Developing human minor salivary glands: morphological parallel relation between the expression of TGF-beta isoforms and cytoskeletal markers of glandular maturation

Morphogenesis of salivary glands involves complex coordinated events. Synchronisation between cell proliferation, polarisation and differentiation, which are dependent on epithelial-mesenchymal interactions and on the microenvironment, is a requirement. Growth factors mediate many of these orchestrated biological processes and transforming growth factor-beta (TGF-beta) appear to be relevant. Using immunohistochemistry and immunofluorescence, we have mapped the distribution of TGF-beta 1, 2 and 3 and compared it with the expression of maturation markers in human salivary glands obtained from foetuses ranging from weeks 4 to 24 of gestation. TGF-beta 1 first appeared during canalisation stage in the surrounding mesenchyme and, in the more differentiated stages, was expressed in the cytoplasm of acinar cells throughout the adult gland. TGF-beta 2 was detected since the bud stage of the salivary gland. Its expression was observed in ductal cells and increased along gland differentiation, TGF-beta 3 was detected from the canalisation stage of the salivary gland, being weakly expressed on ductal cells, and it was the only factor detected on myoepithelial cells. The data suggest that TGF-beta have a role to play in salivary gland development and differentiation.

Claudin expression is dysregulated in prostate adenocarcinomas but does not correlate with main clinicopathological parameters

Aims: Claudins, a large family of essential tight junction (TJ) proteins, are abnormally regulated in human carcinomas. The aim of this study was to determine the expression of claudins 1, 2, 3, 4, 5, 7, and 11 in prostate samples from Brazilian patients and correlate it with the clinicopathological features of prostate cancer. Methods: Using a tissue microarray (TMA) of specimens of prostate adenocarcinoma and benign prostatic hyperplasia (BPH) we analysed the expression of claudins 1, 2, 3, 4, 5, 7, and 11 by immunohistochemistry. Results: Claudin 4 was down-regulated and claudins 2, 3, and 5 were overexpressed in prostate adenocarcinomas compared with BPH samples. Expression of claudins 1 and 7 was similar in tumours and BPH samples. Claudin 11 was absent from all prostate samples. Overexpression of claudin 3 was associated with perineural invasion (p = 0.014) and tended to occur in advanced stages of the disease (p = 0.064). Increased expression of claudin 5 was marginally associated with perineural invasion (p = 0.060). Conclusions: Our results suggest that alterations in claudin expression occur in prostate cancer cells, although we have not found an association with the main clinicopathological parameters.

Human salivary gland morphogenesis: myoepithelial cell maturation assessed by immunohistochemical markers

Aims: Myoepithelial cells are important components of salivary gland structure, aiding the expulsion of saliva from acinar lobules. The aim was to evaluate the expression of smooth muscle actin (SMA), calponin, caldesmon, CD10, CD29, S100 protein, glial fibrillary acidic protein (GFAP) and p63 in myoepithelial cells during salivary gland morphogenesis to understand the maturation process of these cells and their possible use in the diagnosis of salivary gland lesions. Methods and results: Major and minor human salivary glands at various stages of development, derived from fetuses at 8-26 weeks of gestation, were studied immunohistochemically. Fully developed salivary glands were used as controls. The protein p63 was present in all stages of salivary gland morphogenesis from initial bud to terminal bud stage. CD29, S100 and calponin were detected increasingly as salivary gland structure matured and in fully developed salivary gland. Proteins GFAP, CD10 and caldesmon were not observed in myoepithelial cells of salivary glands. Conclusions: The proteins SMA, calponin, CD29, S100 and p63, which are present from the earliest stages of salivary gland maturation, are valuable myoepithelial markers but, although very specific, are not exclusive markers for this cell type.

Oral squamous cell carcinoma: status of tight junction claudins in the different histopathological patterns and relationship with clinical parameters. A tissue-microarray-based study of 136 cases

