Philip C. Kelleher

α1-Fetoprotein: Separation of two molecular variants by affinity chromatography with concanavalin A-agarose

Alpha1-Fetoprotein present in fetal/newborn rat serum and in hepatoma-bearing human serum has been resolved into two molecular variants by concanavalin A-agarose affinity chromatography. The concanavalin A-reactive variant has been purified by gel filtration column chromatography, preparative block electrophoresis and immunoadsorption affinity chromatography.

Rat alpha-fetoprotein: In vitroproduction of four molecular variants by clonal cell lines

Abstract Mass and six clonal cultures derived from Morris hepatoma 7777 by standard tissue culture techniques synthesize and secrete alpha-fetoprotein in vitro . During serial passage, the alpha-fetoprotein which accumulates in the media of these cultures contains two concanavalin A-affinity molecular variants. Each of the concanavalin A-affinity molecular variants shows two electrophoretic variants. Mass and clonal cell populations of hepatoma cells continue to secrete in vitro four molecular variants of rat alpha-fetoprotein known to occur in vivo . These results demonstrate for the first time that individual hepatoma cells have the potential to synthesize four molecular variants of alpha-fetoprotein and that this phenotypic property is maintained during serial subculture in vitro .

CONCANAVALIN‐A‐AFFINITY MOLECULAR HETEROGENEITY OF HUMAN HEPATOMA AFP AND CORD‐SERUM AFP

Glycosylation of alpha-fetoprotein (AFP) by human primary hepatocellular carcinoma (PHC) is abnormal. Concanavalin A (Con A)-affinity molecular variant patterns of serum AFP from patients with PHC are different from those of cord-serum AFP. Different patients with PHC exhibit different Con-A-affinity AFP molecular variant patterns, and the pattern remains constant over time in a given individual. The degree of deviation of the AFP molecular variant pattern from the molecular pattern of AFP secreted by neonatal liver cells is independent of the total serum AFP concentration. We propose that analysis of the AFP lectin-affinity molecular heterogeneity will improve the discrimination between malignant and nonmalignant liver disease in cases when the degree of elevation of the serum AFP concentration is nondiagnostic.

Urinary excretion of hydroxyproline, hydroxylysine and hydroxylysine glycosides by patients with paget's disease of bone and carcinoma with metastases in bone

Patients with carcinoma metastases in bone and Pagents disease of bone have different patterns of collagen metabolite excretion. Both forms of bone disease resulted in an increased excretion of total hydroxyproline and the ratio of glucosylgalactosylhydroxylsine to galactosylhydroxylysine was below normal. The excretion of glucosylgalactosylhydroxylysine and galactosylhydroxylysine was increased in all patients with carcinoma metastases in bone while the excretion of glucosylgalactosylhydroxylysine in patients with Pagets disease. The ratio of total hydroxylysine (free hydroxylysine + glycosylated hydroxylysines) to total hydroxyproline was normal in patients with carcinoma metastases in bone and below normal in patients with Pagets disease bosne. The pattern of urinary collagen metabolite excretion is a more specific indicator of the presence of bone disease than is the measurement of the excretion rate of any individual collagen metabolite. Bone diseases of different etiologies may result in different patterns of urinary collagen metabolite excretion.

The 11-S sow colostral immunoglobulin isolation and physicochemical properties

Abstract 1. 1. An immunoglobulin that is not present in porcine serum has been isolated from sow colostrum by chromatography on Sephadex G-200 and DEAE-Sephadex A-50, followed by preparative ultracentrifugation in a 10–40% sucrose density gradient. 2. 2. Sedimentation velocity studies on this purified protein yield an s ° 20, w value of 11.2 S which is similar to the s ° 20, w of human secretory IgA. 3. 3. The sow colostral 11-S immunoglobulin does not have the resistance to reductive cleavage that is a characteristic of human 11-S secretory IgA. Treatment with 0.2 M β-mercaptoethanol results in almost complete dissociation of the 11.2-S molecule to subunits with an s 20, w of 6.5 S. 4. 4. The relationship of the isolated sow colostral 11-S immunoglobulin to human serum IgA is evidenced by its immunologic reaction with specific antiserum to human IgA and by its lack of immunologic reaction with specific antisera to human IgG and IgM.

