Cytokeratins of Tumorigenic and Highly Malignant Respiratory Tract Epithelial Cells

In malignant airway epithelial cells, structural abnormalities were evident from the cytokeratin organization. To determine whether the cytokeratins themselves were responsible, an in vitro model for bronchogenic carcinoma, consisting of three highly malignant lines and three less tumorigenic lines, was studied. Cytokeratins were evaluated by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). When typical constraints on tumors were relieved by in vitro culture, lines showed profiles resembling normal, primary cells. The CK5/CK14 combination, characteristic of basal epithelial layers, was represented by CK6A/CK14. CK17 was invariably present, while CK5, CK7, CK8, CK19, and CK42 content varied. CK19 appeared to substitute for the rarely observed CK18. While lacking the common CK8/CK18 combination of hyperproliferative cells, an invasive, metastasizing line had CK6A/CK7 or CK8 with CK19 suggesting derivation similar to adenocarcinomas. Bands of CK19 and actin migrated to higher pI in tumorigenic and malignant lines than in normal cells. Ubiquitinated acidic cytokeratins with a low isoelectric point (pI) and high molecular weight (MW) showed no consistent differences in lines that differed in growth potential. Type II made up 49–52% of total cytokeratins in nonmalignant lines, whereas highly malignant lines showed lower levels. Posttranslational modifications were identified but could not explain the shortfall of basic cytokeratins

Cell Edge Features Affected by Microtubule Inhibitor Combinations

The tumor promoter, phorbol 12-myristate 13-acetate (PMA), enhances tumor yield through an epigenetic mechanism. PMA, like another promoter, phosphatase inhibitor okadaic acid, works by maintaining proteins in a phosphorylated state. In order to identify chemicals with promoter and antipromoter effects, this laboratory has developed a standard curve of morphogenetic changes using data from precancerous cell lines that eventually became neoplastic. Using the curve as a basis of comparison, we defined the “signature” phenotype as that adopted when a cell line became neoplastic. The results of solving for signature type disclosed that the microtubule-depolymerizing compound, colchicine, had a promoter-like effect [2]. The opposite effect was found if cells were exposed to paclitaxel (Taxol®) and colchicine [3]. Such findings gave clinicians a rationale to test microtubule inhibitor combinations [4]. Therapy is only effective when agents are administered nearly simultaneously. In the current work, we study the effect of inhibitor combinations on cell features defined by computer-assisted microscopy and classification methods based on latent factors

Activated Protein Kinase C (PKC) Is Persistently Trafficked with Epidermal Growth Factor (EGF) Receptor

Protein kinase Cs (PKCs) are activated by lipids in the plasma membrane and bind to a scaffold assembled on the epidermal growth factor (EGF) receptor (EGFR). Understanding how this complex is routed is important, because this determines whether EGFR is degraded, terminating signaling. Here, cells were preincubated in EGF-tagged gold nanoparticles, then allowed to internalize them in the presence or absence of a phorbol ester PKC activator. PKC colocalized with EGF-tagged nanoparticles within 5 min and migrated with EGFR-bearing vesicles into the cell. Two conformations of PKC-epsilon were distinguished by different primary antibodies. One, thought to be enzymatically active, was on endosomes and displayed a binding site for antibody RR (R&D). The other, recognized by Genetex green (GG), was soluble, on actin-rich structures, and loosely bound to vesicles. During a 15-min chase, EGF-tagged nanoparticles entered large, perinuclear structures. In phorbol ester-treated cells, vesicles bearing EGF-tagged nanoparticles tended to enter this endocytic recycling compartment (ERC) without the GG form. The correlation coefficient between the GG (inactive) and RR conformations on vesicles was also lower. Thus, active PKC has a Charon-like function, ferrying vesicles to the ERC, and inactivation counteracts this function. The advantage conferred on cells by aggregating vesicles in the ERC is unclear

Unraveling the Determinants of Protrusion Formation

A computerized morphometric classification technique based on latent factors reveals major protrusion classes: factors 4, 5, and 7. Previous work showed that factor 4 represented filopodia, 5 the distribution of lamellar cytoplasm, and 7 a blunt protrusion. We explore the relationship of focal contact (FC) characteristics and their integrated actin cables to factors values. The results show that FC maturation/cytoskeletal integration affects factor 5, because FC elongation/integration was correlated with its values. On the contrary, 7 values decreased with maturation, so cable or FC size or their integration must be restricted to form these protrusions. Where integration did occur, the cables showed distinctive size and orientation, as indicated by correlation of 7 values with FC shape. Results obtained with myosin inhibitors support the interpretation that a central, isometric, contractile network puts constraints on both factor 5 and 7 protrusions. We conclude that cells establish functional domains by rearranging the cytoskeleton

Emission and Absorption Properties of Low-Mass Type 2 Active Galaxies with XMM-Newton