Aims Claudins are integral transmembrane proteins of the tight junctions, critical for maintaining cell adhesion and polarity. Alterations in the expression of individual claudins have been detected in carcinomas and appear to correlate with tumour progression. Methods In this study, a panel of anti-claudin antibodies (anti-claudins 1, 2, 3, 4, 5 and 7) was employed to map claudin expression in 136 cases of oral squamous cell carcinoma (OSCC) organised in a tissue microarray. Results Claudins were expressed in a reticular pattern up to the prickle layer in normal mucosal epithelium. In OSCC, claudins were strongly present in well-differentiated tumours, they presented mild and low expression in moderately differentiated OSCC, and were negative in poorly differentiated OSCC; the absences of claudin 1 (p = 0.002) and claudin 4 (p<0.001) were associated with moderately/poorly differentiated tumours. Strong expression of claudin 4 was associated with decreased perineural infiltration (p = 0.024). Claudins 5 and 7 were mostly negative or weakly expressed in all cases studied. Expression of claudin 7 was associated with the early clinical stages of the disease, whereas loss of claudin 7 tended to be more frequent in advanced stages of OSCC (p = 0.054). Absence of claudin 7 was also associated with absent vascular infiltration (p = 0.045) and with presence of recurrence (p = 0.052). Conclusions Claudin expression patterns showed a strong correlation with histological type of OSCC; claudin expression was decreased in areas of invasion, and negative in poorly differentiated tumours. This pattern may be related to evolution and prognosis of these tumours, especially in the case of claudin 7, which seems to be associated with a poor prognosis in OSCC.

Effects of neonatal handling on central noradrenergic and nitric oxidergic systems and reproductive parameters in female rats

Early-life environmental events that disrupt the mother-pup relationship may induce profound long-lasting changes on several behavioral and neuroendocrine systems. The neonatal handling procedure, which involves repeated brief maternal separations followed by experimental manipulations, reduces sexual behavior and induces anovulatory estrous cycles in female rats. On the afternoon of proestrus, neonatally handled females show a reduced surge of luteinizing hormone (LH) and an increased content of gonadotropin-releasing hormone in the medial preoptic area (MPOA). In order to detect the possible causes for the reduced ovulation and sexual behavior, the present study aimed to analyze the effects of neonatal handling on noradrenaline (NA) and nitric oxide (NO) levels in the MPOA on the afternoon of proestrus. Neonatal handling reduced MHPG (NA metabolite) levels and MHPG/NA ratio in the MPOA, indicating decreased NAergic activity. Additionally, neonatal handling decreased NO levels, as measured by the metabolites (NO x), nitrite and nitrate in the same period. We may conclude that the neonatal handling procedure decreased activity of the NAergic and NOergic systems in the MPOA during proestrus, which is involved in the control of LH and FSH secretion, and this may possibly explain the anovulatory estrous cycles and reduced sexual behavior of the neonatally handled female rats. Copyright (c) 2007 S. Karger AG, Basel.

Human osteoblastic cell response to a Ca- and P-enriched titanium surface obtained by anodization

The aim of the present study was to evaluate the in vitro osteogenic potential of subcultured human osteoblastic cells derived from alveolar bone on a titanium (Ti) surface produced by an anodized alkali treatment (BSP-AK). Primary osteoblastic cells were subcultured on BSP-AK and machined Ti discs (control) and grown for periods of up to 21 days under osteogenic conditions. Morphologic and biochemical methods were used to assess important parameters of in vitro bone-like tissue formation. Although no major differences were observed between the BSP-AK and the control Ti surface in terms of cell attachment and mineralized matrix formation, a significant increase in cell population, ALP activity, and collagen content was detected in cultures on BSP-AK surface. Our results demonstrate that human osteoblastic cells are sensitive to the BSP-AK-modified Ti surface during the transitional stage between the end of the proliferative phase and the onset of the differentiation /matrix maturation ones. Together with the good mechanical properties exhibited by the Ca- and P- coating, our findings suggest that BSP-AK treatment could be useful for the development of a new surface for dental and orthopedic implants. (c) 2008 Wiley Periodicals, Inc.J Biomed Mater Res 88A: 841-848, 2009

Ultrastructural Analysis of Radicular Dentine Surface Submitted to CO(2) Laser at Different Parameters

