Prospects of RNA interference therapy for cancer.

RNA interference (RNAi) is a powerful gene-silencing process that holds great promise in the field of cancer therapy. The discovery of RNAi has generated enthusiasm within the scientific community, not only because it has been used to rapidly identify key molecules involved in many disease processes including cancer, but also because RNAi has the potential to be translated into a technology with major therapeutic applications. Our evolving understanding of the molecular pathways important for carcinogenesis has created opportunities for cancer therapy employing RNAi technology to target the key molecules within these pathways. Many gene products involved in carcinogenesis have already been explored as targets for RNAi intervention, and RNAi targeting of molecules crucial for tumor-host interactions and tumor resistance to chemo- or radiotherapy has also been investigated. In most of these studies, the silencing of critical gene products by RNAi technology has generated significant antiproliferative and/or proapoptotic effects in cell-culture systems or in preclinical animal models. Nevertheless, significant obstacles, such as in vivo delivery, incomplete suppression of target genes, nonspecific immune responses and the so-called off-target effects, need to be overcome before this technology can be successfully translated into the clinical arena. Significant progress has already been made in addressing some of these issues, and it is foreseen that early phase clinical trials will be initiated in the very near future.

Endothelial cells potentiate phagocytic killing by macrophages via platelet-activating factor release.

The immunomodulatory function of endothelial cells (EC) includes the initiation of leukocyte margination, diapedesis, and activation through the upregulation of various cell surface-associated molecules. However, the effect that EC have on the phagocytic function of neighboring monocytes and macrophages is less well described. To address this issue, microvascular EC were cocultured with murine peritoneal macrophages, first in direct contact, then in a noncontact coculture system, and macrophage phagocytosis and phagocytic killing were assessed. The presence of increasing concentrations of EC resulted in a dose-dependent increase in macrophage phagocytic killing. This stimulatory effect was inhibited in a dose-dependent manner by the pretreatment of macrophage/EC cocultures with WEB-2086 or CV-6209, specific platelet-activating factor (PAF)-receptor antagonists, but not by anti-tumor necrosis factor-alpha, anti-interleukin (IL)-1alpha, or anti-IL-1beta. Furthermore, the effect was reproduced in the absence of EC by the exogenous administration of nanomolar concentrations of PAF. Microvascular EC potentiate macrophage phagocytic killing via the release of a soluble signal; PAF appears to be an important component of that signal.

Use of leukocyte depletion to decrease injury after lung preservation and rewarming ischemia: an experimental model.

BACKGROUND: Hypothermia is critical for proper lung preservation. Ideally, the lungs should be maintained at the optimal preservation temperature during the entire ischemic interval. Lung rewarming during implantation is commonly observed. This study was undertaken to investigate the severity of rewarming ischemia on preservation injury and the possibility of minimizing this by use of leukocyte depletion during initial reperfusion. METHODS: Four experimental groups were tested as follows: neonatal piglet heart-lung blocks were either (1) placed on an isolated, blood-perfused, working heart-lung circuit without intervening ischemia (control, n = 6), (2) reperfused on the circuit with whole blood (WB, n = 6) after 13 hours of preservation, (3) reperfused with WB after 12 hours of preservation and 1 hour of rewarming (RWB, n = 5), or (4) reperfused with leukocyte-depleted blood for an initial 10 minutes followed by WB, after 12 hours of preservation and 1 hour of rewarming (n = 6). All groups were studied for 4 hours. RESULTS: The partial pressure of arterial oxygen and lung compliance were significantly lower in the RWB group than in controls (113.8+/-33.1 vs 417.3+/-6.2 mm Hg, p < 0.01; and 0.8+/-0.2 vs 2.9+/-0.4 ml/cm H2O, p < 0.05, respectively). Pulmonary vascular resistance and lung wet/dry weight ratios were significantly higher in the RWB group than in controls (15884.1+/-11354.8 vs 6108.3+/-1309.9 dyne x sec x cm[-5], p < 0.05; and 7.13+/-0.24 vs 5.82+/-0.35, p < 0.05, respectively). The WB and leukocyte-depleted groups did not differ significantly from controls for any measured parameter. CONCLUSIONS: This model confirms that rewarming ischemia during lung implantation exacerbates reperfusion injury. Leukocyte-depleted reperfusion as tested for a short period of time (10 minutes) ameliorates this injury and therefore should be considered for clinical lung transplantation.

L-arginine administration during reperfusion improves pulmonary function.

