Alpha 1-acid glycoprotein (AGP): a possible carrier of sialyl lewis X (slewis X) antigen in colorectal carcinoma.

OBJECTIVES: 1- to detect alpha 1-acid glycoprotein (AGP) and sialyl Lewis x (sLex) in colorectal malignant, benign and normal samples; 2- to isolate AGP from colorectal cancer and 3- to study its immunoreactivity with an anti-sLex monoclonal antibody (MAb). MATERIALS AND METHODS: tissue and serum samples from 88 patients with colorectal cancer, 22 adenomas and 23 normal were included. Expression of AGP and sLex was studied by immunohistochemistry (IHC); isolation approach: AGP was precipitated with ammonium sulphate and immunoprecipitated with anti-AGP MAb. The immune complex formed was isolated by protein A-Sepharose CL-4B affinity chromatography and further eluted; fractions were analysed by SDS-PAGE and Western-blot. Statistical analysis was performed by means of Principal Component Analysis. RESULTS: By Western blot employing anti-AGP MAb and sLex MAbs, isolated fractions from malignant samples showed a band at about 45 kD. IHC revealed that AGP was expressed in 70% of colorectal carcinoma samples, 50% of benign and 35% of normals. SLex was detected in 31% of malignant samples, 41% of benign and in one normal sample. In malignant samples, AGP reaction comprised the whole specimen with a strong and homogeneous staining while normal and benign samples showed a restricted reaction. In cancer, sLex expression consisted in an intense reactivity in membrane, cellular debris and some cytoplasmic foci while normal and benign samples were occasionally stained. A statistically significant positive correlation was found between AGP and sLex expression. Serum AGP levels were measured by radial immunodiffusion and statistical comparative analysis with tissue expression did not show a correlation between both parameters. CONCLUSION: AGP may constitute a carrier of sLex in colorectal cancer.

Synthesis, characterization and bioactivity of a new VO2+/aspirin complex.

A new VO2+ complex with salicylic acid acetate (Aspirin) of formula C18H18Cl2O12V2 was synthesized and characterized. Its biological effects upon cell proliferation, differentiation and promotion of tyrosine protein phosphorylation have been tested in two lines of osteoblast-like cells in culture.

Vanadium compounds. Their action on alkaline phosphatase activity.

The direct effect of different vanadium compounds upon alkaline phosphatase (ALP) activity was investigated. Vanadate and vanadyl inhibited both the soluble and particulate ALP activity from UMR.106 cells and from bovine intestinal ALP. We have also shown the inhibition of ALP activity in the soluble fraction of osteoblasts by peroxo and hydroperoxo vanadium compounds. ALP activity in the particulate fraction was not inhibited by these species; nor was the bovine intestinal ALP. Using inhibitors of Tyr-phosphatase (PTPases), the soluble ALP was partially characterized as a PTPase. The major activity in the particulate fraction represents the bone-specific ALP-activity. This study demonstrates that different forms of vanadium are direct inhibitors of ALP activity. This effect is dependent on the enzymatic activity investigated and on the origin of the ALP.

Effect of vanadium compounds on acid phosphatase activity.

The direct effect of different vanadium compounds on acid phosphatase (ACP) activity was investigated. Vanadate and vanadyl but not pervanadate inhibited the wheat germ ACP activity. These vanadium derivatives did not alter the fibroblast Swiss 3T3 soluble fraction ACP activity. Using inhibitors of tyrosine phosphatases (PTPases), the wheat germ ACP was partially characterized as a PTPase. This study suggests that the inhibitory ability of different vanadium derivatives to modulate ACP activity seems to depend on the geometry around the vanadium atom more than on the oxidation state. Our results indicate a correlation between the PTPase activity and the sensitivity to vanadate and vanadyl cation.

Performance assessment of a glucose control protocol in septic patients with an automated intermittent plasma glucose monitoring device.

