Glucocorticoids and the fate of exogenous insulin in insulin-immunized guinea pigs

Diabetologia, Aug 1975

Summary Groups of normal rats and of normal and insulin-immunized guinea pigs were injected intravasculary with mixtures of radio-iodinated and unlabelled porcine insulins. No differences could be discerned between the fates of radio-iodinated and unlabelled insulins in normal or immune guinea pigs except with respect to the kidneys of normal animals, which seemed to accumulate TCA-precipitable radioactive substances devoid of immunoreactivity. The abnormally slow disappearance of insulin from the plasma and its persistent accumulation in the livers and spleens of immune guinea pigs were not affected by prior treatment for three days with large daily doses (ca 20 mg/kg body wt.) of hydrocortisone acetate.

A PDF file should load here. If you do not see its contents the file may be temporarily unavailable at the journal website or you do not have a PDF plug-in installed and enabled in your browser.

Alternatively, you can download the file locally and open with any standalone PDF reader:

https://link.springer.com/content/pdf/10.1007%2FBF00422391.pdf

Glucocorticoids and the fate of exogenous insulin in insulin-immunized guinea pigs

Glucocorticoids and the Fate of E x o g e n o u s Insulin in Insulin-Immunized Guinea Pigs P.H. Wright 0 R.L. Gingerich 0 J.R. Oliver 0 0 Dept. of Pharmacology, Indiana University School of Medicine , Indianapolis, Indiana , USA Summary. Groups of normal rats and of normal and insulin-immunized guinea pigs were injected intravasculary with mixtures of radio-iodinated and unlabelled porcine insulins. No differences could be discerned between the fates of radio-iodinated and unlabelled insulinsin normal or immune guinea pigs except with respect to the kidneys of normal animals, which seemed to accumulate TCA-precipitable radioactive substances devoid of immunoreactivity. The abnormally slow disappearance of insulinfrom the plasma and its persistent accumulation in the livers and spleens of immune guinea pigs were not affected by prior treatment for three days with large daily doses (ca 20 mg/kg body wt.) of hydrocortisone acetate. Insulin injection; insulin recovery; tissue insulin concentration; insulin immunized guinea pigs; hydrocortisone acetate - We have previously shown [ 3 ] that the fate of radio-iodinated porcine insulin injected into normal animals differs markedly from that found in insulinimmunized guinea pigs. In brief, TCA-precipitable radioactive material, presumed to represent unchanged hormone, concentrates transiently in the liver and kidneys of normal animals, but disappears more slowly from the plasma of insulin-immunized animals to accumulate more persistently and to a greater extent in their livers and spleens. In view of convincing evidence that heavily iodinated and biologically inactive insulin is not metabolized by normal rats either as rapidly or in the same manner as lightly labelled and biologically active hormone [ 4 ], the fates of commercially labelled and unlabelled insulins were compared in normal and immune guinea pigs. The influence of hydrocortisone acetate upon the fate of insulin in immune guinea pigs was also studied, since glucocorticoids have been shown to have the most consistent beneficial effects upon insulin resistance seen in some human diabetic patients [ 9 ]. The results show that very similar results can be obtained with the labelled and unlabelled hormones, and that pretreatment o f immune guinea pigs with large doses of hydrocortisone does not appear to have any effect upon the fate of subsequently injected insulin. Materials and Methods Animals Male albino guinea pigs (body wts., 0.83 to 1.64 kg) were either normal or had been immunized over a period of 8 to 9 months with porcine insulin by a standard method [ 13 ]. Levels of insulin-binding antibodies in the sera of these guinea pigs were estimated by the method of Makulu and Wright [ 5 ]. In an additional experiment, male albino rats (body wts., 178 to 216 g) were used. All animals had free access to food and water up to the time of intravascular insulin injection. Intravascular Inocula These consisted of mixtures of radio-iodinated and unlabelled porcine insulins dissolved in solutions of either