Human tryptase fibrinogenolysis is optimal at acidic pH and generates anticoagulant fragments in the presence of the anti-tryptase monoclonal antibody B12

Human tryptase is uniquely regulated by its association with heparin and resists inhibition by biological protease inhibitors. The effects of pH and B12, an IgG anti-tryptase mAb, on cleavage of the synthetic substrate tosyl-Gly-Pro-Lys-p-nitroanilide and of the biological substrate fibrinogen by tryptase were examined. Tosyl-Gly-Pro-Lys-pnitroanilide cleavage was optimal at neutral pH and was inhibited by the B12 mAb at acidic and neutral pH values. At pH 7.5, inhibition was reversible and noncompetitive. In contrast, the optimal pH for tryptase to cleave fibrinogen was acidic. B12 dramatically enhanced the rate and extent that tryptase cleaved all three fibrinogen subunits at pH 6.0 to 6.5, but inhibited these activities at neutral pH. Major fibrinogen cleavage fragments generated at acidic pH by the B12:tryptase complex were identical with those made by plasmin. Thus, at acid pH, tryptase alone destroyed the ability of fibrinogen to clot, while the B12:tryptase complex increased the rate of fibrinogenolysis and also generated the anticoagulant, fragment D. The acidic pH optimum for tryptase fibrinogenolysis may direct this activity to tissue sites of inflammation. A putative biological equivalent to B12 would limit tryptase fibrinogenolytic activity at sites of neutral pH, such as blood, but would augment activity at acidic sites.     

Synthesis and evaluation of novel rhodacyanine dyes that exhibit antitumor activity

Rhodacyanine dyes and several analogous delocalized lipophilic cations (DLCs) were synthesized and evaluated as novel antitumor agents. Rhodacyanine dye consists of two heteroaromatic rings such as thiazoles at both termini of the conjugate systems and 4-oxothiazolidine (rhodanine) in the middle of it. Compounds with such a unique double-conjugate structure were found to inhibit the growth of several tumor cell lines, such as colon carcinoma CX-1, and to exhibit relatively low toxicity against normal kidney cell line CV-1 (e.g., IC50(CX-1) = 50 nM, IC50(CV-1) = 17.3 microM; selectivity index = 346 for compound 5). These compounds were also found to be efficacious in the tumor-bearing nude mice model (e.g., against human melanoma LOX; T/C (%) = 168 for compound 5). Structural modifications on rhodacyanine, including deletion of a heteroaromatic ring involved in the merocyanine conjugate system and replacement of rhodanine with a structurally related moiety such as 4-oxoimidazolidine or 4-oxo-1,3-dithiolane, resulted in a loss of the selectivity and/or the activity. Our current structure-activity studies imply that the double-conjugate system with a rhodanine moiety is essential for the selective activity of rhodacyanine dyes, and we find this class of compounds as unique antitumor agents candidates.     

Severe malformation of one foot from amniocentesis needle injury

An 11-year-old boy is described who was born with a poorly developed right foot. At 16 weeks gestation his mother had had amniocentesis without direct ultrasound guidance. After the insertion of the amniocentesis needle, she felt strong abdominal resistance, which disappeared with slight withdrawal of the needle. Then, real-time ultrasound was used to determine the position of the needle, which was reported to be up against the feet. Nine days later deep purple amniotic fluid was removed by amniocentesis. At birth there was a scab over an oozing hole in the lateral aspect of his right 'foot', a poorly formed structure with five digit-like structures at the tip. It seems most likely that the deformity was caused by tissue injury from needle puncture at 16 weeks gestation.     

Leiomyosarcoma of the kidney in an HIV-infected child

Spindle cell tumors (leiomyoma and leiomyosarcoma) have been described in immunocompromised children after transplantation and in association with HIV infection. Previous reports have described tumors of respiratory (lung, bronchi), gastrointestinal (stomach, bowel, liver), and subcutaneous origins. We report a case of leiomyosarcoma of the kidney in an HIV-infected child.     

Effect of sodium bicarbonate ingestion on exhaustive resistance exercise performance

Six weight trained males were studied prior to, during, and in recovery from exhaustive resistance exercise, 105 min after ingesting 300 mg.kg-1 of either a placebo or NaHCO3. The exercise test consisted of four sets of 12 repetitions with a fifth set to volitional fatigue on a Universal leg press machine at a resistance equaling approximately 70% of the subjects 1-repetition maximum. Arterialized venous blood was analyzed for lactate concentration, blood gas, and acid-base parameters. The ingestion of NaHCO3 produced a significant increase in resting pH (7.39 to 7.46), HCO3- (22.9 to 28.3 mEq.l-1), and oxygenated base excess (-1.3 to 4.4 mEq.l-1). With the completion of each exercise set, a progressive decline in the acid-base status of both groups was observed (pH set 1-5: NaHCO3, 7.40 to 7.31; placebo, 7.34 to 7.25; HCO3- set 1-5: NaHCO3, 25.3 to 17.9; placebo, 21.7 to 15.3 mEq.l-1; base excess set 1-5: NaHCO3, 3.7 to -7.1; placebo, -1.4 to -10.7 mEq.l-1); however, the NaHCO3 condition was significantly more alkaline than the placebo condition. Blood lactate concentration [La] progressively increased with the completion of each exercise set ([La] set 1-5: NaHCO3, 1.37 to 11.15; placebo, 1.31 to 9.81 mM); but were not significantly different between treatments. Repetitions performed in the final exercise set were not significantly different between groups (NaHCO3: 19.6 +/- 1.6, placebo: 18.2 +/- 1.1 repetitions).(ABSTRACT TRUNCATED AT 250 WORDS)     

Electron microscopic and immunomorphologic study of the placenta in genital mycoplasmosis]

The authors have made an analysis of 1500 operations using a unique technology of closure without perfusion of the interventricular septum defects of the heart under conditions of hypothermia. Lethality was 1.7%, frequency of recanalization of the defects was 0.9%.     

