Immunopurification of human plasma kininogens

Human plasma kininogens were purified by immunoadsorption on Sepharose columns using two different approaches, either removing protein impurities with the respective immunospecific polymers or applying an anti-kininogen-specific immunoadsorbent column. An anti-kininogen serum developed and investigated in this laboratory in earlier studies was used. This antiserum recognizes the native conformational determinants in the kininogen heavy chain, the common denominator in plasma kininogens, and reacts with three heterogeneous molecular forms of high mol. wt kininogen (mol. wts 103,000, 92,000 and 90,000) as well as with low mol. wt kininogen. Heterogeneity of kininogens was shown by SDS gel electrophoresis and immunoelectrophoresis. With the antibody-specific polymers the yield was 80-100% compared to 75% or lower when several consecutive immunoadsorption steps were applied to remove impurities. Both methods serve the purpose of preparing immunologically pure kininogens suitable for immunization.     

Rat brain synthesizes two ‘vitamin D-dependent’ calcium-binding proteins

Two proteins from rat brain reacting against anti-chick intestinal vitamin D-dependent calcium-binding protein were characterized in terms of their mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and their molecular size. The proteins were present in the isolated cytoplasm and were produced following translation of brain mRNA in the rabbit reticulocyte lysate system. Their apparent molecular weight was 29,000 and 27,000 daltons whereas rat kidney contained only one protein cross-reacting with this antiserum and with a molecular weight of 27,000 daltons.     

Studies of phenylalanine hydroxylase activity in patients with chronic renal failure: the effect of haemodialysis

The effect of serum from patients with renal failure on phenylalanine hydroxylase activity has been measured in normal individuals, patients with steady-state chronic renal failure and patients undergoing haemodialysis. Significant inhibition of enzyme activity was induced by serum from patients with steady-state chronic renal failure but not from patients undergoing haemodialysis. Inhibitor(s) was readily diffusible in vitro and appeared to have an approximate molecular mass of 800. The results suggest that the low-plasma tyrosine levels observed in patients with chronic renal failure are due, at least in part, to the inhibition of phenylalanine hydroxylase.     

Immuno-suppressive lymphocyte factors—III. Complete purification and partial characterization of human inhibitor of DNA synthesis

Inhibitor of DNA synthesis (IDS) is an immunoregulatory T lymphocyte factor first characterized in the rat. It appears to cause the non-specific immune suppression seen in certain tolerant disease states.We have produced IDS from human peripheral blood lymphocytes, purified it to homogeneity and partiallycharacterized it. Human IDS has an isoelectric point of 3.40 and a molecular weight of 20,000. However, in the supernatants containing IDS activity, it usually exists in an aggregated tetrameric form which is also biologically active. IDS is not cytotoxic and is glycoprotein in nature. The activity depends on an intact carbohydrate moiety. Human IDS is identical to rat IDS except for a slightly higher isoelectric point. The biologic role is undetermined at present, but might be similar to that of rat IDS.     

Modification in messenger RNA population synthesized in rabbit cerebral cortex in the recovery period following convulsions

The labeling pattern of microsomal poly(A)-associated messenger RNA was studied in rabbit cerebral cortex during the postconvulsive period. Labeled uridine was injected into the brain 2 hr after a single electroconvulsive shock and the animals sacrificed 1 hr later. Poly(A) RNA was then extracted from cerebral cortex microsomes, and fractionated by gel electrophoresis. The results indicate that the messenger RNA population synthesized in the cortex in the postconvulsive recovery period is clearly different from that synthesized in the normal controls.     

