It is becoming more and more apparent that monolayer cultures of tumor cells cannot completely represent the characteristics of three-dimensional solid tumors. Consequently, the multicellular tumor spheroid model, which is of intermediate complexity between in vivo tumors and monolayer cultures, was developed. In this review, the major similarities between spheroids and solid tumors are discussed. After a brief survey of the different spheroid culturing techniques, the general morphological and growth characteristics of these systems are examined and compared to solid tumors. Finally, selected studies regarding the use of tumor spheroids to examine cell response to antineoplastic agents and radiation, cell death including both necrosis and apoptosis and cell adhesion in spheroids are reviewed.

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Although apoptosis is considered one of the major mechanisms of CD4+ T cell depletion in HIV-infected patients, the virus-infected cells somehow appear to be protected from apoptosis, which generally occurs in bystander cells. Vpr is an auxiliary HIV-1 protein, which, unlike the other regulatory gene products, is present at high copy number in virus particles. We established stable transfectants of CD4+ T Jurkat cells constitutively expressing low levels of vpr. These clones exhibited cell cycle characteristics similar to those of control-transfected cells. Treatment of control clones with apoptotic stimuli (i.e., cycloheximide/tumor necrosis factor α (TNF-α), anti-Fas antibody, or serum starvation) resulted in a massive cell death by apoptosis. In contrast, all the vpr-expressing clones showed an impressive protection from apoptosis independently of the inducer. Notably, vpr antisense phosphorothioate oligodeoxynucleotides render vpr-expressing cells as susceptible to apoptosis induced by cycloheximide and TNF-α as the control clones. Moreover, the constitutive expression of HIV-1 vpr resulted in the upregulation of bcl-2, an oncogene endowed with antiapoptotic activities, and in the downmodulation of bax, a proapoptotic factor of the bcl-2 family. Altogether, these results suggest that low levels of the endogenous vpr protein can interfere with the physiological turnover of T lymphocytes at early stages of virus infection, thus facilitating HIV persistence and, subsequently, viral spread. This might explain why apoptosis mostly occurs in bystander uninfected cells in AIDS patients.

A specific neuronal vulnerability to amyloid protein toxicity may account for brain susceptibility to protein misfolding diseases. To investigate this issue, we compared the effects induced by oligomers from salmon calcitonin (sCTOs), a neurotoxic amyloid protein, on cells of different histogenesis: mature and immature primary hippocampal neurons, primary astrocytes, MG63 osteoblasts and NIH-3T3 fibroblasts. In mature neurons, sCTOs increased apoptosis and induced neuritic and synaptic damages similar to those caused by amyloid beta oligomers. Immature neurons and the other cell types showed no cytotoxicity. sCTOs caused cytosolic Ca(2+) rise in mature, but not in immature neurons and the other cell types. Comparison of plasma membrane lipid composition showed that mature neurons had the highest content in lipid rafts, suggesting a key role for them in neuronal vulnerability to sCTOs. Consistently, depletion in gangliosides protected against sCTO toxicity. We hypothesize that the high content in lipid rafts makes mature neurons especially vulnerable to amyloid proteins, as compared to other cell types; this may help explain why the brain is a target organ for amyloid-related diseases.

The pathophysiological role of the adenosine A 3 receptor in the central nervous system is largely unknown. We have investigated the effects of the selective A 3 receptor agonist 2-chloro-N 6 -(3-iodobenzyl)-adenosine, Cl-IB-MECA, in cells of the astroglial lineage (human astrocytoma ADF cells). A marked reorganization of the cytoskeleton, with appearance of stress fibers and numerous cell protrusions, was found following exposure of cells to low (nM) concentrations of Cl-IB-MECA. These "trophic" effects were accompanied by induction of the expression of Rho, a small GTP-binding protein, which was virtually absent in control cells, and by changes of the intracellular distribution of the antiapoptotic protein Bcl-x L , that, in agonist-exposed cells, became specifically associated to cell protrusions. This is the first demonstration that the intracellular organization of Bcl-x L can be modulated by the activation of a G-protein-coupled membrane receptor, such as the A 3 adenosine receptor. Moreover, modulation of the astrocytic cytoskeleton by adenosine may have intriguing implications in both nervous system development and in the response of the brain to trauma and ischemia.Adenosine regulates various physiological functions through four distinct G-protein-coupled receptors (the A 1 , A 2A , A 2B and A 3 subtypes) (1) and participates in physiological neurotransmission (2). Activation of the A 1 receptor results in potent neuroprotection (2), white the A 2 receptor (particularly the A 2A subtype) has been implicated in regulation of motor functions (1, 2). On the other hand, very little is known about the A 3 receptor, which has a unique structure-activity relationship profile, tissue distribution and effector coupling (3).Insights into the possible pathophysiological role of the A 3 receptor have recently come from the use of the first really selective A 3 receptor agonists, such as N 6 -(3-iodobenzyl)-adenosine-5′-N-methyluronamide (IB-MECA, which is 50-fold selective for A 3 versus A 1 or A 2A receptor) (4) and its derivative 2-chloro-IB-MECA (Cl-IB-MECA), which shows a selectivity for the A 3 versus A 1 and A 2A receptors of 2500-and 1400-fold, respectively (5). Thanks to these selective compounds, we now know that the A 3 receptor may play a role in inflammation (6), hypotension (7), mast cell degranulation (8), and, most strikingly, in regulation of cell survival, by promoting both cell protection and cell death, depending upon the cell type and the agonist concentration (9-12). Protective effects against ischemia/ hypoxia are induced by low (nM) concentrations of A 3 receptor agonists in both cardiac myocytes and in brain (13,14). The molecular mechanisms at the basis of these effects are totally unknown. In this study, we have investigated the effects of nM Cl-IB-MECA concentrations on human cells of the astroglial lineage, a brain cell type which plays a key role in both brain development and in brain repair and remodelling following trauma and ischemia (15). In particular, based on the hypothes...

Because of their particular architectural characteristics, multicellular spheroids are demonstrated to be extremely useful in testing radiotherapeutic protocols, including dose rate and fractionation, radioimmunotherapy and the effects of combined treatments (e.g. radiation and anti-neoplastic drugs). Further studies should seek not only to continue testing these protocols, but also to investigate the more fundamental questions of radiation-induced apoptotic cell death, cell-cycle events, cell-cell interactions and cell adhesion phenomena.

The literature indicates that there is still no general agreement on the exact biological detrimental effects of ELF fields, on the physical mechanisms that may be behind these effects or on the extent to which these effects may be harmful to humans. Nonetheless, the majority of the in vitro experimental results indicate that ELF fields induce numerous types of changes in cells. Whether or not the perturbations observed at the cellular level can be directly extrapolated to negative effects in humans is still unknown. However, the myriad of effects that ELF fields have on biological systems should not be ignored when evaluating risk to humans from these fields and, consequently, in passing appropriate legislation to safeguard both the general public and professionally-exposed workers. With regard to the positive effects of these fields, the possibility of testing further their efficacy in therapeutic protocols should also not be overlooked.