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Higher serum iron and ferritin levels noted in hepatitis B antigen (HBAg) carriers than in noncarriers suggests that virus might actively replicate in hepatocytes, stimulate ferritin synthesis, and result in increased liver iron stores. A comparative semiquantitative study of immunohistochemical ferritin (0-12) and hemosiderin (0-9) was performed on 54 normal, 13 cirrhotic, and 70 nonneoplastic livers from patients with hepatocellular carcinoma, in each group, comparing amounts in HBAg-positive and HBAg-negative patients. Mean scores for ferritin and hemosiderin were high in all three groups, normal livers averaging 8.3 and 6, respectively, cirrhotic livers, 8.5 and 7.4, respectively, and carcinoma livers, 5.6 and 6.1, respectively. In each group, there was no significant difference in ferritin and hemosiderin mean scores in HBAg-positive and HBAg-negative patients. The large liver iron stores do not appear to be a consequence of hepatitis B virus infection alone. Their role in the development of hepatocellular carcinoma is still to be elucidated.

An assessment was made of the frequency of liver cell dysplasia and the mean age of each group in 56 normal, 13 cirrhotic, and 50 hepatocellular carcinoma (HCC) patients, 40 with cirrhosis, from southern Africa. Dysplasia increased from 7.1% in normal subjects to 38.5% in cirrhotic, 40% in noncirrhotic HCC, and 52.5% in cirrhotic HCC patients, three statistically similar frequencies. Average patient ages were as follows: patients with normal livers, 37.3 years; with cirrhosis, 42.4 years; with noncirrhotic HCC, 36.5 years; and with cirrhotic HCC, 34 years, the mean age with dysplasia being lower than that of the whole group. With no increase in frequency of dysplasia from cirrhosis to HCC with cirrhosis, with a similar high frequency in HCC without cirrhosis, and with a mean age of all HCC patients 8 years less than that of cirrhotics and 3 years less than normals, chronologic evolutionary progression from cirrhosis to dysplasia to HCC in southern Africa cannot be demonstrated.

Three commercial Na125I solutions (Amersham, New England Nuclear, and Union Carbide) have been examined with respect to multiple parameters affecting their use in the radioiodination of three representative peptides (insulin, growth hormone, and gastrin): % of radioiodine incorporation in protein; immunoreactivity and non-specific binding properties of the radiolabeled proteins; pH, volatility, and radionuclidic purity of radioiodine solutions; and vial construction with respect to multidose use. All three commercial Na125I produced radioiodinated proteins of good quality for use in radioligand assays. The radioiodines differed with respect to the amount of iodine released during initial vial opening as a consequence of different pH levels: 15 nCi/mCi (pH 12.5) to 1.0 microCi/mCi (pH 7.5). Two of the three products were shipped in vials with poor construction with respect to multidose use. Selection of a radioiodine was therefore reduced to the secondary considerations of iodine volatility and vial construction. The volatilized radioiodine observed during the spill of millicuries quantities of unbuffered pH 7.5 Na125I was 14 microcuries per millicurie within the first 30 minutes. One thickness of rubber gloves reduced potential skin contamination from an accidental spill to insignificant levels: 20-30 picocuries per microcurie. Common good housekeeping procedures: i.e. rubber gloves, laboratory coat and a fume hood were found to be sufficient protection to eliminate most radioiodine volatility and contamination hazards associated with protein radiolabeling procedures.

Serum ferritin concentrations are elevated in 35% to 100% of patients with hepatocellular carcinoma (HCC). With an immunoperoxidase technique, ferritin was demonstrated in tumor tissue from 32 of 74 (43%) black southern African patients, and from 12 of 19 (63%) American patients with HCC (P greater than 0.1). Ferritin was present in nonneoplastic liver in 82% of African and 100% of American patients (P greater than 0.1). Moderate to large amounts of stainable hepatic storage iron (hemosiderin) were present in 76% of African and 67% of American patients (P greater than 0.1). Fifty-two (70%) African patients had macronodular cirrhosis. In the literature, 80% to 90% of American patients with HCC have cirrhosis. High serum ferritin levels in patients with HCC may be due to ferritin production by the tumor, or related to the associated iron overload and/or cirrhosis.