ISolation of catfish proinsulin messenger rna and synthesis of its complementary DNA

The purpose of these experiments was to obtain proinsulin mRNA from catfish pancreatic islets and synthesize its cDNA. Poly(A)-rich mRNA was electrophoresed on preparative agarose-urea gels. One RNA fraction was obtained which translated predominantly preproinsulin. This mRNA was estimated to be approx. 210 000 M_r (650 nucleotides) when electrophoresed under denaturing conditions. [³H]Proinsulin cDNA was hybridized to excess RNA to monitor purification of mRNA from total islet RNA. Greater than 94% of proinsulin messenger contained poly(A) sequences. [³H]Proinsulin cDNA hybridized to its template mRNA with a $\text{Rot}\text{1}\text{2}$ of 4.4 × 10^?3 mole · sec/1. The overall purification was 80-fold by this type of analysis. Thermal denaturation studies indicated a high degree of fidelity of hybrid formation between [³H]proinsulin cDNA and proinsulin mRNA. Proinsulin comprised 20% of total islet protein when synthesis was measured in vivo (Albert and Permutt, 1979) and 12–20% when total islet mRNA was translated in a cell-free system. Using the [³H]proinsulin cDNA probe it was estimated that proinsulin mRNA accounted for approx. 15% of total islet mRNA.     

Polyurethane-covered silicone gel mammary prosthesis for successful breast reconstruction

A polyurethane-covered silicone gel implant has been used by the author in 150 breast reconstructions and augmentations in the past 9 years. The results have been most gratifying with regard to breast softness, breast compressibility, and esthetics. Only 4 patients have developed a unilateral capsule contracture and firm breast. The reasons are postulated for these satisfying results. Complications have been few and very minor.     

Micro Disc Gradient Gel Electrophoresis of Seminal Plasma Proteins before and after Vasectomy and in Different Split Ejaculate Fractions

Mikro-Disk-Gradienten-Gelelektrophorese von Spermaplasmaproteinen vor und nach Vasektomie sowie in verschiedenen Split-Ejakulat-Fraktionen
Mit Hilfe der Polyacrylamidgelelektrophorese am kontinuierlichen Gradientengel wurden Spermaplasmaproteine von Patienten vor und nach Vasektomie sowie unter Verwendung der Splitejakulattechnik untersucht. Es wurden beträchtliche interindividuelle Schwankungen des Proteinmusters beobachtet. Nach Vasektomie fiel das Verschwinden einer Proteinbande im hochmolekularen Bereich auf. Diese Proteinbande konnte bei Split-ejakulaten jeweils im Bereich der spermatozoenreichen Fraktion lokalisiert werden. Das nachgewiesene Protein stammt offenbar aus dem Hoden-Nebenhodensekret und könnte zur funktionellen Charakterisierung des Hoden-Nebenhodensekretes und für den Nachweis von Samenwegsverschlüssen Bedeutung gewinnen.     

Function of Culturing Monolayer Hepatocytes by Collagen Gel Coating and Coculture with Nonparenchymal Cells

Abstract: Since 1987, we have been developing a bioartificial liver (BAL) using multiplated cultured hepatocyte monolayers. With the goal of promoting hepatic functions of cultured hepatocyte monolayers, we combined the use of a collagen gel layer over the monolayers of hepatocytes and/or cocultured hepatocytes with nonparenchymal cells (NPCs). The study was divided into four groups according to culture configurations: Group 1: hepatocyte monolayer culture (control); Group 2: coculture of hepatocytes and NPCs; Group 3: hepatocyte monolayer with a overlaid collagen gel layer; and Group 4: coculture with a overlaid collagen gel layer. The culture continued for 14 days. Morphological changes and hepatic functions were evaluated by urea and albumin syntheses. The morphological status of the hepatocytes remained for 2 weeks in Groups 3 and 4. Deterioration and detachment of hepatocytes and/or NPCs started in Group 1 and 2 on the third day in culture. Significantly high urea synthesis was noted in Group 4 (p < 0.001 compared with Group 1 and 2: p = 0.0014 compared with Group 3). Although there was no significant difference in albumin synthesis among the four groups, those hepatocytes covered by the collagen gel (Groups 3 and 4) tended to secrete albumin throughout the observation period. These results indicted that the environment, although artificial (but close to the in vivo state), supplied with collagen gel and the coculture, enhanced the activities of the cultured hepatocyte monolayers. We suggest that use of cocultured hepatocytes under a collagen gel is a promising candidate for a bioreactor of multiplated BAL.