normal or anti-insulin sera from guinea pigs. The ~25I-labelled (ca 100 ~tC/ml, 0.8 to 0.9 gg insulin/ ml; Cambridge Nuclear Corporation, Billerica, Mass.) and 131I-labelled (ca 130 gC/ml, 0.65 ~tg insulin/ml; Abbott Laboratories, North Chicago, Illinois) porcine insulins were used within three days of receipt after appropriate dilution. No attempt was made to purify further these samples of labelled hormone. Unlabelled porcine insulin (Lot Nos. PJ4489, 21.1U/mg, and PJ5682, 23.9 U/mg; Eli Lilly and Company, Indianapolis, Indiana) was dissolved in dilute acetic acid (0.6% glacial, v/v), the stock solution (100 U/ml) being stored at 4 ~ for later dilution immediately before use. These hormones were initially diluted in relatively small volumes of a buffered solution (Na2HPOa/KH2PO4, 0.05 M, p H 7.0) containing sodium chloride (0.4%, w/v) and normal guinea pig serum (NGPS; 3%, v/v); this buffered solution is here termed 3% NGPS. Final dilution of the hormones was carried out in relatively larger volumes of undiluted normal guinea pig serum (NGPS) or guinea pig antiinsulin serum (GPAIS). The GPAIS used for the experiment in rats (Lot No. 555, binding 1.53 Units porcine insulin/ml) was allowed to react with the insulin for 30 min at 36 ~ before it was injected. The precise compositions of inocula are given below with the results. Experimental Procedures Some animals were treated with hydrocortisone (Hydrocortone acetate, saline suspension; Merck, Sharpe and Dohme, West Point, PA) for 3 days before injection of insulin. Both rats (5 mg/day) and guinea pigs (20 mg/day) received relatively large sub' cutaneous doses. Labelled and unlabelled insulins which had previously been incubated with GPAIS were injected intravenously (tail vein) into rats which were killed 30 min later by decapitation. Guinea pigs, whether normal or immune, were all given an antihistamine (Chlorpheniramine maleate, 10 mg; Chlortrimeton maleate, Schering Corporation, Bloomfield, N.J'.) by subcutaneous injection, were then lightly anesthetized with ether and given the insulin inoculum (0.5 ml) by intracardiac injection; approximately 10 min elapsed between the two injections. No i m m u n e guinea pig was seen to develop signs of any acute allergic reaction; without prior treatment with an antihistamine, death occurs within a few minutes with signs of severe respiratory distress [ 12 ]. Guinea pigs were killed by decapitation 5 to 60 min after intracardiac injection. After decapitation, blood was collected from the severed necks into heparinized beakers and the plasma rapidly separated by centrifugation. If not used immediately, plasma samples were stored in the frozen state at - 1 0 ~C. The liver, kidneys and spleen of each animal (unless otherwise stated) were rapidly removed from the body and weighed. Weighed portions of each, asindicatedundertheheading"Results", were then homogenized or extracted, initial handling, precipitation or extraction being complete within 2 min: a) Tissue Homogenates. Weighed portions (ca 1.0 g) of liver and kidney were added to ice-cooled distilled water (10 ml) and homogenized (Polytron, Kinematica GMBH, Lucerne, Switzerland). Smaller portions were used for the spleens. The gamma-radioactive contents of aliquots (0.5 ml) of these homogenates were measured in an automatic well-type scintillation counter (Series 300, Model 5052, Packard Instrument Company, LaGrange, Illinois) either (i) without further treatment (Total radioactive content) or (ii) after precipitation of proteins with an equal volume (0.5 ml) of trichloracetic acid (TCA, 20%, w/v) and subsequent washing of the precipitate with more dilute (10%, w/v) T C A (TCA-precipitable radioactive content). Plasma (1.0 ml) was diluted in distilled water (10 ml) and then treated in the same way as the homogenates. The radioactive contents of these precipitates, solutions and homogenates are here expressed with respect to the wet wt. (g) or volume (ml) of tissue. b) Acid-Alcohol Extraction. Plasma (1.0 m l ) o r weighed aliquots (ca. 1.0 g) of liver or kidney were added to an ice-cooled solution (10 ml) of acid-alcohol (2.1%, v/v, concentrated sulfuric acid in 80 %, v/v, aqueous ethanol). The tissue was homogenized and the suspensions allowed to stand at 4 ~ C for 2 days. Precipitates were