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.

Low-dose thioperamide injected into the cerebellar vermis of mice immediately after exposure to the elevated plus-maze impairs their avoidance behavior on re-exposure to the apparatus

The present study investigated the effect of thioperamide (THIO), an H3 histaminergic receptor antagonist, microinjected into the cerebellar vermis on emotional memory consolidation in male Swiss albino mice re-exposed to the elevated plus-maze (EPM). We implanted a guide cannula into the cerebellar vermis using stereotactic surgery. On the third day after surgery, we performed behavioral tests for two consecutive days. On the first day (exposure), the mice (n=10/group) were exposed to the EPM and received THIO (0.06, 0.3, or 1.5 ng/0.1 µL) immediately after the end of the session. Twenty-four hours later, the mice were re-exposed to the EPM under the same experimental conditions, but without drug injection. A reduction in the exploration of the open arms upon re-exposure to the EPM (percentage of number of entries and time spent in open arms) compared with the initial exposure was used as an indicator of learning and memory. One-way analysis of variance (ANOVA) followed by the Duncan post hoc test was used to analyze the data. Upon re-exposure, exploratory activity in the open arms was reduced in the control group, and with the two highest THIO doses: 0.3 and 1.5 ng/0.1 µL. No reduction was seen with the lowest THIO dose (0.06 ng/0.1 µL), indicating inhibition of the consolidation of emotional memory. None of the doses interfered with the animals' locomotor activity. We conclude that THIO at the lowest dose (0.06 ng/0.1 µL) microinjected into the cerebellum impaired emotional memory consolidation in mice.

Liver cancer stem cells are selectively enriched by low-dose cisplatin

Accumulating evidence has indicated the importance of cancer stem cells in carcinogenesis. The goal of the present study was to determine the effect of low-dose cisplatin on enriched liver cancer stem cells (LCSCs). Human hepatoblastoma HepG2 cells were treated with concentrations of cisplatin ranging from 1 to 5 μg/mL. Cell survival and proliferation were evaluated using a tetrazolium dye (MTT) assay. LCSCs were identified using specific markers, namely aldehyde dehydrogenase-1 (ALDH1) and CD133. The percentage of ALDH1+ or CD133+ cells was examined by flow cytometric analysis. The expression of ALDH1 and/or CD133 in HepG2 cells was determined by immunocytochemical analysis. Low-dose cisplatin treatment significantly decreased cell survival in HepG2 cells after 24 or 72 h. However, the percentage of LCSCs in the surviving cells was greatly increased. The percentage of ALDH1+ or CD133+ cells was increased in a time- and dose-dependent manner after treatment with 1-4 μg/mL cisplatin, whereas 5 μg/mL cisplatin exposure slightly reduced the number of positive cells. These findings indicate that low-dose cisplatin treatment may efficiently enrich the LCSC population in HepG2 cells.

Low and high-dose intradermal infection with Leishmania majorand Leishmania amazonensis in C57BL/6 mice

A model of skin infection with Leishmania amazonensiswith low doses of parasites is compared to infection with high doses of L. amazonensis and low and high doses of Leishmania major. C57BL/6 mice were infected with 10³ or 10(6) parasites in the ear and the outcome of infection was assessed. The appearance of lesions in mice infected with 10³ parasites was delayed compared to mice infected with 10(6) Leishmania and parasites were detectable at the infection site before lesions became apparent. Mice infected with L. amazonensisdisplayed persistent lesions, whereas infection with L. major spontaneously healed in all groups, although lymphocytes persisted at the site of infection after healing. Macrophages persisted only in L. amazonensis-infected mice. High-dose L. amazonensis-infected mice produced lower levels of IFN-γ and TNF than mice infected with L. major. No correlation between the persistence of parasites and IL-10 levels and the production of nitric oxide or urea by macrophages was found. We conclude that infection with low doses of L. amazonensisin the dermis changes the course of infection by delaying the appearance of lesions. However, low-dose infection does not change the outcomes of susceptibility and cytokine production described for subcutaneous infection with high numbers of parasites.

