Elevated cyclic AMP suppresses ConA-induced MT1-MMP expression in MDA- MB-231 human breast cancer cells

We have previously reported that induction of MMP-2 activation by Concanavalin A (ConA) in MDA-MB-231 human breast cancer cells involves both transcriptional and post-transcriptional mechanisms, and that the continuous presence of ConA is required for MMP-2 activation (Yu et al. Cancer Res, 55, 3272-7, 1995). In an effort to identify signal transduction pathways which may either contribute to or modulate this mechanism, we found that three different cAMP-inducing agents, cholera toxin (CT), forskolin (FSK), and 3- isobutyl-1-methylxanthine (IBMX) partially inhibited ConA-induced MT1-MMP expression and MMP-2 activation in MDA-MB-231 cells. Combinations of CT or FSK with IBMX exhibited additive effects on reduction of MT1-MMP mRNA expression and MMP-2 activation. Agents which increase cAMP levels appeared to target transcriptional aspects of ConA induction, reducing MT1-MMP mRNA and protein in parallel with the reduced MMP-2 activation. In the absence of ConA, down-regulation of constitutive production of MT1-MMP mRNA and protein was observed, indicating that cAMP acts independently of ConA. These observations may help to elucidate factors regulating MT1-MMP expression, which may be pivotal to the elaboration of invasive machinery on the cell surface.

Sphingolipid distribution changes with age in the human lens

The formation of an internal barrier to the diffusion of small molecules in the lens during middle age is hypothesized to be a key event in the development of age-related nuclear (ARN) cataract. Changes in membrane lipids with age may be responsible. In this study, we investigated the effect of age on the distribution of sphingomyelins, the most abundant lens phospholipids. Human lens sections were initially analyzed by MALDI mass spectrometry imaging. A distinct annular distribution of the dihydrosphingomyelin, DHSM (d18:0/16:0), in the barrier region was observed in 64- and 70-year-old lenses but not in a 23-year-old lens. An increase in the dihydroceramide, DHCer (d18:0/16:0), in the lens nucleus was also observed in the older lenses. These findings were supported by ESI mass spectrometry analysis of lipid extracts from lenses dissected into outer, barrier, and nuclear regions. A subsequent analysis of 18 lenses ages 20-72 years revealed that sphingomyelin levels increased with age in the barrier region until reaching a plateau at approximately 40 years of age. Such changes in lipid composition will have a significant impact on the physical properties of the fiber cell membranes and may be associated with the formation of a barrier.-Deeley, J. M., J. A. Hankin, M. G. Friedrich, R. C. Murphy, R. J. W. Truscott, T. W. Mitchell, and S. J. Blanksby. Sphingolipid distribution changes with age in the human lens. J. Lipid Res. 2010. 51: 2753-2760.

Human lens lipids differ markedly from those of commonly used experimental animals

Electrospray ionisation tandem mass spectrometry has allowed the unambiguous identification and quantification of individual lens phospholipids in human and six animal models. Using this approach ca. 100 unique phospholipids have been characterised. Parallel analysis of the same lens extracts by a novel direct-insertion electron-ionization technique found the cholesterol content of human lenses to be significantly higher (ca. 6 times) than lenses from the other animals. The most abundant phospholipids in all the lenses examined were choline-containing phospholipids. In rat, mouse, sheep, cow, pig and chicken, these were present largely as phosphatidylcholines, in contrast 66% of the total phospholipid in Homo sapiens was sphingomyelin, with the most abundant being dihydrosphingomyelins, in particular SM(d18:0/16:0) and SM(d18:0/24:1). The abundant glycerophospholipids within human lenses were found to be predominantly phosphatidylethanolamines and phosphatidylserines with surprisingly high concentrations of ether-linked alkyl chains identified in both classes. This study is the first to identify the phospholipid class (head-group) and assign the constituent fatty acid(s) for each lipid molecule and to quantify individual lens phospholipids using internal standards. These data clearly indicate marked differences in the membrane lipid composition of the human lens compared to commonly used animal models and thus predict a significant variation in the membrane properties of human lens fibre cells compared to those of other animals. © 2008 Elsevier B.V. All rights reserved.

