (DHT) and epidermal growth factor (EGF) on the growth, morphology and phenotype characterisation of the U285 line obtained from human prostate hyperplastic tissue. Modifications of growth rate induced by these two substances have been evaluated by means of the neutral red assay formulated by Borenfreund and Puerner (1985) as well as by means of Kenacid blue assay described by Knox et al. (1986), culturing cells for 24, 48 and 72 hr with scalar doses of DHT (0.5, 1, 2, 5, 10 microM) and EGF (5, 10, 20, 100 ng/ml). An optical microscope connected to a computer aided system and a scanning electron microscope were used to monitor morphological changes induced by DHT and EGF. The immunophenotype characterisation of the treated and control cells was carried out by using a monoclonal antibody panel. Our results show that the expression of anti-cytokeratin 5+6+18, anti-cytokeratin 8+18+19 and anti-proline-4-hydroxylase antibodies varied in relation to the type of treatment undergone by the cells. Moreover, exogenous DHT does provoke a flattening of the U285 cells without modifying their rate of growth, while EGF both shortens the lag phase reactivating the quiescent cells and regulates the subsequent log growth phase, thus causing no cellular overgrowth.

[Persistent sciatic artery aneurysm. Case report]. / L'aneurisma dell'arteria "ischiatica" persistente. A proposito di un caso.

Persistent sciatic artery is a rare vascular anomaly particularly interesting for possible complications as buttock aneurysm and ischemic or embolic. Authors describes a case of buttock aneurysm of persistent sciatic artery personally observed analyse its anatomic and clinical features, and lay stress on the importance of angiography as essential examination in order to an exact surgical treatment.

[Aneurysms of the internal carotid artery. Apropos of a case]. / Aneurismi dell'arteria carotide interna. A proposito di un caso.

Aetiopathogenetic mechanisms and clinical aspects of carotid artery aneurysms are examined and personal experience of arterial reconstruction in one case reported.

Corneal absorption of a new riboflavin-nanostructured system for transepithelial collagen cross-linking.

Corneal collagen cross-linking (CXL) has been described as a promising therapy for keratoconus. According to standard CXL protocol, epithelium should be debrided before treatment to allow penetration of riboflavin into the corneal stroma. However, removal of the epithelium can increase procedure risks. In this study we aim to evaluate stromal penetration of a biocompatible riboflavin-based nanoemulsion system (riboflavin-5-phosphate and riboflavin-base) in rabbit corneas with intact epithelium. Two riboflavin nanoemulsions were developed. Transmittance and absorption coefficient were measured on corneas with intact epithelia after 30, 60, 120, 180, and 240 minutes following exposure to either the nanoemulsions or standard 0.1% or 1% riboflavin-dextran solutions. For the nanoemulsions, the epithelium was removed after measurements to assure that the riboflavin had passed through the hydrophobic epithelium and retained within the stroma. Results were compared to de-epithelialized corneas exposed to 0.1% riboflavin solution and to the same riboflavin nanoemulsions for 30 minutes (standard protocol). Mean transmittance and absorption measured in epithelialized corneas receiving the standard 0.1% riboflavin solution did not reach the levels found on the debrided corneas using the standard technique. Neither increasing the time of exposure nor the concentration of the riboflavin solution from 0.1% to 1% improved riboflavin penetration through the epithelium. When using riboflavin-5-phosphate nanoemulsion for 240 minutes, we found no difference between the mean absorption coefficients to the standard cross-linking protocol (p = 0.54). Riboflavin nanoemulsion was able to penetrate the corneal epithelium, achieving, after 240 minutes, greater stromal concentration when compared to debrided corneas with the standard protocol (p = 0.002). The riboflavin-5-phosphate nanoemulsion diffused better into the stroma than the riboflavin-base nanoemulsion.

Amphotericin B microemulsion reduces toxicity and maintains the efficacy as an antifungal product.