BACKGROUND & AIMS: The optimal level and modality of glucose control in critically ill patients is still debated. A protocolized approach and the use of nearly-continuous technologies are recommended to manage hyperglycemia, hypoglycemia and glycemic variability. We recently proposed a pato-physiology-based glucose control protocol which takes into account patient glucose/carbohydrate intake and insulin resistance. Aim of the present investigation was to assess the performance of our protocol with an automated intermittent plasma glucose monitoring device (OptiScanner™ 5000). METHODS: OptiScanner™ was used in 6 septic patients, providing glucose measurement every 15' from a side-port of an indwelling central venous catheter. Target level of glucose was 80-150 mg/dL. Insulin infusion and kcal with nutritional support were also recorded. RESULTS: 6 septic patients were studied for 319 h (1277 measurements); 58 [45-65] hours for each patient (measurements/patient: 231 [172-265]). Blood glucose was at target for 93 [90-98]% of study time. Mean plasma glucose was 126 ± 11 mg/dL. Only 3 hypoglycemic episodes (78, 78, 69 mg/dL) were recorded. Glucose variability was limited: plasma glucose coefficient of variation was 11.7 ± 4.0% and plasma glucose standard deviation was 14.3 ± 5.5 mg/dL. CONCLUSIONS: The local glucose control protocol achieved satisfactory glucose control in septic patients along with a high degree of safeness. Automated intermittent plasma glucose monitoring seemed useful to assess the performance of the protocol.

Extracorporeal lung perfusion and ventilation to improve donor lung function and increase the number of organs available for transplantation.

INTRODUCTION: Ex vivo lung perfusion (EVLP) has been validated as a valuable technique to increase the pool of organs available for lung transplantation. MATERIAL AND METHODS: After a preclinical experience, we obtained permission from the Ethics Committee of our institution to transplant lungs after EVLP reconditioning. ABO compatibility, size match, and donor arterial oxygen pressure (PaO(2))/fraction of inspired oxygen (FiO(2)) ≤ 300 mm Hg were considered to be inclusion criteria, whereas the presence of chest trauma and lung contusion, evidence of gastric content aspiration, pneumonia, sepsis, or systemic disease were exclusion criteria. We only considered subjects on an extra corporeal membrane oxygenation (ECMO) bridge to transplantation with rapid functional deterioration. Using Steen solution with packed red blood cells oxygenated with 21% O(2), 5% to 7% CO(2) was delivered, targeted with a blood flow of approximately 40% predicted cardiac output. Once normothermic, the lungs were ventilated with a tidal volume of 7 mL/kg a PEEP of 5 cmH(2)O and a respiratory rate of 7 bpm. Lungs were considered to be suitable for transplantation if well oxygenated [P(v-a) O(2) > 350 mm Hg on FiO(2) 100%], in the absence of deterioration of pulmonary vascular resistance and lung mechanics over the perfusion time. RESULTS: From March to September 2011, six lung transplantations were performed, including two with EVLP. The functional outcomes were similar between groups: at T72 posttransplantation, the median PaO(2)/FiO(2) were 306 mm Hg (range, 282 to 331 mm Hg) and 323 mm Hg (range, 270 to 396 mm Hg) (P = 1, EVLP versus conventional). Intensive care unit ICU and hospital length of stay were similar (P = .533 and P = .663, respectively) with no mortality at 60 days in both groups. EVLP donors were older (49 ± 6 y versus 21 ± 7 y, P < .05), less well oxygenated (184 ± 6 mm Hg versus 570 ± 30, P < .05), displaying higher Oto scores (9.5 ± 0.7 versus 1.7 ± 1.5, P < .05). CONCLUSIONS: The first 6 months of the EVLP program allowed us to increase the number of organs available for transplantation with short-term outcomes comparable to conventional transplantations.

The consumption of glucose during ex vivo lung perfusion correlates with lung edema.

