Josiah Brown

Autoimmune Thyroid Diseases—Graves' and Hashimoto's

Abstract Thyroid-related autoimmune diseases (Graves thyroid disease, Graves ophthalmopathy, and Hashimotos thyroiditis) may occur alone or in any association. The diagnosis of Hashimotos thyro...

Fetal Pancreas Transplantation for Reversal of Streptozotocin-induced Diabetes in Rats

Streptozotocin-induced diabetes in rats was completely reversed by transplantation of syngeneic fetal pancreases placed beneath the kidney capsule. To accomplish complete reversal of diabetes, four or more pancreases were necessary; three resulted in partial reversal, and two produced a slight but significant effect in some recipients. Removal of the transplants resulted in the prompt return of diabetes. The islets of Langerhans in the transplants functioned homeostatically; this was indicated by regular normal blood glucose values, in addition to normal findings in blood IRI response and glucose disappearance rate after glucose injection. Disappearance of exocrine elements, with only ducts and fibrous tissue remaining, resulted in a pure endocrine organ. The advantages of this technic, such as ease of accessibility for placement, observation, and removal, are of great importance for possible application to humans.

Importance of hepatic portal circulation for insulin action in streptozotocin-diabetic rats transplanted with fetal pancreases.

The importance of the hepatic portal circulation in the response to insulin was assessed in streptozotocin-diabetic rats transplanted with syngeneic fetal pancreases. Partial reversal of diabetes was accomplished by transplantation of two or three fetal pancreases beneath the capsule of the kidney; complete reversal followed shunting of the venous drainage from the transplants to the liver. Plasma glucose after streptozotocin of 509+/-31 mg/dl (mean+/-SEM) fell after transplantation to 395+/-23 and after the shunt to 143+/-5 mg/dl. Urine volume fell from 84+/-4 to 50+/-5 ml/d and then to normal (17+/-1 ml/d) after the shunt. Glucose excretion which was 8.1+/-0.3 g/d after streptozotocin fell after transplantation to 4.8+/-0.3 g/d and after the shunt completely disappeared from the urine. The disappearance rate of glucose injected into the circulation, which was 0.50+/-0.07%/min in untreated diabetes, increased to 1.39+/-0.38%/min after transplantation and to 2.52+/-0.31%/min after the shunt, not different from normal controls (2.79+/-0.25). Plasma immunoreactive insulin (IRI) was below normal (25-35 muU/ml) and unresponsive to glucose in untreated diabetic rats. After transplantation IRI levels ranged from 73-223 muU/ml and there was no rise after glucose injection. After the shunt both the basal IRI (36+/-5 muU/ml) and the peak response to glucose at 10 min (58+/-7 muU/ml) were the same as in normal controls (42+/-4 and 62+/-7 muU/ml, respectively). The fall in IRI after the shunt is explained by increased extraction of insulin passing into the liver and also diminished secretion. After removal of the transplants plasma glucose and urine values returned almost to pretransplant levels. Secretion of insulin by transplanted pancreases into the liver enhances the effectiveness probably by increased extraction and action and reveals the importance of the normal route for insulin delivery.

Islet Cells Grow after Transplantation of Fetal Pancreas and Control of Diabetes

The capacity of a fetal rat pancreas to grow and function was assessed after its transplantation into adult diabetic rats. One month after induction of diabetes by streptozotocin (SZ) (72.5 mg/kg) in adult Lewis rats, one syngeneic 17-17½ day fetal pancreas was transplanted under the kidney capsule. Insulin, initially 3 U/day and decreasing to 0.5 U every other day, was given for 27.5 ± 1.7 (× ± SEM) days (total dose, 48 ± 4 U) until the diabetes reversed (45 ± 3 days). Plasma glucose, 481 ± 10 mg/dl after SZ, fell after transplantation and cessation of insulin injections during a 4-mo period to 129 ± 2 mg/dl. Urine volume fell from 82 ± 4 ml per day to normal (12 ± 2), and glucose excretion, which was 7.7 ± 0.4 g per day after SZ, completely disappeared from the urine. The disappearance rate of glucose injected into the circulation, which was 0.50 ± 0.07% per minute in untreated diabetes, reverted to normal, 2.78 ± 0.18% per minute, not different from that in the normal control, 2.55 ± 0.13% per minute. Plasma insulin in control diabetic rats was totally unresponsive to glucose injection, despite a rise in plasma glucose concentration from 471 ± 5 mg/dl to 621 ±18 mg/dl. In transplanted rats, plasma insulin was moderately higher than normal before and at most time points after glucose injection. After removal of the transplanted pancreases, plasma glucose, urine volume, and glucose excretion returned to pretransplant levels and ketones appeared in the urine of all rats. Analysis of the insulin content of the pancreases removed 77 to 279 days after transplantation gave a value of 818 ± 43 mil, which is 22% of the insulin content of normal rats. To achieve optimal growth and function of a transplanted fetal pancreas, careful control of the blood glucose is necessary during the period of growth and development.

