Solid phase red cell adherence immunoassay for anti-HIV 1: a simple, rapid, and accurate method for donor screening

In technically developed countries in which acquired immunodeficiency syndrome is a risk to the recipients of blood or tissue, it is mandatory to screen the donor for evidence of HIV (human immunodeficiency virus) infection. Current tests, based on enzyme-linked immunoassay, are time-consuming and expensive and as such are unsuitable for developing countries. We describe a second generation test using anti-human IgG coupled to red cells as the indicator of antibody having reacted with test antigen (1). The test is complete within ten minutes, simple to perform and to read and has 100% sensitivity and 99% specificity compared with Western blot. It is ideal for the rapid screening of organ donors and for the screening of blood donors where cost is a major consideration.     

Ovarian metastases: computed tomographic appearances

Computed tomographic scans of 34 patients with ovarian metastases were reviewed to assess the radiographic appearances and to correlate these with the primary neoplasms. Primary neoplasms were located in the colon (20 patients), breast (six), stomach (five), small bowel (one), bladder (one), and Wilms tumor of the kidney (one). The radiographic appearance of the metastatic lesions could be described as predominantly cystic (14 lesions), mixed (12 lesions), or solid (seven lesions). There was one false-negative examination which showed no ovarian enlargement, although neoplastic deposits were found on the ovary at laparotomy. The cystic and mixed lesions tended to be larger in overall diameter than the solid. The metastases from gastric carcinoma appeared solid in four of five cases. The metastases from the other neoplasms had variable appearances simulating primary ovarian carcinoma.     

Constitutive expression of platelet glycoproteins by the human leukemia cell line K562

The human leukemia cell line K562 was derived from a patient with chronic granulocytic leukemia. This cell line has subsequently been shown to possess phenotypic markers typical of erythroid and myeloid cells. Using a rabbit antiserum directed against purified platelet glycoproteins (PGPs), we have obtained evidence for the constitutive expression of PGPs on the surface of K562 cells. PGPs expressed have been tentatively identified as IIa and III based on their apparent migration in a 7% sodium dodecylsulfate polyacrylamide gel. K562 may become an important tool for the study of early events involved in megakaryocytic differentiation.     

Cigarette smoke exposure up-regulates endothelin receptor B in human pulmonary artery endothelial cells: molecular and functional consequences

BACKGROUND AND PURPOSE

Pulmonary arteries from smokers and chronic obstructive pulmonary disease patients show abnormal endothelium-dependent vascular reactivity. We studied the effect of cigarette smoke extract (CSE) on endothelin receptor B (ET_B) expression in human pulmonary artery endothelial cells (HPAECs) and its role in endothelial dysfunction.

EXPERIMENTAL APPROACH

ET_B receptor expression was measured by real time RT-PCR, Western blot and immunofluorescence. Cell contraction, intracellular Ca²⁺, F/G-actin, RhoA activity, myosin light chain phosphorylation, ET, NO, thromboxane (Tx)A₂ and reactive oxygen species (ROS) were measured by traction microscopy, fluorescence microscopy, phalloidin fluorescence, colorimetric assay, Western blot, elisa and DCFDA fluorescence respectively.

KEY RESULTS

Cigarette smoke extract dose-dependently increased ET_B receptor expression in HPAECs after 24 h incubation. CSE-induced ET_B expression was attenuated by bosentan, the ET_B receptor antagonist BQ788, the Rho kinase antagonist Y27632 and the antioxidant N-acetylcysteine. A monoclonal antibody to ET-1 prevented CSE-induced ET_B receptor overexpression. Twenty-four hour exposure to ET-1 dose-dependently increased ET_B receptor expression, mimicking the effect of CSE. CSE-induced ET_B receptor overexpression caused greater cell contraction; increased intracellular Ca²⁺; increased F/G-actin and RhoA activity; increased myosin light chain phosphorylation; augmented TxA₂ and ROS production; and decreased NO after acute ET-1 (10 nM). These effects were attenuated by bosentan, BQ788, Y27632 and N-acetylcysteine.

CONCLUSIONS AND IMPLICATION

Cigarette smoke extract induced ET_B receptor overexpression by a feed forward mechanism mediated partly by ET release, promoting HPAEC dysfunction and attenuated by ET_B receptor blockade, Rho kinase and ROS inhibition. These results provide support for the use of bosentan in CS-related endothelial dysfunction.     

Induction of apoptosis by the ginsenoside Rh2 by internalization of lipid rafts and caveolae and inactivation of Akt

Background and purpose:

Lipid rafts and caveolae are membrane microdomains with important roles in cell survival signalling involving the Akt pathway. Cholesterol is important for the structure and function of these microdomains. The ginsenoside Rh2 exhibits anti-tumour activity. Because Rh2 is structurally similar to cholesterol, we investigated the possibility that Rh2 exerted its anti-tumour effect by modulating rafts and caveolae.

Experimental approach:

A431 cells (human epidermoid carcinoma cell line) were treated with Rh2 and the effects on cell apoptosis, raft localization and Akt activation measured. We also examined the effects of over-expression of Akt and active-Akt on Rh2-induced cell death.

Key results:

Rh2 induced apoptosis concentration- and time-dependently. Rh2 reduced the levels of rafts and caveolae in the plasma membrane and increased their internalization. Furthermore, Akt activity was decreased and consequently, Akt-dependent phosphorylation of Bad, a pro-survival protein, was decreased whereas the pro-apoptotic proteins, Bim and Bax, were increased upon Rh2 treatment. Unlike microdomain internalization induce by cholesterol depletion, Rh2-mediated internalization of rafts and caveolae was not reversed by cholesterol addition. Also, cholesterol addition did not restore Akt activation or rescue cells from Rh2-induced cell death. Rh2-induced cell death was attenuated in MDA-MB-231 cells over-expressing either wild-type or dominant-active Akt.

Conclusions and implications:

Rh2 induced internalization of rafts and caveolae, leading to Akt inactivation, and ultimately apoptosis. Because elevated levels of membrane rafts and caveolae, and Akt activation have been correlated with cancer development, internalization of these microdomains by Rh2 could potentially be used as an anti-cancer therapy.