Background: There are no reported studies comparing different parameter settings of the CO(2) laser and irradiation direction considering their effect on the morphology of radicular dentine surface. Purpose: To evaluate the alterations of radicular dentine (cervical, middle, and apical thirds) irradiated with CO(2) laser at different potencies and irradiation directions. Study Design: Roots of 35 canines were prepared and randomly distributed according to the laser potency: GI: no laser treatment (control) (n = 5); GII, 2 W (n = 10); GIII: 4 W (n = 10); GIV: 6 W (n = 10). Each group (excepting GI) was divided in two subgroups according to the irradiation distance (n = 5): (A) parallel and (B) perpendicular to the root canal walls. The roots were splited longitudinally and analyzed by scanning electron microscopy in a qualiquatitative way. The scores were submitted to Kruskal-Wallis and Dunn`s tests. Results: No significant statistical differences were observed among root canal thirds (P > 0.05). The specimens irradiated with 2 W were statistically different (P < 0.05) from those irradiated with 4 and 6 W, which were statistically similar between themselves (P > 0.05). With 2, 4, and 6 W at in parallel irradiation and 2 W in perpendicular direction, the surface showed a fissured aspect. With 4 W in perpendicular direction and 6 W in parallel and perpendicular direction, surface was modified by laser action and exhibited fused areas. Conclusions: The intensity of the effects is dependent on the laser-irradiation dosimetries. Alterations were more intense when higher parameters were used. Microsc. Res. Tech. 72:737-743, 2009. (C) 2009 Wiley-Liss, Inc.

Crystallization and preliminary diffraction studies of native and selenomethionine CcmG (CycY, DsbE)

t Disulfide-bond (Dsb) proteins are a family of redox proteins containing a Cys-X-X-Cys motif. They are essential for disulfide-bond exchange in the bacterial periplasm and are necessary for the correct folding and function of many secreted proteins. CcmG (DsbE) is a reducing Dsb protein required for cytochrome c maturation. Crystals of Bradyrhizobium japonicum CcmG have been obtained that diffract X-rays to 1.14 Angstrom resolution. The crystals are orthorhombic, space group P2(1)2(1)2(1), with unit-cell parameters a = 35.1, b = 48.2, c = 90.2 Angstrom. Selenomethionine CcmG was expressed without using a methionine auxotroph or methionine-pathway inhibition and was purified without reducing agents.

Expression of glycoproteins in the vomeronasal organ reveals a novel spatiotemporal pattern of sensory neurone maturation

The main olfactory and the accessory olfactory systems are both anatomically and functionally distinct chemosensory systems. The primary sensory neurones of the accessory olfactory system are sequestered in the vomeronasal organ (VNO), where they express pheromone receptors, which are unrelated to the odorant receptors expressed in the principal nasal cavity. We have identified a 240 kDa glycoprotein (VNO240) that is selectively expressed by sensory neurones in the VNO but not in the main olfactory neuroepithelium of mouse. VNO240 is first expressed at embryonic day 20.5 by a small subpopulation of sensory neurones residing within the central region of the crescent-shaped VNO, Although VNO240 was detected in neuronal perikarya at this age, it was not observed in the axons in the accessory olfactory bulb until postnatal day 3.5, This delayed appearance in the accessory olfactory bulb suggests that VNO240 is involved in the functional maturation of VNO neurones rather than in axon growth and targeting to the bulb, During the first 2 postnatal weeks, the population of neurones expressing VNO240 spread peripherally, and by adulthood all primary sensory neurones in the VNO appeared to be expressing this molecule. Similar patterns of expression were also observed for NOC-1, a previously characterized glycoform of the neural cell adhesion molecule NCAM, To date, differential expression of VNO-specific molecules has only been reported along the rostrocaudal axis or at different apical-basal levels in the neuroepithelium. This is the first demonstration of a centroperipheral wave of expression of molecules in the VNO, These results indicate that mechanisms controlling the molecular differentiation of VNO neurones must involve spatial cues organised, not only about orthogonal axes, but also about a centroperipheral axis, Moreover, expression about this centroperipheral axis also involves a temporal component because the subpopulation of neurones expressing VNO240 and NOC-1 increases during postnatal maturation. (C) 2001 John Wiley & Sons, Inc.

Assembly and maturation of the flavivirus Kunjin virus appear to occur in the rough endoplasmic reticulum and along the secretory pathway, respectively

The intracellular assembly site for flaviviruses in currently not known but is presumed to be located within the lumen of the rough endoplasmic reticulum (RER), Building on previous studies involving immunofluorescence (IF) and cryoimmunoelectron microscopy of Kunjin virus (KUN)-infected cells, we sought to identify the steps involved in the assembly and maturation of KUN. Thus, using antibodies directed against envelope protein E in IF analysis, we found the accumulation of E within regions coincident with the RER and endosomal compartments. Immunogold labeling of cryosections of infected cells indicated that E and minor envelope protein prM were localized to reticulum membranes continuous with KUN-induced convoluted membranes (CM) or paracrystalline arrays (PC) and that sometimes the RER contained immunogold-labeled virus particles. Both proteins were also observed to be labeled in membranes at the periphery of the induced CIM or PC structures, but the latter were very seldom labeled internally. Utilizing drugs that inhibit protein and/or membrane traffic throughout the cell, we found that the secretion of KUN particles late in infection was significantly affected in the presence of brefeldin A and that the infectivity of secreted particles was severely affected in the presence of monensin and N-nonyl-deoxynojirimycin. Nocodazole did not appear to affect maturation, suggesting that microtubules play no role in assembly or maturation processes. Subsequently, we showed that the exit of intact virions from the RER involves the transport of individual virions within individual vesicles en route to the Golgi apparatus. The results suggest that the assembly of virions occurs within the lumen of the RER and that subsequent maturation occurs via the secretory pathway.

