Gene-expression-based analysis of local and metastatic neuroblastoma variants reveals a set of genes associated with tumor progression in neuroblastoma patients.

Metastasis is the primary cause of mortality in Neuroblastoma (NB) patients, but the metastatic process in NB is poorly understood. Metastsis is a multistep process that requires the coordinated action of many genes. The identification of genes that promote or suppress tumor metastasis can advance our understanding of this process. In the present study, we utilized a human NB xenograft model comprising local and metastatic NB variants, which was recently developed in our laboratory. We set out to identify molecular correlates of NB metastasis and to determine the clinical relevance of these molecules. We first performed genome-wide expression profiles of metastatic and nonmetastatic NB variants that have an identical genetic background. We found that some of the proteins highly expressed in the metastatic NB variants are localized in the cytoplasm and endoplasmic reticulum. Other proteins are linked to metabolic processes and signaling pathways, thereby supporting the invasive and metastatic state of the cells. Subsequently, we intersected the differentially expressed genes in the human xenografted variants with genes differentially expressed in Stage 1 and Stage 4 primary tumors of NB patients. By using the same gene-expression platform, molecular correlates associated with metastatic progression in primary NB tumors were identified. The resulting smaller gene set was clinically relevant as it discriminated between high- and low-risk NB patients.

The effects of vascular intrauterine growth retardation on cortical astrocytes.

OBJECTIVE: We sought to determine the pathogenesis of neurodevelopmental impairments in survivors of intrauterine growth retardation (IUGR). METHODS: We used an experimental rabbit vascular IUGR model. We ligated 25% of uteroplacental vessels (partial ischemia) of one-half of the fetuses on day 25 at the end of the third trimester. We then determined hemispheral DNA and protein levels, and used glial fibrillary acidic protein (GFAP) staining to count the labeled astrocytes at the superficial cortical layers. RESULTS: Ischemic fetuses were significantly smaller than control fetuses and presented a disproportionately small body and a relatively larger head compared with the normal body/head ratio, confirming the study model as that of asymmetric IUGR. Hemispheral DNA was unchanged in IUGR fetuses, but they had decreased brain weight, hemispheral protein content, and a reduced number of mature (GFAP-positive) cortical astrocytes compared with control fetuses. CONCLUSION: Vascular IUGR, as demonstrated in our asymmetric IUGR model, adversely affected brain growth, cell size, and cortical astrocytes maturation. In view of the neurotrophic and neuroprotective functions of astrocytes, a reduced number of mature astrocytes during this critical period of development may be implicated in the pathogenesis of the neurodevelopmental impairments observed in IUGR.

Generation and characterization of novel local and metastatic human neuroblastoma variants.

Neuroblastoma (NB) is the most commonly occurring solid tumor in children. The disease usually arises in the adrenal medulla, and it is characterized by a remarkable heterogeneity in its progression. Most NB patients with an advanced disease have massive bone marrow infiltration at diagnosis. Lung metastasis represents a widely disseminated stage and is typically considered to be a terminal event. Much like other malignancies, NB progression is a complex, multistep process. The expression, function, and significance of the various factors involved in NB progression must be studied in relevant in vivo and in vitro models. Currently, models consisting of metastatic and nonmetastatic cell variants of the same genetic background exist for several types of cancer; however, none exists for NB. In the present study, we describe the generation of a NB metastasis model. SH-SY5Y and MHH-NB-11 NB cells were inoculated orthotopically into the adrenal glands of athymic nude mice. Neuroblastoma cells metastasizing to the lungs were isolated from mice bearing adrenal tumors. Lung metastatic variants were generated by repeated cycles of in vivo passage. Characterization of these variants included cellular morphology and immunophenotyping in vitro, aggressiveness in vivo, and various biologic parameters in vitro. The NB metastatic variant in each model displayed unique properties, and both metastatic variants demonstrated a metastatic phenotype in vivo. These reproducible models of human NB metastasis will serve as an unlimited source of transcriptomic and proteomic material. Such models can facilitate future studies on NB metastasis and the identification of novel NB biomarkers and targets for therapy.

Telmisartan in the treatment of Cohen-Rosenthal Diabetic Hypertensive rats: the benefit of PPAR-gamma agonism.

The antihypertensive and hypoglycemic effects of telmisartan, which has dual angiotensin II antagonist-PPAR-gamma agonist properties, was studied in Cohen-Rosenthal Diabetic Hypertensive rats (CRDH), a model in which hypertension, insulin resistance, and diabetes co-exist. CRDH, Cohen-diabetic rats (CDR), and SHR received telmisartan (3 mg/kg/day in drinking water) for five months. Telmisartan significantly lowered systolic and diastolic BP in SHR and CRDH, independent of body weight, and remained fairly constant in controls throughout the experiment. Blood glucose levels fell rapidly in the treated animals and remained steady in controls. Results indicate that telmisartan is a prototype of a new approach to treating coexisting diabetes and hypertension.

CO(2) Laser Fascia to Dura Soldering for Pig Dural Defect Reconstruction.

