Novel Flurometric Tool to Assess Mitochondrial Redox State of Isolated Perfused Rat Lungs after Exposure to Hyperoxia.

Recently we demonstrated the utility of optical fluorometry to detect a change in the redox status of mitochondrial autofluorescent coenzymes NADH (Nicotinamide Adenine Dinucleotide) and FAD (oxidized form of Flavin Adenine Dinucleotide (FADH2,)) as a measure of mitochondrial function in isolated perfused rat lungs (IPL). The objective of this study was to utilize optical fluorometry to evaluate the effect of rat exposure to hyperoxia (>95% O2 for 48 hours) on lung tissue mitochondrial redox status of NADH and FAD in a nondestructive manner in IPL. Surface NADH and FAD signals were measured before and after lung perfusion with perfusate containing rotenone (ROT, complex I inhibitor), potassium cyanide (KCN, complex IV inhibitor), and/or pentachlorophenol (PCP, uncoupler). ROT- or KCN-induced increase in NADH signal is considered a measure of complex I activity, and KCN-induced decrease in FAD signal is considered a measure of complex II activity. The results show that hyperoxia decreased complex I and II activities by 63% and 55%, respectively, as compared to lungs of rats exposed to room air (normoxic rats). Mitochondrial complex I and II activities in lung homogenates were also lower (77% and 63%, respectively) for hyperoxic than for normoxic lungs. These results suggest that the mitochondrial matrix is more reduced in hyperoxic lungs than in normoxic lungs, and demonstrate the ability of optical fluorometry to detect a change in mitochondrial redox state of hyperoxic lungs prior to histological changes characteristic of hyperoxia.

Hidden minima of the gibbs free energy revealed in a phase separation in polymer/surfactant/water mixture.

We observed a very unusual kinetic pathway in a separating C(12)E(6)/PEG/H(2)O ternary mixture. We let the mixture separate above the spinodal temperature (cloud point temperature) for some time and next cool it into a metastable region of a phase diagram, characterized by two minima of the Gibbs potential, one corresponding to the homogeneous mixture and one to the fully separated PEG-rich and C(12)E(6)-rich phases. Despite the fact that in the metastable region the thermodynamic equilibrium corresponds to the separated phases (global minimum of the Gibbs free energy), we observe perfect mixing of the initially separated phase. The homogeneous state, obtained in this way, does not separate, if left undisturbed. However, many cooling-heating cycles or full separation with visible meniscus above the cloud point temperature induce the phase separation in the metastable region. The metastable region can exist tens of degrees below the cloud point temperature. This effect is not observed in the binary mixture of C(12)E(6)/H(2)O.

Ordering in surfactant mixtures induced by polymers.

We studied ternary mixtures of nonionic surfactant (C12E6, n-dodecyl hexaoxyethylene glycol monoether), polymer (PEG, polyethylene glycol), and water. A small amount of PEG induces demixing into the polymer-rich and surfactant-rich phases in the ternary PEG/C12E6/water mixture. Above a certain concentration and/or molecular weight of PEG, the surfactant-rich phase orders, even in a solution consisting of a few percent of surfactant. The phase boundary acts as a semipermeable membrane, and the equilibrium is determined by the chemical potential of water in two phases. The explicit expression for the amount of PEG needed to order C12E6 water solution is given and verified experimentally. The analysis of the coexistence conditions leads to the conjecture that only two oxygen atoms in the outward part of the hydrophilic surfactant head strongly affect the chemical potential of water. Our methodology is generic, i.e., on the same basis one can design a similar experiment for any surfactant/polymer/water system and find the right proportions of polymer that induce order in a surfactant-rich phase.

Endowing human pancreatic ribonuclease with toxicity for cancer cells.