Vimentin and epithelial-mesenchymal transition in human breast cancer : observations in vitro and in vivo

Breast cancer is a highly prevalent disease among women worldwide. While the expression of certain proteins within these tumours is used for prognosis and selection of therapies, there is a continuing need for additional markers to be identified. A considerable amount of current literature, based predominantly on cell culture systems, suggests that a major mechanism responsible for the progression of breast cancer is due to tumour cells losing their epithelial features and gaining mesenchymal properties. These events are proposed to be very similar to the epithelial-mesenchymal transition (EMT) process that has been well characterised in embryonic development. For the developmental and putative cancer EMT, the cell intermediate filament status changes from a keratin-rich network which connects to adherens junctions and hemidesmosomes, to a vimentin-rich network connecting to focal adhesions. This review summarises observations of vimentin expression in breast cancer model systems, and discusses the potential role of EMT in human breast cancer progression, and the prognostic usefulness of vimentin expression.

The use of uniaxial and triaxial accelerometers to measure children's "free-play" physical activity

In order to effectively measure the physical activity of children, objective monitoring devices must be able to quantify the intermittent and nonlinear movement of free play. The purpose of this study was to investigate the validity of the Computer Science and Applications (CSA) uniaxial accelerometer and the TriTrac-R3D triaxial accelerometer with respect to their ability to measure 8 "free-play" activities of different intensity. The activities ranged from light to very vigorous in intensity and included activities such as throwing and catching, hopscotch, and basketball. Twenty-eight children, ages 9 to 11, wore a CSA and a heart rate monitor while performing the activities. Sixteen children also wore a Tritrac. Counts from the CSA, Tritrac, and heart rates corresponding to the last 3 min of the 5 min spent at each activity were averaged and used in correlation analyses. Across all 8 activities, Tritrac counts were significantly correlated with predicted MET level (r= 0.69) and heart rate (r= 0.73). Correlations between CSA output, predicted MET level (0.43), and heart rate (0.64) were also significant but were lower than those observed for the Tritrac. These data indicate that accelerometers are an appropriate methodology for measuring children's free-play physical activities.

Calibration of the biotrainer pro activity monitor in children

The unique physical and movement characteristics of children necessitate the development of accelerometer equations and cut points that are population specific. The purpose of this study is to develop an ecologically valid cut point for the Biotrainer Pro monitor that reflects a threshold for moderate-intensity physical activity in elementary school children. A sample of 30 children (ages 8-12) wore a Biotrainer monitor while completing a series of 7 movement tasks (calibration phase) and while participating in an organized group activity (cross-validation phase). Videotapes from each session were processed using a computerized direct-observation technique to provide a criterion measure of physical activity. Analyses involved the use of mixed-model regression and receiver operator characteristic (ROC) curves. The results indicated that a cut point of 4 counts/min provides the optimal balance between the related needs for sensitivity (accurately detecting activity) and specificity (limiting misclassification of activity as inactivity). Results with the cross-validation data demonstrated that this value yielded the best overall kappa (.58) and a high classification agreement (84%) for activity determination. The specificity of 93% demonstrates that the proposed cut point can accurately detect activity; however, the lower sensitivity value of 61% suggests that some minutes of activity might be incorrectly classified as inactivity. The cut point of 4 counts/min provides an ecologically valid cut point to capture physical activity in children using the Biotrainer Pro activity monitor.

Validity of the previous day physical activity recall (PDPAR) in fifth-grade children

This study evaluated the validity of the Previous Day Physical Activity Recall (PDPAR) self-report instrument in quantifying after-school physical activity behavior in fifth-grade children. Thirty-eight fifth-grade students (mean age, 10.8 +/- 0.1; 52.6% female; 26.3% African American) from two urban elementary schools completed the PDPAR after wearing a CSA WAM 7164 accelerometer for a day. The mean within-subject correlation between self-reported MET level and total counts for each 30-min block was 0.57 (95% C.I., 0.51-0.62). Self-reported mean MET level during the after-school period and the number of 30-min blocks with activity rated at greater than or equal to 6 METs were significantly correlated with the CSA outcome variables. Validity coefficients for these variables ranged from 0.35 to 0.43 (p <.05). Correlations between the number of 30-min blocks with activity rated at greater than or equal to 3 METs and the CSA variables were positive but failed to reach statistical significance (r = 0.19-0.23). The PDPAR provides moderately valid estimates of relative participation in vigorous activity and mean MET level in fifth-grade children. Caution should be exercised when using the PDPAR to quantify moderate physical activity in preadolescent children.