separated by centrifugation and an aliquot (1.0 ml) of the clear supernatant solution neutralized with a saturated solution of sodium bicarbonate (0.7 ml); the final pH of this solution was usually between 7.0 and 7.5. The insulin contents of these neutralized extracts, after removal of precipitates, were immediately determined using small aliquots (0.05 ml) and standard solutions of porcine insulin dissolved in the same neutralized solution of acid alcohol. The method of insulin immuno-assay in which 125I-labelled and unlabelled porcine insulins were used, has been described elsewhere [ 14 ]. Immunoreactive contents were related to the wet wts. (g) or volumes (ml) of tissue. For experiments in which only the fate of the labelled hormone was to be studied, 12SI-labelled porcine insulin was injected. When the fates of both labelled and unlabelled insulins were to be followed, 13li_labell_ ed porcine insulin was injected so that 12SI-labelled insulin could be used subsequently for the assay of immunoreactive insulin in tissue extracts. Separate pieces of the same tissue were used for assays of radioactive and immunoreactive coiatents. This was done to speed initial processing of the tissues and so reduced the possibility of insulin destruction by enzymes in those tissues. Unless Otherwise stated, all values are given as the m e a n s ( + SEM), differences b e t w e e n m e a n values being considered significant at or below the 5 % level of probability. All statistical analyses were carried out according to m e t h o d s described by Snedecor and Cochran [ 10 ]. Results Fates o f 3lI-Labelled and o f Unlabelled Porcine Insulins in Normal and Immune Guinea Pigs A n inoculum (0.5 ml) containing 131I-labelled (1.1 m U ) and unlabelled (1.0 U) porcine insulins, dissolved in a mixture of 3% N G P S (0.17 ml) and undiluted N G P S (0.33 ml),was injected into four groups of 5 or 7 n o r m a l or i m m u n e guinea pigs which were killed 5 or 30 min later. D e t a i l s concerning the animals used and of the concentrations of T C A - p r e c i p i t a b l e radioactivity and of i m m u n o r e a c t i v e insulin found in their tissues are given in T a b l e 1. In n o r m a l guinea pigs the concentrations of T C A precipitable radioactivity fell by a b o u t half b e t w e e n 5 and 30 min in the plasma, liver and spleen, and by a b o u t one quarter in the kidneys. Only in the kidneys, at b o t h 5 and 30 min, did the concentration of radioactivity exceed that in the plasma. In insulin~immunized guinea pigs, on the other hand, concentrations of T C A - p r e c i p i t a b l e radioactivity decreased m o r e slowly in the plasma but increased by almost 5 0 % in both the livers and spleens b e t w e e n 5 and 30 min. V e r y little radioactivity was r e c o v e r e d at either time f r o m the kidneys of these i m m u n e animals. A f t e r 30 min the m e a n concentration of radioactivity in the livers exceeded and that in the spleens equalled levels found in plasma. A similar, but not identical, p a t t e r n of distribution of injected i m m u n o r e a c t i v e h o r m o n e was o b s e r v e d in these same animals. Thus in n o r m a l guinea pigs the concentrations of r e c o v e r e d i m m u n o r e a c t i v e insulin f e l l by a b o u t half b e t w e e n 5 and 30 min in the p l a s m a and livers, but in i m m u n e guinea pigs it accumulated in the livers as it d i s a p p e a r e d m o r e slowly f r o m the plasma. In these two tissues, p l a s m a and liver, whether r e m o v e d after 5 or 30 min f r o m n o r m a l or i m m u n e animals, the ratios of r e c o v e r e d immunoreactivity to T C A - p r e c i p i t a b l e radioactivity ( m U / 1 0 0 cpm) were constant. Thus the ratios in n o r m a l animals for p l a s m a at 5 (0.177 + 0 . 0 3 1 ) a n d 30 (0.237 _+ 0 . 0 5 5 ) m i n and in i m m u n e animals at 5 (0.182 + 0.007) and 30 (0.216 _+ 0.028) rain were not significantly different f r o m one another; this was also true of livers f r o m these same four groups of animals. T h e m e a n ratio for all these p l a s m a samples (0.201 + 0.016, n = 21) was n o t s i g n i f i c a n t l y d i f f e r e n t f r o m t h a t c a l c u l a t e d for t h e c o r r e s p o n d i n g livers ( 0 . 1 6 9 + 0 . 