We present XMM-Newton observations of four low-redshift Seyfert galaxies selected to have low host luminosities (M_g>-20 mag) and small stellar velocity dispersions (sigma_star<45 km/s), which are among the smallest stellar velocity dispersions found in any active galaxies. These galaxies show weak or no broad optical emission lines and have likely black hole masses <10^6 M_sun. Three out of four objects were detected with >3sigma significance in ~25 ks exposures and two observations had high enough signal-to-noise ratios for rudimentary spectral analysis. We calculate hardness ratios (-0.43 to 0.01) for the three detected objects and use them to estimate photon indices in the range of Gamma=1.1-1.8. Relative to [OIII], the type 2 objects are X-ray faint in comparison with Seyfert 1 galaxies, suggesting that the central engines are obscured. We estimate the intrinsic absorption of each object under the assumption that the [OIII] emission line luminosities are correlated with the unabsorbed X-ray luminosity. The results are consistent with moderate (N_H~10^22 cm^-2) absorption over the Galactic values in three of the four objects, which might explain the non-detection of broad-line emission in optical spectra. One object in our sample, SDSS J110912.40+612346.7, is a near identical type 2 counterpart of the late-type Seyfert 1 galaxy NGC 4395. While the two objects have very similar [OIII] luminosities, the type 2 object has an X-ray/[OIII] flux ratio nearly an order of magnitude lower than NGC 4395. The most plausible explanation for this difference is absorption of the primary X-ray continuum of the type 2 object, providing an indication that obscuration-based unified models of active galaxies can apply even at the lowest luminosities seen among Seyfert nuclei, down to L_bol~10^40-10^41 erg/s.Comment: 5 figures, 3 tables, accepted for publication in Ap

Pattern Analysis of Microtubule-Polymerizing and -Depolymerizing Agent Combinations as Cancer Chemotherapies

Subcellular distribution of mass can be analyzed by a technique that involves culturing cells on interferometers and digitizing their interference contours. Contour sampling resulted in 102 variables per cell, which were predictors of oncogenic transformation. Cell phenotypes can be deconstructed by use of latent factors, which represent the covariance of the real variables. The reversal of the cancertype phenotype by a combination of microtubule- stabilizing and -depolymerizing agents was described previously. The implications of these results have been explored by clinicians who treated patients with the combination of docetaxel and vinorelbine (Navelbine®). The current study was performed to determine the effects of different combinations on phenotype and in phases of the cell cycle other than mitosis. Combinations of paclitaxel with either colchicine, podophyllotoxin, nocodazole, or vinblastine caused phenotype reversal. Paclitaxel analogue, 7-deoxytaxol, by itself caused reversal. Factors #4, (filopodia), #5 (displacement and/or deep invaginations in the periphery), #8, and #12 took on values typical of normal cells, whereas the values of #7 (p21-activated kinase), and #13 (rounding up) shifted toward the cancer-type. All combinations altered microtubule arrangement at the cell edge. Delivery schedules and drug ratios used in clinical studies were subjected to analysis. Clinical response rates were better when the combination was not interspersed with a single agent (P=0.004). The results support the idea that efficacy depends upon simultaneous exposure to both agents, and suggest a novel mechanism for combination therapies. These therapies appear to restore in transformed cells some of the features of a contact-inhibited cell, and to impede progress through the cell cycle even when provided at nanomolar concentrations

ALMA detection of a disc-dominated [C II] emission line at z = 4.6 in the luminous QSO J1554+1937

We present observations and analysis of an unusual [C ii] emission line in the very luminous quasi-stellar object (QSO) SDSS J155426.16+193703.0 at z ~ 4.6. The line is extremely broad (full width at half-maximum 735 km s−1) and seems to have a flat-topped or double-peaked line profile. A velocity map of the line shows a gradient across the source that indicates large-scale rotation of star-forming gas. Together, the velocity map and line profile suggest the presence of a massive rotating disc with a dynamical mass Mdyn ≳5×1010 M☉. Using the assumption of a rotating disc origin, we employ an empirical relation between galaxy disc circular velocity and bulge velocity dispersion (σ) to estimate that σ > 310 km s−1, subject to a correction for the unknown disc inclination. This result implies that this source is consistent with the local M–σ relation, or offset at most by an order of magnitude in black hole mass. In contrast, the assumption of a bulge origin for the [C ii] emission line would lead to a conclusion that the black hole is nearly two orders of magnitude more massive than predicted by the M–σ relation, similar to previous findings for other high-redshift QSOs. As disc rotation may be a common origin for [C ii] emission at high redshifts, these results stress that careful consideration of dynamical origins is required when using observations of this line to derive properties of high-redshift galaxies

Putting episodic disability into context: a qualitative study exploring factors that influence disability experienced by adults living with HIV/AIDS

<p>Abstract</p> <p>Background</p> <p>An increasing number of individuals may be living with the health-related consequences of HIV and its associated treatments, a concept we term disability. However, the context in which disability is experienced from the HIV perspective is not well understood. The purpose of this paper is to describe the contextual factors that influence the experiences of disability from the perspective of adults living with HIV.</p> <p>Methods</p> <p>We conducted four focus groups and 15 face-to-face interviews with 38 men and women living with HIV. We asked participants to describe their health-related challenges, the physical, social and psychological areas of their life affected, and the impact of these challenges on their overall health. We also conducted two validity check focus groups with seven returning participants. We analyzed data using grounded theory techniques to develop a conceptual framework of disability for adults living with HIV, called the Episodic Disability Framework.</p> <p>Results</p> <p>Contextual factors that influenced disability were integral to participants' experiences and emerged as a key component of the framework. Extrinsic contextual factors included social support (support from friends, family, partners, pets and community, support from health care services and personnel, and programme and policy support) and stigma. Intrinsic contextual factors included living strategies (seeking social interaction with others, maintaining a sense of control over life and the illness, "blocking HIV out of the mind", and adopting attitudes and beliefs to help manage living with HIV) and personal attributes (gender and aging). These factors may exacerbate or alleviate dimensions of HIV disability.</p> <p>Conclusion</p> <p>This framework is the first to consider the contextual factors that influence experiences of disability from the perspective of adults living with HIV. Extrinsic factors (level of social support and stigma) and intrinsic factors (living strategies and personal attributes) may exacerbate or alleviate episodes of HIV-related disability. These factors offer a broader understanding of the disability experience and may suggest ways to prevent or reduce disability for adults living with HIV.</p