Onconase is an amphibian protein that is now in Phase III clinical trials as a cancer chemotherapeutic. Human pancreatic ribonuclease (RNase 1) is homologous to Onconase but is not cytotoxic. Here, ERDD RNase 1, which is the L86E/N88R/G89D/R91D variant of RNase 1, is shown to have conformational stability and ribonucleolytic activity similar to that of the wild-type enzyme but > 10(3)-fold less affinity for the endogenous cytosolic ribonuclease inhibitor protein. Most significantly, ERDD RNase 1 is toxic to human leukemia cells. The addition of a non-native disulfide bond to ERDD RNase 1 not only increases the conformational stability of the enzyme but also increases its cytotoxicity such that its IC(50) value is only 8-fold greater than that of Onconase. Thus, only a few amino acid substitutions are necessary to make a human protein toxic to human cancer cells. This finding has significant implications for human cancer chemotherapy.

A synapomorphic disulfide bond is critical for the conformational stability and cytotoxicity of an amphibian ribonuclease.

Onconase((R)) (ONC) is a homolog of ribonuclease A (RNase A) that has unusually high conformational stability and is toxic to human cancer cells in vitro and in vivo. ONC and its amphibian homologs have a C-terminal disulfide bond, which is absent in RNase A. Replacing this cystine with a pair of alanine residues greatly decreases the conformational stability of ONC. In addition, the C87A/C104A variant is 10-fold less toxic to human leukemia cells. These data indicate that the synapomorphic disulfide bond of ONC is an important determinant of its cytotoxicity.

[Imaging in parotid tumors: US versus MRT]. / Bildgebung bei Parotistumoren: US versus MRT.

27 patients, clinically suspected of having parotid tumours, were examined prospectively by sonography (high frequency linear array) and MRI (1.5 T, T1- and T2-weighted SE sequences before, and T1-weighted SE sequences after contrast). In 23 patients, 25 intraglandular tumours were demonstrated (two were multicentric); they consisted of 6 malignant parotid tumours, 11 pleomorphic adenomas, 7 cystadenolymphomas and one oncocytic adenoma. 4 patients had extraglandular tumours. All tumours were demonstrated by either method, but two extraglandular tumours were incorrectly diagnosed by sonography as being intraglandular and 8 tumours (28%) were not completely delineated by sonography. MRI localised the tumours correctly in all cases and demonstrated all lesions accurately, independent of their position or size. In summary, sonography appears suitable as the primary imaging method in the investigation of parotid tumours but in case of large, parapharyngeal or malignant tumours, MRI is clearly superior to sonography.

[Value of ultrasound in follow-up of biliary metal stents--initial results]. / Wertigkeit der Sonographie in der Kontrolle biliärer Metallstents--Erste Ergebnisse.

Self-expandable metallic biliary endoprostheses (= "stents") are an accepted therapeutic alternative in the palliative treatment of malignant biliary obstruction. The physiological direction of bile flow is re-established for a varying period of time. Sonography is an acutely available modality for the evaluation of symptomatic patients who are carriers of such endoprostheses. The cause of recurrent obstructive jaundice can be diagnosed quickly and a new intervention can be planned carefully.

Pituitary adenomas: findings of postoperative MR imaging.

Preoperative and postoperative magnetic resonance (MR) images obtained in 25 patients with pituitary macroadenomas (1.1-5.2 cm in diameter) were evaluated to determine normal (physiologic) and abnormal findings after transsphenoidal or subfrontal surgery. With a 1.5-T unit, T1-weighted sagittal and coronal images were obtained before and after administration of gadopentetate dimeglumine. The physiologic changes after surgery included resorption of packing material and sphenoid sinus opacifications, reexpansion of the pituitary gland, and lowering of the optic chiasm. Implanted gelatin foam appeared as an endosellar, circularly enhancing mass that was seen on follow-up studies to have decreased in size. Muscle-and-fat implants appeared as areas of high signal intensity. In 14 patients, residual tumors were found in the suprasellar (n = 4), retrosellar (n = 3), parasellar (n = 8), and/or endosellar (n = 3) space. Residual tumors were differentiated from implant materials by means of location, characteristic signal intensity, and enhancing pattern, which were identical to those of the corresponding preoperative adenoma in 13 cases (93%). Preoperative studies and clinical information are helpful in the evaluation of postoperative MR imaging examinations.