0 2 1 , n = 22), b u t b o t h r a t i o s w e r e s i g n i f i c a n t l y l o w e r t h a n t h e o b s e r v e d r a t i o in t h e i n o c u l u m ( 0 . 2 6 4 m U / 1 0 0 c p m ) . L i t t l e i m m u n o r e a c t i v e o r r a d i o a c t i v e m a t e r i a l was r e c o v e r e d f r o m t h e k i d n e y s of i m m u n e a n i m a l s . R e l a t i v e l y high c o n c e n t r a t i o n s of b o t h w e r e f o u n d a f t e r 5 m i n in t h e k i d n e y s of n o r m a l a n i m a l s w h e n t h e r a t i o ( 0 . 2 1 8 + 0 . 0 2 2 m U / 1 0 0 0 c p m ) was n o t s i g n i f i c a n t l y d i f f e r e n t f r o m t h o s e q u o t e d a b o v e for t h e livers a n d p l a s m a . A f t e r 30 min, h o w e v e r , t h e i m m u n o r e a c t i v e c o n t e n t s of t h e k i d n e y s of t h e s e n o r m a l a n i m a l s h a d f a l l e n to a m u c h g r e a t e r e x t e n t t h a n h a d t h e i r r a d i o a c t i v e c o n t e n t s , t h e r a t i o ( 0 . 0 9 6 + 0 . 0 2 3 m U / 1 0 0 0 c p m ) t h e n b e i n g s i g n i f i c a n t l y l o w e r t h a n t h e v a l u e f o u n d a f t e r 5 min. Effects o f Treatment with Hydrocortisone upon the Fates o f Unlabelled and lodinated Insulins in Rats and Guinea Pigs G r o u p s of 7 to 9 rats a n d g u i n e a pigs w e r e given d a i l y s u b c u t a n e o u s i n j e c t i o n s of h y d r o c o r t i s o n e for 3 d a y s ; c o n t r o l a n i m a l s r e c e i v e d n o i n j e c t i o n s . T h e rats w e r e t h e n i n j e c t e d i n t r a v e n o u s l y with i n s u l i n - a n t i b o d y c o m p l e x e s f o r m e d b y p r e - i n c u b a t i o n of 125I-lab e l l e d a n d u n l a b e l l e d p o r c i n e insulins with a slight excess of G P A I S . T h e i n s u l i n - i m m u n i z e d g u i n e a pigs, u n d e r light e t h e r a n e s t h e s i a a n d a f t e r a d m i n i s t r a t i o n of an a n t i h i s t a m i n e , r e c e i v e d i n t r a c a r d i a c i n j e c t i o n s of i n o c u l u m c o n t a i n i n g r a d i o - i o d i n a t e d a n d u n l a b e l l e d insulin d i s s o l v e d in N G P S , a n d w e r e k i l l e d b y d e c a p i t a t i o n 30 o r 60 m i n l a t e r . D e t a i l s of e a c h g r o u p of ~, 100t !~Q ~ 40-~ 0 12 8 < N 4 v 0 Fig. 1. TCA-precipitable radioactive and immunoreactive contents (Mean + SEM) of tissues removed from hydrocortisone-treated (cross-hatched) and control (stippled) insulin-immunized guinea pigs killed 30 rain after intracardiac injection of a mixture of 131Ilabelled (1.1 mU) and unlabelled (1.0 U) porcine insulins. Each column represents the mean of observations in 7 or 8 animals. Further experimental details are given in the text and were comparable with those shown in Table 2, Expt. "C" animals, of their tratments and of the TCA-precipitable and immunoreactive contents of their tissues are shown in Table 2 and Fig. 1. In the first experiment (A) normal rats were killed 30 min after injection of heterologous insulin-antibody complexes. As shown in Table 2, the mean concentrations of TCA-precipitable radioactivity found in the plasma, livers and spleens of the hydrocortisone-treated rats were not significantly different from those found in the same tissues of control animals. Based on the mean plasma volume (4.30 + 0.13 ml/100 g body wt.) found in a previous studyof similar rats (15) and the known weights of the organs concerned, it is also seen that total recoveries from plasma and the other tissues do not differ in the two groups. In the second experiment (B) immune guinea pigs were killed 60 min after injection of a mixture of ~25I-labelled and unlabelled (200 mU) porcine insulins. Mean concentrations of TCA-precipitable radioactivity in the plasma and spleens of hydrocortisonetreated animals were not significantly different from those for the controls, but the mean concentration in livers of treated guinea pigs was significantly (p < 0.025) and appreciably (33%) lower than the control value. When the published value [ 7 ] for plasma volume in guinea pigs (3.94 ml/100 g body wt.) and the known weights of the various organs were used f o r calculations, the total recoveries from the plasma and other tissues of treated guinea pigs were not significantly different from the controls. In the third