Amphotericin B remains the drug of choice for the treatment of most of the systemic fungal infections in immunodeficient patients. Because of the high incidence of adverse drug reactions the clinical use of Amphotericin B is rather limited. To reduce its toxicity new drug delivery systems has been suggested. Nevertheless, these carriers present several technological drawbacks that impair the development of a marketable product. The aim of this work was to develop an Amphotericin B microemulsion in order to increase its efficacy and decrease its toxicity compared to Fungizon, the widely know inexpensive micellar system of Amphotericin B. Amphotericin B loaded microemulsion showed an average size close to 300 nm by photon correlation spectroscopy. In the UV spectrum, the observation of the monomeric peak at 405 nm, which was independent of the sample dilution, revealed that the Amphotericin B molecules were strongly and individually bound to the microemulsion droplets. The new microemulsion formulation had the same efficacy than Fungizon against C. albicans. Concerning toxicity, Amphotericin B loaded microemulsion showed lower toxicity against human red blood cells compared to the commercial product. Taken together, these results suggested that microemulsion is an eligible drug carrier for Amphotericin B or other water insoluble molecules, and it has potential applications to targeting fungal cells. Additionally, a novel formulation of Amphotericin B-loaded microemulsion was prepared by a straightforward and fast procedure.

Subconjunctival delivery of antibiotics in a controlled-release system: a novel anti-infective prophylaxis approach for cataract surgery.

OBJECTIVE: To compare the efficacy of subconjunctival injection of a combination of triamcinolone and ciprofloxacin hydrochloride, 2 mg/0.1 mL, in a controlled-release system (DuoCat) with that of ciprofloxacin hydrochloride, 0.3%, eyedrops for infection prophylaxis. METHODS: Rabbit eyes were injected subconjunctivally with a combination of triamcinolone and ciprofloxacin hydrochloride, 2 mg/0.1 mL, or ciprofloxacin hydrochloride, 2 mg/0.1 mL, alone. The aqueous and vitreous humor pharmacokinetic profiles were compared with those of a single drop of ciprofloxacin hydrochloride, 0.3%, 6 times daily. In 45 rabbits, Staphylococcus aureus was injected into the anterior chamber: 15 randomly received 1 drop of ciprofloxacin hydrochloride, 0.3%, every 4 hours during 24 hours; 15 received drops of balanced salt solution; and 15 received a combination of triamcinolone and ciprofloxacin hydrochloride, 2 mg/0.1 mL. After 24 hours, endophthalmitis scores were recorded, aqueous and vitreous humors underwent culture, and histologic analysis was performed. RESULTS: The combined triamcinolone and ciprofloxacin treatment allowed higher intraocular levels of ciprofloxacin. The median endophthalmitis clinical scores for the combination of triamcinolone and ciprofloxacin and ciprofloxacin-only eyedrop groups were equivalent (P = .42) and were significantly lower than those of the balanced salt solution group (P < .001). The culture was negative for S aureus in the combined triamcinolone and ciprofloxacin and ciprofloxacin eyedrop regimens. No adverse effects were observed with either route. CONCLUSIONS: Ciprofloxacin eyedrops and combined triamcinolone and ciprofloxacin were equally tolerated and efficacious. The combined triamcinolone and ciprofloxacin treatment may eliminate noncompliance issues and may prove to be a valuable clinical tool for surgical prophylaxis. CLINICAL RELEVANCE: The combined triamcinolone and ciprofloxacin treatment may be a new useful strategy for surgical prophylaxis.

Controlled transscleral drug delivery formulations to the eye: establishing new concepts and paradigms in ocular anti-inflammatory therapeutics and antibacterial prophylaxis.

IMPORTANCE OF THE FIELD: The use of topical agents poses unique and challenging hurdles for drug delivery. Topical steroids effectively control ocular inflammation, but are associated with the well-recognized dilemma of patient compliance. Although administration of topical antimicrobials as prophylaxis is acceptable among ophthalmologists, this common practice has no sound evidence base. Developing a new antimicrobial agent or delivery strategy with enhanced penetration by considering the anatomical and physiological constraints exerted by the barriers of the eye is not a commonly perceived strategy. Exploiting the permeability of the sclera, subconjunctival routes may offer a promising alternative for enhanced drug delivery and tissue targeting. AREA COVERED IN THIS REVIEW: Ocular drug delivery strategies were reviewed for ocular inflammation and infections clinically adopted for newer class of antimicrobials, which use a multipronged approach to limit risks of endophthalmitis. WHAT THE READER WILL GAIN: The analysis substantiates a new transscleral drug delivery therapeutic approach for cataract surgery. TAKE HOME MESSAGE: A new anti-inflammatory and anti-infective paradigm that frees the patient from the nuisance of topical therapeutics is introduced, opening a large investigative avenue for future improved therapies.