The microvascular endothelial cell monolayer localized at the critical interface between the blood and vessel wall has the vital functions of regulating tissue fluid balance and supplying the essential nutrients needed for the survival of the organism. The endothelial cell is an exquisite "sensor" that responds to diverse signals generated in the blood, subendothelium, and interacting cells. The endothelial cell is able to dynamically regulate its paracellular and transcellular pathways for transport of plasma proteins, solutes, and liquid. The semipermeable characteristic of the endothelium (which distinguishes it from the epithelium) is crucial for establishing the transendothelial protein gradient (the colloid osmotic gradient) required for tissue fluid homeostasis. Interendothelial junctions comprise a complex array of proteins in series with the extracellular matrix constituents and serve to limit the transport of albumin and other plasma proteins by the paracellular pathway. This pathway is highly regulated by the activation of specific extrinsic and intrinsic signaling pathways. Recent evidence has also highlighted the importance of the heretofore enigmatic transcellular pathway in mediating albumin transport via transcytosis. Caveolae, the vesicular carriers filled with receptor-bound and unbound free solutes, have been shown to shuttle between the vascular and extravascular spaces depositing their contents outside the cell. This review summarizes and analyzes the recent data from genetic, physiological, cellular, and morphological studies that have addressed the signaling mechanisms involved in the regulation of both the paracellular and transcellular transport pathways.

https://www.physiology.org/doi/pdf/10.1152/physrev.00012.2005

Signaling Mechanisms Regulating Endothelial Permeability

Peroxidase-modified electrodes: Fundamentals and application

PURPOSE: The purpose of this study was to investigate whether positron emission tomography (PET) with the glucose analog [18F]fluorodeoxyglucose (FDG) and the estrogen analog 16 alpha-[18F]fluoroestradiol-17 beta (FES), performed before and after treatment with tamoxifen, could be used to detect hormone-induced changes in tumor metabolism (metabolic flare) and changes in available levels of estrogen receptor (ER). In addition, we investigated whether these PET findings would predict hormonally responsive breast cancer. PATIENTS AND METHODS: Forty women with biopsy-proved advanced ER-positive (ER+) breast cancer underwent PET with FDG and FES before and 7 to 10 days after initiation of tamoxifen therapy; 70 lesions were evaluated. Tumor FDG and FES uptake were assessed semiquantitatively by the standardized uptake value (SUV) method. The PET results were correlated with response to hormonal therapy. RESULTS: In the responders, the tumor FDG uptake increased after tamoxifen by 28.4% ± 23.3% (mean ± SD); onl...

Metabolic flare: indicator of hormone responsiveness in advanced breast cancer.

Cancer cells show increased metabolism of both glucose and amino acids, which can be monitored with 18F-2-deoxy-2-fluoro-D-glucose (FDG), a glucose analogue, and 11C-L-methionine (Met), respectively. FDG uptake is higher in fast-growing than in slow-growing tumors. FDG uptake is considered to be a good marker of the grade of malignancy. Several studies have indicated that the degree of FDG uptake in primary lung cancer can be used as a prognostic indicator. Differential diagnosis of lung tumors has been studied extensively with both computed tomography (CT) and positron emission tomography (PET). It has been established that FDG-PET is clinically very useful and that its diagnostic accuracy is higher than that of CT. Detection of lymph node or distant metastases in known cancer patients using a whole-body imaging technique with FDG-PET has become a good indication for PET. FDG uptake may be seen in a variety of tissues due to physiological glucose consumption. Also FDG uptake is not specific for cancer. Various types of active inflammation showed FDG uptake to a certain high level. Understanding of the physiological and benign causes of FDG uptake is important for accurate interpretation of FDG-PET. In monitoring radio/chemotherapy, changes in FDG uptake correlate with the number of viable cancer cells, whereas Met is a marker of proliferation. Reduction of FDG uptake is a sensitive marker of viable tissue, preceding necrotic extension and volumetric shrinkage. FDG-PET is useful for the detection of recurrence and for monitoring the therapeutic response of tumor tissues in various cancers, including those of the lung, colon, and head and neck. Thus, PET, particularly with FDG, is effective in monitoring cancer cell viability, and is clinically very useful for the diagnosis and detection of recurrence of lung and other cancers.

