Polyamine patterns in haploid and diploid tobacco tissues and in vitro cultures

The aim of this work was to determine PAs levels in pith tissues and callus cultures from haploid and diploid tobacco plants, explanted from the apical and basal regions of the stem. These explants were cultured in an RM-64 medium supplied with IAA and kinetin, under light or in the dark, during successive subcultures. PAs levels followed a basipetal decrease in diploid and an increase in haploid, pith tissues. A similar pattern of total PAs (free + conjugated) was observed for the callus of diploid and haploid plants maintained in the light, and for the haploid callus in the dark, whereas the diploid callus in the dark showed a constant increase in total PAs levels until the end of culture. The PA increase in the diploid callus in the dark was related to free Put levels increase. The ploidy status of the plants could express different PA gradients together with the plant pith and in vitro callus cultures.

Expression of SMAD proteins, TGF-beta/activin signaling mediators, in human thyroid tissues

OBJECTIVE: To investigate the expression of SMAD proteins in human thyroid tissues since the inactivation of TGF-β/activin signaling components is reported in several types of cancer. Phosphorylated SMAD 2 and SMAD3 (pSMAD2/3) associated with the SMAD4 induce the signal transduction generated by TGF-β and activin, while SMAD7 inhibits this intracellular signaling. Although TGF-β and activin exert antiproliferative roles in thyroid follicular cells, thyroid tumors express high levels of these proteins. MATERIALS AND METHODS: The protein expression of SMADs was evaluated in multinodular goiter, follicular adenoma, papillary and follicular carcinomas by immunohistochemistry. RESULTS: The expression of pSMAD2/3, SMAD4 and SMAD7 was observed in both benign and malignant thyroid tumors. Although pSMAD2/3, SMAD4 and SMAD7 exhibited high cytoplasmic staining in carcinomas, the nuclear staining of pSMAD2/3 was not different between benign and malignant lesions. CONCLUSIONS: The finding of SMADs expression in thyroid cells and the presence of pSMAD2/3 and SMAD4 proteins in the nucleus of tumor cells indicates propagation of TGF-β/activin signaling. However, the high expression of the inhibitory SMAD7, mostly in malignant tumors, could contribute to the attenuation of the SMADs antiproliferative signaling in thyroid carcinomas.

Evaluation of Femtosecond Laser-Induced Breakdown Spectroscopy for Analysis of Animal Tissues

The aim of this work was to evaluate the performance of femtosecond laser-induced breakdown spectroscopy (fs-LIBS) for the determination of elements in animal tissues. Sample pellets were prepared from certified reference materials, such as liver, kidney, muscle, hepatopancreas, and oyster, after cryogenic grinding assisted homogenization. Individual samples were placed in a two-axis computer-controlled translation stage that moved in the plane orthogonal to a beam originating from a Ti:Sapphire chirped-pulse amplification (CPA) laser system operating at 800 mu and producing a train of 840 mu J and 40 fs pulses at 90 Hz. The plasma emission was coupled into the optical fiber of a high-resolution intensified charge-coupled device (ICCD)-echelle spectrometer. Time-resolved characteristics of the laser-produced plasmas showed that the best results were obtained with delay times between 80 and 120 ns. Data obtained indicate both that it is a matrix-independent sampling process and that fs-LIBS can be used for the determination of Ca, Cu, Fe, K, Mg, Na, and P, but efforts must be made to obtain more appropriate detection limits for Al, Sr, and Zn.

Extraction and purification of total RNA from banana tissues (small scale)

Introduction. This protocol aims at preparing total RNA for gene expression analysis by Northern blots, RT-PCR and real-time quantitative PCR; cDNA isolation by RTPCR; and cDNA library construction. The principle, key advantages, starting plant material, time required for obtaining total RNA and expected results are presented. Materials and methods. This part describes the required materials and the 27 steps necessary for preparing RNA from peel and pulp fruit tissue: preparation of plant tissue powder, preparation of the complete RNA extraction buffer and isolation of RNA from ground banana fruit tissue. Results. Extraction of total RNA by the method described makes it possible to achieve electrophoresis under denatured conditions and in vitro reverse transcription. An example for Northern blot analysis is illustrated.

