Telomerase activity in well-differentiated papillary thyroid carcinoma corrrelates with advanced clinical stage of the disease

Clinical relevance and stage correlation of telomerase activity in well-differentiated papillary thyroid carcinoma (WD PTC) has not been well determined, as its reported activity could be due to the analysis of tumors with lymphocytic infiltrates or aggressive variants of papillary carcinomas. We conducted a prospective study of telomerase activity in WD PTC without inflammatory infiltrates and correlated it with clinical stage. Fifty WD PTCs were analyzed for telomerase activity by PCR-based TRAP (telomeric repeat amplification protocol) assay. Results were correlated with stage and other clinicopathologic variables. Twenty-one (42%) WD PTCs demonstrated telomerase activity. The enzyme was detected more frequently in stage III/IVa WD PTCs (p=0.02) and in tumors with extra thyroidal extension (p=0.04). The risk of presenting advanced disease (stage III/IVa) and extrathyridal growth was significantly increased in telomerase-positive tumors (p=0.01; odds ratio [OR] 4.4 [95%Cl 1.3–14.7]) and (p=0.04; OR 3.6 [95%Cl 1.1–11.7]), respectively. Also, a correlation was found between telomerase activity and age. There was no correlation of telomerase activity with gender, histologic variant, tumor size, or cervical lymph node metastasis. Telomerase activity was observed in 42% of WD PTC and was detected more frequently in AJCC TNM stage III/IVa cases. This finding suggests that telomerase deregulation could be involved in tumor progression.     

MIF is essential to the establishment of house dust mite-induced airway inflammation and tissue remodeling in mice

Macrophage migration inhibitory factor (MIF) is present in high amounts in the BALF and serum of asthmatic patients, contributing to the pathogenesis of experimental asthma induced by OVA in mice. Whether MIF contributes to the physiopathology on a more complex and relevant asthma model has not been characterized. Mif-deficient (Mif^−/−) or WT mice treated with anti-MIF antibody were challenged multiple times using house dust mite (HDM) extract by the intranasal route. HDM-challenged Mif^−/− mice presented decreased airway hyperresponsiveness, lung infiltration of eosinophils, mucus hypersecretion, and subepithelial fibrosis compared to HDM-challenged WT mice. Amounts of IL-4, IL-5, and IL-13 were decreased in the lungs of Mif^−/− mice upon HDM challenges, but the increase of CCL11 was preserved, compared to HDM-challenged WT mice. We also observed increased numbers of group 2 innate lymphoid cells and Th2 cells in the BALF and mediastinal LNs (mLN)-induced challenged by HDM of WT mice, but not in HDM-challenged Mif^−/− mice. Anti-MIF treatment abrogated the airway infiltration of eosinophils, mucus hypersecretion, and subepithelial fibrosis in the lungs of HDM-challenged mice. In conclusion, MIF ablation prevents the pathologic hallmarks of asthma in HDM-challenged mice, reinforcing the promising target of MIF for asthma therapy.     

Argininic acid alters markers of cellular oxidative damage in vitro: Protective role of antioxidants

We, herein, investigated the in vitro effects of argininic acid on thiobarbituric acid-reactive substances (TBA-RS), total sulfhydryl content and on the activities of antioxidant enzymes such as catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in the blood, kidney and liver of 60-day-old rats. We also verified the influence of the antioxidants (each at 1.0 mM) trolox and ascorbic acid, as well as of N^G-nitro-l-arginine methyl ester (L-NAME) at 1.0 mM, a nitric oxide synthase inhibitor, on the effects elicited by argininic acid on the parameters tested. The liver, renal cortex and renal medulla were homogenized in 10 vol (1:10w/v) of 20 mM sodium phosphate buffer, pH 7.4, containing 140 mM KCl; and erythrocytes and plasma were prepared from whole blood samples obtained from rats. For in vitro experiments, the samples were pre-incubated for 1 h at 37 °C in the presence of argininic acid at final concentrations of 0.1, 1.0 and 5.0 μM. Control experiments were performed without the addition of argininic acid. Results showed that argininic acid (5.0 μM) enhanced CAT and SOD activities and decreased GSH-Px activity in the erythrocytes, increased CAT and decreased GSH-Px activities in the renal cortex and decreased CAT and SOD activities in the renal medulla of 60-day-old rats, as compared to the control group. Antioxidants and/or L-NAME prevented most of the alterations caused by argininic acid on the oxidative stress parameters evaluated. Data suggest that argininic acid alters antioxidant defenses in the blood and kidney of rats; however, in the presence of antioxidants and L-NAME, most of these alterations in oxidative stress were prevented. These findings suggest that oxidative stress may be make an important contribution to the damage caused by argininic acid in hyperargininemic patients and that treatment with antioxidants may be beneficial in this pathology.     