The lack of sufficient quantities of human parathyroid hormone (PTH) for antibody production is the single most important factor responsible for the slow progress in the development of ideal radioimmunoassays of human PTH in plasma. Use of an assay which employs an antibovine PTH antiserum, however, has proved applicable to clinical problems of parathyroid dysfunction. Further progress in the characterization of heterogeneous forms of plasma PTH has been prevented by the great technical difficulties encountered in the fractionation and separation of low concentrations of PTH from plasma in systems using molecular sieve technics or zone electrophoresis.

THIRTY years ago Marine and Baumann (1) noted that adrenal insufficiency in rabbits led to an increase in heat production and respiratory exchange and to the appearance of a symptom complex resembling exophthalmic goiter. Marine (2) subsequently reviewed the evidence pointing to the importance of a primary adreno-gonadal insufficiency as a possible fundamental cause of Graves' disease. The occurrence of Graves' disease following roentgen ray damage to the adrenal cortex in 2 cases reported by Oppenheimer (3) provided a direct clinical example in support of this concept. The pathologic changes observed in patients who have died of thyrotoxicosis—enlargement of the thymus, diffuse lymphoid hyperplasia and small adrenal cortices—have been cited as evidence suggestive of adrenal cortical insufficiency (2). The endocrine antithesis to the hyperthyroid-hypoadrenocortical state is well demonstrated by the frequent occurrence of an atrophied thyroid gland in Cushing's syndrome (4). The high concentration of serum...

Summary and Conclusions1. The insulin content of each of 2 crystalline glucagon preparations was found to be less than 0.05% by immunochemical assay. 2. Immunologically reactive insulin was not detected in a cysteine treated glucagon preparation.

Electrophoretic and filtration analysis of immunoreactive plasma gastrin extends our previous finding of two immunoreactive components of plasma gastrin. One component has the characteristics of Gregory-Tracy heptadecapeptide gastrin; the other component has a less acidic charge than heptadecapeptide gastrin but emerges from a Sephadex gel column in a zone consistent with a molecular weight of about 7000, which is 3 1/2 times the molecular weight of heptadecapeptide gastrin. The larger, more basic component (BG) usually represents the major fraction of plasma gastrin; the heptadecapeptide-like component (H-LG) is virtually undetectable in some samples. However, only (H-LG) was found in the urine of a patient whose plasma contained nearly equal amounts of both components. There is no spontaneous interconversion of the two components in vitro, and on refractionation each component retains its integrity. However, trypsin converts BG into an immunoreactive component with characteristics similar to those of heptadecapeptide gastrin. The separated plasma components individually are indistinguishable from each other and from heptadecapeptide porcine gastrin I in their immunochemical reaction with guinea pig antiporcine gastrin I antibodies. The secretion of both components is stimulated by feeding.

Plasma gastrin concentrations were determined by radioimmunoassay in 8 subjects given a test meal with and without prior atropinization. Gastrin levels increased in all subjects following feeding. Integrated increments in plasma gastrin responses over the 2-hr period following feeding were always greater under atropine than in the control studies. Particularly marked differences between atropine and control studies were observed in 3 patients with duodenal ulcer. One patient with pernicious anemia and high fasting plasma gastrin concentrations showed no significant sustained changes in concentration in the fasting state over a 2-hr period following atropine administration. It is concluded that the tonic release of gastrin in patients with anacidity is atropine-resistant and that the reduction in gastric acid secretion produced by atropine in acid-secreting subjects serves to blunt the normal inhibitory effect of antral acidification on gastrin secretion, thereby overcoming any possible atropine blockade of gastrin release in response to feeding.

A HYPOPARATHYROID subject acquired resistance to the effects of parathyroid extract (parathyroid, U.S.P.) during treatment. Resistance developed in association with specific antibody detected as gamma globulin with affinity for binding 131I-labeled parathyroid hormone in soluble complexes. The findings are presented below. Case Report A.S. (04–75–11), a 39-year-old woman, first noted thyroid enlargement in 1946. In 1954, the thyroid gland began to increase further in size; she became nervous, and lost 4.5 kg. (10 pounds) in weight. A subtotal thyroidectomy was performed in February, 1955; on the 3d or 4th postoperative day she complained of tingling of the lips and . . .