BACKGROUND: Nitric oxide is crucial to the maintenance of vascular homeostasis. Because nitric oxide levels decline upon lung reperfusion, infusion of L-arginine, a nitric oxide precursor, during reperfusion might prove effective at ameliorating reperfusion injury. METHODS: Neonatal piglet heart-lung blocks were preserved with Euro-Collins solution for 12 hours, rewarmed at room temperature for 1 hour, and reperfused for 10 minutes with either whole blood (n = 5), whole blood containing L-arginine (10 mmol/L; n = 6), or leukocyte-depleted blood (n = 6) on an isolated, blood-perfused, working heart-lung circuit. After the initial 10 minutes, all blocks received whole blood for 4 hours. Control blocks were continuously perfused on the circuit without intervening ischemia (n = 6). RESULTS: The partial pressure of oxygen in the whole blood group (113.8 +/- 33.1 mm Hg) was significantly less than in controls (417.3 +/- 6.2 mm Hg; p < 0.01). Lung compliance was significantly less in the whole blood group (0.8 +/- 0.2 mL/cm H2O) than in controls (2.9 +/- 0.4 mL/cm H2O; p < 0.01). The L-arginine and leukocyte-depleted blood groups showed no significant difference from controls. CONCLUSIONS: L-Arginine infusion during reperfusion improves pulmonary function, making it a simple alternative to leukocyte depletion.

Normocalcemic blood or crystalloid cardioplegia provides better neonatal myocardial protection than does low-calcium cardioplegia.

Although standard blood cardioplegia provides good myocardial protection for cardiac operations in adults, protection of the cyanotic, immature myocardium remains suboptimal. Calcium, which has been implicated in reperfusion injury and in the development of "stone heart" in mature myocardium, is routinely lowered in standard cardioplegic solutions. Immature, neonatal myocardium has lower intracellular calcium stores and is more reliant on extracellular calcium for contraction. To determine if normocalcemic cardioplegia would result in improved cardiac function in the neonatal heart, we conducted a series of experiments using an isolated, blood-perfused working heart model. Thirty-two neonatal piglet hearts (24 to 48 hours) were excised without intervening ischemia and were placed directly on a blood-perfused circuit. Baseline stroke work index was assessed. Hearts were then arrested with cold cardioplegic solution delivered at 45 mm Hg for 2 minutes: group I, low-calcium blood cardioplegic solution (Ca = 0.6 mmol/L); group II, normal-calcium blood cardioplegic solution (Ca = 1.1 mmol/L); group III, University of Wisconsin solution; and group IV, University of Wisconsin solution with added calcium (Ca = 1.0 mmol/L). Cardioplegic solution was administered every 20 minutes for 2 hours and topical hypothermia was used. Hearts were then reperfused with warm whole blood. Functional recovery, expressed as a percentage of control stroke work index, was determined minutes after reperfusion. Hearts preserved with normocalcemic cardioplegic solution (groups II and IV) had complete functional recovery at 60 minutes, whereas hearts preserved with low-calcium cardioplegic solution (groups I and III) achieved functional recoveries of only 80% and 65%, respectively, at a left atrial pressure of 9 mm Hg. Electron micrographs taken 1 hour after reperfusion showed minimal edema and only mild myofibrillar changes. They were identical in both the low-calcium and normocalcemic groups. Complete functional recovery is possible in immature myocardium when calcium is added to either blood or an intracellular crystalloid cardioplegic solution. The addition of calcium does not result in ultrastructural damage and does result in good functional recovery.

Fluosol cardioplegia results in complete functional recovery: a comparison with blood cardioplegia.

Blood cardioplegia is considered by many to be the preferred solution for myocardial protection. Proposed benefits include the ability to deliver oxygen and the ability to maintain metabolic substrate stores. However, the decreased capacity of blood to release oxygen at hypothermic conditions as well as the presence of deleterious leukocytes, platelets, and complement may limit complete functional recovery. Fluosol is an asanguineous solution with the ability to bind and release oxygen linearly at low temperatures. Neonatal piglet hearts (24 to 48 hours old) were excised and supported on an isolated, blood-perfused working heart model. After baseline stroke-work index was determined, hearts were arrested with either normocalcemic blood cardioplegia (group 1, n = 8) or normocalcemic Fluosol cardioplegia (group 2, n = 8). Cold cardioplegia was administered at 45 mm Hg every 20 minutes for 2 hours. Hearts were then reperfused with whole blood. Functional recovery, expressed as percent of control stroke-work index, was determined 60 minutes after reperfusion at left atrial pressures of 3, 6, 9, and 12 mm Hg. Functional recovery at 60 minutes was similar between group 1 (95%, 93%, 93%, 88%) and group 2 (100%, 94%, 94%, 95%) at left atrial pressures of 3, 6, 9, and 12 mm Hg, respectively. Mean lactate consumption 5 minutes after reperfusion was significantly greater (p = 0.0001) in group 1 (31.8 +/- 6.3 micrograms.min-1 x g-1) than in group 2 (-0.59 +/- 0.1 microgram.min-1 x g-1), indicating superior metabolic recovery in the blood cardioplegia hearts. Edema formation, as determined both by water content (group 1, 81.10%; group 2, 81.63%) and by electron microscopy, was not significantly different between groups.(ABSTRACT TRUNCATED AT 250 WORDS)