STUDIES ON THE STATE OF INSULIN IN BLOOD: DISSOCIATION OF PURIFIED HUMAN BLOOD INSULIN COMPLEX(ES) BY INCUBATION WITH ADIPOSE TISSUE EXTRACTSIN VITRO¹

The nature of human serum insulin-like activity (ILA): characterization of ILA in serum and serum fractions obtained by acid-ethanol extraction and adsorption chromatography

“…The protein concentration of this fraction is equivalent to the amount of protein eluted per milliliter of serum applied. which is a two-to three-fold greater specific activity than that reported previously in assay systems employing adipose tissue extracts (34,41,60). Table VI shows the effect on glucose uptake in muscle of serum proteins in the NaCi wash and in the alkaline eluate in the presence of insulin antiserum.…”

“…Certainly, variations in the method of chromatography and in the bioassay procedure may account for this difference in part; since comparative studies with and without ATE were not done, this question remains unanswered. It should be noted, however, that the specific activities of several eluate fractions bioassayed without ATE moderately exceeded values recently reported by others employing ATE (34,41,60). Fractionated human serum ILA prepared by a variety of other methods was also biologically active on muscle in vitro (38).…”

“…From our results, it is difficult to make definite conclusions regarding the ionic nature of the insulin-like substances in human serum since studies of the fundamental mechanism of adsorption of proteins and other macromolecules onto ion exchange surfaces are lacking (67 Although the present studies corroborate the findings of Antoniades and coworkers that ILA is adsorbed onto Dowex, a perplexing discrepancy exists in the results obtained for the biological activity of this ILA on muscle. Based on data derived from the rat hemidiaphragm bioassay of serum and "bound insulin" preparations, with and without the presence of an extract of adipose tissue (ATE), it has been concluded that "bound insulin" is inactive on muscle unless dissociated or activated by a low molecular substance present in adipose tissue extracts (60,68). In our hands, however, the hemidiaphragm bioassay of the resin-adsorbed protein ("bound insulin") consistently yielded considerable biological activity in the absence of ATE.…”

Studies of the Insulin-inhibitory Effect of Human Albumin Fractions*

“…It may be that adipose tissue possesses enzymes capable of altering inhibitors. There is precedence in the reports of Antoniades and Gundersen (26) and Shaw and Shuey (27) for such a hypothesis. These investigators have shown that adipose tissue and adipose tissue extracts release insulin from a complexed form to a free form, but no such activity can be attributed to muscle.…”

Studies on Circulating Free and Bound Insulin

“…In the absence of ATE bound insulin preparations, or whole sera from fasting subjects, were biologically inactive on isolated muscle (9)(10). The possibility was suggested that ATE may stimulate tissue factors on hemidiaphragm, possibly enzymes, which in turn catalyzed the activation and utilzation of bound insulin.…”