Gastric acid secretion and plasma gastrin concentrations were determined during fasting and following sham feeding or feeding in dogs with Pavlov pouches or gastric fistulae. Sham feeding caused immediate elevations of plasma gastrin concentrations that rapidly returned to basal levels following the onset of acid secretion although acid was continuously secreted for several hours. Sham feeding with the gastric fistula open produced higher gastrin concentrations and acid output than with the fistula closed. Feeding raised the plasma gastrin concentrations considerably higher than sham feeding in the same dogs although the peak acid output equaled that following sham feeding with gastric fistula open. The delay in onset of acid secretion was more pronounced during feeding than during sham feeding. Atropinization did not significantly influence basal plasma gastrin concentration but virtually abolished the gastrin response to sham feeding. Plasma gastrin responses were observed following feeding despite atropinization.

Ten patients with advanced Hodgkin's disease were treated with cortisone for periods varying between two weeks and three months. There was considerable but transient subjective improvement in seven. Fever subsided to normal in three. Three patients became worse under therapy, all of whom died. In none was the unfavorable course attributable to the hormone. In only one instance was significant regression of nodal or parenchymal involvement seen. In another patient considerable diminution of a postauricular mass occurred under cortisone therapy twenty-five days after a course of nitrogen mustard. The role of cortisone in this situation is debatable. Studies of total body potassium with K showed a significant decrease in four of six patients which was not reflected by the plasma potassium level. Depression of thyroid activity was found in five of six patients as manifested by decrease in radioiodine uptake. Serial studies of the bone marrow regularly showed a stimulation of marrow elements by cortisone with marked increase in cellularity by the twentieth day of treatment. This was not always correlated with the findings in the peripheral blood. Little effect was noted on eosinophils, blood volume or carbohydrate metabolism as a result of cortisone. The sedimentation rate fell in five patients in whom serial observations were made. Relapses following reduction in dosage to 300 mg. weekly occurred in three patients in whom this was attempted. All three regained the previous limited improvement on restoration of 100 mg. daily dosage. Undesirable effects were edema in five and rounding of the face in eight, acne in three, weakness in three following reduction in dosage, and hypertension in two. Generalized convulsions occurred in one. Gastrointestinal bleeding from a gastric ulcer due to Hodgkin's infiltration occurred in one. Another patient at autopsy eight days following cessation of cortisone therapy had perforated gastric and duodenal ulcers. He had had no symptoms referable to his gastrointestinal tract either before or after the perforation, unlike the case reported recently by Habif et al. We conclude from these observations that cortisone produces symptomatic improvement in some cases of advanced Hodgkin's disease. Other than reduction of fever and sedimentation rate, objective changes are few. There is little or no effect on the neoplastic process. True remission was not achieved with this hormone in any of the ten patients treated.

SummarySerum albumin, serum γ globulin, crystalline glucagon and crystalline insulin were observed to bind to diaphragm in vitro and to be eluted slowly on continuous washing with buffered solutions. A lack of reproducibility of binding of insulin at low concentrations is attributable in part to variable adsorption to glassware. A definite saturation of binding sites on diaphragm was not observed at the highest concentrations employed. Competitive inhibition was observed in the presence of heterologous proteins. Binding capacity was not altered by treatment of diaphragms with formaldehyde. Binding of insulin by isolated rat diaphragm in vitro is not demonstrably of biologic significance but is attributable to nonspecific surface adsorption of proteins.

A high degree of specificity and exquisite sensitivity combine to give radioimmunoassay its great value in detection of many endogenous hormones, among them insulin, growth hormone, parathyroid hormone, ACTH, chorionic gonadotropin, and follicle stimulating hormone. The authors, who pioneered development of the approach, discuss its present and future place in clinical and investigational medicine.