Pituitary deficiency after aneurysmal subarachnoid hemorrhage

OBJECTIVE:

Aneurysmal subarachnoid hemorrhage puts patients at high risk for the development of pituitary insufficiency. We evaluated the incidence of pituitary dysfunction in these patients and its correlation with clinical outcome.

METHODS:

Pituitary function was tested in 66 consecutive patients in the first 15 days after aneurysmal subarachnoid hemorrhage. The following were measured in all patients: thyroid-stimulating hormone, free thyroxine, triiodothyronine, luteinizing hormone, follicle-stimulating hormone, total testosterone (in males), estradiol (in females), prolactin, serum cortisol, plasma adrenocorticotropic hormone, growth hormone and insulin growth factor.

RESULTS:

The endocrine assessment was made at a mean of 7.4 days (standard deviation ±6.6) after subarachnoid hemorrhage. Forty-four (66.7%) female and 22 (33.3%) male patients were evaluated. Thirty-nine patients (59.1%) had some type of pituitary dysfunction. Follicle-stimulating hormone/luteinizing hormone deficiency was the most frequent disorder (34.8%), followed by growth hormone/insulin growth factor (28.7%), adrenocorticotropic hormone (18.1%) and thyroid-stimulating hormone (9%). Seventeen (25.7%) patients showed deficiencies in more than one axis. A greater incidence of hormone deficiency was observed in patients with a Glasgow Coma Scale score ≤13 (t test, p = 0.008), Hunt-Hess grade ≥4 (t test, p<0.001), or Fisher grade 4 (t test, p = 0.039). Hormone deficiency was not significantly associated (p>0.05) with increased hospitalization or clinical outcome.

CONCLUSION:

Pituitary dysfunction was identified in a substantial portion of patients with previous aneurysmal subarachnoid hemorrhage, but no association was found between this dysfunction and poor clinical outcome.     

Development of Doxycycline MIC and Disk Diffusion Interpretive Breakpoints and Revision of Tetracycline Breakpoints for Streptococcus pneumoniae

A study was performed to derive susceptibility testing interpretive breakpoints for doxycycline with Streptococcus pneumoniae and to reassess breakpoints for tetracycline using the requirements defined in Clinical and Laboratory Standards Institute (CLSI) document M23-A3. Tetracycline and doxycycline MICs and disk diffusion zone sizes were determined on 189 isolates selected from the 2009-2010 CDC Active Bacterial Core surveillance strain collection according to the testing methods described in CLSI documents M07-A8 and M02-A10. Tetracycline and doxycycline MICs and zones were compared to each other directly, and the reproducibility of MICs and zone diameters for both drugs was determined. Scattergrams of tetracycline MICs versus corresponding zone diameters and doxycycline MICs versus zones were prepared, and analysis indicated that the present CLSI tetracycline MIC and disk breakpoints did not fit the susceptibility data for doxycycline. Doxycycline was 1 to 3 dilutions more potent than tetracycline, especially in strains harboring the tetM resistance determinant. tetM was detected in ≥90% of isolates having tetracycline MICs of ≥4 μg/ml and in ≥90% with doxycycline MICs of ≥1. Limited pharmacokinetic/pharmacodynamic (PK/PD) data coupled with application of the error-rate bounded method of analysis suggested doxycycline-susceptible breakpoints of either ≤0.25 μg/ml or ≤0.5 μg/ml, with intermediate and resistant breakpoints 1 and 2 dilutions higher, respectively. The disk diffusion zone diameter correlates were susceptible at ≥28 mm, intermediate at 25 to 27 mm, and resistant at ≤24 mm. Revised lower tetracycline MIC breakpoints were suggested as susceptible at ≤1 μg/ml, intermediate at 2 μg/ml, and resistant at ≥4 μg/ml. Suggested tetracycline disk diffusion zones were identical to those of doxycycline.     

Hypoxia and fatigue-induced modification of function and proteins in intact and skinned murine diaphragm muscle

Fatigue studies of isolated, intact muscles typically utilize solutions saturated with O2. However, under in vivo fatiguing conditions, less oxygen is delivered to the muscles and they actually experience hypoxia. No studies to date have correlated the effects of acute hypoxia on the isometric contractile properties of intact muscles, skinned fibers isolated from the same muscles, and the cellular content of specific muscle proteins. Therefore, we have studied the effects of in vitro acute hypoxia on the fatigability of intact diaphragm muscle strips and on the isometric contractile properties of single Triton-skinned fibers isolated from control and hypoxic diaphragm muscles. We found that hypoxia and fatiguing stimulation per se affect the tetanic force of intact muscle strips without exhibiting any significant deleterious effects on the calcium-activated force of skinned muscle fibers dissected from the intact muscles. In contrast, fatiguing stimulation under hypoxic conditions decreased both the tetanic force of muscle strips and the calcium-activated force of skinned muscle fibers. Gel electrophoresis of muscles subjected to hypoxia and hypoxic-fatigue revealed that there is a significant reduction in three protein bands when compared to control muscles. Protein modification may be the underlying mechanism of muscle fatigue under physiologic conditions.     

