Identification and characterization of microRNAs in the screwworm flies Cochliomyia hominivorax and Cochliomyia macellaria (Diptera: Calliphoridae).

MicroRNAs (miRNAs) are small noncoding RNAs that modulate gene expression through post-transcriptional regulation. Here, we report the identification and characterization of miRNAs in two closely related screwworm flies with different feeding habits: Cochliomyia hominivorax and Cochliomyia macellaria. The New World screwworm, C. hominivorax, is an obligatory parasite of warm-blooded vertebrates, whereas the secondary screwworm, C. macellaria, is a free-living organism that feeds on decaying organic matter. Here, the small RNA transcriptomes of adults and third-instar larvae of both species were sequenced. A total of 110 evolutionarily conserved miRNAs were identified, and 10 putative precursor miRNAs (pre-miRNAs) were predicted. The relative expression of six selected miRNAs was further investigated, including miRNAs that are related to reproduction and neural processes in other insects. Mature miRNAs were also characterized across an evolutionary time scale, suggesting that the majority of them have been conserved since the emergence of the Arthropoda [540 million years ago (Ma)], Hexapoda (488 Ma) and Brachycera (195 Ma) lineages. This study is the first report of miRNAs for screwworm flies. We also performed a comparative analysis with the hereby predicted miRNAs from the sheep blowfly, Lucilia cuprina. The results presented may advance our understanding of parasitic habits within Calliphoridae and assist further functional studies in blowflies.

Pipeline for macro- and microarray analyses.

The pipeline for macro- and microarray analyses (PMmA) is a set of scripts with a web interface developed to analyze DNA array data generated by array image quantification software. PMmA is designed for use with single- or double-color array data and to work as a pipeline in five classes (data format, normalization, data analysis, clustering, and array maps). It can also be used as a plugin in the BioArray Software Environment, an open-source database for array analysis, or used in a local version of the web service. All scripts in PMmA were developed in the PERL programming language and statistical analysis functions were implemented in the R statistical language. Consequently, our package is a platform-independent software. Our algorithms can correctly select almost 90% of the differentially expressed genes, showing a superior performance compared to other methods of analysis. The pipeline software has been applied to 1536 expressed sequence tags macroarray public data of sugarcane exposed to cold for 3 to 48 h. PMmA identified thirty cold-responsive genes previously unidentified in this public dataset. Fourteen genes were up-regulated, two had a variable expression and the other fourteen were down-regulated in the treatments. These new findings certainly were a consequence of using a superior statistical analysis approach, since the original study did not take into account the dependence of data variability on the average signal intensity of each gene. The web interface, supplementary information, and the package source code are available, free, to non-commercial users at http://ipe.cbmeg.unicamp.br/pub/PMmA.

Effects of a photic input on the human cortico-motoneuron connection.

OBJECTIVES: Disease manifestations such as photic cortical reflex myoclonus or myoclonus due to intermittent light stimulation rely on a pathologic interaction between non-structured visual inputs and the corticospinal system. We wanted to assess the normal interaction, if any, between a prior photic input and the output of the cortico-motoneuron connection. METHODS: In 9 consenting healthy subjects we quantified the changes exerted by a sudden, unexpected bright light flash on (i) the motor potentials (MEPs) evoked in the right first dorsal interosseous muscle (FDI) by transcranial magnetic or electrical stimulation (TMS/TES) of the primary motor cortex, (ii) the FDI F-waves and (iii) the soleus H-wave. Separately, we measured the simple reaction times to the flash itself. All determinations were repeated twice with an interval of 2-24 months. RESULTS: When the flash preceded TMS by 55-70 ms, the MEP size was reduced, while at interstimulus intervals (ISIs) of 90-130 ms it was enlarged. Statistical significance (P<0.05) emerged at ISIs of 55, 70, 100, 105 and 120 ms. Conversely, the MEP latency was prolonged at ISIs of 55-70 ms and shortened at ISIs of 90-130 ms (P<0.05 at ISIs of 55, 110 and 130 ms). Electrical MEPs were enhanced at an ISI of 120 ms. The F-wave size showed a non-significant trend of enhancement at ISIs of 90-130 ms. The soleus H-wave showed significant enlargement at ISIs of 90-130 ms (P<0.05 at ISIs of 100 and 105 ms). The minimum reaction time was on average 120 ms. CONCLUSIONS: An unexpected photic input, to which no reaction is planned, can cause an early inhibition of the responses to TMS. We think its origin lies within the primary motor cortex, since it is not associated with changes in spinal excitability or electrical MEPs. A later facilitation persists using TES and has a temporal relationship with an enlargement of the soleus H-wave. Thus, it likely results from activation of descending (possibly reticulospinal) fibers that excite the spinal motor nucleus.