Maturation studies of pecan nuts grown in Queensland

Changes in chemical composition, physical and sensory characteristics were followed in two pecan cultivars Wichita and Western Schley harvested from a commercial orchard at Gatton in Queensland seven times during 1996. Testa colour of both pecan cultivars darkened and opalescence decreased as the nuts matured. Bitterness of Western Schley pecans decreased with maturity. Colour of shuck, shell and kernel of both cultivars developed as the nuts matured. Wichita pecans were larger than Western Schley at all harvest times. Both nut-in-shell and kernel moisture decreased with maturity, whereas oil and sucrose contents increased. Both pecan cultivars had reached advanced maturation by the first harvest on March 18.

The investigation of optimal bombardment parameters for transient and stable transgene expression in sorghum

This report outlines the development of optimized particle inflow gun (PIG) parameters for producing transgenic sorghum (Sorghum bicolor (L.) Moench). Both transient and stable expression were examined when determining these parameters. The uidA reporter gene (GUS) encoding beta -glucuronidase was used in transient experiments and the green fluorescent protein (GFP) used to monitor stable expression. Initially, optimization was conducted using leaf segments, as the generation of sorghum callus in sufficiently large quantities is time-consuming. Following leaf optimization, experiments were conducted using callus, identifying a high similarity between the two tissue types (r(s) = 0.83). High levels of GUS expression were observed in both leaf and callus material when most distant from the DNA expulsion point, and using a pressure greater than 1800 kPa. A higher level of expression was also observed when the aperture of the helium inlet valve was constricted. Using the optimized conditions (pressure of 2200 kPa, distance to target tissue of 15 cm from the expulsion point, and the aperture of the helium inlet valve at one full turn), three promoters (Ubiquitin, Actin1 and CaMV 35S) were evaluated over a 72-h period using GUS as the reporter gene. A significantly higher number of GUS foci were counted with the Ubiquitin construct over this period, compared to the Actin1 and CaMV 35S constructs. Stable callus sectors (on 2 mg l(-1) bialaphos) with GFP expression were visualized for as long as 6 wk post-bombardment. Using this optimized protocol, several plants were regenerated after having been bombarded with the pAHC20 construct (containing the bar gene), with molecular evidence confirming integration.

Associations involving delays (particularly long delays) between certain weather parameters and geomagnetic activity

Four sunspot-minimum periods (1963-1966, 1971-1977, 1983-1987 and 1992-1997) have been examined for the results which are presented. Using several different weather parameters, tropospheric gravity waves, enhanced cold fronts and two rainfall data sets in Eastern Australia, associations at reasonably high levels of significance have been found with enhanced geomagnetic activity (EGA). Statistically this EGA involved either short delays of several days or long delays of about 20 days. The geomagnetic parameters used were (a) the AE index (b) the hourly H component for a number of stations and (c) the daily K-P-sum value. The K-P-sum analyses have shown that the EGA associated with the delays form part of four or five cycles of recurrent geomagnetic activity for 27-day periodicities. Furthermore statistically two recurrent cycles are found to exist concurrently, one apparently related to the short delays and the other to the long delays. Periodicities of 13.5 days are created because the two sets are displaced from each other by approximately this interval. A brief reference is made to the 13.5 periodicity known to exist for geomagnetic activity and the evidence in the literature for active regions on the sun to be displaced by 180 degrees of solar longitude.

The influence of posture on perineal ultrasound imaging parameters

A prospective clinical study was carried out to evaluate the influence of posture on perineal ultrasound imaging parameters. One hundred and thirty-two consecutive women presenting with symptoms of lower urinary tract dysfunction were examined by multichannel videourodynamics and perineal ultrasound, both supine and standing. Ultrasound included color Doppler imaging when available, i.e. in a subgroup of 99 patients. The position of the bladder neck at rest was higher in the supine position (P