From tumor biology to clinical Pet: a review of positron emission tomography (PET) in oncology.

The biophysical mechanism(s) underlying diffusion-weighted MRI contrast following brain injury remains to be elucidated. Although it is generally accepted that water apparent diffusion coefficient (ADC) decreases after brain injury, it is unknown whether this is associated with a decrease in intracellular or extracellular water displacement, or both. To address this question, 2-[ 19 F]luoro-2-deoxyglucose-6-phosphate (2FDG-6P) was employed as a compartment-specific marker in normal and globally ischemic rat brain. Through judicious choice of routes of administration, 2FDG-6P was confined to the intra- or extracellular space. There was no statistical difference between intra- and extracellular 2FDG-6P ADCs in normal or in globally ischemic brain (p > 0.16), suggesting that water ADCs in both compartments are similar. However, ischemia did result in a 40% ADC decrease in both compartments (P < 0.001). Assuming that 2FDG-6P reflects water motion, this study shows that water ADC decreases in both spaces after ischemia, with the reduction of intracellular water motion being the primary source of diffusion-weighted contrast.

Evaluation of extra- and intracellular apparent diffusion in normal and globally ischemic rat brain via 19F NMR

Circulating biogenic amines, such as serotonin or norepinephrine, are cleared by the lung and can be considered as indicators of pulmonary endothelial integrity. An iodinated norepinephrine analogue, metaiodo-benzylguanidine (MIBG), is extracted in the lung by an active, saturable transport system. In order to investigate the use of MIBG lung extraction to detect minimal,lung endothelial cell lesions, an experimental model of initial pulmonary vascular lesions was developed in rats using repeated daily intraperitoneal injection of bleomycin (BLM: 2 U/100 g body weight) over a 5-day period. At this time, a significant decrease of serum angiotensin-converting enzyme activity, an index of endothelial cell metabolism, was observed (214±11 U/ml in controls as compared with 182±11 U/ml in BLM-treated rats,P < 0.05); electron microscopy showed the presence of typical but minimal endothelial cell lesions (blebbing). MIBG extraction (%EMIBG) was measured using the isolated perfused lung model after a 10-min steady-state period and 2 min of perfusion with123I-MIBG (0.1 μCi/ml) and125I-labelled human serum albumin (HSA) (0.2 μCi/ml). Intraperitoneal administration of BLM to rats for 5 days resulted in a significant decrease in pulmonary extraction of MIBG. %EMIBG declined from a control value of 24.7%±1.1% in controls to 19.3%±1.6% in BLM-treated rats (n=27,P<0.05; -21.2%). HSA lung extraction, used as an estimate of nonspecific residual lung activity, was not different in the lungs of BLM-treated rats as compared with controls. Because measurements of MIBG lung extraction (%EMIBG) were able to demonstrate the presence of minimal endothelial cell lesions due to the BLM toxicity, MIBG is an index of pulmonary endothelial cell function.

123I-MIBG pulmonary removal: a biochemical marker of minimal lung endothelial cell lesions.