Intravenous leiomyomatosis of the uterus. A report of three cases.

Three cases of intravenous leiomyomatosis (IVL) of the uterus, a rare benign smooth-muscle tumor, are described. A preoperative diagnosis of IVL was not made in any of the patients, all of which presented with a pelvic mass with the presumptive diagnosis of leiomyoma. Surgical exploration confirmed the presence of uterine mass and two of the three cases showed extra-uterine extension into the ovarian or uterine veins. Histological examination demonstrated a fascicular pattern of bland spindle-shaped smooth-muscle cells, which extended to veins inside the myometrium or to extrauterine veins. This was confirmed by immunohistochemical stain for desmin and factor VIII. Despite their histological benignity, these lesions have a tendency to metastasize and are closely related to the conditions called "benign metastasizing leiomyoma" and "intracaval mass and cardiac extension". The primary treatment of IVL is hysterectomy and excision of any extrauterine tumor, when technically feasible. Anti-estrogenic therapy has been suggested as potentially useful in controlling of unresectable tumor. According to the literature, the follow-up must be long and periodic postoperative ultrasonic or magnetic nuclear resonance imaging studies may be useful in detecting growth of residual intravascular tumor.

[Value of echography in the measurement of changes in the total and visceral adipose mass]. / Validità dell'ecografia nella valutazione delle modificazioni della massa adiposa totale e viscerale.

Various methods to assess lean and fat body masses and abdominal-visceral adipose tissue were compared in 26 obese women on extremely hypocaloric diets. The following anthropometric parameters were measured before and after 15 days of extreme calorie restriction: arm circumference (A), waist circumference (W), hip circumference (H) and thigh circumference (T); W/H, W/T, A/H and A/T ratios; subcutaneous biceps , triceps , subscapular, abdominal and thigh folds; echographic thickness of abdominal muscle-aorta; area of visceral (VAT), total (AT) and subcutaneous adipose tissue measured using computed tomography, lean and fat body masses assessed by impedance measurement. Under standard conditions it was shown that echography, like impedance measurement, was the most useful method for assessing lean and fat body masses in obese subjects, whereas plicometry was not found reliable. On extremely hypocaloric diets, echography is able to assess the variations of lean and fat body masses, whereas impedance measurement overestimated the lean body mass. Localisation indexes of visceral fat based on body circumferences were shown not to be suitable for the evaluation of changes in visceral adipose tissue, at least when these were slight. Echography was also the most useful method to assess slight changes in visceral adipose tissue.

The sentinel lymph node in T1N0 breast cancer.

The authors report their experience gathered from December 1998 to December 1999 in the use of the sentinel lymph node (SN) method in breast cancer treatment. In 20 out of 21 cases (95%) localization of the SN was obtained by scintigraphy while in 19 cases (90.5%) the SN was found during surgery. Histological examination of the axillary lymph nodes gave a 95% accuracy with only one negative SN associated with positive axillary lymph nodes out of a total of 19. However, the authors have subjected all patients to a complete three-level axillary dissection since they believe the method applied has not yet been fully validated.

[Diagnosis and treatment of non-palpable breast lesions: from the biopsy to the microhistology results. Our experience]. / La diagnosi ed il trattamento delle lesioni non palpabili della mammella: dalla biopsia su repere alla microistologia. Nostra esperienza.