Intravenous glucose and tolbutamide tolerance tests were performed in six nonobese, nondiabetic control subjects, in six untreated nonobese maturity-onset diabetic patients without pancreatic diabetes and in one insulin-non-requiring patient with diabetes secondary to chronic pancreatitis. Following injection of glucose the control group responded with a sharp rise in plasma insulin from a fasting level of 8 ± 2 μU/ml. to a peak value of 216 ± 29 μU/ml. at 2 minutes whereas the maturity-onset diabetics showed a very slight increase in plasma insulin from a fasting level of 19 ± 4 μU/ml. to 24 ± 10 μU/ml. at 2 minutes and a peak value of 35 ± 9 μU/ml. at 30 minutes. In contrast initial insulin secretory responses of both groups to intravenous tolbutamide were essentially the same with peak values of 69 ± 1 μU/ml. and 72 ± 20 μU/ml. being obtained at 2 minutes from fasting levels of 6 ± 1 μU/ml. and 13 ± 5 μU/ml. in the control and diabetic groups, respectively. The response of the maturity-onset diabetics was biphasic and there appeared to be a sustained insulin release for over 30 minutes. The subject with pancreatic diabetes had virtually no response to either glucose or tolbutamide. The rate of disappearance of crystalline beef insulin from the plasma of two subjects with maturity-onset diabetes, two control subjects and the subject with pancreatic diabetes was about the same for all, with half-times ranging from 9–11 minutes after the initial distribution phase. The differences in plasma insulin concentration therefore reflect differences in secretory rate rather than differences in metabolic turnover. The possibility is suggested that the early lesion in β cell function in the maturity-onset diabetics studied here is expressed specifically in a failure of insulin release to be triggered promptly by glucose rather than in a generally defective mechanism for synthesizing and storing insulin.

The preparation of I131-labeled human insulin from a lot of human insulin containing approximately 25% insulin by weight and its purification from labeled contaminants are described. The reaction of human insulin-I133 with insulinbinding antibodies in the serums of human subjects treated with commercial mixtures of animal insulins is demonstrated directly. Comparison of the binding of human insulin and beef insulin in antiserums from eight insulinresistant and nonresistant diabetic subjects revealed a lesser affinity' of antibody for human than for beef insulin in most cases, but considerable variability in this respect was encountered among different antiserums.

Extracts of mucosa from human antrum, duodenum, and proximal jejunum obtained at surgery and at autopsy contain two immunoreactive components of gastrin, separable on the basis of size and charge. These components exhibit the same characteristics as the two plasma gastrin components identified earlier. The major plasma component (BG) has a less acidic charge and larger molecular size than heptadecapeptide gastrin; the minor plasma component (H-LG) resembles heptadecapeptide gastrin in both respects. H-LG predominates in most extracts of antral mucosa, but the relative abundance of BG increases distally, being the only detectable component (although at very low concentration) in proximal jejunal mucosal extracts. Both components are immunochemically indistinguishable from the two plasma components and from heptadecapeptide porcine gastrin.

Among the disputed questions considered in this presentation are (1) Relationships between a) the known metabolic abnormalities in diabetes and the development of microangiopathy, and b) insulin antibodies and nodular glomerulosclerosis; (2) Sensitivity to insulin and availability of insulin in idiopathic diabetes mellitus and in other conditions associated with impaired glucose tolerance; (3) The role of pituitary growth hormone in the etiology of idiopathic diabetes; (4) The status of “bound insulin,” ILA, etc.; and (5) The identification of the “synalbumin” insulin antagonist as the B chain of insulin. Also included are the presentation of data on insulin and growth hormone-secretory responses to glucose loading in obese and nonobese diabetic and nondiabetic subjects and a discussion of the interrelationships among the peptide hormones, glucose and free fatty acids in blood.