Circulating biogenic amines are known to be cleared by the mammalian lung Their lung uptake is considered as an indicator of pulmonary endothelial integrity Unfortunately, their use as markers of pulmonary metabolic function in human pathology is precluded by their biological effects and by the type of radiolabeling (3H and 14C), making them harmful for repeat injections and unfit for scintigraphy Metaiodobenzylguanidine (MIBG) is structurally related to the neuron blocking agent guanethidine, devoid of significant biological effects, and has been shown to be extracted by the same active sodium dependent, saturable transport as norepinephrine in perfused rat lungs in vitro We studied the single pass lung extraction of 131I-MIBG in five awake and five anaesthetised sheep using the standard double indicator dilution technique with 99mTc-human serum albumin (HSA) as an intravascular reference tracer Intravenous bolus injection of increasing doses of MIBG up to 400 nmol resulted in a significant (F ratio=7778, P<00001) dose dependent decrease of MIBG extraction in both awake and anaesthetised sheep, without significant differences of extraction values between the two groups For the 10 sheep, the averaged percentage single pass pulmonary uptake of MIBG at the peak of the dilution curve decreased from 32%±3% (mean±SE, n=27 measurements) with 20 nmol to 18%±2% (n=32) with 400 nmol Estimates of the apparent Michaelis-Menten constant (Km) averaged 2±12 μM (n=7), whereas estimates of the apparent maximum velocity of removal (Vmax) was 11±05 μmol/min (n=7) In contrast to some intersubject variability, the pharmacokinetic parameters showed little intra subject variation No correlation was found between MIBG extraction, Km or Vmax values and haemodynamic or gas exchange parameters These data indicate that using a standard double dilution technique, lung extraction of MIBG may be determined in vivo Its lung removal is dose limited and may be characterised by the Michaelis-Menten kinetic constants suggesting a saturable process In contrast to norepinephrine, the gamma emitter labeled MIBG could therefore be a suitable compound to monitor pulmonary endothelial cell function in vivo using a non invasive scintigraphic method

Lung uptake of 131I-metaiodobenzylguanidine in sheep. An in vivo measurement of pulmonary metabolic function.

Radioiodinated metaiodobenzylguanidine (MIBG) has been shown to be extracted in the lung by an active, sodium-dependent, saturable transport system similar to that which extracts norepinephrine. Accordingly, first transit uptake of MIBG has been investigated as a method for in vivo evaluation of pulmonary biogenic amine metabolism. Issues that still must be resolved include the degree to which MIBG extraction is affected by simple loss of available vascular surface (in distinction to changes in amine metabolism). If MIBG lung extraction is altered by loss of pulmonary vascular surface alone, then a tracer must be found that can serve as a simultaneous monitor of vascular surface loss in order to allow normalization of the MIBG data. One tracer with such potential is radioiodinated iodoamphetamine (IMP). Accordingly, an experimental model of pulmonary vascular surface loss (lobar ligatures) was applied in an isolated-perfused lung model with simultaneous administration of MIBG and IMP. A linear relationship was found between MIBG extraction and percent lung surface loss by weight (n=21, r=0.75, P0.05). Addition of IMP (1 or 10 μM) to media containing MIBG (1 μM) (n=28) inhibited overall MIBG extraction, but decreases in MIBG uptake with vascular surface loss were otherwise parallel to those in experiments without IMP. IMP, however, did not adequately reflect vascular surface loss at the concentrations used in this study (extraction of IMP 1 μM=31.2±12.9% in control vs 32.7%±7.7% with greater than 50% lung surface loss, n=18, P=NS). The results indicate that a correction for vascular surface loss is needed to interpret MIBG results in vivo, but suggest that IMP is not able to serve this purpose.

131I-metaiodobenzylguanidine and 125I-iodoamphetamine. Parameters of lung endothelial cell function and pulmonary vascular area.