INTRODUCTION: Ex vivo lung perfusion (EVLP) has been recently proposed to recondition organs before transplantation from donors with marginal or unacceptable features. The aim of our investigation was to explore glucose consumption during EVLP. MATERIALS AND METHODS: We investigated 8 domestic pigs (mean weight, 21 ± 0.8 kg). After perfusion with Perfadex, retrieval, and back table surgery, we initiated EVLP. The lungs were perfused with Steen solution with added methylprednisolone, cefazoline, and heparin. The blood flow was gradually increased with a target of 40% of the estimated cardiac output (or less if the pulmonary artery pressure was >15 mm Hg), while keeping the left atrial pressure between 3 and 5 mm Hg. The temperature of the perfusate was increased from 25 °C to 37 °C. Once the temperature of the lung outflow was >32 °C, we began gas flow (4 L/min, 5%-8% CO(2) in air) and mechanical ventilation. EVLP parameters and blood gases were measured throughout the experiment; glucose consumption was calculated as (glucose initial-glucose final)/time. The wet to dry ratio was also calculated as an index of lung edema. RESULTS: When stratified by median glucose consumption (0.237 mg/min), high glucose consumers (0.588 ± 0.17) were characterized by worse lung function, as assessed by oxygenation (partial pressure of oxygen/inspiratory fraction of oxygen [PaO(2)/FiO(2)] 326 ± 63 mm Hg vs 218 ± 84; P=.083 low vs high, respectively), and lung edema (wet/dry ratio 6.5 ± 0.7 vs 8.6 ± 0.9; P=.012). Glucose consumption correlated with wet to dry ratio (R(2)=0.663; P=.014). CONCLUSIONS: We found that the worse the lung function, the greater the consumption of glucose during EVLP. This observation suggests the need to explore lung metabolism during EVLP to possibly obtain metrics for evaluation.

Proliferative and morphological changes induced by vanadium compounds on Swiss 3T3 fibroblasts.

Vanadium compounds are shown to have a mitogenic effect on fibroblast cells. The effects of vanadate, vanadyl and pervanadate on the proliferation and morphological changes of Swiss 3T3 cells in culture are compared. Vanadium derivatives induced cell proliferation in a biphasic manner, with a toxic-like effect at doses over 50 microM, after 24 h of incubation. Vanadyl and vanadate were equally potent at 2.5-10 microM. At 50 microM vanadate inhibited cell proliferation, whereas slight inhibition was observed at 100 microM of vanadyl. At 10 microM pervanadate was as potent as vanadate and vanadyl in stimulating fibroblast proliferation, but no effect was observed at lower concentrations. A pronounced cytotoxic-like effect was induced by pervanadate at 50 microM. All of these effects were accompanied by morphological changes: transformation of fibroblast shape from polygonal to fusiform; retraction with cytoplasm condensation; and loss of lamellar processes. The magnitude of these transformations correlates with the potency of vanadium derivatives to induce a cytotoxic-like effect: pervanadate > vanadate > vanadyl. These data suggest that the oxidation state and coordination geometry of vanadium determine the degree of the cytotoxicity.

Tyrosine phosphorylation and morphological transformation induced by four vanadium compounds on MC3T3E1 cells.

The present study was performed to determine the phosphotyrosine-protein levels induced by insulin and by four vanadium derivatives in MC3T3E1 osteoblast-like cells. We have also attempted to associate these patterns with the vanadium-induced growth and morphological changes of such cells. Vanadate (Vi), vanadyl (VO), bis(maltolato)oxovanadium (IV) (BMOV) and bis(maltolato)dioxovanadium (V) (BMV) stimulate cell growth in a narrow range of concentration, but are also inhibitors for the cells at high concentrations. Vanadium-treated cells displayed clear changes in their morphology after overnight incubation. However, BMV was the least cytotoxic and the weakest inducer of morphological changes. All the compounds promote the phosphorylation of tyrosine residues in several proteins. This effect was more pronounced at low than at high doses. At low doses (10 microM), BMV showed a phosphorylation pattern similar to that of insulin, while Vi, VO and BMOV induced strong phosphorylation of cell proteins. The present findings suggest that the vanadium-induced growth regulation and morphological changes in MC3T3E1 osteoblast-like cells are associated with the ability of these agents to increase the phosphotyrosine protein levels and to inhibit phosphotyrosine phosphatases. These properties are dependent on the oxidation state as well as on the organic ligand which coordinates the vanadium atom.