Active immunization against hepatitis B virus (HBV) with low-doses of plasma-derived vaccine by intradermal route

Schedule for vaccination against HBV infection has usually been based on three separate injections of 20 meg of the vaccine by intramuscular route. One of the main shortcomings to its use in large scale programs has been its high cost. Ninety out of 300 health workers were submitted to three injections of 2 meg of plasma-derived vaccine (PDV) by intradermal (ID) route on days 0, 30, and 180. Anti-HBs was detected in 74 (82.2%) after the second dose and in 80 (88.9%) after the third dose, a non-significant difference. However, levels above 10 times the cut-off were observed in 29 (32.2%) and 77 (85.5%), respectively (p < 0.001). The results showed that a low-dose schedule is effective when used in health workers and should be tried with other risk groups.

Intradermal vaccination of adults with three low doses (2 µg) of recombinant hepatitis B vaccine. I. Seroconversion rate and adverse effects

A total of 250 dentists (53.6% men and 46.4% women), with a mean age of 35.1 ± 9.8 years, were submitted to serological tests for the diagnosis of hepatitis B (HB) - HBsAg, anti-HBs, anti-HBc, HBeAg, and anti-HBe - using a radioimmunoassay. One or more of these markers were detected in 78 individuals (31.2%) who were excluded from the group to be vaccinated. Of the 172 HB-susceptible individuals, 135 (78.5%) responded to the call and were intradermally injected with three 2 µg doses of the Belgian HB recombinant vaccine, applied at an interval of one month between the 1st and 2nd dose and of five months between the 2nd and 3rd dose. A new determination of HB markers carried out 50 days after the 3rd dose showed that 110 (81.5%) individuals had become anti-HBs positive (65.5% good responders and 34.5% poor responders). Mean serum anti-HBs titer of these 110 dentists was 42.4 U S/N, similar in both sexes. The adverse effects analyzed in 106 dentists were: (a) local: pain (12.3%), burning sensation (14.1%), pruritus (25.5%), erythema (28.3%), local heat (18.9%), and a hypochromic spot (32.1%); (b) systemic (4.7%): discomfort in two patients, and fever, anorexia, and asthenia in one patient each. Intradermal administration of a fourth 2 µg vaccine dose to 39 dentists (poor or non-responders) increased the total number of anti-HBs-positive individuals from 110 (81.5%) to 114 (84.4%), with the number of good responders increasing from 72 (65.5%) to 85 (74.6%). We conclude that the Belgian recombinant vaccine applied in the scheme used here induces a high rate of seroconversion and causes only mild and transitory adverse effects.

Possible benefit of consolidation therapy with high-dose cytarabine on overall survival of adults with non-promyelocytic acute myeloid leukemia

In adults with non-promyelocytic acute myeloid leukemia (AML), high-dose cytarabine consolidation therapy has been shown to influence survival in selected patients, although the appropriate doses and schemes have not been defined. We evaluated survival after calculating the actual dose of cytarabine that patients received for consolidation therapy and divided them into 3 groups according to dose. We conducted a single-center, retrospective study involving 311 non-promyelocytic AML patients with a median age of 36 years (16-79 years) who received curative treatment between 1978 and 2007. The 131 patients who received cytarabine consolidation were assigned to study groups by their cytarabine dose protocol. Group 1 (n=69) received <1.5 g/m2 every 12 h on 3 alternate days for up to 4 cycles. The remaining patients received high-dose cytarabine (≥1.5 g/m2 every 12 h on 3 alternate days for up to 4 cycles). The actual dose received during the entire consolidation period in these patients was calculated, allowing us to divide these patients into 2 additional groups. Group 2 (n=27) received an intermediate-high-dose (<27 g/m2), and group 3 (n=35) received a very-high-dose (≥27 g/m2). Among the 311 patients receiving curative treatment, the 5-year survival rate was 20.2% (63 patients). The cytarabine consolidation dose was an independent determinant of survival in multivariate analysis; age, karyotype, induction protocol, French-American-British classification, and de novo leukemia were not. Comparisons showed that the risk of death was higher in the intermediate-high-dose group 2 (hazard ratio [HR]=4.51; 95% confidence interval [CI]: 1.81-11.21) and the low-dose group 1 (HR=4.43; 95% CI: 1.97-9.96) than in the very-high-dose group 3, with no significant difference between those two groups. Our findings indicated that very-high-dose cytarabine during consolidation in adults with non-promyelocytic AML may improve survival.