experiment (C) immune guinea pigs were killed 30 min after injection of 131I-labelled insulin mixed with t h e large dose (1.0 U) of unlabelled hormone. On this occasion, no significant differences were found between the concentrations of TCA-precipitable radioactivity in the plasma, livers, spleens or kidneys of the control and hydrocortisone-treated animals; the same applied to calculated total recovery rates. Similarly, and as illustrated on Fig. 1, no distinction could be made between treated and control animals on the basis of recovered concentrations of immunoreactive insulin in these same tissues. Discussion Before considering possible implications of these findings, three brief comments are indicated. First, total recoveries of radioactive and immunoreactive insulin from the various tissues have been calculated on the basis of assumed plasma volumes or on the assumption that the weighed organs contained no trapped blood. As a result, for example, it is found that total recoveries of TCA-precipitable radioactivity from the plasma, livers, kidneys, and spleens of immune guinea pigs killed 5 (134%) and 30 (111%) min after injection (Table I) exceed 100%. In the absence of specific knowledge of the plasma volumes and of the blood or plasma contents of the tissues, any calculated values for total recovery can only be considered as approximate. Secondly, the low but constant measured ratio of immunoreactive to radioactive contents of tissues removed from injected animals (0.17-0.20 mU/1000 cpm) compared to that (0.26 mU/1000 cpm) of the inoculum probably reflects a low but uniform rate of insulin recovery (60-70%) during acid-alcohol extraction. Finally, the low ratio of immunoreactive to radioactive contents in kidneys from normal guinea pigs killed 30 rain after injection (0.096 mU/1000 cpm) was not further investigated. It is known, however, that iodinated insulin can retain TCA-precipitability whilst losing its immunoreactive and/or its biological functions. Thus Narahara et al. [ 8 ] could only convert 60 to 70% of the TCA-precipitable material in radio-iodinated insulin to a TCA-soluble form by incubation in vitro with liver homogenates. Similarly, Malaisse et al. [ 6 ] could only achieve about 70% conversion to TCA-soluble material by incubation of radio-iodinated insulin with pieces of pancreatic tissue. During the same time, however, ability of the labelled hormone to adsorb onto cellulose powder was completely lost. These latter observations suggest that the TCA-precipitable material persisting in the kidneys of normal guinea pigs (Table 1) probably includes degradation products of the hormone which have lost immunoreactivity but have retained TCAprecipitability. The results themselves show that radio-iodinated and unlabelled exogenous insulins are utilized in vivo in much the same ways. Theyparticularlydemonstrate that insulin is utilized in a markedly abnormal way by immune animals, concentrating as it does in those tissues, the liver and spleen, which contain large numbers of fixed tissue macrophages. Such an abnormal fate was also envisaged by Berson and Yalow [ 1 ] for insulin injected into human diabetic patients having high levels of circulating insulin antibodies and high requirements for insulin. They postulated that "owing to its degradation with antibody, a large fraction of the bound (injected) insulin never becomes available to the tissues." The nature of this degradation process is not known, but if it resembles that involved in the destruction of other antigen/antibody complexes then, as Berson and Yalow later suggested [ 2 ], injected insulin might be destroyed by such patients at rates of the order of 500 to 1000 Units/day. Such resistance to therapy with insulin is not common but "the adrenal steroids and ACTH," wrote Shipp et al. [ 9 ], "have most consistently altered insulin resistance;" the doses of insulin needed to maintain metabolic control often drop dramatically within a matter of days. The reason for the success of this treatment is not known, but glucocorticoids are known to influence immune processes. Vernon-Roberts [ 11 ] in a recent review has pointed out that small doses of glucocorticoids stimulate and large doses inhibit phagocytic activity in guinea pigs, rats and mice. On the assumption that