Prevention of Descending Pneumonia in Rats with Perflubron-delivered Tobramycin

OBJECTIVES: Patients undergoing emergent endotracheal intubation are at increased risk for developing pneumonia. Although numerous strategies have been investigated to reduce ventilator-associated pneumonia (VAP), the incidence of VAP and its associated mortality remains high. This investigation tested the hypothesis that LiquiVent (Alliance Pharmaceutical, San Diego, CA-LV) delivered antibiotics (via spray-dried microspheres-SDM) would improve survival in a rat model of descending gram-negative pneumonia. METHODS: Wistar rats (n = 49) were randomized to receive prophylaxis with 1). nothing (controls); 2). intramuscular (IM) tobramycin, 3). intratracheal LV plus SDM shells (vehicle), 4). intratracheal LV plus SDM shells plus IM tobramycin, or 5). intratracheal LV plus SDM containing 1 mg/kg of tobramycin. All interventions were given 24 hours before a bacterial challenge with 10(8) colony-forming units of intratracheal Klebsiella pneumoniae. Mortality at ten days was the sole outcome measure. Survival in individual groups was compared with controls by Fisher's exact test with Bonferroni correction for multiple comparisons. RESULTS: All animals in the control group died of pneumonia within ten days of bacterial inoculation (0% survival). Prophylaxis with either IM tobramycin or SDM vehicle plus IM tobramycin provided no protection (0% survival). This is in sharp contrast to the cohort receiving pretreatment with tobramycin-containing SDM delivered via LV, in which 60% of the animals survived to study completion (p < 0.05). CONCLUSIONS: Prophylaxis with SDM containing antibiotics delivered in low-dose LV provided significant protection in a rat model of descending gram-negative pneumonia. These data support the hypothesis that perfluorocarbon-delivered intratracheal antimicrobials may be useful in the prevention of VAP.     

Indium-labeled presensitized T cells for diagnosis of graft rejection.

The aim of this experiment was to test a safe, noninvasive method for necessary, accurate diagnosis of early allograft rejection. Heart-lung allograft was performed heterotopically using Brown Norway (BN) rats as the donor and Lewis (LEW) rats as the recipient. T cell suspensions were prepared from lymphnodes of specifically sensitized LEW rats that had acutely rejected full-thickness BN skin graft. Cell count was adjusted 50 x 10(6) cells/ml. The suspension was incubated in vitro with 111I oxide (1 m Ci-ml). An aliquot of labeled cell suspension containing 40 x 10(6) cells and a total radioactivity of 200 mCi was administered intravenously to each animal 3 and 6 days after heart-lung transplant. The traffic of T cells was followed in vivo and in isolated organs under large field view gamma camera. The gamma camera revealed radioactivity on the graft starting Postoperative Day 5 when the heart was actively beating; no radioactivity was revealed at the site of the isografted organs. The histology showed mild to moderate cellular infiltration parallel to the grade of radioimaging intensity. The injection of indium-labeled presensitized T cells was able to detect the rejection process in an early phase when there are no clinical symptoms of rejection and/or the rejection cascade can be reversed. These results suggest that a similar method can be used in human organ transplantation for early diagnosis of rejection.     

High affinity neurotrophin receptors in the human pre-term newborn, infant, and adult cerebellum

The immunohistochemical occurrence of the high affinity neurotrophin (NT) receptors trkA, trkB, and trkC is shown in the pre-term newborn, infant, and adult human post-mortem cerebellum. Immunoreactive neuronal perikarya and processes were observed in all specimens examined, where they appeared unevenly distributed in the cerebellar cortical layers and deep nuclei, and showed regional differences among cerebellar lobules and folia. The trk receptor-antibodies, tested by Western blot on human cerebellum homogenates, revealed multiple immunoreactive bands for trkA and single bands for trkB and trkC. The results obtained show the tissue localization of the trk receptor-like immunoreactivity in the human cerebellum from prenatal to adult age. The analysis for codistribution of the receptors with the relevant ligand and among the receptors in discrete cortical and deep nuclei tissue fields shows a wide variety of conditions, from a good similarity in terms of type and density of labeled structures, to a lack of correspondence, and suggests the possibility of colocalization of trk receptors with the relevant neurotrophin and among them in the cerebellar cortex. These results sustain the concept that the neurotrophin trophic system participates in the development, differentiation, and maintenance of the human cerebellar connectivity and support the possibility of a multifactorial trophic support for the neurotrophins through target-derived and local mechanisms.     

Localization and characterization of epidermal growth-factor receptors in the developing rat medial septal area in culture

The presence and binding properties of epidermal growth-factor receptors (EGF-Rs) in different cell types purified from the rat medial septal area in culture were investigated. We report that astrocytes, oligodendrocytes and neurons from this area possess EGF-Rs while microglia do not. EGF-binding sites are detectable on astrocytes derived from the medial septum of both embryonic and neonatal rats. Scatchard analysis of the data for astrocytes from the fetal rats show that EGF specifically binds to both high- (K_d = 7.21 × 10⁻¹⁰ M, B_max = 3602 receptors/cell) and low-affinity (K_d = 3.99 × 10⁻⁸ 10⁻⁸ M, B_max = 6,265 receptors/cell) receptors on these cells. On the other hand, astrocytes purified from neonatal tissue possess a greater number of high-affinity receptors (B_max = 10,938 receptors/cell) when compared with the embryonic astroglia. With time in culture, the number of both types of receptors on neonatal astrocytes decreases. Oligodendrocytes also possess high- and low-affinity EGF-Rs with dissociation constants of 3.25 × 10⁻¹⁰ M and 3.85 × 10⁻⁸ M, respectively. The number of receptors on oligodendrocytes is significantly lower than those on neonatal astrocytes (B_max = 1185 and 25,081 receptors/cell for high- and low-affinity binding sites, respectively). Finally, neurons from this area also exhibit two different EGF-R types with dissociation constants similar to those described for astrocytes. As the number of receptors/neuron (B_max = 136 and 1159 receptors/cell for high- and low-affinity binding sites, respectively) appears to be extremely low, it is possible that EGF specifically binds only to a subpopulation of neurons from this area. These studies demonstrate which cell types in the developing medial sepal area posses EGF-Rs and provide a detailed characterization of these binding sites. These EGF-R-bearing cells may be potential targets for this growth factor or for transforming growth factor α in this brain area.