Regional lymph node metastasis from cutaneous squamous cell carcinoma

The phenotypes of various STAT knockout mice reveal an unexpected specificity in the biological roles of these molecules. The mechanisms involved in generating selectivity and modulating STAT activity have been the focus of intense studies. This work has led to the discovery of novel families of proteins that regulate Jak-STAT signaling. Recently, the structures of a STAT dimer/DNA complex and of the amino-terminal domain have been solved, providing new insights into the function of these versatile proteins.     

Trauma and posttraumatic stress disorder in severe mental illness

The objective of this study was to evaluate the performance of a new, compact, dynamic diffusion cell for in vitro transdermal permeation. These so-called Kelder-cells were developed as an automated alternative to the static Franz diffusion cells. The new cells were used in combination with the ASPEC-system (automatic sample preparation with extraction columns) which was initially designed for the automation of solid-phase extractions. Three variables were tested to optimize the performance of the new cell system: injection height into the inlet compartment, volume flowing through the receptor compartment and temperature. Experiments were performed using the tritium labelled anticholinergic [3H]dexetimide permeating through an artificial membrane (Silastic). The injection height of the needle into the inlet compartment of the cell should be programmed at -34 mm to ensure complete air tightness, thus forcing the buffer to flow through the cell. The volume of buffer flow through the receptor compartment is important in maintaining sink conditions: a volume of 117 microliters was chosen to replace the total content of the cell (84 microliters) every 2 min. The temperature was precisely controlled in a thermostatic cabinet to minimize variations in experimental conditions. For [3H]dexetimide, an increase in temperature of 20 degrees C reduced the lag time by a factor of approximately two, however the influence on the flux was negligible. The data for the Kelder-cells were comparable with static Franz diffusion cells at a pseudo-steady state, however Kelder-cells have the advantage of automatic sampling, continuous replacement of the receptor solution, and unattended operation over at least 24 h.     

Brain-derived neurotrophic factor stimulates hindlimb stepping and sprouting of cholinergic fibers after spinal cord injury

Neurotrophic factors have been proposed as a therapeutic treatment for traumatic brain and spinal cord injury. The present study determined whether exogenous administration of one such factor, brain-derived neurotrophic factor (BDNF), could effect behavioral recovery and/or histopathological changes after spinal cord injury. Adult rats received a mild or moderate contusion injury or complete transection of the mid-thoracic spinal cord. Immediately thereafter, they were infused intrathecally with vehicle or BDNF for 28 days. Behavioral recovery was evaluated for 6 weeks after injury, at which time the rats were sacrificed and the spinal cord tissue was examined histologically. The infusion of BDNF resulted in acute stimulation of hindlimb activity. These effects included activation of alternating airstepping in injured rats when the hindlimbs were unloaded as well as slight improvements in the rate of recovery in open field locomotion score. BDNF infusion was also associated with enhanced growth of cholinergic fibers at the injury epicenter, but did not affect white matter sparing or density of serotonergic axons at or below the injury site. Based on immunohistochemical detection of BDNF protein distribution, these described effects are likely to be mediated by the activation of cells and axons within the central injury region and the along the peripheral rim of the spinal cord. Together, these findings demonstrate that the exogenous infusion of BDNF after spinal trauma can influence postinjury outcome through mechanisms that include acute stimulation of hindlimb activity and neuritogenesis at the injury site.     

Brain sites involved in the antinociceptive effect of bradykinin in rats

The localization of brain sites where bradykinin (BK) induces its antinociceptive effect in rats, was studied using as index the threshold for the jaw-opening reflex elicited by the dental pulp electrical stimulation test (DPEST). The microinjection of BK into the lateral or fourth cerebral ventricles induced an antinociceptive effect, with Index of Antinociception (IA) of 0.51+/-0.03 and 0.68+/-0.05, respectively. However, microinjections of the peptide into the third ventricle induced a less marked antinociception (IA = 0.28+/-0.08). The brain sites where the microinjection of BK caused an antinociceptive effect were: locus coeruleus, principal nucleus, oral part of the spinal sensorial trigeminal nucleus, and the sensory root of the trigeminal nerve. The antinociceptive effect was more intense when BK (4-16 nmol) was injected into the locus coeruleus. Microinjection of BK (4 nmol) into the fourth ventricle, but not into the locus coeruleus, induced an increase in blood pressure. The microinjection of the peptide into the nucleus tractus solitarius, a site that is also involved in the pressor effect of BK, did not induce an antinociceptive effect. These results indicate that the antinociceptive effect of BK is not related to blood pressure changes. The microinjection of BK into some of the sites involved in the mechanisms of analgaesia, including the periaqueductal gray matter (dorsal, lateral and ventrolateral) and the dorsal raphe nucleus did not induce an antinociceptive effect. The results suggest that the most likely brain sites involved in the antinociceptive effect of BK are the locus coeruleus and the principal sensory trigeminal nucleus. The present results did not exclude the involvement of other brain sites surrounding the lateral and the third ventricles.