The aim of modern senology lies in the diagnosis and treatment of non-palpable breast lesions (NPBLs). Through the diffusion of regional mammography screening the lesions being observed are continuously smaller, thus calling for more and more accurate methodology. Our experience in this area is based on the use of certain methods for retrieval and removal of NPBLs, such as Kopan's sec. philo-guide, ultrasound and advanced breast biopsy instrumentation. In our opinion methods allowing total removal of lesions in order to obtain complete histopathological characterization and enabling adequate therapeutic programs are to be preferred. In reviewing case studies a noteworthy increase of initial carcinoma (DCIS or LCIS), from 19.5% to 57.1%, has been observed in the last three years due to the extensive use of the aforementioned methods.

[Can analysis of the molecular status of the p53 gene contribute to improving the therapeutic strategy for breast carcinoma?]. / L'analisi dello stato molecolare del gene p53 può contribuire ad innovare le strategie terapeutiche nel carcinoma della mammella?

The occurrence of mutations in the p53 tumor suppressor gene is a specific and recurring genetic event in solid tumors. P53 plays a pivotal role in multiple cellular processes such as cell growth control, DNA repair and programmed cell death. Genotoxic damage, also induced by chemotherapy or radiotherapy, induces p53 overexpression in order to control the rate of proliferating damaged cells, thus triggering the mismatch repair or apoptotic pathways. P53 inactivation determines a condition of genetic instability, justifying the subsequent susceptibility to acquire mutations of different other genes. P53 mutations are associated with worse prognosis and with chemo/radioresistance, due to the inability to trigger p53-dependent programmed cell death. Molecular diagnostic strategies show 32% p53 mutations in breast cancer. The analysis of the p53 gene performed by FAMA (Fluorescence Assisted Mismatch Analysis) in high-risk breast cancer patients with > or = 10 involved axillary nodes may help identify a subset of very high risk BC patients (vHR-BC) with poorer prognosis and a subset with better prognosis, potentially responsive to medical treatments. The accurate evaluation of the p53 status can predict prognosis and sensitivity to chemotherapy, thus representing the first step toward better definition of therapeutic strategies according to the molecular characterization of the individual patient.

[Noise generated from driving-in foundation piles]. / Rilievi di rumore generato dalla battitura di pali di consolidamento del terreno.

Results of sound pressure measurements during driving-in of foundation piles are reported. The results are then compared with latest recommendation from the standpoint of hearing impairment due to continuous or impulsive noise.

[Non-palpable lesions of the breast: retrospective clinical study]. / Lesioni non palpabili della mammella: studio clinico retrospettivo.

A non-palpable breast lesion (NPBL) is a disease of the mammary gland that cannot be detected during clinical examination but that can be visualized by mammography and/or ultrasonography, either during screening programs or sometimes in asymptomatic women. These small lesions require an adequate diagnostic-therapeutic approach to ensure correct treatment. The aim of the present study was to analyse a series of NPBLs retrospectively in order to define them nosologically and establish an adequate diagnostic-therapeutic work-up for such cases. Ninety-three patients with a total of 99 NPBLs were observed from January 1989 to December 1999. The 99 NPBLs were submitted to ultrasonography and 31 (31.3%) were also submitted to US-guided fine needle aspiration biopsy (FNAB). Later on the diagnostic-therapeutic procedure involved surgical biopsy after radiological centering and, in the case of malignant neoplastic lesions, surgical intervention and adjuvant therapy. Ultrasonography confirmed the presence of NPBL in 45 cases of the 99 detected at mammography (45.4%). Cytological examination of the 31 FNABs yielded the following results: unreliable 19.3%, suspected malignancy 42%, negative for neoplastic cytology 6.5%, positive for carcinoma 32.3%. The histological diagnosis was one of mammary carcinoma in 41 patients (43%). Among the 41 carcinomas there were 8 (19.5%) carcinomas in situ, 24 (58.5%) invasive ductal carcinomas, 8 (19.5%) invasive lobular carcinomas, and 1 (2.5%) medullary carcinoma. In the 32 (80%) patients submitted to lymphadenectomy for 33 invasive carcinomas, 6 patients (18.7%) presented positive lymph nodes (N1). The Authors conclude that NPBLs are an important clinical entity because they may be the expression of a malignant lesion; most NPBLs are diagnosed during screening programs or sometimes in asymptomatic women by means of mammography, which is the only standardised method for their identification. The poor diagnostic capability of non-invasive methods and the potential malignancy of NPBL justify the indication for surgical excisional biopsies; in cases of histological findings of malignancy it is often possible to perform conservative surgery with similar results to radical surgery in terms of survival. When NPBLs turn out to be invasive carcinomas, a concomitant lymphadenectomy is mandatory.