Schistosoma mansoni: inflammatory foci around larvae in the peritoneal cavity of naive mice is radiosensitive

Innate attack to Schistosoma mansoni cercariae was evaluated in irradiated mice. It was observed that 70% of the larvae from mice sacrificed one day after whole body irradiation with 400 or 800 rads were surrounded by cluster reactivities, without difference from controls. Differences were apparent on day 5 after irradiation with sub lethal (400 rads) or lethal doses (800 rads) suggesting that innate defence to infection take at least 5 days to be affected by low dose whole-body radiation.

FACTORS ASSOCIATED TO ADHERENCE TO DIFFERENT TREATMENT SCHEMES WITH MEGLUMINE ANTIMONIATE IN A CLINICAL TRIAL FOR CUTANEOUS LEISHMANIASIS

The favorable outcome of the treatment of a disease is influenced by the adherence to therapy. Our objective was to assess factors associated with adherence to treatment of patients included in a clinical trial of equivalence between the standard and alternative treatment schemes with meglumine antimoniate (MA) in the treatment of cutaneous leishmaniasis (CL), in the state of Rio de Janeiro. Between 2008 and 2011, 57 patients with CL were interviewed using a questionnaire to collect socioeconomic data. The following methods were used for adherence monitoring: counting of vial surplus, monitoring card, Morisky test and modified Morisky test (without the question regarding the schedule); we observed 82.1% (vial return), 86.0% (monitoring card), 66.7% (Morisky test) and 86.0% (modified Morisky test) adherence. There was a strong correlation between the method of vial counting and the monitoring card and modified Morisky test. A significant association was observed between greater adherence to treatment and low dose of MA, as well as with a lower number of people sleeping in the same room. We recommend the use of the modified Morisky test to assess adherence to treatment of CL with MA, because it is a simple method and with a good performance, when compared to other methods.

RESOLUTION OF CUTANEOUS LEISHMANIASIS AFTER ACUTE ECZEMA DUE TO INTRALESIONAL MEGLUMINE ANTIMONIATE

We report a case of a 42 year-old female, who came to a leishmaniasis reference center in Rio de Janeiro, Brazil, presenting a cutaneous leishmaniasis lesion in the right forearm. Treatment with low-dose intramuscular meglumine antimoniate (MA) (5 mg Sb5+/kg/day) was initiated, with improvement after 28 days, although with the development of generalized eczema. After 87 days, the lesion worsened. Patient refused treatment with amphotericin B. MA was then infiltrated in the lesion, in two sessions, resulting in local eczema, with bullae formation; however, twenty days after, both the ulcer and eczema receded. Intralesional administration of MA should be used carefully when previous cutaneous hypersensitivity is detected.

An alternative antimonial schedule to be used in cutaneous leishmaniasis when high doses of antimony are undesirable

Despite more than half a century of use in leishmaniasis, antimony therapy still presents serious problems concerning dosage and toxicity. Low and high doses have been shown to be equally effective. In this paper, the feasibility of injecting one ampoule of meglumine antimoniate intramuscularly every other day until clinical cure is demonstrated, while studying a series of 40 cutaneous leishmaniasis cases. Total dose used varied from 1,822.5 to 12,150mg of pentavalent antimony and total time of treatment varied from 3 to 10 weeks, with 86% efficacy. Thirty-six out of the 40 patients are still on follow-up with a mean time of 10.7 ± 7 months and a median of 9 months. No relapse or mucosal lesions have been noted so far. The schedule showed good tolerance and easy application and its efficacy was comparable to the officially recommended WHO schedule. Therefore, such a schedule represents a valuable alternative for the cases with high toxicicity to antimony or daily injections are an obstacle to the treatment.

Clinical variables associated with leprosy reactions and persistence of physical impairment