BACKGROUND AND OBJECTIVES: The purposes of this study were to demonstrate that laser soldering is safe and effective for tissue bonding in dural reconstruction and to compare this new reconstruction technique to an established one. STUDY DESIGN: A temperature-controlled fiberoptic CO(2) laser system or fibrin glue were used for in vitro dural defect reconstruction in two groups of pigs. The CO(2) laser technique was also used for dural reconstruction in live pigs. RESULTS: The burst pressure of the reconstructed dura by the laser system was significantly higher than that of fibrin glue (mean pressure 258.5 +/- 117.3 cm H(2)O and 76.8 +/- 47.2 cm H(2)O, respectively). There were no postoperative complications and no signs of thermal damage to the dura, fascia, or underlying tissue on histological analysis following the in vivo CO(2) laser experiments. CONCLUSIONS: Temperature-controlled laser soldering is an effective technique for dural repair. It creates a strong tissue bonding with no thermal damage to the tissue. The burst pressure of the reconstructed dura done with laser soldering is significantly higher than that of fibrin glue.

NAP enhances neurodevelopment of newborn apolipoprotein E-deficient mice subjected to hypoxia.

Perinatal hypoxic injury is associated with significant neonatal morbidity and long-term neurodevelopmental complications. NAP, a peptide derived from ADNP (activity-dependent neuroprotective protein), has previously shown neuroprotective abilities in various adult animal models. To evaluate its neuroprotective role in neonatal hypoxic-ischemic injury, we evaluated the neurodevelopmental outcome in apolipoprotein E (ApoE)-deficient (knockout) mice (a breed prone to brain damage during hypoxic insult) exposed to postnatal global hypoxic damage with and without treatment with NAP. ApoE-deficient (n = 80) and control (C57B6) mice pups (n = 81) were exposed to postnatal global hypoxia (35 min of 8% O(2) within 24 h of birth) or room air with or without subsequent subcutaneous NAP treatment during postnatal days 1 to 14. Pups were then evaluated for neonatal motor reflex attainment, spatial learning ability in the Morris water maze, and locomotor open-field activity. The C57B6 and ApoE-deficient anoxic groups showed significantly slower achievement of neonatal reflexes, diminished locomotor activity, and diminished spatial learning ability compared with their control groups. This was more pronounced in the anoxic ApoE-deficient pups. NAP treatment had a pronounced effect on neurodevelopmental outcome in both breeds, particularly in the ApoE-deficient mice. ApoE-deficient and control mouse pups exposed to postnatal hypoxia and treated with NAP showed improvement in neurodevelopmental outcome compared with nontreated mice pups. ApoE-deficient mice show a greater susceptibility to hypoxic damage and better response to NAP treatment.

Dural defect repair with fascia by a CO2 laser system in a porcine model.

HYPOTHESIS/OBJECTIVES: We tested the effectiveness of a temperature-controlled CO2 laser soldering system on a porcine model for dural defect reconstruction using a fascial patch. METHODS: A dural patch was excised and then reconstructed with fascia by a CO2 laser system in vitro in 27 animals and in vivo in five animals. RESULTS: After dural reconstruction, the average burst pressure of the soldered patch in vitro, as measured by a custom-made pressure detector, was 258.5 cm H2O. All five pigs in the in vivo group showed no neurological complications or cerebrospinal fluid leak, and the underlying brain tissue showed no thermal injury. CONCLUSION: The CO2 laser system created a watertight bond and did not cause thermal injury to the brain. The procedure is potentially faster than conventional repair, and wound healing may also be more rapid.

The pregnant spontaneously hypertensive rat as a model of asymmetric intrauterine growth retardation and neurodevelopmental delay.

INTRODUCTION: Hypertension in pregnancy and vascular placental insufficiency are considered common pathogenic factors in human intrauterine growth retardation (IUGR). IUGR neonates experience higher mortality, and the surviving infants have a higher incidence of neurological and intellectual impairment. METHODS: To mimic this condition, we used pregnant spontaneously hypertensive rats (SHR) and performed biometric measurements on Embryonic Day 20, postnatal developmental reflexes, and locomotor activity evaluations. RESULTS: SHR fetuses had significant decreased body weight compared to the Wistar-Kyoto control fetuses (1.51+/-0.02 g vs. 2.05+/-0.01 g, respectively; p<0.0001), and were relatively microcephalic (2.86+/-0.04 cm vs. 3.3+/-0.03 cm, respectively; p<0.0001). Their cephalization index (head circumference/body weight) was increased (1.88+/-0.03 vs. 1.62+/-0.02, respectively; p<0.0001), indicating a "brain-sparing" process. The disproportional ratio indicated that the IUGR type in this model is asymmetric. The SHR pups exhibited a significant (p<0.04) neurodevelopmental delay in the acquisition of neonatal reflexes (righting, negative geotaxis, placing), but they spontaneously caught up with the control pups after approximately 10 days. On Day 30, the SHR pups exhibited significantly increased walking speed and distance and spent less time in quadrant than the controls (p<0.002). CONCLUSION: We speculate that the model of pregnant SHR closely simulate human IUGR caused by hypertension in pregnancy and should enable investigation of mechanisms of hypertension-mediated placenta-vascular injury as well as provide a system for preclinical evaluations of future preventive neuroprotective treatments.