Hexokinase Isoenzymes in Liver and Adipose Tissue of Man and Dog

A hexokinase, with a low Michaelis constant, not previously described, has been found in extracts of human and dog liver but not of rat liver. Earlier reports are contradicted in that glucokinase occurs in extracts of liver from well-nourished humans and dogs; it is absent, or almost so, during states of poor nutrition.

Cryopreservation of human fetal pancreas.

To determine the optimal conditions for successful cryopreservation of the human fetal pancreas, techniques developed for the rat organ were modified. The parameters studied were the conditions of exposure to the cryopreservation agent dimethyl sulfoxide (DMSO), the rate of cooling and thawing, and the effect of in vitro culture. A total of 33 pancreases, obtained after fetal death due to prostaglandin-induced abortion, was studied. Survival of the pancreas is based on incorporation of 3H-amino acids into protein by pancreas pieces (2 mm3) during a 4-h incubation compared with nonfrozen control pieces from the same pancreas. Toxicity of DMSO at 37°C was found to be severe after a 4-h exposure. Varying the effects of temperature, time of exposure, and concentrations of DMSO on survival after freeze-thaw revealed that 1.5 M DMSO for 1 h at room temperature was optimal. The cooling rate was 0.22°C/min and thawing was at room temperature. Since these conditions resulted in only 50% survival, a period of culture before exposure to DMSO was added. The optimal duration of culture was 12–16 h. Using this method with addition of culture for 24 h after thawing, survival has varied from 70 to 120% of control. If a functional test for growth and insulin production by the frozen-thawed pancreas is positive, permanent shortage of the human fetal pancreas will be possible.

Reversal of diabetes in rats using fetal pancreases stored at -196 C.

Fetal rat pancreases frozen to and stored at -196 C were used for transplantation into streptozotocin-induced diabetic syngeneic adult recipients. Transplantation was carried out either directly after thawing from -196 C, or after a 21-day growth period in a syngeneic, normoglycemic adult carrier. All transplants were placed under the kidney capsule. A single, frozen fetal rudiment was sufficient to restore blood glucose, urine volume, and urine glucose to normal, provided it had first been grown for 21 days in a normal carrier. It vitro perfusion studies showed that fetal pancreases stored at -196 C were equivalent to fresh rudiments in their responses to a glucose stimulus.

Thyroid Physiology in Health and Disease

Abstract The pituitary thyrotrophin reserve in 55 patients was tested with thyrotrophin-releasing hormone, and only 28% with deficient responses were hypothyroid; in three patients with hypothalami...

Pancreas Transplantation for Diabetes Mellitus

Abstract Despite the best efforts of physicians and diabetic patients in the use of insulin for control of juvenile-onset (insulin-deficient) diabetes, vascular complications occur in most patients...