Introduction Leprosy is a chronic disease that affects skin and peripheral nerves. Disease complications include reactional episodes and physical impairment. One World Health Organization (WHO) goal of leprosy programs is to decrease the number of grade 2 impairment diagnoses by 2015. This study aims to evaluate clinical factors associated with the occurrence of leprosy reactions and physical impairment in leprosy patients. Methods We conducted a retrospective study of data from medical records of patients followed in two important centers for the treatment of leprosy in Aracaju, Sergipe, Brazil, from 2005 to 2011. We used the chi-square test to analyze associations between the following categorical variables: gender, age, operational classification, clinical forms, leprosy reactions, corticosteroid treatment, and physical impairment at the diagnosis and after cure. Clinical variables associated with multibacillary leprosy and/or reactional episodes and the presence of any grade of physical impairment after cure were evaluated using the logistic regression model. Results We found that men were more affected by multibacillary forms, reactional episodes, and grade 2 physical impairment at diagnosis. Leprosy reactions were detected in a total of 40% of patients and all were treated with corticosteroids. However, physical impairment was observed in 29.8% of the patients analyzed at the end of the treatment and our multivariate analysis associated a low dose and short period of corticosteroid treatment with persistence of physical impairments. Conclusions Physical impairment should receive an increased attention before and after treatment, and adequate treatment should be emphasized.

Diffuse panbronchiolitis: an underdiagnosed disease? Study of 4 cases in Brazil

BACKGROUND: Diffuse panbronchiolitis is a clinical pathologic condition characterized by chronic inflammation of respiratory bronchioles, with clinical features that position it as a differential diagnosis among the sinopulmonary syndromes. METHODS AND RESULTS: We present 4 cases (1 Black, 2 Japanese descendants, and 1 Japanese), living in Brazil, in which the diagnosis was made by the clinical and radiological features and confirmed by transbronchial biopsy. The clinical findings included chronic sinusitis, productive cough, rhonchi, and wheezes. The pulmonary function tests showed an obstructive pattern. High resolution computerized tomography showed a diffuse nodular pattern, airway ectasia, and airway wall thickening. The biopsy showed interstitial accumulation of foam cells and lymphoid cells in the walls of respiratory bronchioles: 2 of our cases had bronchus-associated lymphoid tissue hyperplasia. We searched for the HLA Bw54 in all of our patients, but only 1 was positive. A low dose macrolide treatment was introduced, resulting in with clinical and functional improvement. A score that rated the extent of nodules, airway ectasia, mucus plugging, and airway wall thickening was applied on pre- and post-treatment High resolution computerized tomography results, revealing an improvement in tomographic pattern related to that observed in the pulmonary function tests. CONCLUSION: We conclude that diffuse panbronchiolitis is a systemic disease that is not exclusive to the Asian population, whose clinical and radiological features should be better known by occidental pulmonary physicians.

Short and long-term effects of azadirachtin A on development and egg production of Rhodnius prolixus

Azadirachtin A was given through a blood meal to 4th-instar larvae and to adult females of Rhodnius prolixus. Development (ecdysis) and egg production were inhibited in a dose-dependent manner. Long-term experiments with subsequent four feedings on azadirachtin-free blood were performed with 4th-instar larvae and with adult females. Only in the low-dose azadirachtin larval groups (0.01 and 0.1 microng/ml of blood), development was partially restored; after a single 1.0 microng/ml treatment about 50% of the treated larvae were still alive 120 days later without any adult emergence. Similarly fed females had a dose-dependent lower survival and egg deposition rate. The results are discussed in relation to the mode of azadirachtin A action.

Optimization of a mouse immunization protocol with Paracoccidioides brasiliensis antigens

The objectives of the present study were to optimize the protocol of mouse immunization with Paracoccidioides brasiliensis antigens (Rifkind's protocol) and to test the modulation effect of cyclophosphamide (Cy) on the delayed hypersensitivity response (DHR) of immunized animals. Experiments were carried out using one to four immunizing doses of either crude particulate P. brasiliensis antigen or yeast-cell antigen, followed by DHR test four or seven days after the last immunizing dose. The data demonstrated that an immunizing dose already elicited response; higher DHR indices were obtained with two or three immunizing doses; there were no differences between DHR indices of animals challenged four or seven days after the last dose. Overall the inoculation of two or three doses of the yeast-cell antigen, which is easier to prepare, and DHR test at day 4 simplify the original Rifkind's immunization protocol and shorten the duration of the experiments. The modulation effect of Cy on DHR was assayed with administration of 2.5, 20 and 100 mg/kg weight at seven day intervals starting from day 4 prior to the first immunizing dose. Only the treatment with 2.5 mg Cy increased the DHR indices. Treatment with 100 mg Cy inhibited the DHR, whereas 20 mg Cy did not affect the DHR indices. Results suggest an immunostimulating effect of low dose of Cy on the DHR of mice immunized with P. brasiliensis antigens.