α-Fetoprotein glycosylation is abnormal in some hepatocellular carcinomas, including white patients with a normal α-fetoprotein concentration

Abstract Lectin-affinity analyses with Lens culinaris agglutinin (LCA) and other lectins have demonstrated that the glycosylation of α-fetoprotein (AFP) secreted by hepatocellular carcinomas (HCC) is frequently altered when the serum AFP concentration is increased. To determine if AFP LCA-binding properties are altered in patients with HCC whose serum AFP concentration is normal, the percentage of LCA-binding AFP in serum from white newborns, white normal adults, white patients with chronic hepatitis and hereditary tyrosinemia and white and black patients with HCC were determined. The serum LCA-binding AFP fraction was low in newborns (1–4%) and normal adults (1–8%). There was a significant increase in LCA-binding AFP in patients with chronic hepatitis (10–24%) and hereditary tyrosinemia (5–35%). The AFP LCA-binding fraction was clearly abnormal (greater than 40%) in three of the white patients with an HCC and a normal serum AFP concentration, and the range of values (10–63%) in these HCC patients was similar to that seen in both white and black patients with HCC accompanied by increased AFP concentrations.

A method for measuring hydroxylysine and glycosylated hydroxylysines in urine and protein hydrolysates

A method for measuring hydroxylysine and glycosylated hydroxylysines is described, based on the separation of the 3 compounds by ion-exchange chromatography, followed by spectrophotometric analysis of the hydroxylysine present as glucosylgalactosylhydroxylysine, galactosylhydroxylysine and free hydroxylysine. The method does not require prior preparation of the urine sample nor the use of high-resolution ion-exchange systems. The method is applicable to the determination of the glycosylated hydroxylysine and hydroxylysine content of urine or alkaline hydrolysates of proteins.

ORIGIN OF AMNIOCENTESIS‐INDUCED RISES OF ALPHA‐FETOPROTEIN CONCENTRATIONS IN MATERNAL SERUM

An elevation of alpha‐fetoprotein concentration in maternal serum occurred in 7 of 65 patients (11 per cent) following amniocentesis. Analysis of the molecular heterogeneity of the alpha‐fetoprotein by chromatography on concanavalin A‐agarose indicated that the source of the alpha‐fetoprotein was fetal serum in three patients and amniotic fluid in four patients.

Plasma fucose and sialic acid concentrations during oral glucose tolerance tests in normal and diabetic (mellitus) humans

1. 1. The plasma fucose and sialic acid concentrations were determined during oral glucose tolerance tests in normal humans and in hospitalized patients with normal and impaired glucose tolerance. 2. 2. Measurement of the plasma fucose concentration by means of a specific thin layer chromatography—isotope dilution method and the method of Dische and Shettles demonstrates that no rise in fucose levels occurs in human subjects with normal or impaired glucose tolerance during an oral glucose tolerance test. 3. 3. The plasma sialic acid concentration does not change in either normal or diabetic (mellitus) individuals during an oral glucose tolerance test. 4. 4. The apparent rise in plasma fucose concentration during oral glucose tolerance tests in patients with impaired glucose tolerance which was reported by Shaw, Kryston and Mills is not due to a rise in true plasma fucose concentration.

Prolyl hydroxylase in pulmonary alveolar macrophages

It has been reported that cell lines which did not orginate from connective tissue cells either synthesize hydroxyproline-containing polypeptides [l-6] or possess prolyl hydroxylase (EC 1.14.11.2) activity [2,3,7]. Since these cell lines had been maintained in culture for some time, the relationship of these observations to the in vivo function(s) of the cells is uncertain [ 11. Few instances of either the formation of hydroxyproline-containing polypeptides [8] or the presence of prolyl hydroxylase in homogeneous cell populations obtained directly from the intact animal have been reported [9,10]. Pulmonary lavage in humans [ 111 and rabbits [ 12,131 yields a cell population which consists primarily of pulmonary alveolar macrophages (PAM). It has been reported that PAM do not either synthesize collagen [ 141 or contain prolyl hydroxylase [7]. The present study demonstrated that rabbit and human PAM do contain significant amounts of prolyl hydroxylase activity.