The presence of a new form of immunoreactive insulin, “big, big insulin”, was first detected in the plasma of an insuloma suspect. On Sephadex gel filtration the elution volume of this insulin is approximately that of 125I-γ-globulin. Its sedimentation velocity on ultracentrifugation is between that of labeled albumin and labeled γ-globulin. Thus, it appears to have a molecular weight of about 100,000. Studies with starch gel and starch block electrophoresis reveal that it is a more basic protein than porcine or crystalline human insulin. It is immunochemically identical with crystalline human insulin, maintains its identity on refractionation on Sephadex gel filtration and is rapidly transformed by trypsin to an insulin-like component. “Big, big insulin” is a minor component (

Separation of Antibody-Bound and Unbound Peptide Hormones Labelled with Iodine-131 by Talcum Powder and Precipitated Silica

Five patients with active Hodgkin’s disease were treated for a period varying from four to eighteen days with an antiguanine preparation, guanazolo (5-amino-7-hydroxy-1H-v-triazolo[d]pyrimidine) with a total dosage of 400 mg. to 800 mg. intramuscularly. No therapeutic effect was observed. The only toxic manifestations were pain and tenderness at the site of injection in 4 of the 5 cases. The effect of guanazolo on tissue cultures of an involved node was tested. No inhibition of growth was observed, thus paralleling the lack of in vivo effect.

In a recent communication to this Journal, Bauer and Goodwin (1) reported 3 cases of gynecomastia appearing in patients treated with radioactive iodine for Graves’ disease. This symptom was considered as a complication of such therapy. We wish to call attention to tender breast enlargement as one manifestation of the symptomatology of Graves’ disease itself and to present 4 such cases in males observed in the past one and a half years among a total of 45 cases of hyperthyroidism examined during this period. In 2 of the cases, the appearance of breast symptoms antedated therapy of any kind. In 1 case, gynecomastia was first noted shortly after institution of therapy with Tapazole (1 methyl 2 mercaptoimidazole) but before significant clinical response was evident. In the fourth case, tender breast swelling was first noted later in the course of therapy, at a time when the patient had improved considerably. In all cases, remission of breast symptoms coincided with the return of function to the euthyroid state.

A sensitive and specific assay method has been used to demonstrate acute changes in plasma HCH in normal and diabetic subjects. HGH secretion is suppressed by glucose administration and markedly stimulated by hypoglycemia, by a rapid fall in blood glucose without hypoglycemia, and by interference with intracellular glucose utilization. High levels of HGH were also observed during prolonged fasting, after exercise, and four to six hours following oral glucose administration. Abnormalities in HGH secretion were found in association with acromegaly and obesity, and following section of the hypophyseal stalk. The half time for disappearance of endogenous plasma HGH is twenty to thirty minutes.

TO understand fully the role of any hormone in the bodily economy it is essential to know not only the effects produced by the hormone and the mechanism of action by which these effects are produce...

The role of serum albumin in the causation of cardiac edema has been disputed. Starling (1) postulated that circulatory stasis resulting from cardiac insufficiency is responsible for anoxia and increased permeability of the capillary membrane, the consequence of which is the passage of albumin and water into the extravascular spaces. Although Smirk (2) observed evidence of increased capillary permeability to water and crystalloids in congestive heart failure, Stead and Warren (3) demonstrated, by direct analysis, that the protein concentration of cardiac edema fluid was not elevated above that of the edema fluid produced by tourniquet stasis in normal subjects. Other studies have directed attention to the low serum albumin concentrations occasionally encountered in heart failure (4-7), and have suggested or implied (4, 8) a causal relationship to the pathogenesis of cardiac edema. Abnormalities in liver function tests and morphologic evidence of liver injury in severe congestive failure (9-14) have appeared to strengthen the possibility that hepatic synthesis of albumin may be impaired in this condition. Alternatively, low serum albumin concentrations have been attributed to inadequate dietary protein intake or poor absorption from a congested gastrointestinal tract (15, 16). Since the plasma concentration of albumin is not an adequate measure of the total quantity of circulating and extravascular albumin stores, it was of interest to measure the amount of total exchangeable albumin in subjects with heart failure employing I'8l labeled albumin as a tracer. The rate of metabolism of I18l labeled albumin was also studied with the aim of evaluating the ability of the subject in heart failure to synthesize albumin. Although there may yet remain some reservations regarding the validity of this tracer in the