Epidermal growth factor in liposomes may enhance osteoclast recruitment during tooth movement in rats

Objective: To evaluate the effects of local administration of epidermal growth factor (EGF) located within liposomes on recruitment of osteoclasts during mechanical force in rats. Materials and Methods: An orthodontic elastic band was inserted between the left upper first and second molars, to move mesially the first molar. Rats were randomly divided into 4 groups (n = 8): EGF (2 ng/mu L) located within liposomes (group 1), liposomes only (group 2), soluble EGF (2 ng/mu L; group 3), or vehicle alone (group 4). The solutions were injected into the region of the root furcation of the left first molar after elastic band insertion. Tooth movement was measured using a plaster model of the maxilla, and the number of osteoclasts recruited at the pressure side of the first molar was histologically evaluated. Results: Intergroup analysis showed that there was no significant difference between group 2 and group 4 (P >.05) and between group 1 and group 3 (P >.05). However, group 1 and group 3 exhibited greater differences in tooth movement than group 2 and group 4 (P <.05). On the other hand, group 1 showed greater tooth movement than groups 2 and 4 with statistical significance (P <.01). The increase in the number of osteoclasts in group 1 was significantly higher than in the other groups (P <.05). Conclusion: Exogenous EGF-liposome administration has an additive effect when compared with soluble EGF on the rate of osteoclast recruitment, producing faster bone resorption and tooth movement.

Identification of Neoplasias of Breast Tissues Using a Powder Diffractometer

An investigation was carried out to study the potential use of the angular distribution of scattered photons by human breast samples for a rapid identification of neoplasias of breast tissues. This technique has possible applications as diagnostic aid for breast cancer. In this work, a commercial powder diffractometer was used to obtain the scattering profiles from breast tissues histopathologically classified as normal breast tissues, fibroadenomas (benign breast diseases) and carcinomas (malignant breast diseases), in the interval 0.02 angstrom(-1) < x < 0.62 angstrom(-1). The experimental methods and data corrections are discussed in detail, and they included background subtraction, polarization, self-attenuation and geometric effects. The validation of the experimental procedure was achieved through an analysis of water sample. The results showed that the scattering profile is a unique impression of each type of tissue, being correlated with their microscopic morphological features. Multivariate analysis was applied to these profiles in order to verify if the information carried by these scattering profiles allow the differentiation between normal, benign and malignant breast tissues. The statistical analysis results showed that a correct identification of 75% of the analyzed samples is accomplished. The values of sensibility and specificity of this method in correctly differentiating between normal and neoplastic samples were 95.6% and 82.3%, respectively, while the values for differentiation between benign and malignant neoplasias were 78.6% and 62.5%. These initial results indicate the feasible use of commercial powder diffractometer to provide a rapid diagnostic with a high sensitivity.

Toxic Epidermal Necrolysis-like Cutaneous Lupus Erythematosus: A Series of Three Patients

Toxic epidermal necrolysis-like lesions have been described in the setting of lupus erythematosus, and have been considered as a specific hyperacute variant of cutaneous lupus erythematosus, with features different from classical drug-related toxic epidermal necrolysis. We report here a series of three patients with lupus erythematosus who presented with severe worsening of their cutaneous disease in a toxic epidermal necrolysis-like fashion. We compared these cases with cases reported previously. Based on this discussion, we speculate that some of these patients may have classical drug-related toxic epidermal necrolysis rather than lupus erythematosus-related toxic epidermal necrolysis.

Mesenchymal progenitor cells from canine fetal tissues: yolk sac, liver, and bone marrow

During fetal development, mesenchymal progenitor (MP) cells are co-localized in major hematopoietic territories, such as yolk sac (YS), bone marrow (BM), liver (LV), and others. Studies using mouse and human MP cells isolated from fetus have shown that these cells are very similar but not identical to adult mesenchymal stem cells (MSC). Their differentiation potential is usually restricted to production of highly committed osteogenic and chondrogenic precursors. Such properties of fetal MP cells can be very useful for tissue regeneration, when a great number of committed precursors are required. The objectives of this study were to isolate and characterize MP cells from canine YS, BM, and LV in early and late stages of fetal development. Gestational stage was identified, and cell culture conditions were evaluated for efficient isolation of canine MP cells. All canine fetal MP cells expressed vimentin, nestin, and CD44 proteins. Cytokeratin 18 expression was observed in BM-and LV-MP cells, and vascular endothelial (VE)-cadherin expression was observed only in YS-MP cells. A small number of MP cells (5%) from LV and YS expressed Oct3/4 protein. The differentiation potential of canine fetal MP cells varied significantly: YS- and BM-MP cells differentiated into bone and cartilage, whereas LV-MP cells differentiation was limited to osteogenic fate. None of the canine fetal MP cells were able to differentiate into adipose cells. Our data suggest that canine fetal MP cells are an appropriate in vitro model to study MP biology from hematopoietic territories and they are a source of committed osteogenic and chondrogenic precursors for regenerative medicine.