The LCC15-MB human breast cancer cell line expresses osteopontin and exhibits an invasive and metastatic phenotype

We have characterized the LCC15-MB cell line which was recently derived from a breast carcinoma metastasis resected from the femur of a 29-year-old woman. LCC15-MB cells are vimentin (VIM) positive, exhibit a stellate morphology in routine cell culture, and form penetrating colonies when embedded in three-dimensional gels of Matrigel or fibrillar collagen. They show high levels of activity in the Boyden chamber chemomigration and chemoinvasion assays, and like other invasive human breast cancer (HBC) cell lines, LCC15-MB cells activate matrix-metalloproteinase-2 in response to treatment with concanavalin A. In addition, these cells are tumorigenic when implanted subcutaneously in nude mice and recolonize bone after arterial injection. Interestingly, both the primary lesion and the bone metastasis from which LCC15-MB were derived, as well as the resultant cell line, abundantly express the bone matrix protein osteopontin (OPN). OPN is also expressed by the highly metastatic MDA-MB-435 cells, but not other invasive or noninvasive HBC cell lines. Expression of OPN is retained in the subcutaneous xenograft and intraosseous metastases of LCC15-MB as detected by immunohistochemistry. Both VIM and OPN expression have been associated with breast cancer invasion and metastasis, and their expression by the LCC15-MB cell line is consistent with its derivation from a highly aggressive breast cancer. These cells provide a useful model for studying molecular mechanisms important for breast cancer metastasis to bone and, in particular, the implication(s) of OPN and VIM expression in this process.

Involvement of focal adhesion kinase in inhibition of motility of human breast cancer cells by sphingosine 1-phosphate

Sphingosine 1-phosphate (SPP), a bioactive sphingolipid metabolite, inhibits chemoinvasiveness of the aggressive, estrogen-independent MDA-MB-231 human breast cancer cell line. As in many other cell types, SPP stimulated proliferation of MDA-MB-231 cells, albeit to a lesser extent. Treatment of MDA-MB-231 cells with SPP had no significant effect on their adhesiveness to Matrigel, and only high concentrations of SPP partially inhibited matrix metalloproteinase-2 activation induced by Con A. However, SPP at a concentration that strongly inhibited invasiveness also markedly reduced chemotactic motility. To investigate the molecular mechanisms by which SPP interferes with cell motility, we examined tyrosine phosphorylation of focal adhesion kinase (FAK) and paxillin, which are important for organization of focal adhesions and cell motility. SPP rapidly increased tyrosine phosphorylation of FAK and paxillin and of the paxillin-associated protein Crk. Overexpression of FAK and kinase-defective FAK in MDA-MB-231 cells resulted in a slight increase in motility without affecting the inhibitory effect of SPP, whereas expression of FAK with a mutation of the major autophosphorylation site (F397) abolished the inhibitory effect of SPP on cell motility. In contrast, the phosphoinositide 3'-kinase inhibitor, wortmannin, inhibited chemotactic motility in both vector and FAK-F397- transfected cells. Our results suggest that autophosphorylation of FAK on Y397 may play an important role in SPP signaling leading to decreased cell motility.

Human trafficking involving marriage and partner migration to Australia

In this report, what is known about human trafficking involving marriage and partner migration to Australia is described, drawing on primary information obtained from victim/survivor testimonies, stakeholder knowledge and expertise, and reported cases that progressed through the Australian justice system. It confirms what some stakeholders in the human trafficking area have long suspected—that marriage and partner migration have been used to facilitate the trafficking of people into Australia.

Towards the development of 1-to-n human machine interfaces for unmanned aerial vehicles

Multi-touch interfaces across a wide range of hardware platforms are becoming pervasive. This is due to the adoption of smart phones and tablets in both the consumer and corporate market place. This paper proposes a human-machine interface to interact with unmanned aerial systems based on the philosophy of multi-touch hardware-independent high-level interaction with multiple systems simultaneously. Our approach incorporates emerging development methods for multi-touch interfaces on mobile platforms. A framework is defined for supporting multiple protocols. An open source solution is presented that demonstrates: architecture supporting different communications hardware; an extensible approach for supporting multiple protocols; and the ability to monitor and interact with multiple UAVs from multiple clients simultaneously. Validation tests were conducted to assess the performance, scalability and impact on packet latency under different client configurations.