Closure of skin incisions in rabbits by laser soldering: I: Wound healing pattern.

BACKGROUND AND OBJECTIVES: Temperature-controlled tissue laser soldering is an innovative sutureless technique awaiting only solid experimental data to become the gold-standard surgical procedure for incision closure. The goals of the current study were: (1) to define the optimal laser soldering conditions, (2) to explore the immediate skin reparative healing events after sealing the wound, and (3) to determine the long-term trajectory of skin wound healing. STUDY DESIGN/MATERIALS AND METHODS: Skin incisions were generated over rabbit dorsa and were closed using different wound-closure interventions, in three groups: (a) closure, using a temperature-controlled infrared fiberoptic CO2 laser system, employing 47% bovine serum albumin as a solder; (b) wound closure by cyanoacrylate glues; and (c) wound closure by sutures. The reparative outcomes were evaluated macroscopically and microscopically, employing semi-quantitative grading indices. RESULTS: Laser soldering of incisions at T = 65 degrees C emerged as the optimal method achieving immediate wound sealing. This in turn induced accelerated reparative events characterized by a reduced inflammatory reaction, followed by minimal scarring and leading to a fine quality healing. CONCLUSIONS: Temperature-controlled laser soldering offers an accelerated wound reparative process with numerous advantages over the conventional methods. Further investigations may reveal additional benefits in the spectrum of advantages that this innovative surgical technology has to offer. This can introduce new scientific insight that will pave the way for clinical use.

Closure of skin incisions in rabbits by laser soldering II: Tensile strength.

BACKGROUND AND OBJECTIVES: The basic characteristic property of wound closure is the immediate and long-term tensile strength (LTS). The objective of the current study was to compare tissue laser soldering to other available methods (i.e., cyanoacrylate glues and sutures) in the performance and outcome of wound closure and reparative healing process, with an emphasis on the immediate and LTS. STUDY DESIGN/MATERIALS AND METHODS: The animals were divided into three groups according to the type and details of the closure procedure. Group A: laser treatments at different temperatures were compared to sutured incisions, emphasizing the LTS after 10 days. Group B: laser soldering at 65 +/- 5 degrees C was compared to chemical glues (i.e., Histoacryl and Dermabond), emphasizing the immediate tensile strength (ITS). Group C: LTS of laser soldered incisions was compared to that of sutured incisions at various time intervals emphasizing LTS (3, 7, 14, 28 days). RESULTS: Group A: LTS at 60 degrees C exhibited the highest values (0.48 MPa). Group B: no ITS difference was detected between laser soldering and chemical glues. Group C: soldered incisions at 65 degrees C exhibited higher LTS (1.81 MPa) than that of sutured incisions (1.08 MPa) (P < 0.043). CONCLUSIONS: Temperature-controlled laser soldering at 65 degrees C provided sufficient ITS and higher bonding LTS values compared with sutures, resulting in better wound healing characteristics. The laser soldering system presented here should be tested on larger animal models before adopting it for clinical usage.

Immunomodulatory and protective effects of moxifloxacin against Candida albicans-induced bronchopneumonia in mice injected with cyclophosphamide.

In a previous study, moxifloxacin was shown to ameliorate immunosuppression and enhance cytokine production in several tissues, including the lungs of cyclophosphamide-injected mice. We examined here the effects of moxifloxacin on Candida albicans lung infection in cyclophosphamide-injected mice. Mice were injected on day 0 with 250 mg of cyclophosphamide/kg, and on days 1 to 4 they were given moxifloxacin at 22.5 mg/kg/day compared to controls given ceftazidime at 75 mg/kg/day or saline. On day 6, C. albicans (10 7 CFU/mouse) was inoculated intratracheally, and animals were observed for the development of bronchopneumonia, weight loss, mortality, the presence of C. albicans, and lung cytokine production. Histopathology on day 10 postinoculation revealed bronchopneumonia in 50, 67, and 0% of saline-, ceftazidime-, and moxifloxacin-treated mice, respectively (P < 0.05). The mortality rates were 28, 17, and 5%, respectively (P < 0.05), and weight loss occurred at 20, 32, and 0%, respectively (P < 0.05). By day 15, C. albicans was eliminated from all moxifloxacin-treated mice but was still isolated from lung homogenates of 50 to 60% of the saline- and ceftazidime-treated groups. Among the cytokines tested on days 0 to 15, we found an increased production of tumor necrosis factor alpha, KC (functional interleukin-8), and gamma interferon in the lungs of ceftazidime- and saline-treated controls compared to the moxifloxacin pretreatment that abolished their secretion. In conclusion, moxifloxacin protected cyclophosphamide-injected mice from C. albicans-induced lung infection and significantly reduced pneumonia, weight loss, and mortality despite the lack of direct antifungal activity. This is most likely due to an immunomodulating activity conferred by moxifloxacin, as shown in this model and in our previous studies. Its potential protective role should be studied in patients undergoing chemotherapy and immune suppression.