Ultrastructure of the Interface between Periodontal Tissues and Titanium Mini-Implants

Objective: To describe the ultrastructure of the interface between periodontal tissues and titanium mini-implants in rat mandibles. Materials and Methods: A titanium mini-implant was placed between the buccal roots of the mandibular first molar of 24 adult rats. After 21, 30, 45, 60, 90, and 120 days of implantation, the mandibular portion was removed and fixed in cacodylate-buffered 2% glutaraldehyde + 2.5% formaldehyde. The material was decalcified and processed for scanning and transmission electron microscopy. Results: Ultrastructural analysis revealed a thin cementum-like layer at longer times after implantation at the areas in which the periodontal ligament was in contact with the implant. Conclusions: The alveolar bone and the periodontal ligament reorganized their constituents around the implant, and a thin cementum-like layer was formed at longer times after implantation at the areas in which the periodontal ligament was in contact with the implant. (Angle Orthod. 2010;80:459-435.)

Melatonin reduces LH, 17 beta-estradiol and induces differential regulation of sex steroid receptors in reproductive tissues during rat ovulation

Background: Melatonin is associated with direct or indirect actions upon female reproductive function. However, its effects on sex hormones and steroid receptors during ovulation are not clearly defined. This study aimed to verify whether exposure to long-term melatonin is able to cause reproductive hormonal disturbances as well as their role on sex steroid receptors in the rat ovary, oviduct and uterus during ovulation. Methods: Twenty-four adult Wistar rats, 60 days old (+/-250 g) were randomly divided into two groups. Control group (Co): received 0.9% NaCl 0.3 mL + 95% ethanol 0.04 mL as vehicle; Melatonin-treated group (MEL): received vehicle + melatonin [ 100 mu g/100 g BW/day] both intraperitoneally during 60 days. All animals were euthanized by decapitation during the morning estrus at 4 a. m. Results: Melatonin significantly reduced the plasma levels of LH and 17 beta-estradiol, while urinary 6-sulfatoximelatonin (STM) was increased at the morning estrus. In addition, melatonin promoted differential regulation of the estrogen receptor (ER), progesterone receptor (PR), androgen receptor (AR) and melatonin receptor (MTR) along the reproductive tissues. In ovary, melatonin induced a down-regulation of ER-alpha and PRB levels. Conversely, it was observed that PRA and MT1R were up-regulated. In oviduct, AR and ER-alpha levels were down-regulated, in contrast to high expression of both PRA and PRB. Finally, the ER-beta and PRB levels were down-regulated in uterus tissue and only MT1R was up-regulated. Conclusions: We suggest that melatonin partially suppress the hypothalamus-pituitary-ovarian axis, in addition, it induces differential regulation of sex steroid receptors in the ovary, oviduct and uterus during ovulation.

Three-dimensional finite element thermal analysis of dental tissues irradiated with Er,Cr:YSGG laser

In the present study, a finite element model of a half-sectioned molar tooth was developed in order to understand the thermal behavior of dental hard tissues (both enamel and dentin) under laser irradiation. The model was validated by comparing it with an in vitro experiment where a sound molar tooth was irradiated by an Er,Cr:YSGG pulsed laser. The numerical tooth model was conceived to simulate the in vitro experiment, reproducing the dimensions and physical conditions of the typical molar sound tooth, considering laser energy absorption and calculating the heat transfer through the dental tissues in three dimensions. The numerical assay considered the same three laser energy densities at the same wavelength (2.79 mu m) used in the experiment. A thermographic camera was used to perform the in vitro experiment, in which an Er, Cr: YSGG laser (2.79 mu m) was used to irradiate tooth samples and the infrared images obtained were stored and analyzed. The temperature increments in both the finite element model and the in vitro experiment were compared. The distribution of temperature inside the tooth versus time plotted for two critical points showed a relatively good agreement between the results of the experiment and model. The three dimensional model allows one to understand how the heat propagates through the dentin and enamel and to relate the amount of energy applied, width of the laser pulses, and temperature inside the tooth. (C) 2008 American Institute of Physics. [DOI: 10.1063/1.2953526]

Hormone-regulated expression and distribution of versican in mouse uterine tissues

Background: Remodeling of the extracellular matrix is one of the most striking features observed in the uterus during the estrous cycle and after hormone replacement. Versican (VER) is a hyaluronan-binding proteoglycan that undergoes RNA alternative splicing, generating four distinct isoforms. This study analyzed the synthesis and distribution of VER in mouse uterine tissues during the estrous cycle, in ovariectomized (OVX) animals and after 17beta-estradiol (E2) and medroxyprogesterone (MPA) treatments, either alone or in combination. Methods: Uteri from mice in all phases of the estrous cycle, and animals subjected to ovariectomy and hormone replacement were collected for immunoperoxidase staining for versican, as well as PCR and quantitative Real Time PCR. Results: In diestrus and proestrus, VER was exclusively expressed in the endometrial stroma. In estrus and metaestrus, VER was present in both endometrial stroma and myometrium. In OVX mice, VER immunoreaction was abolished in all uterine tissues. VER expression was restored by E2, MPA and E2+MPA treatments. Real Time PCR analysis showed that VER expression increases considerably in the MPA-treated group. Analysis of mRNA identified isoforms V0, V1 and V3 in the mouse uterus. Conclusion: These results show that the expression of versican in uterine tissues is modulated by ovarian steroid hormones, in a tissue-specific manner. VER is induced in the myometrium exclusively by E2, whereas MPA induces VER deposition only in the endometrial stroma.