Fetal pancreas as a donor organ

AbstractThe use of the fetal pancreas as a donor organ for the treatment of diabetes has been explored in the rat and currently is being tried in minipigs. Growth and development of a fetal pancreas under the kidney capsule of a diabetic rat is sufficient to completely reverse streptozotocin diabetes; the organ contains 20–25% of the insulin content of a normal adult pancreas. The fetal pancreas grows and develops most fully in normoglycemic environment. Cryopreservation results in nearly complete survival of the pancreas and provides time for tissue and blood typing and testing for compatibility of donor and recipient lymphocytes using the Mixed Leucocyte Reaction. These methods have been developed in human systems. Removal of the pancreas from the fetus and transplantation prior to a critical period of exocrine development (18 days in the rat, 55 in the pig) are followed by atrophy of exocrine elements and a pure endocrine organ results. This obviates the problem of exocrine enzymes. Venous drainage of insulin from the transplanted pancreas into the liver provides the most effective response when the supply of insulin is limited. Metabolic studies in successfully-transplanted rats indicate complete reversal of the diabetic state to normal. These include blood glucose and insulin levels and responses to stimuli, the activity levels of 6 enzymes in the liver, glucagon levels, and the response to arginine stimulation and glucose metabolism during and after pregnancy.nMethods that have been used to prevent rejection, all of which have been partially effective, include donor liver pretreatment and short-term immunosuppression, total lymphoid irradiation combined with donor bone marrow, and treatment of the donor organ to eliminate immune cells by digestion and culture. Additional research will be required to develop an effective and safe method for prevention of allograft rejection in the minipig prior to application to diabetic humans.RésuméLe pancréas foetal est utilisé comme organe donneur pour le traitement du diabète chez le rat et est actuellement testé chez le cochon miniature. Un pancréas foetal greffé sous la capsule rénale dun rat rendu diabétique croît et se différencie suffisamment pour faire régresser complètement le diabète induit par la streptozotocine; le contenu en insuline du pancréas foetal atteint 20 à 25% de celui dun organe adulte normal. Cest en milieu normoglycémique que la croissance et la différenciation du pancréas foetal sont le plus complètes. La cryoconservation assure une survie de la quasi-totalité du pancréas et laisse un temps suffisant pour le typage sanguin et tissulaire et les tests de compatibilité des lymphocytes du donneur et du receveur en culture mixte de lymphocytes. Ces méthodes sont en cours délaboration pour les tissus humains. Lexérèse du pancréas foetal et sa greffe avant la période critique du développement exocrine (18 jours chez le rat, 55 chez le porc) provoque latrophie des éléments exocrines et il en résulte un organe purement endocrine, ce qui résoud le problème des enzymes des sécrétions exocrines. Cest le drainage portal de linsuline du greffon qui donne le meilleur résultat lorsque la production dinsuline est limitée. Les paramètres métaboliques des rats diabétiques transplantés avec succès reviennent complètement à la normale; en particulier la glycémie et linsulinémie et leur réponse aux stimuli, les activités de 6 enzymes hépatiques, les taux de glucagon et leur réponse à la stimulation par larginine et le métabolisme du glucose pendant et après une grossesse.Parmi les méthodes utilisées pour prévenir le rejet, et qui toutes ont montré une efficacité partielle, figurent le prétraitement du foie du donneur et limmunosuppression, lirradiation lymphoïde totale associée à la moëlle osseuse de donneur, et le traitement de lorgane donneur pour éliminer les cellules immunitaires par digestion et mise en culture. Des recherches supplémentaires sont nécessaires pour mettre au point une méthode efficace et sûre de prévention du rejet des hétérogreffes chez le porc miniature avant lapplication au diabète humain.ResumenEl uso del páncreas fetal como órgano donante en el tratamiento de la diabetes ha sido explorado en la rata y actualmente está siendo ensayado en cerdos. El crecimiento y desarrollo del páncreas fetal colocado bajo la cápsula del riñón de una rata diabética son suficientes para revertir en forma completa la diabetes inducida por estreptozotocina; el órgano así transplantado exhibe un 20–25% del contenido de insulina de un páncreas adulto normal. El páncreas fetal se desarrolla al máximo en un ambiente normoglicémico. La criopreservación resulta en supervivencia casi completa del páncreas y representa ganancia de tiempo para la tipificación tisular y sanguínea y para realizar pruebas de compatibilidad de los linfocitos donantes y recipientes mediante la Reacción de Linfocitos Mixtos. Estos métodos han sido desarrollados en sistemas humanos. La excisión del pancreas del feto y su transplante antes del período crítico de desarrollo exocrino (18 días en la rata, 55 en el cerdo) produce atrofia de los elementos exocrinos y da como resultado un órgano endocrino puro, lo cual elimina el problema de la enzimas exocrinas. El drenaje venoso de la insulina que secreta el páncreas transplantado hacia el hígado hace posible la más efectiva respuesta en condiciones en las cuales la provisión de insulina es limitada. Estudios metabólicos en ratas exitosamente trasplantadas indican la reversión completa de la diabetes a un estado normal. Tales estudios incluyen niveles de glucosa y de insulina sanguínea y su respuesta a estímulos, niveles de actividad de 6 enzimas en el hígado, niveles de glucagón y su respuesta a estimulación con arginina y el metabolismo de glucosa durante y después del embarazo.Los métodos utilizados para prevenir el rechazo, los cuales son parcialmente efectivos, incluyen el pretratamiento con inyecciones intravenosas de extracto de hígado adulto donante e inmunosupresión a corto término, la irradiación linfocítica total combinada con inyección de médula ósea adulta donante y el tratamiento del órgano donante, o sea el páncreas fetal, por medio de la digestión y el cultivo para eliminar las células inmunes. Se requiere realizar investigación adicional para el desarrollo de un método efectivo y seguro que logre prevenir el rechazo del alotransplante en el cerdo antes de su aplicación a diabéticos humanos.