Sex ratio and size structure of Micropogonias furnieri (Desmarest, 1823) (Perciformes, Sciaenidae) in Sepetiba Bay, Rio de Janeiro, Brazil.

Sex ratio and size structure constitute basic information in assessing reproductive potential and estimating stock size in fish populations. One hundred fifty-one individuals of Micropogonias furnieri caught by experimental otter trawls, in three zones (inner, central, and outer) of Sepetiba Bay between October 1998 and September 1999 were examined. Males outnumbered females (1.3:1.0) in all zones but no significant differences were detected. Only in the outer zone (5.0:1.0) were male/female rates significantly different according to the chi-square test. No temporal differences were observed in sex ratio. Fish size ranged from 81 to 244 mm total length (TL) with significant differences in 155 to 185 mm TL size classes, where males predominated, and a slightly higher number of females were observed for the smaller size class (TL = 95-150 mm). Size distribution varied according to the zone, with juveniles predominating in the inner and adults in the outer zone. Spatial difference in size structure observed in this study indicates that the inner bay is a rearing ground during the first life-cycle period, and movement toward the sea occurs as fish increase in size.

Resting metabolic rate, body composition and thyroid hormones. Short term effects of very low calorie diet.

27 obese women (relative body weight 191 +/- 27%), aged 35 +/- 11, underwent a 15 day very low calorie diet (VLCD: 1337 kJ/day). Mean body weight loss was 6 +/- 1.5 kg. Resting metabolic rate (RMR), measured by indirect calorimetry, showed a significant decrease both in absolute values (from 1793 +/- 225 to 1569 +/- 231 kcal/day; P less than 0.001), and in keal/kg body weight/day (from 17 +/- 2, to 16 +/- 2, P less than 0.001). Serum triiodothyronine (T3) significantly decreased from 1.1 +/- 0.2 to 1 +/- 0.2 microgram/1 (P less than 0.001). Before VLCD the differences between measured RMR and theoretical RMR (RMR-T) were not significant. After VLCD RMR values were significantly (P less than 0.001) lower than RMR-T. It was concluded that fat-free mass loss, combined with periferic thyroxin (T4) reduced monodeiodination in its more active form mainly due to low calorie intake T3 could be responsible for the important reduction observed in the resting metabolic rate.

Cortical excitability in drug-naive patients with partial epilepsy: a cross-sectional study.

OBJECTIVE: To use paired-pulse transcranial magnetic stimulation (TMS) to investigate cortical excitability in drug-naive patients with partial epilepsy. METHODS: Twenty-one drug-naive patients with partial epilepsy and 15 control subjects were studied. The relaxed threshold to TMS, the central silent period, and the intracortical inhibition/facilitation were measured. Statistics implied cluster analysis methods. Also assessed were the patient interictal EEG epileptiform abnormalities (EAs) on a semiquantitative basis. Then the TMS was contrasted to the clinical and EEG findings, using chi2 or Fisher exact tests. RESULTS: One-third of the patients made up a "pathologic" cluster with a disrupted intracortical inhibition (p < 0.01). Two-thirds had a normal inhibition. Interictal EAs predominated in the pathologic cluster, for frequency (p < 0.04), duration (p < 0.04), and focality (p < 0.02). CONCLUSIONS: Intracortical inhibition, which was impaired in one-third of the patients, reflects gamma-aminobutyric acid (GABA) activity within cortical area 4. Defective GABA inhibition is a typical pathogenic factor in partial epilepsy. Transcranial magnetic stimulation proved able to detect it. The weaker cortical inhibition had a direct relation to the severity of interictal epileptiform abnormalities.

