Endogenous tumor necrosis factor exerts its protective function intracellularly against the cytotoxicity of exogenous tumor necrosis factor.

Based on the findings that expression of endogenous tumor necrosis factor (enTNF), which is not present in TNF-susceptible cells, was generally observed in TNF-resistant cells and that TNF gene transfection gives rise to TNF resistance, the assumption was made that enTNF may be a protective protein against the cytotoxicity of exogenous TNF. However, it remains unknown whether the protection by enTNF is exerted in an intracellular or extracellular (autocrine) manner. We therefore transfected a nonsecretory human TNF gene (pTNF delta pro) into highly TNF-sensitive mouse tumorigenic fibroblasts (L-M cells) and investigated their TNF susceptibility. The transfectants expressed enTNF which was not secreted into the medium and acquired an appreciable degree of resistance to exogenous TNF. A significant increase in the manganous superoxide dismutase level was also noted in the transfectants. These findings suggest that enTNF exerts its protective function intracellularly by inducing manganous superoxide dismutase production.     

Endogenous tumor necrosis factor functions as a resistant factor against hyperthermic cytotoxicity.

One of the mechanisms of cytotoxicity by tumor necrosis factor (TNF) is the induction of reactive oxygen molecules. Cells producing endogenous tumor necrosis factor (enTNF) show resistance to the cytotoxicity of exogenous TNF by scavenging the reactive oxygen molecules. The intracellular hydroxyl radical production is also known to be involved in the heat-induced cytotoxicity. In the present study, we therefore examined the possibility that enTNF may act as a protective protein against the heat-induced cytotoxicity in a manner similar to that of exogenous TNF. Heat-sensitive L-M (mouse tumorigenic fibroblast) cells, originally expressing no enTNF, were transfected with a human TNF expression vector to produce enTNF. The stable transfectants showed apparent resistance to heat treatment. Conversely, when HeLa (human uterine cervical cancer) cells, originally producing an appreciable amount of enTNF, were transfected with an antisense TNF mRNA expression vector to inhibit enTNF synthesis, their heat sensitivity was enhanced. Furthermore, L-M cells which were transfected with nonsecretory human TNF expression vector also acquired resistance to heat treatment. In these cells, heat resistance correlated well with expression of enTNF and intracellular levels of manganous superoxide dismutase. These results indicate that enTNF exerts its intracellular protective effect against the heat-induced cytotoxicity by scavenging reactive oxygen with induced manganous superoxide dismutase in a manner similar to that found in cells treated with exogenous TNF.     

Development of a new perfusion system for pharmacological study on rabbit basilar arteries

We developed a constant-flow perfusion system to measure vascular responses to vasoactive agents applied intraluminally or extraluminally. The intraluminal and extraluminal sides of a cylindrical section of rabbit basilar artery were isolated completely. By loading with 0.75 g of tension, the resting condition of each preparation was made constant. The intraluminal side was perfused at a constant flow of 8 ml/min and under an intraluminal pressure of 8 mm Hg. When 30 mM KCl was administered intraluminally the preparation showed marked contraction, whereas only slight contraction was observed with extraluminal administration. When 2 x 10(-7) M 5-hydroxytryptamine was administered, no significant differences in contraction could be detected between the intraluminal and extraluminal routes. Application of 10(-6) to 10(-4) M acetylcholine after precontraction with 30 mM KCl resulted in much stronger dilatation upon intraluminal application. Thus, it was demonstrated that under certain conditions significant differences exist in the responses of rabbit basilar arteries to vasoactive agents applied intraluminally or extraluminally. This system can detect the effects of vasoactive agents administered intraluminally and extraluminally at a high level of sensitivity and shows good reproducibility as a means of analyzing vascular functions and characteristics.     

Genetic alteration in carcinoid tumors of the lung.

Surgically resected specimens of 13 carcinoid tumors of the lung including nine typical carcinoids and four atypical carcinoids, and eight salivary gland type carcinomas (six mucoepidermoid carcinomas and two adenoid cystic carcinomas) were analyzed regarding p53 expression, loss of heterozygosity (LOH) in chromosome 3p, 9p, and K-ras mutation. The overexpression of p53 was identified in four atypical carcinoid tumors, one mucoepidermoid carcinoma, and one adenoid cystic carcinoma, however, none of typical carcinoids showed p53 immunoreactivity. LOH in 3p14 was demonstrated in three of seven informative cases in all tumors. LOH in 9p was demonstrated in two of five informative cases in all tumors. Two of three cases with LOH at 3p14 had a poor prognosis, one of which also had LOH at 9p. No mutation of the K-ras gene was observed in any of these tumors. These data thus indicate that p53 overexpression might distinguish atypical carcinoid tumors from typical tumors and might therefore be useful as an adjunct modality in the differential diagnosis of pulmonary carcinoid tumors. The presence of LOH at 3p14 or 9p may thus help to identify lung cancer patients with a poor prognosis.     

In-vivo blood-brain barrier transport of a novel adrenocorticotropic hormone analogue, ebiratide, demonstrated by brain microdialysis and capillary depletion methods.