Interaction of Leishmania mexicana promastigotes with the plasminogen–plasmin system

The binding of human plasminogen and plasmin to the promastigote form of Leishmania mexicana was investigated. L. mexicana was capable to bind both molecules, the binding being inhibited by -aminocaproic acid. Scatchard plot analysis revealed a dissociation constant (K_d) value of 2.4±0.8 μM and 0.9±0.1×10⁴ binding sites per cell for plasminogen and a K_d value of 1.2±0.4 μM and 1.6±0.2×10⁵ binding sites per cell for plasmin. C-terminal lysine residues are involved in plasminogen binding to cells, since carboxypeptidase B treatment reduced this binding by 34%. Ligand blotting analysis showed a group of proteins, with molecular masses between 105 and 115 kDa, capable to interact with plasminogen. Zymogram analysis showed that the protease activity acquired by L. mexicana, due to the interaction with either plasminogen or plasmin, comprises an important fraction of the total protease activity at pH 7.7. Plasminogen activation by tissue-type plasminogen activator (t-PA) was enhanced by the presence of L. mexicana promastigotes. These results raise the question whether the interaction of L. mexicana with components of the fibrinolytic system is involved in the virulence of the parasite.     

Changes in β-giardin sequence of <Emphasis Type="Italic">Giardia intestinalis</Emphasis> sensitive and resistant to albendazole strains

Giardia intestinalis can develop resistance to albendazole, although the molecular mechanism is not understood. The aim of this study was to investigate the differences and permanent mutation in the β-giardin gene of G. intestinalis strains: sensitive, resistant, or recovered-resistance to albendazole. The β-giardin gene was amplified by nested polymerase chain reaction. The IC₅₀ values varied from 0.29 to 0.38 μg/mL for strains sensitive to albendazole. For resistant strains, the IC₅₀ range was 1.31–2.12 μg/mL. Recovered-sensitivity albendazole strains’ IC₅₀ values were 0.33–0.49 μg/mL, and for strains with recovered-resistance, the IC₅₀ was 1.42–2.74 μg/mL. β-giardin amplicon (720 bp) was sequenced and analysis sequence revealed several amino acid mutations from resistant and recovered-sensitive strains of G. intestinalis. Most of the mutations were located in the ROD domain of β-giardin with a change from the sequence “TIARERA” in sensitive strains instead “IDRPRE” in resistant strains. A comparative sequence analysis in resistant, recovered-sensitive, and resistant-recovered strains revealed permanent mutation. This is the first report of combinatorial serine–proline–arginine repeats in the ROD domain of β-giardin, whereas such repeats have been reported previously in the HEAD domain of SF-assemblin proteins. This is the first time that the resistance to albendazole correlates with genetics but it is not necessarily caused by mutations in the β-giardin gene of G. intestinalis.     

Durian (Durio zibethinus Murr.) cultivars as nutritional supplementation to rat's diets.

The properties of Mon Thong, Chani and Kan Yao durian (Durio zibethinus Murr.) cultivars were compared in vitro and in vivo studies in order to find the best one as a supplement to antiatherosclerotic diet. Total polyphenols (361.4+/-35.3 mgGAE/100g FW), flavonoids (93.9+/-8.9 mgCE/100g FW) and total antioxidant capacity determined by DPPH and beta-carotene-linoleic acid assays (261.3+/-25.3 microMTE/100g FW and 77.8+/-7.8% of inhibition) were maximal in Mon Thong in comparison with Chani and Kan Yao and showed a good correlation between these three variables (R(2)=0.9859). Five groups of rats were fed diets supplemented with cholesterol and different durian cultivars. Diets supplemented with Mon Thong and to a lesser degree with Chani and Kan Yao significantly hindered the rise in the plasma lipids (TC - 8.7%, 16.1% and 10.3% and (b) LDL-C - 20.1%, 31.3% and 23.5% for the Chol/Kan Yao, Chol/Mon Thong and Chol/Chani, respectively) and the decrease in plasma antioxidant activity (P<0.05). Nitrogen retention remained significantly higher in Chol/Mon Thong than in other diet groups. Diet supplemented with Mon Thong affected the composition of plasma fibrinogen in rats and showed more intensity in protein bands around 47 kDa. No lesions were found in the examined tissue of heart and brains. Mon Thong cultivar is preferable for the supplementation of the diet as positively influenced the lipid, antioxidant, protein and metabolic status. The durian fruit till now was not investigated extensively, therefore based on the results of this study durian cultivars can be used as a relatively new source of antioxidants.