phagocytosis might also be playing some part in the removal of insulin-antibody complexes from the plasma of immune guinea pigs, and that a similar phenomenon might be responsible for insulin resistance in the human diabetic, large doses of hydrocortisone were given to the present immune animals. However, as is clearly shown in Table 2 and Fig. 1, prior treatment of guinea pigs and rats for 3 days did not influence the fates of injected insulin or of insulinantibody complexes. There was a suggestion that uptake of insulin by the livers of one group of immune guinea pigs (Expt. No. B) was reduced, but this was not confirmed in the second study (Expt. No. C). It is conceivable that the experimental conditions were not ideal, that the period of treatment with cortisone was too short, or that the species of animal used here was not suitable, but the present results do not support the contention that glucocorticoids inhibit immunological destruction of exogenous insulin in the immune animal. Acknowledgements. This investigation was supported by the U.S. Public Health Service (Grant No. PHS AM 16534-02). The authors wish to thank Eli Lillyand Company, Indianapolis, Indiana, for insulinpreparations; and Mrs. Vivian Williams and Mrs. Debra Stricklin for skilled technical assistance. 1. Berson , S.A. , Yalow , R.S.: Insulin antagonists, insulin antibodies and insulin resistance . Amer. J. Med . 25 , 155 - 159 ( 1958 ) 2. Berson , S.A. , Yalow , R.S.: The present status of insulin antagonists in plasma . Diabetes 13 , 247 - 259 ( 1964 ) 3. Frikke , M.J. , Gingerich , R.L. , Stranahan , P.D. , Carter , G. , Bauman , A.K. , Greider , M.H. , Wright , P.H. , Lacy , P.E. : Distribution of injected insulin and insulin-antibody complexes in normal and insulin-immunized animals . Diabetologia 10 , 345 - 351 ( 1974 ) 4. Izzo , J.L. , Bartlett , J.W. , Roncone , A. , Izzo , M.J. , Bale , W.F. : Physiologicalprocesses and dynamicsin the disposition of small and large doses of biologicallyactive and inactive 131I-insulinin the rat . J. biol. Chem . 242 , 2343 - 2355 ( 1967 ) 5. Makulu , D.R. , Wright , P.H. : Immune response to insulin in guinea pigs: some quantitative studies . Metabolism 20 , 770 - 781 ( 1971 ) 6. Malaisse , W. , Malaisse-Lagae , F. , Wright , P.H.: A new method for the measurement in vitro of pancreatic insulin secretion . Endocrinology 80 , 99 - 108 ( 1967 ) 7. Masouredis , S.P. , Melcher , L.R. : Blood, plasma and "Globulin" space of guinea pigs determined with ITM rabbit globulin . Proc. Soc. exp. Biol . (N.Y.) 78 , 264 - 266 ( 1951 ) 8. Narahara , H.T. , Tomizawa , H.H. , Miller , A. , Williams , R.H. : Intracellular distribution of an insulin-inactivating system of liver . J. biol. Chem . 217 , 675 - 684 ( 1955 ) 9. Shipp , J.C. , Cunningham , R.W. , Russell , R.O. , Marble , A. : Insulinresistance: clinicalfeatures, naturalcourse and effectsof adrenal steroid treatment . Medicine (Baltimore) 44 , 165 - 186 ( 1965 ) 10. Snedecor , G.W. , Cochran , W.G.: Statistical Methods . Iowa State University Press 1967 11. Vernon-Roberts , B. : The effects of steroid hormones on macrophage activity . Int. Rev. Cytol . 25 , 131 - 159 ( 1969 ) 12. Wright , P.H. : The effect of insulinantibodies on glucoseuptake by the isolated rat diaphragm, biochem . J . 71 , 633 - 638 ( 1959 ) 13. Wright , P.H. , Makulu , D.R. , Posey , I.J.: Guineapig anti-insulin serum; adjuvant effect of H. pertussis vaccine . Diabetes 17 , 513 - 516 ( 1968 ) 14. Wright , P.H. , Makulu , D.R. , Vichiek , D. , Sussman , K.E. : Insulin immunoassay by back-titration; some characteristics of tl~e techniques and the insulin precipitant action of alcohol . Diabetes 20 , 33 - 45 ( 1971 ) 15. Wright , P.H. , Rivera-Calimlim , L. , Malaisse , W.J.: Endogenous insulinsecretion in the rat followinginjectionof anti-insulin serum . Amer. J. Physiol . 211 , 1089 - 1094 ( 1966 ) P.H. Wright , M. D. Dept. of Pharmacology Indiana Univ. School of Medicine 1100 West Michigan Street Indianapolis, Indiana 46202 USA


This is a preview of a remote PDF: https://link.springer.com/content/pdf/10.1007%2FBF00422391.pdf

P. H. Wright, R. L. Gingerich, J. R. Oliver. Glucocorticoids and the fate of exogenous insulin in insulin-immunized guinea pigs, Diabetologia, 1975, 273-278, DOI: 10.1007/BF00422391