The transport of ebiratide, a novel adrenocorticotropic hormone (ACTH) analogue, [H-Met-(O2)-Glu-His-Phe-D-Lys-Phe-NH(CH2)8-NH2], through the blood-brain barrier was directly demonstrated in-vivo. [125I]Ebiratide (16.9 MBq mL-1) or [14C]sucrose (29.2 MBq mL-1) known to be restrictively transported through the blood-brain barrier was infused into the rat internal carotid artery at a flow rate of 50 microL min-1 for 10 min, and after 15 min infusion the distribution volume of each compound in the brain parenchyma was determined by the capillary depletion method. The distribution volume of [125I]ebiratide was 167.8 +/- 62.2 microL (g brain)-1, which was about seven times higher than that of [14C]sucrose (24.9 +/- 4.0 microL g brain)-1, indicating the uptake of ebiratide into brain parenchymal cells. During the infusion into the internal carotid artery, brain microdialysis was simultaneously performed to directly collect the brain interstitial fluid as the dialysate. Radioactivity was detected in the dialysate during the [125I]ebiratide infusion and HPLC analysis of the dialysate revealed that the intact ebiratide accounted for greater than or equal to 80% total radioactivity. The concentrations of [125I]ebiratide and [14C]sucrose in the brain interstitial fluid were estimated based on the relative recovery obtained in the in-vitro recovery study. The brain interstitial fluid/internal carotid arterial blood concentration ratio for [125I]ebiratide was determined to be 1.47 x 10(-2) +/- 0.17 x 10(-2) and was about eight times higher than that for [14C]sucrose (1.92 x 10(-3) +/- 0.36 x 10(-3)), indicating significant transport of ebiratide to the brain interstitial fluid.(ABSTRACT TRUNCATED AT 250 WORDS)     

MR appearance of parasymphyseal insufficiency fractures of the os pubis

 Objective. To clarify the MRI features of parasymphyseal insufficiency fractures of the os pubis. Design and patients. MRI was performed in four postmenopausal women with parasymphyseal insufficiency fractures. The diagnosis was confirmed with plain films in every patient. T1-weighted and T2-weighted images were obtained in four patients using a 1.5-T unit. Postcontrast T1-weighted imaging was also done in three patients. Results and conclusions. MRI of pubic parasymphyseal insufficiency fracture characteristically demonstrates a hyperintense mass lesion with a hypointense rim on T2-weighted imaging, showing peripheral and septal enhancement after contrast administration. It is important to have this entity in mind in patients with osteoporosis, especially in patients with a history of pelvic irradiation for malignant disease, so as not to misinterpret it as a chondroid tumor or bone metastasis.     

Species and sex differences of aflatoxin B1-induced glutathione S-transferase placental form in single hepatocytes.

Species and sex differences of aflatoxin B1 (AFB1)-induced glutathione S-transferase placental form (GST-P) positive single hepatocytes have been investigated 48 h after an intraperitoneal injection of AFB1 to young male and female Fischer rats (2 mg AFB1/kg body wt) and male Syrian golden hamsters (6 mg AFB1/kg body wt). The presence of GST-P positive hepatocytes was examined by the immunohistochemical method. Male rats formed three times as many AFB1-induced GST-P positive hepatocytes as females. Pretreatment of both male and female rats with an inhibitor of GSH synthesis, buthionine sulfoximine (BSO) (4 mmol/kg body wt), 2 h and 4 h before AFB1 injection increased AFB1-induced GST-P positive hepatocytes by about 120% above the controls. Male hamsters formed several-fold less AFB1-induced GST-P positive hepatocytes than male rats. Pretreatment with BSO did not increase AFB1-induced GST-P positive hepatocytes in hamsters even though it produced an increase in hepatic necrosis. It appears that GSH and GSH S-transferases play an important role in modulating hepatic AFB1-DNA binding and AFB1-induced GST-P positive hepatocytes in rats and hamsters.     

Effect of epidural analgesia on postoperative insulin resistance as evaluated by insulin clamp technique

The influence of epidural neural blockade on postoperative insulin resistance was studied using the euglycaemic insulin clamp technique. Eighteen patients undergoing elective upper abdominal surgery of moderate severity were allocated to two groups: group G patients underwent operation under general anaesthesia, and postoperative pain was relieved by systemic administration of analgesia; and group E patients received epidural analgesia during surgery and epidural morphine postoperatively. In each patient the euglycaemic insulin clamp test was performed twice: several days before surgery and on postoperative day 1. Peroperative catecholamine and cortisol responses were also measured to investigate possible endocrine mechanisms of the insulin resistance. Glucose disposal (M) decreased in both groups on postoperative day 1 at plasma insulin concentrations ranging from 1.2 to 10.0 milliunits ml-1, resulting in the downward shift of dose-response curves. However, this downward shift was significantly smaller in group E than in group G patients. Urinary adrenaline excretion increased markedly on the day of operation in group G, but was significantly inhibited in group E. Urinary noradrenaline excretion increased mainly on postoperative day 1 in group G, but was significantly inhibited in group E. Plasma cortisol response was lower in group E than in group G during and shortly after operation, and was significantly inhibited in group E on postoperative day 1. These results indicate that insulin resistance after elective abdominal surgery is due to a postreceptor deficit in glucose utilization, as indicated by the downward shift of the dose-response curves. This disturbance in glucose metabolism was reduced by epidural analgesia, the results being associated with inhibited catecholamine and cortisol responses.