Interleukin 1 activity in normal human urine

Human leukocyte dialysates contain components capable of amplifying cutaneous delayed-type hypersensitivity (DTH) reactions. In the present study, two such amplifiers, both less than 3500 m.w., were partially purified from human leukocyte dialysates by gel filtration on Sephadex G-10 followed by high pressure reverse-phase liquid chromatography. These amplifiers of DTH were examined for their effects on production of the migration inhibitory lymphokines leukocyte migration inhibition factor (LIF) and macrophage migration inhibition factor (MIF). The amplifiers were found to increase LIF and MIF production by antigen- or alloantigen-stimulated human peripheral blood lymphocytes in a dose-dependent fashion. Further analysis demonstrated that although antigen-stimulated T4 and T8 cell subpopulations could produce LIF activity under the assay conditions employed, amplification of lymphokine production by modulator was only observed with the T4 subset.     

Proteomic characterization of serine hydrolase activity and composition in normal urine

Background

Serine hydrolases constitute a large enzyme family involved in a diversity of proteolytic and metabolic processes which are essential for many aspects of normal physiology. The roles of serine hydrolases in renal function are largely unknown and monitoring their activity may provide important insights into renal physiology. The goal of this study was to profile urinary serine hydrolases with activity-based protein profiling (ABPP) and to perform an in-depth compositional analysis.

Methods

Eighteen healthy individuals provided random, mid-stream urine samples. ABPP was performed by reacting urines (n = 18) with a rhodamine-tagged fluorophosphonate probe and visualizing on SDS-PAGE. Active serine hydrolases were isolated with affinity purification and identified on MS-MS. Enzyme activity was confirmed with substrate specific assays. A complementary 2D LC/MS-MS analysis was performed to evaluate the composition of serine hydrolases in urine.

Results

Enzyme activity was closely, but not exclusively, correlated with protein quantity. Affinity purification and MS/MS identified 13 active serine hydrolases. The epithelial sodium channel (ENaC) and calcium channel (TRPV5) regulators, tissue kallikrein and plasmin were identified in active forms, suggesting a potential role in regulating sodium and calcium reabsorption in a healthy human model. Complement C1r subcomponent-like protein, mannan binding lectin serine protease 2 and myeloblastin (proteinase 3) were also identified in active forms. The in-depth compositional analysis identified 62 serine hydrolases in urine independent of activity state.

Conclusions

This study identified luminal regulators of electrolyte homeostasis in an active state in the urine, which suggests tissue kallikrein and plasmin may be functionally relevant in healthy individuals. Additional serine hydrolases were identified in an active form that may contribute to regulating innate immunity of the urinary tract. Finally, the optimized ABPP technique in urine demonstrates its feasibility, reproducibility and potential applicability to profiling urinary enzyme activity in different renal physiological and pathophysiological conditions.     

Radioimmunological quantitation of the urinary trypsin inhibitor in normal blood and urine

A radioimmunoassay for measurement of the urinary trypsin inhibitor in human serum and urine is described. Because of the immunological cross-reactivity between the inter-alpha-trypsin inhibitor and the urinary trypsin inhibitor the plasma and serum were treated with perchloric acid to precipitate the inter-alpha-trypsin inhibitor. Gel filtration of serum before and after acid treatment showed identical peaks corresponding to the urinary trypsin inhibitor. The normal level of the urinary trypsin inhibitor in fresh plasma from 30 blood donors was 6.38 +/- 0.33 mg/l (SEM), and in sera from 24 healthy volunteers 7.14 +/- 0.27 mg/l (SEM). In urine from 23 healthy volunteers the normal excretion was 8.17 +/- 1.18 mg/24 h (SEM).     

Comparative study of alanine-amino-peptidase and gamma-glutamyl-transferase activity in normal Wistar rat urine

24 h urinary alanine-amino-peptidase (AAP) and gamma-glutamyl transferase (GGT) activities were studied from the 3rd-7th month of life in male Wistar rats. A close relationship was found between AAP and GGT activity, except at the beginning and at the end of this period. At the end there appear 2 subgroups, the larger (70%) showing a strong correlation between AAP and GGT activity and the other (30%), demonstrating no correlation at all. A good correlation between AAP and GGT activities, creatinine, 24 h urine volume and 24 h creatinine output was found.     

Relation between the level of conditioned reflex activity and concentration of serotonin in the blood of normal and neurotic dogs

Higher nervous activity of dogs was studied using Pavlovian method of alimentary salivary conditioned reflexes. Serotonin (5- hydroxytryptamine) content in the venous blood was defined by means of fluorescent spectrofluorimeter using orthophthalic dialdehyde. A high level of conditioned and unconditioned reflexes was accompanied by a low serotonin blood content, while a low level of reflexes was accompanied by a high serotonin content. Negative correlation is supposed to exist between the degree of activation of nervous processes and blood serotonin content. Blood serotonin content increased in dogs in experimental neurosis with expressed reduction of the level on alimentary conditioned and unconditioned reflexes.