Photochemistry of anthocyanins and their biological role in plant tissues

Anthocyanins, the major red, purple, and blue pigments of plants, absorb visible as well as UV radiation and are effective antioxidants and scavengers of active oxygen species. In plant leaves, one of the functional roles proposed for anthocyanins is protection of the photosynthetic apparatus from the effects of excess incident visible or UV-B radiation and photooxidative stress. In essence, a photoprotective role requires that the excited singlet states of both complexed and uncomplexed anthocyanins deactivate back to the ground state so quickly that intersystem crossing, photoreaction, and diffusion-controlled quenching processes cannot compete. Studies of the photochemical properties of synthetic analogs of anthocyanins and of several naturally occurring anthocyanins show that this is indeed the case, uncomplexed anthocyanins decaying back to the ground state via fast (subnanosecond) excited-state proton transfer (ESPT) and anthocyanin-copigment complexes by fast (sub-picosecond) charge-transfer-mediated internal conversion.

Characterization of electrical penetration graphs of Bucephalogonia xanthophis, a vector of Xylella fastidiosa in citrus

The sharpshooter Bucephalogonia xanthophis (Berg) (Homoptera: Cicadellidae) is a vector of the xylem-limited bacterium, Xylella fastidiosa (Wells, Raju, Hung, Weisburg, Mandelco-Paul, and Brenner), which causes citrus variegated chlorosis. Despite the importance of citrus variegated chlorosis, the probing behavior of vectors on citrus and its implications for transmission of X. fastidiosa have not been studied. Here we studied electrical penetration graph (EPG-DC system) waveforms produced by B. xanthophis on Citrus sinensis (L.) Osbeck (Rutaceae), and their relationships with stylet activities and xylem ingestion. Electrical penetration graph waveforms were described based on amplitude, frequency, voltage level, and electrical origin of the observed traces during stylet penetration on plant tissues. The main waveforms were correlated with histological observations of salivary sheaths in plant tissues and excretion analysis, in order to determine stylet activities and their precise position. Six waveforms and associated activities are described: (S) secretion of salivary sheath and intracellular stylet pathway, (R) resting during stylet pathway, (Xc) contact of stylets with xylem vessels, (Xi) active xylem ingestion, (N) interruption within the xylem phase (during Xc or Xi), and (W) withdrawal of stylet from the plant. The sharpshooter spent 91.8% of its probing time with its stylet in the xylem, where the main activity was ingestion (Xi: 97.5%). During a probe, the most likely sequence of events is secretion of salivary sheath and pathway (S) through epidermal and parenchyma cells (all individuals), followed by contact with xylem (Xc) (67.6% of all individuals) and ingestion (Xi) (88.3% of those that exhibit waveform Xc). The mean time to contact the xylem (Xc) and initiate ingestion (Xi) after onset of the first probe was 27.8 and 34.2 min, respectively. However, sustained xylem ingestion (Xi > 5 min) was established after 39.8 min, on average. This information is basic for future studies on the transmission mechanisms of X. fastidiosa and in order to establish control strategies aimed at interfering with this process.

Discriminant analysis of trace elements in normal, benign and malignant breast tissues measured by total reflection X-ray fluorescence

In this work total reflection X-ray fluorescence spectrometry has been employed to determine trace element concentrations in different human breast tissues (normal, normal adjacent, benign and malignant). A multivariate discriminant analysis of observed levels was performed in order to build a predictive model and perform tissue-type classifications. A total of 83 breast tissue samples were studied. Results showed the presence of Ca, Ti, Fe, Cu and Zn in all analyzed samples. All trace elements, except Ti, were found in higher concentrations in both malignant and benign tissues, when compared to normal tissues and normal adjacent tissues. In addition, the concentration of Fe was higher in malignant tissues than in benign neoplastic tissues. An opposite behavior was observed for Ca, Cu and Zn. Results have shown that discriminant analysis was able to successfully identify differences between trace element distributions from normal and malignant tissues with an overall accuracy of 80% and 65% for independent and paired breast samples respectively, and of 87% for benign and malignant tissues. (C) 2009 Elsevier B.V. All rights reserved.