Cortical excitability and sleep deprivation: a transcranial magnetic stimulation study.

The objective was to assess the changes in cortical excitability after sleep deprivation in normal subjects. Sleep deprivation activates EEG epileptiform activity in an unknown way. Transcranial magnetic stimulation (TMS) can inform on the excitability of the primary motor cortex. Eight healthy subjects (four men and four women) were studied. Transcranial magnetic stimulation (single and paired) was performed by a focal coil over the primary motor cortex, at the "hot spot" for the right first dorsal interosseous muscle. The following motor evoked potential features were measured: (a) active and resting threshold to stimulation; (b) duration of the silent period; (c) amount of intracortical inhibition on paired TMS at the interstimulus intervals of 2 and 3 ms and amount of facilitation at interstimulus intervals of 14 and 16 ms. The whole TMS session was repeated after a sleep deprivation of at least 24 hours. After the sleep deprivation, the threshold to stimulation (in the active and resting muscle), as well as the silent period, did not change significantly. By contrast, the paired stimulus study showed a significant (p<0.05) reduction in both intracortical inhibition and facilitation. Thus, TMS showed that sleep deprivation is associated with changes in inhibition-facilitation balance in the primary motor cortex of normal subjects. These changes might have a link with the background factors of the "activating" effects of sleep deprivation.

Pipeline for macro- and microarray analyses

The pipeline for macro- and microarray analyses (PMmA) is a set of scripts with a web interface developed to analyze DNA array data generated by array image quantification software. PMmA is designed for use with single- or double-color array data and to work as a pipeline in five classes (data format, normalization, data analysis, clustering, and array maps). It can also be used as a plugin in the BioArray Software Environment, an open-source database for array analysis, or used in a local version of the web service. All scripts in PMmA were developed in the PERL programming language and statistical analysis functions were implemented in the R statistical language. Consequently, our package is a platform-independent software. Our algorithms can correctly select almost 90 percent of the differentially expressed genes, showing a superior performance compared to other methods of analysis. The pipeline software has been applied to 1536 expressed sequence tags macroarray public data of sugarcane exposed to cold for 3 to 48 h. PMmA identified thirty cold-responsive genes previously unidentified in this public dataset. Fourteen genes were up-regulated, two had a variable expression and the other fourteen were down-regulated in the treatments. These new findings certainly were a consequence of using a superior statistical analysis approach, since the original study did not take into account the dependence of data variability on the average signal intensity of each gene. The web interface, supplementary information, and the package source code are available, free, to non-commercial users at http://ipe.cbmeg.unicamp.br/pub/PMmA.

Sex ratio and size structure of Micropogonias furnieri (Desmarest, 1823) (Perciformes, Sciaenidae) in Sepetiba Bay, Rio de Janeiro, Brazil

Sex ratio and size structure constitute basic information in assessing reproductive potential and estimating stock size in fish populations. One hundred fifty-one individuals of Micropogonias furnieri caught by experimental otter trawls, in three zones (inner, central, and outer) of Sepetiba Bay between October 1998 and September 1999 were examined. Males outnumbered females (1.3:1.0) in all zones but no significant differences were detected. Only in the outer zone (5.0:1.0) were male/female rates significantly different according to the chi-square test. No temporal differences were observed in sex ratio. Fish size ranged from 81 to 244 mm total length (TL) with significant differences in 155 to 185 mm TL size classes, where males predominated, and a slightly higher number of females were observed for the smaller size class (TL = 95-150 mm). Size distribution varied according to the zone, with juveniles predominating in the inner and adults in the outer zone. Spatial difference in size structure observed in this study indicates that the inner bay is a rearing ground during the first life-cycle period, and movement toward the sea occurs as fish increase in size.