The use of fluorodeoxyglucose (FDG) and positron emission tomography (PET) is recognized as an accurate tool for the specific diagnosis and staging of cancer. It has also been proposed for the monitoring of anticancer therapy. FDG cell incorporation reflects glycolytic activity whereas inhibition of cell proliferation corresponds to an efficient cancer treatment. The relationship between FDG incorporation and cell proliferation has yet to be demonstrated. Therefore, we aimed to correlate the effects of the toxic agents bleomycin and unlabelled meta-iodobenzylguanidine (mIBG) on cellular metabolism and proliferation. We determined the in vitro metabolic and cytotoxic effects of bleomycin and mIBG by measuring the incorporation of fluorine-18 FDG (%UFDG) and hydrogen-3 thymidine (%UTHY) in cells of the human premonocytic line U937 in the presence of increasing concentrations of these agents. Proliferation rate of these cells was studied by means of limiting dilution analysis. %UTHY appeared more sensitive to bleomycin or mIBG-mediated cell injury than %UFDG. After 1 h of exposure to 0.5 μM bleomycin, %UTHY was significantly reduced to 62.0% ± 10.4% of control value whereas %UFDG remained unchanged (91.6% ± 5.3%). Similar results were obtained after 1 h of exposure to increasing concentrations of mIBG (1 μM to 1 mM). After 20 h of exposure to bleomycin, %UTHY and %UFDG were significantly reduced as a function of concentration. After 20 h of exposure to mIBG, a transient increase in %UFDG up to 149.3% ± 11.2% with 50 μM mIBG was further followed by a reduction to 20.1% ± 6.7% with 0.5 mM (P < 0.001). The clonogenic efficiency was reduced as a function of bleomycin (ANOVA, n=255, P) or mIBG concentration (n=80, P) and nearly abolished with 0.1 μM bleomycin or 0.1 mM mIBG. In conclusion, %UTHY appears to be a more sensitive index of cytotoxicity in vitro and more accurately relates to cell proliferation than %UFDG.

Fluorodeoxyglucose cell incorporation as an index of cell proliferation: evaluation of accuracy in cell culture

OBJECTIVES To determine a) whether hydrogen peroxide-induced, early lung endothelial cell dysfunction can be detected in an isolated, perfused, rat lung model; and b) whether the organic phosphothioate N-(2-mercaptoethyl)-1,3-propanediamine, which protects cells in culture against hydrogen peroxide-mediated damage, can exert the same protection in this model. DESIGN Intervention study; before-after trial. SETTING Research laboratory. MODEL Isolated, perfused, rat lung model. INTERVENTION Continuous hydrogen peroxide infusion at increasing concentrations and infusion times, preceded or not by a N-(2-mercaptoethyl)-1,3-propanediamine infusion. MEASUREMENTS AND MAIN RESULTS Early pulmonary endothelial cell alterations, assessed by the lung extraction (% extraction) of 123I-metaiodobenzylguanidine. Permeability edema by % extraction of 125I-human serum albumin and the lung dry-to-wet weight ratio. Control experiments: % extraction-123I-metaiodobenzylguanidine: 21.7 +/- 3.8% (n = 7). With increasing concentrations of hydrogen peroxide (0.025, 0.125, 0.5, and 2 mmol), % extraction-123I-metaiodobenzylguanidine was progressively depressed (n = 28, ANOVA, p < .05), significantly decreased from controls at 2 mmol (10.2 +/- 5.0%, n = 7, p < .05). When the 2-mmol hydrogen peroxide infusion was preceded by the N-(2-mercaptoethyl)-1,3-propanediamine (2 mmol) infusion, % extraction-123I-metaiodobenzylguanidine (19.9 +/- 2.9%, n = 5) was not significantly different from controls (n = 7) and was significantly greater than after the 2-mmol hydrogen peroxide infusion alone (8.7 +/- 7.4%, p < .05, n = 8). In all experiments, % extraction of human serum albumin ratio and dry-to-wet weight ratio were not significantly different from that of controls. CONCLUSIONS a) Hydrogen peroxide-induced lung endothelial cell dysfunction was detected at an early stage, before any permeability defect appeared; b) N-(2-mercaptoethyl)-1,3-propanediamine protected against such damage.

Alfred Donath

Papers

123I-MIBG pulmonary removal: a biochemical marker of minimal lung endothelial cell lesions.

Lung uptake of 131I-metaiodobenzylguanidine in sheep. An in vivo measurement of pulmonary metabolic function.

131I-metaiodobenzylguanidine and 125I-iodoamphetamine. Parameters of lung endothelial cell function and pulmonary vascular area.

Fluorodeoxyglucose cell incorporation as an index of cell proliferation: evaluation of accuracy in cell culture

Early hydrogen peroxide-induced pulmonary endothelial cell dysfunction: detection and prevention.